1 00:00:04,922 --> 00:00:06,657 GOOD MORNING, EVERYBODY. 2 00:00:06,657 --> 00:00:08,526 WE'RE GOING TO TRY TO START ON 3 00:00:08,526 --> 00:00:09,627 TIME BECAUSE WE HAVE A FULL 4 00:00:09,627 --> 00:00:11,395 SCHEDULE AND WE'RE VERY EXCITED. 5 00:00:11,395 --> 00:00:17,301 SO ON BEHALF LF PARK MY 6 00:00:17,301 --> 00:00:19,103 CO-CHAIR, WHICH HE BASICALLY HAS 7 00:00:19,103 --> 00:00:20,738 DONE ALL THE WORK AND SAID OH 8 00:00:20,738 --> 00:00:23,107 YOU SHOULD STAND UP HERE AND I'M 9 00:00:23,107 --> 00:00:25,409 NAOMI TAYLOR, YOU SHOULD ALL 10 00:00:25,409 --> 00:00:28,579 KNOW ME HAD U NETS A HUGE 11 00:00:28,579 --> 00:00:29,613 PLEASURE TO HAVE YOU HERE. 12 00:00:29,613 --> 00:00:31,315 I'LL SAY A COUPLE WORDS BUT I 13 00:00:31,315 --> 00:00:34,685 WANTED TO TAKE THE OPPORTUNITY 14 00:00:34,685 --> 00:00:38,422 TO INTRODUCE GLENN MERLINO AND 15 00:00:38,422 --> 00:00:40,825 THANK GLENN MERLINO AND ALL THE 16 00:00:40,825 --> 00:00:41,092 DIRECTION. 17 00:00:41,092 --> 00:00:42,293 CENTER FOR CANCER RESEARCH FOR 18 00:00:42,293 --> 00:00:43,361 THEIR INCREDIBLE SUPPORT AND 19 00:00:43,361 --> 00:00:44,462 REALLY A HUGE THANKS FOR 20 00:00:44,462 --> 00:00:47,698 STARTING THIS MEETING. 21 00:00:47,698 --> 00:00:49,300 >> THANKS, NAOMI. 22 00:00:49,300 --> 00:00:53,004 I AM REALLY EXCITED TO BE HERE. 23 00:00:53,004 --> 00:00:55,006 THE IMMUNOLOGY MEETINGS HERE ARE 24 00:00:55,006 --> 00:00:56,674 ALWAYS THE BEST, IN MY OPINION, 25 00:00:56,674 --> 00:00:58,209 AND THIS I'M SURE LOOKING AT THE 26 00:00:58,209 --> 00:00:59,410 SCHEDULE AND SEEING WHO IS IN 27 00:00:59,410 --> 00:01:01,579 THE AUDIENCE, THIS WILL BE NO 28 00:01:01,579 --> 00:01:01,846 DIFFERENT. 29 00:01:01,846 --> 00:01:03,881 I WOULD LIKE TO WELCOME YOU TO 30 00:01:03,881 --> 00:01:04,982 BETHESDA, FOR THOSE OF YOU 31 00:01:04,982 --> 00:01:06,517 DHEENT LIVE HERE, AND TO THIS 32 00:01:06,517 --> 00:01:07,618 WONDERFUL MEETING, AND I WOULD 33 00:01:07,618 --> 00:01:11,322 ALSO LIKE TO DO SO ON BEHALF 34 00:01:11,322 --> 00:01:13,491 THOMAS BELLY AND CCR RESEARCH. 35 00:01:13,491 --> 00:01:15,393 I'M GOING TO SPEND A COUPLE 36 00:01:15,393 --> 00:01:16,961 MINUTES SO YOU CAN GET RIGHT TO 37 00:01:16,961 --> 00:01:18,262 T BUT THIS IS GOING TO BE A 38 00:01:18,262 --> 00:01:20,598 LITTLE BIT OF A CCR-CENTRIC 39 00:01:20,598 --> 00:01:20,931 INTRODUCTION. 40 00:01:20,931 --> 00:01:22,199 FOR THOSE ON THE OUTSIDE, YOU 41 00:01:22,199 --> 00:01:23,300 GET TO SEE A LITTLE BIT ABOUT 42 00:01:23,300 --> 00:01:24,668 WHAT WE'RE DOING HERE AND WHAT 43 00:01:24,668 --> 00:01:24,969 WE'RE ABOUT. 44 00:01:24,969 --> 00:01:26,170 FOR THOSE OF YOU WHO LIVE AND 45 00:01:26,170 --> 00:01:27,571 WORK HERE, MAYBE YOU'LL REMEMBER 46 00:01:27,571 --> 00:01:29,273 HOW GREAT THIS PLACE REALLY IS 47 00:01:29,273 --> 00:01:31,509 FROM THE INTRODUCTION. 48 00:01:31,509 --> 00:01:33,344 AND THANK YOU NAOMI AND I'LL 49 00:01:33,344 --> 00:01:36,113 COME BACK TO THAT FOR ALL YOU 50 00:01:36,113 --> 00:01:38,182 ORGANIZERS AT THE END. 51 00:01:38,182 --> 00:01:38,449 OKAY. 52 00:01:38,449 --> 00:01:40,418 SO BECAUSE OF OUR SITUATION HERE 53 00:01:40,418 --> 00:01:45,523 IN TERMS OF HOW WE DO BUSINESS 54 00:01:45,523 --> 00:01:47,758 AND HOW WE GET FUNDED AND THE 55 00:01:47,758 --> 00:01:49,293 RESOURCES THAT WE HAVE, IT GIVES 56 00:01:49,293 --> 00:01:53,431 US REALLY SCIENTIFIC -- GIVES US 57 00:01:53,431 --> 00:01:56,934 REALLY AMAZING SCIENTIFIC 58 00:01:56,934 --> 00:01:58,702 FREEDOM, BY SOLVING THE MOST 59 00:01:58,702 --> 00:01:59,904 IMPORTANT CHALLENGING AND 60 00:01:59,904 --> 00:02:02,073 NEGLECTED PROBLEMS IN CANCER 61 00:02:02,073 --> 00:02:03,174 RESEARCH, PREVENTION AND ALSO 62 00:02:03,174 --> 00:02:04,942 PATIENT CARE, AND OUR VISION IS 63 00:02:04,942 --> 00:02:06,577 SIMPLY TO CREATE THE MEDICINES 64 00:02:06,577 --> 00:02:08,212 OF TOMORROW. 65 00:02:08,212 --> 00:02:11,649 SO CCR IS A BIG PLACE. 66 00:02:11,649 --> 00:02:13,117 NCI IS A BIGGER PLACE. 67 00:02:13,117 --> 00:02:16,153 BUT WE ARE THE LARGEST OF THE 68 00:02:16,153 --> 00:02:19,190 INTRAMURAL COMPONENTS, AND WE DO 69 00:02:19,190 --> 00:02:20,791 FROM SOUP TO NUTS. 70 00:02:20,791 --> 00:02:21,725 WE HAVE VERY STRONG BASIC 71 00:02:21,725 --> 00:02:23,694 RESEARCH PROGRAM, WE DO 72 00:02:23,694 --> 00:02:25,930 TRANSLATION, CLINICAL CARE, AND 73 00:02:25,930 --> 00:02:28,265 WE HAVE, AS YOU'LL SEE, THE 74 00:02:28,265 --> 00:02:30,401 LEARNLGST CLINICAL RESEARCH 75 00:02:30,401 --> 00:02:32,236 CENTER IN THE -- LARGEST 76 00:02:32,236 --> 00:02:34,171 CLINICAL VERCH IN THE WORLD. 77 00:02:34,171 --> 00:02:39,009 TODAY COURAGE WE HAVE 233 PI'S 78 00:02:39,009 --> 00:02:40,878 ORGANIZED INTO FOUR FOUR 79 00:02:40,878 --> 00:02:42,179 BRANCHES, CENTERS AND PROGRAMS 80 00:02:42,179 --> 00:02:43,681 MOSTLY DPEENLDING ON OUR MODE I 81 00:02:43,681 --> 00:02:44,281 THINK MORE THAN ANYTHING ELSE, 82 00:02:44,281 --> 00:02:48,185 WE HAVE TWO WIG BIG CAMPUSES, 83 00:02:48,185 --> 00:02:49,820 BETHESDA, ABOUT 50 MINUTES NORTH 84 00:02:49,820 --> 00:02:51,689 IN FREDERICK WHICH HAS ABOUT 25% 85 00:02:51,689 --> 00:02:55,826 OF OUR PI'S, 3300 STAFF 86 00:02:55,826 --> 00:02:57,528 ALTOGETHER AND 40% TRAINEES, SO 87 00:02:57,528 --> 00:02:59,897 I WANT TO EMPHASIZE THAT WE DO 88 00:02:59,897 --> 00:03:00,998 THE FULL SPECTRUM. 89 00:03:00,998 --> 00:03:01,999 AND FOR THE RESEARCH THAT WE'LL 90 00:03:01,999 --> 00:03:03,434 BE TALKING ABOUT TODAY, I WANT 91 00:03:03,434 --> 00:03:06,737 TO EMPHASIZE THE 92 00:03:06,737 --> 00:03:07,705 DISCOVERY-DRIVEN BASIC RESEARCH 93 00:03:07,705 --> 00:03:09,373 COMPONENT BECAUSE TOM AND I AND 94 00:03:09,373 --> 00:03:10,641 ALL OF THE LEADERSHIP BELIEVES 95 00:03:10,641 --> 00:03:12,276 THAT THIS IS THE FOUNDATION OF 96 00:03:12,276 --> 00:03:17,548 THANKS WE DO HERE AND LEADS TO 97 00:03:17,548 --> 00:03:18,449 TRANSLATION, CLINICAL RESEARCH 98 00:03:18,449 --> 00:03:19,783 AND OF COURSE EVENTUALLY PATIENT 99 00:03:19,783 --> 00:03:20,818 CARE BUT THE FOUNDATION IS 100 00:03:20,818 --> 00:03:23,354 REALLY THE BASIC RESEARCH. 101 00:03:23,354 --> 00:03:24,755 SOY MENTION THIS HAD BEFORE BUT 102 00:03:24,755 --> 00:03:26,056 REALLY WHAT'S UNIQUE ABOUT OUR 103 00:03:26,056 --> 00:03:27,124 PROGRAM IS THE SCIENTIFIC 104 00:03:27,124 --> 00:03:30,761 FREEDOM THAT ALLOWS US TO ALMOST 105 00:03:30,761 --> 00:03:32,196 LITERALLY WAKE UP, GO INTO THE 106 00:03:32,196 --> 00:03:33,664 SHOWER, HAVE AN IDEA AND IF YOU 107 00:03:33,664 --> 00:03:36,133 CAN CONVINCE YOUR POOR GULLIBLE 108 00:03:36,133 --> 00:03:37,501 POST-DOC THAT YOU SHOULD DO THIS 109 00:03:37,501 --> 00:03:39,003 NEW PROJECT, YOU'RE ON YOUR WAY. 110 00:03:39,003 --> 00:03:40,671 AND WE WANT YOU TO TAKE RISKS. 111 00:03:40,671 --> 00:03:43,707 WE ACTUALLY CHALLENGE YOU TO 112 00:03:43,707 --> 00:03:45,910 TAKE RISKS AND WEKS DO THINGS 113 00:03:45,910 --> 00:03:48,212 HERE THAT TAKE -- AND WE CAN DO 114 00:03:48,212 --> 00:03:50,014 THINGS THAT TAKE A LONG TIME. 115 00:03:50,014 --> 00:03:54,418 FOR EXAMPLE, WAS RTDING TO BUILD 116 00:03:54,418 --> 00:03:56,287 MEL LANOMA MOUSE MODEL SAYS, I'M 117 00:03:56,287 --> 00:03:58,389 PRETTY SURE HAD THERE I COULD DO 118 00:03:58,389 --> 00:04:01,125 THE FIRST BUT WOULDN'T GET THE 119 00:04:01,125 --> 00:04:02,226 COMPETITIVE MODEL AFTER THREE OR 120 00:04:02,226 --> 00:04:02,760 FOUR YEARS. 121 00:04:02,760 --> 00:04:03,961 BUT YOU HAVE THE TIME AND IT 122 00:04:03,961 --> 00:04:05,196 WORKS OUT REALLY WELL AND THE 123 00:04:05,196 --> 00:04:07,831 HEART OF OUR PLACE HERE IN 124 00:04:07,831 --> 00:04:10,968 BETHESDA IS NIH CLINICAL CENTER, 125 00:04:10,968 --> 00:04:11,969 LARGEST CLINICAL CENTER IN THE 126 00:04:11,969 --> 00:04:14,171 WORLD, WE HAVE A FULLY DEDICATED 127 00:04:14,171 --> 00:04:17,208 STAFF, HOSPITAL, AND BASICALLY 128 00:04:17,208 --> 00:04:19,009 ALL THE PHYSICIAN SCIENTIST HERE 129 00:04:19,009 --> 00:04:20,077 HAVE ACTIVE RESEARCH 130 00:04:20,077 --> 00:04:21,378 LABORATORIES AS WELL AS THEIR 131 00:04:21,378 --> 00:04:23,247 CLINICAL TRIALS, AND WE TRY TO 132 00:04:23,247 --> 00:04:29,019 HAVE SEAMLESS TRANSLATION SAYS 133 00:04:29,019 --> 00:04:30,487 AND FIRST IN HUMAN TRIALS. 134 00:04:30,487 --> 00:04:32,556 THESE ARE SOME OF THE NUMBERS. 135 00:04:32,556 --> 00:04:34,525 LAST YEAR WE HAD ELEVEN HUNDRED 136 00:04:34,525 --> 00:04:37,027 ARTICLES AND PEER REVIEWED 137 00:04:37,027 --> 00:04:39,763 JOURNALS, CURRENTLY 346 OPEN 138 00:04:39,763 --> 00:04:40,564 TRIALS, CLINICAL TRIALS COMING 139 00:04:40,564 --> 00:04:42,967 ALL THE TIME, 27,000 PATIENTS IN 140 00:04:42,967 --> 00:04:45,869 VAROUS STUDIES, THOUSANDS OF 141 00:04:45,869 --> 00:04:47,538 COLLABORATIONS IN ALL THE STATES 142 00:04:47,538 --> 00:04:48,973 AND MANY COUNTRIES. 143 00:04:48,973 --> 00:04:50,507 OH, SINCE I BEGAN TO TALK RS WE 144 00:04:50,507 --> 00:04:53,577 HAVE TWO MORE P -- WE HAVE TWO 145 00:04:53,577 --> 00:04:56,880 MORE PI'S THAN WE HAD BEFORE, 146 00:04:56,880 --> 00:04:58,515 YOU SEE HOW ACTIVE OUR PROGRAM 147 00:04:58,515 --> 00:04:58,716 IS. 148 00:04:58,716 --> 00:04:59,817 BOIRKS I'VE GOT TO TALK TO 149 00:04:59,817 --> 00:05:00,784 SOMEBODY ABOUT THAT. 150 00:05:00,784 --> 00:05:03,520 18 NEW FACULTY RECRUITS JUST 151 00:05:03,520 --> 00:05:06,824 THIS YEAR AND THREE NEWLY 152 00:05:06,824 --> 00:05:08,993 TENURED INVESTIGATORS, STAFF 153 00:05:08,993 --> 00:05:11,295 SCIENCE IS ACTUAL BACKBONE OF 154 00:05:11,295 --> 00:05:15,466 OUR PROGRAM AND FRMENTS OUTSIDE 155 00:05:15,466 --> 00:05:16,867 AND REALLY THE HEARTBEAT OF WHAT 156 00:05:16,867 --> 00:05:17,968 GOES ON HERE AND OF COURSE WE 157 00:05:17,968 --> 00:05:21,372 HAVE HUNDREDS OF POSTDOCTORAL 158 00:05:21,372 --> 00:05:23,107 FELLOWS, POSTBACK STUDENTS 159 00:05:23,107 --> 00:05:24,541 REALLY INTO MENTORING AND 160 00:05:24,541 --> 00:05:27,144 PROMOTING CAREERS OF YOUNG 161 00:05:27,144 --> 00:05:28,579 INVESTIGATORS, LOTS OF PATENTS 162 00:05:28,579 --> 00:05:35,486 WHAT, WE CALL CRADA'S AND ACTIVE 163 00:05:35,486 --> 00:05:35,919 LICENSES. 164 00:05:35,919 --> 00:05:40,524 AS ANY AT THAT THIS ROGERS AND 165 00:05:40,524 --> 00:05:42,192 MIKE BETA, CLOSE TO THEM IN 166 00:05:42,192 --> 00:05:44,361 THEIR LAB, LOWER RIGHT-HAND 167 00:05:44,361 --> 00:05:48,198 CORNER, PIONEERING IMMUNOTHERAPY 168 00:05:48,198 --> 00:05:50,134 FROM STEVE ROSENBERG ON WE'VE 169 00:05:50,134 --> 00:05:57,141 HAD A LOT TO DO WITH -- YEARS 170 00:05:57,141 --> 00:05:59,743 AGO AND IT TURNED OUT IT WAS 171 00:05:59,743 --> 00:06:00,577 ABSOLUTELY ESSENTIAL IN THE 172 00:06:00,577 --> 00:06:01,211 CLINICAL TRIALS. 173 00:06:01,211 --> 00:06:02,880 AS I SAID THE PEOPLE IN HERE ARE 174 00:06:02,880 --> 00:06:04,882 GOING TO SET THE STAGE FOR THAT. 175 00:06:04,882 --> 00:06:06,583 TO JUST DRIVE THAT HOME, THESE 176 00:06:06,583 --> 00:06:09,353 ARE SOME OF THE EXISTING 177 00:06:09,353 --> 00:06:10,321 STRENGTHS, WE HAVE OF COURSE 178 00:06:10,321 --> 00:06:12,189 DIFFERENT KINDS OF CANCER 179 00:06:12,189 --> 00:06:13,757 MECHANISMS, WE HAVE A LOT OF 180 00:06:13,757 --> 00:06:16,126 WORK ON RARE CANCERS IN 181 00:06:16,126 --> 00:06:18,195 PARTICULAR, WE REALLY FOCUS ON 182 00:06:18,195 --> 00:06:19,096 THOSE. 183 00:06:19,096 --> 00:06:20,597 EPIGENETICS, SOME OF THE FIRST 184 00:06:20,597 --> 00:06:24,968 MICROBIOME WORK WAS DONE HERE, 185 00:06:24,968 --> 00:06:26,737 DRUG DISCOVERY ALL THE TIME, OF 186 00:06:26,737 --> 00:06:28,272 COURSE PRECISION MEDICINE, OF 187 00:06:28,272 --> 00:06:29,139 COURSE HIGHLIGHTING IN 188 00:06:29,139 --> 00:06:30,974 IMMUNOLOGY AND HOW IMPORTANT THE 189 00:06:30,974 --> 00:06:32,209 FUNDAMENTAL IMMUNOLOGY IS TO THE 190 00:06:32,209 --> 00:06:33,177 ENTIRE PROGRAM AND LEADS TO 191 00:06:33,177 --> 00:06:34,945 EVERYTHING ELSE THAT YOU SEE 192 00:06:34,945 --> 00:06:35,346 HERE. 193 00:06:35,346 --> 00:06:37,014 AND AS I SAID, WE'RE VERY PROUD 194 00:06:37,014 --> 00:06:38,816 OF THE IMMUNOLOGY MEETINGS AND 195 00:06:38,816 --> 00:06:41,952 SEMINARS AND SYMPOSIUMS WE'VE 196 00:06:41,952 --> 00:06:42,619 HAD OVER THE YEARS. 197 00:06:42,619 --> 00:06:45,222 SO THIS IS JUST FROM 2017, OUR 198 00:06:45,222 --> 00:06:47,491 RECENT FDA DRUG APPROVALS, YOU 199 00:06:47,491 --> 00:06:49,193 CAN SEE THERE'S QUITE A FEW OF 200 00:06:49,193 --> 00:06:51,028 THEM IN ALL KINDS DIFFERENT 201 00:06:51,028 --> 00:06:51,261 AREAS. 202 00:06:51,261 --> 00:06:53,097 I'M VERY PLEASED IN PARTICULAR 203 00:06:53,097 --> 00:06:57,568 ABOUT THE 2018 HAIRY CELL 204 00:06:57,568 --> 00:06:59,803 LEUKEMIA BECAUSE I WORKED FOR 205 00:06:59,803 --> 00:07:02,639 IRA PAST SOME TIME AGO AND HIS 206 00:07:02,639 --> 00:07:03,507 IMMUNOTOXINS REALLY WERE THE KEY 207 00:07:03,507 --> 00:07:04,575 TO THAT DRUG AND YOU CAN SEE 208 00:07:04,575 --> 00:07:07,778 THEY CONTINUE ALL THE WAY INTO 209 00:07:07,778 --> 00:07:07,978 2023. 210 00:07:07,978 --> 00:07:10,013 IN TERM OF GLOBAL REACH, WE HAVE 211 00:07:10,013 --> 00:07:11,849 THOUSANDS OF COLLABORATIONS 212 00:07:11,849 --> 00:07:12,049 THERE. 213 00:07:12,049 --> 00:07:16,253 WE WORK WITH MANY, MANY 214 00:07:16,253 --> 00:07:19,089 INSTITUTIONS AND MANY, MANY 215 00:07:19,089 --> 00:07:19,623 DIFFERENT TRAINING 216 00:07:19,623 --> 00:07:19,957 OPPORTUNITIES. 217 00:07:19,957 --> 00:07:21,225 WE TRY TO BE AS INFLUENTIAL AS 218 00:07:21,225 --> 00:07:22,393 WE CAN THROUGHOUT THE WORLD. 219 00:07:22,393 --> 00:07:23,994 JUST LET ME QUICKLY, I'M NOT GRG 220 00:07:23,994 --> 00:07:25,095 TO GO THROUGH THE SCHEDULE, BUT 221 00:07:25,095 --> 00:07:29,633 THE SCHEDULE IS REALLY QUITE 222 00:07:29,633 --> 00:07:29,933 AMAZING. 223 00:07:29,933 --> 00:07:32,836 HAD YOU HAVE LUMINARIES FROM 224 00:07:32,836 --> 00:07:34,438 INTRAMURALS AND EXTRAMURALS, 225 00:07:34,438 --> 00:07:35,572 SCIENTIFIC COUNCILMEMBERS, I 226 00:07:35,572 --> 00:07:37,074 THINK THE ORGANIZERS DID AN 227 00:07:37,074 --> 00:07:38,308 AMAZING JOB OF PUTTING THIS 228 00:07:38,308 --> 00:07:40,043 PROGRAM TOGETHERS. 229 00:07:40,043 --> 00:07:42,880 SO ON DAY ONE YOU'LL BE TALKING 230 00:07:42,880 --> 00:07:45,182 ABOUT THE THYMUS MOSTLY AND 231 00:07:45,182 --> 00:07:50,187 MOSTLY IMMUNITY FRRKS 232 00:07:50,187 --> 00:07:51,522 CROSS-TALK, DICIALT KINDS OF 233 00:07:51,522 --> 00:07:54,024 SIGNALLING, DAY TWO INNATE 234 00:07:54,024 --> 00:07:55,993 IMMUNITY AND MORE ON CELL 235 00:07:55,993 --> 00:07:58,061 SIGNALLING AND CELL FUNCTION AND 236 00:07:58,061 --> 00:07:59,062 SECOND PART FINALLY OF THE 237 00:07:59,062 --> 00:08:00,731 CANCER IMMUNOLOGY PROGRAM. 238 00:08:00,731 --> 00:08:02,566 IT LOOKS LIKE NAOMI IS GOING TO 239 00:08:02,566 --> 00:08:04,001 WRAP THINGS UP FOR US, WHICH 240 00:08:04,001 --> 00:08:09,907 TAKES ME TRITED -- TAKES ME 241 00:08:09,907 --> 00:08:11,542 RIGHT TO WHERE I WANT TO BE, IN 242 00:08:11,542 --> 00:08:13,076 THANKING EVERYBODY FOR PUTTING 243 00:08:13,076 --> 00:08:14,745 THIS WONDERFUL SYMPOSIUM 244 00:08:14,745 --> 00:08:15,012 TOGETHER. 245 00:08:15,012 --> 00:08:19,416 I WANT TO PARTICULARLY THAT I 246 00:08:19,416 --> 00:08:22,186 NAOMI AND HYUN AND OTHERS ALSO 247 00:08:22,186 --> 00:08:23,487 AND I WOULD LIKE TO SHOUT OUT 248 00:08:23,487 --> 00:08:25,155 THE PEOPLE WHO REALLY MAKE THIS 249 00:08:25,155 --> 00:08:28,192 STUFF HAPPEN, JULIA LAMB, 250 00:08:28,192 --> 00:08:29,493 CRYSTAL -- 251 00:08:29,493 --> 00:08:31,595 [CHEERING] 252 00:08:31,595 --> 00:08:35,666 [APPLAUSE] 253 00:08:35,666 --> 00:08:36,834 HAD BECAUSE OTHERWISE WE WOULD 254 00:08:36,834 --> 00:08:38,168 ALL BE SITTING HERE AND HAVING 255 00:08:38,168 --> 00:08:39,470 COFFEE AND THAT WOULD BE IT. 256 00:08:39,470 --> 00:08:39,703 OKAY. 257 00:08:39,703 --> 00:08:42,906 SO THAT'S REALLY ALL I HAD TO 258 00:08:42,906 --> 00:08:43,073 SAY. 259 00:08:43,073 --> 00:08:44,775 I'M GOING TO HAVE NAOMI COME UP 260 00:08:44,775 --> 00:08:46,210 AND REALLY KICK THINGS OFF RIGHT 261 00:08:46,210 --> 00:08:46,543 NOW. 262 00:08:46,543 --> 00:08:47,811 BUT THANK YOU AGAIN FOR THE 263 00:08:47,811 --> 00:08:48,812 INVITATION TO TALK, AND PLEASE 264 00:08:48,812 --> 00:08:52,082 HAVE A WONDERFUL SYMPOSIUM. 265 00:08:52,082 --> 00:08:58,655 [APPLAUSE] 266 00:08:58,655 --> 00:09:00,157 >> THANK YOU, GLENN, FOR 267 00:09:00,157 --> 00:09:03,060 STARTING THAT OFF ON A WONDERFUL 268 00:09:03,060 --> 00:09:03,494 FOOTING. 269 00:09:03,494 --> 00:09:05,596 IT'S REALLY A HUGE PLEASURE TO 270 00:09:05,596 --> 00:09:06,763 BE HERE. 271 00:09:06,763 --> 00:09:08,131 AND I THINK WHAT GLENN TALKED 272 00:09:08,131 --> 00:09:09,333 ABOUT IS THAT THIS IS A MEETING 273 00:09:09,333 --> 00:09:11,368 SPORCH SOARED BY THE CENTER FOR 274 00:09:11,368 --> 00:09:13,237 CANCER RESEARCH, BUT IT REALLY 275 00:09:13,237 --> 00:09:15,339 IS SUPPOSED TO BE -- SPONSORED 276 00:09:15,339 --> 00:09:17,307 BY THE CENTERED FOR CANCER 277 00:09:17,307 --> 00:09:18,308 RESEARCH, REALM IS SUPPOSED TO 278 00:09:18,308 --> 00:09:20,477 BE TO SHOWCASE IMMUNOLOGY. 279 00:09:20,477 --> 00:09:21,778 I SPENT MOST OF MY CAREER IN 280 00:09:21,778 --> 00:09:22,880 FRANCE AND HAD THE OPPORTUNITY 281 00:09:22,880 --> 00:09:24,648 TO COME BACK TO U.S. A COUPLE 282 00:09:24,648 --> 00:09:26,383 YEARS TO JOIN THE PEDIATRIC 283 00:09:26,383 --> 00:09:27,050 ONCOLOGY BRANCH. 284 00:09:27,050 --> 00:09:28,819 LAST WEEK WE HAD SOME 285 00:09:28,819 --> 00:09:31,989 INDIVIDUALS COME FROM CENTER FOR 286 00:09:31,989 --> 00:09:33,090 CHILDHOOD CANCER ALLIANCE COME 287 00:09:33,090 --> 00:09:37,094 TO VISIT THE LAB, AND, YOU KNOW, 288 00:09:37,094 --> 00:09:38,395 CLEARLY PEOPLE WHO ARE NOT 289 00:09:38,395 --> 00:09:39,830 NECESSARILY INTERESTED IN BASIC 290 00:09:39,830 --> 00:09:40,697 IMMUNOLOGY, THEY'RE THERE 291 00:09:40,697 --> 00:09:42,900 BECAUSE THEY WANT A CURE -- THEY 292 00:09:42,900 --> 00:09:44,535 WANT TO CURE CHILDREN WITH 293 00:09:44,535 --> 00:09:44,768 CANCER. 294 00:09:44,768 --> 00:09:46,069 BUT I THINK WHEN WE SHOW THEM 295 00:09:46,069 --> 00:09:46,937 THE PICTURES OF WHAT WE WERE 296 00:09:46,937 --> 00:09:49,339 DOING IN THE LAB AND WHAT WAS 297 00:09:49,339 --> 00:09:53,176 IMPORTANT, YOU KNOW, ONE OF THE 298 00:09:53,176 --> 00:09:59,783 THINGS THAT WAS ON THE SCREEN IS 299 00:09:59,783 --> 00:10:00,951 PEOPLE HADN'T SPENT A HUGE 300 00:10:00,951 --> 00:10:04,988 AMOUNT OF TIME REALIZING THAT 301 00:10:04,988 --> 00:10:06,823 GREEN FLUORESCENT PROTEIN FROM A 302 00:10:06,823 --> 00:10:07,691 JELLYFISH COULD BE USED TO 303 00:10:07,691 --> 00:10:09,026 FIGURE OUT WHERE CELLS WERE 304 00:10:09,026 --> 00:10:09,226 GOING. 305 00:10:09,226 --> 00:10:10,861 WE COULDN'T BE ABLE TO LOOK AT 306 00:10:10,861 --> 00:10:12,763 CANCERS IN MICE IF WE DIDN'T 307 00:10:12,763 --> 00:10:14,631 KNOW HOW LUCIFERASE WORKED AND 308 00:10:14,631 --> 00:10:16,433 TO BE ABLE TO STUDY FIREFLIES. 309 00:10:16,433 --> 00:10:17,367 WHILE SOME OF THESE THINGS YOU 310 00:10:17,367 --> 00:10:19,770 WOULD MAKE THE FRONT PAGE OF THE 311 00:10:19,770 --> 00:10:21,572 WASHINGTON POST AND SAY WHY ARE 312 00:10:21,572 --> 00:10:22,639 WE SPENDING MONEY ON THIS? 313 00:10:22,639 --> 00:10:24,841 THE POINT IS WE WHERE THOSE 314 00:10:24,841 --> 00:10:26,143 FRONTIERS COME, I THINK THAT'S 315 00:10:26,143 --> 00:10:26,810 WHY WE'RE HERE. 316 00:10:26,810 --> 00:10:28,912 IT'S A HUGE PLEASURE TOWF ALL 317 00:10:28,912 --> 00:10:30,614 HERE IN PERSON BECAUSE I THINK 318 00:10:30,614 --> 00:10:32,049 MAKING THOSE LEAPS AND 319 00:10:32,049 --> 00:10:33,150 BOUNDARIES CAN'T HAPPEN IF 320 00:10:33,150 --> 00:10:34,585 PEOPLE DON'T TALK TO EACH OTHER. 321 00:10:34,585 --> 00:10:36,420 THE PLEASURE OF BEING HERE ON 322 00:10:36,420 --> 00:10:37,421 SABBATICAL WOULD SAY THE MOST 323 00:10:37,421 --> 00:10:38,956 IMPORTANT THINGS YOU LEARN YOU 324 00:10:38,956 --> 00:10:40,390 LEARN FROM GOING AND HAVING 325 00:10:40,390 --> 00:10:41,625 COFFEE AND TALKING AND I THINK 326 00:10:41,625 --> 00:10:42,793 THAT WITH COVID AND WITH THE 327 00:10:42,793 --> 00:10:44,561 FACT THAT PEOPLE WEREN'T HERE IN 328 00:10:44,561 --> 00:10:45,896 PERSON, WE REALLY LOST SOME OF 329 00:10:45,896 --> 00:10:46,096 THAT. 330 00:10:46,096 --> 00:10:47,364 SO IT'S REALLY A HUGE PLEASURE 331 00:10:47,364 --> 00:10:50,033 TO SEE SO MANY SMILING FACES AND 332 00:10:50,033 --> 00:10:53,203 WE HAVE TRIED TO KEEP THE TALKS 333 00:10:53,203 --> 00:10:55,839 DOWN TO A MINIMUM BECAUSE WE 334 00:10:55,839 --> 00:10:57,040 WOULD LIKE TO MAKE SURE THAT WE 335 00:10:57,040 --> 00:10:57,774 HAVE QUESTIONS. 336 00:10:57,774 --> 00:10:58,875 SO PLEASE ASK QUESTIONS. 337 00:10:58,875 --> 00:11:00,744 WE WILL GIVE AWARDS FOR TRAINEES 338 00:11:00,744 --> 00:11:02,179 WHO ASK QUESTIONS BECAUSE THAT'S 339 00:11:02,179 --> 00:11:04,381 WHERE WE WANT THE QUESTIONS TO 340 00:11:04,381 --> 00:11:05,248 COME FROM. 341 00:11:05,248 --> 00:11:06,783 SO IF YOU SEE SOMEBODY WHO LOOKS 342 00:11:06,783 --> 00:11:08,118 OLDER AND MORE GRAY HAIRS IN 343 00:11:08,118 --> 00:11:09,286 FRONT OF YOU, YOU CAN GO IN 344 00:11:09,286 --> 00:11:09,886 FRONT OF THEM. 345 00:11:09,886 --> 00:11:11,488 IF THERE'S SOMEONE BEHIND YOU 346 00:11:11,488 --> 00:11:13,490 WHO HAS LESS GRAY HAIRS THAN 347 00:11:13,490 --> 00:11:15,025 YOU, THEY GET TO GO FIRST. 348 00:11:15,025 --> 00:11:16,727 THAT GOES FOR YOU TOO. 349 00:11:16,727 --> 00:11:16,994 SERIOUSLY. 350 00:11:16,994 --> 00:11:18,962 THE POINT IS REALLY TO HAVE IT 351 00:11:18,962 --> 00:11:20,697 BE AN INTERACTIVE ENVIRONMENT. 352 00:11:20,697 --> 00:11:21,798 WE REALLY WANT THIS TO BE A 353 00:11:21,798 --> 00:11:23,433 PLACE WHERE WE CAN DISCUSS, 354 00:11:23,433 --> 00:11:25,669 WHERE WE CAN INTERACT. 355 00:11:25,669 --> 00:11:27,371 SO IT'S A HUGE PLEASURE TO BE 356 00:11:27,371 --> 00:11:28,939 ABLE TO DO THIS, TO THINK ABOUT 357 00:11:28,939 --> 00:11:30,240 WHERE WE'RE GOING TO GO AND TO 358 00:11:30,240 --> 00:11:32,442 NOT HAVE ANY IDEA WHAT THAT 359 00:11:32,442 --> 00:11:32,676 MEANS. 360 00:11:32,676 --> 00:11:34,611 SO WE HAVE SPEAKERS WHO ARE 361 00:11:34,611 --> 00:11:37,648 HERE, WHO WILL BE SHOWCASING THE 362 00:11:37,648 --> 00:11:39,216 CCR, AND THAT IS TOGETHER WITH 363 00:11:39,216 --> 00:11:40,684 PEOPLE FROM ALL OVER THE WORLD, 364 00:11:40,684 --> 00:11:42,719 AND IT'S REALLY A HUGE PLEASURE 365 00:11:42,719 --> 00:11:44,121 TO HAVE PEOPLE WHO COME HERE 366 00:11:44,121 --> 00:11:45,422 FROM QUITE SOME NUMBER OF 367 00:11:45,422 --> 00:11:46,390 DIFFERENT CONTINENTS TO REALLY 368 00:11:46,390 --> 00:11:49,593 BE ABLE TO SEE WHERE THE 369 00:11:49,593 --> 00:11:50,794 EX-QUESTIONS ARE. 370 00:11:50,794 --> 00:11:52,429 AND IT'S -- WHERE THE NEXT 371 00:11:52,429 --> 00:11:53,397 QUESTIONS ARE. 372 00:11:53,397 --> 00:11:55,165 THE QUESTIONS ARE IMPORTANT 373 00:11:55,165 --> 00:11:56,466 IMRRKS THEN WE CAN DEVISE THE 374 00:11:56,466 --> 00:11:57,668 EXPERIMENTS TO ANSWER THEM ONCE 375 00:11:57,668 --> 00:11:58,769 WN WHAT THE QUESTIONS ARE. 376 00:11:58,769 --> 00:12:00,404 IT WOULD BE REALLY PATE TI NOT 377 00:12:00,404 --> 00:12:01,605 TO BE DISCUSSING THOSE 378 00:12:01,605 --> 00:12:01,872 QUESTIONS. 379 00:12:01,872 --> 00:12:02,839 PLEASE DO ASK THAT. 380 00:12:02,839 --> 00:12:03,907 SO I'M STANDING HERE BECAUSE 381 00:12:03,907 --> 00:12:05,542 LOTS OF PEOPLE CAME BEFORE ME TO 382 00:12:05,542 --> 00:12:07,177 PUT THIS IN PLACE. 383 00:12:07,177 --> 00:12:09,379 THIS FRONTIERS IN BASIC 384 00:12:09,379 --> 00:12:09,846 IMMUNOLOGY PROGRAM. 385 00:12:09,846 --> 00:12:12,549 THIS IS P THE SEVENTH ONE. 386 00:12:12,549 --> 00:12:14,184 I WASN'T EVEN BORN FOR THE FIRST 387 00:12:14,184 --> 00:12:14,484 ONE. 388 00:12:14,484 --> 00:12:16,386 I'M KIDDING. 389 00:12:16,386 --> 00:12:18,555 BUT IT WAS STARTED BY AL SINGER 390 00:12:18,555 --> 00:12:20,157 AND IT WAS STARTED BY ALTOGETHER 391 00:12:20,157 --> 00:12:29,299 WITH LARRY SAMULSON, JOHN ASHWA 392 00:12:29,299 --> 00:12:31,501 AND RAINY, A SHOUT OUT TO ALL OF 393 00:12:31,501 --> 00:12:33,036 YOU TO COME BACK AND ORGANIZE 394 00:12:33,036 --> 00:12:33,236 THIS. 395 00:12:33,236 --> 00:12:34,971 SO WITHOUT FURTHER ADO, WELCOME, 396 00:12:34,971 --> 00:12:35,972 AND THANK YOU AGAIN. 397 00:12:35,972 --> 00:12:40,444 PLEASE ASK QUESTIONS. 398 00:12:40,444 --> 00:12:43,880 [APPLAUSE] 399 00:12:43,880 --> 00:12:48,351 >> GOOD MORNING TION EVERYONE, 400 00:12:48,351 --> 00:12:50,754 WELCOME AGAIN. 401 00:12:50,754 --> 00:12:54,491 I'M STANLEY ADORO TOGETHER WITH 402 00:12:54,491 --> 00:12:57,527 CHRISTIAN MAYER WE'LL BE 403 00:12:57,527 --> 00:12:59,863 STARTING THE MORNING QUESTION. 404 00:12:59,863 --> 00:13:01,832 I WOULD LIKE TO WELCOME AL 405 00:13:01,832 --> 00:13:03,800 SINGER TO GIVE THE FIRST TALK 406 00:13:03,800 --> 00:13:05,335 TIED TO TRANSCRIPTIONAL 407 00:13:05,335 --> 00:13:08,138 DETERMINATION OF T LINEAGE FATE. 408 00:13:08,138 --> 00:13:09,306 WELCOME, AL. 409 00:13:09,306 --> 00:13:19,549 >> THANK YOU. 410 00:13:20,250 --> 00:13:20,717 SOPHIA. 411 00:13:20,717 --> 00:13:21,818 I'VE ACTUALLY CHANGED THE TOPIC 412 00:13:21,818 --> 00:13:23,987 OF MY TALK THIS MORNING. 413 00:13:23,987 --> 00:13:30,694 AND I'LL DESCRIBE IT TO YOU IN A 414 00:13:30,694 --> 00:13:40,837 MOMENT. 415 00:13:43,540 --> 00:13:43,707 THANKS. 416 00:13:43,707 --> 00:13:45,375 THANK YOU. 417 00:13:45,375 --> 00:13:46,810 WELL, LISTEN, SO GOOD MORNING 418 00:13:46,810 --> 00:13:47,711 EVERYONE AND WELCOME OF THE 419 00:13:47,711 --> 00:13:48,645 WE'VE BEEN LOOKING FORWARD TO 420 00:13:48,645 --> 00:13:49,780 HAVING YOU ALL HERE. 421 00:13:49,780 --> 00:13:57,354 SO IT'S GREAT TO SEE YOU. 422 00:13:57,354 --> 00:14:00,624 AS MECHANISM OF CD8 T CELL 423 00:14:00,624 --> 00:14:02,826 TOLERANCE WHICH WE CAME UP WITH 424 00:14:02,826 --> 00:14:03,794 REALLY BASED ON OUR 425 00:14:03,794 --> 00:14:05,862 UNDERSTANDING OF HOW T LYNN 426 00:14:05,862 --> 00:14:08,331 LINEAGE FATE IS DMERND THYMUS. 427 00:14:08,331 --> 00:14:09,266 I WAS ORIGINALLY GOING TO TALK 428 00:14:09,266 --> 00:14:11,802 TO YOU ABOUT HOW IT IS 429 00:14:11,802 --> 00:14:12,769 DETERMINED IN THE THYMUS I 430 00:14:12,769 --> 00:14:14,104 THOUGHT I WOULD TELL YOU ABOUT 431 00:14:14,104 --> 00:14:15,105 THIS NEXT GENERATION OF STUDY 432 00:14:15,105 --> 00:14:16,373 ABOUT WHAT WE'VE LEARNED AS A 433 00:14:16,373 --> 00:14:17,674 CONSEQUENCE OF IT. 434 00:14:17,674 --> 00:14:20,310 SO AGAIN, I'M AL SINGER FROM THE 435 00:14:20,310 --> 00:14:21,444 EXPERIMENTAL IMMUNOLOGY BRANCH 436 00:14:21,444 --> 00:14:23,079 HERE IN THE INTRAMURAL PROGRAM 437 00:14:23,079 --> 00:14:28,752 OF THE CANCER INSTITUTE AND -- 438 00:14:28,752 --> 00:14:34,157 WAS DONE BY SHARIFF BODDER WHO 439 00:14:34,157 --> 00:14:35,559 PUSHED THIS FROM THE BEGINNING 440 00:14:35,559 --> 00:14:37,527 TO THE END AND YOU'LL (AWAY FROM 441 00:14:37,527 --> 00:14:41,565 MIC) ABOUT DLSZ THESE INSIGHTS. 442 00:14:41,565 --> 00:14:42,165 SHERIF BADR. 443 00:14:42,165 --> 00:14:43,533 YOU'LL LEARN ABOUT A NEW 444 00:14:43,533 --> 00:14:44,534 MECHANISM OF T CELL TOLERANCE 445 00:14:44,534 --> 00:14:47,904 WHICH WE THINK IS THE ACTIVE 446 00:14:47,904 --> 00:14:50,006 MECHANISM OF T CELL TOLERANCE 447 00:14:50,006 --> 00:14:51,341 NAMED CLONAL EVICTION. 448 00:14:51,341 --> 00:14:55,645 AS YOU KNOW IN THE THYMUS, 449 00:14:55,645 --> 00:14:59,616 IMMATURE THYMUS SITES DEVELOP 450 00:14:59,616 --> 00:15:01,384 FROM STEM CELL PRECURSORS AND 451 00:15:01,384 --> 00:15:03,019 THESE GUYS THROUGH AN ORDERED 452 00:15:03,019 --> 00:15:04,654 SERIES OF DEVELOPMENTAL EVENTS 453 00:15:04,654 --> 00:15:05,856 IN THE THYMUS. 454 00:15:05,856 --> 00:15:08,491 THOSE THAT BECOME ALPHA-BETA T 455 00:15:08,491 --> 00:15:10,961 CELLS EXPRESS THE ALPHA-BETA T 456 00:15:10,961 --> 00:15:12,796 CELL RECEPTOR AND THEY FIRST 457 00:15:12,796 --> 00:15:14,865 EXPRESS THE ALPHA-BETA T CELL 458 00:15:14,865 --> 00:15:18,134 RECEPTOR AS AN IMMATURE DOUBLE 459 00:15:18,134 --> 00:15:19,236 POSITIVE THYMUS SITE AND IT'S 460 00:15:19,236 --> 00:15:20,971 DOUBLE POSITIVE BECAUSE IT HAS 461 00:15:20,971 --> 00:15:24,541 BOTH CD4 AND CD8 CODE RECEPTORS. 462 00:15:24,541 --> 00:15:26,877 DEVELOPMENT STOPS AT THIS POINT 463 00:15:26,877 --> 00:15:28,945 UNLESS THE DOUBLE POSITIVE 464 00:15:28,945 --> 00:15:31,181 THYMUS SITES FIND A LIGAND FOR 465 00:15:31,181 --> 00:15:33,083 THE T CELL RECEPTOR THAT SIGNALS 466 00:15:33,083 --> 00:15:34,284 THEM TO GO FURTHER. 467 00:15:34,284 --> 00:15:36,152 SO TO GET SIGNALED, THE CELLS 468 00:15:36,152 --> 00:15:39,122 NEED TO GET, NEED TO SOMEHOW 469 00:15:39,122 --> 00:15:42,192 ENCOUNTER LCK AND THE LCK IS 470 00:15:42,192 --> 00:15:45,028 SEQUESTERED BY THE CD4 AND CD8 471 00:15:45,028 --> 00:15:45,996 CO-VEEP TORS. 472 00:15:45,996 --> 00:15:47,964 CD4 ENGAGES CLASS TWO AS YOU ALL 473 00:15:47,964 --> 00:15:49,165 KNOW, SAILT ENGAGES CLASS ONE 474 00:15:49,165 --> 00:15:51,635 THAT, BRINGS IN THE LCK 475 00:15:51,635 --> 00:15:52,869 ASSOCIATED WITH THE TAIL, YOU 476 00:15:52,869 --> 00:15:53,603 GET A SIGNAL. 477 00:15:53,603 --> 00:15:55,405 SO THE FIRST STEP IN SELECTION 478 00:15:55,405 --> 00:15:57,874 OF THE THYMUS IS THIS TCR 479 00:15:57,874 --> 00:15:58,975 SIGNALLING STEP. 480 00:15:58,975 --> 00:16:01,811 AS A CONSEQUENCE OF THAT T -- OF 481 00:16:01,811 --> 00:16:03,413 THOSE TCR SIGNALS AND AS I'LL 482 00:16:03,413 --> 00:16:05,115 TELL YOU IN A MOMENT, DEPENDING 483 00:16:05,115 --> 00:16:07,284 ON THE QUALITY OF THOSE TCR 484 00:16:07,284 --> 00:16:12,522 SIGNALS, THE DOUBLE POSITIVE 485 00:16:12,522 --> 00:16:15,125 THYMUS CYTES ARE INDUCED TO 486 00:16:15,125 --> 00:16:16,893 EXPRESS ONE OR THE OTHER OF THE 487 00:16:16,893 --> 00:16:18,328 LINEAGE FACTORS THAT DETERMINE 488 00:16:18,328 --> 00:16:20,263 THEIR T CELL LINEAGE FATE, SO 489 00:16:20,263 --> 00:16:23,266 THERE'S A TCH CRZ FACTOR YOU'LL 490 00:16:23,266 --> 00:16:25,101 HEAR ABOUT A LOT TODAY THAT 491 00:16:25,101 --> 00:16:28,305 HELPS HELPER LINEAGE FATE ON THE 492 00:16:28,305 --> 00:16:34,511 THERE CELLS OR RUNX3D, D WHICH 493 00:16:34,511 --> 00:16:37,380 INDUCES THE CD8 CYTOTOXIC TELZ. 494 00:16:37,380 --> 00:16:39,716 WE'VE BEEN STUDYING HOW THOSE 495 00:16:39,716 --> 00:16:41,318 EVENTS HAPPEN FOR MANY YEARS NOW 496 00:16:41,318 --> 00:16:44,387 AND A LONG TIME AGO, ACTUALLY IN 497 00:16:44,387 --> 00:16:46,256 2000, WE THERE PUBLIC PUBLISHED 498 00:16:46,256 --> 00:16:48,558 A MODEL CALMED THE KINETIC 499 00:16:48,558 --> 00:16:51,361 SIGNALLING MODEL OF T LIJAL FATE 500 00:16:51,361 --> 00:16:51,695 DETERMINATION. 501 00:16:51,695 --> 00:16:53,330 THE FUNNY THING ABOUT THE MODEL 502 00:16:53,330 --> 00:16:58,501 WAS THAT IT WAS BASED NOT ON THE 503 00:16:58,501 --> 00:17:01,471 CO-RECEPTOR PROTEINS BUT ON THE 504 00:17:01,471 --> 00:17:02,505 CO-REACCEPTOR GENES AND IT CAME 505 00:17:02,505 --> 00:17:05,542 FROM THE OBSERVATION WHICH WAS A 506 00:17:05,542 --> 00:17:06,977 SURPRISE TO US AND WITH THOUGHT 507 00:17:06,977 --> 00:17:08,111 FROM THE MOMENT WE SAW IT THAT 508 00:17:08,111 --> 00:17:09,779 THERE WAS SOMETHING IMPORTANT 509 00:17:09,779 --> 00:17:11,681 HERE, BUT IT'S TAKEN A WHILE TO 510 00:17:11,681 --> 00:17:13,216 FIGURE OUT WHAT THAT WAS, THE 511 00:17:13,216 --> 00:17:16,252 TCR SIGNALLING, SO THIS 512 00:17:16,252 --> 00:17:18,254 SIGNALLING EVENT IN THYMIC 513 00:17:18,254 --> 00:17:20,223 SELECTION HAS A CONSEQUENCE ON 514 00:17:20,223 --> 00:17:21,624 CD4 AND CD8 GENE EXPRESSION, 515 00:17:21,624 --> 00:17:23,960 THAT IS, THAT TCR SIGNALLING 516 00:17:23,960 --> 00:17:25,261 EVENTS SELECTIVELY TERM NAILTS 517 00:17:25,261 --> 00:17:30,500 THE TRANSCRIPTION OF CD8 GENE. 518 00:17:30,500 --> 00:17:32,902 SO YOU GET A TCR SIGNAL, THE CD8 519 00:17:32,902 --> 00:17:34,137 GENE IS SHUT OFF. 520 00:17:34,137 --> 00:17:36,439 SO TCR SIGNALLING THAT'S 521 00:17:36,439 --> 00:17:40,477 DEPENDENT ON THE CD8 ENCODED 522 00:17:40,477 --> 00:17:41,678 CO-RECEPTOR PROTEIN, THOSE 523 00:17:41,678 --> 00:17:43,646 SIGNALS GRADUALLY WEAKEN, 524 00:17:43,646 --> 00:17:45,181 STEADILY WEAKEN EARN UNTIL THE 525 00:17:45,181 --> 00:17:47,584 SIGNAL IS FINALLY DISRUPTED 526 00:17:47,584 --> 00:17:48,985 DURING SELECTION. 527 00:17:48,985 --> 00:17:53,356 THAT DISRUPTION OF TCR ARE 528 00:17:53,356 --> 00:17:56,226 SIGNALLING FLRKS CYTOKINE, WE'VE 529 00:17:56,226 --> 00:17:57,627 IDENTIFIED SIX OR SEVEN OF THEM, 530 00:17:57,627 --> 00:18:00,463 THAT WILL INDUCE SIGNAL OF THESE 531 00:18:00,463 --> 00:18:06,669 CELLS TO EXPRESS LLZ -- ON THE 532 00:18:06,669 --> 00:18:08,605 OTHER HAND IF THE TCR SIGNAL IS 533 00:18:08,605 --> 00:18:12,208 DEPENDENT NOT ON THE CD8 ENCODED 534 00:18:12,208 --> 00:18:13,877 CO-RECEPTOR BUT ON THE CD4, 535 00:18:13,877 --> 00:18:16,279 NOTHING HAPPENS TO IT, IN FACT, 536 00:18:16,279 --> 00:18:17,814 THAT SIGNAL INCREASES AND 537 00:18:17,814 --> 00:18:18,815 PERSISTS THROUGH OUR POSITIVE 538 00:18:18,815 --> 00:18:21,017 SELECTION AND THAT PERSISTS IN 539 00:18:21,017 --> 00:18:27,457 TCR SIGNAL AND THIS IS THPOK TO 540 00:18:27,457 --> 00:18:29,192 DEPENDENT THE HELPER CELL. 541 00:18:29,192 --> 00:18:31,294 THIS IS HOW LYNN YAM FATE IS 542 00:18:31,294 --> 00:18:32,062 DECIDED, DEPENDENT ON WHETHER 543 00:18:32,062 --> 00:18:34,264 THE SIGNAL PERSIST OR CREASES 544 00:18:34,264 --> 00:18:39,602 AND THE CD8 DEPENDENT TCR SIGNAL 545 00:18:39,602 --> 00:18:41,704 CEASES. 546 00:18:41,704 --> 00:18:44,774 SO IT WILL CD8 RECEPTOR SIGNALS 547 00:18:44,774 --> 00:18:46,176 ARE DISRUPTED AND CEASE DURING 548 00:18:46,176 --> 00:18:49,112 THE PROCESS OF POSITIVE 549 00:18:49,112 --> 00:18:49,379 SELECTION. 550 00:18:49,379 --> 00:18:50,880 SO WE'VE BEEN INTERESTED IN 551 00:18:50,880 --> 00:18:52,215 UNDERSTANDING HOW THIS CAN I 552 00:18:52,215 --> 00:18:54,717 KNELT I CAN SIGNALLING MODEL 553 00:18:54,717 --> 00:18:56,719 IMABLGHTS ON T CELL TOLERANCE -- 554 00:18:56,719 --> 00:18:57,987 IMPACTS ON T CELL TOLERANCE. 555 00:18:57,987 --> 00:19:00,090 THIS IS WHAT I'LL TELL YOU ABOUT 556 00:19:00,090 --> 00:19:01,291 TODAY, BECAUSE IF YOU THINK 557 00:19:01,291 --> 00:19:03,259 ABOUT WHAT THIS KINETIC 558 00:19:03,259 --> 00:19:05,628 CERTAINTY MODEL PREDICTS FOR CD8 559 00:19:05,628 --> 00:19:07,330 T CELL TOLERANCE THERE'S QUITE 560 00:19:07,330 --> 00:19:08,431 AN INTERESTING PREDICTION H THAT 561 00:19:08,431 --> 00:19:11,601 IS THAT THE CD8 DEPENDENT 562 00:19:11,601 --> 00:19:12,702 SIGNALS IN THE THYMUS BECAUSE 563 00:19:12,702 --> 00:19:14,437 THEY PROGRESSIVELY WEAKEN UNTIL 564 00:19:14,437 --> 00:19:17,273 THEY'RE DISRUPTED, DEVELOPING 565 00:19:17,273 --> 00:19:20,009 CD8 T CELLS MAY NOT BE SIGNALED 566 00:19:20,009 --> 00:19:21,778 STRONGLY ENOUGH AND MAY NOT BE 567 00:19:21,778 --> 00:19:24,180 SIGNALED LONG ENOUGH TO BE ABLE 568 00:19:24,180 --> 00:19:27,684 TO INDUCE CLONAL DELETION DURING 569 00:19:27,684 --> 00:19:28,751 THYMIC SELECTION, TONT SAY 570 00:19:28,751 --> 00:19:30,720 THERE'S NO CLONAL DELETION, BUT 571 00:19:30,720 --> 00:19:32,155 THERE CAN'T BE ENOUGH CLONAL 572 00:19:32,155 --> 00:19:33,556 DELETION TO ACCOUNT FOR T CELL 573 00:19:33,556 --> 00:19:35,425 TOLERANCE SO WE WONDERED, WHAT'S 574 00:19:35,425 --> 00:19:37,627 THE BASIS FOR CD8 T CELL 575 00:19:37,627 --> 00:19:37,894 TOLERANCE? 576 00:19:37,894 --> 00:19:39,262 IS IT REALLY CLONAL DELETION THE 577 00:19:39,262 --> 00:19:41,231 WAY WE ALWAYS THOUGHT ABOUT 578 00:19:41,231 --> 00:19:41,498 ABOUT IT? 579 00:19:41,498 --> 00:19:43,733 I MEAN, EVERYBODY THINKS THAT 580 00:19:43,733 --> 00:19:48,705 CLONAL -- AND CAN TELL YOU THAT 581 00:19:48,705 --> 00:19:51,207 NO, IT'S ACTUALLY NOT TRUE FOR 582 00:19:51,207 --> 00:19:52,108 CD8 T CELLS. 583 00:19:52,108 --> 00:19:56,346 THE STUDY I'LL TELL YOU ABOUT 584 00:19:56,346 --> 00:19:57,814 THAT WAS PURSUED MADE USE OF 585 00:19:57,814 --> 00:19:59,649 THIS MOUSE, TRANSGENIC MOUSE 586 00:19:59,649 --> 00:20:02,919 THAT WAS GENERATED BY TROY 587 00:20:02,919 --> 00:20:07,624 BALDWIN AND CHRIS CHRIS HEDCRIS, 588 00:20:07,624 --> 00:20:13,696 MADE USE OF THE HY TRANSGENIC 589 00:20:13,696 --> 00:20:16,299 GENE, THIS IS A CLASSIC T CELL 590 00:20:16,299 --> 00:20:18,568 RECEPTOR SPECIFICITY. 591 00:20:18,568 --> 00:20:22,472 WELL, WHAT CHRIS HOGQUIST AND 592 00:20:22,472 --> 00:20:24,040 BALDWIN MODIFIED IN TERMS OF 593 00:20:24,040 --> 00:20:26,409 THIS TRANSGENIC MOUSE WAS THEY 594 00:20:26,409 --> 00:20:28,278 PREVENTED EXPRESSION OF THE TCR 595 00:20:28,278 --> 00:20:30,079 TRANSGENE SO THAT IT WASN'T 596 00:20:30,079 --> 00:20:31,714 EXPRESSED UNTIL THE DOUBLE 597 00:20:31,714 --> 00:20:32,482 POSITIVE STAGE. 598 00:20:32,482 --> 00:20:33,816 SO THIS REFLECTED EXACTLY WHAT 599 00:20:33,816 --> 00:20:35,985 HAPPENED IN THE NORMAL MOUSE. 600 00:20:35,985 --> 00:20:37,954 T CELLS DEVELOPED WHEN YOU HAVE 601 00:20:37,954 --> 00:20:42,425 THE DOUBLE POSITIVE IT THYMUS 602 00:20:42,425 --> 00:20:44,861 CYTE STATES AND FINALLY 603 00:20:44,861 --> 00:20:47,797 EXPRESSED T CELL RECEPTOR. 604 00:20:47,797 --> 00:20:49,332 SPECIFIC HY T CELLS AND YOU CAN 605 00:20:49,332 --> 00:20:51,834 SEE IN THE FEMALE MOUSE THESE 606 00:20:51,834 --> 00:20:54,938 CELLS UNDERGO POSITIVE SELECTION 607 00:20:54,938 --> 00:20:56,873 AND GRURND CD8 POSITIVE. 608 00:20:56,873 --> 00:20:58,641 IN THE MALE MOUSE THERE ARE NO 609 00:20:58,641 --> 00:20:59,742 CD8 POSITIVE CELLS AND THE 610 00:20:59,742 --> 00:21:01,477 PRESUMPTION AS TO WHY THERE WAS 611 00:21:01,477 --> 00:21:03,379 NEVER ANY CD8 SIGNAL POSITIVE 612 00:21:03,379 --> 00:21:06,683 CELLS SINCE THE VERY FIRST 613 00:21:06,683 --> 00:21:10,587 TRANSGENIC MOUSE, VERY FIRST 614 00:21:10,587 --> 00:21:15,425 TRANSGENIC MOWD MADE BY BOB 615 00:21:15,425 --> 00:21:18,828 CLOMER THOUGHT DUE TO DELETION. 616 00:21:18,828 --> 00:21:20,897 ARE THESE CELLS MISSING BECAUSE 617 00:21:20,897 --> 00:21:22,365 THEY AND THE PRECURSORS HAVE 618 00:21:22,365 --> 00:21:23,633 BEEN CLONALLY DELETED? 619 00:21:23,633 --> 00:21:24,734 BUT IN FACT IF YOU LOOK IN THE 620 00:21:24,734 --> 00:21:27,570 PERIPHERY OF THESE MALE MICE, 621 00:21:27,570 --> 00:21:31,975 THE PERIPHERY HAVE LOTS OF STAT. 622 00:21:31,975 --> 00:21:35,278 AND IN FACT, THE MALE MOUSE HAS 623 00:21:35,278 --> 00:21:36,746 MANY MORE CD8 SIGNAL POSITIVE 624 00:21:36,746 --> 00:21:39,082 CELLS IN THE PERIPHERY THAN THE 625 00:21:39,082 --> 00:21:42,585 POSITIVELY SELECTING MALE -- 626 00:21:42,585 --> 00:21:43,219 FEMALE MOUSE. 627 00:21:43,219 --> 00:21:45,989 WE THOUGHT THIS CANNOT BE CLONAL 628 00:21:45,989 --> 00:21:46,556 DELETION. 629 00:21:46,556 --> 00:21:50,860 SOS CLONAL DELETION IN FACT 630 00:21:50,860 --> 00:21:51,427 EXPLAINS HOW IT IS THAT CD8 631 00:21:51,427 --> 00:21:53,763 CELLS ARE MISSING F YET THOSE 632 00:21:53,763 --> 00:21:54,864 SAME CELLS APPEAR IN THE PRIFER 633 00:21:54,864 --> 00:21:55,031 RI. 634 00:21:55,031 --> 00:21:56,866 HOW DOES THIS WORK? 635 00:21:56,866 --> 00:21:58,601 SO WE LOOKED IN THE THYMUS AT 636 00:21:58,601 --> 00:22:01,304 THESE HY CD4 MALE CELLS AND WE 637 00:22:01,304 --> 00:22:02,705 FOUND OF COURSE AS MINK OF YOU 638 00:22:02,705 --> 00:22:03,840 KNOW THAT IN THE THYMUS WHAT YOU 639 00:22:03,840 --> 00:22:11,748 SEE IS A POPULATION OF CD4 CD8 640 00:22:11,748 --> 00:22:13,049 DOUBLE DULL CELLS WHICH HAVE 641 00:22:13,049 --> 00:22:14,817 BEEN STRONGLY SIGNALLED BECAUSE 642 00:22:14,817 --> 00:22:16,119 THEY'VE UNDERGONE NEGATIVE 643 00:22:16,119 --> 00:22:17,654 SELECTION AFTER ALL THEY'RE HY 644 00:22:17,654 --> 00:22:19,722 SPECIFIC AND ENCOUNTER THE HY 645 00:22:19,722 --> 00:22:22,458 MALE ANTIGEN OF THE THYMUS SO 646 00:22:22,458 --> 00:22:25,428 THAT STRONG SIGNALLING 647 00:22:25,428 --> 00:22:28,731 UPREGULATES EXPRESSION OF THE 648 00:22:28,731 --> 00:22:31,000 PD1 WHICH IS A FACT OF TCR 649 00:22:31,000 --> 00:22:32,935 SIGNALLING AND ATTENUATOR. 650 00:22:32,935 --> 00:22:34,837 SO IT'S AN ATTEMPT BY THE CELL 651 00:22:34,837 --> 00:22:37,807 TO RESPOND IT THAT STRONG TCR 652 00:22:37,807 --> 00:22:42,111 SIGNAL AND UPREGULATES PD1 AND 653 00:22:42,111 --> 00:22:43,546 THESE CELLS ARE ACTUALLY 654 00:22:43,546 --> 00:22:47,116 IMMATURE SO THEY DON'T EXPRESS 655 00:22:47,116 --> 00:22:48,518 CD24, THEY DON'T DOWN REGULATE, 656 00:22:48,518 --> 00:22:50,720 THEY EXPRESS HIGH LEVELS OF CD 657 00:22:50,720 --> 00:22:52,789 24-RBGS THEY DON'T DOWN REGULATE 658 00:22:52,789 --> 00:22:55,625 CD4 AND THEY DON'T EXPRESS QA2, 659 00:22:55,625 --> 00:22:58,928 THESE ARE IMMATURE PD1 POSITIVE 660 00:22:58,928 --> 00:23:00,863 DOUBLE DULL CELLS SO WE WONDERED 661 00:23:00,863 --> 00:23:02,031 WHAT HAPPENS TO THESE CELLS? 662 00:23:02,031 --> 00:23:03,633 IS IT POSSIBLE THESE GUYS 663 00:23:03,633 --> 00:23:05,635 ACTUALLY GET OUT OF THE THYMUS? 664 00:23:05,635 --> 00:23:07,470 NORMALLY THE ONLY CELLS IN THE 665 00:23:07,470 --> 00:23:09,005 THYMUS THAT LEAVE THE THYMUS ARE 666 00:23:09,005 --> 00:23:09,706 MATURE CELLS. 667 00:23:09,706 --> 00:23:10,807 THESE RNLT MATURE CELLS. 668 00:23:10,807 --> 00:23:12,542 AS YOU SAW, THESE ARE IMMATURE 669 00:23:12,542 --> 00:23:14,077 CELLS BUT WE WONDERED, MAYBE 670 00:23:14,077 --> 00:23:19,816 THEY DO LEAVE THE THYMUS ANYWAY. 671 00:23:19,816 --> 00:23:24,620 TO TEST THAT WE INJECTED THESE 672 00:23:24,620 --> 00:23:27,256 MICE WITH A CONJUGATED 673 00:23:27,256 --> 00:23:28,057 MONOCLONAL ANTIBODY AND LOOKED 674 00:23:28,057 --> 00:23:28,891 FIVE MINUTES LATER. 675 00:23:28,891 --> 00:23:30,727 THE IDEA BEING THAT IN FIVE 676 00:23:30,727 --> 00:23:32,195 MINUTES THE ONLY CELLS THESE 677 00:23:32,195 --> 00:23:33,196 ANTIBODIES WOULD ENCOUNTER WOULD 678 00:23:33,196 --> 00:23:34,263 BE THOSE THAT WERE IN CONTACT 679 00:23:34,263 --> 00:23:38,835 WITH THE BLOODSTREAM BECAUSE THE 680 00:23:38,835 --> 00:23:39,802 ANTIBODY DIDN'T HAVE ENOUGH TIME 681 00:23:39,802 --> 00:23:41,070 TO GET INTO TISSUES. 682 00:23:41,070 --> 00:23:47,477 IN FACT, IF YOU LOOK AND SEE, 683 00:23:47,477 --> 00:23:49,245 YOU SEE THAT THE CELLS THAT GOT 684 00:23:49,245 --> 00:23:51,214 LABELED BY THE ANTIBODIES ARE 685 00:23:51,214 --> 00:23:52,615 THESE DOUBLE DULL CELLS EVEN 686 00:23:52,615 --> 00:23:53,616 THOUGH MOST OF THE CELLS IN THE 687 00:23:53,616 --> 00:23:56,386 THYMUS ARE ACTUALLY PRESELECTION 688 00:23:56,386 --> 00:23:59,455 DOUBLE POSITIVE THYMOSIGHTS SO 689 00:23:59,455 --> 00:24:02,392 THEY DON'T GET LABELED. 690 00:24:02,392 --> 00:24:03,826 SUGGEST THESE TELZ ARE IN 691 00:24:03,826 --> 00:24:04,894 CONTACT WITH THE BLOODSTREAM 692 00:24:04,894 --> 00:24:06,129 BECAUSE THEY'RE IN PROCESS OF 693 00:24:06,129 --> 00:24:08,965 LEAFING THE THYMUS IT. 694 00:24:08,965 --> 00:24:09,999 IF THEY LEAVE THE THYMUS WE 695 00:24:09,999 --> 00:24:11,167 SHOULD BE ABLE TO FIND THEM IN 696 00:24:11,167 --> 00:24:12,902 THE PERIPHERY, YOU SHOULD 697 00:24:12,902 --> 00:24:15,204 REFLECT BOTH IN THE LYMPH NODE 698 00:24:15,204 --> 00:24:17,740 AND SPLEEN AND YOU SEE IN THE 699 00:24:17,740 --> 00:24:18,941 MALE YOU IN FACT FIND THESE 700 00:24:18,941 --> 00:24:22,078 CELLS, YOU FIND THESE DOUBLE 701 00:24:22,078 --> 00:24:24,747 DULL CELLS PRESENT IN THE MALE 702 00:24:24,747 --> 00:24:25,014 PERIPHERY. 703 00:24:25,014 --> 00:24:26,416 AND THESE ARE ACCIDENT OCCURLY 704 00:24:26,416 --> 00:24:30,219 LIKE THE DOUBLE DULL CELLS IN 705 00:24:30,219 --> 00:24:34,056 THE THIEMSZ, CD24 OHIO, THEY'RE 706 00:24:34,056 --> 00:24:36,592 IMMATURE AND QA2 NEGATIVE. 707 00:24:36,592 --> 00:24:38,227 THESE CELLS APPEAR IN THE 708 00:24:38,227 --> 00:24:44,767 PERIPHERY AS IMMATURE CELLS SO 709 00:24:44,767 --> 00:24:48,137 IS IT POSSIBLE THESE CELLS 710 00:24:48,137 --> 00:24:51,641 ARE -- OUTSIDE OF THE THYMUS TO 711 00:24:51,641 --> 00:24:53,509 BECOME IMMATURE CELLS? 712 00:24:53,509 --> 00:24:59,549 TO TEST THAT WE TOOK THE CD4/8 713 00:24:59,549 --> 00:25:01,317 MICE PARK THEM IN A RAG 714 00:25:01,317 --> 00:25:02,285 KNOCK-OUT MICE HERE AND YOU SEE 715 00:25:02,285 --> 00:25:04,353 HERE IN TWO OR TWO AND A HALF 716 00:25:04,353 --> 00:25:07,223 DAY IN THE RAG KNOCK-OUT MOUSE 717 00:25:07,223 --> 00:25:09,192 THESE DOUBLE DULL CELLS ALL BACK 718 00:25:09,192 --> 00:25:09,759 CD8 POSITIVE. 719 00:25:09,759 --> 00:25:11,360 YOU CAN ALSO DO THAT BY SIMPLY 720 00:25:11,360 --> 00:25:13,029 TAKING THESE CELLS AND PUT THEM 721 00:25:13,029 --> 00:25:16,399 INTO CULTURE WITH IL SEVEN IN 722 00:25:16,399 --> 00:25:18,067 VITRO FOR A COUPLE DAYS, SAME 723 00:25:18,067 --> 00:25:21,471 THING HAPPENS, THESE CELLS GO 724 00:25:21,471 --> 00:25:23,873 FROM BEING DOUBLE DULL TO 725 00:25:23,873 --> 00:25:25,508 BECOMING CD8 POSITIVE N THE 726 00:25:25,508 --> 00:25:29,445 REACT IS PD1 POSITIVE NEGATIVELY 727 00:25:29,445 --> 00:25:35,651 SELECTED CD4 CD8 THYMOCYTES, 728 00:25:35,651 --> 00:25:41,691 PREMATURELY EVICTED AS CONTINUE 729 00:25:41,691 --> 00:25:43,359 ARE DIFFERENTIATION IN THE 730 00:25:43,359 --> 00:25:44,427 PERIPHERY, CALLED CLONAL 731 00:25:44,427 --> 00:25:44,694 EVICTION. 732 00:25:44,694 --> 00:25:47,363 HOW DO THESE CELLS GET EVICTED? 733 00:25:47,363 --> 00:25:48,364 HOW IN THE WORLD IS THIS 734 00:25:48,364 --> 00:25:49,565 ACTUALLY POSSIBLE THAT THESE 735 00:25:49,565 --> 00:25:50,666 IMMATURE CELLS GET EE VICK 736 00:25:50,666 --> 00:25:51,868 UNDERSTAND FROM THE THYMUS? 737 00:25:51,868 --> 00:25:55,238 WE KNOW THAT MATURE CELLS 738 00:25:55,238 --> 00:25:58,941 NORMALLY LEAVE THE THYMUS BY 739 00:25:58,941 --> 00:26:01,310 EXPRESSION OF THESE PHOSPHATE 740 00:26:01,310 --> 00:26:06,182 TYPE ONE RECEPTOR, KNOWN AS S1P1 741 00:26:06,182 --> 00:26:10,353 THIS IS THE GENE THAT ENCODES 742 00:26:10,353 --> 00:26:14,323 S1P1 AND YOU CAN SEE NORMALLY 743 00:26:14,323 --> 00:26:17,260 MATURES DON'T NORMALLY EXPRESS 744 00:26:17,260 --> 00:26:19,462 S1PR1, ONLY MATURE, BUT IN 745 00:26:19,462 --> 00:26:20,429 NEGATIVELY SELECTED MALE MOUSE 746 00:26:20,429 --> 00:26:24,033 WE SEE THE CD8 DULL IMMATURE 747 00:26:24,033 --> 00:26:26,335 PRECURSOR CELLS THAT HAVE BEEN 748 00:26:26,335 --> 00:26:28,971 NEGATIVELY SELECTED ALSO EXPRESS 749 00:26:28,971 --> 00:26:29,305 S1P1. 750 00:26:29,305 --> 00:26:31,474 IS THIS IMPORTANT FOR EXPRESSION 751 00:26:31,474 --> 00:26:32,808 OF DEPARTURE LEAVING THE THYMUS? 752 00:26:32,808 --> 00:26:36,178 TO TEST THAT, WE MADE THE HY CD4 753 00:26:36,178 --> 00:26:38,581 MALE MOUSE S1P1 DEFICIENT BY 754 00:26:38,581 --> 00:26:42,184 MAKING THEM S1P1 CONDITIONAL 755 00:26:42,184 --> 00:26:42,451 KNOCK-OUT. 756 00:26:42,451 --> 00:26:43,853 SO WHEREAS THE MALE MOUSE 757 00:26:43,853 --> 00:26:46,689 NORMALLY DOESN'T HAVE ANY CD8 758 00:26:46,689 --> 00:26:50,626 SIGNAL POSITIVE THYNOCYTES WE 759 00:26:50,626 --> 00:26:51,861 FOUND SOMETHING REALLY DRAMATIC, 760 00:26:51,861 --> 00:26:53,129 THAT IS, IF YOU PREVENT THE 761 00:26:53,129 --> 00:26:55,431 CELLS FROM LEAVING THE THYMUS BY 762 00:26:55,431 --> 00:26:57,199 REMOVING S1P1 EXPRESSION, NOW 763 00:26:57,199 --> 00:26:59,435 YOU SUDDENLY SEE CD8 POSITIVE 764 00:26:59,435 --> 00:27:00,703 CELLS APPEARING IN THE 765 00:27:00,703 --> 00:27:04,173 NEGATIVELY SELECTING MALE MOUSE. 766 00:27:04,173 --> 00:27:05,942 SO THIS IS NOT CLONAL DELETION. 767 00:27:05,942 --> 00:27:08,778 THE CELLS HAVEN'T BEEN DELETED 768 00:27:08,778 --> 00:27:10,079 HERE, BECAUSE IF YOU SIMPLE STOP 769 00:27:10,079 --> 00:27:12,848 THEM FROM LEECHG THE THYMUS, THE 770 00:27:12,848 --> 00:27:13,916 CELLS APPEAR. 771 00:27:13,916 --> 00:27:16,419 AND THESE CELLS IN FACT NOW HAVE 772 00:27:16,419 --> 00:27:18,220 BECOME MATURE, THEY'VE MATURED 773 00:27:18,220 --> 00:27:21,891 IN THE THYMUS TO BECOME MATURE 774 00:27:21,891 --> 00:27:24,427 QA2 POSITIVE CD8 SIGNAL POSITIVE 775 00:27:24,427 --> 00:27:25,394 CELLS, THE WAY THAT HAPPENS IS 776 00:27:25,394 --> 00:27:27,163 YOU JUST HAVE TO KEEP THEM FROM 777 00:27:27,163 --> 00:27:28,364 LEAVING THE THYMUS. 778 00:27:28,364 --> 00:27:31,200 SO I THINK THIS EXPERIMENT IN 779 00:27:31,200 --> 00:27:32,668 ITSELF PROVIDES I THINK, I DON'T 780 00:27:32,668 --> 00:27:34,604 KNOW IF IT'S PROOF, BUT IT'S 781 00:27:34,604 --> 00:27:36,238 CERTAINLY COMPELLING EVIDENCE 782 00:27:36,238 --> 00:27:39,208 THAT THE ABSENCE OF CD8 SIGNAL 783 00:27:39,208 --> 00:27:40,610 POSITIVE CELLS IN THESE 784 00:27:40,610 --> 00:27:49,685 NEGATIVELY SELECTING MICE, 785 00:27:49,685 --> 00:27:51,554 EVICTION, THERE THEY ARE. 786 00:27:51,554 --> 00:27:51,787 OKAY. 787 00:27:51,787 --> 00:27:53,089 SO THAT'S OUR CONCLUSION FOR 788 00:27:53,089 --> 00:27:55,925 THIS PART, THAT THE ABSENCE OF 789 00:27:55,925 --> 00:27:59,629 CD8 POSITIVE THYMOCYTES IS DUE 790 00:27:59,629 --> 00:28:01,197 TO CLONAL EVICTION, NOT 791 00:28:01,197 --> 00:28:01,464 DELETION. 792 00:28:01,464 --> 00:28:03,599 HOW IS IT THAT S1P1 WHICH IS 793 00:28:03,599 --> 00:28:04,934 NORMALLY EXPRESSED ONLY ON 794 00:28:04,934 --> 00:28:06,669 MATURE CELLS, HOW IS IT THAT 795 00:28:06,669 --> 00:28:10,940 THESE GUYS, THAT IMMATURE 796 00:28:10,940 --> 00:28:12,808 THYMOCYT HE S ARE INDUCED TO 797 00:28:12,808 --> 00:28:14,777 EXPRESS S1P1? 798 00:28:14,777 --> 00:28:16,045 THERE WAS REALLY A REMARKABLE 799 00:28:16,045 --> 00:28:20,750 SET OF EXPERIMENTS WHAT SHERIF 800 00:28:20,750 --> 00:28:25,488 FOUND, THESE ARE IMMATURE 801 00:28:25,488 --> 00:28:29,291 THYMOCYTES STIMULATED IN VITRO 802 00:28:29,291 --> 00:28:31,193 FLRKS FOR ANTIBODY. 803 00:28:31,193 --> 00:28:35,998 WITHOUT STIMULATION, THYMOCYTES 804 00:28:35,998 --> 00:28:42,004 DON'T EXPRESS S1P1. 805 00:28:42,004 --> 00:28:47,510 SO IMMATURE TRRLS THYMOCYTES 806 00:28:47,510 --> 00:28:49,045 ACTUALLY REQUIRE SIGNAL TO 807 00:28:49,045 --> 00:28:50,012 UPREGULATE AND IF YOU SIGNAL 808 00:28:50,012 --> 00:28:52,648 THEM YOU GET S1P1 EXPRESSION. 809 00:28:52,648 --> 00:28:54,083 IN MATURE CELLS, THE EXACT 810 00:28:54,083 --> 00:28:55,284 OPPOSITE HAPPENS. 811 00:28:55,284 --> 00:28:57,153 BUT HOW IS THIS WORKING? 812 00:28:57,153 --> 00:29:01,157 THE WAY IT WORKS IN FACT IS THAT 813 00:29:01,157 --> 00:29:05,861 UPSTREAM OF S1P1 IS 814 00:29:05,861 --> 00:29:08,330 TRANSCRIPTIONAL PRSES REPRESSOR 815 00:29:08,330 --> 00:29:11,567 PRRKS EXPRESSED AT HIGH LEVEL 816 00:29:11,567 --> 00:29:16,605 IMMATURE THYMOCYTES, RESPONSIBLE 817 00:29:16,605 --> 00:29:22,578 FOR PREVENTING S1P1 BEING 818 00:29:22,578 --> 00:29:24,647 EXPRESSED. 819 00:29:24,647 --> 00:29:33,189 ARE. 820 00:29:33,189 --> 00:29:34,924 SO HOW DO WE PROVE THAT? 821 00:29:34,924 --> 00:29:37,259 REALLY N A REMARKABLE EXPERIMENT 822 00:29:37,259 --> 00:29:41,097 THAT SHARIF CAN HE USED THIS 823 00:29:41,097 --> 00:29:43,733 FLEX SWITCH MS SYSTEM TO TRY TO 824 00:29:43,733 --> 00:29:46,669 INDUCE EXPRESSION OF GFI1 IN 825 00:29:46,669 --> 00:29:49,839 THESE IMMATURE THYME SIGHTS. 826 00:29:49,839 --> 00:29:57,313 SO -- THYMOCYTES, IT WE HAD A 827 00:29:57,313 --> 00:30:00,950 LENTIVIRUS, WE LOOKED AND THE 828 00:30:00,950 --> 00:30:05,554 CONTROL LENTIVIRUS DIDN'T HAVE 829 00:30:05,554 --> 00:30:07,990 CD8 CELLS. 830 00:30:07,990 --> 00:30:18,501 SO HAD YOU CAN SEE THESE CELLS 831 00:30:21,203 --> 00:30:23,873 EXPRESSED HIGH LEVEL OF GFI1 832 00:30:23,873 --> 00:30:27,610 FROM LENTIVIRUS, THESE CELLS 833 00:30:27,610 --> 00:30:29,879 RETAINED IN THYMUS AND DEVELOPED 834 00:30:29,879 --> 00:30:31,647 INTO MATURE CD8 T CELLS. 835 00:30:31,647 --> 00:30:31,847 OKAY. 836 00:30:31,847 --> 00:30:33,649 SO AT THIS POINT, WE WOULD SAY 837 00:30:33,649 --> 00:30:35,017 THAT HERE IS OUR UNDERSTANDING 838 00:30:35,017 --> 00:30:36,919 OF IT. 839 00:30:36,919 --> 00:30:40,322 AND THAT IS THAT STRONG TCR 840 00:30:40,322 --> 00:30:41,524 SIGNALLING DURING NEGATIVE 841 00:30:41,524 --> 00:30:43,993 SELECTION DOWN REGULATES GFI ONE 842 00:30:43,993 --> 00:30:45,895 IN THE NEGATIVELY SELECTED 843 00:30:45,895 --> 00:30:47,263 IMMATURE THYMOCYTES WHICH ALLOWS 844 00:30:47,263 --> 00:30:50,299 THESE CELLS TO EXPRESS S1T1 THAT 845 00:30:50,299 --> 00:30:51,934 CAUSES THESE IMMATURE THYMOCYTES 846 00:30:51,934 --> 00:30:53,135 TO LEAVE THE THYMUS AND GO INTO 847 00:30:53,135 --> 00:30:54,670 THE PERIPHERY. 848 00:30:54,670 --> 00:30:56,205 AND IN THE PERIPHERY, THESE 849 00:30:56,205 --> 00:30:57,873 CELLS ENCOUNTER IL7, PERHAPS 850 00:30:57,873 --> 00:30:59,375 OTHER FACTORS AS WELL, AND THAT 851 00:30:59,375 --> 00:31:01,043 INDUCES THE DIFFERENTIATION FROM 852 00:31:01,043 --> 00:31:03,979 THE DOUBLE DULL CELLS TO BECOME 853 00:31:03,979 --> 00:31:04,980 MATURE CD8 T CELLS. 854 00:31:04,980 --> 00:31:08,250 SO THE NEXT QUESTION WAS, ARE 855 00:31:08,250 --> 00:31:09,685 THESE MATURE CD8 T CELLS THAT 856 00:31:09,685 --> 00:31:11,420 NOW ARISE IN THE PERIPHERY FROM 857 00:31:11,420 --> 00:31:13,289 THESE PRECURSOR CELLS, DO THESE 858 00:31:13,289 --> 00:31:16,125 CELLS R THESE CELLS 859 00:31:16,125 --> 00:31:16,458 SELF-TOLERANT? 860 00:31:16,458 --> 00:31:19,195 AND THE ANSWER IS, YES. 861 00:31:19,195 --> 00:31:22,164 AND THE WAY THIS IS SHOWN WAS 862 00:31:22,164 --> 00:31:27,670 THAT WE TOOK EXPHAIL FEMALE HYC 863 00:31:27,670 --> 00:31:31,574 CD4 MOUSE AND INJECTED THEM WITH 864 00:31:31,574 --> 00:31:35,744 PEPTIDE AND FEMALES RESPONDED BY 865 00:31:35,744 --> 00:31:38,280 UPREGULATING CD107A, YOU CAN SEE 866 00:31:38,280 --> 00:31:40,983 THE FEMALES RESPONDED BUT MALE 867 00:31:40,983 --> 00:31:47,056 CD8 CELLS DID NOT RESPOND. 868 00:31:47,056 --> 00:31:48,390 SO TO SHOW THAT THEY'RE TOLERANT 869 00:31:48,390 --> 00:31:51,026 AS WELL, THEY'RE NONRESPONSIVE. 870 00:31:51,026 --> 00:31:51,260 OKAY. 871 00:31:51,260 --> 00:31:53,028 SO WE THINK THE SOLUTION TO THE 872 00:31:53,028 --> 00:31:55,731 PROBLEM OF HOW YOU GET CELLS 873 00:31:55,731 --> 00:31:56,732 MATURE CD8 CELLS IN THE 874 00:31:56,732 --> 00:31:58,067 PERIPHERY WHEN NONE OF THEM ARE 875 00:31:58,067 --> 00:32:00,002 PRESENT IN THE THYMUS IS THAT 876 00:32:00,002 --> 00:32:02,104 PRECURSORS GET PREMATURELY 877 00:32:02,104 --> 00:32:04,373 CLONEALLY EVICTED BY THE STRONG 878 00:32:04,373 --> 00:32:05,574 TCR SIGNALLING DOWN REGULATION 879 00:32:05,574 --> 00:32:08,878 OF GF IN I1 AND EXPRESSION OF 880 00:32:08,878 --> 00:32:10,613 S1P1 AND THEN COMPLETE THE 881 00:32:10,613 --> 00:32:11,614 DIFFERENTIATION IN THE PERIPHERY 882 00:32:11,614 --> 00:32:18,187 IN THE PRESENCE OF THEIR EXTRA 883 00:32:18,187 --> 00:32:20,556 THYMIC LIGAND, AUTO REACTIVE 884 00:32:20,556 --> 00:32:22,658 LIGAND TO BECOME SELF-TOLERANT 885 00:32:22,658 --> 00:32:23,325 CD8 T CELLS. 886 00:32:23,325 --> 00:32:25,160 IN THE LAST MINUTE OR SO I WANT 887 00:32:25,160 --> 00:32:26,829 TO SHOW YOU THAT THIS IS NOT 888 00:32:26,829 --> 00:32:31,901 SOMETHING UNIQUE TO THE CD8 889 00:32:31,901 --> 00:32:35,371 TRANSGENE OR ANY TRANSGENE 890 00:32:35,371 --> 00:32:37,139 BECAUSE WE LOOKED TO SEE IF THIS 891 00:32:37,139 --> 00:32:39,875 WAS HAPPENING IN NORMAL 892 00:32:39,875 --> 00:32:41,377 POLYCLONAL MICE. 893 00:32:41,377 --> 00:32:43,612 THESE ARE ALL A SET OF QUITE 894 00:32:43,612 --> 00:32:45,247 SURPRISING EXPERIMENTS. 895 00:32:45,247 --> 00:32:51,687 TO LOOK AT POLYCLONAL SELECTED 896 00:32:51,687 --> 00:32:53,689 THYMOCYTES, WE LOOKED IN 897 00:32:53,689 --> 00:32:55,658 KNOCK-OUT MOUSE, HERE NORMAL 898 00:32:55,658 --> 00:32:58,527 MODEL DOESN'T HAVE CD4 T CELLS, 899 00:32:58,527 --> 00:33:00,729 BUT IT DOES HAVE CD8 T CELLS. 900 00:33:00,729 --> 00:33:02,998 THERE'S A SMALL SUBSET OF CELLS 901 00:33:02,998 --> 00:33:04,967 EXPRESS PD1 INDICATING THEY'VE 902 00:33:04,967 --> 00:33:06,502 BEEN STRONGLY SIGNALLED DURING 903 00:33:06,502 --> 00:33:07,736 NEGATIVE SELECTION. 904 00:33:07,736 --> 00:33:08,904 WHAT'S INTERESTING IS THAT IF 905 00:33:08,904 --> 00:33:11,006 YOU LOOK AT THE PD1 NEGATIVE 906 00:33:11,006 --> 00:33:12,641 CELLS THE VAST MAJORITY OF THE 907 00:33:12,641 --> 00:33:15,044 CELLS YOU SEE IT LOOKS JUST LIKE 908 00:33:15,044 --> 00:33:17,913 NORMAL CD8 SIGNAL POSITIVE 909 00:33:17,913 --> 00:33:18,180 SELECTION. 910 00:33:18,180 --> 00:33:19,615 IT LOOKS JUST LIKE THE FEMALE 911 00:33:19,615 --> 00:33:24,687 MOUSE, IF YOU WILL, IN THE 912 00:33:24,687 --> 00:33:27,423 TRANS-- BUT IF YOU LOOK AT THE 913 00:33:27,423 --> 00:33:29,391 PD1 POSITIVE CELLS, DRAMATIC 914 00:33:29,391 --> 00:33:30,693 OBSERVATION, YOU SEE DULL CELLS 915 00:33:30,693 --> 00:33:32,695 IN THE POLYCLONAL MOUSE AND IT 916 00:33:32,695 --> 00:33:36,966 LOOKS JUST LIKE THE HYCD4 917 00:33:36,966 --> 00:33:38,367 NEGATIVELY SELECTING MALE MOUSE 918 00:33:38,367 --> 00:33:41,103 AND THOSE -- OH, SORRY. 919 00:33:41,103 --> 00:33:44,740 AND THOSE DOUBLE DULL CELLS IN 920 00:33:44,740 --> 00:33:49,178 FACT HAVE DOWN REGULATED GFI1 921 00:33:49,178 --> 00:33:54,216 AND UPREGULATED LOGICAL S1P1. 922 00:33:54,216 --> 00:33:56,118 YOU SEE THE SAME IN THE 923 00:33:56,118 --> 00:33:57,519 PERIPHERY AS YOU SEE ON 924 00:33:57,519 --> 00:33:59,421 TRANSGENIC AND THOSE DOUBLE DULL 925 00:33:59,421 --> 00:34:01,023 CELLS IF YOU CULTURE THEM IN 926 00:34:01,023 --> 00:34:03,359 FROM IL7 GO FROM BEING DOUBLE 927 00:34:03,359 --> 00:34:05,661 DULL TO MATURING INTO CD8 T 928 00:34:05,661 --> 00:34:07,796 CELLS JUST AS WE SAW IN THE 929 00:34:07,796 --> 00:34:08,163 TRANSGENIC. 930 00:34:08,163 --> 00:34:12,234 SO THESE CELLS DIFFERENTIATE 931 00:34:12,234 --> 00:34:14,503 INTO THE MATURE CD8 CELLS. 932 00:34:14,503 --> 00:34:16,472 MY LAST DATA SLIDE HERE IS THAT 933 00:34:16,472 --> 00:34:22,478 IF WE LOOK AT A NORMAL B6 MOUSE, 934 00:34:22,478 --> 00:34:24,880 LOOKING AT MALE AND FEMALE MICE 935 00:34:24,880 --> 00:34:27,016 AND LOOK AT HY SPECIFIC CLASS 2 936 00:34:27,016 --> 00:34:30,185 CELLS OR HY SPECIFIC CD8 CLASS 937 00:34:30,185 --> 00:34:31,987 ONE CELLS, AND WHAT YOU CAN SEE 938 00:34:31,987 --> 00:34:34,490 IS IF YOU ACTUALLY USE STAIN TO 939 00:34:34,490 --> 00:34:37,459 GO IDENTIFY HY SPECIFIC T CELL, 940 00:34:37,459 --> 00:34:39,428 YOU SEE IN THE MALE CELLS 941 00:34:39,428 --> 00:34:41,530 THERE'S A LOSS BECAUSE OF CLONAL 942 00:34:41,530 --> 00:34:44,366 DELETION OF THE HY SPECIFIC 943 00:34:44,366 --> 00:34:45,367 CLASS 2 T CELLS. 944 00:34:45,367 --> 00:34:47,870 BUT IF YOU LOOK AT THE CD8 CELLS 945 00:34:47,870 --> 00:34:49,872 IN BOTH EXPHAIL FEMALE THERE'S 946 00:34:49,872 --> 00:34:51,473 NO -- MALE AND FEMALE THERE'S NO 947 00:34:51,473 --> 00:34:53,142 LOSS WHATSOEVER, THESE GUYS HAVE 948 00:34:53,142 --> 00:34:56,311 NOT UNDERGONE CLONAL DLEEKS ARE 949 00:34:56,311 --> 00:34:58,614 THEY'VE UNDERGONE CLONAL 950 00:34:58,614 --> 00:34:58,881 EVICTION. 951 00:34:58,881 --> 00:35:02,084 CONCLUSION WE THINK THATS 952 00:35:02,084 --> 00:35:03,185 POLYCLONAL THYMOCYTES ARE 953 00:35:03,185 --> 00:35:04,953 EVICTED DURING NEGATIVE 954 00:35:04,953 --> 00:35:06,021 SELECTION, COMPLETED 955 00:35:06,021 --> 00:35:06,922 DIFFERENTIATION OF THE TOLERANT 956 00:35:06,922 --> 00:35:07,923 CELLS IN PERIPHERY AND I'VE 957 00:35:07,923 --> 00:35:10,659 SHOWN YOU NOW STUDIES IN BOTH 958 00:35:10,659 --> 00:35:12,528 TCR TRANSGENIC AND NORMAL 959 00:35:12,528 --> 00:35:13,629 POLYCLONAL MOUSE REVEAL THAT 960 00:35:13,629 --> 00:35:15,931 CLONAL EVICTION, NOT CLONAL 961 00:35:15,931 --> 00:35:17,800 DELETION, IS THE BASIS OF CD8 T 962 00:35:17,800 --> 00:35:19,868 CELL TOLERANCE. 963 00:35:19,868 --> 00:35:20,069 OKAY. 964 00:35:20,069 --> 00:35:23,572 SO HERE IS MY LAB. 965 00:35:23,572 --> 00:35:27,342 HERE IS SHERIF, AND MIHO WHO DID 966 00:35:27,342 --> 00:35:29,078 THE FAMOUS FLIP-FLOP MOUSE AND 967 00:35:29,078 --> 00:35:30,579 LAID THE INTELLECTUAL GROUND FOR 968 00:35:30,579 --> 00:35:31,346 THESE EXPERIMENTS AND THIS IS 969 00:35:31,346 --> 00:35:35,484 JUST A PICTURE OF OUR LITTLE 970 00:35:35,484 --> 00:35:36,485 EXPERIMENTAL IMMUNOLOGY BRANCH 971 00:35:36,485 --> 00:35:39,054 AND OUR PICK KNACK LAST YEAR. 972 00:35:39,054 --> 00:35:40,355 SO THANK YOU VERY MUCH -- LAST 973 00:35:40,355 --> 00:35:40,589 JUNE. 974 00:35:40,589 --> 00:35:43,292 SO THANK YOU VERY MUCH. 975 00:35:43,292 --> 00:35:53,469 [APPLAUSE] 976 00:35:57,906 --> 00:35:59,541 >> CAN YOU HEAR NECESSITY? 977 00:35:59,541 --> 00:36:01,944 DO THESE CELLS EXPRESS, THOSE 978 00:36:01,944 --> 00:36:04,379 EVICTED CELLS, THE ONES THAT YOU 979 00:36:04,379 --> 00:36:07,749 FIND, DO THEY EXPRESS CD8 BAY 980 00:36:07,749 --> 00:36:08,851 TASS, THAT'S THE FIRST PART OF 981 00:36:08,851 --> 00:36:12,254 THE QUESTION? 982 00:36:12,254 --> 00:36:13,889 DO THEY EXPRESS BETA? 983 00:36:13,889 --> 00:36:16,191 >> YES, ALPHA ALPHA-BETA T 984 00:36:16,191 --> 00:36:16,391 CELLS. 985 00:36:16,391 --> 00:36:19,495 >> ALSO DO YOU FIND SOME THAT 986 00:36:19,495 --> 00:36:21,530 ARE AM ALPHA IN THE GUT? 987 00:36:21,530 --> 00:36:21,730 >> NO. 988 00:36:21,730 --> 00:36:23,699 >> SO THAT'S REALLY SOMETHING 989 00:36:23,699 --> 00:36:25,801 ENTIRELY DIFFERENT FROM 990 00:36:25,801 --> 00:36:27,336 WHATLY -- AND OTHERS HAVE FOR 991 00:36:27,336 --> 00:36:30,472 INSTANCE PROPOSED. 992 00:36:30,472 --> 00:36:37,346 >> THANK YOU, AL. 993 00:36:37,346 --> 00:36:38,981 DO YOU HEAR ME FROM THE 994 00:36:38,981 --> 00:36:39,281 MICROPHONE? 995 00:36:39,281 --> 00:36:39,848 IS IT ON? 996 00:36:39,848 --> 00:36:40,816 >> YES. 997 00:36:40,816 --> 00:36:47,523 >> FIRST, BOTH REMI AND I BOTH 998 00:36:47,523 --> 00:36:48,590 HAVE TOO MANY GRAY HAIRS TO BE 999 00:36:48,590 --> 00:36:50,592 THE ONLY ONES ASKING QUESTIONS. 1000 00:36:50,592 --> 00:36:53,195 TWO QUICK QUESTIONS, AL, FIRST, 1001 00:36:53,195 --> 00:36:56,865 IT'S A GREAT STORY, CONGRATS, SO 1002 00:36:56,865 --> 00:36:57,799 I DON'T UNDERSTAND HOW YOU SEE 1003 00:36:57,799 --> 00:37:00,169 SUCH A DOWN REGULATION OF CD4 1004 00:37:00,169 --> 00:37:01,637 WHEN THE HALF-LIFE OF CD4 IS 1005 00:37:01,637 --> 00:37:02,738 ACTUALLY QUITE LONG, THAT'S THE 1006 00:37:02,738 --> 00:37:03,405 FIRST PART. 1007 00:37:03,405 --> 00:37:04,473 THE SECOND PART IS THAT WHEN YOU 1008 00:37:04,473 --> 00:37:08,310 LOOKED AT THOSE CELLS THAT WERE 1009 00:37:08,310 --> 00:37:10,445 LOW IN THE CD5 ANTIBODY STAINING 1010 00:37:10,445 --> 00:37:11,914 THERE WERE A BUNCH OF CELLS THAT 1011 00:37:11,914 --> 00:37:15,417 WERE NEITHER CD8 NOR CD4 THAT 1012 00:37:15,417 --> 00:37:16,752 YOU MANAGED TO NOT TALK ABOUT AT 1013 00:37:16,752 --> 00:37:16,919 ALL. 1014 00:37:16,919 --> 00:37:20,989 WHAT WERE THOSE? 1015 00:37:20,989 --> 00:37:24,092 >> SO THE DOUBLE-NEGATIVE CELLS 1016 00:37:24,092 --> 00:37:25,694 THAT EXPRESS NEITHER CD4 OR CD8 1017 00:37:25,694 --> 00:37:27,229 REMAIN TO BE WORKED OUT STILL. 1018 00:37:27,229 --> 00:37:30,332 IT'S NOT CLEAR WHAT THOSE ARE 1019 00:37:30,332 --> 00:37:30,599 ENTIRELY. 1020 00:37:30,599 --> 00:37:32,167 >> AND WHY DO YOU LOSE CD4? 1021 00:37:32,167 --> 00:37:34,136 I LOOKED IT UP, THE HALF-LIFE IS 1022 00:37:34,136 --> 00:37:35,437 SIX TO TWELVE HOURS. 1023 00:37:35,437 --> 00:37:36,705 WHY AFTER AN OVERNIGHT CULTURE 1024 00:37:36,705 --> 00:37:38,073 WHY WOULD YOU SEE NO CD4? 1025 00:37:38,073 --> 00:37:39,608 >> IT'S A TWO DAY CULTURE 1026 00:37:39,608 --> 00:37:39,875 ACTUALLY. 1027 00:37:39,875 --> 00:37:42,444 >> OH, OKAY. 1028 00:37:42,444 --> 00:37:44,680 >> SO IT TAKES A COUPLE OF DAYS 1029 00:37:44,680 --> 00:37:53,322 TO LOSE ALL THE CD4. 1030 00:37:53,322 --> 00:37:55,023 >> IT'S ALL GOOD. 1031 00:37:55,023 --> 00:37:55,390 HELLO. 1032 00:37:55,390 --> 00:37:57,125 HOW DO YOU EXPLAIN THE 1033 00:37:57,125 --> 00:37:58,627 APPEARANCE, SINGLE POSITIVE CD4 1034 00:37:58,627 --> 00:38:00,429 T CELL SUBPOPULATION IN THE 1035 00:38:00,429 --> 00:38:01,296 FEMALE MICE? 1036 00:38:01,296 --> 00:38:03,265 ARE THEY BEING CLONEALLY EVICT 1037 00:38:03,265 --> 00:38:04,666 IN A SIMILAR FASHION? 1038 00:38:04,666 --> 00:38:05,901 >> NO. 1039 00:38:05,901 --> 00:38:11,173 SO WHEN YOU SAY -- SO THESE 1040 00:38:11,173 --> 00:38:14,209 ARE -- SO IN THESE MICE, YOU 1041 00:38:14,209 --> 00:38:18,046 HAVE THESE RAG POSITIVE MICE 1042 00:38:18,046 --> 00:38:19,348 BECAUSE FOR A NUMBER OF REALLY 1043 00:38:19,348 --> 00:38:21,116 JUST I THINK TECHNICAL REASONS, 1044 00:38:21,116 --> 00:38:22,317 THERE MAY BE SOMETHING MORE 1045 00:38:22,317 --> 00:38:23,051 BEHIND AT THAT. 1046 00:38:23,051 --> 00:38:26,355 SO THE CD4 CELLS THAT YOU SEE 1047 00:38:26,355 --> 00:38:28,957 ARE ACTUALLY PROBABLY EXPRESSING 1048 00:38:28,957 --> 00:38:29,958 ENDOGENOUS ALPHA THAT GIVES 1049 00:38:29,958 --> 00:38:31,793 THEM, THAT ALLOWS THEM TO SEE 1050 00:38:31,793 --> 00:38:33,528 CLASS TWO AND BECOME A CD4 T 1051 00:38:33,528 --> 00:38:33,862 CELL. 1052 00:38:33,862 --> 00:38:36,632 SO THOSE ARE NOT EVICTED. 1053 00:38:36,632 --> 00:38:38,200 ARE THE CELLS WE KNOW SO FAR 1054 00:38:38,200 --> 00:38:40,002 FROM THIS STUDY ARE EVICTED ARE 1055 00:38:40,002 --> 00:38:43,105 THE CELLS THAT GET A STRONG TCR 1056 00:38:43,105 --> 00:38:44,006 SIGNAL IN THE THYMUS WHICH WOULD 1057 00:38:44,006 --> 00:38:48,243 BE THE NEGATIVELY SELECTED 1058 00:38:48,243 --> 00:38:48,443 CELLS. 1059 00:38:48,443 --> 00:38:50,012 IF YOU THINK ABOUT IT FOR A 1060 00:38:50,012 --> 00:38:52,114 WHILE, YOU REALIZE THIS MAY 1061 00:38:52,114 --> 00:38:55,250 APPLY TO OTHER THYMIC SUBSETS AS 1062 00:38:55,250 --> 00:38:55,584 WELL. 1063 00:38:55,584 --> 00:38:57,352 THESE ARE THE ONLY ONES WE KNOW 1064 00:38:57,352 --> 00:38:58,020 ABOUT SO FAR. 1065 00:38:58,020 --> 00:38:58,754 IS THAT OKAY? 1066 00:38:58,754 --> 00:39:01,923 >> THANK YOU. 1067 00:39:01,923 --> 00:39:02,090 YES. 1068 00:39:02,090 --> 00:39:05,093 >> THANK YOU, AL. 1069 00:39:05,093 --> 00:39:09,398 [APPLAUSE] 1070 00:39:09,398 --> 00:39:10,866 >> JUST TO QUICKLY REMIND ALL 1071 00:39:10,866 --> 00:39:12,768 THE SPEAKERS TO ALLOW TIME FOR 1072 00:39:12,768 --> 00:39:13,902 QUESTIONS AND SOME DISCUSSION, 1073 00:39:13,902 --> 00:39:18,607 PLEASE KEEP YOUR TALK TO A MAX 1074 00:39:18,607 --> 00:39:20,309 15 MINUTES SO WE HAVE AT LEAST 1075 00:39:20,309 --> 00:39:21,376 FIVE MINUTES FOR QUESTIONS. 1076 00:39:21,376 --> 00:39:23,045 SO I'M GOING TO GO AHEAD AND 1077 00:39:23,045 --> 00:39:30,285 INTRODUCE OUR NEXT SPEAKER, 1078 00:39:30,285 --> 00:39:32,688 REMY BOSSELUT TO TELL US ABOUT 1079 00:39:32,688 --> 00:39:36,558 CD4 T CELL DIFFERENTIATION. 1080 00:39:36,558 --> 00:39:42,831 >> IT'S A TOUGH THING TO DO, 1081 00:39:42,831 --> 00:39:47,602 I'LL TRY MY BEST HAD -- I'LL TRY 1082 00:39:47,602 --> 00:39:50,238 MY BEST. 1083 00:39:50,238 --> 00:40:00,349 OKAY. 1084 00:40:34,516 --> 00:40:35,684 S. 1085 00:40:35,684 --> 00:40:37,319 >> OKAY. 1086 00:40:37,319 --> 00:40:39,254 YES, IP THE -- 1087 00:40:39,254 --> 00:40:41,323 >> YES, I WANT THE POINTER, 1088 00:40:41,323 --> 00:40:41,556 PLEASE. 1089 00:40:41,556 --> 00:40:51,733 THANK YOU. 1090 00:41:26,234 --> 00:41:28,770 THANK YOU VERY MUCH FOR YOUR 1091 00:41:28,770 --> 00:41:29,838 TIME WAITING FOR ME THSM I WOULD 1092 00:41:29,838 --> 00:41:32,174 LIKE TO START BY THANKING THE 1093 00:41:32,174 --> 00:41:40,182 COMMITTEE FOR INVITING ME TODAY. 1094 00:41:40,182 --> 00:41:41,450 MOST OF THE WORK I AM GOING TO 1095 00:41:41,450 --> 00:41:51,493 SHOW WAS DONE BY HIGH -- 1096 00:41:51,493 --> 00:41:54,229 STUDENT, NOW IN MICHIGAN IN 1097 00:41:54,229 --> 00:41:55,764 MEDICAL SCHOOL, SHE'S VERY 1098 00:41:55,764 --> 00:41:56,731 TALENTED ALTOGETHER. 1099 00:41:56,731 --> 00:42:07,709 THIS WAS REALLY, WITH HELP BE -D 1100 00:42:11,079 --> 00:42:12,113 THERE WERE PLENTIFUL PEOPLE IN 1101 00:42:12,113 --> 00:42:14,349 THE LAB WHO HELPED WITH THIS AND 1102 00:42:14,349 --> 00:42:19,287 ARE GOING ON SHOW A -- DRIVING 1103 00:42:19,287 --> 00:42:22,257 FORCE BEHIND THIS, WHEN WE 1104 00:42:22,257 --> 00:42:22,491 STARTED. 1105 00:42:22,491 --> 00:42:26,428 WE HAD FANTASTIC COLLABORATOR 1106 00:42:26,428 --> 00:42:33,401 JEFF CHU, MIKE KELLY, AND WE GOT 1107 00:42:33,401 --> 00:42:34,970 MICE FROM -- AND NONE OF THIS 1108 00:42:34,970 --> 00:42:35,971 WOULD HAVE BEEN POSSIBLE WITHOUT 1109 00:42:35,971 --> 00:42:37,806 THE ASSISTANCE OF THE CENTER FOR 1110 00:42:37,806 --> 00:42:41,743 CANCER RESEARCH FOR FUNDING NEW 1111 00:42:41,743 --> 00:42:43,144 TECHNOLOGY AND INSTRUMENTAL FOR 1112 00:42:43,144 --> 00:42:43,345 THIS. 1113 00:42:43,345 --> 00:42:46,915 SO THIS IS WHAT I AM GOING TO 1114 00:42:46,915 --> 00:42:48,884 ADDRESS, REALLY TWO SHORT 1115 00:42:48,884 --> 00:42:51,152 VIGNETTES ON SINGLE CELL 1116 00:42:51,152 --> 00:42:53,455 TRANSCRIPT TORE THE ROLE OF 1117 00:42:53,455 --> 00:42:55,524 THESE TWO FACTORS, IN THE THYMUS 1118 00:42:55,524 --> 00:42:56,258 LEADING TO THE TRANSITION. 1119 00:42:56,258 --> 00:42:57,826 SO AS MANY PEOPLE HERE ARE 1120 00:42:57,826 --> 00:42:59,594 INTERESTED IN THE FUNCTION OF T 1121 00:42:59,594 --> 00:43:01,763 CELL AND DIFFERENTIATION IN 1122 00:43:01,763 --> 00:43:05,500 PARTICULAR CONVENTIONAL T CELLS 1123 00:43:05,500 --> 00:43:09,204 THAT BIND WITH PEPTIDES AND 1124 00:43:09,204 --> 00:43:12,140 EXPRESS CD4, BIND CLASS ONE 1125 00:43:12,140 --> 00:43:15,677 EXPRESS CD8 SO EMERGE FROM 1126 00:43:15,677 --> 00:43:16,978 THYMUS I THINK I ALREADY 1127 00:43:16,978 --> 00:43:18,613 INTRODUCED THIS EXPRESS BOTH CD4 1128 00:43:18,613 --> 00:43:22,050 AND CD8 AND SO BECOME CD4 1129 00:43:22,050 --> 00:43:23,552 POSITIVE AND THIS IS A VERY 1130 00:43:23,552 --> 00:43:26,288 SIMPLIFIED VIEW OF WHAT WE THINK 1131 00:43:26,288 --> 00:43:29,357 HOW THIS IS HAPPENING. 1132 00:43:29,357 --> 00:43:32,961 THE CD4 THYMUS EXPRESS 1133 00:43:32,961 --> 00:43:33,929 TRANSCRIPTION FACTOR, EXPRESSION 1134 00:43:33,929 --> 00:43:35,697 OF THIS OTHER FACTOR EXPRESSED 1135 00:43:35,697 --> 00:43:39,000 IN CD8 CELLS, SO THAT THE CELL 1136 00:43:39,000 --> 00:43:42,504 EXPRESSED NORMALLY -- SIGNAL 1137 00:43:42,504 --> 00:43:44,039 BECOMES CD4 AND DON'T EXPRESSES 1138 00:43:44,039 --> 00:43:46,775 ARE CLASS ONE SIGNAL, BECOME 1139 00:43:46,775 --> 00:43:48,343 CD8. 1140 00:43:48,343 --> 00:43:52,213 NOW, IF NOT COMPLETE AT THIS, 1141 00:43:52,213 --> 00:43:55,850 WHAT HAPPENS THERE BEFORE THE 1142 00:43:55,850 --> 00:43:57,319 CELLS EXPRESS BECAUSE POSITIVE 1143 00:43:57,319 --> 00:44:00,455 EXPRESS NEITHER OF THEM, DOESN'T 1144 00:44:00,455 --> 00:44:01,756 MEAN MUCH LESS CLEAR IN PART 1145 00:44:01,756 --> 00:44:03,258 BECAUSE THE CELLS THAT MAKE THIS 1146 00:44:03,258 --> 00:44:06,328 DECISION ARE IN THIS -- MY MOUSE 1147 00:44:06,328 --> 00:44:08,697 ISN'T MAKING IT, BUT WHERE THEY 1148 00:44:08,697 --> 00:44:12,801 ARE CELLS THAT ARE POSITIVE AND 1149 00:44:12,801 --> 00:44:15,837 SELECTION ARE PRECURSOR AND OF 1150 00:44:15,837 --> 00:44:17,272 COURSE THEY ARE EXPRESSING THE 1151 00:44:17,272 --> 00:44:17,472 CELLS. 1152 00:44:17,472 --> 00:44:20,008 SO TRY TO DISENTANGLE THIS 1153 00:44:20,008 --> 00:44:22,744 COMPLEXITY SOME YEARS AGO, THIS 1154 00:44:22,744 --> 00:44:23,678 IDEA WHAT WAS AT THE TIME IN 1155 00:44:23,678 --> 00:44:26,948 THIS TECHNOLOGY, SINGLE CELL 1156 00:44:26,948 --> 00:44:29,117 SEQUENCING, AND STARTED FROM 1157 00:44:29,117 --> 00:44:32,621 CLASS ONE TO CLASS TWO KNOCK-OUT 1158 00:44:32,621 --> 00:44:36,891 MOUSE AND THE CD69 POSITIVE 1159 00:44:36,891 --> 00:44:39,160 THYMOCYTES, ALREADY A LOOSE 1160 00:44:39,160 --> 00:44:39,361 GATE. 1161 00:44:39,361 --> 00:44:41,930 SO THIS IS A KNOCK-OUT RECEPTOR 1162 00:44:41,930 --> 00:44:44,733 SIGNALLING, THE CELL EXPRESSED 1163 00:44:44,733 --> 00:44:47,802 IN 69 I'VE SEEN SOME SIGNALLING. 1164 00:44:47,802 --> 00:44:50,005 SHE STARTED THOSE CELLS, 1165 00:44:50,005 --> 00:44:53,074 CAPTURED THEM WITH THIS 10X 1166 00:44:53,074 --> 00:44:55,243 DROPLET SYSTEM AND DID A NUMBER 1167 00:44:55,243 --> 00:44:56,578 OF BIOINFORMATICS. 1168 00:44:56,578 --> 00:44:58,446 I WILL NOW DETAIL. 1169 00:44:58,446 --> 00:44:59,748 THIS WAS PUBLISHED A COUPLE 1170 00:44:59,748 --> 00:45:01,516 YEARS AGO. 1171 00:45:01,516 --> 00:45:04,552 IT AND THIS IS WHAT YOU SEE THE 1172 00:45:04,552 --> 00:45:08,923 WHOLE FAMILY, SHOWING CELLS, AND 1173 00:45:08,923 --> 00:45:10,925 THE CELLS ARE ARRANGED IN SUCH A 1174 00:45:10,925 --> 00:45:12,894 WAY THAT THEIR DISTANCE IN THE 1175 00:45:12,894 --> 00:45:14,295 BLOOD MORE OR LESS MATCHES THEIR 1176 00:45:14,295 --> 00:45:19,534 TRANSCRIPT TORE SIMILARITY. 1177 00:45:19,534 --> 00:45:22,737 NOW, PUTTING TOGETHER CELLS THAT 1178 00:45:22,737 --> 00:45:26,675 ARE CLOSE TO EACH OTHER, THESE 1179 00:45:26,675 --> 00:45:27,909 BROWN-RED CELLS ARE CLOSER TO 1180 00:45:27,909 --> 00:45:29,177 EACH OTHER THAN THE ORANGE 1181 00:45:29,177 --> 00:45:29,377 CELLS. 1182 00:45:29,377 --> 00:45:31,079 WE CAN IDENTIFY THOSE CRYSTALS 1183 00:45:31,079 --> 00:45:41,623 BY GENE EXPRESSION, -- THE ONES 1184 00:45:44,759 --> 00:45:46,961 CD4 AND CD8 AND SO ON AND SO 1185 00:45:46,961 --> 00:45:47,262 FORTH. 1186 00:45:47,262 --> 00:45:48,496 THIS AS I'VE BEEN TOLD IN THE 1187 00:45:48,496 --> 00:45:51,132 PAST IS A LITTLE BIT, HAS A 1188 00:45:51,132 --> 00:45:54,969 LITTLE BIT FEELING OF DEJA VU. 1189 00:45:54,969 --> 00:45:56,905 NOW, WE CAN DIETLE MORE THING, 1190 00:45:56,905 --> 00:45:59,340 WE CAN TRY TO USABLE CIS TO DO 1191 00:45:59,340 --> 00:46:01,876 TRAJECTORY MAPPING AND FIND A 1192 00:46:01,876 --> 00:46:03,712 WAY -- FIND WHERE THE LINEAGES 1193 00:46:03,712 --> 00:46:07,649 BRANCH OUT FROM EACH OTHER AND 1194 00:46:07,649 --> 00:46:09,517 ALSO INDEED WHAT HAPPENS VERY 1195 00:46:09,517 --> 00:46:11,686 EARLY, SO THOSE ARE THE DOUBLE 1196 00:46:11,686 --> 00:46:15,223 POSITIVE THYMOCYTES AND WE CAN 1197 00:46:15,223 --> 00:46:21,262 TRY TO LOOK AT THIS SH TO SEE 1198 00:46:21,262 --> 00:46:23,131 OUR TRY TO SEE HOW GENE 1199 00:46:23,131 --> 00:46:24,532 EXPRESSION WORKS AND IF WE DO 1200 00:46:24,532 --> 00:46:25,767 THIS, YOU CAN SEE WE REALLY 1201 00:46:25,767 --> 00:46:28,303 CATCH THE CELLS IN THE ACT OF 1202 00:46:28,303 --> 00:46:32,207 SIGNALLING, THESE ARE CD5 AND 1203 00:46:32,207 --> 00:46:33,408 CD69 TO SIGNALLING AND WE CAN 1204 00:46:33,408 --> 00:46:35,443 SEE THIS INCREASE AS THE CELL 1205 00:46:35,443 --> 00:46:38,613 MOVES FROM DP1 TO DP2 AND 1206 00:46:38,613 --> 00:46:41,249 CONVERSELY IF WE LOOK AT GENES 1207 00:46:41,249 --> 00:46:43,752 EXPRESSED BY TCR ORANGE CELLS, 1208 00:46:43,752 --> 00:46:46,387 WE SEE THEM GOING DOWN AS THE 1209 00:46:46,387 --> 00:46:48,156 CELLS STOP SIGNALLING. 1210 00:46:48,156 --> 00:46:51,426 AND WE CAN ALSO TRACK THE 1211 00:46:51,426 --> 00:46:53,695 EXPRESSION AT LEAST IN THIS 1212 00:46:53,695 --> 00:46:55,563 EARLY PHASE, MORE OR LESS THE 1213 00:46:55,563 --> 00:46:58,533 SAME WAY REGARDLESS OF IMAGE 1214 00:46:58,533 --> 00:46:58,833 SPECIFICITY. 1215 00:46:58,833 --> 00:47:00,535 WITH THIS, WE CAN TRY TO LOOK 1216 00:47:00,535 --> 00:47:02,637 INTO THOSE CELLS INTO THEIR 1217 00:47:02,637 --> 00:47:03,938 PROGENY AND SEE WHICH 1218 00:47:03,938 --> 00:47:05,607 TRANSCRIPTION FACTORS COULD BE 1219 00:47:05,607 --> 00:47:07,375 IMPORTANT FOR LINEAGE 1220 00:47:07,375 --> 00:47:08,676 DIFFERENTIATION AND WE FIND A 1221 00:47:08,676 --> 00:47:10,011 NUMBER OF PATTERNS I'M GOING ON 1222 00:47:10,011 --> 00:47:11,446 SHOW YOU EVERYTHING, OF COURSE, 1223 00:47:11,446 --> 00:47:13,148 THIS WOULD BE WAY TOO BIG. 1224 00:47:13,148 --> 00:47:18,720 BUT THIS PATTERN, THE GENE 1225 00:47:18,720 --> 00:47:20,889 EXPRESSION IN CLASS ONE SIGNAL 1226 00:47:20,889 --> 00:47:22,557 CELL, NO EXPRESSION IN CLASS ONE 1227 00:47:22,557 --> 00:47:24,092 SIGNAL CELL, AND EXPRESSION AS 1228 00:47:24,092 --> 00:47:27,162 THE CELLS BECOME IMMATURE MATURE 1229 00:47:27,162 --> 00:47:27,328 CD4. 1230 00:47:27,328 --> 00:47:28,696 THERE ARE OTHER FACTORS THAT ARE 1231 00:47:28,696 --> 00:47:31,132 EXPRESSED MUCH EARLIER THAT WE 1232 00:47:31,132 --> 00:47:34,569 CAN SEE IN THESE DP2 CELLS IN 1233 00:47:34,569 --> 00:47:37,672 WHICH THERE ARE STILL ARRANGING 1234 00:47:37,672 --> 00:47:39,541 CELLS AND WHAT I AM GOING TO 1235 00:47:39,541 --> 00:47:43,778 DISCUSS TODAY IS THIS ADDITIONAL 1236 00:47:43,778 --> 00:47:46,014 FACTOR, 281 THAT'S UNIQUE TO 1237 00:47:46,014 --> 00:47:48,950 SPECIFIC AND COMES EARLY IN THE 1238 00:47:48,950 --> 00:47:49,184 PROCESS. 1239 00:47:49,184 --> 00:47:51,119 SO THIS IS JUST TO SUMMARIZE 1240 00:47:51,119 --> 00:47:52,654 WHAT I JUST SAID. 1241 00:47:52,654 --> 00:47:56,257 AND THE QUESTION WE'RE ASKING IS 1242 00:47:56,257 --> 00:47:58,993 WHETHER, YOU KNOW, 281 IS 1243 00:47:58,993 --> 00:48:01,396 IMPORTANT FOR DIFFERENTIATION. 1244 00:48:01,396 --> 00:48:03,264 SO WE CONTACTED, WE DID SOME 1245 00:48:03,264 --> 00:48:04,766 RESEARCH AROUND TO SEE WHAT WAS 1246 00:48:04,766 --> 00:48:10,572 DONE ON 281 AND REALIZED THAT IN 1247 00:48:10,572 --> 00:48:13,007 CHINA THEY HAD IN FACT MADE A 1248 00:48:13,007 --> 00:48:15,677 KNOCK-OUT OF 281 SO WE CONTACTED 1249 00:48:15,677 --> 00:48:17,745 LI AND OF COURSE TO SOME EXTENT 1250 00:48:17,745 --> 00:48:21,449 WE USE A HOAX BECAUSE 281 WAS 1251 00:48:21,449 --> 00:48:23,518 INDEED IMPORTANT FOR T CELL 1252 00:48:23,518 --> 00:48:25,820 FUNCTION BUT DIDN'T FIND MUCH IN 1253 00:48:25,820 --> 00:48:26,821 APPROXIMATE T CELL 1254 00:48:26,821 --> 00:48:27,488 DIFFERENTIATION IN THE THYMUS. 1255 00:48:27,488 --> 00:48:33,061 SO IT TOOK MORE THAN THIS AND 1256 00:48:33,061 --> 00:48:35,330 REALIZE THAT IN FACT IN ADDITION 1257 00:48:35,330 --> 00:48:37,732 TO 281 THERE IS THIS OTHER 1258 00:48:37,732 --> 00:48:40,235 FACTOR 148, VERY SIMILAR TO 281 1259 00:48:40,235 --> 00:48:44,339 AND THIS IS HOW IN FACT, THERE 1260 00:48:44,339 --> 00:48:49,143 IS NO -- OUTSIDE ITSELF BETWEEN 1261 00:48:49,143 --> 00:48:50,044 THESE TWO FACTORS AND ANYTHING 1262 00:48:50,044 --> 00:48:54,515 ELSE. SO WE OBTAINED FROM 1263 00:48:54,515 --> 00:48:56,017 JUANITA WHO WAS AT MICHIGAN THE 1264 00:48:56,017 --> 00:48:57,919 COMMISSIONER MOUSE FOR 148 AND 1265 00:48:57,919 --> 00:49:00,421 WE DECIDED TO ALSO THIS IS JUST 1266 00:49:00,421 --> 00:49:02,790 SORRY THE EXPRESSION FACTOR OF 1267 00:49:02,790 --> 00:49:04,893 281 THAT'S EXPRESSED IN TWO 1268 00:49:04,893 --> 00:49:09,063 SIGNAL CELLS, 148 IS EXPRESSED 1269 00:49:09,063 --> 00:49:11,132 IN THYMOCYTES REGARDLESS OF I AM 1270 00:49:11,132 --> 00:49:13,534 MARGE SPECIFICITY. 1271 00:49:13,534 --> 00:49:23,177 SO 148 AND 281 USING CD4 THYMUS 1272 00:49:23,177 --> 00:49:27,882 AND FB DISPAISH WE DELETED THESE 1273 00:49:27,882 --> 00:49:37,125 TWO GENES WITH THE PDK. I'LL BE 1274 00:49:37,125 --> 00:49:39,527 BRIEF. 1275 00:49:39,527 --> 00:49:43,197 SO IN SHOWING THIS, OKAY, THE 1276 00:49:43,197 --> 00:49:46,034 281 DOESN'T HAVE MUCH OF -- THE 1277 00:49:46,034 --> 00:49:49,270 148 DOESN'T HAVE MUCH. 1278 00:49:49,270 --> 00:49:50,905 BOTH THOSE DHOINGTZ SEEM TO GO 1279 00:49:50,905 --> 00:49:52,006 VERY WELL, YOU HAVE THIS 1280 00:49:52,006 --> 00:49:53,174 POPULATION OF DOUBLE POSITIVE 1281 00:49:53,174 --> 00:49:57,712 CELLS AND THEY'RE ALL INDEED 1282 00:49:57,712 --> 00:49:59,881 DOUBLE POSITIVE THYMOCYTES, BUT 1283 00:49:59,881 --> 00:50:03,217 HERE WE SEE THAT THE EXPRESSION 1284 00:50:03,217 --> 00:50:05,186 GOES DOWN BY 50%, BOTH IN THYMUS 1285 00:50:05,186 --> 00:50:06,821 AND WHAT I'M SHOWING YOU ON 1286 00:50:06,821 --> 00:50:14,362 THYMUS IS ALSO TRUE IN THE -- SO 1287 00:50:14,362 --> 00:50:16,030 THAT'S SOMETHING BUT NOT 1288 00:50:16,030 --> 00:50:16,297 GIGANTIC. 1289 00:50:16,297 --> 00:50:18,433 WE LOOK AT SINGLE CELL AND THERE 1290 00:50:18,433 --> 00:50:20,735 IS MUCH BOLDER IN FACT THAT 1291 00:50:20,735 --> 00:50:21,869 STAINING CAN TELL US. 1292 00:50:21,869 --> 00:50:23,571 SO THIS IS SINGLE CELL 1293 00:50:23,571 --> 00:50:25,239 SEQUENCING ON WILD-TYPE CELLS 1294 00:50:25,239 --> 00:50:29,844 WITH THOSE SAME IN THE 1295 00:50:29,844 --> 00:50:30,478 ORGANIZATIONS. 1296 00:50:30,478 --> 00:50:31,546 AND I WANT YOU TO PAY ATTENTION 1297 00:50:31,546 --> 00:50:33,314 ON THESE ORANGE CELLS, IMMATURE 1298 00:50:33,314 --> 00:50:34,816 CD4 IN THE KNOCK-OUT. 1299 00:50:34,816 --> 00:50:37,485 THERE IS NO ORANGE CELL INSIDE 1300 00:50:37,485 --> 00:50:42,657 THE CD4 AREA, THERE ARE CRYSTAL 1301 00:50:42,657 --> 00:50:45,927 SEPARATING, SO THERE'S A MAJOR 1302 00:50:45,927 --> 00:50:46,894 TRANSCRIPTOR CHANGE BETWEEN THIS 1303 00:50:46,894 --> 00:50:49,097 AND THIS AS WE REMOVE THOSE TWO 1304 00:50:49,097 --> 00:50:49,330 FACTORS. 1305 00:50:49,330 --> 00:50:50,598 WE TRIED TO UNDERSTAND HOW THIS 1306 00:50:50,598 --> 00:50:58,139 WAS WORKING, SO ANALYSIS ON THE 1307 00:50:58,139 --> 00:51:02,510 THPOK GENE AND FOUND THAT 1308 00:51:02,510 --> 00:51:05,179 THPOK 281 BINDING TWO TWO -- 1309 00:51:05,179 --> 00:51:08,016 BINDING TO TWO SITES, THOSE RED 1310 00:51:08,016 --> 00:51:09,784 BLOTS, YEARS AGO, THIS WAS 1311 00:51:09,784 --> 00:51:11,619 BINDING TO THE TRANSCRIPTION 1312 00:51:11,619 --> 00:51:14,022 FACTOR AS I DISCUSSED EARLIER. 1313 00:51:14,022 --> 00:51:15,556 SO WHAT THIS SUGGESTED TO US IS 1314 00:51:15,556 --> 00:51:18,393 THAT THERE MIGHT BE AN 1315 00:51:18,393 --> 00:51:24,866 INTERACTION BETWEEN WHAT THE BE 1316 00:51:24,866 --> 00:51:26,701 GATA3 AND 218 CHT IF YOU EXPRESS 1317 00:51:26,701 --> 00:51:29,437 BOTH IN CELL LINE YOU WILL FIND 1318 00:51:29,437 --> 00:51:34,242 281 WILL BIND TO GATA3 AND WE 1319 00:51:34,242 --> 00:51:36,110 FOUND TREATMENT OF THE MICE AND 1320 00:51:36,110 --> 00:51:37,879 DOESN'T SIMPLY AFFECT THE 1321 00:51:37,879 --> 00:51:39,847 BINDING DNA IN A MANNER THAT 1322 00:51:39,847 --> 00:51:41,816 DOESN'T INVOLVE PROTEIN-PROTEIN 1323 00:51:41,816 --> 00:51:42,150 INTERACTIONS. 1324 00:51:42,150 --> 00:51:44,886 SO THIS TO SUMMARIZE THIS, SO 1325 00:51:44,886 --> 00:51:49,390 THESE TWO FACTORS, 1326 00:51:49,390 --> 00:51:51,259 DIFFERENTIATION WITH THYMUS, 1327 00:51:51,259 --> 00:51:55,229 BIND THPOK AND EXPRESSION. 1328 00:51:55,229 --> 00:52:00,568 THE FACT THEY BIND TO 3, I WAS 1329 00:52:00,568 --> 00:52:01,436 THINKING ABOUT WHAT ELSE THEY 1330 00:52:01,436 --> 00:52:03,871 COULD DO TOGETHER, AND THIS IS 1331 00:52:03,871 --> 00:52:05,606 HOW WE GOT INTERESTED INTO THE 1332 00:52:05,606 --> 00:52:09,110 POTENTIAL FUNCTION OF TH2 1333 00:52:09,110 --> 00:52:09,477 DIFFERENTIATION. 1334 00:52:09,477 --> 00:52:12,280 SO WE DID A NUMBER OF 1335 00:52:12,280 --> 00:52:13,514 EXPERIMENTS, IN THE INTEREST OF 1336 00:52:13,514 --> 00:52:15,583 TIME I'M NOT GOING TO SHOW, BUT 1337 00:52:15,583 --> 00:52:21,055 WHAT WE DID WITH TH2 IS LOOK AT 1338 00:52:21,055 --> 00:52:26,561 THESE MICE ASK SEE HOW THEY 1339 00:52:26,561 --> 00:52:31,265 RESPOND WITH MEUNTD, PRODUCES AN 1340 00:52:31,265 --> 00:52:35,069 AIRWAYLY P -- IN A WILD-TYPE 1341 00:52:35,069 --> 00:52:36,838 MOUSE YOU CAN FIND IN THE LUNG 1342 00:52:36,838 --> 00:52:40,241 OR LYMPH NODE SOME CELLS THAT 1343 00:52:40,241 --> 00:52:43,878 DIFFERENTIATE INTO TH2 AND 1344 00:52:43,878 --> 00:52:46,380 PRODUCE INTO CYTOKINES. 1345 00:52:46,380 --> 00:52:48,749 AND THIS IS STRONGLY AFFECTED IN 1346 00:52:48,749 --> 00:52:52,120 THE PDKO MOUSE IN WHICH THE T 1347 00:52:52,120 --> 00:52:54,722 CELLS IS NOT EXPRESSED, MUCH 1348 00:52:54,722 --> 00:52:56,591 LESS OF THESE TWO CYTOKINES, 1349 00:52:56,591 --> 00:53:00,528 MUCH LESS AS FOUR AS WELL. 1350 00:53:00,528 --> 00:53:02,497 WHAT IS SOMEWHAT REMARKABLE, THE 1351 00:53:02,497 --> 00:53:06,767 FACTOR IS CRITICAL TO MS 1352 00:53:06,767 --> 00:53:07,535 DIFFERENTIATION, NOT MUCH 1353 00:53:07,535 --> 00:53:08,736 AFFECTED BY THE KNOCK-OUT. 1354 00:53:08,736 --> 00:53:11,272 WHAT WE SUSPECTED IS THAT THE 1355 00:53:11,272 --> 00:53:15,109 CELLS PERHAPS DIDN'T MAKE THE 1356 00:53:15,109 --> 00:53:18,946 CYTOKINES, THAT MICE ABLE TO 1357 00:53:18,946 --> 00:53:20,348 DIFFERENTIATE INTO SOMETHING 1358 00:53:20,348 --> 00:53:21,649 LIKE -- SO TO TRY TO UNDERSTAND 1359 00:53:21,649 --> 00:53:24,085 WHAT WAS GOING ON IN THOSE MICE, 1360 00:53:24,085 --> 00:53:26,154 AGAIN SIGNIFICANT HE WILL CELL 1361 00:53:26,154 --> 00:53:28,256 STUDIES, THIS TIME SHE DID 1362 00:53:28,256 --> 00:53:33,928 TECHNOLOGY THAT DOES TOGETHER ON 1363 00:53:33,928 --> 00:53:43,004 THE SAME SLIDE. 1364 00:53:43,004 --> 00:53:44,872 THIS WAS DONE. 1365 00:53:44,872 --> 00:53:47,642 AND THE SAME KIND AS SHE DID 1366 00:53:47,642 --> 00:53:47,875 BEFORE. 1367 00:53:47,875 --> 00:53:51,245 AGAIN, SO WE GET THIS MAP AND 1368 00:53:51,245 --> 00:53:52,146 CRYSTALS THAT WE CAN NUMBER AND 1369 00:53:52,146 --> 00:53:53,614 WE CAN TRY TO IDENTIFY THOSE 1370 00:53:53,614 --> 00:53:58,553 CRYSTALS AND YOU CAN SEE 1371 00:53:58,553 --> 00:54:01,289 EXPRESSED, IN4 IS EXPRESSED THIS 1372 00:54:01,289 --> 00:54:03,357 IN THIS BRANCH, AND WE FIND 1373 00:54:03,357 --> 00:54:10,264 EXPRESSION OF L21 OR ASCL2 ON 1374 00:54:10,264 --> 00:54:13,000 THE VERY BOTTOM OF THIS WING SO 1375 00:54:13,000 --> 00:54:21,876 WITH THIS SOME CRYSTAL AT LEASTW 1376 00:54:21,876 --> 00:54:27,915 YOU, THIS EXPRESS. 1377 00:54:27,915 --> 00:54:31,219 WHAT HAPPENS IF WE LOOK NOW, 1378 00:54:31,219 --> 00:54:32,620 WELL, NOT MUCH, AND THAT WAS 1379 00:54:32,620 --> 00:54:33,721 SOMETHING THAT WAS A SURPRISE, 1380 00:54:33,721 --> 00:54:34,655 BECAUSE IF YOU REMEMBER ALREADY 1381 00:54:34,655 --> 00:54:36,023 IN THE THYMUS THERE WAS A CLEAR 1382 00:54:36,023 --> 00:54:37,992 SHIFT OF THE KNOCK-OUT CONTROL 1383 00:54:37,992 --> 00:54:39,393 CELLS, HERE THERE IS NOT MUCH 1384 00:54:39,393 --> 00:54:40,127 THAT HAPPENS. 1385 00:54:40,127 --> 00:54:41,362 IN FACT, THERE IS INDEED A 1386 00:54:41,362 --> 00:54:43,864 CHANGE IN L4 EXPRESSION, THAT 1387 00:54:43,864 --> 00:54:45,866 EXPRESSION IS NOT AFFECTED IN 1388 00:54:45,866 --> 00:54:48,369 WHICH WE DO A BOLD SEARCH WE CAN 1389 00:54:48,369 --> 00:54:49,937 SHOW YOU, THERE ARE PROBABLY A 1390 00:54:49,937 --> 00:54:52,006 FEW GENES THAT DIFFERENTIATE 1391 00:54:52,006 --> 00:54:53,007 EXPRESSION BUT NOT MANY AND 1392 00:54:53,007 --> 00:54:54,208 THERE'S REALLY NOT MUCH THAT'S 1393 00:54:54,208 --> 00:55:00,348 GOING ON. 1394 00:55:00,348 --> 00:55:02,717 IF YOU DO ATTACK SIGNAL THINGS 1395 00:55:02,717 --> 00:55:03,551 ARE SLIGHTLY DIFFERENT. 1396 00:55:03,551 --> 00:55:05,553 WHAT YOU CAN SEE IS THAT WE CAN 1397 00:55:05,553 --> 00:55:08,289 PROJECT THE TRANSCRIPTOME ON 1398 00:55:08,289 --> 00:55:10,258 THIS MAP L WE CAN SEE A VERY 1399 00:55:10,258 --> 00:55:12,426 GOOD CAUSE BETWEEN THE 1400 00:55:12,426 --> 00:55:14,762 TRANSCRIPTOME TRK THE RAFN AND 1401 00:55:14,762 --> 00:55:17,098 THE WAY THE CELLS ARE AND YOU 1402 00:55:17,098 --> 00:55:23,871 CAN ALSO SEE THAT THE TH2 CELLS 1403 00:55:23,871 --> 00:55:28,576 ARE SOMEWHAT MOVED AND YOU FIND 1404 00:55:28,576 --> 00:55:29,610 THE CRYSTALLINE DIFFERENT, I 1405 00:55:29,610 --> 00:55:31,646 HOPE YOU CAN SEE THIS BLUISH. 1406 00:55:31,646 --> 00:55:34,048 THIS SUGGESTED TO US THAT IN 1407 00:55:34,048 --> 00:55:36,784 FACT THERE IS A DIFFERENCE, IT'S 1408 00:55:36,784 --> 00:55:37,985 NOT SO MUCH TRIPTD TORE 1409 00:55:37,985 --> 00:55:39,987 DIFFERENCE BUT THERE IS A 1410 00:55:39,987 --> 00:55:42,290 DIFFERENCE AT THE LEVEL OF 1411 00:55:42,290 --> 00:55:44,158 CONNECTING ACCESSIBILITY. 1412 00:55:44,158 --> 00:55:51,032 SO TO LOOK INTO THIS, TH2 1413 00:55:51,032 --> 00:55:55,403 EXPRESSION, WE DO THIS WITH 1414 00:55:55,403 --> 00:55:58,072 CRYSTALS IT, THIS IS WHERE 1415 00:55:58,072 --> 00:56:00,908 TISSUE CYTOKINE WILL BE ALL THE 1416 00:56:00,908 --> 00:56:03,577 WAY ON YOUR RIGHT ARE ENCODED. 1417 00:56:03,577 --> 00:56:05,846 THIS IS A CRYSTAL THEETS NOT A 1418 00:56:05,846 --> 00:56:07,982 TISSUE CRYSTAL, TH1 CELLS AND 1419 00:56:07,982 --> 00:56:11,185 YOU CAN SEE THAT THERE ARE A FEW 1420 00:56:11,185 --> 00:56:12,953 PEAKS THAT HAPPEN. 1421 00:56:12,953 --> 00:56:14,121 BUT THERE ARE OTHER AREAS IN 1422 00:56:14,121 --> 00:56:16,223 WHICH THERE IS A MUCH LESS 1423 00:56:16,223 --> 00:56:22,463 COMPARED TO THIS WILD-TYPE TH2 1424 00:56:22,463 --> 00:56:24,865 CRYSTAL -- CLUSTER, AND THIS IS 1425 00:56:24,865 --> 00:56:27,368 PARTICULARLY TRUE AT A SITE 1426 00:56:27,368 --> 00:56:29,570 THAT'S BEEN IDENTIFIED YEARS AGO 1427 00:56:29,570 --> 00:56:32,973 AS BEING IMPORTANT FOR TH2 1428 00:56:32,973 --> 00:56:42,516 CYTOKINE EXPRESSION, CGRE, CNS1 1429 00:56:42,516 --> 00:56:42,850 AND 4. 1430 00:56:42,850 --> 00:56:45,486 YOU CAN SEE THERE IS DEFINITELY 1431 00:56:45,486 --> 00:56:47,355 AN CRERKS HERE, HERE, HERE. 1432 00:56:47,355 --> 00:56:49,757 SO THE ACCESSIBILITY OF THE 1433 00:56:49,757 --> 00:56:51,025 CONNECTION IS NOT NORMAL AND WE 1434 00:56:51,025 --> 00:56:54,195 BELIEVE THAT THIS IS IN FACT IN 1435 00:56:54,195 --> 00:56:55,629 IN THE EXPRESSION OF THE 1436 00:56:55,629 --> 00:56:55,896 CYTOKINES. 1437 00:56:55,896 --> 00:56:58,399 SO IF WE TRY TO UNDERSTAND A 1438 00:56:58,399 --> 00:57:00,134 LITTLE BIT MORE WHAT'S 1439 00:57:00,134 --> 00:57:02,670 HAPPENING, THIS IS THE SAME BUT 1440 00:57:02,670 --> 00:57:06,974 SMALLER, WE NEED TO SEEK 1441 00:57:06,974 --> 00:57:11,746 ANALYSIS FOR THE FP281 -- 1442 00:57:11,746 --> 00:57:14,181 ZFP281, THIS CAN BE DONE IN 1443 00:57:14,181 --> 00:57:14,382 VIVO. 1444 00:57:14,382 --> 00:57:17,518 WE CAN FIND BINDING TO MOST OF 1445 00:57:17,518 --> 00:57:18,819 THESE PEAKS IN FACT, THERE'S NO 1446 00:57:18,819 --> 00:57:22,757 BINDING, NO NEGATIVE CONTROL 1447 00:57:22,757 --> 00:57:22,990 BINDING. 1448 00:57:22,990 --> 00:57:26,460 THOSE PEAKS CORRESPOND TO AREAS 1449 00:57:26,460 --> 00:57:27,561 WHERE -- CAN BIND AS WELL AS 1450 00:57:27,561 --> 00:57:28,329 SHOWN HERE. 1451 00:57:28,329 --> 00:57:29,530 ONE THANK WE THOUGHT MIGHT BE 1452 00:57:29,530 --> 00:57:36,670 POSSIBLE IS THAT 281 WILL BRING 1453 00:57:36,670 --> 00:57:39,640 THAT BINDING, BUT THAT'S 1454 00:57:39,640 --> 00:57:41,675 ACTUALLY N THE CASE, AS YOU CAN 1455 00:57:41,675 --> 00:57:44,712 SEE, THE BINDING EXACTLY THE 1456 00:57:44,712 --> 00:57:46,414 SAME, WHETHER WE HAVE 281 OR 1457 00:57:46,414 --> 00:57:47,982 WHETHER WE DON'T HAVE IT. 1458 00:57:47,982 --> 00:57:49,617 SO OUR CURRENT NODLE EXPLAIN HOW 1459 00:57:49,617 --> 00:57:52,219 THIS WORKS IS THAT THOSE TWO -- 1460 00:57:52,219 --> 00:57:53,854 CURRENT NODLE EXPLAIN HOW THIS 1461 00:57:53,854 --> 00:57:55,055 WORKS, THOSE TWO OR THREE 1462 00:57:55,055 --> 00:57:57,925 FACTORS HAVE TO BIND TOGETHER TO 1463 00:57:57,925 --> 00:58:00,661 PROMOTE NORMAL CONNECTING AND 1464 00:58:00,661 --> 00:58:01,962 TH2 CYTOKINE EXPRESSION. 1465 00:58:01,962 --> 00:58:04,598 IF ONLY HERE, THE CELLS CAN 1466 00:58:04,598 --> 00:58:07,435 STILL ADOPT A TH2 FATE SO THEY 1467 00:58:07,435 --> 00:58:09,203 ARE NOT BECOMING TH1 OR ANYTHING 1468 00:58:09,203 --> 00:58:11,071 ELSE BUT THEY WILL NOT PROPERLY 1469 00:58:11,071 --> 00:58:14,241 EXPRESS THE CYTOKINES THE TH2 1470 00:58:14,241 --> 00:58:14,909 CYTOKINES. 1471 00:58:14,909 --> 00:58:16,310 ONE POSSIBILITY WE CONSIDERED 1472 00:58:16,310 --> 00:58:17,745 WAS THAT THIS WAS IN FACT A 1473 00:58:17,745 --> 00:58:20,714 GENERIC MECHANISM AND THAT EVERY 1474 00:58:20,714 --> 00:58:24,318 GENE WOULD ALSO DEPEND ON 281, 1475 00:58:24,318 --> 00:58:25,519 BUT WE DON'T THINK THAT'S THE 1476 00:58:25,519 --> 00:58:28,823 CASE BECAUSE WHEN WE LOOK AT 1477 00:58:28,823 --> 00:58:34,395 BINDING ON THE GENOME BASIS, 1478 00:58:34,395 --> 00:58:37,031 CHIPSEQ WE CAN FIND PLENTY OF 1479 00:58:37,031 --> 00:58:38,766 ZFP281 PEAKS BUT RELATIVELY FEW. 1480 00:58:38,766 --> 00:58:43,771 AND AMONG THOSE -- CONTROLLED 1481 00:58:43,771 --> 00:58:47,408 BOTH BY GATA3 AND BY ZFP281. 1482 00:58:47,408 --> 00:58:49,043 WITH THIS I'LL STOP HERE AND TO 1483 00:58:49,043 --> 00:58:53,214 SUMMARIZE WHAT I SAID, WE THINK 1484 00:58:53,214 --> 00:58:56,283 ZFP281 -- DIFFERENTIATION IN THE 1485 00:58:56,283 --> 00:58:57,918 THYMUS, THAT THOSE TWO FACTORS 1486 00:58:57,918 --> 00:59:02,356 ARE IMPORTANT FOR P THE 1487 00:59:02,356 --> 00:59:04,592 EXPRESSION OF CYTOKINES BUT WE 1488 00:59:04,592 --> 00:59:05,793 DON'T THINK THEY'RE ESSENTIAL 1489 00:59:05,793 --> 00:59:14,835 FOR T CELLS IT TO ADOPT A TH2 1490 00:59:14,835 --> 00:59:16,136 TRANSCRIPTOME THERE. 1491 00:59:16,136 --> 00:59:20,107 MAKES US SUSPECT THAT THEY 1492 00:59:20,107 --> 00:59:23,143 FUNCTION AS GATA3 COFACTORS AT 1493 00:59:23,143 --> 00:59:26,647 SPECIFIC GENE LOCI IN DEVELOPING 1494 00:59:26,647 --> 00:59:28,249 AND MATURE CD4 T CELLS. 1495 00:59:28,249 --> 00:59:31,018 THANK YOU VERY MUCH. 1496 00:59:31,018 --> 00:59:41,195 [APPLAUSE] 1497 00:59:44,732 --> 00:59:45,533 >> HELLO. 1498 00:59:45,533 --> 00:59:48,068 >> HELLO. 1499 00:59:48,068 --> 00:59:48,235 HAD. 1500 00:59:48,235 --> 00:59:49,770 >> I WILL SPEAK LOUD. 1501 00:59:49,770 --> 00:59:52,740 HAVE YOU LOOKED AT THE EFFECT OF 1502 00:59:52,740 --> 00:59:57,811 ZFP DEFICIENCY IN BINDING OF 1503 00:59:57,811 --> 01:00:00,781 FIVE IN FM CYTOKINE ORGANISMS? 1504 01:00:00,781 --> 01:00:02,283 >> YOU'RE ASKING ME WHETHER IT 1505 01:00:02,283 --> 01:00:06,220 AFFECT THE BINDING OF STAT5? 1506 01:00:06,220 --> 01:00:07,121 >> AFFECT SND. 1507 01:00:07,121 --> 01:00:10,958 >> NO, WE HAVE NOT LOOKED. 1508 01:00:10,958 --> 01:00:13,794 >> FIVE PROBABLY WITH EACH OTHER 1509 01:00:13,794 --> 01:00:16,297 TO INDUCE TH2 CYTOKINES SPHWHR 1510 01:00:16,297 --> 01:00:18,832 IT'S ABSOLUTELY POSSIBLE, BUT WE 1511 01:00:18,832 --> 01:00:20,200 HAVE NOT LOOKED INTO THAT. 1512 01:00:20,200 --> 01:00:23,871 >> CAN YOU HEAR ME, REMY? 1513 01:00:23,871 --> 01:00:24,171 >> YES. 1514 01:00:24,171 --> 01:00:25,839 >> VERY NICE TALK. 1515 01:00:25,839 --> 01:00:30,544 SO IF YOU LOOK AT TRRMS 1516 01:00:30,544 --> 01:00:32,079 CHROMATIN YOU'LL SEE THAT MASTER 1517 01:00:32,079 --> 01:00:34,148 TRANSCRIPTION FACTORS LIKE 1518 01:00:34,148 --> 01:00:35,916 GATA3, IF YOU LOOK AT THOSE LOCI 1519 01:00:35,916 --> 01:00:37,985 THAT YOU SEE BOTH POSITIVE AND 1520 01:00:37,985 --> 01:00:41,689 REPRESSIVE MARKS IN TH1 AND TH2 1521 01:00:41,689 --> 01:00:44,325 WHEREAS THE CYTOKINES ONLY SHOW 1522 01:00:44,325 --> 01:00:51,865 OPEN CHROMATIN OR H3 WHATEVER, 1523 01:00:51,865 --> 01:00:53,200 THE POSITIVE MARKS, THE 1524 01:00:53,200 --> 01:00:54,068 APPROPRIATE LINEAGE. 1525 01:00:54,068 --> 01:00:56,370 SO WHAT DO YOU SEE, HAVE YOU 1526 01:00:56,370 --> 01:01:00,841 LOOKED AT ZPF281 AND 148 IN 1527 01:01:00,841 --> 01:01:03,010 TERMS OF THEIR CHROMATIN 1528 01:01:03,010 --> 01:01:03,577 MODIFICATION? 1529 01:01:03,577 --> 01:01:06,213 >> SO WE HAVEN'T LOOKED AT 1530 01:01:06,213 --> 01:01:09,183 WHETHER -- SO THE SITE I WAS 1531 01:01:09,183 --> 01:01:14,655 POINTING AT ARE MARKS BY -- AND 1532 01:01:14,655 --> 01:01:18,492 OTHERS, BY ITS 27, SO THIS 1533 01:01:18,492 --> 01:01:21,328 ACTIVITY MARKER, AND WE HAVEN'T 1534 01:01:21,328 --> 01:01:24,531 LOOKED IF THAT IS AFFECTED BY 1535 01:01:24,531 --> 01:01:26,000 THE 281 AND 148 KNOCK-OUT, IF 1536 01:01:26,000 --> 01:01:26,967 THAT WAS YOUR QUESTION. 1537 01:01:26,967 --> 01:01:30,104 >> THANKS. 1538 01:01:30,104 --> 01:01:31,639 >> HELLO. 1539 01:01:31,639 --> 01:01:31,839 SO -- 1540 01:01:31,839 --> 01:01:33,707 >> I LIKE THAT -- 1541 01:01:33,707 --> 01:01:35,643 >> I WOULD LIKE TO ASK A 1542 01:01:35,643 --> 01:01:36,744 QUESTION ABOUT TRANSCRIPTION 1543 01:01:36,744 --> 01:01:38,946 FACTOR AND THE -- CELLS, WHY 1544 01:01:38,946 --> 01:01:40,147 SPECIFICALLY UPREGULATED IN 1545 01:01:40,147 --> 01:01:41,882 CLUSTER SELECTED NOT CLASS ONE, 1546 01:01:41,882 --> 01:01:43,317 WHAT SIGNAL INDUCES THEIR 1547 01:01:43,317 --> 01:01:47,888 EXPRESSION OR BINDING TO GATA3? 1548 01:01:47,888 --> 01:01:49,189 >> IT'S A GREAT QUESTION. 1549 01:01:49,189 --> 01:01:51,759 YOU KNOW, I THINK TO BE FAIR, IF 1550 01:01:51,759 --> 01:01:54,361 YOU ASK -- WOULD BECOME A 1551 01:01:54,361 --> 01:01:55,596 DIFFERENT ANSWER. 1552 01:01:55,596 --> 01:01:56,764 BUT I THINK THE FAIR ANSWER IS 1553 01:01:56,764 --> 01:02:00,968 THAT SO FAR, IT'S NOT ENTIRELY 1554 01:02:00,968 --> 01:02:01,168 KNOWN. 1555 01:02:01,168 --> 01:02:03,570 ONE OF THE PROBLEMS HERE IS THAT 1556 01:02:03,570 --> 01:02:06,106 THE SIGNALS THAT WOULD 1557 01:02:06,106 --> 01:02:08,709 UPREGULATE THESE FACTORS ARE 1558 01:02:08,709 --> 01:02:10,110 MISSING TO -- ARE SIMILAR TO 1559 01:02:10,110 --> 01:02:12,079 THOSE THAT DRIVE POSITIVE 1560 01:02:12,079 --> 01:02:13,280 SELECTION IN THE THYMUS SO THAT 1561 01:02:13,280 --> 01:02:18,352 IT'S GENETICALLY VERY DIFFICULT 1562 01:02:18,352 --> 01:02:19,987 FOR ONE AND THE OTHER TO TRY TO 1563 01:02:19,987 --> 01:02:25,059 ADDRESS THIS QUESTION IN THE 1564 01:02:25,059 --> 01:02:28,662 2020 STUDY TWRM WE USED SINGLE 1565 01:02:28,662 --> 01:02:30,197 CELL ATTACK AND RNA SEQUENCING 1566 01:02:30,197 --> 01:02:32,566 TO TRY TO PREDICT FACTORS THAT 1567 01:02:32,566 --> 01:02:37,071 WOULD BIND THE ZFP281 AND 148 1568 01:02:37,071 --> 01:02:38,372 WAS ONE OF THEM. 1569 01:02:38,372 --> 01:02:44,278 AND OTHER FACTORS, IT LIKE -- 1570 01:02:44,278 --> 01:02:46,547 WERE ALSO, SO IT IS POSSIBLE 1571 01:02:46,547 --> 01:02:48,549 THAT THESE FACTORS ARE INVOLVED, 1572 01:02:48,549 --> 01:02:49,650 BUT THE TRUTH OF THE MATTER IS 1573 01:02:49,650 --> 01:02:51,251 THAT WE DON'T KNOW. 1574 01:02:51,251 --> 01:02:53,454 >> OKAY. 1575 01:02:53,454 --> 01:02:53,987 THANK YOU. 1576 01:02:53,987 --> 01:02:56,523 >> WRONG ANSWER TO SAY NOTHING, 1577 01:02:56,523 --> 01:02:56,724 RIGHT? 1578 01:02:56,724 --> 01:02:59,393 >> THANKS, REMY. 1579 01:02:59,393 --> 01:03:01,895 DO YOU HAVE ANY NOAKSZ ABOUT 1580 01:03:01,895 --> 01:03:02,930 STABILITY -- DO YOU HAVE ANY 1581 01:03:02,930 --> 01:03:04,832 NOTIONS ABOUT STABILITY OF 1582 01:03:04,832 --> 01:03:05,032 GATA3? 1583 01:03:05,032 --> 01:03:09,369 YOU SHOWED THAT IT'S THERE, 1584 01:03:09,369 --> 01:03:12,873 BUT -- EVEN AT THE LOCUS, IF YOU 1585 01:03:12,873 --> 01:03:16,143 HAVE A STABLE EFFECT OF GATA3'S 1586 01:03:16,143 --> 01:03:16,410 FUNCTION. 1587 01:03:16,410 --> 01:03:18,579 SO IF YOU WOULD LOOK AT OTHER 1588 01:03:18,579 --> 01:03:20,547 KINDS THIS PERTUBATIONS WHERE 1589 01:03:20,547 --> 01:03:22,549 YOU MIGHT SEE PLASTICITY OR 1590 01:03:22,549 --> 01:03:32,359 THINGS LIKE THAT. 1591 01:03:32,359 --> 01:03:34,928 >> YOU MEAN THE STABLGHT OF 1592 01:03:34,928 --> 01:03:36,630 GATA3 TO THE LOCUS? 1593 01:03:36,630 --> 01:03:38,398 I DON'T KNOW HOW EASY THAT WOULD 1594 01:03:38,398 --> 01:03:38,699 BE. 1595 01:03:38,699 --> 01:03:44,738 WHAT I CAN SAY IS THAT IF WE DO 1596 01:03:44,738 --> 01:03:47,040 -- EXPERIMENTING WE DON'T 1597 01:03:47,040 --> 01:03:49,276 FIND A SHIFT TO TH1 OR GVMENT 1598 01:03:49,276 --> 01:03:52,479 THE CELLS REMAIN GATA3 1599 01:03:52,479 --> 01:03:54,715 EXPRESSING, THEY DON'T MAKE THE 1600 01:03:54,715 --> 01:03:56,016 CYTOKINES VERY WELL, BUT THEY 1601 01:03:56,016 --> 01:03:58,519 DON'T BECOME TH1. 1602 01:03:58,519 --> 01:04:03,490 TO SNREAS WILL NOT BECOME TH1. 1603 01:04:03,490 --> 01:04:06,160 THANK YOU VERY MUCH. 1604 01:04:06,160 --> 01:04:11,832 [APPLAUSE] 1605 01:04:11,832 --> 01:04:19,239 >> OUR NEXT SPEAK ARE IS YOUSUKE 1606 01:04:19,239 --> 01:04:22,509 TAKAHAMA, TITLE OF HIS TALK IS 1607 01:04:22,509 --> 01:04:28,649 THYMIC FORMATION OF CD8 T CELL 1608 01:04:28,649 --> 01:04:31,752 REPERTOIRE. 1609 01:04:31,752 --> 01:04:34,454 >> THANK YOU VERY MUCH. 1610 01:04:34,454 --> 01:04:41,728 MY NAME IS YOUSUKE TAKAHAMA HAD, 1611 01:04:41,728 --> 01:04:44,231 EXPERIMENTAL IMMUNOLOGY BRANCH. 1612 01:04:44,231 --> 01:04:45,866 TALKING ABOUT SELECTION OF CD8 T 1613 01:04:45,866 --> 01:04:48,068 CELLS BUT I DECIDE TO DO CHANGE 1614 01:04:48,068 --> 01:04:49,903 MY MIND. 1615 01:04:49,903 --> 01:04:55,275 BUT I DECIDED TO CHANGE MY 1616 01:04:55,275 --> 01:04:56,810 MIND. 1617 01:04:56,810 --> 01:04:58,011 DOES IT WORK? 1618 01:04:58,011 --> 01:04:58,545 THIS SORRY. 1619 01:04:58,545 --> 01:05:01,081 I WOULD LIKE TO FOCUS ON MY 1620 01:05:01,081 --> 01:05:04,384 RECENT WORK ON THE MAJORITY OF T 1621 01:05:04,384 --> 01:05:07,421 CELL 21 WHICH IS CHEMOKINE WHICH 1622 01:05:07,421 --> 01:05:09,723 IS IMPORTANT FOR T CELLS. 1623 01:05:09,723 --> 01:05:13,760 T CELL SELECTION, THYMIC 1624 01:05:13,760 --> 01:05:16,396 CONTEXT, THE CORTICAL 1625 01:05:16,396 --> 01:05:18,031 MICROENVIRONMENT, MAINLY VIA 1626 01:05:18,031 --> 01:05:23,070 CORTICAL THYMIC CELLS IN THE 1627 01:05:23,070 --> 01:05:25,739 REGION OF THYMOCYTES SELECTED 1628 01:05:25,739 --> 01:05:31,645 AND IN THIS ENVIRONMENT, SLECTDZ 1629 01:05:31,645 --> 01:05:35,449 THYMOCYTES MOVE TO THE 1630 01:05:35,449 --> 01:05:40,821 ENVIRONMENT TO ESTABLISH ARE THE 1631 01:05:40,821 --> 01:05:41,121 REPERTOIRE. 1632 01:05:41,121 --> 01:05:43,690 IN THE REGION WHAT IS -- IN THE 1633 01:05:43,690 --> 01:05:47,394 MEDULLA REGION WHAT IS IMPORTANT 1634 01:05:47,394 --> 01:05:49,796 IS THYMIC EPITHELIAL CELLS ARE 1635 01:05:49,796 --> 01:05:52,099 IMPORTANT FOR T CELLS TO 1636 01:05:52,099 --> 01:05:59,873 ESTABLISH SELF-TOLERANCE. 1637 01:05:59,873 --> 01:06:02,743 MTEC'S FOR INSTANCE IN 1638 01:06:02,743 --> 01:06:04,478 TRANSCRIPTION FACTOR CAUSE 1639 01:06:04,478 --> 01:06:06,346 SEVERE AUTISM UNITY, AND NOT 1640 01:06:06,346 --> 01:06:09,516 ONLY THAT, NUCLEAR PROTEIN 1641 01:06:09,516 --> 01:06:14,021 CALLED AIRE, AUTO IRREGULAR 1642 01:06:14,021 --> 01:06:17,724 EXPRESSED IN EPITHELIAL CELLS 1643 01:06:17,724 --> 01:06:22,963 PROMOTE GENOME-WIDE EXPRESSION, 1644 01:06:22,963 --> 01:06:27,000 CONTRIBUTING TO NEWLY GENERATED 1645 01:06:27,000 --> 01:06:28,168 T CELLS. 1646 01:06:28,168 --> 01:06:34,374 AND RECENTLY, THE IT IS 1647 01:06:34,374 --> 01:06:39,546 HIGHLIGHTED TO ENHANCE THE 1648 01:06:39,546 --> 01:06:40,514 DISPLAY. 1649 01:06:40,514 --> 01:06:43,283 ANYWAY, IT'S IMPORTANT TO 1650 01:06:43,283 --> 01:06:49,823 PROVIDE VAST MAJORITY OF 1651 01:06:49,823 --> 01:06:52,459 REMEMBER BE SELF-ANTIGENS IN THE 1652 01:06:52,459 --> 01:06:53,593 THYMUS FOR SELF-TOLL RANGS. 1653 01:06:53,593 --> 01:06:54,461 THE QUESTION THAT I WANT TO 1654 01:06:54,461 --> 01:06:56,830 ADDRESS HERE AND I HAVE BEEN 1655 01:06:56,830 --> 01:06:58,598 ADDRESSING IS HOW IT'S POSSIBLE 1656 01:06:58,598 --> 01:07:01,101 TO SET UP THE THYMOCYTES TO BE 1657 01:07:01,101 --> 01:07:03,070 GENERATED IN THE CORTEX MIGRATE 1658 01:07:03,070 --> 01:07:05,072 FROM THE CORTICAL ENVIRONMENT TO 1659 01:07:05,072 --> 01:07:06,373 THE MEDULLARY ENVIRONMENT. 1660 01:07:06,373 --> 01:07:07,541 THAT IS A QUESTION THAT I HAVE 1661 01:07:07,541 --> 01:07:13,180 BEEN ADDRESSING, AND WE HAVE 1662 01:07:13,180 --> 01:07:16,583 FOUND THAT THE ARE 7 CHEMICAL 1663 01:07:16,583 --> 01:07:18,018 RECEPTOR IS IMPORTANT TO THE 1664 01:07:18,018 --> 01:07:19,219 MIGRATION OF T CELLS AND THAT IS 1665 01:07:19,219 --> 01:07:22,923 IMPORTANT TO ESTABLISH 1666 01:07:22,923 --> 01:07:31,565 EVENTUALLY ESTABLISH CENTRAL 1667 01:07:31,565 --> 01:07:33,000 TOLERANCE, WITH SEVERAL 1668 01:07:33,000 --> 01:07:35,302 EXPERIMENTAL RESULTS, CCR7 IS 1669 01:07:35,302 --> 01:07:38,038 EXPRESSED BY MATURE MATURE 1670 01:07:38,038 --> 01:07:40,474 THYMOCYTES BUT NOT BY IMMATURE 1671 01:07:40,474 --> 01:07:44,745 THYMOCYTES AND IN IMMATURE 1672 01:07:44,745 --> 01:07:48,482 THYMOCYTES THIS IS INDUCED FOR 1673 01:07:48,482 --> 01:07:50,650 SELECTION, SIGNALS INDUCE OR 1674 01:07:50,650 --> 01:07:56,323 PROMOTE THE EXPRESSION OF CCR7. 1675 01:07:56,323 --> 01:07:58,091 MEDULLARY ACCUMULATION OF MATURE 1676 01:07:58,091 --> 01:08:01,695 THYMOCYTES IS DEFECTIVE IN CCR7 1677 01:08:01,695 --> 01:08:02,696 ANIMALS AND MORE IMPORTANTLY 1678 01:08:02,696 --> 01:08:06,967 THEY FAIL TO ESTABLISH T CELLS 1679 01:08:06,967 --> 01:08:10,437 WITHOUT T CELL 7 MEDULLARY 1680 01:08:10,437 --> 01:08:11,671 MIGRATION COULD AUTOIMMUNE 1681 01:08:11,671 --> 01:08:17,577 TISSUE LESIONS. 1682 01:08:17,577 --> 01:08:19,179 SO WHAT IS THE REASON FOR THIS? 1683 01:08:19,179 --> 01:08:20,647 THERE ARE THREE POTENTIAL 1684 01:08:20,647 --> 01:08:26,953 PROTEINS ENCODED, ONE IS CCR21 A 1685 01:08:26,953 --> 01:08:30,791 GENE, ENCODE TO CCL21 PROTEIN 1686 01:08:30,791 --> 01:08:35,529 AND MULTIPLE GENE ELM CODE IN 1687 01:08:35,529 --> 01:08:40,400 CCL21 PROTEIN, TO LOU SEEN. 1688 01:08:40,400 --> 01:08:42,502 -- TO LEUCINE. 1689 01:08:42,502 --> 01:08:45,572 THE GENE ENCODED, THE CCL19 1690 01:08:45,572 --> 01:08:46,640 PROTEIN, AND NOT ONLY THAT, BY 1691 01:08:46,640 --> 01:08:48,842 MAKING A SERIES OF KNOCK-OUT 1692 01:08:48,842 --> 01:08:51,344 MICE WE HAVE IDENTIFIED THAT 1693 01:08:51,344 --> 01:08:55,082 CCR21A ENCODED IN CCR21 1694 01:08:55,082 --> 01:08:57,150 PROTEIN -- CCL21 SPREENT MOST 1695 01:08:57,150 --> 01:08:59,786 IMPORTANT ONE IN THIS CONTEXT. 1696 01:08:59,786 --> 01:09:03,957 HOW CCL21A PROTEIN WORKS IN THIS 1697 01:09:03,957 --> 01:09:08,428 CONTEXT, IN CCL21 DEFICIENT 1698 01:09:08,428 --> 01:09:11,498 DEFICIENT MICE, IN THE MEDULLA. 1699 01:09:11,498 --> 01:09:14,334 MEDULLARY MIGRATION OF 1700 01:09:14,334 --> 01:09:17,704 THYMOCYTES DEFECTIVE IN CCL21A 1701 01:09:17,704 --> 01:09:22,876 DEFICIENT MICE, KNOCK-OUT 1702 01:09:22,876 --> 01:09:26,613 MYSELF, BUT NOT CCL19 KNOCK-OUT 1703 01:09:26,613 --> 01:09:30,317 MICE, DEFECTIVE IS THIS CCL21A 1704 01:09:30,317 --> 01:09:31,084 KNOCK-OUT MICE AND WHAT IS 1705 01:09:31,084 --> 01:09:34,254 INTERESTING HERE IS THAT CCL21A 1706 01:09:34,254 --> 01:09:35,589 EXPRESSED IN THE CELLS WITHIN 1707 01:09:35,589 --> 01:09:38,492 THE THYMUS ARE PRIMARILY 1708 01:09:38,492 --> 01:09:42,195 MEDULLARY THYMIC CELLS OR 1709 01:09:42,195 --> 01:09:42,429 MTEC'S. 1710 01:09:42,429 --> 01:09:44,164 WHAT ARE THOSE MTEC'S? 1711 01:09:44,164 --> 01:09:47,701 AS I MENTIONED THAT MTEC'S ARE 1712 01:09:47,701 --> 01:09:50,337 SO POPULAR IN TERMS DISPLAYING 1713 01:09:50,337 --> 01:09:55,142 CELL FUNCTIONS BUT NOW MTEC'S DO 1714 01:09:55,142 --> 01:09:57,310 EXPRESS CERTAIN PARTICULAR AND 1715 01:09:57,310 --> 01:10:01,281 WE FOUND THAT CCL21-EXPRESSING 1716 01:10:01,281 --> 01:10:04,784 MTEC'S WITHIN MTEC POPULATION, 1717 01:10:04,784 --> 01:10:06,119 CCL21 EXPRESSING IN THE CELLS 1718 01:10:06,119 --> 01:10:09,589 ARE LARGELY DISTINCT FROM AIRE 1719 01:10:09,589 --> 01:10:14,394 EXPRESSING CELLS WHICH IS NOT 1720 01:10:14,394 --> 01:10:16,530 JUST BY THE POPULATION 1721 01:10:16,530 --> 01:10:23,069 EXPERIMENT ANALYSIS, BUT 1722 01:10:23,069 --> 01:10:24,738 EXPRESSING THE EXPRESSION. 1723 01:10:24,738 --> 01:10:28,208 SO MTEC'S ARE NOT JUST 1724 01:10:28,208 --> 01:10:29,776 HOMOGENEOUS POPULATION TO 1725 01:10:29,776 --> 01:10:32,245 DISPLAY CELL FUNCTIONS. 1726 01:10:32,245 --> 01:10:34,681 MTEC'S INCLUDE FUNCTIONALLY 1727 01:10:34,681 --> 01:10:39,719 HETEROGENEOUS SUBSET, THAT IS 1728 01:10:39,719 --> 01:10:41,221 IMPORTANT FOR CENTRAL 1729 01:10:41,221 --> 01:10:44,891 SELF-TOLERANCE, ONE IS THE 1730 01:10:44,891 --> 01:10:48,161 FUNCTION AND THYME SIETSZ 1731 01:10:48,161 --> 01:10:50,664 ATTRACTING FMS PRODUCING CELLS. 1732 01:10:50,664 --> 01:10:51,932 I WOULD LIKE TO SHOW TODAY SOME 1733 01:10:51,932 --> 01:10:54,467 OF THE RESULTS OF HOW THOSE 1734 01:10:54,467 --> 01:10:58,205 DIFFERENT HETEROGENEOUS SUBSET 1735 01:10:58,205 --> 01:11:00,807 CONTRIBUTE TO ESTABLISH CENTRAL 1736 01:11:00,807 --> 01:11:01,374 SELF-TOLERANCE. 1737 01:11:01,374 --> 01:11:05,879 WE ACTUALLY MADE CCL21 DOUBLE 1738 01:11:05,879 --> 01:11:08,148 DEFICIENT MICE TO CHARACTERIZE 1739 01:11:08,148 --> 01:11:08,715 HERE. 1740 01:11:08,715 --> 01:11:11,451 INITIALLY WE FOUND THAT OF 1741 01:11:11,451 --> 01:11:13,753 COURSE IN DEFICIENT MICE AS HAS 1742 01:11:13,753 --> 01:11:16,690 BEEN VERY WELL KNOWN ABOUT THE 1743 01:11:16,690 --> 01:11:23,296 TISSUE LESIONS IN MULTIPLE 1744 01:11:23,296 --> 01:11:29,536 TISSUES, SO THE CCL21 MUCH MORE 1745 01:11:29,536 --> 01:11:32,539 SEVERE TISSUE LESIONS IN A 1746 01:11:32,539 --> 01:11:34,241 VARIETY OF TISSUES. 1747 01:11:34,241 --> 01:11:37,010 HERE ON THE RIGHT SIDE IS AN 1748 01:11:37,010 --> 01:11:39,079 EXAMPLE OF THE LIVER SHOWING 1749 01:11:39,079 --> 01:11:43,283 THAT AIRE DWICIALT SINGLE CELL 1750 01:11:43,283 --> 01:11:46,219 KNOCK-OUT, CCL21 KNOCK-OUT MICE 1751 01:11:46,219 --> 01:11:47,854 SHOWED MINOR TISSUE LESIONS BUT 1752 01:11:47,854 --> 01:11:49,623 DOUBLE KNOCK-OUT MICE SHOWED 1753 01:11:49,623 --> 01:11:51,391 QUITE SEVERE TISSUE ISSUES. 1754 01:11:51,391 --> 01:11:54,794 AND THOSE TISSUE LESIONS IS NOT 1755 01:11:54,794 --> 01:12:00,900 ONLY TISSUE PROBLEMS BUT ALSO 1756 01:12:00,900 --> 01:12:02,469 THE LIVER ENZYMES TO THE 1757 01:12:02,469 --> 01:12:05,705 BLOODSTREAM AND ALSO THE 1758 01:12:05,705 --> 01:12:09,442 OBSTRUCTION CAWDZED HERE 1759 01:12:09,442 --> 01:12:12,479 SPLENOMEGALY, NOT JUST LIMITED 1760 01:12:12,479 --> 01:12:14,781 TO THE LIVER BUT PRODUCING THE 1761 01:12:14,781 --> 01:12:21,955 ORGAN IS ALSO SEVERE IN HERE, IN 1762 01:12:21,955 --> 01:12:27,027 T CELL IT CCL21 DEFICIENT MICE. 1763 01:12:27,027 --> 01:12:30,764 HOWEVER, CCL21 EXPRESSED 1764 01:12:30,764 --> 01:12:35,669 SPECIFICALLY IN UNTIL MTEC'S BUT 1765 01:12:35,669 --> 01:12:37,537 THE EXPRESSION OF THOSE ARE NOT 1766 01:12:37,537 --> 01:12:40,273 FLIMENTD THYMUS, FOR EXAMPLE, 1767 01:12:40,273 --> 01:12:44,110 THE EXTRA THYMIC EXPRESSED IN 1768 01:12:44,110 --> 01:12:47,247 THE CELLS CONTRIBUTING TO THE 1769 01:12:47,247 --> 01:12:47,614 SELF-TOLERANCE. 1770 01:12:47,614 --> 01:12:51,518 SO DO THOSE MTEC'S COOPERATE TO 1771 01:12:51,518 --> 01:12:53,520 PREVENT THE TISSUE LESIONS? 1772 01:12:53,520 --> 01:13:02,595 TO ADDRESS IT, THE THYMUS IT 1773 01:13:02,595 --> 01:13:09,302 TRANSPLANTATION INTO AM ATHYMIC 1774 01:13:09,302 --> 01:13:11,404 MICE FRMS NUDE MICE TO 1775 01:13:11,404 --> 01:13:16,643 RECONSTITUTE T CELL DEVELOPMENT, 1776 01:13:16,643 --> 01:13:20,213 TO FIND THAT THE LOSS OF THOSE 1777 01:13:20,213 --> 01:13:24,818 TWO POPULATIONS CAWDZED MOST 1778 01:13:24,818 --> 01:13:31,524 SEVERE AUTOIMMUNE DISEASE TR, 1779 01:13:31,524 --> 01:13:36,863 EXPRESSING MTEC'S IT, WHICH IS 1780 01:13:36,863 --> 01:13:40,834 CLEAR AND ALSO THE SCORING OF 1781 01:13:40,834 --> 01:13:49,476 THE IN VIVO LUNG, SO ARE SHOWING 1782 01:13:49,476 --> 01:13:55,715 MUCH MORE SEVERE PROBLEMS IN THE 1783 01:13:55,715 --> 01:13:57,016 BODIES. 1784 01:13:57,016 --> 01:14:00,086 SO THIS RESULT SHOWS THAT LIKE 1785 01:14:00,086 --> 01:14:03,123 AIRE POSITIVE ARE MTEC'S 1786 01:14:03,123 --> 01:14:04,657 CONTRIBUTE TO SELF-TOLERANCE, 1787 01:14:04,657 --> 01:14:07,594 AND MORE IMPORTANTLY, THOSE TWO 1788 01:14:07,594 --> 01:14:12,999 DIFFERENT SUBSETS COOMENT TO -- 1789 01:14:12,999 --> 01:14:16,269 COOPERATE TO ESTABLISH CENTRAL 1790 01:14:16,269 --> 01:14:16,536 TOLERANCE. 1791 01:14:16,536 --> 01:14:18,972 WHAT IS MOST INTERESTING WITH 1792 01:14:18,972 --> 01:14:20,940 THAT, THAT COOPERATION SEEMS TO 1793 01:14:20,940 --> 01:14:25,545 BE ATTRIBUTED ON THE LOSS OF 1794 01:14:25,545 --> 01:14:26,846 ANTI-MEDULLARY REGION WHICH IS 1795 01:14:26,846 --> 01:14:30,150 WHAT'S SHOWN HERE ON THE RIGHT 1796 01:14:30,150 --> 01:14:36,389 SIDE BY RELB KNOCK-OUT MICE, 1797 01:14:36,389 --> 01:14:37,257 APPROXIMATE SHOWING SIGNIFICANT 1798 01:14:37,257 --> 01:14:43,930 PROBLEMS WITH THEIR AIRE CCL21 1799 01:14:43,930 --> 01:14:45,365 THYMIC ENVIRONMENT. 1800 01:14:45,365 --> 01:14:47,200 SO THE COMBINATION RESULTS IN 1801 01:14:47,200 --> 01:14:51,704 DIFFERENT SUBSET, FUNCTIONALLY 1802 01:14:51,704 --> 01:14:58,378 EQUIVALENT TO MTEC TOLERANCE. 1803 01:14:58,378 --> 01:14:59,813 HOW DO THEY COOPERATE? 1804 01:14:59,813 --> 01:15:04,184 DO THEY CONTRIBUTE TO ELIMINATE 1805 01:15:04,184 --> 01:15:05,552 LY THE REGULATORY T 1806 01:15:05,552 --> 01:15:08,021 CELLS AND FOR THE ELIMINATION OF 1807 01:15:08,021 --> 01:15:10,990 A SENSE OF REACTIVITY AS WE 1808 01:15:10,990 --> 01:15:14,828 MEASURE THE NUMBER OF THE TWO 1809 01:15:14,828 --> 01:15:19,098 DIFFERENT IT SELF-REACTIVE T 1810 01:15:19,098 --> 01:15:26,506 CELLS TO REACTIVE TO THOSE CELL 1811 01:15:26,506 --> 01:15:33,980 FUNCTIONS IN THE ACTIVATION OF 1812 01:15:33,980 --> 01:15:35,715 SELF-REACTIVE T CELLS. 1813 01:15:35,715 --> 01:15:38,651 ALSO HAD IN THE DEVELOPMENT OF 1814 01:15:38,651 --> 01:15:39,886 REGULATORY T CELLS FROM THE 1815 01:15:39,886 --> 01:15:42,155 MATURE STATE -- IMMATURE STAGE 1816 01:15:42,155 --> 01:15:44,257 TO THE MATURE STAGE, DOUBLE 1817 01:15:44,257 --> 01:15:46,759 DEFICIENT ANIMALS SHOW MUCH MORE 1818 01:15:46,759 --> 01:15:50,129 SEVERE PROBLEMS THAN THE SINGLE 1819 01:15:50,129 --> 01:15:52,765 POSITIVE HAD SINGLE DEFICIENT 1820 01:15:52,765 --> 01:15:53,032 ANIMALS. 1821 01:15:53,032 --> 01:15:57,136 SO THE CONCLUSION HERE IS THAT 1822 01:15:57,136 --> 01:16:01,307 AIRE DEFICIENT MTEC'S AND CCL21 1823 01:16:01,307 --> 01:16:03,910 POSITIVE COOPERATE TO ESTABLISH 1824 01:16:03,910 --> 01:16:07,847 SELF-TOLERANCE BY OPTIMIZING 1825 01:16:07,847 --> 01:16:11,484 NEGATIVE SELECTION AND TREG 1826 01:16:11,484 --> 01:16:11,951 GENERATION. 1827 01:16:11,951 --> 01:16:14,787 SO THEY COOPERATE, HIGHLIGHTING 1828 01:16:14,787 --> 01:16:15,989 THE FUNCTIONAL IMPORTANCE OF 1829 01:16:15,989 --> 01:16:18,958 THOSE CHEMICAL IN CCL21 1830 01:16:18,958 --> 01:16:24,097 EXPRESSING MTEC'S IN CENTRAL 1831 01:16:24,097 --> 01:16:24,497 TOLERANCE. 1832 01:16:24,497 --> 01:16:25,732 IN THE LAST FEW MINUTES I WOULD 1833 01:16:25,732 --> 01:16:27,367 LIKE TO ADD THAT THE RECENT 1834 01:16:27,367 --> 01:16:30,770 RESULTS OF SHOWING HOW THOSE, TO 1835 01:16:30,770 --> 01:16:33,940 ANALYZE HOW CCL21 THYMOCYTES ARE 1836 01:16:33,940 --> 01:16:36,342 INTERACTING WITH MTEC'S CAN BE 1837 01:16:36,342 --> 01:16:42,015 INTEGRATED INTO THE DEVELOPMENT 1838 01:16:42,015 --> 01:16:44,317 OF WELL ESTABLISHED FOR THE 1839 01:16:44,317 --> 01:16:46,386 DEVELOPMENT OF SELF-ANTIGEN 1840 01:16:46,386 --> 01:16:50,390 DISPLAYING MT HE C'S, THEN TO 1841 01:16:50,390 --> 01:16:53,026 BECOME THE POST AIR HE 1842 01:16:53,026 --> 01:16:53,359 POPULATION. 1843 01:16:53,359 --> 01:16:57,163 HOW DO THOSE CCL21 POSITIVE 1844 01:16:57,163 --> 01:17:00,867 CELLS COULD BE INTEGRATED INTO 1845 01:17:00,867 --> 01:17:04,170 THIS AIRE MTEC PATHWAY? 1846 01:17:04,170 --> 01:17:07,440 TO ADDRESS IT, WE CREATED TWO 1847 01:17:07,440 --> 01:17:09,409 DIFFERENT IT MICE TO MEASURE THE 1848 01:17:09,409 --> 01:17:15,748 PRESENT EXPRESSION OF CCL21 GENE 1849 01:17:15,748 --> 01:17:20,019 AND PAST CCL21 GENE BY CROSSING 1850 01:17:20,019 --> 01:17:29,529 CCL21 MICE WITH LOXP ANIMALS, SO 1851 01:17:29,529 --> 01:17:32,899 IN THE MICE YOU CAN MEASURE THE 1852 01:17:32,899 --> 01:17:36,970 PRESENT EXPRESSION IN THE MICE, 1853 01:17:36,970 --> 01:17:39,439 PRESENT AND PAST EXPRESSION OF 1854 01:17:39,439 --> 01:17:40,206 CCL21 GENE. 1855 01:17:40,206 --> 01:17:47,680 TO FIND THAT, CCL21A ATDTOMATO 1856 01:17:47,680 --> 01:17:52,418 EXPRESSED BY 40% MTEC'S WHICH IS 1857 01:17:52,418 --> 01:17:57,256 DIFFERENT FROM AIRE POSITIVE 1858 01:17:57,256 --> 01:18:04,364 CELLS, OKAY, THIS IS THE 1859 01:18:04,364 --> 01:18:06,733 PROTOTYPIC CAN AIRE, THEY ARE 1860 01:18:06,733 --> 01:18:09,002 DIFFERENT CELLS, THE CELLS ARE 1861 01:18:09,002 --> 01:18:12,538 DIFFERENT, BUT CCL21 CRE SHOWS 1862 01:18:12,538 --> 01:18:15,742 THAT THE VAST MAJORITY OF MTEC'S 1863 01:18:15,742 --> 01:18:18,811 ARE LABELED BY EGFP INCLUDING 1864 01:18:18,811 --> 01:18:25,551 THE AIRE POSITIVE CELLS ANDLY -- 1865 01:18:25,551 --> 01:18:28,454 POSITIVE CELLS. 1866 01:18:28,454 --> 01:18:35,628 SO DERIVED FROM CCL21 1867 01:18:35,628 --> 01:18:36,295 TRANSCRIBING CELLS. 1868 01:18:36,295 --> 01:18:37,497 THOSE ARE JUST GENE EXPRESSION 1869 01:18:37,497 --> 01:18:40,867 OF CCL21 LOCUS, THE PROTEIN 1870 01:18:40,867 --> 01:18:43,403 PRODUCED IN THYMOCYTES ARE 1871 01:18:43,403 --> 01:18:45,438 INTERACTING FUNCTIONAL INDEX 1872 01:18:45,438 --> 01:18:47,206 COULD RETAIN THAT CAPABILITY TO 1873 01:18:47,206 --> 01:18:49,509 GIVE RISE TO SELF-ANTIGEN 1874 01:18:49,509 --> 01:18:51,411 DISPLAYING MTEC'S? 1875 01:18:51,411 --> 01:18:53,379 TO ANALYZE THAT, WE 1876 01:18:53,379 --> 01:18:58,317 RECONSTITUTED CCR21 PROTEIN 1877 01:18:58,317 --> 01:19:03,423 POSITIVE ARE TDTOMATO RE: 1878 01:19:03,423 --> 01:19:06,492 AGGREGATED WITH DEFICIENT IT -- 1879 01:19:06,492 --> 01:19:10,196 TO MAKE REAGGREGATED THYMUS TO 1880 01:19:10,196 --> 01:19:15,268 TRANSPLANT INTO THE KIDNEY 1881 01:19:15,268 --> 01:19:15,568 CAPSULE. 1882 01:19:15,568 --> 01:19:21,007 SOME FIVE WEEKS LATER WE FOUND 1883 01:19:21,007 --> 01:19:28,214 THAT THYMUS REAGGREGATED WITH 1884 01:19:28,214 --> 01:19:31,517 TDTOMATO. 1885 01:19:31,517 --> 01:19:34,654 SHOWING THE DEVELOPMENT OF IT 1886 01:19:34,654 --> 01:19:37,724 AIRE POSITIVE CELLS, THAT'S 1887 01:19:37,724 --> 01:19:40,026 POSITIVE, AND ALSO THE MAJORITY 1888 01:19:40,026 --> 01:19:41,427 REAGENT IS DEVELOPED. 1889 01:19:41,427 --> 01:19:44,063 SO TO FINALIZE MY TALK, I JUST 1890 01:19:44,063 --> 01:19:47,433 WOULD LIKE TO GIVE YOU A 1891 01:19:47,433 --> 01:19:52,038 CONCLUSION THAT T CELL IF -- 1892 01:19:52,038 --> 01:19:56,743 THAT CCL21 POSITIVE SLS ATTAIN 1893 01:19:56,743 --> 01:19:58,177 DEVELOPMENT THE CAPABILITY TO 1894 01:19:58,177 --> 01:20:02,482 DIFFERENTIATE INTO AIRE POSITIVE 1895 01:20:02,482 --> 01:20:02,815 MTEC'S. 1896 01:20:02,815 --> 01:20:03,783 SO ONE OF THE BIGGEST 1897 01:20:03,783 --> 01:20:07,620 CONCLUSIONS HERE IS THAT CCL21 1898 01:20:07,620 --> 01:20:08,654 POSITIVE FUNCTIONAL INDEX 1899 01:20:08,654 --> 01:20:10,690 INCLUDE THE CAPABILITY TO 1900 01:20:10,690 --> 01:20:12,892 FUNCTIONALLY CONVERT FROM CCL21 1901 01:20:12,892 --> 01:20:15,628 POSITIVE CELLS TO AIRE POSITIVE 1902 01:20:15,628 --> 01:20:19,465 CELLS TO ESTABLISH CENTRAL 1903 01:20:19,465 --> 01:20:21,367 TOLERANCE AND THAT IS IMPORTANT, 1904 01:20:21,367 --> 01:20:23,169 WE BELIEVE, TO ASSEMBLE THE 1905 01:20:23,169 --> 01:20:25,438 FUNCTIONAL DIVERSITY IN THE 1906 01:20:25,438 --> 01:20:31,244 THYMUS MEDULLA. 1907 01:20:31,244 --> 01:20:33,780 LY THANK YOU VERY MUCH FOR YOUR 1908 01:20:33,780 --> 01:20:35,848 ATTENTION. 1909 01:20:35,848 --> 01:20:42,855 [APPLAUSE] 1910 01:20:42,855 --> 01:20:45,024 SORRY, FMS I SHOULD THANK MY 1911 01:20:45,024 --> 01:20:45,358 COLLABORATORS. 1912 01:20:45,358 --> 01:20:49,061 THIS STURDY WAS INITIATED IN 1913 01:20:49,061 --> 01:20:57,370 JAPAN, ANDLY BY MY TEAM. 1914 01:20:57,370 --> 01:20:59,238 >> I HAVE A QUESTION ON WHAT THE 1915 01:20:59,238 --> 01:21:00,573 FUNCTION OF THIS. 1916 01:21:00,573 --> 01:21:01,574 SO DO YOU THINK THAT THE -- 1917 01:21:01,574 --> 01:21:02,308 >> SPEAK UP. 1918 01:21:02,308 --> 01:21:03,276 I CANNOT HEAR YOU MUCH. 1919 01:21:03,276 --> 01:21:05,478 >> IS THIS BETTER? 1920 01:21:05,478 --> 01:21:07,613 SO DO YOU THINK THAT THE 21 1921 01:21:07,613 --> 01:21:10,516 EXPRESSING MTECS ARE 1922 01:21:10,516 --> 01:21:12,051 FUNCTIONALLY THERE TO BRING 1923 01:21:12,051 --> 01:21:14,887 THYMOCYTES INTO A POCKET OF 1924 01:21:14,887 --> 01:21:16,222 OTHER PRESENTING CELLS THAT ARE 1925 01:21:16,222 --> 01:21:17,857 GOING TO ACTUALLY DO THE 1926 01:21:17,857 --> 01:21:18,758 TOLERANCE INDUCTION, OR DO YOU 1927 01:21:18,758 --> 01:21:21,260 THINK THAT THE ANTIGENS 1928 01:21:21,260 --> 01:21:22,128 PRESENTED ARE -- 1929 01:21:22,128 --> 01:21:23,296 >> THAT'S A VERY GOOD QUESTION. 1930 01:21:23,296 --> 01:21:25,565 ONE THING THAT WE HAVE RECENTLY 1931 01:21:25,565 --> 01:21:29,468 NOTICED FROM P -- FROM THE 1932 01:21:29,468 --> 01:21:30,803 SINGLE TYPE OF EXPERIMENT IS 1933 01:21:30,803 --> 01:21:37,243 THAT THEY DO EXPRESS A TYPICAL 1934 01:21:37,243 --> 01:21:39,645 CELL FUNCTION CALLED C-REACTIVE 1935 01:21:39,645 --> 01:21:40,780 PROTEIN. 1936 01:21:40,780 --> 01:21:43,049 CCL21 POSITIVE CELLS DO EXPRESS 1937 01:21:43,049 --> 01:21:45,785 OR THE MAJOR SOURCE FOR THE 1938 01:21:45,785 --> 01:21:53,726 C-REACTIVE PROTEIN, WHICH IS 1939 01:21:53,726 --> 01:21:53,960 QUITE -- 1940 01:21:53,960 --> 01:21:54,227 [LAUGHTER] 1941 01:21:54,227 --> 01:21:54,427 WHAT? 1942 01:21:54,427 --> 01:21:54,594 OH? 1943 01:21:54,594 --> 01:21:56,028 YES. 1944 01:21:56,028 --> 01:21:59,832 ANYWAY, SO THEY COULD PROVIDE 1945 01:21:59,832 --> 01:22:03,135 SOME SELF-SO-CALLED TISSUE 1946 01:22:03,135 --> 01:22:05,104 RESECTOR CELL FUNCTIONS INTO THE 1947 01:22:05,104 --> 01:22:06,305 MEDULLARY REGION. 1948 01:22:06,305 --> 01:22:07,840 BUT WE BELIEVE THAT THEIR MAJOR 1949 01:22:07,840 --> 01:22:10,009 JOB IS TO ATTRACT AND ACCUMULATE 1950 01:22:10,009 --> 01:22:14,914 AND PERHAPS HOLD THOSE 1951 01:22:14,914 --> 01:22:17,116 DEVELOPING THYMOCYTES FOR THEM 1952 01:22:17,116 --> 01:22:19,752 TO ESTABLISH OR FOR THEM TO 1953 01:22:19,752 --> 01:22:22,255 INTERACT WITH A VAST MAJORITY OF 1954 01:22:22,255 --> 01:22:26,092 AIRE POSITIVE OR THYMIC CELL 1955 01:22:26,092 --> 01:22:27,526 FUNCTIONS PROVIDED BY OTHER 1956 01:22:27,526 --> 01:22:29,028 POPULATIONS IN MT HE C'S. 1957 01:22:29,028 --> 01:22:29,562 >> THANK YOU. 1958 01:22:29,562 --> 01:22:33,065 >> THANK YOU. 1959 01:22:33,065 --> 01:22:37,603 >> REMY WAS FIRST BUT HE'S BEING 1960 01:22:37,603 --> 01:22:39,972 ULTRA CHIVALROUS. 1961 01:22:39,972 --> 01:22:41,407 I WANT TO ASK YOU ABOUT AGING. 1962 01:22:41,407 --> 01:22:44,710 WHAT HAPPENS TO WANT CCL21 1963 01:22:44,710 --> 01:22:46,479 POSITIVITY AS THE ANIMAL 1964 01:22:46,479 --> 01:22:47,980 EXPAIDGES BY EXTENSION PEOPLE 1965 01:22:47,980 --> 01:22:48,881 LIKE OURSELVES? 1966 01:22:48,881 --> 01:22:52,151 >> THAT'S, WELL, I WOULD BE 1967 01:22:52,151 --> 01:22:55,888 HAPPY TO EXAMINE CCL21 1968 01:22:55,888 --> 01:22:57,290 EXPRESSION, BUT, YOU KNOW, 1969 01:22:57,290 --> 01:22:59,158 THAT'S A GREAT QUESTION. 1970 01:22:59,158 --> 01:22:59,926 WE DON'T KNOW. 1971 01:22:59,926 --> 01:23:04,497 I MEAN, I DON'T KNOW MUCH. 1972 01:23:04,497 --> 01:23:07,600 DEPENDING ON -- SO WE HAVEN'T 1973 01:23:07,600 --> 01:23:11,938 EXAMINED ANYTHING MORE THAN DID 1974 01:23:11,938 --> 01:23:15,574 HOW YEAR OLD OF AGE IN ANIMALS, 1975 01:23:15,574 --> 01:23:18,077 SO AS WELL AS WE TALK ABOUT 1976 01:23:18,077 --> 01:23:21,247 MIDDLE-AGED ANIMALS, WE DON'T 1977 01:23:21,247 --> 01:23:24,083 SEE MUCH OF THE REDUCTION IN 1978 01:23:24,083 --> 01:23:26,719 AIRE POSITIVE CELLS OR CCL21 1979 01:23:26,719 --> 01:23:28,254 POSITIVE CELLS, BUT I UNDERSTAND 1980 01:23:28,254 --> 01:23:32,224 THAT THE AIRE, SHOWING THAT THE 1981 01:23:32,224 --> 01:23:37,063 EXPRESSION OF AIRE IS QUITE 1982 01:23:37,063 --> 01:23:40,199 IMPORTANT AT THE PERINATAL 1983 01:23:40,199 --> 01:23:41,667 STAGE -- LATER LIFE. 1984 01:23:41,667 --> 01:23:44,937 MAYBE FOR MATURE T CELLS FOR 1985 01:23:44,937 --> 01:23:47,673 THEIR DEVELOPMENT, PERINATAL 1986 01:23:47,673 --> 01:23:48,441 DEVELOPMENT, MAYBE THE KEY 1987 01:23:48,441 --> 01:23:50,843 PERIOD FOR THEM TO ESTABLISH 1988 01:23:50,843 --> 01:23:51,210 SELF-TOLERANCE. 1989 01:23:51,210 --> 01:23:54,447 IF THAT IS THE CASE, CCL21 IN 1990 01:23:54,447 --> 01:23:58,117 THE THYMUS MAY BE MOST IMPORTANT 1991 01:23:58,117 --> 01:24:02,888 IN EARLY STAGE OR OF THE THYMUS 1992 01:24:02,888 --> 01:24:04,090 DEVELOPMENT PERINATALLY. 1993 01:24:04,090 --> 01:24:05,558 >> I'M JUST THINKING ABOUT THE 1994 01:24:05,558 --> 01:24:07,360 DEFECTS IN TOLERANCE ESTABLISHED 1995 01:24:07,360 --> 01:24:13,165 IN ADULTS WHEN THEY HAVE 1996 01:24:13,165 --> 01:24:16,469 TRANSPLANTATION AND IF THE T 1997 01:24:16,469 --> 01:24:22,808 CELLS NEWLY DEVELOPED GET GET TF 1998 01:24:22,808 --> 01:24:27,279 MEDULLA BECAUSE OF T CELLS BE 1999 01:24:27,279 --> 01:24:27,680 COULDN'T DEVELOP. 2000 01:24:27,680 --> 01:24:28,948 >> THAT'S EXIENTING TOPIC. 2001 01:24:28,948 --> 01:24:31,684 >> I'LL ASK -- SO ASKED ALL THE 2002 01:24:31,684 --> 01:24:33,753 QUESTIONS I WANTED TO ASK BUT 2003 01:24:33,753 --> 01:24:35,755 ONE THEY DIDN'T HAVE, THAT IS IF 2004 01:24:35,755 --> 01:24:37,456 YOU YOU'VE ALSO LOOKED AT ANY -- 2005 01:24:37,456 --> 01:24:38,324 SETTINGS AND YOU KNOW WHAT 2006 01:24:38,324 --> 01:24:41,060 HAPPENS DURING TIMES OF 2007 01:24:41,060 --> 01:24:45,031 REGENERATION WITH THESE 2008 01:24:45,031 --> 01:24:46,632 DIFFERENT -- 2009 01:24:46,632 --> 01:24:47,900 >> THAT'S A GREAT QUESTION. 2010 01:24:47,900 --> 01:24:50,403 WE HAVE NO ANSWERS ACTUALLY TO 2011 01:24:50,403 --> 01:24:54,206 YOU, BUT -- I MIDDLE EASTERN, TO 2012 01:24:54,206 --> 01:24:55,508 ANYBODY, BUT WE'LL BE HAPPY TO 2013 01:24:55,508 --> 01:24:58,044 LEARN MORE ABOUT THE REGION AND 2014 01:24:58,044 --> 01:25:00,446 PROCESS IN TERMS OF THOSE 2015 01:25:00,446 --> 01:25:04,116 CHEMOKINE OR CHEMOKINE RECEPTOR 2016 01:25:04,116 --> 01:25:04,650 INTERACTION. 2017 01:25:04,650 --> 01:25:04,950 THANK YOU. 2018 01:25:04,950 --> 01:25:06,185 >> GREAT. 2019 01:25:06,185 --> 01:25:06,485 THANK YOU. 2020 01:25:06,485 --> 01:25:09,655 >> IS THERE ANY TRANSCRIPTION 2021 01:25:09,655 --> 01:25:19,598 SIMILARITY WITH FRC EXPRESSED -- 2022 01:25:19,598 --> 01:25:22,334 >> YES, COMMON CHARACTER, BUT 2023 01:25:22,334 --> 01:25:25,171 THERE WHAT IS MOST IMPORTANT IS 2024 01:25:25,171 --> 01:25:30,209 THAT FRC'S IN LYMPH NODE THEY DO 2025 01:25:30,209 --> 01:25:33,045 PRODUCE A LOT OF CHEMOKINE 2026 01:25:33,045 --> 01:25:36,982 CALLED CCL19, WHICH FROM THE 2027 01:25:36,982 --> 01:25:40,052 COMPARISON OF KNOCK-OUT MICE IN 2028 01:25:40,052 --> 01:25:43,489 ANIMALS CCL19 SEEMS TO PLAY MUCH 2029 01:25:43,489 --> 01:25:48,627 MORE IMPORTANT ROLE THAN CCL21 2030 01:25:48,627 --> 01:25:55,634 IN MAJORITY OF LYMPH NODE. 2031 01:25:55,634 --> 01:25:56,902 SO DEPENDING ON THE ORGANS. 2032 01:25:56,902 --> 01:25:58,804 >> JUST A QUICK QUESTION. 2033 01:25:58,804 --> 01:26:01,774 SO TRANSCRIPTIONALLY ALL THE 2034 01:26:01,774 --> 01:26:03,409 CCL21-EXPRESSING CELLS, ALL THE 2035 01:26:03,409 --> 01:26:05,578 CELLS EXPRESS CCL21 AT SOME 2036 01:26:05,578 --> 01:26:06,879 POINT AND EVENTUALLY SOME OF 2037 01:26:06,879 --> 01:26:08,414 THESE WILL BECOME AIRE 2038 01:26:08,414 --> 01:26:09,081 EXPRESSING CELLS. 2039 01:26:09,081 --> 01:26:10,382 WHAT TRIGGERS THIS CONVERSION? 2040 01:26:10,382 --> 01:26:11,784 IS THERE ANYTHING SPECIFIC? 2041 01:26:11,784 --> 01:26:12,751 WHY WOULD THERE BECOME -- 2042 01:26:12,751 --> 01:26:15,855 >> WHAT TRIGGERS THE CONVERSION? 2043 01:26:15,855 --> 01:26:18,357 YEAH, EXCELLENT QUESTION FROM 2044 01:26:18,357 --> 01:26:22,094 REVIEW OF NUMBER ONE. 2045 01:26:22,094 --> 01:26:25,731 [LAUGHTER] 2046 01:26:25,731 --> 01:26:26,565 WE DON'T KNOW YET. 2047 01:26:26,565 --> 01:26:27,900 WE WOULD LOVE TO LEARN HOW THIS 2048 01:26:27,900 --> 01:26:28,767 CONVERSION OR WHATEVER THE 2049 01:26:28,767 --> 01:26:29,602 DEVELOPMENT PROCESS IS OPERATED. 2050 01:26:29,602 --> 01:26:40,079 THANK YOU FOR YOUR INTEREST. 2051 01:26:41,780 --> 01:26:49,455 APPLAUSE PLU. 2052 01:26:49,455 --> 01:26:50,089 >>> 2053 01:26:50,089 --> 01:26:50,356 [APPLAUSE] 2054 01:26:50,356 --> 01:26:54,460 >> OUR NEXT SPEAKER IS JONATHAN 2055 01:26:54,460 --> 01:26:58,297 ASHWELL AND HE WILL TALK ABOUT 2056 01:26:58,297 --> 01:27:08,807 GLUCOCORTICOIDS IN THE THYMUS. 2057 01:27:52,418 --> 01:27:53,652 I'VE GOT IT. 2058 01:27:53,652 --> 01:27:53,852 OKAY. 2059 01:27:53,852 --> 01:27:56,422 THANK YOU. 2060 01:27:56,422 --> 01:28:00,125 OKAY H WELCOME. 2061 01:28:00,125 --> 01:28:02,661 THINGS ARE RUNNING A LITTLE 2062 01:28:02,661 --> 01:28:02,861 LATE. 2063 01:28:02,861 --> 01:28:04,997 A BREAK IS NEXT, SO THERE'S 2064 01:28:04,997 --> 01:28:06,131 RELIEF COMING. 2065 01:28:06,131 --> 01:28:08,901 I'LL BE TALKING ABOUT THE 2066 01:28:08,901 --> 01:28:10,436 ANATOMY OF THE -- SORRY. 2067 01:28:10,436 --> 01:28:12,171 THE ANATOMY, REGULATION AND 2068 01:28:12,171 --> 01:28:14,340 FUNCTION OF GLUCOCORTICOIDS 2069 01:28:14,340 --> 01:28:16,875 PRODUCED IN THE THYMUS, AND I'LL 2070 01:28:16,875 --> 01:28:18,210 START LIKE EVERYBODY ELSE BY 2071 01:28:18,210 --> 01:28:20,913 REVIEWING VERY QUICKLY THYMUS 2072 01:28:20,913 --> 01:28:23,215 SELECTION -- THYMOCYTES 2073 01:28:23,215 --> 01:28:24,083 SELECTION AND THIS IS OUR REVIEW 2074 01:28:24,083 --> 01:28:26,619 OF IT, MOST PEOPLE'S REVIEW OF 2075 01:28:26,619 --> 01:28:29,788 IT, DURING EVICTION -- IGNORING 2076 01:28:29,788 --> 01:28:31,223 EVICTION, THAT THIS SHOWS THE 2077 01:28:31,223 --> 01:28:32,091 RELATIONSHIP BETWEEN THE 2078 01:28:32,091 --> 01:28:34,526 AFFINITY OF T CELL RECEPTORS 2079 01:28:34,526 --> 01:28:36,362 THAT ARE BETA RECEPTORS 2080 01:28:36,362 --> 01:28:37,329 GENERATED IN THE THIEMTION AND 2081 01:28:37,329 --> 01:28:40,633 THE FATE OF THE CELLS. 2082 01:28:40,633 --> 01:28:43,802 THE NOTION IS SHOWN HERE 2083 01:28:43,802 --> 01:28:44,770 SCHEMATICALLY ON THE RIGHT THAT 2084 01:28:44,770 --> 01:28:47,740 THOSE CELLS THAT HAVE 2085 01:28:47,740 --> 01:28:49,942 SUBTHRESHOLD AFFINITY FOR 2086 01:28:49,942 --> 01:28:54,747 SELF-PEPTIDE MHC, DEATH BY 2087 01:28:54,747 --> 01:28:57,149 NEGLECT, THE THYMOCYTES 2088 01:28:57,149 --> 01:28:58,651 ALPHA-BETA RECEPTORS WITH VERY 2089 01:28:58,651 --> 01:29:01,287 HIGH AFFINITY FOR SELF AND ARE 2090 01:29:01,287 --> 01:29:02,921 POTENTIALLY DANGEROUS ARE 2091 01:29:02,921 --> 01:29:05,024 ELIMINATED VIA AN ACTIVE PATHWAY 2092 01:29:05,024 --> 01:29:10,596 THAT INDUCES APOPTOSIS, 2093 01:29:10,596 --> 01:29:11,697 SELECTION, CELLS SHOWN IN GREEN 2094 01:29:11,697 --> 01:29:14,333 THAT THREE OR FOUR PERCENT OF 2095 01:29:14,333 --> 01:29:19,471 CELLS IN THE THYMUS THAT HAVE -- 2096 01:29:19,471 --> 01:29:21,040 AND THAT'S POSITIVE SELECTION. 2097 01:29:21,040 --> 01:29:22,775 THESE CELLS GO ON TO THE 2098 01:29:22,775 --> 01:29:24,510 PERIPHERY WHERE THEY FORM THE T 2099 01:29:24,510 --> 01:29:27,346 CELL REPERTOIRE, THAT IS THE 2100 01:29:27,346 --> 01:29:28,614 UNIVERSE ADVANTAGE TO WHICH THE 2101 01:29:28,614 --> 01:29:30,783 T CELLS CAN RESPOND. 2102 01:29:30,783 --> 01:29:32,851 NOW, OTHER STORY GOES BACK EVEN 2103 01:29:32,851 --> 01:29:35,888 FURTHER THAN AL'S STORY TO THE 2104 01:29:35,888 --> 01:29:37,523 1990S, WHERE WE HAD DISCOVERED 2105 01:29:37,523 --> 01:29:41,994 THAT ACTIVATION DUE TO T CELL 2106 01:29:41,994 --> 01:29:42,961 APOPTOSIS AND WE HAD THOUGHT AT 2107 01:29:42,961 --> 01:29:44,196 THE TIME THAT THIS MIGHT BE A 2108 01:29:44,196 --> 01:29:46,265 MODEL FOR NEGATIVE SELECTION OF 2109 01:29:46,265 --> 01:29:47,132 THYMUS BECAUSE AT THE TIME THAT 2110 01:29:47,132 --> 01:29:50,202 WAS THE ONLY SITUATION WE KNEW 2111 01:29:50,202 --> 01:29:51,870 IN WHICH TCR OCCUPANCY WOULD 2112 01:29:51,870 --> 01:29:52,938 RESULT IN DEATH OF T CELLS 2113 01:29:52,938 --> 01:29:54,473 RATHER THAN ACTIVATION. 2114 01:29:54,473 --> 01:29:56,875 WE ALSO KNEW THAT DOUBLE 2115 01:29:56,875 --> 01:30:00,312 POSITIVE THYMOCYTES JUST LIKE T 2116 01:30:00,312 --> 01:30:02,481 CELL -- WAS SENSITIVE TO 2117 01:30:02,481 --> 01:30:05,417 APOPTOSIS INDUCED BY 2118 01:30:05,417 --> 01:30:06,618 GLUCOCORTICOIDS AND POST-DOC IN 2119 01:30:06,618 --> 01:30:08,287 THE LAB WANTED TO KNOW WHETHER 2120 01:30:08,287 --> 01:30:09,722 THESE TWO DEATH STIMULI WERE 2121 01:30:09,722 --> 01:30:11,457 ADDITIVE OR SYNERGISTIC AND 2122 01:30:11,457 --> 01:30:12,358 SURPRISINGLY THEY WERE NEITHER. 2123 01:30:12,358 --> 01:30:14,760 THAT WHEN BOTH THE CELLS WERE 2124 01:30:14,760 --> 01:30:16,962 STIMULATED BOTH VIA TCR AND 2125 01:30:16,962 --> 01:30:17,863 GLUCOCORTICOIDS THE CELL 2126 01:30:17,863 --> 01:30:20,232 SURVIVED AND WE CALLED THAT MEW 2127 01:30:20,232 --> 01:30:22,301 TUT ANTAGONISM AND IMMEDIATELY 2128 01:30:22,301 --> 01:30:27,239 OF COURSE -- MUTE ACTUAL AND -- 2129 01:30:27,239 --> 01:30:27,473 MUTUAL. 2130 01:30:27,473 --> 01:30:28,874 AGAIN THIS IS THE CONVENTIONAL 2131 01:30:28,874 --> 01:30:29,875 SLIDE SHOWN ON THE LEFT SHOWING 2132 01:30:29,875 --> 01:30:31,176 THE RELATIONSHIP BETWEEN 2133 01:30:31,176 --> 01:30:33,345 AFFINITY AND FATE, AND WHAT WE 2134 01:30:33,345 --> 01:30:36,415 POSTULATED WAS THAT 2135 01:30:36,415 --> 01:30:37,950 GLUCOCORTICOIDS ACTUALLY WERE 2136 01:30:37,950 --> 01:30:40,352 IMPORTANT FOR SETTING THAT RIGHT 2137 01:30:40,352 --> 01:30:41,253 THRESHOLD, THE GATE BETWEEN 2138 01:30:41,253 --> 01:30:43,222 POSITIVE AND NEGATIVE SELECTION, 2139 01:30:43,222 --> 01:30:45,657 THAT TCR SIGNALLED THYMOCYTES 2140 01:30:45,657 --> 01:30:47,726 WOULD UNDERGO DEATH NEGATIVE 2141 01:30:47,726 --> 01:30:48,927 SELECTION BUT AT THE LOWER RANGE 2142 01:30:48,927 --> 01:30:50,429 OF THAT HIGH AFFINITY, THAT 2143 01:30:50,429 --> 01:30:52,631 WOULD BE ANTAGONIZED BY 2144 01:30:52,631 --> 01:30:53,866 GLUCOCORTICOIDS AND THOSE CELLS 2145 01:30:53,866 --> 01:30:55,267 WOULD SURVIVE, THEY WOULD BE 2146 01:30:55,267 --> 01:30:57,035 RESCUED FROM NEGATIVE SELECTION. 2147 01:30:57,035 --> 01:30:57,236 OKAY? 2148 01:30:57,236 --> 01:30:59,671 THAT WOULD MAINTAIN A ROBUST AND 2149 01:30:59,671 --> 01:31:02,841 FUNCTIONAL TCR REPERTOIRE. 2150 01:31:02,841 --> 01:31:04,076 AND NECESSARY AND TESTABLE 2151 01:31:04,076 --> 01:31:06,245 PREDICTION OF THIS MODEL IS THAT 2152 01:31:06,245 --> 01:31:08,013 IF ONE WERE TO DECREASE 2153 01:31:08,013 --> 01:31:09,648 SENSITIVITY OF THYME SIETSZ TO 2154 01:31:09,648 --> 01:31:11,617 GLUCOCORTICOIDS THAT NOW THOSE 2155 01:31:11,617 --> 01:31:14,787 CELLS THAT HAVE BEEN RESCUED BY 2156 01:31:14,787 --> 01:31:15,988 GLUCOCORTICOIDS WOULD GO ON TO 2157 01:31:15,988 --> 01:31:17,956 GO NEGATIVELY SELECTED, NO MORE 2158 01:31:17,956 --> 01:31:19,158 AN TAG NISM AND THEREFORE THAT 2159 01:31:19,158 --> 01:31:20,159 GREEN POPULATION THAT POOL THAT 2160 01:31:20,159 --> 01:31:22,161 GOES TO THE PERIPHERY IS GOING 2161 01:31:22,161 --> 01:31:23,996 TO BE REDUCED IN NUMBER AND 2162 01:31:23,996 --> 01:31:24,296 COMPLEXITY. 2163 01:31:24,296 --> 01:31:26,398 SO YOU WOULD END UP BECAUSE 2164 01:31:26,398 --> 01:31:27,466 THOSE CLONES HAVE BEEN LOST IN 2165 01:31:27,466 --> 01:31:28,333 THE REPERTOIRE. 2166 01:31:28,333 --> 01:31:28,534 OKAY? 2167 01:31:28,534 --> 01:31:32,404 SO YOU END UP WITH BOTH A LESS 2168 01:31:32,404 --> 01:31:35,574 COMPLEX REPERTOIRE AND ALSO A 2169 01:31:35,574 --> 01:31:36,975 LESS -- A LOWER AFFINITY 2170 01:31:36,975 --> 01:31:37,976 REPERTOIRE BECAUSE YOU'RE 2171 01:31:37,976 --> 01:31:39,011 ELIMINATING THOSE CELLS THAT ARE 2172 01:31:39,011 --> 01:31:41,480 IN THE HIGHER END OF AFFINITY 2173 01:31:41,480 --> 01:31:43,515 FOR SELF-PEPTIDE PLUS MHC. 2174 01:31:43,515 --> 01:31:45,551 THERE WAS A PROBLEM WITH THIS 2175 01:31:45,551 --> 01:31:46,919 MODEL WE KNEW RIGHT AWAY, AND 2176 01:31:46,919 --> 01:31:48,854 THAT WAS THAT GLUCOCORTICOIDS 2177 01:31:48,854 --> 01:31:51,223 ARE MADE BY THE ADRENALS, RIGHT, 2178 01:31:51,223 --> 01:31:52,624 AND THEY'RE DISTRIBUTED IN THE 2179 01:31:52,624 --> 01:31:55,494 BLOOD AS A HORMONE. 2180 01:31:55,494 --> 01:31:58,430 BUT FETAL MICE OR FETAL MAMMALS 2181 01:31:58,430 --> 01:32:00,165 AND POSTNATAL MICE AND HUMANS 2182 01:32:00,165 --> 01:32:01,834 HAVE VERY LOW LEVELS OF SICK 2183 01:32:01,834 --> 01:32:02,768 LATING GLUCOCORTICOIDS. 2184 01:32:02,768 --> 01:32:04,570 IN MICE THEY DON'T ACHIEVE ADULT 2185 01:32:04,570 --> 01:32:05,737 LEVELS UNTIL ABOUT FOUR WEEKS OF 2186 01:32:05,737 --> 01:32:07,072 AGE AND MUMS I THINK IT TAKES 2187 01:32:07,072 --> 01:32:08,907 ABOUT A YEAR BEFORE THEY REACH 2188 01:32:08,907 --> 01:32:12,077 LEVELS SIMILAR TO OLDER 2189 01:32:12,077 --> 01:32:13,412 INDIVIDUALS, IN HUMANS. 2190 01:32:13,412 --> 01:32:16,682 IF THIS MODEL HAD ANY CHANCE OF 2191 01:32:16,682 --> 01:32:19,351 BEING CORRECT THE -- ITSELF 2192 01:32:19,351 --> 01:32:23,055 WOULD HAVE TO BE A SOURCE OF THE 2193 01:32:23,055 --> 01:32:23,422 GLUCOCORTICOIDS. 2194 01:32:23,422 --> 01:32:23,822 WE'LL TESTS THAT. 2195 01:32:23,822 --> 01:32:25,557 ON THE LEFT, ALL YOU SEE RIGHT 2196 01:32:25,557 --> 01:32:28,427 NOW IS THE SCHEME SHOWING THE 2197 01:32:28,427 --> 01:32:31,163 PATHWAY OF THE NOVA SYNTHESIS 2198 01:32:31,163 --> 01:32:32,030 GLUCOCORTICOIDS ALL THE WAY FROM 2199 01:32:32,030 --> 01:32:34,032 CHOLESTEROL TO CORTICOSTERONE 2200 01:32:34,032 --> 01:32:36,935 WHICH IS THE PROMINENT 2201 01:32:36,935 --> 01:32:38,837 GLUCOCORTICOID MOUSE, WHAT YOU 2202 01:32:38,837 --> 01:32:39,771 REMEMBER FROM THE SLIDE HOWEVER 2203 01:32:39,771 --> 01:32:41,874 IS THIS LAST STEP, AND THAT'S 2204 01:32:41,874 --> 01:32:46,979 THE CONVERSION OF THE OXY 2205 01:32:46,979 --> 01:32:48,747 CORTICOSTERONE WHICH IS INACTIVE 2206 01:32:48,747 --> 01:32:52,651 TO THE GLUCOCORTICOID STER OWN, 2207 01:32:52,651 --> 01:32:54,653 AND FROM THIS SLIDE WHAT I WANT 2208 01:32:54,653 --> 01:32:57,956 YOU TO REMEMBER IS SIP IS 11B1 2209 01:32:57,956 --> 01:32:59,725 BECAUSE THIS WILL PLAY A 2210 01:32:59,725 --> 01:33:00,893 DOMINANT ROLE IN THE REST OF 2211 01:33:00,893 --> 01:33:01,460 THIS TALK. 2212 01:33:01,460 --> 01:33:06,698 SO WE TESTED IF THE THYMUS 2213 01:33:06,698 --> 01:33:08,133 GLUCOCORTICOIDS, A POST-DOC IN 2214 01:33:08,133 --> 01:33:09,968 THE LAB DID A SIMPLE EXPERIMENT, 2215 01:33:09,968 --> 01:33:15,574 TOOK THYMUS, GROUND IT UP, 2216 01:33:15,574 --> 01:33:19,878 COLLAGENOUS TREATED, AND FOR THE 2217 01:33:19,878 --> 01:33:21,813 PRESENCE OF ACTIVE 2218 01:33:21,813 --> 01:33:24,783 CORTICOSTERONE AND WE DIDN'T SEE 2219 01:33:24,783 --> 01:33:25,050 ANYTHING. 2220 01:33:25,050 --> 01:33:28,153 BUT IF SHE THYMOCYTES DEPLETED 2221 01:33:28,153 --> 01:33:29,721 FIRST AND THEN GROUND UP THE 2222 01:33:29,721 --> 01:33:31,757 THYMUS ET CETERA AND NOW WE 2223 01:33:31,757 --> 01:33:34,393 ENRICHED FOR THE HE EPITHELIAL 2224 01:33:34,393 --> 01:33:35,360 COMPONENT IS PUT THE SAME NUMBER 2225 01:33:35,360 --> 01:33:37,095 OF CELLS ABOUT 5 MILLION CELLS 2226 01:33:37,095 --> 01:33:39,665 IN THE WELL NOW SHE COULD SEESLY 2227 01:33:39,665 --> 01:33:46,104 SEE THE PRODUCTION OF DEOCTY. 2228 01:33:46,104 --> 01:33:50,509 SO DOES PRODUCE GLUCOCO-COURT 2229 01:33:50,509 --> 01:33:53,145 STOID T -- GLUCOCORTICOID, NEXT 2230 01:33:53,145 --> 01:33:54,446 FEW SLIDES. 2231 01:33:54,446 --> 01:33:56,415 LET'S TEST OUR MODEL. 2232 01:33:56,415 --> 01:33:58,684 VERY FIRST PREDICTION F WE 2233 01:33:58,684 --> 01:34:01,353 REDUCE RESPONSE WE SHOULD ALTER 2234 01:34:01,353 --> 01:34:02,421 SELECTION AND REPERTOIRE. 2235 01:34:02,421 --> 01:34:03,589 IS THAT THE CASE? 2236 01:34:03,589 --> 01:34:04,656 MANY DIFFERENT STUDIES 2237 01:34:04,656 --> 01:34:05,857 ADDRESSING THIS, BUT I THINK THE 2238 01:34:05,857 --> 01:34:06,725 MOST CONVINCE AND GO WHAT I'LL 2239 01:34:06,725 --> 01:34:08,260 SHOW YOU A FEW EXAMPLES OF IS 2240 01:34:08,260 --> 01:34:09,795 THIS STUDY HERE THRAFS DONE BY 2241 01:34:09,795 --> 01:34:12,397 PAUL MIDDLE DEM STAT A STAFF -- 2242 01:34:12,397 --> 01:34:14,499 MIDDLE DEM STAT A STAFF 2243 01:34:14,499 --> 01:34:16,768 SCIENTIST, KNOCKED OUT THE 2244 01:34:16,768 --> 01:34:20,005 GLUCOCOURTED COID RECEPTOR HAD 2245 01:34:20,005 --> 01:34:21,607 OR AND KNOCKED OUT AND MAKING 2246 01:34:21,607 --> 01:34:23,675 MICE AND CROSSING THEM TO 2247 01:34:23,675 --> 01:34:25,677 APPROXIMATE CRE MICE, NOW ONLY 2248 01:34:25,677 --> 01:34:27,045 THYMOCYTES ARE GOING TO BE 2249 01:34:27,045 --> 01:34:29,181 AFFECTED AND WE SHOWED THAT IN 2250 01:34:29,181 --> 01:34:33,719 FACT THEY DO THIS GR -- EXCUSE 2251 01:34:33,719 --> 01:34:33,885 ME. 2252 01:34:33,885 --> 01:34:36,121 AND THE ADRENAL IS PERFECTLY 2253 01:34:36,121 --> 01:34:37,889 NORMAL, CIRCULATING PERFECTLY 2254 01:34:37,889 --> 01:34:38,123 NORMAL. 2255 01:34:38,123 --> 01:34:40,258 THYMUS IN THESE MICE IS ABOUT A 2256 01:34:40,258 --> 01:34:43,128 THIRD HAD OR SMALLER THAN 2257 01:34:43,128 --> 01:34:43,495 WILD-TYPE. 2258 01:34:43,495 --> 01:34:45,197 DESPITE THAT THE PERIPHERAL T 2259 01:34:45,197 --> 01:34:46,431 CELL POPULATION IN THE POOL 2260 01:34:46,431 --> 01:34:48,300 LOOKS GROSSLY NORMAL, NORMAL 2261 01:34:48,300 --> 01:34:50,936 NUMBERS OF LYMPH NODE T CELLS, 2262 01:34:50,936 --> 01:34:53,038 CD4 CD8 DISTRIBUTIONS ARE VERY 2263 01:34:53,038 --> 01:34:53,438 SIMILAR. 2264 01:34:53,438 --> 01:34:53,639 OKAY? 2265 01:34:53,639 --> 01:34:54,973 SO AMONG THE MANY EXPERIMENTS 2266 01:34:54,973 --> 01:34:56,775 THESE ARE I'LL SHOW YOU JUST A 2267 01:34:56,775 --> 01:34:57,009 COUPLE. 2268 01:34:57,009 --> 01:34:58,744 THIS IS ONE WHERE PAUL IMMUNIZED 2269 01:34:58,744 --> 01:35:02,981 THESE MICE WITH PIGEON 2270 01:35:02,981 --> 01:35:04,850 CYTOCHROME C CHOSEN FOR A 2271 01:35:04,850 --> 01:35:07,352 REASON, THE RESPONSE IN B10A 2272 01:35:07,352 --> 01:35:11,189 MICE DEPICTED IN C IS QUITE 2273 01:35:11,189 --> 01:35:12,391 TYPICAL, CD4 RESPONSE I SHOULD 2274 01:35:12,391 --> 01:35:15,560 SAY, THEY'RE ALL RESTRICTED BY 2275 01:35:15,560 --> 01:35:17,796 IBFK, THEY ALL SEE THE FRAGMENTS 2276 01:35:17,796 --> 01:35:20,298 81 TO 104 AND TEFNED ON USE A T 2277 01:35:20,298 --> 01:35:24,736 CELL RECEPTOR CONTAINING D 2278 01:35:24,736 --> 01:35:27,472 ALPHA-1 1 BETA 3, WE IMMUNIZED 2279 01:35:27,472 --> 01:35:29,141 MICE, WAITED A WEEK, TEN DAYS, 2280 01:35:29,141 --> 01:35:30,876 TOOK OUT DRAINING LYMPH NODES 2281 01:35:30,876 --> 01:35:32,177 AND ASSAYS WITH INCREASING 2282 01:35:32,177 --> 01:35:33,512 AMOUNT OF PEPTIDE AND YOU SEE 2283 01:35:33,512 --> 01:35:36,014 THE WILD-TYPE T CELLS RESPONDED 2284 01:35:36,014 --> 01:35:39,217 PERFECTLY NORMALLY TO ANTIGEN 2285 01:35:39,217 --> 01:35:43,155 BUT THE MICE FROM THE GR 2286 01:35:43,155 --> 01:35:46,358 KNOCK-OUT MICE, GR KNOCK-OUT 2287 01:35:46,358 --> 01:35:47,893 THYMOCYTES MICE THE RESPONSE WAS 2288 01:35:47,893 --> 01:35:50,095 VERY POOR, AT LEAST 100 FOLD 2289 01:35:50,095 --> 01:35:51,730 SHIFT IN RESPONSE TO THE RIGHT. 2290 01:35:51,730 --> 01:35:53,198 NOT BECAUSE THE T CELLS WERE 2291 01:35:53,198 --> 01:35:54,933 SICK OR HAD SOME PROBLEM FROM 2292 01:35:54,933 --> 01:35:56,101 DIVIDING, ET CETERA, BECAUSE IF 2293 01:35:56,101 --> 01:35:58,070 WE CROSS LINK T CELL RECEPTOR, 2294 01:35:58,070 --> 01:36:00,038 TO BYPASS THE SPECIFICITY OF THE 2295 01:36:00,038 --> 01:36:02,541 T CELL RECEPTORS WE GET 2296 01:36:02,541 --> 01:36:03,875 PROLIFERATION AND EVEN BETTER 2297 01:36:03,875 --> 01:36:05,010 EXPERIMENT I THINK WAS THAT WHEN 2298 01:36:05,010 --> 01:36:09,781 WE CROSS THESE NIES THE AND 2299 01:36:09,781 --> 01:36:12,184 TRANSGENIC MOUSE, WHICH HAS A 2300 01:36:12,184 --> 01:36:15,487 RECEPTOR THAT SEES PIGEON 2301 01:36:15,487 --> 01:36:16,688 CYTOKINE C, ALL THE T CELLS 2302 01:36:16,688 --> 01:36:19,091 THERE'S NO POLYCLONAL REPERTOIRE 2303 01:36:19,091 --> 01:36:20,726 NOW, ALL THE T CELLS SEE 2304 01:36:20,726 --> 01:36:22,494 CYTOCHROME C AND NOW ACTIVATE 2305 01:36:22,494 --> 01:36:24,563 WITH ANTIGEN WE SEE THEY'RE 2306 01:36:24,563 --> 01:36:25,731 IDENTICAL TO THE WILD-TYPE T 2307 01:36:25,731 --> 01:36:27,065 CELLS SO THIS IS A REPERTOIRE 2308 01:36:27,065 --> 01:36:30,402 ISSUE, IT'S NOT AN ISSUE WITH 2309 01:36:30,402 --> 01:36:31,570 SPECIFICITY -- OF THE T CELLS 2310 01:36:31,570 --> 01:36:33,138 THEMSELVES, BUT THIS WOULD BE I 2311 01:36:33,138 --> 01:36:35,073 WOULD CONSIDER A CLASSIC KIND OF 2312 01:36:35,073 --> 01:36:36,575 HOLE IN THE REPERTOIRE AND WE 2313 01:36:36,575 --> 01:36:38,910 FOUND MANY OTHER EXAMPLES OF 2314 01:36:38,910 --> 01:36:39,144 THIS. 2315 01:36:39,144 --> 01:36:42,514 WE ALSO HAVE SOME EVIDENCE-BASED 2316 01:36:42,514 --> 01:36:44,483 ON THE REPERTOIRE CHANGE MORE A 2317 01:36:44,483 --> 01:36:46,118 PHYSICAL BASIS, THIS IS A WHILE 2318 01:36:46,118 --> 01:36:48,553 AGO IN THE EARLY DAYS OF HIGH 2319 01:36:48,553 --> 01:36:49,521 THROUGH PRESIDENT SEQUENCING, 2320 01:36:49,521 --> 01:36:56,094 BUT WE SEQUENCED THE CDR3 REGION 2321 01:36:56,094 --> 01:36:58,597 OF NAIVE T CELLS THAT EXPRESS V 2322 01:36:58,597 --> 01:37:00,465 ALPHA ELEVEN BETA 3, I WANT TO 2323 01:37:00,465 --> 01:37:02,434 EXPRESS WE SORTED FOR NAIVE T 2324 01:37:02,434 --> 01:37:03,969 CELLS AND WE EVENTUALLY LOOKED 2325 01:37:03,969 --> 01:37:07,405 AT THE POST THYMIC REPERTOIRE 2326 01:37:07,405 --> 01:37:09,574 PRIOR TO ANY PERTUBATIONS BY 2327 01:37:09,574 --> 01:37:11,676 EXPOSURE TO ANTIGEN, THIS IS THE 2328 01:37:11,676 --> 01:37:14,579 NAIVE REPERTOIRE. 2329 01:37:14,579 --> 01:37:24,189 AND ON THE FREQUENCY OF THE 100 2330 01:37:24,189 --> 01:37:27,359 OR 110 MOST USED CDR3'S IN THE 2331 01:37:27,359 --> 01:37:28,360 WILD-TYPE MICE AND THAT'S IN THE 2332 01:37:28,360 --> 01:37:29,361 CLOSED DOTLE IF YOU CAN SEE 2333 01:37:29,361 --> 01:37:31,863 THAT, AND ASKED WHETHER THE GR 2334 01:37:31,863 --> 01:37:34,032 KNOCK-OUT MICE HAD THE SAME 2335 01:37:34,032 --> 01:37:37,269 REPERTOIRE BASED ON CDR3 AND THE 2336 01:37:37,269 --> 01:37:38,403 ANSWER IS NO IT'S VERY 2337 01:37:38,403 --> 01:37:39,838 DIFFERENT, WE SAW IF WE DID THE 2338 01:37:39,838 --> 01:37:41,039 OPPOSITE EXPERIMENT WE SAW THE 2339 01:37:41,039 --> 01:37:43,341 SAME THING THAT, THE USE OF CDR3 2340 01:37:43,341 --> 01:37:44,543 IS QUITE DIFFERENT. 2341 01:37:44,543 --> 01:37:47,946 SO THIS ACTUALLY, THAT MODEL 2342 01:37:47,946 --> 01:37:50,048 ACTUALLY, THIS TEST OF THAT 2343 01:37:50,048 --> 01:37:52,551 MODEL IS PASSED, IF YOU KNOCK 2344 01:37:52,551 --> 01:37:54,186 OUT GLUCOCORTICOID 2345 01:37:54,186 --> 01:37:56,788 RESPONSIVENESS YOU GET MICE THAT 2346 01:37:56,788 --> 01:37:59,658 ARE IMMUNOLOGICALLY UNFIT, HAVE 2347 01:37:59,658 --> 01:38:01,193 AN ALTERED REPERTOIRE AND 2348 01:38:01,193 --> 01:38:02,828 INEFFECTIVE REPERTOIRE. 2349 01:38:02,828 --> 01:38:04,563 SO LET'S GOT BACK TO THE THYMUS 2350 01:38:04,563 --> 01:38:06,464 AND GLUCOCORTICOID PRODUCTION. 2351 01:38:06,464 --> 01:38:08,667 WE NOW KNOW THAT GLUCOCORTICOID 2352 01:38:08,667 --> 01:38:10,101 IS MADE IN THE THYMUS BUT WE 2353 01:38:10,101 --> 01:38:11,603 DON'T KNOW WHETHER THEY'RE 2354 01:38:11,603 --> 01:38:12,804 BIOLOGICALLY RELEVANT R THEY 2355 01:38:12,804 --> 01:38:14,372 MADE IN A HIGH ENOUGH QUANTITY, 2356 01:38:14,372 --> 01:38:15,473 IN THE RIGHT LOCATION, ET 2357 01:38:15,473 --> 01:38:18,243 CETERA, OR IS THE EFFECT OF 2358 01:38:18,243 --> 01:38:21,213 GLUCOCORTICOIDS MEDIATED VIA 2359 01:38:21,213 --> 01:38:22,080 CIRCULATED ADRENAL SUPPLY POOL. 2360 01:38:22,080 --> 01:38:23,515 TO ANSWER THIS QUESTION, PAUL 2361 01:38:23,515 --> 01:38:25,050 MADE ANOTHER KNOCK-OUT MOUSE, 2362 01:38:25,050 --> 01:38:29,087 THIS TIME KNOCKED OUT CIF11B1 2363 01:38:29,087 --> 01:38:31,690 THE ENZYME THAT SYNTHESIZES 2364 01:38:31,690 --> 01:38:33,491 GLUCOCORTICOIDS AND KNOCKED OUT 2365 01:38:33,491 --> 01:38:36,428 EPITHELIUM, CROTIONED TO FOX 2366 01:38:36,428 --> 01:38:39,731 N1CRE MOUSE SO ONLY THYMIC 2367 01:38:39,731 --> 01:38:41,166 EPITHELIUM WILL BE ABLE TO MAKE 2368 01:38:41,166 --> 01:38:43,902 THE -- CIRCULATING PERFECTLY 2369 01:38:43,902 --> 01:38:44,536 NORMAL, STRETCH RESPONSE IN 2370 01:38:44,536 --> 01:38:46,972 THESE MICE IS PERFECTLY NORMAL. 2371 01:38:46,972 --> 01:38:48,940 HIGHLY LOCALIZED KNOCK YOWLT OF 2372 01:38:48,940 --> 01:38:49,307 GLUCOCORTICOIDS. 2373 01:38:49,307 --> 01:38:50,675 THIS SHOWS YOU THAT THE MICE ARE 2374 01:38:50,675 --> 01:38:54,279 WHAT WE SAY THEY ARE, THIS IS 2375 01:38:54,279 --> 01:38:57,482 DONE BY CALIN DONAHUE A GRAD 2376 01:38:57,482 --> 01:38:59,351 STUDENT IN THE LAB WHO CULTURED 2377 01:38:59,351 --> 01:39:03,088 THYMUS AND NOW OUR SAAS SAYS -- 2378 01:39:03,088 --> 01:39:04,956 NOW OUR ASSAYS ARE BETTER NOW 2379 01:39:04,956 --> 01:39:06,791 AND CAN MEASURE IF 2380 01:39:06,791 --> 01:39:07,459 GLUCOCORTICOID DIRECT FROM 2381 01:39:07,459 --> 01:39:08,793 THYMUS AND SEE IN WILD-TYPE 2382 01:39:08,793 --> 01:39:13,465 MOUSE THEY MAKE PLENTY OF, BUT 2383 01:39:13,465 --> 01:39:15,367 IN THE KNOCK YOWLT MICE YOU 2384 01:39:15,367 --> 01:39:16,801 DON'T SEE IT. 2385 01:39:16,801 --> 01:39:19,804 SO WE CAN NOW ELIMINATE THYME 2386 01:39:19,804 --> 01:39:20,839 SIETSZ PRODUCTION -- THYMUS 2387 01:39:20,839 --> 01:39:22,107 PRODUCTION OF GLUCOCORTICOIDS. 2388 01:39:22,107 --> 01:39:23,708 WHAT HAPPENS NOW? 2389 01:39:23,708 --> 01:39:25,443 DOES THIS HAVE ANY -- WHAT'S THE 2390 01:39:25,443 --> 01:39:26,645 EFFECT OF THIS? 2391 01:39:26,645 --> 01:39:28,980 DO WE SEE ANY DIFFERENCE DPIERTD 2392 01:39:28,980 --> 01:39:30,382 WILD-TYPE MICE? 2393 01:39:30,382 --> 01:39:32,017 SO -- COMPARED TO THE WILD-TYPE 2394 01:39:32,017 --> 01:39:32,217 MICE? 2395 01:39:32,217 --> 01:39:34,085 TO ADDRESS THIS, WE ASKED, WHAT 2396 01:39:34,085 --> 01:39:35,420 DO THYMOCYTES PERCEIVE? 2397 01:39:35,420 --> 01:39:38,590 SO WE LOOKED FROM AT STEROID 2398 01:39:38,590 --> 01:39:41,893 RESPONSIVE GENE, GLUCOCORTICOID 2399 01:39:41,893 --> 01:39:44,162 RESPONSIVE GENES OF THYMOCYTES 2400 01:39:44,162 --> 01:39:45,497 FROM THESE DIFFERENT MICE. 2401 01:39:45,497 --> 01:39:48,133 WE SHOW IN LAB ONE, PAUL DID OUR 2402 01:39:48,133 --> 01:39:51,736 PCR TO QUANTITATE THEM AND GIVE 2403 01:39:51,736 --> 01:39:52,938 WILD-TYPE MICE THYMOCYTES THE 2404 01:39:52,938 --> 01:39:54,372 LEVEL OF ONE AND FIRST LOOKED AT 2405 01:39:54,372 --> 01:39:57,075 THE GR1 KNOCK-OUT THYMOCYTES, 2406 01:39:57,075 --> 01:39:58,076 THYMOCYTES THAT CANNOT RESPOND 2407 01:39:58,076 --> 01:39:59,611 TO GLUCOCORTICOIDS AT ALL 2408 01:39:59,611 --> 01:40:00,612 REGARDLESS OF THEIR SOURCE AND 2409 01:40:00,612 --> 01:40:04,349 WE SEE A 40 TO 50 PEST REDUCTION 2410 01:40:04,349 --> 01:40:05,984 IN -- 40 TO 50% REDUCTION IN 2411 01:40:05,984 --> 01:40:08,053 EXPRESSION OF THESE GENES, BASAL 2412 01:40:08,053 --> 01:40:08,353 PRODUCTION. 2413 01:40:08,353 --> 01:40:09,721 WHAT WAS IMPORTANT HERE IS WHEN 2414 01:40:09,721 --> 01:40:16,728 WE LOOKED AT THE CIF -- AT THE 2415 01:40:16,728 --> 01:40:18,563 CIF -- WE SEE THE EXACT SAME 2416 01:40:18,563 --> 01:40:21,866 REDUCTION IN EXPRESSION OF THESE 2417 01:40:21,866 --> 01:40:22,834 GLUCOCORTICOID RESPONSIVE GENES 2418 01:40:22,834 --> 01:40:23,969 MEANING EFFECTIVELY THE ONLY 2419 01:40:23,969 --> 01:40:25,603 SOURCE OF STEROIDS THAT ARE 2420 01:40:25,603 --> 01:40:27,238 PERCEIVED BY THYMOCYTES AT LEAST 2421 01:40:27,238 --> 01:40:30,408 TO THE LEVEL OF UPREGULATING 2422 01:40:30,408 --> 01:40:32,377 THESE GENES AND OTHERS IS 2423 01:40:32,377 --> 01:40:34,112 PRODUCED LOCALLY, THAT SYSTEMIC 2424 01:40:34,112 --> 01:40:36,214 LEVELS DON'T MAKE A DIFFERENCE, 2425 01:40:36,214 --> 01:40:38,817 IT IS ONLY LOCAL PRODUCTION THAT 2426 01:40:38,817 --> 01:40:39,584 MATTERS. 2427 01:40:39,584 --> 01:40:41,987 AND COMFORTINGLY, THESE MICE 2428 01:40:41,987 --> 01:40:45,590 COPIED THE GENE KNOCK-OUT MICE, 2429 01:40:45,590 --> 01:40:48,360 T CELL RESPONSES WERE POOR, 2430 01:40:48,360 --> 01:40:49,227 SHOWING -- INTOWNTS THE T CELLS 2431 01:40:49,227 --> 01:40:50,996 ARE FINE BECAUSE IF YOU CROSS T 2432 01:40:50,996 --> 01:40:53,031 CELL RECEPTOR, THEY RESPOND 2433 01:40:53,031 --> 01:40:53,298 NORMALLY. 2434 01:40:53,298 --> 01:40:55,333 SO WHO IS MAKING THE 2435 01:40:55,333 --> 01:40:56,134 GLUCOCORTICOIDS? 2436 01:40:56,134 --> 01:40:59,070 CORTICAL EPITHELIUM OR THYMIC 2437 01:40:59,070 --> 01:41:00,705 EPITHELIUM, MEDULLARY 2438 01:41:00,705 --> 01:41:01,740 EPITHELIUM? 2439 01:41:01,740 --> 01:41:01,940 SORRY. 2440 01:41:01,940 --> 01:41:02,907 THANK YOU FOR GOING THROUGH ALL 2441 01:41:02,907 --> 01:41:04,542 THAT SO I CAN JUST IGNORE THE 2442 01:41:04,542 --> 01:41:06,444 DIFFERENCES BETWEEN THESE CELLS, 2443 01:41:06,444 --> 01:41:08,279 WITH THE EXCEPTION I'M UNDER 2444 01:41:08,279 --> 01:41:10,482 STRESS, THE POINT THAT AIRE IS 2445 01:41:10,482 --> 01:41:13,551 EXPRESSED BY MEDULLARY TEC AND 2446 01:41:13,551 --> 01:41:24,095 THAT AIRE DRIVES -- AS FAMOUSLY 2447 01:41:24,462 --> 01:41:28,133 PROCESSED TO ANTIGENS THAT ARE 2448 01:41:28,133 --> 01:41:32,170 THEN USED TO DELETE CAUSE 2449 01:41:32,170 --> 01:41:35,040 NEGATIVE SELECTION OF CELLS THAT 2450 01:41:35,040 --> 01:41:36,341 BEAR -- RECEPTORS THAT RECOGNIZE 2451 01:41:36,341 --> 01:41:37,342 THEM, IMPORTANT IN THE 2452 01:41:37,342 --> 01:41:39,077 ESTABLISHMENT OF CENTRAL 2453 01:41:39,077 --> 01:41:39,411 TOLERANCE. 2454 01:41:39,411 --> 01:41:43,181 SO WE COULDN'T USE ANTIBODIES TO 2455 01:41:43,181 --> 01:41:44,082 CIF112B1 TO IDENTIFY THE CELLS 2456 01:41:44,082 --> 01:41:45,583 OF INTEREST BECAUSE THE 2457 01:41:45,583 --> 01:41:47,018 ANTIBODIES CROSS-REACT, THESE 2458 01:41:47,018 --> 01:41:48,887 ENZYMES ARE VERY SIMILAR IN 2459 01:41:48,887 --> 01:41:53,725 THESE PATHWAYS, AND THE ANTIBODY 2460 01:41:53,725 --> 01:41:54,059 CROSS-REACTS. 2461 01:41:54,059 --> 01:41:57,195 SO MATT TAVES A POST-DOC IN THE 2462 01:41:57,195 --> 01:41:59,164 LAB DECIDES TO MAKE A REPORTER 2463 01:41:59,164 --> 01:42:01,900 MOUSE, SCARLET TO CIP11B1 SHOWN 2464 01:42:01,900 --> 01:42:04,102 HERE AND KNOCK THAT INTO THE 2465 01:42:04,102 --> 01:42:05,970 ENDOGENOUS LOCUS IN B6 MICE AND 2466 01:42:05,970 --> 01:42:07,138 YOU CAN SEE ON THE FAR RIGHT 2467 01:42:07,138 --> 01:42:09,307 THAT IT WORKED BECAUSE THAT'S A 2468 01:42:09,307 --> 01:42:12,277 CROSS-SECTION OF THE ADRENAL AND 2469 01:42:12,277 --> 01:42:19,384 YOU CAN SEE THE ZONAFOCICULADA 2470 01:42:19,384 --> 01:42:21,619 LIGHTS UP, SO TO ADDRESS THE 2471 01:42:21,619 --> 01:42:23,054 QUESTION WE'VE BEEN THINKING 2472 01:42:23,054 --> 01:42:25,457 ABOUT SINCE EARLY 190S AND THAT 2473 01:42:25,457 --> 01:42:27,625 IS WHICH CELL IS ACTUALLY MAKING 2474 01:42:27,625 --> 01:42:27,992 GLUCOCORTICOIDS? 2475 01:42:27,992 --> 01:42:30,361 IF YOU LOOK AT THE RIGHT, LOWER 2476 01:42:30,361 --> 01:42:33,264 RIGHT, THAT'S FOLK AM MICROSCOPY 2477 01:42:33,264 --> 01:42:34,999 THAT KAITLYN DONAHUE PERFORMED 2478 01:42:34,999 --> 01:42:37,902 IN WHICH SHE STAINED THE 2479 01:42:37,902 --> 01:42:42,073 CORTICAL IF TEC'S WITH L51 2480 01:42:42,073 --> 01:42:44,576 THAT'S BLUE NO, RED IN THAT 2481 01:42:44,576 --> 01:42:46,778 REGION, REGULAR TEXT IS STAINED 2482 01:42:46,778 --> 01:42:49,380 IN GREEN UEA1 AND MEDULLA 2483 01:42:49,380 --> 01:42:50,482 EXPRESSION I HOPE YOU CAN SEE T 2484 01:42:50,482 --> 01:42:51,649 CAPABILITY SEE IT FROM THIS 2485 01:42:51,649 --> 01:42:53,351 ANGLE, THAT QUIEPT A FEW RED 2486 01:42:53,351 --> 01:42:55,220 CELLS, YELLOW CELLS, ET CETERA. 2487 01:42:55,220 --> 01:42:57,155 SO IT'S THE MEDULLARY TEXT THAT 2488 01:42:57,155 --> 01:43:02,760 ARE MAKING FROM CYP11B1 2489 01:43:02,760 --> 01:43:05,296 CONFIRMED ON THE RIGHT BY FLOW 2490 01:43:05,296 --> 01:43:06,731 CYTOMETRY WHERE KAITLYN NUND IF 2491 01:43:06,731 --> 01:43:08,800 YOU LOOK AT THE BOTTOM -- FOUND 2492 01:43:08,800 --> 01:43:09,968 IF YOU LOOK AT THE BOTTOM, THE 2493 01:43:09,968 --> 01:43:11,503 REPORTER MICE, THAT THYMOCYTES 2494 01:43:11,503 --> 01:43:15,473 DON'T MAKE IT, APC, MACROPHAGES 2495 01:43:15,473 --> 01:43:19,310 DON'T MAKE TCTEC DON'T MAKE IT, 2496 01:43:19,310 --> 01:43:21,713 MTEC LOWS WHICH ARE THE MORE 2497 01:43:21,713 --> 01:43:28,153 MAIM TOUR MTEC'S START TO MAKE 2498 01:43:28,153 --> 01:43:32,290 CYP11B1 AND ABOUT TEN PERCENT OF 2499 01:43:32,290 --> 01:43:34,092 THE CELLS ARE POSITIVE. 2500 01:43:34,092 --> 01:43:36,461 AT THE TIME IN 2022 THE PAPER 2501 01:43:36,461 --> 01:43:40,832 FROM NICHOLSON AND FROM 2502 01:43:40,832 --> 01:43:41,933 MATHESON'S LAB, I'VE REFERRED 2503 01:43:41,933 --> 01:43:45,336 TO, WHERE THEY SORTED MHC LOW 2504 01:43:45,336 --> 01:43:52,243 CELLS AND LOOKED AT CELLLY SEQ, 2505 01:43:52,243 --> 01:43:53,444 WE PREFERRED CELL HIGH, WE WOULD 2506 01:43:53,444 --> 01:43:54,879 TAKE WHAT WE WOULD GERKTS PLEASE 2507 01:43:54,879 --> 01:43:57,348 TO DO SEE WHEN WE INTERROGATED 2508 01:43:57,348 --> 01:43:59,918 THEIR DATASET, WE COULD SEE A 2509 01:43:59,918 --> 01:44:02,854 POPULATION OF CYP11B1 THAT 2510 01:44:02,854 --> 01:44:06,591 CLUSTERED IN THIS ANALYSIS. 2511 01:44:06,591 --> 01:44:08,993 IMPORTANTLY FOR US, IT TURNED 2512 01:44:08,993 --> 01:44:11,296 OUT THAT THAT CLUSTER ALSO WAS 2513 01:44:11,296 --> 01:44:12,463 ENRICHED IN AIRE. 2514 01:44:12,463 --> 01:44:14,933 SO WE ASKED THE QUESTION, IS 2515 01:44:14,933 --> 01:44:16,901 AIRE RELATED IN FACT TO THE 2516 01:44:16,901 --> 01:44:20,705 EXPRESSION OF CYP11B1? 2517 01:44:20,705 --> 01:44:23,208 KAITLYN LOOKED AT CYP11B1 2518 01:44:23,208 --> 01:44:24,209 POSITIVE CELLS AND COUNT THE 2519 01:44:24,209 --> 01:44:26,711 NUMBER THAT ARE EITHER IF AIRE 2520 01:44:26,711 --> 01:44:28,012 POSITIVE OR NEGATIVE AND AS YOU 2521 01:44:28,012 --> 01:44:30,548 CAN SEE ABOUT 60% OF THOSE CELLS 2522 01:44:30,548 --> 01:44:32,951 THAT MAKE GLUCOCORTICOIDS ARE 2523 01:44:32,951 --> 01:44:34,385 AIRE POSITIVE. 2524 01:44:34,385 --> 01:44:35,920 AN EXAMPLE SHOWN THE CONFOCAL 2525 01:44:35,920 --> 01:44:39,757 EXAMPLE I SHOWED THERE SHOWS AN 2526 01:44:39,757 --> 01:44:42,393 AIRE POSITIVE CELL IS ALSO 2527 01:44:42,393 --> 01:44:43,228 CYP11B1 POSITIVE. 2528 01:44:43,228 --> 01:44:45,697 WHAT ABOUT THE 40% AIRE 2529 01:44:45,697 --> 01:44:46,130 NEGLECTITY? 2530 01:44:46,130 --> 01:44:46,965 DOES THAT MEAN IT'S NOT 2531 01:44:46,965 --> 01:44:49,267 NECESSARILY INVOLVED? 2532 01:44:49,267 --> 01:44:50,435 MTEC'S DON'T STOP 2533 01:44:50,435 --> 01:44:51,369 DIFFERENTIATING AFTER THEY 2534 01:44:51,369 --> 01:44:54,505 BECOME AIRE POSITIVE, THEY FORM 2535 01:44:54,505 --> 01:44:56,574 A STATE WHERE THEY LOSE AIRE 2536 01:44:56,574 --> 01:44:58,109 EXPRESSION AND GAIN OTHER 2537 01:44:58,109 --> 01:45:00,078 ANTIGENS LIKE OR MOLECULES LIKE 2538 01:45:00,078 --> 01:45:00,612 KERATIN 10. 2539 01:45:00,612 --> 01:45:03,047 THESE ARE CALLED POST AIRE 2540 01:45:03,047 --> 01:45:03,581 CELLS. 2541 01:45:03,581 --> 01:45:04,449 KAITLYN THEN ASKED WHAT 2542 01:45:04,449 --> 01:45:07,652 PERCENTAGE OF THESE 40%, WHAT 2543 01:45:07,652 --> 01:45:09,120 PERCENTAGE ALSO EXPRESS KERATIN 2544 01:45:09,120 --> 01:45:10,521 10 AND THE ANSWER WAS 80%, 2545 01:45:10,521 --> 01:45:12,056 THAT'S IN THE UPPER RIGHT, MY 2546 01:45:12,056 --> 01:45:13,124 POINTER IS NOT WORKING SO I 2547 01:45:13,124 --> 01:45:15,760 CAN'T POINT TO T BUT THE ANSWER 2548 01:45:15,760 --> 01:45:18,096 IS THAT BY THIS ANALYSIS, ALMOST 2549 01:45:18,096 --> 01:45:22,467 ALL THE CELLS THAT MAKE UP 2550 01:45:22,467 --> 01:45:23,801 CYP11B1 ARE EITHER POSITIVE AT 2551 01:45:23,801 --> 01:45:25,470 THAT TIME OR MORE RECENTLY AIR 2552 01:45:25,470 --> 01:45:27,305 HE POSITIVE, CONFIRMED WHEN 2553 01:45:27,305 --> 01:45:29,741 KAITLYN CROSSED THE AIRE 2554 01:45:29,741 --> 01:45:31,075 KNOCK-OUT MICE TO THE REPORTER 2555 01:45:31,075 --> 01:45:33,111 MICE AND YOU CAN SEE THAT IF YOU 2556 01:45:33,111 --> 01:45:36,180 LOOK AT THE BOTTOM LEFT YOU CAN 2557 01:45:36,180 --> 01:45:38,349 SEE THAT THE AIRE KNOCKOUTS LOSE 2558 01:45:38,349 --> 01:45:42,954 EXPRESSION OF CYP11B1. 2559 01:45:42,954 --> 01:45:46,124 NOW, CYP11B1 BY ITSELF DOESN'T 2560 01:45:46,124 --> 01:45:48,559 MAKE GLUCOCORTICOIDS, NEEDS ALL 2561 01:45:48,559 --> 01:45:49,961 THE OTHER ENZYME PS IN THE SUITE 2562 01:45:49,961 --> 01:45:53,798 AS SHOWN THERE, SO LOOKED AT THE 2563 01:45:53,798 --> 01:45:54,999 SEQ DATA IN THE THYMUS WHICH 2564 01:45:54,999 --> 01:45:56,668 POPULATION CAN MAKE ALL THE 2565 01:45:56,668 --> 01:45:57,502 ENZYMES AND THE ONLY POPULATION 2566 01:45:57,502 --> 01:45:59,470 THAT COULD MAKE ALL THE ENZYMES 2567 01:45:59,470 --> 01:46:01,005 IS THE TEC'S. 2568 01:46:01,005 --> 01:46:03,408 IF YOU LOOK TO THE RIGHT OF 2569 01:46:03,408 --> 01:46:05,376 THAT, HE ALSO ASKED NOW IF YOU 2570 01:46:05,376 --> 01:46:07,145 SUBDIVIDE THE TEC'S INTO 2571 01:46:07,145 --> 01:46:08,112 DIFFERENT POPULATIONS, WHAT 2572 01:46:08,112 --> 01:46:09,547 ENZYME DID THEY EXPRESS, AND 2573 01:46:09,547 --> 01:46:12,583 ONLY THE MTEC HIGHS MADE ALL THE 2574 01:46:12,583 --> 01:46:14,686 ENZYME SAYS REQUIRED FOR 2575 01:46:14,686 --> 01:46:15,987 GLUCOCORTICOID SYNTHESIS. 2576 01:46:15,987 --> 01:46:17,088 THAT DOESN'T MEAN THEY'RE ALL ON 2577 01:46:17,088 --> 01:46:18,289 THE SAME CELL, OF COURSE, 2578 01:46:18,289 --> 01:46:19,624 HOWEVER, THEY ARE ON THE SAME 2579 01:46:19,624 --> 01:46:19,824 CELL. 2580 01:46:19,824 --> 01:46:21,225 IF YOU LOOK TO THE FAR RIGHT AS 2581 01:46:21,225 --> 01:46:23,094 I'VE ALREADY SHOWN YOU WE CAN 2582 01:46:23,094 --> 01:46:25,396 DETECT THYMOCYTES PRODUCTION OR 2583 01:46:25,396 --> 01:46:26,464 GLUCOCORTICOID PRUK FROM THE 2584 01:46:26,464 --> 01:46:29,667 THYMUS AND AS YOU SEE HERE, WHEN 2585 01:46:29,667 --> 01:46:32,170 YOU KNOCK OUT AIR HE IT MOSTLY 2586 01:46:32,170 --> 01:46:33,738 GOES AWAISM THIS IS THE LAST 2587 01:46:33,738 --> 01:46:34,672 DATA SLIDE. 2588 01:46:34,672 --> 01:46:37,342 WE THEN ASKED WHAT OTHER 2589 01:46:37,342 --> 01:46:40,078 PATHWAYS MIGHT BE EXPRESSED IN 2590 01:46:40,078 --> 01:46:41,546 MTEC'S? 2591 01:46:41,546 --> 01:46:46,751 KAITLYN DID RPCR ON MTEC'S AND 2592 01:46:46,751 --> 01:46:48,853 INVOLVING BIOSUN THINK CIS, NOT 2593 01:46:48,853 --> 01:46:52,123 JUST GLUCOCORTICOIDS BUTS 2594 01:46:52,123 --> 01:46:53,024 PROGESTERONE AND SEX STEROIDS 2595 01:46:53,024 --> 01:46:54,859 AND YOU CAN SEE THEY ALL THESE 2596 01:46:54,859 --> 01:46:56,627 ENZYMES ARE PRESENT IN MTEC'S 2597 01:46:56,627 --> 01:46:59,364 AND THEY'RE ALL DEPENDENT ON AIR 2598 01:46:59,364 --> 01:46:59,530 HE. 2599 01:46:59,530 --> 01:47:01,332 WHEN YOU KNOCK OUT AIRE THEY ALL 2600 01:47:01,332 --> 01:47:04,602 GO DOWN, PROMPTING US TO 2601 01:47:04,602 --> 01:47:05,903 EXPERIMENT WE NEVER HAD THOUGHT 2602 01:47:05,903 --> 01:47:11,809 OF DOING BEFORE, AND THAT IS 2603 01:47:11,809 --> 01:47:14,879 TO -- PROGESTERONE, TESTOSTERONE 2604 01:47:14,879 --> 01:47:17,281 AND HE IS STROH DIAL AND -- 2605 01:47:17,281 --> 01:47:19,283 ESTRADIOL AND THEY'RE ALL 2606 01:47:19,283 --> 01:47:21,152 MARKEDLY REDUCED IN THE ABSENCE 2607 01:47:21,152 --> 01:47:22,253 OF AIRE. 2608 01:47:22,253 --> 01:47:24,522 TAKE AWAY, CONCLUSION IS THAT 2609 01:47:24,522 --> 01:47:25,890 AIRE DRIVES EXPRESSION OF NOT 2610 01:47:25,890 --> 01:47:29,827 JUST TRA'S BUT ALSO ENTIRE 2611 01:47:29,827 --> 01:47:31,863 SYNTHETIC PATHWAYS, GENES THAT 2612 01:47:31,863 --> 01:47:33,231 CAN PRODUCE SECONDARY REAGENTS 2613 01:47:33,231 --> 01:47:35,500 THAT ARE INDIRECTLY REGULATED BY 2614 01:47:35,500 --> 01:47:37,135 AIRE BUT ARE ACTUALLY PRODUCED 2615 01:47:37,135 --> 01:47:41,272 BY SYNTHETIC PATHWAYS. 2616 01:47:41,272 --> 01:47:44,409 NOW, THERE'S A PARADOX HERE, AND 2617 01:47:44,409 --> 01:47:47,545 THAT IS WHY WOULD AIRE PROMOTE 2618 01:47:47,545 --> 01:47:49,781 BOTH TRA MEDIATED DELETION, 2619 01:47:49,781 --> 01:47:51,449 WHICH IS REQUIRED FOR TOLERANCE, 2620 01:47:51,449 --> 01:47:53,451 AS WELL AS GLUCOCORTICOID 2621 01:47:53,451 --> 01:47:54,252 MEDIATED POSITIVE SELECTION, 2622 01:47:54,252 --> 01:47:57,922 WHICH IS REQUIRED FOR EFFECTIVE 2623 01:47:57,922 --> 01:48:00,358 IMMUNITY? 2624 01:48:00,358 --> 01:48:01,859 I THINK THE ANSWER ACTUALLY IS 2625 01:48:01,859 --> 01:48:04,262 PRETTY CLEAR, AND THAT IS TO 2626 01:48:04,262 --> 01:48:06,330 EX-PABD THE T CELL RECEPTOR. 2627 01:48:06,330 --> 01:48:08,332 IF MT HE C'S WERE ONLY DELETING 2628 01:48:08,332 --> 01:48:10,501 AND EXPRESSING THOUSANDS AND 2629 01:48:10,501 --> 01:48:11,702 THOUSANDS OF -- GENERAL OUGHTING 2630 01:48:11,702 --> 01:48:13,337 MANY THOUSANDS OF SPECIFICITIES 2631 01:48:13,337 --> 01:48:14,739 THEN YOU WOULD LOSE ALL THOSE 2632 01:48:14,739 --> 01:48:15,573 CELLS FROM THE REPERTOIRE AND 2633 01:48:15,573 --> 01:48:17,308 YOU WOULD HAVE A RESTRICTED 2634 01:48:17,308 --> 01:48:17,608 REPERTOIRE. 2635 01:48:17,608 --> 01:48:20,678 BUT BECAUSE THE SAME CELL THAT'S 2636 01:48:20,678 --> 01:48:22,180 PRESENTING THE ANTIGEN THAT 2637 01:48:22,180 --> 01:48:23,648 WOULD OTHERWISE DLEELT AND 2638 01:48:23,648 --> 01:48:25,583 PRODUCES GLUCOCORTICOIDS THAT 2639 01:48:25,583 --> 01:48:27,752 ARE THERE THAT SYNAPSE OR IN 2640 01:48:27,752 --> 01:48:30,054 THAT IF YOU WILL IN THAT REGION, 2641 01:48:30,054 --> 01:48:30,922 NOTICES CELLS CAN BE SELECTED 2642 01:48:30,922 --> 01:48:33,324 AND GO ON -- THOSE CELLS CAN BE 2643 01:48:33,324 --> 01:48:35,126 SELECTED AND GO ON TO PROMOTE 2644 01:48:35,126 --> 01:48:35,793 POSITIVE IMMUNE RESPONSE AND 2645 01:48:35,793 --> 01:48:37,595 THIS IS EXACTLY WHAT WE SAW WHEN 2646 01:48:37,595 --> 01:48:39,697 WE KNOCKED OUT GLUCOCORTICOID 2647 01:48:39,697 --> 01:48:45,136 RECEPTOR IN THYME SIETSZ, WE SAW 2648 01:48:45,136 --> 01:48:47,138 ARE REPERTOIRE AND WEAK IMMUNE 2649 01:48:47,138 --> 01:48:47,371 SYSTEM. 2650 01:48:47,371 --> 01:48:48,372 I'VE ALREADY MENTIONED I THINK 2651 01:48:48,372 --> 01:48:49,574 PEOPLE INVOLVED IN THIS WORK. 2652 01:48:49,574 --> 01:48:52,009 I WANT FOINT OUT AGAIN THAT MATT 2653 01:48:52,009 --> 01:48:54,579 TASE AND KAITLYN WERE INVOLVED 2654 01:48:54,579 --> 01:48:58,583 FROM THE POINT FROM THE -- AND 2655 01:48:58,583 --> 01:48:59,584 DID ALL THE WORK FROM THE 2656 01:48:59,584 --> 01:49:02,186 REPORTER MOUSE ON, MATT IS AT 2657 01:49:02,186 --> 01:49:03,287 CORNELL WRITING GRANTS AND 2658 01:49:03,287 --> 01:49:05,256 KAITLYN IS IN THE LAB BUSY 2659 01:49:05,256 --> 01:49:06,924 WRITING HER THESIS. 2660 01:49:06,924 --> 01:49:11,128 SO THANK YOU VERY MUCH. 2661 01:49:11,128 --> 01:49:20,371 [APPLAUSE] 2662 01:49:20,371 --> 01:49:22,640 >> THIS TIME I THINK WE'LL ASK 2663 01:49:22,640 --> 01:49:23,841 EXACTLY THE SAME QUESTION. 2664 01:49:23,841 --> 01:49:25,676 YOU MIGHT KNOW THE OLD WORK BY 2665 01:49:25,676 --> 01:49:27,979 RICHARD BOYTES THAT HAS 2666 01:49:27,979 --> 01:49:30,081 DEMONSTRATED THAT SEX STEROIDS 2667 01:49:30,081 --> 01:49:31,382 DURING AGING MIGHT BE ONE OF THE 2668 01:49:31,382 --> 01:49:31,649 REASONS -- 2669 01:49:31,649 --> 01:49:33,150 >> IGD HARDLY HEAR, I'M SORRY. 2670 01:49:33,150 --> 01:49:34,652 >> I'M SURE IF I CAN GET THIS 2671 01:49:34,652 --> 01:49:42,026 ANY LOUDER. 2672 01:49:42,026 --> 01:49:42,426 OKAY. 2673 01:49:42,426 --> 01:49:43,995 SO ACTUALLY DEMONSTRATING THAT 2674 01:49:43,995 --> 01:49:46,597 SEX STEROIDS BLOCK AIDS CAN LEAD 2675 01:49:46,597 --> 01:49:49,100 TO IMPROVED THYMIC FUNCTION H HE 2676 01:49:49,100 --> 01:49:52,837 ALSO STIPULATED THAT WITH AGE 2677 01:49:52,837 --> 01:49:54,372 THAT SEX STEROID LEVELS GO UP 2678 01:49:54,372 --> 01:49:56,173 AND THAT THAT MIGHT BE ONE OF 2679 01:49:56,173 --> 01:50:00,811 THE REASONS FOR THE EVOLUTION. 2680 01:50:00,811 --> 01:50:01,679 -- INVOLUTION. 2681 01:50:01,679 --> 01:50:02,780 MY QUESTION IS OF COURSE IF YOU 2682 01:50:02,780 --> 01:50:04,315 SEE ANY CHANGES IN THE 2683 01:50:04,315 --> 01:50:07,485 GENERATION OF THESE SEX STEROIDS 2684 01:50:07,485 --> 01:50:11,355 WITH AGE BY THESE TEC'S. 2685 01:50:11,355 --> 01:50:14,625 >> GOOD QUESTION. 2686 01:50:14,625 --> 01:50:16,227 IT'S IMPORTANT TO DO THE WORK. 2687 01:50:16,227 --> 01:50:17,795 >> WE CAN'T HEAR YOU ON THE 2688 01:50:17,795 --> 01:50:19,530 ZIESH OH, I CAN SPEAK LOUDER. 2689 01:50:19,530 --> 01:50:20,865 YES, THAT'S A VERY GOOD 2690 01:50:20,865 --> 01:50:22,600 QUESTION, AND YES, OF COURSE THE 2691 01:50:22,600 --> 01:50:25,770 WORK, IN FACT, MATT AND I WROTE 2692 01:50:25,770 --> 01:50:27,738 A REVIEW LAST YEAR ON THE ROLE 2693 01:50:27,738 --> 01:50:29,273 OF SEX STEROIDS AND THYMUS AND 2694 01:50:29,273 --> 01:50:30,808 IT TURNS OUT THEY MOSTLY 2695 01:50:30,808 --> 01:50:33,110 REGULATE THE EPITHELIUM, RATHER 2696 01:50:33,110 --> 01:50:36,714 THAN THE THYMUS ACTUALLY 2697 01:50:36,714 --> 01:50:37,014 THEMSELVES. 2698 01:50:37,014 --> 01:50:38,583 THIS INFORMATION ABOUT THE SEX 2699 01:50:38,583 --> 01:50:39,584 STEROIDS IS RATHER NEW. 2700 01:50:39,584 --> 01:50:42,553 WE HAVE NOT EXPLORED THAT, 2701 01:50:42,553 --> 01:50:44,188 ANYWHERE CLOSE TO BEING DONE 2702 01:50:44,188 --> 01:50:45,723 WITH GLUCOCORTICOIDS HAD. 2703 01:50:45,723 --> 01:50:48,359 WHAT I CAN TELL YOU IS THAT 2704 01:50:48,359 --> 01:50:50,428 GLUCOCORTICOID SYNTHESIS IS 2705 01:50:50,428 --> 01:50:52,597 HIGHEST IN FEEBLGHTS LIFE AND 2706 01:50:52,597 --> 01:50:56,934 NEONATAL LIFE, AND IT GOES DOWN 2707 01:50:56,934 --> 01:50:58,002 POST-BIRTH AND REACHES ADULT 2708 01:50:58,002 --> 01:50:59,937 LEVELS ALTHOUGH ABOUT 50% AT 2709 01:50:59,937 --> 01:51:06,310 ABOUT FOUR WEEKS OF AGE, WHICH 2710 01:51:06,310 --> 01:51:10,815 YOU MENTIONED THE ROLE OF AGING 2711 01:51:10,815 --> 01:51:11,782 AND GLUB CORTICOIDS. 2712 01:51:11,782 --> 01:51:13,851 I SUSPECT THAT'S GOING TO BE 2713 01:51:13,851 --> 01:51:15,386 TRUE FOR THE OTHER PATHWAYS 2714 01:51:15,386 --> 01:51:16,587 INVOLVED BECAUSE THEY ARE 2715 01:51:16,587 --> 01:51:18,689 DEPENDENT. 2716 01:51:18,689 --> 01:51:19,023 OKAY? 2717 01:51:19,023 --> 01:51:24,462 BUT WE HAVE NOT INVESTIGATED 2718 01:51:24,462 --> 01:51:24,662 THAT. 2719 01:51:24,662 --> 01:51:26,330 >> I ACTUALLY AGAIN WANTED TO 2720 01:51:26,330 --> 01:51:27,798 ASK A RELATED QUESTION, BUT THIS 2721 01:51:27,798 --> 01:51:32,870 IS REALLY ABOUT DIOXINS AND IT'S 2722 01:51:32,870 --> 01:51:34,205 VERY OLD LITERATURE ABOUT 2723 01:51:34,205 --> 01:51:36,073 BASICALLY THE ENDOCRINE 2724 01:51:36,073 --> 01:51:38,275 DISRUPTING EFFECT OF DIOXINS ON 2725 01:51:38,275 --> 01:51:40,011 THE THYMUS AND IT SEEMS TO ME 2726 01:51:40,011 --> 01:51:40,845 THAT WITH THE TOOLS THAT YOU 2727 01:51:40,845 --> 01:51:41,746 HAVE RIGHT NOW IT WOULD BE 2728 01:51:41,746 --> 01:51:43,280 REALLY INTERESTING TO REEXAMINE 2729 01:51:43,280 --> 01:51:46,684 THIS WHOLE ISSUE AND TO ASK 2730 01:51:46,684 --> 01:51:48,552 WHICH CELLS ROLES ARE REALLY 2731 01:51:48,552 --> 01:51:50,054 BEING EITHER MIMICKED OR SUB 2732 01:51:50,054 --> 01:51:51,956 VERLTED BY THE DIOXINS. 2733 01:51:51,956 --> 01:51:55,359 >> YOU SEE WITH AHR SIGNALLING? 2734 01:51:55,359 --> 01:51:56,927 >> YES, ALSO THERE WAS SOME 2735 01:51:56,927 --> 01:51:59,230 POSSIBILITY THAT IT HAD AN 2736 01:51:59,230 --> 01:52:01,198 ESTROGEN EFFECTED AND I WAS 2737 01:52:01,198 --> 01:52:02,967 CURIOUS IF THAT HAD COME UP IN 2738 01:52:02,967 --> 01:52:03,834 ANY OF THE CONNECTIONS OF THE 2739 01:52:03,834 --> 01:52:06,337 WORK THAT YOU'RE SEEING. 2740 01:52:06,337 --> 01:52:08,839 >> THAT'S POSSIBLE. 2741 01:52:08,839 --> 01:52:11,275 WE DO THINK, I WANT TO JUST 2742 01:52:11,275 --> 01:52:14,779 MENTION, WE SAW OTHER PALT WAYS 2743 01:52:14,779 --> 01:52:17,281 THAT WEEFNLT INVESTIGATED YET -- 2744 01:52:17,281 --> 01:52:18,482 PATHWAYS THAT WE'VE NOT 2745 01:52:18,482 --> 01:52:20,151 INVESTIGATED YET, ENZYME 2746 01:52:20,151 --> 01:52:21,352 PATHWAYS IN FEALT ACID SIFN 2747 01:52:21,352 --> 01:52:22,787 THINK CIS AND PHOSPHATE 2748 01:52:22,787 --> 01:52:24,522 GENERATION SO WE THINK LOTS OF 2749 01:52:24,522 --> 01:52:25,956 THESE PALT WAYS THAT MAY BE 2750 01:52:25,956 --> 01:52:28,559 INVOLVED HERE. 2751 01:52:28,559 --> 01:52:34,699 >> JOHN, COULD YOU JUST SIMPLY 2752 01:52:34,699 --> 01:52:37,334 -- THE MTEC'S, THE 2753 01:52:37,334 --> 01:52:37,568 SUBSET? 2754 01:52:37,568 --> 01:52:39,303 >> YOU'RE TALKING ABOUT THE 2755 01:52:39,303 --> 01:52:42,907 TISSUE MTEC'S THAT MATHIS WAS 2756 01:52:42,907 --> 01:52:43,774 HERE TALKING ABOUT RECENTLY? 2757 01:52:43,774 --> 01:52:44,408 >> YES. 2758 01:52:44,408 --> 01:52:48,846 >> THOSE WOULD BE THE POST 2759 01:52:48,846 --> 01:52:50,915 AIRE MTEC'S. 2760 01:52:50,915 --> 01:52:53,851 WE MAPPED OUR CLUSTER WITH THE 2761 01:52:53,851 --> 01:52:55,152 CLUSTER SHE HAD DEFINED AND THE 2762 01:52:55,152 --> 01:52:59,123 CLUSTER THAT CONTAINS CYP11B1 IS 2763 01:52:59,123 --> 01:53:01,125 DIFFERENT, THE NEUROENDOCRINE 2764 01:53:01,125 --> 01:53:04,128 CLUSTERS OR THE DIFFERENT TISSUE 2765 01:53:04,128 --> 01:53:05,629 SHE FOUND SO IT SEEMS LIKE IT 2766 01:53:05,629 --> 01:53:07,832 DOESN'T CLUSTER WITH HERS, BUT 2767 01:53:07,832 --> 01:53:09,834 THEN WE'RE LOOKING AT, WE SEE 2768 01:53:09,834 --> 01:53:12,136 THIS IN MTEC HIGHS AND SHE'S 2769 01:53:12,136 --> 01:53:14,872 REALLY LOOKING AT MTEC LOWS, A 2770 01:53:14,872 --> 01:53:16,173 POPULATION THAT'S NOT AS 2771 01:53:16,173 --> 01:53:17,475 RELEVANT HERE FOR 2772 01:53:17,475 --> 01:53:17,842 GLUCOCORTICOIDS. 2773 01:53:17,842 --> 01:53:19,243 >> BUT DID YOU LOOK AT LIKE 2774 01:53:19,243 --> 01:53:24,281 TRANSCRIPTION FACTORS THAT ARE 2775 01:53:24,281 --> 01:53:30,454 SPECIFIC FOR -- THE WAY SHE 2776 01:53:30,454 --> 01:53:30,788 IDENTIFIES -- 2777 01:53:30,788 --> 01:53:33,190 >> I'M SAYING WE GO DO ON THE 2778 01:53:33,190 --> 01:53:35,025 SINGLE CELL SEQ WE DID, YES, AND 2779 01:53:35,025 --> 01:53:37,328 THERE'S NO OVERLAP WITH THOSE 2780 01:53:37,328 --> 01:53:38,229 POPULATIONS, WITH HER GROUPS 2781 01:53:38,229 --> 01:53:39,430 THAT SHE HAD DEFINED. 2782 01:53:39,430 --> 01:53:40,631 IT'S A NEW, UNIQUE GROUP. 2783 01:53:40,631 --> 01:53:44,435 >> THANK YOU. 2784 01:53:44,435 --> 01:53:45,870 >> HAVE YOU LOOKED TO SEE IF 2785 01:53:45,870 --> 01:53:50,040 THERE'S ANY CHANGES IN HAD T REG 2786 01:53:50,040 --> 01:53:51,308 POPULATIONS DEPENDENT ON 2787 01:53:51,308 --> 01:53:52,409 GLUCOCORTICOID LEVELS? 2788 01:53:52,409 --> 01:53:54,845 >> THAT'S A GREAT QUESTION, AND 2789 01:53:54,845 --> 01:53:57,348 WE THOUGHT THRMED BE -- AND WE 2790 01:53:57,348 --> 01:53:58,549 THOUGHT THERE WOULD BE, IN FACT, 2791 01:53:58,549 --> 01:53:59,784 WE DIDN'T. 2792 01:53:59,784 --> 01:54:02,286 THE T REGS, IN NUMBER THEY 2793 01:54:02,286 --> 01:54:04,021 AREN'T CHANGED. 2794 01:54:04,021 --> 01:54:06,223 WE DID NOT -- WE DID ONE 2795 01:54:06,223 --> 01:54:10,661 EXPERIMENT WHERE WE TRIED TO SEE 2796 01:54:10,661 --> 01:54:12,663 IF THE FUNCTION NORMALLY, WITH 2797 01:54:12,663 --> 01:54:16,167 THE ADOPTED COLLIDUS MODEL, THEY 2798 01:54:16,167 --> 01:54:18,135 LOOKED PRETTY NORMAL SO WE HAVE 2799 01:54:18,135 --> 01:54:18,702 NOT PURSUED THAT. 2800 01:54:18,702 --> 01:54:21,505 >> YOU ONLY LOOKED AT IN FOX # 2801 01:54:21,505 --> 01:54:21,772 POSITIVE? 2802 01:54:21,772 --> 01:54:24,475 >> YES, WE ONLY LOOKED AT FOX, 2803 01:54:24,475 --> 01:54:24,642 YES. 2804 01:54:24,642 --> 01:54:26,744 >> IT SOUNDS LIKE YOUR MUTANT 2805 01:54:26,744 --> 01:54:28,312 MICE MIGHT HAVE A MORE SEVERE 2806 01:54:28,312 --> 01:54:30,147 PHENOTYPE EARLIER IN LIFE. 2807 01:54:30,147 --> 01:54:35,553 HAVE YOU LOOKED IN NEONATAL 2808 01:54:35,553 --> 01:54:36,420 MICE? 2809 01:54:36,420 --> 01:54:37,221 >> IN WHAT REGARD? 2810 01:54:37,221 --> 01:54:40,691 >> WELL, YOU SAID THE CIRCLE A 2811 01:54:40,691 --> 01:54:45,729 AND GLUCOCORTICOIDS DON'T GO 2812 01:54:45,729 --> 01:54:47,565 THERE EARLY IN LIFE, RIGHT? 2813 01:54:47,565 --> 01:54:48,098 >> NELLER LIFE. 2814 01:54:48,098 --> 01:54:50,167 >> YOU SAID THAT THE 2815 01:54:50,167 --> 01:54:51,902 GLUCOCORTICOIDS ARE HIGHER IN 2816 01:54:51,902 --> 01:54:52,369 FETAL THYMUS. 2817 01:54:52,369 --> 01:54:53,137 >> RIGHT. 2818 01:54:53,137 --> 01:54:55,339 >> AS OPPOSED TO ADULT THYMUS. 2819 01:54:55,339 --> 01:54:57,641 >> IN THE THYMUS, YEAH. 2820 01:54:57,641 --> 01:54:59,944 >> THAT MADE ME THINK THAT THE 2821 01:54:59,944 --> 01:55:01,545 PHENOTYPE MIGHT BE MORE SEVERE 2822 01:55:01,545 --> 01:55:04,181 EARLIER IN LIFE AS OPPOSED ZIESH 2823 01:55:04,181 --> 01:55:09,920 THE PHENOTYPE WE'RE LOOKING 2824 01:55:09,920 --> 01:55:13,090 AT -- YOU WOULD HAVE TO LOOK AT, 2825 01:55:13,090 --> 01:55:14,191 TAKE UP -- 2826 01:55:14,191 --> 01:55:17,828 >> IF YOU LOOK UP THE 2827 01:55:17,828 --> 01:55:19,263 REPERTOIRE, FOR EXAMPLE. 2828 01:55:19,263 --> 01:55:20,731 >> WE HAVE LOOKED AT THYMUS AT 2829 01:55:20,731 --> 01:55:23,367 DIFFERENTIATION AND AS I SAID 2830 01:55:23,367 --> 01:55:25,336 QUALITATIVELY WE HAVE A SMALL 2831 01:55:25,336 --> 01:55:27,404 THYMUS, POPULATIONS FROM DOUBLE 2832 01:55:27,404 --> 01:55:29,740 POSITIVE ON DOWN ARE REDUCED, 2833 01:55:29,740 --> 01:55:31,242 OKAY, DOUBLE-NEGATIVES ARE 2834 01:55:31,242 --> 01:55:33,110 PRESERVED, WE SEE PERFECTLY 2835 01:55:33,110 --> 01:55:34,778 NORMAL DOUBLE-NEGATIVE 2836 01:55:34,778 --> 01:55:35,079 GENERATION. 2837 01:55:35,079 --> 01:55:37,047 SO WE THINK IT'S MOSTLY 2838 01:55:37,047 --> 01:55:40,351 AFFECTING ONLY TCR SIGNAL CELLS, 2839 01:55:40,351 --> 01:55:42,419 AND WE CAN, WE'VE ALSO STHEEN 2840 01:55:42,419 --> 01:55:46,357 THE MHC BACKGROUND MAKES A BIG 2841 01:55:46,357 --> 01:55:46,657 DIFFERENCE. 2842 01:55:46,657 --> 01:55:48,225 THE BACKGROUND HAPLOTYPE MAKES A 2843 01:55:48,225 --> 01:55:49,660 BIG DIFFERENCE. 2844 01:55:49,660 --> 01:55:52,363 MORE COMPLEX HAPLOTYPE THE 2845 01:55:52,363 --> 01:55:54,365 SMALLER THE THYMUS BECOMES, 2846 01:55:54,365 --> 01:55:57,635 BECAUSE WE THINK BECAUSE THERE'S 2847 01:55:57,635 --> 01:56:00,137 MORE CELL PEPTIDE AND MORE 2848 01:56:00,137 --> 01:56:01,572 DIVERSITY PEPTIDE AND MORE 2849 01:56:01,572 --> 01:56:02,773 RECEPTORS ARE GOING TO BE 2850 01:56:02,773 --> 01:56:03,040 OCCUPIED. 2851 01:56:03,040 --> 01:56:07,811 >> JOHN, IT'S A WONDERFUL STORY, 2852 01:56:07,811 --> 01:56:11,782 I'M GLAD IT'S BEEN RESURRECTED. 2853 01:56:11,782 --> 01:56:12,716 TWO QUICK QUESTIONS. 2854 01:56:12,716 --> 01:56:14,752 ONE IS, DO YOU KNOW WHEN YOU SEE 2855 01:56:14,752 --> 01:56:19,089 ALL THESE DIFFERENT GENES 2856 01:56:19,089 --> 01:56:20,958 EXPRESSED IN THE CORTICOSTEROID 2857 01:56:20,958 --> 01:56:22,459 PATHWAY, IS IT JUST THE GENES 2858 01:56:22,459 --> 01:56:23,827 EXPRESSED OR DO YOU KNOW IF ALL 2859 01:56:23,827 --> 01:56:25,229 THE PROTEINS ARE EXPRESSED AS 2860 01:56:25,229 --> 01:56:26,864 WELL ALONG THE WAY? 2861 01:56:26,864 --> 01:56:30,768 >> WELL, BY CYP11B1 REPORTER 2862 01:56:30,768 --> 01:56:32,436 IT'S A FUSION PROTEIN, RIGHT? 2863 01:56:32,436 --> 01:56:34,772 SO IF YOU SEE IN SCARLET YOU'RE 2864 01:56:34,772 --> 01:56:38,342 SEEING CYP1 <!0> B1 AND WE ALSO 2865 01:56:38,342 --> 01:56:41,645 KNOW THAT TRP THE CYP11B1 IS 2866 01:56:41,645 --> 01:56:43,347 FUNCTIONAL BECAUSE THE ADRENALS 2867 01:56:43,347 --> 01:56:44,682 ARE PERFECTLY FUNCTIONAL, 2868 01:56:44,682 --> 01:56:47,651 OTHERWISE THE MICE WOULD BE 2869 01:56:47,651 --> 01:56:50,254 HYPOADRENAL AND THEY'RE NOT. 2870 01:56:50,254 --> 01:56:51,121 CIRCULATING LEVELS ARE NORMAL, 2871 01:56:51,121 --> 01:56:52,489 STRESS RESPONSE IS NORMAL. 2872 01:56:52,489 --> 01:56:53,757 THAT'S IDENTIFYING THE PROTEIN. 2873 01:56:53,757 --> 01:56:54,091 OKAY? 2874 01:56:54,091 --> 01:56:57,328 LOOKING AT THE OTHER PROTEINS, 2875 01:56:57,328 --> 01:56:59,897 THERE ARE OTHER PEOPLE WHO HAVE 2876 01:56:59,897 --> 01:57:01,231 IDENTIFIED -- NOT MT HE C'S OF 2877 01:57:01,231 --> 01:57:02,733 COURSE BUT IN THE THYMUS SOME 2878 01:57:02,733 --> 01:57:05,936 PEOPLE YES HAVE BY ANTIBODY 2879 01:57:05,936 --> 01:57:08,572 MEDIATED MECHANISMS TECHNIQUES 2880 01:57:08,572 --> 01:57:09,740 HAVE SEEN SOME OF THE OTHERS 2881 01:57:09,740 --> 01:57:10,007 EXPRESSED. 2882 01:57:10,007 --> 01:57:13,677 >> THE OTHER QUESTION WAS, THE 2883 01:57:13,677 --> 01:57:13,978 ORIGINAL -- 2884 01:57:13,978 --> 01:57:16,547 >> YOU REALIZE THEY'RE MAKING 2885 01:57:16,547 --> 01:57:16,914 GLUCOCORTICOIDS. 2886 01:57:16,914 --> 01:57:18,515 SO GENES DON'T MAKE 2887 01:57:18,515 --> 01:57:20,184 GLUCOCORTICOIDS, JUST TO 2888 01:57:20,184 --> 01:57:21,585 CLARIFY, RESURRECT THAT NOTION. 2889 01:57:21,585 --> 01:57:24,755 OKAY. 2890 01:57:24,755 --> 01:57:25,322 >> THANKS. 2891 01:57:25,322 --> 01:57:26,490 SO ORIGINALLY AS I RECALL YOU 2892 01:57:26,490 --> 01:57:28,625 THOUGHT THAT THE GLUCOCORTICOIDS 2893 01:57:28,625 --> 01:57:30,861 MIGHT BE THE MECHANISM OF DEATH 2894 01:57:30,861 --> 01:57:33,864 BY NEGLECT. 2895 01:57:33,864 --> 01:57:35,132 IS THAT STILL THE CASE? 2896 01:57:35,132 --> 01:57:36,800 DO YOU THINK THAT IT'S -- 2897 01:57:36,800 --> 01:57:38,869 >> I CAN'T QUITE HEAR. 2898 01:57:38,869 --> 01:57:41,839 >> SO DO YOU KNOW IF THE 2899 01:57:41,839 --> 01:57:42,773 GLUCOCORTICOID -- ORIGINALLY YOU 2900 01:57:42,773 --> 01:57:44,041 THOUGHT THAT GLUCOCORTICOIDS 2901 01:57:44,041 --> 01:57:46,844 WERE THE INDUCERS OF DEATH BY 2902 01:57:46,844 --> 01:57:47,077 NEGLECT. 2903 01:57:47,077 --> 01:57:47,711 >> YES. 2904 01:57:47,711 --> 01:57:49,513 >> DO YOU STILL THINK THAT? 2905 01:57:49,513 --> 01:57:49,947 THAT THEY -- 2906 01:57:49,947 --> 01:57:51,015 >> NO. 2907 01:57:51,015 --> 01:57:51,181 NO. 2908 01:57:51,181 --> 01:57:53,117 THAT WAS DISPROVED LONG AGO. 2909 01:57:53,117 --> 01:57:55,052 BACK IN THE EARLY DAYS, WE KNEW 2910 01:57:55,052 --> 01:57:57,454 NOTHING ABOUT, YOU KNOW, ALL THE 2911 01:57:57,454 --> 01:57:59,123 MECHANISMS WE KNOW TODAY AND WE 2912 01:57:59,123 --> 01:58:02,292 THOUGHT CONCEIVABLY THAT 2913 01:58:02,292 --> 01:58:03,494 GLUCOCORTICOIDS WOULD -- WERE 2914 01:58:03,494 --> 01:58:05,029 RESPONSIBLE FOR THE DEATH OF 2915 01:58:05,029 --> 01:58:07,865 CELLS, THE UNAN TAG NIETZED BY 2916 01:58:07,865 --> 01:58:10,134 ARE TCR REGULATION AND THE 2917 01:58:10,134 --> 01:58:12,569 ANSWER IS NO THE GR KNOCK-OUT 2918 01:58:12,569 --> 01:58:13,771 THYMOCYTES HAVE NO CHANGES IN 2919 01:58:13,771 --> 01:58:15,139 DEATH BY NEGLECT. 2920 01:58:15,139 --> 01:58:16,407 AND WE'RE NOT THE FIRST ONE TO 2921 01:58:16,407 --> 01:58:19,143 SHOW THAT. 2922 01:58:19,143 --> 01:58:22,846 LAST QUESTION? 2923 01:58:22,846 --> 01:58:25,482 >> SO YOU SHOWED THAT IT'S A 2924 01:58:25,482 --> 01:58:27,818 PROBLEM IN THE TCR REPERTOIRE IN 2925 01:58:27,818 --> 01:58:31,622 RESPONSE TO STIMULATION, SO 2926 01:58:31,622 --> 01:58:32,289 POST-SELECTION, I GUESS. 2927 01:58:32,289 --> 01:58:34,324 THE REPORTER MOUSE SHOWED THAT 2928 01:58:34,324 --> 01:58:40,464 THE CYP11B1 IS EXPRESSED IN 2929 01:58:40,464 --> 01:58:42,132 APPROXIMATE MINOR -- HOW DOES 2930 01:58:42,132 --> 01:58:43,233 THAT AFFECT POSITIVE SELECTION? 2931 01:58:43,233 --> 01:58:44,868 >> WE DON'T THINK IT HAS 2932 01:58:44,868 --> 01:58:46,837 ANYTHING TO DO WITH POSITIVE 2933 01:58:46,837 --> 01:58:47,104 SELECTION. 2934 01:58:47,104 --> 01:58:48,405 IN THE CORTEX, WE DON'T THINK 2935 01:58:48,405 --> 01:58:49,473 IT'S GOT TO DO WITH POSITIVE 2936 01:58:49,473 --> 01:58:51,208 SELECTION OF THE CORTEX. 2937 01:58:51,208 --> 01:58:52,943 WE THINK IT HAS TO DO WITH THE 2938 01:58:52,943 --> 01:58:55,145 RES RESCUE OF CELLS THAT WOULD 2939 01:58:55,145 --> 01:58:56,113 UNDERGO NEGATIVE SELECTION GOO 2940 01:58:56,113 --> 01:58:58,315 CELLS FOR ANTIGEN UNDERGO 2941 01:58:58,315 --> 01:59:00,050 NEGATIVE SELECTION IF THE 2942 01:59:00,050 --> 01:59:01,585 GLUCOCORTICOID SIGNALLING IS NOT 2943 01:59:01,585 --> 01:59:01,919 THERE? 2944 01:59:01,919 --> 01:59:05,322 >> THE IDEA IS THAT YOU EXPRESS 2945 01:59:05,322 --> 01:59:12,996 IT -- AND THAT T CELLS THAT 2946 01:59:12,996 --> 01:59:14,731 RECOGNIZE THOSE ANTIGENS, THAT 2947 01:59:14,731 --> 01:59:15,999 MADE IT THROUGH THE CORTEX, 2948 01:59:15,999 --> 01:59:17,901 THEY'RE STILL AROUND, THEY'RE 2949 01:59:17,901 --> 01:59:20,337 GOING TO BE DELETED BUT THEY'RE 2950 01:59:20,337 --> 01:59:23,040 RESCUED BY THE GLUCOCORTICOIDS. 2951 01:59:23,040 --> 01:59:24,475 AND THAT EXPANDS THE REPERTOIRE 2952 01:59:24,475 --> 01:59:26,577 TO INCLUDE ALL THOSE CELLS THAT 2953 01:59:26,577 --> 01:59:28,445 NOW SEE ALL THE DIFFERENT TRA'S, 2954 01:59:28,445 --> 01:59:30,848 THAT'S THE NOTION. 2955 01:59:30,848 --> 01:59:32,483 AND WORK DIDN'T HAVE A CHANCE TO 2956 01:59:32,483 --> 01:59:34,685 SHOW BUT AT THE TIME IS THAT 2957 01:59:34,685 --> 01:59:36,086 MATT ACTUALLY DEVELOPED AN ASSAY 2958 01:59:36,086 --> 01:59:38,055 WHERE WE COULD ACTUALLY MEASURE 2959 01:59:38,055 --> 01:59:39,923 GR INTERACTIONS WITH CHROMATIN, 2960 01:59:39,923 --> 01:59:41,692 SO THAT'S THE SIGNAL CELLS AND 2961 01:59:41,692 --> 01:59:43,193 WE SHOWED THE ONLY CELLS THAT 2962 01:59:43,193 --> 01:59:46,497 ARE RECEIVING THE SIGNAL ARE 2963 01:59:46,497 --> 01:59:49,366 THIS SIGNAL AMONG THE THYMIC 2964 01:59:49,366 --> 01:59:51,101 THYMOCYTES POPULATION ARE THE 2965 01:59:51,101 --> 01:59:52,669 TCR HIGH DOUBLE POSITIVES, THE 2966 01:59:52,669 --> 01:59:54,838 TRIGGER DOUBLE POSITIVES. 2967 01:59:54,838 --> 01:59:55,072 OKAY? 2968 01:59:55,072 --> 01:59:57,474 TO SOME EXTENT POST THAT. 2969 01:59:57,474 --> 01:59:59,543 SO THESE ARE VERY LOCALIZED 2970 01:59:59,543 --> 02:00:04,047 DELIVER RIFF GLUCOCORTICOIDS 2971 02:00:04,047 --> 02:00:07,651 FROM THE FROM THE PRESENTING 2972 02:00:07,651 --> 02:00:09,153 MTEC'S SO RESPONGE ON THE 2973 02:00:09,153 --> 02:00:09,386 SURFACE. 2974 02:00:09,386 --> 02:00:11,054 >> AND -- DOES ANYTHING THERE. 2975 02:00:11,054 --> 02:00:13,223 >> SORRY, WE'RE BEING TOLD IT'S 2976 02:00:13,223 --> 02:00:13,991 TIME FOR A BREAK. 2977 02:00:13,991 --> 02:00:14,391 >> OKAY. 2978 02:00:14,391 --> 02:00:15,626 >> OKAY, THANK YOU MPLETS. 2979 02:00:15,626 --> 02:00:16,960 [APPLAUSE] 2980 02:00:16,960 --> 02:00:17,895 >> THANK YOU. 2981 02:00:17,895 --> 02:00:20,464 [APPLAUSE] 2982 02:00:20,464 --> 02:00:22,232 >> OKAY RS WE'LL HAVE A 20 2983 02:00:22,232 --> 02:00:23,133 MINUTE BREAK AND WE'LL START 2984 02:00:23,133 --> 02:00:27,867 WITH THE NEXT TALK AT 10:50. 2985 02:00:27,867 --> 02:00:29,602 SO WE'LL GET STARTED IN THIS 2986 02:00:29,602 --> 02:00:31,471 SECOND PART OF THE MORNING 2987 02:00:31,471 --> 02:00:33,139 SESSION WITH DR. DINAH SINGER 2988 02:00:33,139 --> 02:00:39,245 FROM THE NCI TO TELL US ABOUT 2989 02:00:39,245 --> 02:00:43,383 THE DEVELOPMENT AND ROLE OF THE 2990 02:00:43,383 --> 02:00:44,584 FACTOR BRD4. 2991 02:00:44,584 --> 02:00:46,653 >> THANK YOU, STANLEY. SO IF I 2992 02:00:46,653 --> 02:00:48,321 RESIGN, DIE GET AN EXTRA FIVE 2993 02:00:48,321 --> 02:00:48,688 MINUTES? 2994 02:00:48,688 --> 02:00:49,222 [LAUGHTER] 2995 02:00:49,222 --> 02:00:49,422 OKAY. 2996 02:00:49,422 --> 02:00:53,359 IT'S A DEAL. 2997 02:00:53,359 --> 02:00:55,295 I WANT TO START BY THANKING THE 2998 02:00:55,295 --> 02:00:56,763 ORGANIZERS TOIN VOOTING ME TO 2999 02:00:56,763 --> 02:00:58,064 SPEAK, I'M DELIGHTED TO BE ABLE 3000 02:00:58,064 --> 02:00:59,399 TO TELL YOU ABOUT SOME OF OUR 3001 02:00:59,399 --> 02:01:01,901 RECENT STUDIES ON THE REGULATION 3002 02:01:01,901 --> 02:01:04,637 OF THYMIC DEVELOPMENT BY THE 3003 02:01:04,637 --> 02:01:07,273 TRANSCRIPTION FACTOR BRD4. 3004 02:01:07,273 --> 02:01:09,976 SO WHY BRD4? 3005 02:01:09,976 --> 02:01:12,178 BRD4 HAS BEEN STUDIED FOR SOME 3006 02:01:12,178 --> 02:01:17,216 TIME NOW AS AN ACTIVE REGULATOR 3007 02:01:17,216 --> 02:01:18,885 IN INFLAMMATORY DISEASE AS IN T 3008 02:01:18,885 --> 02:01:20,920 CELL FUNCTION, AND IT'S BEEN 3009 02:01:20,920 --> 02:01:23,122 PARTICULARLY STUDIED AS A DRIVER 3010 02:01:23,122 --> 02:01:24,991 IN A VARIETY OF DIFFERENT BOTH 3011 02:01:24,991 --> 02:01:26,726 LIQUID AND SOLID TUMORS, SO IT'S 3012 02:01:26,726 --> 02:01:29,362 GOTTEN A LOT OF ATTENTION AS A 3013 02:01:29,362 --> 02:01:32,231 POSSIBLE THERAPEUTIC TARGET. 3014 02:01:32,231 --> 02:01:34,734 FROM THE BIOCHEMICAL STANDPOINT, 3015 02:01:34,734 --> 02:01:37,904 BRD4 IS A DOMAIN PROTEIN WE'VE 3016 02:01:37,904 --> 02:01:40,340 SHOWN OVER THE YEARS HAS 3017 02:01:40,340 --> 02:01:42,408 PLEOTROPIC FUNCTIONS, REGULATES 3018 02:01:42,408 --> 02:01:44,177 CHROMATIN STRUCTURE THROUGH ITS 3019 02:01:44,177 --> 02:01:45,445 INTRINSIC TRANSFERASE ACTIVITY 3020 02:01:45,445 --> 02:01:47,814 HA REMODELS CHROMATIN, IT 3021 02:01:47,814 --> 02:01:48,581 REGULATES TRANSCRIPTION THROUGH 3022 02:01:48,581 --> 02:01:53,653 ITS KINASE ACTIVITY WHERE IT 3023 02:01:53,653 --> 02:01:56,589 PHOSPHORYLATES POLYMERASE 2 AND 3024 02:01:56,589 --> 02:01:58,925 NIC EXPREENTLY SHOWED THAT BRD4 3025 02:01:58,925 --> 02:02:00,460 REGULATES RNA SPLICING AND IN 3026 02:02:00,460 --> 02:02:01,661 SPITE OF ALL THE WORK THAT'S 3027 02:02:01,661 --> 02:02:03,763 BEEN DONE ON BRD4, VERY LITTLE 3028 02:02:03,763 --> 02:02:06,265 IS KNOWN ABOUT ITS ROLE IN 3029 02:02:06,265 --> 02:02:09,335 DEVELOPMENT AND DIFFERENTIATION 3030 02:02:09,335 --> 02:02:10,269 GENERALLY, BUT PARTICULARLY IN 3031 02:02:10,269 --> 02:02:11,104 THE THYMUS. 3032 02:02:11,104 --> 02:02:15,341 SO WE WERE INTERESTED IN ASKING 3033 02:02:15,341 --> 02:02:18,077 DOES BRD4 HAVE AN ACTIVE ROLE IN 3034 02:02:18,077 --> 02:02:19,278 THYMIC DIFFERENTIATION, AND I'M 3035 02:02:19,278 --> 02:02:21,147 GOING TO LEAVE YOU WITH THREE 3036 02:02:21,147 --> 02:02:22,582 TAKE-HOME MESSAGES IN CASE 3037 02:02:22,582 --> 02:02:25,652 STANLEY CUTS ME OFF. 3038 02:02:25,652 --> 02:02:29,255 ONE IS, THE FIRST ONE IS THAT IN 3039 02:02:29,255 --> 02:02:31,357 SINGLE POSITIVE THYMOCYTES -- 3040 02:02:31,357 --> 02:02:33,226 IMMATURE POSITIVE THYMOCYTES ARE 3041 02:02:33,226 --> 02:02:38,665 UNIQUELY DEPENDENT ON BRD4 FOR 3042 02:02:38,665 --> 02:02:41,534 THEIR DIFFERENTIATION, THAT BRD4 3043 02:02:41,534 --> 02:02:42,268 REGULATES ALTERNATIVE RNA 3044 02:02:42,268 --> 02:02:43,970 SPLICING IN ALL THYMOCYTE 3045 02:02:43,970 --> 02:02:45,471 SUBSETS AND POSITIVE SELECTION 3046 02:02:45,471 --> 02:02:47,040 OF DOUBLE POSITIVE THYMOCYTES IS 3047 02:02:47,040 --> 02:02:48,274 UNIQUELY DEPENDENT ON BRD4 3048 02:02:48,274 --> 02:02:50,910 EXPRESSION IN THYMIC EPITHELIAL 3049 02:02:50,910 --> 02:02:51,210 CELLS. 3050 02:02:51,210 --> 02:02:54,180 SO WE'VE SEEN VARIOUS CARTOONS 3051 02:02:54,180 --> 02:02:58,718 OF THIS, THIS MORNING ALREADY, 3052 02:02:58,718 --> 02:03:02,488 OF THE SEQUENCE OF THYMIC 3053 02:03:02,488 --> 02:03:04,323 DIFFERENTIATION WHICH GOES IN 3054 02:03:04,323 --> 02:03:06,526 WELL-DEFINED SERIES OF STEPS, 3055 02:03:06,526 --> 02:03:09,395 STARTING WITH THE ARRIVAL OF THE 3056 02:03:09,395 --> 02:03:10,997 IMMATURE THYMOCYTES INTO THE 3057 02:03:10,997 --> 02:03:13,933 THYMUS THAT EXPRESS NEITHER SOAR 3058 02:03:13,933 --> 02:03:15,735 NOR CD8, THE DOUBLE-NEGATIVES 3059 02:03:15,735 --> 02:03:19,772 THAT THEN UNDERGO -- NEITHER CD4 3060 02:03:19,772 --> 02:03:22,041 NOR CD8, THE DOUBLE-NEGATIVES 3061 02:03:22,041 --> 02:03:24,310 THAT UNDERGO PROCEED LIFT UNITED 3062 02:03:24,310 --> 02:03:29,282 NATIONS DURING WHICH ARE 3063 02:03:29,282 --> 02:03:31,184 EXPRESSED, AND FINALLY THEIR 3064 02:03:31,184 --> 02:03:32,385 SIGNAL TO EXPRESS CD8 ON THE 3065 02:03:32,385 --> 02:03:34,454 SURFACE AND BECOME WHAT ARE 3066 02:03:34,454 --> 02:03:35,555 CALLED MAIM TOUR SINGLE 3067 02:03:35,555 --> 02:03:37,223 POSITIVES THAT EXPRESS CD8 ONLY. 3068 02:03:37,223 --> 02:03:39,492 THESE ARE TRANSITIONAL CELLS 3069 02:03:39,492 --> 02:03:41,728 BETWEEN THE VERY PROLIFERATIVE 3070 02:03:41,728 --> 02:03:42,895 DOUBLE-NEGATIVE AND THE 3071 02:03:42,895 --> 02:03:44,397 QUIESCENT DOUBLE POSITIVE, AND 3072 02:03:44,397 --> 02:03:47,500 THEY HAVE TO JUSHED GO -- 3073 02:03:47,500 --> 02:03:49,469 UNDERGO ONE FINAL ROUND OF CELL 3074 02:03:49,469 --> 02:03:51,003 DIVISION IN ORDER TO BECOME 3075 02:03:51,003 --> 02:03:52,905 DOUBLE POSITIVES THAT THEN GO ON 3076 02:03:52,905 --> 02:03:55,041 TO MATURE TO BECOME EITHER THE 3077 02:03:55,041 --> 02:03:57,443 CD4 OR THE CD8 SINGLE POSITIVES. 3078 02:03:57,443 --> 02:03:58,911 SO THE QUESTION THAT WE WERE 3079 02:03:58,911 --> 02:04:00,980 INTERESTED IN ASKING IS, WHAT IS 3080 02:04:00,980 --> 02:04:04,016 THE ROLE OF BRD4, IF ANY, IN 3081 02:04:04,016 --> 02:04:05,785 EACH THESE STEPS MANY ONE OF THE 3082 02:04:05,785 --> 02:04:08,621 REASONS FOR ASKING THIS QUESTION 3083 02:04:08,621 --> 02:04:11,124 WAS THAT BRD4 IS ACTUALLY 3084 02:04:11,124 --> 02:04:12,892 EXPRESSED THROUGHOUT THYMIC 3085 02:04:12,892 --> 02:04:13,960 DIFFERENTIATION, SO WE WANTED TO 3086 02:04:13,960 --> 02:04:18,131 KNOW WHAT ROLE IT PLAYS IN EACH 3087 02:04:18,131 --> 02:04:18,831 THOSE STEPS. 3088 02:04:18,831 --> 02:04:27,440 TO DO THAT, WE CONDITIONED -- 3089 02:04:27,440 --> 02:04:28,241 CONDITIONALLY DELETED AS EARLY 3090 02:04:28,241 --> 02:04:30,576 AS WE CAN TODAY, AND THAT IS 3091 02:04:30,576 --> 02:04:35,348 WITH LCK KCRE WHICH DELETES 3092 02:04:35,348 --> 02:04:36,682 AROUND 29 STAGE AND FOLLOW WHAT 3093 02:04:36,682 --> 02:04:38,017 HAPPENS TO BRD4 EXPRESSION AND 3094 02:04:38,017 --> 02:04:41,053 YOU CAN SEE IT'S ALREADY ACUTELY 3095 02:04:41,053 --> 02:04:42,922 DEPLETED IN THE DN STAGE 3096 02:04:42,922 --> 02:04:45,758 COMPLETELY DELETED BY THE ISP'S 3097 02:04:45,758 --> 02:04:47,994 AND CONTINUES TO BE COMPLETELY 3098 02:04:47,994 --> 02:04:49,395 DELETED FOR THE REST OF THE 3099 02:04:49,395 --> 02:04:50,396 DEVELOPMENTAL SEQUENCE. 3100 02:04:50,396 --> 02:04:52,231 SO WITH THAT IN MIND, WE ASKED, 3101 02:04:52,231 --> 02:04:54,667 WHAT IS THE EFFECT OF THIS 3102 02:04:54,667 --> 02:04:56,736 DELETION ON THYMIC 3103 02:04:56,736 --> 02:04:57,103 DIFFERENTIATION? 3104 02:04:57,103 --> 02:05:00,339 AND WHEN WE LOOKED AT CD4 CD8 3105 02:05:00,339 --> 02:05:01,340 PROFILES WHAT WE WERE ABLE TO 3106 02:05:01,340 --> 02:05:03,810 SEE WAS AN ACUTE DEPLETION IN 3107 02:05:03,810 --> 02:05:05,611 THE NUMBER OF DOUBLE POSITIVE 3108 02:05:05,611 --> 02:05:07,580 CELLS AND FREQUENCY OF DOUBLE 3109 02:05:07,580 --> 02:05:09,482 POSITIVE CELLS AND AN INCREASE 3110 02:05:09,482 --> 02:05:11,717 IN THE CD8 SINGLE POSITIVE 3111 02:05:11,717 --> 02:05:12,051 CELLS. 3112 02:05:12,051 --> 02:05:14,020 THE CELLS THAT ARE IN THIS 3113 02:05:14,020 --> 02:05:15,888 COMPARTMENT ARE BOTH THE MATURE 3114 02:05:15,888 --> 02:05:19,192 CD8 SINGLE POSITIVES BUT ALSO 3115 02:05:19,192 --> 02:05:21,594 THE IMMATURE CD8 SINGLE 3116 02:05:21,594 --> 02:05:23,896 POSITIVES, THOSE TRANSITIONAL 3117 02:05:23,896 --> 02:05:24,096 CELLS. 3118 02:05:24,096 --> 02:05:25,932 THEY CAN BE DISTINGUISHED BASED 3119 02:05:25,932 --> 02:05:28,234 ON THEIR EXPRESSION OF TCR BETA 3120 02:05:28,234 --> 02:05:30,002 ON THE SURFACE, AND WHEN YOU 3121 02:05:30,002 --> 02:05:32,738 LOOK AT TCR BETA EXPRESSION IN 3122 02:05:32,738 --> 02:05:34,807 THE CD8 QUADRANT, WHAT YOU SEE 3123 02:05:34,807 --> 02:05:36,108 HERE IN THE LIGHT GRAY IS THAT 3124 02:05:36,108 --> 02:05:39,178 THE WILD-TYPE CELLS 3125 02:05:39,178 --> 02:05:41,914 PREDOMINANTLY EXPRESS THE MATURE 3126 02:05:41,914 --> 02:05:45,084 CD8 SINGLE POSITIVES THAT ARE 3127 02:05:45,084 --> 02:05:46,686 EXPRESSED TCR BETA WHEREAS IN 3128 02:05:46,686 --> 02:05:47,920 THE KNOCK-OUT MOST OF THE CD8 3129 02:05:47,920 --> 02:05:50,256 CELLS THAT ARE IN THIS QUADRANT 3130 02:05:50,256 --> 02:05:52,525 DO NOT EXPRESS TCR BETA AND 3131 02:05:52,525 --> 02:05:53,960 THEREFORE ARE THE IMMATURE 3132 02:05:53,960 --> 02:05:57,463 SINGLE POSITIVES, THOSE 3133 02:05:57,463 --> 02:05:59,532 TRANSITIONAL CELLS. 3134 02:05:59,532 --> 02:06:01,500 WHEN WE QUANTITATE WE SEE THAT 3135 02:06:01,500 --> 02:06:02,602 THE DOUBLE-NEGATIVE POPULATIONS 3136 02:06:02,602 --> 02:06:04,136 ARE NOT AFFECTED BUT THERE IS A 3137 02:06:04,136 --> 02:06:05,238 HUGE INCREASE IN THE NUMBER OF 3138 02:06:05,238 --> 02:06:07,740 THIS ISP'S THAT ACCUMULATE IN 3139 02:06:07,740 --> 02:06:10,543 THE ABSENCE OF BRD4 AND OF 3140 02:06:10,543 --> 02:06:11,878 COURSE THAT THEN IS PROPAGATED 3141 02:06:11,878 --> 02:06:13,779 INTO THE DOUBLE POSITIVES AND 3142 02:06:13,779 --> 02:06:16,382 THE SINGLE POSITIVES BUT TELLS 3143 02:06:16,382 --> 02:06:19,352 US THAT BRD4 DELETION ARRESTS 3144 02:06:19,352 --> 02:06:22,054 THE TRANSITION OF ISP'S TO THE 3145 02:06:22,054 --> 02:06:27,360 DOUBLE POSITIVE STAGE. 3146 02:06:27,360 --> 02:06:29,862 I MENTIONED EARLIER THAT FOR AN 3147 02:06:29,862 --> 02:06:32,398 ISP TO MATURE TO A DP IT HAS TO 3148 02:06:32,398 --> 02:06:34,133 UNDERGO ONE LAST ROUND OF CELL 3149 02:06:34,133 --> 02:06:35,534 CYCLE AND ASKED IF PART OF THE 3150 02:06:35,534 --> 02:06:37,069 DEFECT WE'RE SEEING HERE IS THE 3151 02:06:37,069 --> 02:06:38,404 INABILITY OF THE CELLS TO 3152 02:06:38,404 --> 02:06:40,373 UNDERGO THAT LAST CELL CYCLE. 3153 02:06:40,373 --> 02:06:41,707 TO ANSWER THAT QUESTION, WE 3154 02:06:41,707 --> 02:06:44,977 TURNED ON AN IN VITRO SYSTEM IN 3155 02:06:44,977 --> 02:06:47,513 WHICH YOU CAN TAKE PURIFIED 3156 02:06:47,513 --> 02:06:49,715 ISP'S, CULTURE THEM IN VITRO 3157 02:06:49,715 --> 02:06:52,018 OVERNIGHT AND NOW THEY GO FROM 3158 02:06:52,018 --> 02:06:53,886 BEING ENTIRELY CD8 SINGLE 3159 02:06:53,886 --> 02:06:55,955 POSITIVE TO BECOMING DOUBLE 3160 02:06:55,955 --> 02:06:57,590 POSITIVE AND DOUBLE IN CELL 3161 02:06:57,590 --> 02:06:57,823 NUMBER. 3162 02:06:57,823 --> 02:07:00,326 WHEN YOU DO THAT NOW WITH THE 3163 02:07:00,326 --> 02:07:02,962 BRD4 KNOCK-OUT, TAKE PURIFIED 3164 02:07:02,962 --> 02:07:07,633 ISP'S, WHAT YOU SEE IS THAT THEY 3165 02:07:07,633 --> 02:07:08,634 HAVE REAL DIFFICULTY IN 3166 02:07:08,634 --> 02:07:10,269 UNDERGOING THAT MATURATION AND 3167 02:07:10,269 --> 02:07:12,571 THAT CELL DWIKS, SO THAT THE 3168 02:07:12,571 --> 02:07:14,640 DEFECT HERE IS IN 3169 02:07:14,640 --> 02:07:16,309 DIFFERENTIATION AND 3170 02:07:16,309 --> 02:07:19,045 PROLIFERATION DURING IN VITRO 3171 02:07:19,045 --> 02:07:19,612 CULTURE. 3172 02:07:19,612 --> 02:07:20,546 NOW, WE CAN LOOK AT THE 3173 02:07:20,546 --> 02:07:22,548 MOLECULAR BASIS FOR THIS DEFECT 3174 02:07:22,548 --> 02:07:26,352 BY DOING RNA SE E QUANL CIS -- 3175 02:07:26,352 --> 02:07:27,920 SEQ ANALYSIS OF THE THYMOCYTES 3176 02:07:27,920 --> 02:07:30,656 AND WE DO THAT AND WE SEE 3177 02:07:30,656 --> 02:07:32,058 SURPRISINGLY IS VERY LITTLE 3178 02:07:32,058 --> 02:07:34,593 EFFECT ON RNA SEQ PROFILES OF 3179 02:07:34,593 --> 02:07:38,864 EITHER THE DN'S OR DP 3180 02:07:38,864 --> 02:07:40,933 POPULATIONS WHEN MSZ BRD4 IS 3181 02:07:40,933 --> 02:07:43,369 DELETED BUT YOU SEE ENORMOUS 3182 02:07:43,369 --> 02:07:45,771 CHANGE IN THE PROFILES WHEN YOU 3183 02:07:45,771 --> 02:07:47,640 DELETE BRD4 FROM THE ISP'S. 3184 02:07:47,640 --> 02:07:49,375 WHEN WE DO PATHWAY ANALYSIS, 3185 02:07:49,375 --> 02:07:52,345 MAJOR PATHWAYS AFFECTED ARE NIC 3186 02:07:52,345 --> 02:07:54,747 PATHWAYS AND GLYCOLYTIC PATHWAYS 3187 02:07:54,747 --> 02:07:56,182 AND WE'VE VALIDATED BOTH OF 3188 02:07:56,182 --> 02:07:58,150 THOSE PATHWAYS EXPERIMENTALLY 3189 02:07:58,150 --> 02:08:00,453 AND I'M SHOWING YOU HERE JUST 3190 02:08:00,453 --> 02:08:04,690 NIC IT'S AN IG -- MYC IT'S A 3191 02:08:04,690 --> 02:08:05,691 BROWSER VIEW LOOKING AT 3192 02:08:05,691 --> 02:08:07,660 EXPRESSION OF MYC IN WILD-TYPE 3193 02:08:07,660 --> 02:08:09,628 OR KNOCK-OUT ISP'S AND SEE THE 3194 02:08:09,628 --> 02:08:11,497 COMPLETE LOSS OF MYC WHEN YOU 3195 02:08:11,497 --> 02:08:12,598 LOSE BRD4. 3196 02:08:12,598 --> 02:08:13,899 WHAT'S REALLY SURPRISING THOUGH 3197 02:08:13,899 --> 02:08:16,635 IS YOU DON'T LOSE MYC IN THE 3198 02:08:16,635 --> 02:08:18,070 DOUBLE-NEGATIVES OR IN ANY OF 3199 02:08:18,070 --> 02:08:19,472 THE SINGLE POSITIVE. 3200 02:08:19,472 --> 02:08:22,541 SO BRD4 SELECTIVELY REGULATING 3201 02:08:22,541 --> 02:08:25,378 MYC IN THE IS P'S. 3202 02:08:25,378 --> 02:08:29,448 -- IN THE ISP'S. 3203 02:08:29,448 --> 02:08:31,384 FROM THESE STUDIES, WHAT WE 3204 02:08:31,384 --> 02:08:34,587 CONCLUDE IS THAT THE ISP 3205 02:08:34,587 --> 02:08:36,789 POPULATION IS UNIQUELY DEPENDENT 3206 02:08:36,789 --> 02:08:38,391 ON BRD4. 3207 02:08:38,391 --> 02:08:41,260 IN THE EARLY DN STAGES, 3208 02:08:41,260 --> 02:08:42,995 PROLIFERATION AND 3209 02:08:42,995 --> 02:08:44,964 DIFFERENTIATION OF BRD4 3210 02:08:44,964 --> 02:08:47,266 INDEPENDENT BASED ON THE RNA 3211 02:08:47,266 --> 02:08:48,701 PROFILING AND ALSO OTHER DATA 3212 02:08:48,701 --> 02:08:51,170 THAT I HAVEN'T SHOWN YOU, WE 3213 02:08:51,170 --> 02:08:53,105 KNOW THAT ONCE CELLS BECOME DP 3214 02:08:53,105 --> 02:08:56,275 AND BEYOND, THEY NOW BECOME BRD4 3215 02:08:56,275 --> 02:08:57,576 INDEPENDENT AND IT'S ONLY IN 3216 02:08:57,576 --> 02:09:00,546 THIS ISP STAGE THAT THE CELLS 3217 02:09:00,546 --> 02:09:03,149 ARE BRD4 DEPENDENT FOR BOTH CELL 3218 02:09:03,149 --> 02:09:07,953 CYCLE AND DIFFERENTIATION. 3219 02:09:07,953 --> 02:09:09,955 SO WE FOUND THAT TO BE A VERY 3220 02:09:09,955 --> 02:09:11,190 INTERESTING OBSERVATION, QUITE 3221 02:09:11,190 --> 02:09:12,691 SURPRISING, BUT WE WERE ALSO 3222 02:09:12,691 --> 02:09:16,095 PUZZLED BY THE FACT THAT BRD4 IS 3223 02:09:16,095 --> 02:09:18,631 NOW EXPRESSED IN ALL OF THE CELL 3224 02:09:18,631 --> 02:09:21,233 TYPES EVEN THOUGH WE DON'T SEE A 3225 02:09:21,233 --> 02:09:23,869 DEPENDENCE AT LEAST AS ASSESSED 3226 02:09:23,869 --> 02:09:26,205 BY RNA PROFILING. 3227 02:09:26,205 --> 02:09:28,340 SO WE WONDERED, IS IT POSSIBLE 3228 02:09:28,340 --> 02:09:30,876 THAT BRD4 IS REGULATING 3229 02:09:30,876 --> 02:09:32,745 ALTERNATIVE SPLICING IN THESE 3230 02:09:32,745 --> 02:09:34,146 THYMOCYTE POPULATIONS? 3231 02:09:34,146 --> 02:09:35,147 WHERE YOU WOULDN'T NECESSARILY 3232 02:09:35,147 --> 02:09:37,583 SEE A CHANGE IN THE OVERALL RNA 3233 02:09:37,583 --> 02:09:39,985 PROFILE, BUT YOU MIGHT SEE 3234 02:09:39,985 --> 02:09:42,288 CHANGES IN SPLICING. 3235 02:09:42,288 --> 02:09:44,090 SO WE WENT BACK AND REANALYZED 3236 02:09:44,090 --> 02:09:46,158 OUR DATA TO ASK THAT SPECIFIC 3237 02:09:46,158 --> 02:09:50,629 QUESTION AND IN FACT OF WE FIND 3238 02:09:50,629 --> 02:09:52,364 THAT BRD4 REGULATES ALTERNATIVE 3239 02:09:52,364 --> 02:09:53,999 SPLICING IN ALL LIKE SIGHT 3240 02:09:53,999 --> 02:09:55,134 SUBSETS AND WHAT THE PIE CHART 3241 02:09:55,134 --> 02:09:58,404 IS SHOWING YOU HERE IS THE 3242 02:09:58,404 --> 02:09:59,605 COMPARISON OF THE SPLICING 3243 02:09:59,605 --> 02:10:01,240 PATTERNS IN THE WILD-TYPE AND 3244 02:10:01,240 --> 02:10:03,976 BRD4 KNOCK-OUT ASKING WHAT 3245 02:10:03,976 --> 02:10:04,643 FRACTION DIFFER. 3246 02:10:04,643 --> 02:10:06,278 AND YOU CAN SEE IT'S A VERY 3247 02:10:06,278 --> 02:10:08,447 LARGE FRACTION, IT'S HIGHEST IN 3248 02:10:08,447 --> 02:10:11,851 ISP'S, MAYBE NOT SURPRISING 3249 02:10:11,851 --> 02:10:17,323 THERE BECAUSE -- YOU HAVE LARGE 3250 02:10:17,323 --> 02:10:18,724 SCALE DIFFERENCE IN THE PATTERNS 3251 02:10:18,724 --> 02:10:20,693 OF ALTERNATIVE SPLICING, WHICH 3252 02:10:20,693 --> 02:10:24,296 NOW ARE GOING TO BE REFLECTED IN 3253 02:10:24,296 --> 02:10:25,197 DIFFERENCES IN THE PROTEIN 3254 02:10:25,197 --> 02:10:28,601 PROFILE OF THOSE CELLS. 3255 02:10:28,601 --> 02:10:30,769 IN FACT, WHEN WE LOOK TO SEE 3256 02:10:30,769 --> 02:10:32,972 WHAT THE GENES MOST AFFECTED BY 3257 02:10:32,972 --> 02:10:34,773 ALTERNATIVE SPLICING IN BRD4 3258 02:10:34,773 --> 02:10:37,042 DEFICIENT THYMOCYTES H MANY OF 3259 02:10:37,042 --> 02:10:38,544 THOSE GENES AND THIS IS ONLY A 3260 02:10:38,544 --> 02:10:40,980 PARTIAL LIST ARE SOMEHOW RELATED 3261 02:10:40,980 --> 02:10:43,949 WITH IMMUNE RESPONSES. 3262 02:10:43,949 --> 02:10:45,017 EVERYTHING I'VE SHOWN YOU UP 3263 02:10:45,017 --> 02:10:47,753 UNTIL NOW IS BASED ON ANALYSIS 3264 02:10:47,753 --> 02:10:50,789 OF RNA SEQ DATA AND NEEDS TO BE 3265 02:10:50,789 --> 02:10:51,957 VALIDATED EXPERIMENTALLY. 3266 02:10:51,957 --> 02:10:54,226 SO I'M GOING TO SHOW YOU TWO 3267 02:10:54,226 --> 02:10:58,164 EXPERIMENTAL VALIDATIONS FOR CD 3268 02:10:58,164 --> 02:10:59,798 4R5 AND FOR THE COMMON GAMMA 3269 02:10:59,798 --> 02:11:01,267 CHAIN ALTHOUGH WE'VE VALIDATED 3270 02:11:01,267 --> 02:11:03,002 ALL OF THESE IN OTHER 3271 02:11:03,002 --> 02:11:03,302 EXPERIMENTS. 3272 02:11:03,302 --> 02:11:05,171 SO LET'S LOOK FIRST AT THE 3273 02:11:05,171 --> 02:11:07,506 ALTERNATIVE SPLICING OF CD45. 3274 02:11:07,506 --> 02:11:13,145 IT'S FAIRLY STRAIGHTFORWARD. 3275 02:11:13,145 --> 02:11:14,680 XR5 IS ALTERNATIVELY SPLICED 3276 02:11:14,680 --> 02:11:16,549 SUCH THAT IN THE ABSENCE OF BRD4 3277 02:11:16,549 --> 02:11:20,486 YOU HAVE GREATER RETENTION OF 3278 02:11:20,486 --> 02:11:21,921 AXON 5 AND WHAT THAT RESULTS IN 3279 02:11:21,921 --> 02:11:24,423 IS A LARGER TRANSCRIPT AND WE 3280 02:11:24,423 --> 02:11:28,861 CAN IDENTIFY IT BY SELECTIVE 3281 02:11:28,861 --> 02:11:32,998 RT PCR AND WOULD GENERATE AN 3282 02:11:32,998 --> 02:11:35,534 AMPLICON OF AROUND 265 BASE 3283 02:11:35,534 --> 02:11:40,306 PAIRS WHEN THE EXON IS EXCLUDED. 3284 02:11:40,306 --> 02:11:41,640 ININCLUDED, ONLY 118 BASE PAIRS 3285 02:11:41,640 --> 02:11:43,042 AND LOOKING AT THOSE TWO 3286 02:11:43,042 --> 02:11:44,376 PRODUCTS WE CAN SEE IF THERE ARE 3287 02:11:44,376 --> 02:11:46,245 CHANGES IN THE RATIO IS AS A 3288 02:11:46,245 --> 02:11:47,780 RESULT OF DELETING BRD4 AND IN 3289 02:11:47,780 --> 02:11:51,450 FACT THAT'S EXACT WHAT WILL WE 3290 02:11:51,450 --> 02:11:51,684 SEE. 3291 02:11:51,684 --> 02:11:53,419 THERE'S ABOUT A ONE TO ONE RATIO 3292 02:11:53,419 --> 02:11:54,887 IN THE WILD-TYPE BUT WHEN YOU 3293 02:11:54,887 --> 02:11:56,622 KNOCK OUT BRD4 YOU HAVE 3294 02:11:56,622 --> 02:11:58,958 INCREASED RETENTION OF EXON5 3295 02:11:58,958 --> 02:12:02,328 WHICH IS GOING TO LEAD YOU TO A 3296 02:12:02,328 --> 02:12:04,463 DIFFERENT PRODUCT THAN YOU GET 3297 02:12:04,463 --> 02:12:07,066 IN THE ABSENCE OF EXON5. 3298 02:12:07,066 --> 02:12:09,101 THEN YOU CAN LOOK AT THE COMMON 3299 02:12:09,101 --> 02:12:10,736 GAMMA CHAIN WHICH IS MUCH MORE 3300 02:12:10,736 --> 02:12:12,938 COMPLICATED BUT POSSIBLY MORE 3301 02:12:12,938 --> 02:12:13,472 INTERESTING EXAMPLE. 3302 02:12:13,472 --> 02:12:15,874 IN THIS CASE, THE COMMON GAMA 3303 02:12:15,874 --> 02:12:17,743 CHAIN UNDERGOES A LOT OF 3304 02:12:17,743 --> 02:12:19,511 DIFFERENT ALTERNATIVE SPLICING, 3305 02:12:19,511 --> 02:12:20,813 BUT THE ONE WE'RE PARTICULARLY 3306 02:12:20,813 --> 02:12:23,215 GOING ON LOOK AT IS EXON5 WHICH 3307 02:12:23,215 --> 02:12:25,618 SEEMS TO BE THE MOST AFFECTED BY 3308 02:12:25,618 --> 02:12:27,686 BRD4 DELETION, AND IN THIS CASE, 3309 02:12:27,686 --> 02:12:29,355 WHEN YOU DELETE BRD4 THE 3310 02:12:29,355 --> 02:12:31,023 PREDICTION IS YOU'LL GET 3311 02:12:31,023 --> 02:12:33,425 INCREASED EXON SKIPPING AND THE 3312 02:12:33,425 --> 02:12:34,860 REASON THAT'S PARTICULARLY 3313 02:12:34,860 --> 02:12:38,364 INTERESTING IS THAT EXON 3314 02:12:38,364 --> 02:12:40,532 SKIPPING INTRODUCES TGA STOP 3315 02:12:40,532 --> 02:12:42,401 CODON INTO THE PROTEIN SUCH THAT 3316 02:12:42,401 --> 02:12:45,437 YOU DON'T MAKE ANY COMMON GAMA 3317 02:12:45,437 --> 02:12:49,208 CHAIN. SO LET'S SEE WHAT 3318 02:12:49,208 --> 02:12:51,143 HAPPENS EXPERIMENTALLY WHEN WE 3319 02:12:51,143 --> 02:12:52,144 LOOK AT THE PARTICULAR 3320 02:12:52,144 --> 02:12:53,445 TRANSCRIPTS WE'RE LOOKING AT ARE 3321 02:12:53,445 --> 02:12:55,881 THOSE THAT HAVE SKIPPED EXON5 3322 02:12:55,881 --> 02:12:59,018 AND YOU CAN SEE THEM HERE, 3323 02:12:59,018 --> 02:13:01,320 LOOKING AT THE DIFFERENT STAGES, 3324 02:13:01,320 --> 02:13:03,255 KNOCK-OUT VERSUS WILD-TYPE, YOU 3325 02:13:03,255 --> 02:13:04,823 CAN SEE DISCRETE DIFFERENCES. 3326 02:13:04,823 --> 02:13:07,126 IT'S EASIER TO SEE IN THE 3327 02:13:07,126 --> 02:13:08,427 QUANTITATION WHERE WE'VE 3328 02:13:08,427 --> 02:13:10,429 CALCULATED 9 PERCENT OF EXON 3329 02:13:10,429 --> 02:13:12,931 SKIPPING ACROSS THE DIFFERENT 3330 02:13:12,931 --> 02:13:13,799 THYMOCYTE POPULATIONS, AND WHAT 3331 02:13:13,799 --> 02:13:17,436 YOU CAN SEE IS PRETTY EXTENSIVE 3332 02:13:17,436 --> 02:13:18,971 EXON SKIPPING IN THE 3333 02:13:18,971 --> 02:13:22,274 DOUBLE-NEGATIVES AND ISP'S IN 3334 02:13:22,274 --> 02:13:25,544 THE H WILD-TYPE THAT DROPS PRE 3335 02:13:25,544 --> 02:13:26,845 PRECIPITOUSLY WHEN CELLS BECOME 3336 02:13:26,845 --> 02:13:28,314 DP AND FURTHER DIFFERENTIATE. 3337 02:13:28,314 --> 02:13:30,449 IN CONTRAST, ALTHOUGH WE DO SEE 3338 02:13:30,449 --> 02:13:31,850 EXON SKIPPING IN THE KNOCK-OUT, 3339 02:13:31,850 --> 02:13:35,287 IT STAYS PRETTY CONSTANT, IT 3340 02:13:35,287 --> 02:13:36,155 DIMINISHES A LITTLE BUT IN FACT 3341 02:13:36,155 --> 02:13:38,657 YOU CAN SEE IT'S GREATER IN THE 3342 02:13:38,657 --> 02:13:40,659 DP'S AND IN THE SINGLE POSITIVES 3343 02:13:40,659 --> 02:13:44,263 IN THE KNOCK-OUT THAN IN THE 3344 02:13:44,263 --> 02:13:44,530 WILD-TYPE. 3345 02:13:44,530 --> 02:13:46,065 SO PUTTING ALL THIS TOGETHER, WE 3346 02:13:46,065 --> 02:13:48,734 CAN CONCLUDE THAT 3347 02:13:48,734 --> 02:13:49,601 ALTHOUGHTIVE -- THAT ALTERNATIVE 3348 02:13:49,601 --> 02:13:52,104 SPLICING IS REGULATED BY BRD4 IN 3349 02:13:52,104 --> 02:13:53,439 ALL THYMOCYTE SUBSETS LEADING 3350 02:13:53,439 --> 02:13:55,307 EITHER TO A LOSS OF PROTEIN OR A 3351 02:13:55,307 --> 02:13:57,142 DIVERSE SET OF PROTEIN PRODUCTS 3352 02:13:57,142 --> 02:13:59,678 IN THE DIFFERENT CONDITIONS. 3353 02:13:59,678 --> 02:14:00,012 OKAY. 3354 02:14:00,012 --> 02:14:02,514 SO IN THE LAST FEW MINIMUM 3355 02:14:02,514 --> 02:14:04,049 ITS -- SO IN THE LAST FEW 3356 02:14:04,049 --> 02:14:04,917 MINUTES I WOULD LIKE TO TURN TO 3357 02:14:04,917 --> 02:14:06,218 THE QUESTION OF WHAT IS THE ROLE 3358 02:14:06,218 --> 02:14:08,320 OF THE THYMIC EPITHELIUM IN THE 3359 02:14:08,320 --> 02:14:16,528 DEVELOPMENT OF THE THYMUS? 3360 02:14:16,528 --> 02:14:19,264 SO THYMIC DEVELOPMENT VALLEY A 3361 02:14:19,264 --> 02:14:20,666 REFLECTION OF THE CROSS-TALK 3362 02:14:20,666 --> 02:14:22,868 BETWEEN THYMOCYTES AND THYMIC 3363 02:14:22,868 --> 02:14:24,069 EPITHELIUM, SO WE WANTED TO KNOW 3364 02:14:24,069 --> 02:14:26,905 WHAT ROLE DOES BRD4 PLAY WHEN WE 3365 02:14:26,905 --> 02:14:28,807 DELETE OR IN THE THYMIC 3366 02:14:28,807 --> 02:14:30,442 EPITHELIUM WHAT HAPPENS WHEN YOU 3367 02:14:30,442 --> 02:14:33,278 DELETE IT. 3368 02:14:33,278 --> 02:14:35,681 SO AS WAS MENTIONED THIS 3369 02:14:35,681 --> 02:14:38,851 MORNING, THE TWO MAJOR THYMIC 3370 02:14:38,851 --> 02:14:42,221 EPITHELIAL SUB SUBSETS ARE THE 3371 02:14:42,221 --> 02:14:44,089 CT HE C'S WHICH REGULATE 3372 02:14:44,089 --> 02:14:46,058 POSITIVE SELECTION AND MTEC'S 3373 02:14:46,058 --> 02:14:47,059 WHICH REGULATE NEGATIVE 3374 02:14:47,059 --> 02:14:48,627 SELECTION, THE TWO CELL 3375 02:14:48,627 --> 02:14:50,028 POPULATIONS AS WE'VE HEARD HAVE 3376 02:14:50,028 --> 02:14:51,530 A VERY DIFFERENT EXPRESSION 3377 02:14:51,530 --> 02:14:52,831 PROFILE BUT WHAT THEY DO HAVE IN 3378 02:14:52,831 --> 02:14:55,467 COMMON IS THE EXPRESSION OF FOX 3379 02:14:55,467 --> 02:14:58,370 N1 THE TRANSCRIPTION FACTOR, SO 3380 02:14:58,370 --> 02:15:04,009 WE CAN DELETE BRD4 FROM THYMIC 3381 02:15:04,009 --> 02:15:09,148 EPITHELIAL CELLS WITH FOX N1 AND 3382 02:15:09,148 --> 02:15:09,448 CRE. 3383 02:15:09,448 --> 02:15:11,083 WE DO THAT AND SEE A DRAMATIC 3384 02:15:11,083 --> 02:15:13,619 LOSS IN CELLULARITY IN THE 3385 02:15:13,619 --> 02:15:15,220 TEC'S, YOU CAN SEE HERE IS THE 3386 02:15:15,220 --> 02:15:18,557 WILD-TYPE TE C'S, KNOCK-OUT 3387 02:15:18,557 --> 02:15:19,391 TEC'S, WHAT'S PARTICULARLY 3388 02:15:19,391 --> 02:15:21,493 INTERESTING IS WE'RE SELECT TISM 3389 02:15:21,493 --> 02:15:24,029 LOSING MTEC'S, THE TOTAL NUMBER 3390 02:15:24,029 --> 02:15:25,597 OF CT HE C'S DOESN'T CHANGE AND 3391 02:15:25,597 --> 02:15:26,632 YOU CAN SEE CHANGES IN THE 3392 02:15:26,632 --> 02:15:28,801 FREQUENCY THAT REFLECT THAT. 3393 02:15:28,801 --> 02:15:30,369 WE CAN NOW DRILL DOWN AND ASK 3394 02:15:30,369 --> 02:15:32,538 WHAT THOOPPED THE SINGLE CELL 3395 02:15:32,538 --> 02:15:34,406 LEVEL AND WHAT WE FIND WHEN WE 3396 02:15:34,406 --> 02:15:36,041 DO SINGLE CELL SEQUENCE SG THAT 3397 02:15:36,041 --> 02:15:39,111 WE HAVE FOUR CLUSTERS OF TEC'S, 3398 02:15:39,111 --> 02:15:42,481 THREE MTEC'S AND ONE CTEC, AND 3399 02:15:42,481 --> 02:15:45,451 THE THYMOCYTE POPULATION CD4 CD8 3400 02:15:45,451 --> 02:15:48,954 ARE COMPLETELY DISTINCT. 3401 02:15:48,954 --> 02:15:50,989 WHEN WE LOOK FURTHER AT THE CT 3402 02:15:50,989 --> 02:15:52,758 HE C'S WE CAN ASK WHAT HAPPENS 3403 02:15:52,758 --> 02:15:55,093 WHEN WE KNOCK OUT BRD4 AND WHAT 3404 02:15:55,093 --> 02:15:56,462 WE SEE IS QUITE DRAMATIC, 3405 02:15:56,462 --> 02:15:58,664 THERE'S A HUGE LOSS IN THOSE 3406 02:15:58,664 --> 02:16:00,532 THREE MTEC POPULATIONS AND 3407 02:16:00,532 --> 02:16:03,068 VIRTUALLY NO LOSS IN THE CTECS, 3408 02:16:03,068 --> 02:16:04,603 THAT'S QUANTITATED HERE. 3409 02:16:04,603 --> 02:16:07,673 SO IT REALLY LOOK AS THOUGH BRD4 3410 02:16:07,673 --> 02:16:10,275 SELECTIVELY AFFECTING MTEC 3411 02:16:10,275 --> 02:16:10,576 POPULATION. 3412 02:16:10,576 --> 02:16:14,446 WE CAN VISUALIZE THAT, INCLUDING 3413 02:16:14,446 --> 02:16:15,681 HISTOCHEMISTRY IF WE LOOK AT 3414 02:16:15,681 --> 02:16:16,815 AIRE EXPRESSION IN THE WILD-TYPE 3415 02:16:16,815 --> 02:16:19,184 WE SEE VERY NICE AIRE 3416 02:16:19,184 --> 02:16:21,820 EXPRESSION, MC HE T'S RUA1 IS 3417 02:16:21,820 --> 02:16:23,555 ALSO SHOWING VERY NICE STAINING 3418 02:16:23,555 --> 02:16:25,224 IN THE WILD-TYPE, YOU LOSE IT IN 3419 02:16:25,224 --> 02:16:29,828 THE KNOCK-OUT, IN CONTRAST BETA 3420 02:16:29,828 --> 02:16:31,997 5T MR FOR CTEC'S DOESN'T 3421 02:16:31,997 --> 02:16:33,632 INCREASE, IF ANYTHING WE MAY SEE 3422 02:16:33,632 --> 02:16:34,733 A RELATIVE INCREASE. 3423 02:16:34,733 --> 02:16:35,701 THAT'S ALSO REFLECT UNDERSTAND 3424 02:16:35,701 --> 02:16:37,236 IN THE SINGLE CELL EXPRESSION 3425 02:16:37,236 --> 02:16:38,370 LEVELS OF THE DIFFERENT MARKERS, 3426 02:16:38,370 --> 02:16:41,306 SO IF WE LOOK AT MTEC MARKERS IN 3427 02:16:41,306 --> 02:16:43,375 CLUSTER ZERO AND CLUSTER 4, WE 3428 02:16:43,375 --> 02:16:47,446 SEE AN ACUTE LOSS OF AIRE AND 3429 02:16:47,446 --> 02:16:50,082 RANK, IN THE ABSENCE OF BRD4 3430 02:16:50,082 --> 02:16:52,251 INDICATING THAT BRD4 IS 3431 02:16:52,251 --> 02:16:53,785 REGULATING EXPRESSION OF AIRE 3432 02:16:53,785 --> 02:16:55,988 AND RANK BUT WE DON'T SEE THOSE 3433 02:16:55,988 --> 02:16:57,623 CHANGES IN THE CTEC. 3434 02:16:57,623 --> 02:16:59,758 NOW IN MY LAST SLIDE, STANLEY, 3435 02:16:59,758 --> 02:17:01,293 WE'RE GOING TO ASK WHAT HAPPENS 3436 02:17:01,293 --> 02:17:03,228 AT THE LEVEL OF THE THYMOCYTES 3437 02:17:03,228 --> 02:17:04,863 DISRKS THE LOSS OF BIRD -- DOES 3438 02:17:04,863 --> 02:17:08,267 THE LOSS OF BRD4 IN THE T HE C'S 3439 02:17:08,267 --> 02:17:09,635 AFFECT THYMOCYTE 3440 02:17:09,635 --> 02:17:10,302 DIFFERENTIATION, AND THE ANSWER 3441 02:17:10,302 --> 02:17:10,802 IS YES. 3442 02:17:10,802 --> 02:17:13,739 WE DON'T SEE ANY CHANGE IN THE 3443 02:17:13,739 --> 02:17:15,073 DOUBLE-NEGATIVES OR THE ISP'S 3444 02:17:15,073 --> 02:17:17,809 BUT WE SEE A SIGNIFICANT 3445 02:17:17,809 --> 02:17:18,744 ACCUMULATION OF DOUBLE POSITIVES 3446 02:17:18,744 --> 02:17:21,680 AND THEN A SUBSEQUENT LOSS IN 3447 02:17:21,680 --> 02:17:25,384 THE SINGLE POSITIVE CD4 CD8 3448 02:17:25,384 --> 02:17:28,020 SUBSETS AS WELL AS IN T REGS 3449 02:17:28,020 --> 02:17:29,121 ALLOWING US TO INITIALLY 3450 02:17:29,121 --> 02:17:32,624 CONCLUDE THAT DELETION OF BRD4 3451 02:17:32,624 --> 02:17:35,127 IN C TEST CS BLOCKS THYMIC 3452 02:17:35,127 --> 02:17:37,729 SELECTION OF DOUBLE POSITIVE 3453 02:17:37,729 --> 02:17:38,630 THYMOCYTES, ALTHOUGH REMEMBER WE 3454 02:17:38,630 --> 02:17:40,599 SEE NO DIFFERENCE IN THE NUMBER 3455 02:17:40,599 --> 02:17:42,834 OF CT HE C'S, THERE APPEARS TO 3456 02:17:42,834 --> 02:17:44,636 BE A FUNCTIONAL DEFECT. 3457 02:17:44,636 --> 02:17:46,171 SO THE CONCLUSION FROM THIS 3458 02:17:46,171 --> 02:17:48,807 SECTION IS THAT MTEC DEVELOPMENT 3459 02:17:48,807 --> 02:17:51,076 DEPENDS ON BRD4 EXPRESSION BUT 3460 02:17:51,076 --> 02:17:52,744 POSITIVE SELECTION OF DOUBLE 3461 02:17:52,744 --> 02:17:56,048 POSITIVE THYMOCYTES DEPENDS ON 3462 02:17:56,048 --> 02:17:59,084 BRD4 EXPRESSION IN TEST C'S. 3463 02:17:59,084 --> 02:18:00,586 AND WE'VE HEARD ALL OF THIS, BUT 3464 02:18:00,586 --> 02:18:02,154 I DO TO WANT TAKE ONE MINUTE TO 3465 02:18:02,154 --> 02:18:03,789 GIVE A SPECIAL THANKS ON ALL THE 3466 02:18:03,789 --> 02:18:05,891 PEOPLE WHO WORKED ON THIS 3467 02:18:05,891 --> 02:18:16,401 PROJECT IN MY LAB, JAY MU, ANG I 3468 02:18:16,401 --> 02:18:17,402 GA -- 3469 02:18:17,402 --> 02:18:23,642 IT ANNE, SHEETAL, DEVAIAH, AND 3470 02:18:23,642 --> 02:18:25,210 OUR COLLABORATORS. 3471 02:18:25,210 --> 02:18:27,546 THANK YOU. 3472 02:18:27,546 --> 02:18:37,723 [APPLAUSE] 3473 02:18:38,357 --> 02:18:38,857 >> HI. 3474 02:18:38,857 --> 02:18:39,925 I HAD A QUICK QUESTION. 3475 02:18:39,925 --> 02:18:43,061 WHEN YOU LOOK AT JUST H & E 3476 02:18:43,061 --> 02:18:44,396 HISTOLOGY DO YOU STILL HAVE 3477 02:18:44,396 --> 02:18:47,766 MEDULLARY ISLANDS OR IS THERE 3478 02:18:47,766 --> 02:18:48,667 JUST KNOCK-OUT -- 3479 02:18:48,667 --> 02:18:49,134 >> NO. 3480 02:18:49,134 --> 02:18:50,869 >> THERE'S NO MEDULLARY ISLANDS 3481 02:18:50,869 --> 02:18:53,472 AT ALL? 3482 02:18:53,472 --> 02:18:54,306 >> RESIDUAL AT BEST. 3483 02:18:54,306 --> 02:18:55,874 >> THE RESIDUAL? 3484 02:18:55,874 --> 02:18:57,175 FOR YOUR SINGLE CELL, HAVE YOU 3485 02:18:57,175 --> 02:18:59,077 LOOKED AT THOSE MTEC CLUSTERS 3486 02:18:59,077 --> 02:19:00,579 AND IT SEEMED LIKE IT WAS 3487 02:19:00,579 --> 02:19:03,749 EQUALLY REDUCED FOR ALL OF THEM? 3488 02:19:03,749 --> 02:19:06,051 WERE THERE SPECIFIC CLUSTERS OF 3489 02:19:06,051 --> 02:19:06,485 MTEC? 3490 02:19:06,485 --> 02:19:06,685 OR -- 3491 02:19:06,685 --> 02:19:11,289 >> SO THESE ARE REALLY NEW DATA, 3492 02:19:11,289 --> 02:19:12,924 SWREANT LOOKED AT THEM IN GREAT 3493 02:19:12,924 --> 02:19:14,359 DETAIL, BUT OVERALL, THE 3494 02:19:14,359 --> 02:19:16,094 REDUCTION SEEMS TO BE ABOUT 3495 02:19:16,094 --> 02:19:18,630 EQUIVALENT IN ALL THE THREE 3496 02:19:18,630 --> 02:19:18,897 SUBTYPES. 3497 02:19:18,897 --> 02:19:20,065 >> VERY INTERESTING. 3498 02:19:20,065 --> 02:19:20,332 THANK YOU. 3499 02:19:20,332 --> 02:19:22,167 >> OR SUB CLUSTERS. 3500 02:19:22,167 --> 02:19:23,869 >> CAN YOU HEAR ME? 3501 02:19:23,869 --> 02:19:24,202 OKAY. 3502 02:19:24,202 --> 02:19:29,708 SO I HAVE TWO QUESTIONS TO THE 3503 02:19:29,708 --> 02:19:30,642 THYMOCYTES, FIRST DO YOU SEE IF 3504 02:19:30,642 --> 02:19:34,613 THERE IS ANY CHANGE IN LIKE 3505 02:19:34,613 --> 02:19:36,615 SPLICING OF MACON? 3506 02:19:36,615 --> 02:19:38,717 >> NO. 3507 02:19:38,717 --> 02:19:40,552 EVERYTHING -- THE MYC GENE IS 3508 02:19:40,552 --> 02:19:41,086 NOT TRANSCRIBED. 3509 02:19:41,086 --> 02:19:41,420 >> OKAY. 3510 02:19:41,420 --> 02:19:42,320 >> SO NOTHING IS THERE. 3511 02:19:42,320 --> 02:19:44,756 >> NOTHING, OKAY. 3512 02:19:44,756 --> 02:19:45,157 >> ISP'S. 3513 02:19:45,157 --> 02:19:46,358 THE OTHERS ARE EXPRESSED 3514 02:19:46,358 --> 02:19:46,625 NORMALLY. 3515 02:19:46,625 --> 02:19:47,559 >> OKAY. 3516 02:19:47,559 --> 02:19:50,062 AND THEN DID YOU CHECK FOR HAD 3517 02:19:50,062 --> 02:19:51,563 THE I7 SUB EXPRESSION? 3518 02:19:51,563 --> 02:19:53,832 I KNOW THE COMMON GAMA CHAIN 3519 02:19:53,832 --> 02:19:55,067 WOULD AFFECT IT. 3520 02:19:55,067 --> 02:19:56,001 DO YOU THINK THAT THIS IS THE 3521 02:19:56,001 --> 02:19:57,636 REASON WHY YOU DON'T MOVE ON 3522 02:19:57,636 --> 02:20:01,206 INTO THE DIFFERENTIATION? 3523 02:20:01,206 --> 02:20:03,675 >> SO WE HAVEN'T LOOKED BEYOND 3524 02:20:03,675 --> 02:20:05,610 THE COMMON GAMA CHAIN IN TERMS 3525 02:20:05,610 --> 02:20:06,445 OF SPLICING. 3526 02:20:06,445 --> 02:20:07,846 SO YEAH, I DON'T KNOW. 3527 02:20:07,846 --> 02:20:08,814 GOOD QUESTION THOUGH. 3528 02:20:08,814 --> 02:20:09,915 THANK YOU. 3529 02:20:09,915 --> 02:20:12,751 >> HEY, DINAH, THAT WAS A GREAT 3530 02:20:12,751 --> 02:20:12,951 STORY. 3531 02:20:12,951 --> 02:20:14,920 MY QUESTION IS ACTUALLY A 3532 02:20:14,920 --> 02:20:19,424 CONTINUATION OF ROXANE'S. 3533 02:20:19,424 --> 02:20:20,959 SO YOU CAN ALWAYS GO ONE STEP 3534 02:20:20,959 --> 02:20:21,293 FURTHER. 3535 02:20:21,293 --> 02:20:23,628 YOU SHOWED THAT YOU HAD MUCH 3536 02:20:23,628 --> 02:20:25,130 MORE ALTERNATIVE SPLICING IN 3537 02:20:25,130 --> 02:20:29,401 ISP'S AS COMPARED TO OTHER 3538 02:20:29,401 --> 02:20:30,268 SUBSETS. 3539 02:20:30,268 --> 02:20:31,403 GIVEN SOME OF THE OTHER PEOPLE'S 3540 02:20:31,403 --> 02:20:35,307 WORK HERE, THE ISP'S ARE THE 3541 02:20:35,307 --> 02:20:37,175 MOST METABOLICALLY ACTIVE THYME 3542 02:20:37,175 --> 02:20:37,876 SIGHTED SUBSETS. 3543 02:20:37,876 --> 02:20:39,377 I WAS WONDERING IS IT BECAUSE 3544 02:20:39,377 --> 02:20:42,180 THAT THAT THE ISP'S HAVE MORE 3545 02:20:42,180 --> 02:20:43,215 ALTERNATIVE SPLICING? 3546 02:20:43,215 --> 02:20:44,916 WHAT IS YOUR HYPOTHESIS FOR WHY 3547 02:20:44,916 --> 02:20:46,485 YOU SEE MORE ALTERNATIVE SPLICE 3548 02:20:46,485 --> 02:20:47,552 NG ISP'S? 3549 02:20:47,552 --> 02:20:49,087 ONE THING I WAS TALKING ABOUT 3550 02:20:49,087 --> 02:20:50,288 WAS METABOLISM AND THE OTHER IS 3551 02:20:50,288 --> 02:20:51,490 IS THE DIVERSITY OF TRANSCRIPTS 3552 02:20:51,490 --> 02:20:53,458 IN AN ISP HIGHER THAN THE 3553 02:20:53,458 --> 02:20:55,427 DIVERSITY OF TRIPTSZ IN OTHER 3554 02:20:55,427 --> 02:20:55,660 SUBSETS? 3555 02:20:55,660 --> 02:20:57,362 -- TRANSCRIPTS IN OTHER SUBSETS? 3556 02:20:57,362 --> 02:20:58,964 >> LET ME TRY TO ANSWER ALL 3557 02:20:58,964 --> 02:21:02,033 PARTS OF THE QUESTION. 3558 02:21:02,033 --> 02:21:04,836 WE LOOKED AT GLYCOLYSIS AND IN 3559 02:21:04,836 --> 02:21:08,039 THE ABSENCE OF BRD4 YOU HAVE 3560 02:21:08,039 --> 02:21:09,608 EXTRAORDINARILY SUPPRESSED 3561 02:21:09,608 --> 02:21:10,208 GLYCOLYTIC FUNCTIONS. 3562 02:21:10,208 --> 02:21:13,178 IN TERMS OF ALTERNATIVE 3563 02:21:13,178 --> 02:21:15,247 SPLICING, THE TRIVIAL ANSWER IS 3564 02:21:15,247 --> 02:21:17,349 THAT THE RNA PROFILES OF THE 3565 02:21:17,349 --> 02:21:19,618 WILD-TYPE AND BRD4 ARE SO 3566 02:21:19,618 --> 02:21:22,521 DIFFERENT THAT ALMOST YOU HAVE 3567 02:21:22,521 --> 02:21:23,588 TO HAVE DIFFERENCES IN 3568 02:21:23,588 --> 02:21:24,990 ALTERNATIVE SPLICING BECAUSE YOU 3569 02:21:24,990 --> 02:21:25,891 HAVE DIFFERENT TRANSCRIPTS THAT 3570 02:21:25,891 --> 02:21:28,760 YOU'RE STARTING WITH. 3571 02:21:28,760 --> 02:21:31,496 THE TRANSCRIPTS OF THE ISP'S 3572 02:21:31,496 --> 02:21:32,564 COMPARED TO THE SINGLE POSITIVES 3573 02:21:32,564 --> 02:21:36,935 OR THE DOUBLE-NEGATIVES, THE 3574 02:21:36,935 --> 02:21:40,038 ALTERNATIVE SPLICING PATTERNS DO 3575 02:21:40,038 --> 02:21:44,042 NOT SIGNIFICANTLY OVERLAP. 3576 02:21:44,042 --> 02:21:47,345 THERE ARE IT HUGE GENES THAT 3577 02:21:47,345 --> 02:21:49,414 OVERLAP BUT LI AND LARGE THEY 3578 02:21:49,414 --> 02:21:49,614 DON'T. 3579 02:21:49,614 --> 02:21:53,151 >> GLYCOLYSIS FOR EXAMPLE, THE 3580 02:21:53,151 --> 02:21:55,987 SPLICING FOR RA VERSUS RO IS 3581 02:21:55,987 --> 02:21:56,888 CLEARLY ASSOCIATED WITH THE 3582 02:21:56,888 --> 02:21:57,989 ACTIVATION OF THOSE CELLS. 3583 02:21:57,989 --> 02:21:59,491 >> RIGHT Z SO IT MIGHT 3584 02:21:59,491 --> 02:21:59,791 ACTUALLY -- 3585 02:21:59,791 --> 02:22:00,926 >> YEAH. 3586 02:22:00,926 --> 02:22:02,561 SO WE'RE SEEING THAT ALTERNATIVE 3587 02:22:02,561 --> 02:22:05,297 SPLICING IN ALL OF THE DIFFERENT 3588 02:22:05,297 --> 02:22:07,833 SUBSETS FOR TR CD45 -- SORRY, 3589 02:22:07,833 --> 02:22:10,669 WE'RE ONLY SEEING IT IN CD4'S 3590 02:22:10,669 --> 02:22:11,870 FOR CD45. 3591 02:22:11,870 --> 02:22:18,009 YEAH. 3592 02:22:18,009 --> 02:22:20,178 >> DO YOU THINK HOW THE BRD4 3593 02:22:20,178 --> 02:22:22,914 LIKE REGULATES THE SPLICING 3594 02:22:22,914 --> 02:22:26,218 INTERACTS WITH ANY SPLICEOSOME 3595 02:22:26,218 --> 02:22:26,852 COMPONENTS OR -- 3596 02:22:26,852 --> 02:22:28,286 >> DIDN'T HAVE TIME TO SHOW YOU 3597 02:22:28,286 --> 02:22:31,890 THE DATA, BUT BRD4 ACTUALLY 3598 02:22:31,890 --> 02:22:32,891 PHYSICALLY INTERACTS WITH 3599 02:22:32,891 --> 02:22:35,227 SPLICING FACTORS AND IT APPEARS 3600 02:22:35,227 --> 02:22:39,231 TO BE A REGULATORY FACTOR OF 3601 02:22:39,231 --> 02:22:39,531 SPLICING. 3602 02:22:39,531 --> 02:22:42,100 >> DO YOU KNOW WHICH STEPS, LIKE 3603 02:22:42,100 --> 02:22:44,803 THE COMPLEX B OR C OR ANY STEPS 3604 02:22:44,803 --> 02:22:46,905 OF THE SPLICEOSOME OR IT'S JUST 3605 02:22:46,905 --> 02:22:47,138 LIKE -- 3606 02:22:47,138 --> 02:22:52,477 >> SO ACTUALLY THE U1 RFP IT 3607 02:22:52,477 --> 02:22:54,579 INTERACTS WITH FUS, SO IT'S 3608 02:22:54,579 --> 02:22:57,515 INTERACTING WITH THE REGULATORY 3609 02:22:57,515 --> 02:23:00,051 FACTORS AS WELL AS WITH THE CORE 3610 02:23:00,051 --> 02:23:00,952 SPLICEOSOMAL SUBUNITS. 3611 02:23:00,952 --> 02:23:02,454 >> ANOTHER QUICK QUESTION IS 3612 02:23:02,454 --> 02:23:08,994 THAT YOU MENTIONED THE -- WHAT 3613 02:23:08,994 --> 02:23:12,163 ABOUT OTHER TYPES OF SPLICING 3614 02:23:12,163 --> 02:23:17,002 EVENTS -- HAVE YOU EVER CHECKED 3615 02:23:17,002 --> 02:23:20,405 THOSE ZOOR WE'VE ONLY LOOKED AT 3616 02:23:20,405 --> 02:23:21,172 THE ALTERNATIVE. 3617 02:23:21,172 --> 02:23:22,707 >> WE'VE ONLY LOOKED AT THE 3618 02:23:22,707 --> 02:23:23,341 ALTERNATIVE -- IF I UNDERSTOOD 3619 02:23:23,341 --> 02:23:24,776 YOUR QUESTION CORRECTLY, SWREANT 3620 02:23:24,776 --> 02:23:26,645 LOOKED AT OTHER KINDS OF 3621 02:23:26,645 --> 02:23:26,912 SPLICING. 3622 02:23:26,912 --> 02:23:29,347 >> THANK YOU. 3623 02:23:29,347 --> 02:23:30,749 >> ONE MORE QUICK QUESTION. 3624 02:23:30,749 --> 02:23:32,183 >> HEY, CARROLL. 3625 02:23:32,183 --> 02:23:33,852 >> I WAS JUST WONDERING, HOW 3626 02:23:33,852 --> 02:23:36,955 MUCH OF THESE ACTIVITIES FOR 3627 02:23:36,955 --> 02:23:40,258 YOUR LOSS OF BRD4 WOULD BE 3628 02:23:40,258 --> 02:23:42,694 RESCUED BY MYC AND THE OVERLAP? 3629 02:23:42,694 --> 02:23:47,365 >> YEAH, YEAH. 3630 02:23:47,365 --> 02:23:48,767 WE'VE TALKED ABOUT DOING THOSE 3631 02:23:48,767 --> 02:23:50,435 EXPERIMENTS BUT WE HAVEN'T DONE 3632 02:23:50,435 --> 02:23:50,635 THEM. 3633 02:23:50,635 --> 02:23:51,836 I THINK SOME OF THAT MIGHT BE 3634 02:23:51,836 --> 02:23:54,906 RESCUED BUT NOT ALL OF IT 3635 02:23:54,906 --> 02:23:56,541 BECAUSE OTHER PATHWAY RS AFFECT 3636 02:23:56,541 --> 02:23:58,610 AS WELL. 3637 02:23:58,610 --> 02:24:00,812 IN TERM OF THE SPLICING REMEMBER 3638 02:24:00,812 --> 02:24:02,547 MYC ISN'T AFFECTED IN ANY OF THE 3639 02:24:02,547 --> 02:24:03,181 OTHER SUBSETS. 3640 02:24:03,181 --> 02:24:05,850 >> I KNOW MYC CAN AFFECT 3641 02:24:05,850 --> 02:24:08,787 SPLICING TOO, SO LOSS OF MYC CA- 3642 02:24:08,787 --> 02:24:10,989 >> IN THE ISP'S, YEAH, BUT THOSE 3643 02:24:10,989 --> 02:24:12,357 ARE THE ONLY CELL TYPES WHERE 3644 02:24:12,357 --> 02:24:14,693 YOU SEE AN EFFECT. 3645 02:24:14,693 --> 02:24:15,327 >> YEAH. 3646 02:24:15,327 --> 02:24:25,570 >> THANK YOU. 3647 02:24:25,804 --> 02:24:30,275 [APPLAUSE] 3648 02:24:30,275 --> 02:24:34,846 >> OUR NEXT SPEAKER IS ROXANE 3649 02:24:34,846 --> 02:24:36,181 TUSSIWAND AND SHE WILL TALK 3650 02:24:36,181 --> 02:24:41,953 ABOUT NOTCH1 IN T CELL 3651 02:24:41,953 --> 02:24:50,862 DEVELOPMENT. 3652 02:24:50,862 --> 02:24:53,865 >> GOOD MORNING, EVERYONE. 3653 02:24:53,865 --> 02:24:58,636 SO THIS IS THE WORK OF AN 3654 02:24:58,636 --> 02:25:00,372 EXTREMELY TALENTED Ph.D. 3655 02:25:00,372 --> 02:25:02,107 STUDENT IN THE LAB DURING HIS 3656 02:25:02,107 --> 02:25:04,976 MASTER AND Ph.D. 3657 02:25:04,976 --> 02:25:07,812 SO WE'VE HEARD A LOT ABOUT T 3658 02:25:07,812 --> 02:25:09,714 CELL DEVELOPMENT, SO I WASN'T 3659 02:25:09,714 --> 02:25:12,984 HERE EARLIER, BUT SO NOTCH IS 3660 02:25:12,984 --> 02:25:19,124 DEFINITELY LIKE A KEY FACTOR IN 3661 02:25:19,124 --> 02:25:23,828 T CELL DEVELOPMENT, SO -- WHICH 3662 02:25:23,828 --> 02:25:27,565 WOULD INTERACTED WITH NOTCH ONE 3663 02:25:27,565 --> 02:25:32,904 SUPPRESSOR EXPRESSED BY -- THE 3664 02:25:32,904 --> 02:25:35,507 NEEDS OF THE ANY CD OR NOTCH 3665 02:25:35,507 --> 02:25:36,841 INTRACELLULAR DOMAIN AND WHAT IS 3666 02:25:36,841 --> 02:25:41,346 THOUGHT TO HAPPEN IS THAT THE 3667 02:25:41,346 --> 02:25:47,485 NOTCH WILL BIND TO RPJ 3668 02:25:47,485 --> 02:25:49,354 DISSOMEPLACE THE ARE -- DISPLACE 3669 02:25:49,354 --> 02:25:51,990 THE COMPLEX AND LEAD TO 3670 02:25:51,990 --> 02:25:53,858 INDUCTION OF TRANSCRIPTION. 3671 02:25:53,858 --> 02:25:54,959 SO WHAT WE'RE TRYING TO 3672 02:25:54,959 --> 02:25:56,828 UNDERSTAND IS REALLY LIKE HOW 3673 02:25:56,828 --> 02:26:01,099 THIS MECHANISM WORKS AND WHAT WE 3674 02:26:01,099 --> 02:26:03,068 CAN UNDERSTAND FROM T CELL 3675 02:26:03,068 --> 02:26:03,435 DEVELOPMENT. 3676 02:26:03,435 --> 02:26:07,238 SO WHAT WE KNOW IS THAT WHAT WAS 3677 02:26:07,238 --> 02:26:10,842 RECENTLY SHOWN IS THAT THE NOTCH 3678 02:26:10,842 --> 02:26:17,082 SIGNALLING ALREADY STARTED, IS 3679 02:26:17,082 --> 02:26:18,416 MULTIPLE PROGENITORS WHICH HAVE 3680 02:26:18,416 --> 02:26:20,819 THE CAPACITY TO DEVELOP INTO ALL 3681 02:26:20,819 --> 02:26:21,786 SUBSETS ARE WHICH RECEIVE THE 3682 02:26:21,786 --> 02:26:23,021 SIGNAL AND IT IS THE SIGNAL 3683 02:26:23,021 --> 02:26:24,122 CLEARLY THAT THIS IS THE SIGNAL 3684 02:26:24,122 --> 02:26:29,127 WHICH LEADS TO THE BONE MARROW 3685 02:26:29,127 --> 02:26:31,729 AND TWHEAN THE PROGENITORS WILL 3686 02:26:31,729 --> 02:26:33,064 SOMEHOW REACH THE THYMUS AS 3687 02:26:33,064 --> 02:26:35,667 EARLY THYMIC PROGENITORS, 3688 02:26:35,667 --> 02:26:37,869 RECEIVING AT THIS POINT AN 3689 02:26:37,869 --> 02:26:40,505 ACCIDENT TREEMLY STRONG NOTCH 3690 02:26:40,505 --> 02:26:43,341 SIGNALLING THROUGH THE 3691 02:26:43,341 --> 02:26:48,379 EXPRESSION OF CTEC'S LIKE 3692 02:26:48,379 --> 02:26:49,380 DELTALIKE 4. 3693 02:26:49,380 --> 02:26:51,015 IN THIS PROCESS THIS DEPENDENCY 3694 02:26:51,015 --> 02:26:53,418 OF NOTCH IS REALLY OCCURRING 3695 02:26:53,418 --> 02:26:55,386 THROUGHOUT ALL THIS EARLY 3696 02:26:55,386 --> 02:26:58,123 DIFFERENTIATION STAGES, UNTIL 3697 02:26:58,123 --> 02:27:01,159 THE T CELL RECEPTOR IS INDUCED, 3698 02:27:01,159 --> 02:27:03,161 WHICH THEN LEADS THEN TO THE 3699 02:27:03,161 --> 02:27:05,663 FURTHER MATURATION OF T CELLS AS 3700 02:27:05,663 --> 02:27:09,501 WE'VE HEARD JUST NOW. 3701 02:27:09,501 --> 02:27:12,036 SO WHEN WE LOOK EX-VIVO WE WERE 3702 02:27:12,036 --> 02:27:13,571 INTERESTED IN DOUBLE-NEGATIVE 3703 02:27:13,571 --> 02:27:15,039 SUBSETS WHICH IS STILL NEGATIVE 3704 02:27:15,039 --> 02:27:16,941 FOR CD4 AND CD8 AND THIS IS HOW 3705 02:27:16,941 --> 02:27:20,612 IT CAN BE SUBDIVIDED INTO DN1 3706 02:27:20,612 --> 02:27:24,182 WHICH IS EXPRESSED, DN2, DN3 AND 3707 02:27:24,182 --> 02:27:27,252 DN4 AND THIS IS BASED ON 44 AND 3708 02:27:27,252 --> 02:27:30,188 25 EXPRESSION. 3709 02:27:30,188 --> 02:27:32,590 SO THIS THINK THAT NOTCH IS 3710 02:27:32,590 --> 02:27:36,528 ESSENTIAL WAS SHOWN MANY YEARS 3711 02:27:36,528 --> 02:27:39,063 AGO BECAUSE DELETION OF NOTCH 3712 02:27:39,063 --> 02:27:41,900 ONE LEADS TO A COMPLETE BLOCK AT 3713 02:27:41,900 --> 02:27:44,536 THE DN1 STAGE WITH ECTOPIC BASAL 3714 02:27:44,536 --> 02:27:48,573 DEVELOPMENT WITHIN THE THYMUS. 3715 02:27:48,573 --> 02:27:51,309 OVERSPREFTION AN ICD LEADS TO T 3716 02:27:51,309 --> 02:27:52,610 CELL DEVELOPMENT IN THE BONE 3717 02:27:52,610 --> 02:27:54,179 MARROW WHERE NORMALLY THERE IS 3718 02:27:54,179 --> 02:27:56,014 NO NOTCH SIGNALLING OR NOT 3719 02:27:56,014 --> 02:27:59,017 ENOUGH NOTCH SIGNALLING TO DRIVE 3720 02:27:59,017 --> 02:28:00,585 T CELL DEVELOPMENT. 3721 02:28:00,585 --> 02:28:03,555 SO IT IS AN INSTRUCTIVE SIGNAL 3722 02:28:03,555 --> 02:28:04,756 WHICH SUPPRESSES THE OTHER FATE 3723 02:28:04,756 --> 02:28:08,126 AND LEADS TO T CELL COMMITMENT 3724 02:28:08,126 --> 02:28:12,597 AND SPECIFICAT I ON. 3725 02:28:12,597 --> 02:28:14,499 WE KNOW WITHIN ALL THOSE STAGES 3726 02:28:14,499 --> 02:28:16,134 OF T CELL DEVELOPMENT THERE IS A 3727 02:28:16,134 --> 02:28:18,536 CERTAIN DEGREE OF MULL 3728 02:28:18,536 --> 02:28:18,870 MULTI-POTENCY. 3729 02:28:18,870 --> 02:28:21,039 WHEN WE PUT THE CELLS UNDER 3730 02:28:21,039 --> 02:28:22,807 APPROPRIATE CONDITION WHICH CAN 3731 02:28:22,807 --> 02:28:25,210 GENERATE MACROPHAGES IN K CELLS 3732 02:28:25,210 --> 02:28:28,479 AND B CELLS BUT IT'S ONLY P AT 3733 02:28:28,479 --> 02:28:31,883 THIS STAGE THAT THE T CELL 3734 02:28:31,883 --> 02:28:34,185 SPECIFICA TIRKS ON HAS OCCURRED 3735 02:28:34,185 --> 02:28:38,923 SO THE PROGRESS INTO THE T CELL 3736 02:28:38,923 --> 02:28:40,225 FATE IS WITH DIENDZ. 3737 02:28:40,225 --> 02:28:41,392 NOW, BECAUSE OF THE DEFECT OR 3738 02:28:41,392 --> 02:28:45,230 THE BLOCK OF THE DN1 STAGE, WE 3739 02:28:45,230 --> 02:28:48,299 WERE WONDERING WHAT HAPPENS AND 3740 02:28:48,299 --> 02:28:50,835 HOW IS NOTCH IMPORTANT 3741 02:28:50,835 --> 02:28:54,872 THROUGHOUT ALL THOSE STAGES? 3742 02:28:54,872 --> 02:28:56,407 AND THIS IS -- TO ADDRESS THIS 3743 02:28:56,407 --> 02:28:58,176 QUESTION, WE WORKED A T CELL 3744 02:28:58,176 --> 02:29:03,114 CULTURE. SO WE USED NOTCH 3745 02:29:03,114 --> 02:29:04,649 LIGAND IS PROGENITORS IN THE 3746 02:29:04,649 --> 02:29:06,951 BRNS OF I7 AND STEM CELL FACTOR 3747 02:29:06,951 --> 02:29:11,389 AND WE CAN GENERATED A LARGE 3748 02:29:11,389 --> 02:29:14,058 NUMBER OF THE N2 CELLS, EXPRESS 3749 02:29:14,058 --> 02:29:15,793 24 AND 25, IN THE SUBSET BLOCKED 3750 02:29:15,793 --> 02:29:20,732 THIS STAGE, AND SO WHEN WE LOOK 3751 02:29:20,732 --> 02:29:24,168 AT EX-VIVO THYMOCYTES, MOST OF 3752 02:29:24,168 --> 02:29:26,204 THE CELLS PROGRESS VERY FAST 3753 02:29:26,204 --> 02:29:30,575 FROM N1 TO N2 AND WE HAVE MOSTLY 3754 02:29:30,575 --> 02:29:33,645 BN3 OR 4 CELLS. 3755 02:29:33,645 --> 02:29:38,116 SO WHEN WE PUT THE CELLS, THESE 3756 02:29:38,116 --> 02:29:41,552 PRO T CELLS IN VIVO, WE HAVE A 3757 02:29:41,552 --> 02:29:43,154 SINGLE WAVE OF T CELL 3758 02:29:43,154 --> 02:29:45,590 DEVELOPMENT AND HERE IT IS 3759 02:29:45,590 --> 02:29:48,259 SHOWN, CD4 AND CD8, THEY END UP 3760 02:29:48,259 --> 02:29:49,427 BEING PRESENT WITHIN THE THYMUS 3761 02:29:49,427 --> 02:29:51,095 AND THEN THEY CAN LEAVE THE 3762 02:29:51,095 --> 02:29:54,599 THYMUS AND PERFORM T CELL 3763 02:29:54,599 --> 02:29:55,066 FUNCTIONS. 3764 02:29:55,066 --> 02:29:57,535 SO THE CULTURE CELLS SEEMS TO BE 3765 02:29:57,535 --> 02:30:00,138 COMPLETELY FUNCTIONAL. 3766 02:30:00,138 --> 02:30:03,541 WHEN WE LOOK AT RNA EXPRESSION 3767 02:30:03,541 --> 02:30:06,811 AND COMPARE IT WITH SINGLE CELL 3768 02:30:06,811 --> 02:30:09,213 PROFILES OF THYMOCYTES, THEY 3769 02:30:09,213 --> 02:30:12,417 APPEAR TO CORRESPOND TO CYCLING 3770 02:30:12,417 --> 02:30:13,951 DN2 WHICH IS EXACT WHAT WILL WE 3771 02:30:13,951 --> 02:30:15,920 SEE IN CULTURES TOUR, THESE 3772 02:30:15,920 --> 02:30:18,656 CELLS ARE MOSTLY DN2 BY PROFILE 3773 02:30:18,656 --> 02:30:22,293 AND ACTIVELY PROLIFERATING. 3774 02:30:22,293 --> 02:30:24,696 WHEN WE LOOK AT THE MUCH MORE IN 3775 02:30:24,696 --> 02:30:26,964 DETAIL INTO THIS RNA SEQ DATA 3776 02:30:26,964 --> 02:30:32,570 WHAT WE SEE IS THAT THIS 3777 02:30:32,570 --> 02:30:34,072 PROGRESSION OF THE PRO T CELLS 3778 02:30:34,072 --> 02:30:35,640 THEY REALLY APPEAR TO BE BETWEEN 3779 02:30:35,640 --> 02:30:41,212 THE DN1 AND THE BACK EX-VIVO DN3 3780 02:30:41,212 --> 02:30:43,314 CELLS ALLOWING PC1 LINE WHICH 3781 02:30:43,314 --> 02:30:47,018 SEEMS TO BE THE DEVELOPMENTAL 3782 02:30:47,018 --> 02:30:49,721 PATHWAY WHICH DOES AN ESCAPE OR 3783 02:30:49,721 --> 02:30:52,590 IMMATURE CELLS INTO THE DOUBLE 3784 02:30:52,590 --> 02:30:54,525 POSITIVE THYMOCYTES. 3785 02:30:54,525 --> 02:30:57,061 SO WHAT WE SEE HERE IS REALLY 3786 02:30:57,061 --> 02:31:00,498 LIKE THERE IS AN UPREGULATION OF 3787 02:31:00,498 --> 02:31:02,800 WHAT THE TYPICAL T CELL 3788 02:31:02,800 --> 02:31:04,435 TRANSCRIPTION FACTORS ARE AND 3789 02:31:04,435 --> 02:31:07,805 DOWN REGULATION OF THE LINEAGES. 3790 02:31:07,805 --> 02:31:09,640 WHEN WE LOOK AT THE PATHWAY WE 3791 02:31:09,640 --> 02:31:13,945 REALLY HAVE LSK VERSUS PRO T 3792 02:31:13,945 --> 02:31:15,913 CELLS UPREGULATE ALL THE 3793 02:31:15,913 --> 02:31:16,981 PATHWAYS YOU WOULD EXPECT 3794 02:31:16,981 --> 02:31:18,816 INCLUDING T CELL DIFFERENTIATION 3795 02:31:18,816 --> 02:31:20,385 AND THEY ARE PRETTY MUCH 3796 02:31:20,385 --> 02:31:24,322 CORRESPONDING TO LSK TO DN2 AND 3797 02:31:24,322 --> 02:31:26,090 DN3 SUGGESTING THAT WHAT WE SEE 3798 02:31:26,090 --> 02:31:27,959 HAPPENING IF HAVE I TROA 3799 02:31:27,959 --> 02:31:28,760 CORRESPONDS MOACIALS TO WHAT WE 3800 02:31:28,760 --> 02:31:30,661 HAVE HAPPENING IN VIVO. 3801 02:31:30,661 --> 02:31:31,963 NOW, THE FACT THAT WE WERE ABLE 3802 02:31:31,963 --> 02:31:34,165 TO GENERATE THIS LARGE AMOUNT OF 3803 02:31:34,165 --> 02:31:38,636 CELLS GIVE US THE OPPORTUNITY TO 3804 02:31:38,636 --> 02:31:39,637 ADDRESS THE REQUIREMENTS FOR 3805 02:31:39,637 --> 02:31:44,442 NOTCH AND WHAT IS NOTCH DOING 3806 02:31:44,442 --> 02:31:48,613 BILY CRIP SEC AND MASS SPEC IT 3807 02:31:48,613 --> 02:31:51,649 WILL CHIP AS SPECTD AND HIGH C. 3808 02:31:51,649 --> 02:31:55,720 CHIP SEC WAS USED -- SEQ WAS 3809 02:31:55,720 --> 02:31:58,456 USED USING NOTCH TO CONFIRM ALL 3810 02:31:58,456 --> 02:32:01,292 THE NOTCH TARGETS ALSO HAVE THE 3811 02:32:01,292 --> 02:32:09,000 RBJ BINDING AND THE HISTOMARKS. 3812 02:32:09,000 --> 02:32:10,968 SO THE DATA WAS PRETTY GOOD 3813 02:32:10,968 --> 02:32:12,703 BECAUSE MOST OF THE PEAKS 3814 02:32:12,703 --> 02:32:14,138 CORRESPONDED TO NOTCH PEAK 3815 02:32:14,138 --> 02:32:19,710 RESPONDED TULS OUR RBJ BOOKS 3816 02:32:19,710 --> 02:32:20,912 WITH 99% OVERLAP AND THE OTHER 3817 02:32:20,912 --> 02:32:23,214 THING WAS THAT MOST OF THEM WERE 3818 02:32:23,214 --> 02:32:29,053 REFLECTING HAD ACTIVE CHROMATIN 3819 02:32:29,053 --> 02:32:34,959 ARE ACTIVE MARGE R RL BPJ VORKS 3820 02:32:34,959 --> 02:32:36,594 THEY CORRESPONDED TO SHIITES 3821 02:32:36,594 --> 02:32:39,797 WHERE WE EITHER HAD K27 3822 02:32:39,797 --> 02:32:43,868 ASSIMILATION OR K3 MUTATION. 3823 02:32:43,868 --> 02:32:46,337 SO AGAIN WHEN WE PLOTTED P, WHAT 3824 02:32:46,337 --> 02:32:52,710 WE SAW IS REALLY LIKE MOST OF 3825 02:32:52,710 --> 02:32:55,980 THE FLL THIS NRC SITES WERE 3826 02:32:55,980 --> 02:32:57,381 BINDING, ENHANCER SITES, RATHER 3827 02:32:57,381 --> 02:32:58,883 THAN PROMOTER SITES. 3828 02:32:58,883 --> 02:33:02,420 THE ONE WHICH WERE NOT ACTIVE SO 3829 02:33:02,420 --> 02:33:04,522 WHICH DID NOT OVERLAP WITH 3830 02:33:04,522 --> 02:33:07,258 EITHER HISTONE MARKS, ACTIVE 3831 02:33:07,258 --> 02:33:09,894 HISTONE MARKS WERE REALLY 3832 02:33:09,894 --> 02:33:13,631 DEPLETED BY PROMOTER SITES, 3833 02:33:13,631 --> 02:33:14,432 AGAIN AN ANSWER. 3834 02:33:14,432 --> 02:33:15,833 ONE THING WHICH BECAME VERY 3835 02:33:15,833 --> 02:33:18,236 OBVIOUS IS WHEN WE LOOKED AT ALL 3836 02:33:18,236 --> 02:33:21,339 THE TARGETS BY CHIP SEQ, WHAT WE 3837 02:33:21,339 --> 02:33:23,274 OBSERVED WAS THAT THE PATTERN OF 3838 02:33:23,274 --> 02:33:25,910 THE TARGETS, WHEN WE PROJECTED 3839 02:33:25,910 --> 02:33:28,412 THEM INTO A TIMELINE WAS VERY 3840 02:33:28,412 --> 02:33:28,980 DIFFERENT. 3841 02:33:28,980 --> 02:33:31,282 SO WE HAD TARGETS WHICH WERE 3842 02:33:31,282 --> 02:33:34,552 PROGRESSIVELY UPREGULATED, 3843 02:33:34,552 --> 02:33:36,954 UPREGULATED ONLY AT LATER 3844 02:33:36,954 --> 02:33:38,856 STAGES, OR TRANSIENTSLY INDUCED, 3845 02:33:38,856 --> 02:33:43,194 AS WELL AS DOWN REGULATED. 3846 02:33:43,194 --> 02:33:45,062 SO WHAT WAS CLEAR IS THAT THE T 3847 02:33:45,062 --> 02:33:46,898 CELL IDENTITY GENE WERE THE ONES 3848 02:33:46,898 --> 02:33:48,566 WHICH WERE MOSTLY UPREGULATED 3849 02:33:48,566 --> 02:33:51,936 AND MOSTLY UPREGULATED AT LATER 3850 02:33:51,936 --> 02:33:52,603 STAGES. 3851 02:33:52,603 --> 02:33:57,408 WHEREAS THE GENES, HERE ARE SOME 3852 02:33:57,408 --> 02:34:04,115 EXAMPLES, SO TCF ACTIVELY F 3853 02:34:04,115 --> 02:34:08,586 ELEVEN AND B AND OTHER COMPLEX 3854 02:34:08,586 --> 02:34:09,921 AND CD3. 3855 02:34:09,921 --> 02:34:12,323 SO ON THE OTHER SIDE, WHAT WAS 3856 02:34:12,323 --> 02:34:14,759 OBVIOUS THAT THE DOWN REGULATED 3857 02:34:14,759 --> 02:34:17,728 GENES WERE ASSOCIATED WITH 3858 02:34:17,728 --> 02:34:20,231 STEMNESS OR ALTERNATIVE 3859 02:34:20,231 --> 02:34:22,533 PATHWAYS, LIKE L1 OR I FORGOT 3860 02:34:22,533 --> 02:34:28,105 WHAT I HAVE HERE, SPI1. 3861 02:34:28,105 --> 02:34:30,207 SO WHAT WAS SURPRISING IS LIKE 3862 02:34:30,207 --> 02:34:33,010 NOTCH IS NORMALLY RELATED TO 3863 02:34:33,010 --> 02:34:34,245 INDUCTION OF TRANSCRIPTION AND 3864 02:34:34,245 --> 02:34:35,746 NOT REPRESSION OF TRANSCRIPTION. 3865 02:34:35,746 --> 02:34:37,748 SO WE WERE QUITE PUZZLED BY THE 3866 02:34:37,748 --> 02:34:40,918 FACT THAT WE OBSERVED 3867 02:34:40,918 --> 02:34:41,218 REPRESSION. 3868 02:34:41,218 --> 02:34:44,522 SO BASED ON OUR DATA, WE DECIDED 3869 02:34:44,522 --> 02:34:46,691 TO LOOK AT SUPER HYPHEN HANSERS 3870 02:34:46,691 --> 02:34:50,294 BECAUSE WE KNOW THAT SUPER 3871 02:34:50,294 --> 02:34:52,530 ENHANCER REGULATE IMAGE, SUPER 3872 02:34:52,530 --> 02:34:55,032 ENHANCER WE IDENTIFIED 680 SUPER 3873 02:34:55,032 --> 02:34:56,467 ENHANCER AND AGAIN WE SEE THE 3874 02:34:56,467 --> 02:34:58,903 SAME PATTERN, WE SEE GENES WHICH 3875 02:34:58,903 --> 02:35:01,072 ARE REGULATED BY SUPER ENHANCER 3876 02:35:01,072 --> 02:35:03,074 WHICH ARE ASSOCIATED WITH THE T 3877 02:35:03,074 --> 02:35:05,977 CELL PROGRAM AND GENES WHICH ARE 3878 02:35:05,977 --> 02:35:08,946 DOWN REGULATED WHICH AGAIN ARE 3879 02:35:08,946 --> 02:35:10,581 REGULATED BY SUPER ENHANCER 3880 02:35:10,581 --> 02:35:15,152 WHICH ARE LINKED TO STEMNESS AND 3881 02:35:15,152 --> 02:35:16,354 ALTERNATIVE LINEAGES. 3882 02:35:16,354 --> 02:35:20,191 SO WHEN WE LOOKED MORE CAREFULLY 3883 02:35:20,191 --> 02:35:22,159 AT THE FACTORS WHAT WE OBSERVE 3884 02:35:22,159 --> 02:35:24,161 IS THAT THERE IS A LATE NETWORK 3885 02:35:24,161 --> 02:35:25,863 OR TRANSCRIPTIONAL NETWORK WHICH 3886 02:35:25,863 --> 02:35:27,898 SEEMS TO BE RELATED TO THE T 3887 02:35:27,898 --> 02:35:29,867 CELL PROGRAM, AND THERE IS AN 3888 02:35:29,867 --> 02:35:31,302 EARLY NETWORK WHICH SEEMS TO BE 3889 02:35:31,302 --> 02:35:34,472 RELATED TOP THE DOWN REGULATION 3890 02:35:34,472 --> 02:35:37,441 OF ALTERNATIVE LINEAGES AND 3891 02:35:37,441 --> 02:35:37,708 STEMNESS. 3892 02:35:37,708 --> 02:35:39,977 AND THIS IS THE ONE WHICH STARTS 3893 02:35:39,977 --> 02:35:42,380 TO, SO IT'S EXPRESSED EARLIER, 3894 02:35:42,380 --> 02:35:47,985 AND THIS WILL BE DELAYED 3895 02:35:47,985 --> 02:35:50,054 NETWORK. 3896 02:35:50,054 --> 02:35:53,224 SO WE TOOK ADVANTAGE OF THE THIS 3897 02:35:53,224 --> 02:35:55,559 SEQUENCING DATA WHICH WAS 3898 02:35:55,559 --> 02:36:01,499 GENERATED BY ELLEN, AND SO WE 3899 02:36:01,499 --> 02:36:10,875 PLOTTED ALL THE, LIKE AGAIN N A 3900 02:36:10,875 --> 02:36:12,510 PSEUDOTIMELINE FROM PROGENITORS 3901 02:36:12,510 --> 02:36:16,247 DOWN TO N3, WE HAVE TARGETS 3902 02:36:16,247 --> 02:36:20,084 WHICH ARE DOWN REGULATED AND 3903 02:36:20,084 --> 02:36:21,952 TARGETS UPREGULATED IN TERMS OF 3904 02:36:21,952 --> 02:36:23,587 OR OPENED UP BY ACCESSIBILITY. 3905 02:36:23,587 --> 02:36:25,756 SO WHAT WE HYPOTHESIZED IS THAT 3906 02:36:25,756 --> 02:36:28,292 THERE IS SOMEHOW AN EARLY 3907 02:36:28,292 --> 02:36:30,361 NETWORK WHICH IS ACCESSIBLE AND 3908 02:36:30,361 --> 02:36:34,065 NEEDS TO BE LOCKED, THERE IS 3909 02:36:34,065 --> 02:36:35,733 NETWORKS FOR CLUSTER ONE WHICH 3910 02:36:35,733 --> 02:36:37,568 REMAINS OPEN ALL THE TIME, WHICH 3911 02:36:37,568 --> 02:36:39,103 IS LIKELY LINKED TO GENES WHICH 3912 02:36:39,103 --> 02:36:42,740 ARE REQUIRED ALL THE TIME, AND 3913 02:36:42,740 --> 02:36:44,475 HAD THERE ARE SITES OR TARGETS 3914 02:36:44,475 --> 02:36:47,211 WHICH TBLONG THE LATE NETWORK 3915 02:36:47,211 --> 02:36:50,081 WHICH ARE INCLUDED AT THESE 3916 02:36:50,081 --> 02:36:52,450 LATER POINTS AND THEN ARE OPENED 3917 02:36:52,450 --> 02:36:53,050 UP. 3918 02:36:53,050 --> 02:36:54,652 AND WHAT WAS SURPRISING IS WHEN 3919 02:36:54,652 --> 02:36:56,620 WE LOOKED AT WHAT ARE THE 3920 02:36:56,620 --> 02:36:59,056 MOTIVES FOR WHEN WE PERFORMED 3921 02:36:59,056 --> 02:37:02,093 THE MOTIVE ANALYSIS, IT'S REALLY 3922 02:37:02,093 --> 02:37:04,428 LIKE WE HAVE EDS FACTORS WHICH 3923 02:37:04,428 --> 02:37:05,696 ARE LINKED TO THEY RECALL 3924 02:37:05,696 --> 02:37:07,231 NETWORK OR CLUSTER TWO WHICH 3925 02:37:07,231 --> 02:37:11,302 GETS DOWN REGULATED AND WE HAVE 3926 02:37:11,302 --> 02:37:13,270 TCF AND START TARGETS WHICH ARE 3927 02:37:13,270 --> 02:37:14,905 RELATED TO THIS LATE NETWORK 3928 02:37:14,905 --> 02:37:17,541 WHICH BECOMES OPEN P AT LATER 3929 02:37:17,541 --> 02:37:19,944 TIME POINTS. 3930 02:37:19,944 --> 02:37:21,378 LY WE WANTED TO CON NATIVE 3931 02:37:21,378 --> 02:37:23,714 AMERICAN THE TARGETS -- WE 3932 02:37:23,714 --> 02:37:26,717 WANTED TO CONFIRM THAT THE 3933 02:37:26,717 --> 02:37:27,952 TARGETS OF THE TRANSCRIPTION 3934 02:37:27,952 --> 02:37:29,453 FACTOR WE IDENTIFIED WERE ALSO 3935 02:37:29,453 --> 02:37:31,355 PRESENT OF PART OF THE NOTCH 3936 02:37:31,355 --> 02:37:34,425 COMPLEX, WE PERFORMED MASS SPEC, 3937 02:37:34,425 --> 02:37:37,027 IT'S NOT HERE, HERE, MASS SPEC 3938 02:37:37,027 --> 02:37:40,765 ANALYSIS, AND WE INDEED 3939 02:37:40,765 --> 02:37:51,709 IDENTIFIED TCF7 FACTOR, AND WILD 3940 02:37:51,976 --> 02:37:54,311 RBPJ FACTORS SUGGESTING THAT EGS 3941 02:37:54,311 --> 02:37:56,046 FACTORS WERE LINKED TO THIS 3942 02:37:56,046 --> 02:37:57,381 EARLY NETWORK AND DOWN 3943 02:37:57,381 --> 02:37:58,582 REGULATION OF TARGETS WHICH ARE 3944 02:37:58,582 --> 02:38:02,319 RELATED TO STEM CELLS AND 3945 02:38:02,319 --> 02:38:06,023 PROLIFERATION AND TCF IF IN 3946 02:38:06,023 --> 02:38:08,125 RANKS GENE OR START GENES WERE 3947 02:38:08,125 --> 02:38:10,394 RELATED TO TARGETS THAT WERE 3948 02:38:10,394 --> 02:38:12,997 INDUCED UPON DIFFERENTIATION. 3949 02:38:12,997 --> 02:38:15,032 TO VALIDATE, AGAIN, BECAUSE WE 3950 02:38:15,032 --> 02:38:16,567 HAD THE ADVANTAGE OF LARGE 3951 02:38:16,567 --> 02:38:20,171 NUMBER OF CELLS WE PERFORMED A 3952 02:38:20,171 --> 02:38:22,473 CAS9 DELETION AND BECAUSE OF THE 3953 02:38:22,473 --> 02:38:26,443 RANKS GENES WE KNOW THAT RANKS 3954 02:38:26,443 --> 02:38:31,015 FUNCTION, WE THOUGHT ECF MALE 3955 02:38:31,015 --> 02:38:32,216 OVERLAPPING FUNCTIONS, WE 3956 02:38:32,216 --> 02:38:34,618 DECIDED TO DO EITHER SINGLY OR 3957 02:38:34,618 --> 02:38:35,953 DOUBLE KNOCKOUTS AND WE LOOKED 3958 02:38:35,953 --> 02:38:38,556 AT THE OUTPUT OF CELLS. 3959 02:38:38,556 --> 02:38:41,425 SO SURPRISINGLY, SO THIS IS THE 3960 02:38:41,425 --> 02:38:42,593 CONTROL, THIS IS WHAT WE EXPECT 3961 02:38:42,593 --> 02:38:44,228 AFTER THREE DAYS OF CULTURE, 3962 02:38:44,228 --> 02:38:47,164 LIKE JUST THE DN1 WHICH STARTED 3963 02:38:47,164 --> 02:38:50,067 USING DN2 AND THE NOTCH1 CONTROL 3964 02:38:50,067 --> 02:38:52,536 WHICH IS BLOCKED AT THE DN1 3965 02:38:52,536 --> 02:38:53,971 STAGE, SURPRISINGLY THE DELETION 3966 02:38:53,971 --> 02:38:59,310 OF THIS ETS FACTOR LED TO AN 3967 02:38:59,310 --> 02:39:01,846 INCREASE INTO P THE -- OR A 3968 02:39:01,846 --> 02:39:05,349 FASTER PROGRESSION INTO THIS 3969 02:39:05,349 --> 02:39:05,649 DEVELOPMENT. 3970 02:39:05,649 --> 02:39:13,791 AM ARE WE DONE? 3971 02:39:13,791 --> 02:39:16,293 SO SHOWED -- SO HE WILL -- SO 3972 02:39:16,293 --> 02:39:17,628 ELLEN SHOWED BEAUTIFULLY THAT IT 3973 02:39:17,628 --> 02:39:19,697 TAKES TIME TO GET THE ENTIRE 3974 02:39:19,697 --> 02:39:22,333 PROGRAM READY TO BECOME T CELLS, 3975 02:39:22,333 --> 02:39:24,735 AND SINCE TIME IS NOT PRESENT 3976 02:39:24,735 --> 02:39:28,772 THIS THOSE CELLS, AND SO LIKE 3977 02:39:28,772 --> 02:39:30,507 NOW LIKE I DON'T WANT TO GO, 3978 02:39:30,507 --> 02:39:31,809 BECAUSE WE HAVE ONLY ONE MINUTE, 3979 02:39:31,809 --> 02:39:33,244 SO THERE IS A LOT OF OTHER GENES 3980 02:39:33,244 --> 02:39:34,912 WHICH ARE SUPPOSED TO BE 3981 02:39:34,912 --> 02:39:39,183 EXPRESSED WHICH ARE DISTURBED OR 3982 02:39:39,183 --> 02:39:42,119 ALTERED IN THEIR EXPRESSION 3983 02:39:42,119 --> 02:39:42,653 PROFILES. 3984 02:39:42,653 --> 02:39:44,421 ONE THING WHICH WAS STRIKING, WE 3985 02:39:44,421 --> 02:39:46,957 PERFORMED INJECTION, INTRATHYMIC 3986 02:39:46,957 --> 02:39:50,227 INJECTION USING ULTRASOUND TO 3987 02:39:50,227 --> 02:39:51,428 LOOK AT THE CELLS, SO THIS IS 3988 02:39:51,428 --> 02:39:53,097 WHAT HAPPENS. 3989 02:39:53,097 --> 02:39:56,133 WE INJECT THEM H THE CAS9 AND WE 3990 02:39:56,133 --> 02:39:58,435 END UP HAVING DOUBLE POSITIVE 3991 02:39:58,435 --> 02:40:00,971 AND DOUBLE-NEGATIVE CELLS. 3992 02:40:00,971 --> 02:40:07,845 THE DOUBLE-NEGATIVE REALLY, WE 3993 02:40:07,845 --> 02:40:09,713 HAVE VERY FEW CELLS SURVIVING 3994 02:40:09,713 --> 02:40:11,782 THIS PROCESS, AND THEY ARE 3995 02:40:11,782 --> 02:40:13,751 REALLY DOUBLE-NEGATIVES. 3996 02:40:13,751 --> 02:40:19,189 WHEN WE TAKE OUR ETS DOUBLE 3997 02:40:19,189 --> 02:40:20,391 KNOCK-OUTS, WE SEE THE SAME 3998 02:40:20,391 --> 02:40:22,593 THING, THE CELLS WHICH ARE GUIDE 3999 02:40:22,593 --> 02:40:26,297 POSITIVE DO NOT SURVIVE. 4000 02:40:26,297 --> 02:40:28,465 NOW AND THIS IS LIKE QUANTIFIED 4001 02:40:28,465 --> 02:40:29,233 HERE. 4002 02:40:29,233 --> 02:40:30,901 SO WHEN YOU DELETE BOTH GENES, 4003 02:40:30,901 --> 02:40:33,304 IT'S NOT THAT THEY GROW INTO 4004 02:40:33,304 --> 02:40:34,571 ALTERNATIVE LINEAGES AS THEY 4005 02:40:34,571 --> 02:40:37,908 WOULD DO FOR NOTCH1, THEY REALLY 4006 02:40:37,908 --> 02:40:40,110 DON'T SURVIVE IN VIVO. 4007 02:40:40,110 --> 02:40:43,781 SO NOW WHAT HAPPENS IS THAT THEY 4008 02:40:43,781 --> 02:40:48,519 HAVE A DEFECT IN THE 4009 02:40:48,519 --> 02:40:51,655 PROLIFERATION, SO THE GENES, SO 4010 02:40:51,655 --> 02:40:53,624 THIS INCREASE IN THE PROGRESSION 4011 02:40:53,624 --> 02:40:57,127 SEEMS TO BE RELATED TO INCREASED 4012 02:40:57,127 --> 02:40:59,463 IN PROLIFERATION. 4013 02:40:59,463 --> 02:41:00,564 NOW, WE IDENTIFIED TWO OF THE 4014 02:41:00,564 --> 02:41:03,834 GENES WHICH ARE RESPONSIBLE FOR 4015 02:41:03,834 --> 02:41:06,270 THIS DEFECT AND THEY ARE REALLY 4016 02:41:06,270 --> 02:41:08,439 RELATED TO STEMNESS. 4017 02:41:08,439 --> 02:41:13,043 AND THIS IS KLF2 AND KLF4. 4018 02:41:13,043 --> 02:41:15,012 NOW, BECAUSE WE DON'T HAVE TIME, 4019 02:41:15,012 --> 02:41:18,382 I WILL JUST SKIP TO THIS SLIDE. 4020 02:41:18,382 --> 02:41:22,119 WHAT WE SEE LIKE WHEN WE PUT THE 4021 02:41:22,119 --> 02:41:23,954 CULTURES, THE PRO T CELLS IN 4022 02:41:23,954 --> 02:41:25,589 CULTURE, WE STIMULATE THEM ON 4023 02:41:25,589 --> 02:41:27,257 NOTCH AND THEN WE REMOVE NOTCH 4024 02:41:27,257 --> 02:41:28,859 FOR FOUR HOURS AND THEN WE LOOK 4025 02:41:28,859 --> 02:41:32,062 AT THE CHIP SEQ, IT'S THIS 4026 02:41:32,062 --> 02:41:36,200 DIRECT REGULATION OF NOTCH OF 4027 02:41:36,200 --> 02:41:39,370 THESE GENES WHICH ARE MEDIATED 4028 02:41:39,370 --> 02:41:41,572 THROUGH FREE DISMRG WE SEE HERE 4029 02:41:41,572 --> 02:41:44,408 THAT INDEED THE REMOVAL OF NOTCH 4030 02:41:44,408 --> 02:41:45,743 FROM NOTCH SIGNALLING LEADS TO 4031 02:41:45,743 --> 02:41:49,246 AN INDUCTION OF KLF2 AND ALSO 4032 02:41:49,246 --> 02:41:50,447 KLF4. 4033 02:41:50,447 --> 02:41:52,316 THERE'S ABOUT 25 FOLD INCREASE 4034 02:41:52,316 --> 02:41:54,718 OF THOSE GENES, AS EXPECTED 4035 02:41:54,718 --> 02:41:57,454 NOTCH TARGETS LIKE IL2 RECEPTOR 4036 02:41:57,454 --> 02:41:59,023 IS DOWN REGULATED. 4037 02:41:59,023 --> 02:42:03,160 AND THE ONLY GENE WHICH SEEMS TO 4038 02:42:03,160 --> 02:42:06,463 BE REMOVED IS REALLY NOTCH. 4039 02:42:06,463 --> 02:42:12,436 SO LBJ REMAINS ATOOCHED THE KF2 4040 02:42:12,436 --> 02:42:15,272 GENE AS FLE1 AND RFG, WE BELIEVE 4041 02:42:15,272 --> 02:42:17,107 THERE IS A COMPLEX FORMED 4042 02:42:17,107 --> 02:42:21,011 BETWEEN NOTCH RBPJ AND FLEET 4043 02:42:21,011 --> 02:42:23,614 FACTOR WHICH MEDIATES THE 4044 02:42:23,614 --> 02:42:25,349 REPRESSION OF KLF GENES AND 4045 02:42:25,349 --> 02:42:28,752 LEADS TO THIS DIFFERENTIATION OR 4046 02:42:28,752 --> 02:42:31,355 THIS FAST DIFFERENTIATION, AND 4047 02:42:31,355 --> 02:42:33,791 THIS IS, SO THIS IS THE MODEL 4048 02:42:33,791 --> 02:42:36,860 WHICH WE THINK IS LIKE, SO UNDER 4049 02:42:36,860 --> 02:42:39,797 NORMAL CONDITION, NOTCH RECRUITS 4050 02:42:39,797 --> 02:42:41,098 THE EARLY NETWORK, WHICH 4051 02:42:41,098 --> 02:42:43,300 INCLUDES ETS FACTORS, AND THIS 4052 02:42:43,300 --> 02:42:46,170 LEADS TO THE REPRESSION HAD OF 4053 02:42:46,170 --> 02:42:48,472 GENES WHICH ARE LINKED TO 4054 02:42:48,472 --> 02:42:50,808 PROLIFERATION AND STEMNESS. 4055 02:42:50,808 --> 02:42:52,709 AND THE EARLY T CELL PROGRAM CAN 4056 02:42:52,709 --> 02:42:55,446 START AND THIS LEADS TO THE 4057 02:42:55,446 --> 02:42:55,879 PROGRESSION. 4058 02:42:55,879 --> 02:42:58,882 WHEN WE HAVE LACK OF NOTCH 4059 02:42:58,882 --> 02:43:00,951 SIGNALLING, THIS T CELL PROGRAM 4060 02:43:00,951 --> 02:43:03,353 CM NEVER OCCUR SO WE HAVE 4061 02:43:03,353 --> 02:43:06,623 DIVERSION INTO MYELOID OR WHYLY 4062 02:43:06,623 --> 02:43:07,825 ALTERNATIVE LINEAGES. 4063 02:43:07,825 --> 02:43:12,296 BUT WHEN WE HAVE THE WSM LAC 4064 02:43:12,296 --> 02:43:15,265 FACTOR WE SEE BOTH PROGRAMS ARE 4065 02:43:15,265 --> 02:43:16,700 INDUCED, NOTCH IS ACTIVE, THEY 4066 02:43:16,700 --> 02:43:18,669 RECEIVE THE SIGNAL, BUT THE 4067 02:43:18,669 --> 02:43:19,870 STEMNESS AND ALTERNATIVE 4068 02:43:19,870 --> 02:43:22,372 PROGRAMS CAN NOT BE DOWN 4069 02:43:22,372 --> 02:43:22,639 REGULATED. 4070 02:43:22,639 --> 02:43:25,209 SO CAN SURVIVE IN VITRO WHERE WE 4071 02:43:25,209 --> 02:43:26,543 GIVE THEM EVERYTHING THEY NEED, 4072 02:43:26,543 --> 02:43:28,278 BUT AS SOON AS WE PUT THEM BACK 4073 02:43:28,278 --> 02:43:31,482 IN VIVO, THEY SOMEHOW CRASH 4074 02:43:31,482 --> 02:43:37,054 TRANSCRIPTIONALLY AND CANNOT 4075 02:43:37,054 --> 02:43:40,891 SURVIVE THIS PHENOMENON. 4076 02:43:40,891 --> 02:43:42,192 SO WITH THAT, AND I'M PROBABLY 4077 02:43:42,192 --> 02:43:43,427 OVER TIME AND I APOLOGIZE, I 4078 02:43:43,427 --> 02:43:44,595 WOULD LIKE TO ACKNOWLEDGE THE 4079 02:43:44,595 --> 02:43:48,132 PERSON WHO DID MOST OF THE WORK, 4080 02:43:48,132 --> 02:43:52,269 LADEN, WITH THE HELP OF GOLGI 4081 02:43:52,269 --> 02:43:54,338 FOR SOME OF THE INFORMATIC 4082 02:43:54,338 --> 02:43:58,008 ANALYSIS, PEOPLE IN LAB AND WILL 4083 02:43:58,008 --> 02:43:59,376 THERE ARE MORE NOW WHO HAVE 4084 02:43:59,376 --> 02:44:01,278 JOINED AND ALL THE OTHER 4085 02:44:01,278 --> 02:44:10,988 COLLABORATOR HIGH SCHOOL. -- AND 4086 02:44:10,988 --> 02:44:12,823 ALL THE OTHER COLLABORATORS. 4087 02:44:12,823 --> 02:44:14,825 I'M HAPPY TO TAKE QUESTIONS. 4088 02:44:14,825 --> 02:44:15,092 [APPLAUSE] 4089 02:44:15,092 --> 02:44:16,994 >> ROXANE, I THOUGHT YOU STUDIED 4090 02:44:16,994 --> 02:44:17,794 DEN DRITD I CAN CELLS. 4091 02:44:17,794 --> 02:44:18,862 >> I. 4092 02:44:18,862 --> 02:44:19,129 [LAUGHTER] 4093 02:44:19,129 --> 02:44:21,398 >> I HAVE A QUESTION ABOUT THE 4094 02:44:21,398 --> 02:44:21,798 MPP EXPRESSION. 4095 02:44:21,798 --> 02:44:23,233 >> SURE. 4096 02:44:23,233 --> 02:44:26,637 >> IS NOTCH EXPRESSION 4097 02:44:26,637 --> 02:44:28,939 STOCHASTIC AT THAT POINT? 4098 02:44:28,939 --> 02:44:30,140 DELTA LIGAND? 4099 02:44:30,140 --> 02:44:31,441 >> IN THE BONE MARROW? 4100 02:44:31,441 --> 02:44:33,443 GROO BONE MARROW, TO PUSH IT 4101 02:44:33,443 --> 02:44:34,444 MORE TOWARD T CELL AT THAT 4102 02:44:34,444 --> 02:44:34,745 POINT? 4103 02:44:34,745 --> 02:44:36,146 >> IT'S UNCLEAR TO ME YET. 4104 02:44:36,146 --> 02:44:38,682 I THINK IT'S STILL UNCLEAR. 4105 02:44:38,682 --> 02:44:40,851 LESS SHOWED THAT THERE IS A VERY 4106 02:44:40,851 --> 02:44:42,286 TRANSIENT INDUCTION OF NOTCH 4107 02:44:42,286 --> 02:44:43,420 SIGNALLING WHICH HAPPENS IN THE 4108 02:44:43,420 --> 02:44:45,022 BONE MARROW. 4109 02:44:45,022 --> 02:44:46,223 >> AND -- 4110 02:44:46,223 --> 02:44:46,623 >> YEAH. 4111 02:44:46,623 --> 02:44:49,660 >> IS THAT STOCHASTIC OR 4112 02:44:49,660 --> 02:44:51,061 WHETHER, YOU KNOW, IT'S GOING TO 4113 02:44:51,061 --> 02:44:51,795 GO B OR -- 4114 02:44:51,795 --> 02:44:56,433 >> IT MUST BE SOME STROMA WHICH 4115 02:44:56,433 --> 02:44:58,936 EXPRESSES IT, WHICH DRIVES IT, 4116 02:44:58,936 --> 02:45:00,137 THE DELTA. 4117 02:45:00,137 --> 02:45:03,440 SO YEAH, I DON'T KNOW. 4118 02:45:03,440 --> 02:45:03,640 YEAH. 4119 02:45:03,640 --> 02:45:06,710 >> SO VERY IMPRESSIVE TALK. 4120 02:45:06,710 --> 02:45:09,112 TWO QUESTIONS. 4121 02:45:09,112 --> 02:45:11,982 FIRST, THERE IS A IF FLY ONE IN 4122 02:45:11,982 --> 02:45:12,950 EARLY KNOCK-OUT TORQUES THEY 4123 02:45:12,950 --> 02:45:16,453 HAVE ANY DEFECT FRP ON THE 4124 02:45:16,453 --> 02:45:16,687 BLOCK? 4125 02:45:16,687 --> 02:45:18,088 >> SO LIKE THE PROBLEM IS LIKE 4126 02:45:18,088 --> 02:45:21,692 THE TIMING WHEN IT'S DELETED. 4127 02:45:21,692 --> 02:45:23,527 SO IN THE ONE, THE MOUSE WHICH 4128 02:45:23,527 --> 02:45:26,096 WAS GENERATED WAS A FLOX AND IT 4129 02:45:26,096 --> 02:45:28,498 WAS A CD4 SO IT'S TOO LATE. 4130 02:45:28,498 --> 02:45:33,203 >> BUT DOESN'T NIRS CK CRE OR -- 4131 02:45:33,203 --> 02:45:34,738 INTLO YEAH, IT WAS KNOCKED DOWN 4132 02:45:34,738 --> 02:45:34,972 EARLIER. 4133 02:45:34,972 --> 02:45:37,774 >> VERY SIMILAR TO S1 AND S2, 4134 02:45:37,774 --> 02:45:38,642 TWO GENES ON -- 4135 02:45:38,642 --> 02:45:39,109 >> YEAH. 4136 02:45:39,109 --> 02:45:41,945 >> SO S12 IS NECESSARY FOR CD4 4137 02:45:41,945 --> 02:45:42,913 DEVELOPMENT AND DOES THAT BLOCK 4138 02:45:42,913 --> 02:45:44,248 AT THE DOUBLE-NEGATIVE STAGE, 4139 02:45:44,248 --> 02:45:49,186 HAVE YOU LOOKED IN YOUR DOUBLE 4140 02:45:49,186 --> 02:45:52,356 DEFICIENT, ONE OR TWO STATE 4141 02:45:52,356 --> 02:45:52,556 OR -- 4142 02:45:52,556 --> 02:45:54,858 >> SO BECAUSE I DIDN'T SHOW IT 4143 02:45:54,858 --> 02:45:57,694 BUT WE HAVE DONE THE RNA SEQ TO 4144 02:45:57,694 --> 02:45:58,929 TRY TO UNDERSTAND AND THAT'S HOW 4145 02:45:58,929 --> 02:46:02,766 WE IDENTIFY THELY FACTORS, SO WE 4146 02:46:02,766 --> 02:46:06,036 HAVE ALMOST COMPLETE DELETION OF 4147 02:46:06,036 --> 02:46:08,872 ERG BUT WE DON'T HAVE, WE HAVE 4148 02:46:08,872 --> 02:46:11,608 HALF PERCENT DELETION OF FLE ONE 4149 02:46:11,608 --> 02:46:13,343 SO IT'S A BIPOPULATION, SO WE 4150 02:46:13,343 --> 02:46:15,679 WERE WORK WITH BOC MAKES IT VERY 4151 02:46:15,679 --> 02:46:16,880 COMPLICATED TO ADDRESS YOUR 4152 02:46:16,880 --> 02:46:17,814 QUESTION PRECISELY. 4153 02:46:17,814 --> 02:46:20,917 >> SO YOU DEENTD DO IT AS A 4154 02:46:20,917 --> 02:46:21,285 COMPENSATORY -- 4155 02:46:21,285 --> 02:46:22,352 >> WEBLES. 4156 02:46:22,352 --> 02:46:24,187 WEBLES BECAUSE LIKE EVERYTHING 4157 02:46:24,187 --> 02:46:28,392 WE SEE REQUIRES DELETION OF BOTH 4158 02:46:28,392 --> 02:46:31,561 SO I BELIEVE THERE IS SOME SORT 4159 02:46:31,561 --> 02:46:32,095 OF OVERLAP. 4160 02:46:32,095 --> 02:46:33,497 >> OKAY H. 4161 02:46:33,497 --> 02:46:35,098 >> AND PROBABLY, YEAH -- 4162 02:46:35,098 --> 02:46:36,366 >> BETWEEN ONE AND TWO. 4163 02:46:36,366 --> 02:46:37,367 >> YEAH. 4164 02:46:37,367 --> 02:46:39,436 >> OKAY. 4165 02:46:39,436 --> 02:46:39,736 THANK YOU. 4166 02:46:39,736 --> 02:46:40,304 >> THANK YOU. 4167 02:46:40,304 --> 02:46:41,204 THANK YOU. 4168 02:46:41,204 --> 02:46:43,740 >> SO BEFORE ROXANE FINISHES, 4169 02:46:43,740 --> 02:46:45,008 ROXANE, YOU GET TO COME BACK, 4170 02:46:45,008 --> 02:46:46,877 BECAUSE I THINK THAT THIS IS P 4171 02:46:46,877 --> 02:46:48,645 THE -- ROXANE DESERVES A SPECIAL 4172 02:46:48,645 --> 02:46:50,213 AWARD AND WE ARE REALLY GRATEFUL 4173 02:46:50,213 --> 02:46:51,481 TO HER BECAUSE FOR THOSE OF YOU 4174 02:46:51,481 --> 02:46:52,382 WHO ARE NOT AWARE ACCIDENT SHE 4175 02:46:52,382 --> 02:46:54,551 HAD A BABY TWO AND A HALF WEEKS 4176 02:46:54,551 --> 02:46:55,185 AGO. 4177 02:46:55,185 --> 02:46:55,452 [APPLAUSE] 4178 02:46:55,452 --> 02:46:59,122 TWO AND A HALF WEEKS AGO! 4179 02:46:59,122 --> 02:47:09,299 [APPLAUSE] 4180 02:47:33,390 --> 02:47:34,825 >> WE'RE GOING TO HAVE THE LAST 4181 02:47:34,825 --> 02:47:38,095 SPEAKER FOR THIS SESSION, ELLEN 4182 02:47:38,095 --> 02:47:41,064 ROTHENBERG FROM CAL TECH, AND 4183 02:47:41,064 --> 02:47:42,999 TMS FOR BEING INVITED FROM CAL 4184 02:47:42,999 --> 02:47:44,468 TECH, YOU GET AN EXTRA FIVE 4185 02:47:44,468 --> 02:47:45,102 MINUTES TO TALK. 4186 02:47:45,102 --> 02:47:47,170 >> THANK YOU. 4187 02:47:47,170 --> 02:47:48,004 >> WELCOME. 4188 02:47:48,004 --> 02:47:50,240 >> THANK YOU SO MUCH. 4189 02:47:50,240 --> 02:47:50,507 [APPLAUSE] 4190 02:47:50,507 --> 02:47:54,277 I JUST WANT TO ALSO CONGRATULATE 4191 02:47:54,277 --> 02:47:55,512 ROXANE BECAUSE SHE DID A 4192 02:47:55,512 --> 02:47:56,146 BEAUTIFUL JOB. 4193 02:47:56,146 --> 02:47:59,449 SHE HAS MUCH MORE BEAUTIFUL 4194 02:47:59,449 --> 02:48:06,890 NOTCH CHIP SEQ DATA THAN WE DID 4195 02:48:06,890 --> 02:48:09,192 AND OF NOT BEING TOO TERRIBLE 4196 02:48:09,192 --> 02:48:09,893 OVER TIME. 4197 02:48:09,893 --> 02:48:11,962 THANK YOU SO MUCH FOR HAVING ME 4198 02:48:11,962 --> 02:48:12,162 HERE. 4199 02:48:12,162 --> 02:48:13,397 I WANT TO PICK UP ON SOME OF THE 4200 02:48:13,397 --> 02:48:14,564 SAME THINGS THAT ROXANE STARTED 4201 02:48:14,564 --> 02:48:15,899 TO TALK ABOUT AND REALLY DISCUSS 4202 02:48:15,899 --> 02:48:17,200 WHAT HAPPENS IN THE EARLIEST 4203 02:48:17,200 --> 02:48:18,969 PARTS OF T CELL DEVELOPMENT. 4204 02:48:18,969 --> 02:48:24,074 SO THIS IS JUST TOINT OUT THAT 4205 02:48:24,074 --> 02:48:27,411 ALL OF IMMUNOLOGY BEGINS TO 4206 02:48:27,411 --> 02:48:28,678 HAPPEN FROM THE TIME THAT T 4207 02:48:28,678 --> 02:48:31,114 CELLS BEGIN TO BE EXPRESSED IN T 4208 02:48:31,114 --> 02:48:34,184 CELLS WE'RE REALLY INTERESTED IN 4209 02:48:34,184 --> 02:48:36,920 THE FROM DEVELOPMENT OF T CELLS 4210 02:48:36,920 --> 02:48:39,256 IN STAGES LEADING COMMITMENT TO 4211 02:48:39,256 --> 02:48:40,590 BECOME T CELLS BEFORE T CELLS 4212 02:48:40,590 --> 02:48:43,093 ARE EVEN LOWED TO BEGIN TO DO 4213 02:48:43,093 --> 02:48:45,128 FULL REARRANGEMENTS AND AS 4214 02:48:45,128 --> 02:48:46,696 ROXANE JUST TOLD YOU, THIS IS 4215 02:48:46,696 --> 02:48:48,432 INTENSELY DEPENDENT ON NOTCH 4216 02:48:48,432 --> 02:48:49,332 SIGNALLING BOTH FOR THE 4217 02:48:49,332 --> 02:48:50,200 ACQUISITION OF T CELL PROPERTIES 4218 02:48:50,200 --> 02:48:53,036 AND FOR THE LOSS OF POTENTIAL 4219 02:48:53,036 --> 02:48:55,138 FOR B GRUN LOW SIGHT PREDICT 4220 02:48:55,138 --> 02:48:58,708 CELL ILC AND THEN K CELL 4221 02:48:58,708 --> 02:48:59,142 DEVELOPMENT. 4222 02:48:59,142 --> 02:49:01,912 SO WHAT'S HAPPENING IN THE CELLS 4223 02:49:01,912 --> 02:49:03,213 AT THIS TIME SM YOU GOT A SENSE 4224 02:49:03,213 --> 02:49:04,981 OF THIS FROM ROXANE, WE WANT TO 4225 02:49:04,981 --> 02:49:06,082 SIMPLIFY THIS A LITTLE BIT TO 4226 02:49:06,082 --> 02:49:07,384 SAY THERE'S THREE GROUPS OF 4227 02:49:07,384 --> 02:49:09,453 TRANSCRIPTION FACTORS IN TERMS 4228 02:49:09,453 --> 02:49:12,055 OF DEVELOPMENTAL EXPRESSION 4229 02:49:12,055 --> 02:49:13,957 PATTERNS THAT ARE ALL ACTUALLY 4230 02:49:13,957 --> 02:49:15,392 ESSENTIAL IN VIVO FOR MAKING T 4231 02:49:15,392 --> 02:49:16,560 CELLS, THAT INCLUDES THE OBVIOUS 4232 02:49:16,560 --> 02:49:19,529 SUSPECTS THAT ARE UPREGULATED 4233 02:49:19,529 --> 02:49:22,365 SPECIFICALLY IN THE THYMUS LIKE 4234 02:49:22,365 --> 02:49:23,967 BCL11B WHICH WE'LL TALK ABOUT A 4235 02:49:23,967 --> 02:49:27,704 FAIR BIT, GATA3 AND TCF7 WHICH 4236 02:49:27,704 --> 02:49:29,139 IS DRAMATICALLY UPREGULATED 4237 02:49:29,139 --> 02:49:30,907 EARLY ON AND ALSO DOWN 4238 02:49:30,907 --> 02:49:32,142 REGULATION OF STEM AND 4239 02:49:32,142 --> 02:49:33,009 PROGENITOR TRANSCRIPTION FACTORS 4240 02:49:33,009 --> 02:49:34,311 THAT YOU JUST HEARD INTRODUCED 4241 02:49:34,311 --> 02:49:39,282 BY ROXANE AND AT THE SAME TIME A 4242 02:49:39,282 --> 02:49:41,251 WHOLE SET OF TRANSCRIPTION 4243 02:49:41,251 --> 02:49:42,552 FACTORS INHERITED FROM THE STEM 4244 02:49:42,552 --> 02:49:43,487 AND PROGENITOR CELLS AND 4245 02:49:43,487 --> 02:49:45,288 CONTINUE TO BE EXPRESSED ALMOST 4246 02:49:45,288 --> 02:49:46,823 AT STABLE LEVELS BUT WHICH END 4247 02:49:46,823 --> 02:49:48,058 UP HAVING ABSOLUTELY ESSENTIAL 4248 02:49:48,058 --> 02:49:49,926 ROLES IN T CELL DEVELOPMENT. 4249 02:49:49,926 --> 02:49:51,328 ONE THING I'LL SAY ABOUT THIS 4250 02:49:51,328 --> 02:49:53,196 GROUP OF PROGENITOR FACTORS 4251 02:49:53,196 --> 02:49:57,033 RIGHT AT THE START IS THAT THEY 4252 02:49:57,033 --> 02:49:58,668 HAVE TO BE SILENCED AT LEAST BY 4253 02:49:58,668 --> 02:50:00,971 THE TIME OF T LINEAGE COMMITMENT 4254 02:50:00,971 --> 02:50:02,138 OR A LITTLE LATER AS YOU CAN SEE 4255 02:50:02,138 --> 02:50:04,040 FOR ERG AND IF THEY'RE NOT THEY 4256 02:50:04,040 --> 02:50:06,676 CAN ACTUALLY CAUSE A TALL SO THE 4257 02:50:06,676 --> 02:50:08,078 RE REPRESSION MECHANISMS HAVE TO 4258 02:50:08,078 --> 02:50:09,513 BE VERY, VERY GOOD. 4259 02:50:09,513 --> 02:50:10,847 AND THERE ARE ACTUALLY TWO 4260 02:50:10,847 --> 02:50:14,184 MOLECULAR LANDMARKS FOR THE 4261 02:50:14,184 --> 02:50:16,286 COMMITMENT PROCESS INSIDE THE 4262 02:50:16,286 --> 02:50:16,486 CELL. 4263 02:50:16,486 --> 02:50:18,688 ONE OF THEM IS A MAJOR LANDMARK 4264 02:50:18,688 --> 02:50:21,892 IS THE SILENCING OF PO1 ENCODED 4265 02:50:21,892 --> 02:50:24,928 BY THE SPE1 GENE, ITS EXPRESSION 4266 02:50:24,928 --> 02:50:29,699 PRETTY WELL COINCIDES WITH 4267 02:50:29,699 --> 02:50:30,567 COMMITMENT AND WE'VE BEEN STURD 4268 02:50:30,567 --> 02:50:32,469 DID IING THIS FOR QUITE A FEW 4269 02:50:32,469 --> 02:50:33,637 YEARS, WE KNOW QUITE A LOT ABOUT 4270 02:50:33,637 --> 02:50:35,739 THE DRIVERS OF REPRESSION, 4271 02:50:35,739 --> 02:50:40,010 INCLUDE GATA3 AND R YOU NX1 4272 02:50:40,010 --> 02:50:40,677 POTENTIALLY ADDITIONAL 4273 02:50:40,677 --> 02:50:42,078 MECHANISMS AND KNOW SOMETHING 4274 02:50:42,078 --> 02:50:43,513 ABOUT THE SITES INVOLVED BUT 4275 02:50:43,513 --> 02:50:45,348 IT'S STAGE SPECIFIC. 4276 02:50:45,348 --> 02:50:49,519 THE OTHER IS THE ACTIVATION OF 4277 02:50:49,519 --> 02:50:51,922 BCL11B EXPRESSION, WHICH IS 4278 02:50:51,922 --> 02:50:52,722 CRUCIAL TRANSCRIPTION FACTOR 4279 02:50:52,722 --> 02:50:54,124 WHICH IS GOING TO BE EXPRESSED 4280 02:50:54,124 --> 02:50:56,426 BY BASICALLY ALL T CELLS 4281 02:50:56,426 --> 02:50:57,494 AFTERWARDS BUT WHICH TAKES A 4282 02:50:57,494 --> 02:50:59,896 LONG TIME TO TURN ON IN TERMS OF 4283 02:50:59,896 --> 02:51:01,431 ABSOLUTE NUMBERS OF CELL CYCLES 4284 02:51:01,431 --> 02:51:03,300 THAT THE CELLS UNDERGO IN THE 4285 02:51:03,300 --> 02:51:04,668 THYMUS, AND WE'LL TALK A LITTLE 4286 02:51:04,668 --> 02:51:06,469 BIT ABOUT THIS, BUT WE'VE 4287 02:51:06,469 --> 02:51:09,773 STUDIED THIS AS WELL, IT'S 4288 02:51:09,773 --> 02:51:11,641 DRIVEN BY SPEED FORWARD NETWORK 4289 02:51:11,641 --> 02:51:15,378 OF TCF, GATA3, RUNX AND NOTCH 4290 02:51:15,378 --> 02:51:17,013 SIGNALS, ALL FOUR ARE NEEDED, 4291 02:51:17,013 --> 02:51:19,516 IT'S ALSO DEPENDENT ON A SLOW 4292 02:51:19,516 --> 02:51:23,653 CHROMATIN OPENING STEP WHICH IS 4293 02:51:23,653 --> 02:51:24,888 QUITE STOCHASTICALLY REGULATED 4294 02:51:24,888 --> 02:51:26,856 AND WE'VE DISCUSSED THIS IN THE 4295 02:51:26,856 --> 02:51:28,058 PAST. 4296 02:51:28,058 --> 02:51:30,994 NOW, BCL11B IS A LOVELY GENE IN 4297 02:51:30,994 --> 02:51:33,196 THIS PROCESS, AND BECAUSE OF ITS 4298 02:51:33,196 --> 02:51:35,699 SPECIFICITY OF FUNCTION AND THE 4299 02:51:35,699 --> 02:51:37,033 CLARITY OF ITS TIMING, WE WERE 4300 02:51:37,033 --> 02:51:42,172 ABLE TO MAKE A NONDISRUPTIVE 4301 02:51:42,172 --> 02:51:42,939 FLUORESCENT PROTEIN KNOCK ALLELE 4302 02:51:42,939 --> 02:51:45,108 FOR IT, WE MADE A YELLOW AND RED 4303 02:51:45,108 --> 02:51:46,443 VERSION AND THEY'RE BOTH VERY 4304 02:51:46,443 --> 02:51:50,280 USEFUL AND THE POINT IS THAT THE 4305 02:51:50,280 --> 02:51:52,015 BCL11B ONSET OF EXPRESSION 4306 02:51:52,015 --> 02:51:53,850 MARKED BY THESE REPORTERS 4307 02:51:53,850 --> 02:51:55,118 COINCIDES ALMOST PRECISELY AT 4308 02:51:55,118 --> 02:51:56,586 THE SINGLE CELL LEVEL WITH WHEN 4309 02:51:56,586 --> 02:51:58,254 THE CELLS LOSE THEIR ACCESS TO 4310 02:51:58,254 --> 02:51:59,255 THESE ALTERNATIVE LINEAGE FATES 4311 02:51:59,255 --> 02:52:02,659 SO WE CAN USE IT AS A ONE MARKER 4312 02:52:02,659 --> 02:52:04,494 APPROXIMATION FOR THE COMMITMENT 4313 02:52:04,494 --> 02:52:05,962 STATUS AT THE SINGLE CELL LEVEL. 4314 02:52:05,962 --> 02:52:08,031 WHEN YOU DO THAT, YOU CAN 4315 02:52:08,031 --> 02:52:09,633 ACTUALLY LOOK AT SINGLE CELL 4316 02:52:09,633 --> 02:52:14,371 CLONES OF ETP'S, EARLY T CELL 4317 02:52:14,371 --> 02:52:16,806 PRECURSOR DN1'S DEVELOPING AND 4318 02:52:16,806 --> 02:52:19,643 TRACK THEIR PROGRESSION IN 4319 02:52:19,643 --> 02:52:21,277 REALTIME ACROSS DIFFERENT 4320 02:52:21,277 --> 02:52:23,546 MILESTONES OF T CELL 4321 02:52:23,546 --> 02:52:25,749 DEVELOPMENT, MICROSCOPICALLY BY 4322 02:52:25,749 --> 02:52:27,283 LIVE IMAGING OVER A PERIOD OF A 4323 02:52:27,283 --> 02:52:28,051 WEEK OR SO. 4324 02:52:28,051 --> 02:52:29,919 WHAT YOU CAN SEE IS THAT THE 4325 02:52:29,919 --> 02:52:32,555 TIMING IS REALLY, REALLY, REALLY 4326 02:52:32,555 --> 02:52:32,889 NOISY. 4327 02:52:32,889 --> 02:52:34,958 NOW, WITH A STRONG NOTCH SIGNAL, 4328 02:52:34,958 --> 02:52:37,260 THE DESTINATION IS NOT IN DOUBT. 4329 02:52:37,260 --> 02:52:39,996 THE NOTCH SIGNALLING IS GOING TO 4330 02:52:39,996 --> 02:52:41,398 WHAM THOSE CELLS THROUGH INTO 4331 02:52:41,398 --> 02:52:42,532 THE T CELL PROGRAM, BUT THE 4332 02:52:42,532 --> 02:52:45,235 ACTUAL TIMING THROUGH WHICH OF 4333 02:52:45,235 --> 02:52:46,469 THEIR PASSAGE THROUGH THESE 4334 02:52:46,469 --> 02:52:47,771 DIFFERENT MILESTONES IS VERY 4335 02:52:47,771 --> 02:52:49,072 DIFFERENT CLONE TO CLONE AND 4336 02:52:49,072 --> 02:52:53,009 EVEN WITHIN A CLONE. 4337 02:52:53,009 --> 02:52:55,111 SO WE HAVE DIFFERENCES OF UP TO 4338 02:52:55,111 --> 02:52:56,746 FOUR DAYS IN THE LENGTH OF TIME 4339 02:52:56,746 --> 02:52:58,515 IT TAKES INDIVIDUAL ETP'S TO GO 4340 02:52:58,515 --> 02:53:01,117 TO THE DN2A STAGE MARKED BY THE 4341 02:53:01,117 --> 02:53:03,987 EXPRESSION ON THE SURFACE OF 4342 02:53:03,987 --> 02:53:06,156 CD25 BUT IT TAKES AN ADDITIONAL 4343 02:53:06,156 --> 02:53:07,257 ANYWHERE BETWEEN ZERO AND FOUR 4344 02:53:07,257 --> 02:53:10,660 DAYS FOR THOSE CELLS TO TURN ON 4345 02:53:10,660 --> 02:53:11,628 BCL11B AND THAT'S WITHIN THE 4346 02:53:11,628 --> 02:53:14,264 SAME THING, WHICH SAYS THAT IT'S 4347 02:53:14,264 --> 02:53:15,598 NOT JUST DIDN'T KNOW THE 4348 02:53:15,598 --> 02:53:16,933 STARTING POINT IN THE PIPELINE, 4349 02:53:16,933 --> 02:53:19,402 IT MEANS THAT THE PIPELINE 4350 02:53:19,402 --> 02:53:21,171 KINETICS THEMSELVES ARE 4351 02:53:21,171 --> 02:53:22,072 INTRINSICALLY STOCHASTIC. 4352 02:53:22,072 --> 02:53:24,941 WHAT IS ACTUALLY GOING ON? 4353 02:53:24,941 --> 02:53:27,177 NOW, I MADE AN OFF HAND COMMENT 4354 02:53:27,177 --> 02:53:28,311 ABOUT SOMETHING THAT I'VE TALKED 4355 02:53:28,311 --> 02:53:30,280 ABOUT IN THE PAST, BUT NOT 4356 02:53:30,280 --> 02:53:32,348 TODAY, ABOUT THE CHROMATIN 4357 02:53:32,348 --> 02:53:35,752 CONSTRAINTS ON THE BCL11B LOW 4358 02:53:35,752 --> 02:53:36,619 LOCUS AND WE KNOW THAT THE 4359 02:53:36,619 --> 02:53:38,722 OPENING OF THAT CHROMATIN IS 4360 02:53:38,722 --> 02:53:38,922 NOISY. 4361 02:53:38,922 --> 02:53:39,789 SO IT'S POSSIBLE THAT THAT'S 4362 02:53:39,789 --> 02:53:40,690 ACTUALLY ONE OF THE MAJOR 4363 02:53:40,690 --> 02:53:42,192 CONTRIBUTORS TO THE NOISE IN OUR 4364 02:53:42,192 --> 02:53:44,828 ABILITY TO DETECT THE BCL11B 4365 02:53:44,828 --> 02:53:46,596 BEING TURNED ON, SURPRISE, 4366 02:53:46,596 --> 02:53:46,896 SURPRISE. 4367 02:53:46,896 --> 02:53:48,164 BUT THERE COULD BE SOMETHING 4368 02:53:48,164 --> 02:53:50,100 ELSE AS WELL, AND WITH THE 4369 02:53:50,100 --> 02:53:57,874 ADVENT OF SINGLE CELL RNA SEQ -- 4370 02:53:57,874 --> 02:54:01,244 IMAGING ALSO IN THE TEMPORALLY 4371 02:54:01,244 --> 02:54:02,345 STOCHASTIC WAY OR IF IT'S REALLY 4372 02:54:02,345 --> 02:54:03,780 JUST A ONE MARKER ISSUE. 4373 02:54:03,780 --> 02:54:06,182 AND THE ALTERNATIVE TO JUST THIS 4374 02:54:06,182 --> 02:54:08,818 SINGLE GENE EPIGENETIC BARRIER 4375 02:54:08,818 --> 02:54:10,353 IS THAT IN FACT THERE'S 4376 02:54:10,353 --> 02:54:12,522 SOMETHING ABOUT THE OVERLAP 4377 02:54:12,522 --> 02:54:14,691 BETWEEN THE EXPRESSION PATTERNS 4378 02:54:14,691 --> 02:54:16,326 OF THE STEM AND PROGENITOR 4379 02:54:16,326 --> 02:54:18,061 TRANSCRIPTION FACTORS AND THE 4380 02:54:18,061 --> 02:54:18,828 ONSET OF THE T CELL 4381 02:54:18,828 --> 02:54:21,030 TRANSCRIPTION FACTORS WITHIN THE 4382 02:54:21,030 --> 02:54:22,465 SAME CELLS WHICH WE'VE ACTUALLY 4383 02:54:22,465 --> 02:54:25,635 BEEN ABLE TO VALIDATE ON 4384 02:54:25,635 --> 02:54:26,803 MULTIPLEX SINGLE MOLECULE FISH. 4385 02:54:26,803 --> 02:54:28,371 WE KNOW THEY'RE OVERLAPPING IN 4386 02:54:28,371 --> 02:54:28,671 TIME. 4387 02:54:28,671 --> 02:54:30,240 THE QUESTION IS, HOW DO THEY 4388 02:54:30,240 --> 02:54:32,308 INTERACT WITH EACH OTHER IN THE 4389 02:54:32,308 --> 02:54:33,843 SAME CELLS? 4390 02:54:33,843 --> 02:54:35,478 AND ONE WAY TO LOOK AT IT IS 4391 02:54:35,478 --> 02:54:37,247 EXACTLY BY ADOPTING THE APPROACH 4392 02:54:37,247 --> 02:54:39,983 THAT ROXANE POINTED OUT TO YOU 4393 02:54:39,983 --> 02:54:41,518 ALREADY, WHERE YOU CAN ACTUALLY 4394 02:54:41,518 --> 02:54:43,286 DELETE SOME OF THESE REGULATORS 4395 02:54:43,286 --> 02:54:46,156 AND SEE WHAT EFFECT THAT HAS ON 4396 02:54:46,156 --> 02:54:48,792 THE PROGRESSION KINETICS OF T 4397 02:54:48,792 --> 02:54:49,993 CELL DEVELOPMENT. 4398 02:54:49,993 --> 02:54:53,696 SO FOR EXAMPLE H ONE OF THE 4399 02:54:53,696 --> 02:54:55,198 FIRST STEM AND PROGENITOR GENES 4400 02:54:55,198 --> 02:54:58,401 TO TURN OFF AS THE CELLS COME 4401 02:54:58,401 --> 02:55:01,104 UNDER THE INFLUENCE OF THE NOTCH 4402 02:55:01,104 --> 02:55:02,438 SIGNALLING FIELD IN THE THYMUS 4403 02:55:02,438 --> 02:55:05,408 IS LMO2, IT'S AN INCREDIBLY 4404 02:55:05,408 --> 02:55:07,677 IMPORTANT STEM AND PROGENITOR 4405 02:55:07,677 --> 02:55:12,182 CELL TRANSCRIPTION FACTOR. 4406 02:55:12,182 --> 02:55:14,584 FLING AM LI HAS DONE VERY GOOD 4407 02:55:14,584 --> 02:55:20,456 WORK ON THE LMO ONE COMPLEX 4408 02:55:20,456 --> 02:55:21,991 INSIDE GOMENT HEE MATT TOPO 4409 02:55:21,991 --> 02:55:25,495 EASES, SHOULD BE AN OLD FRIEND. 4410 02:55:25,495 --> 02:55:27,063 LM 12R-8 GOES OFF SO QUICKLY YOU 4411 02:55:27,063 --> 02:55:28,264 MIGHT THINK IT'S ONLY 4412 02:55:28,264 --> 02:55:30,567 REPRESENTING THE PRESENCE OF 4413 02:55:30,567 --> 02:55:32,068 CONTAMINANT CELL IN THE THYMUS 4414 02:55:32,068 --> 02:55:32,969 EARLY T CELL POPULATIONS BUT 4415 02:55:32,969 --> 02:55:34,304 THAT'S NOT TRUE BECAUSE IN FACT 4416 02:55:34,304 --> 02:55:36,840 IF YOU KNOCK OUT LMO2 THE CELLS 4417 02:55:36,840 --> 02:55:38,608 SHOOT FORWARD FASTER, AND THIS 4418 02:55:38,608 --> 02:55:40,510 IS AN EXPERIMENT BY A NEW GRAD 4419 02:55:40,510 --> 02:55:43,346 STUDENT IN THE LAB, SAMANTHA 4420 02:55:43,346 --> 02:55:47,083 CHANG USING POLYVINYL ALCOHOL 4421 02:55:47,083 --> 02:55:48,484 CULTURE SYSTEMS, SERUM FREE 4422 02:55:48,484 --> 02:55:50,119 CULTURE SYSTEM TO EXPAND THE 4423 02:55:50,119 --> 02:55:51,921 PROGENITOR SO WE CAN GET A GREAT 4424 02:55:51,921 --> 02:55:54,157 BIG WAVE OF SYNCHRONIZED 4425 02:55:54,157 --> 02:55:56,593 PROGENITORS AND THEN DOING THE 4426 02:55:56,593 --> 02:55:57,994 CAS9 DELETIONS ON THEM AND 4427 02:55:57,994 --> 02:55:59,662 TESTING THEIR SPEED OF 4428 02:55:59,662 --> 02:56:00,964 DIFFERENTIATION OVER THREE, 4429 02:56:00,964 --> 02:56:02,165 FIVE, SEVEN DAYS, AND YOU CAN 4430 02:56:02,165 --> 02:56:04,801 SEE THAT WHEN THE EMPTY VECTOR 4431 02:56:04,801 --> 02:56:08,404 CONTROLS ARE BASICALLY STILL IN 4432 02:56:08,404 --> 02:56:10,840 DN1 ETP STAGE, THE LMO2 4433 02:56:10,840 --> 02:56:12,141 KNOCKOUTS SPURT FORWARD, SPURT 4434 02:56:12,141 --> 02:56:13,376 FORWARD AND THEN ACTUALLY 4435 02:56:13,376 --> 02:56:16,179 ALREADY GOING INTO DN3 WHILE THE 4436 02:56:16,179 --> 02:56:19,682 CONTROLS ARE STILL IN DN1. 4437 02:56:19,682 --> 02:56:21,084 LMO2 IS ONLY ONE OF THIS GROUP 4438 02:56:21,084 --> 02:56:21,918 OF FACTORS. 4439 02:56:21,918 --> 02:56:24,053 ANOTHER ONE THAT TURNS OFF AT 4440 02:56:24,053 --> 02:56:31,728 ABOUT THE TIME OF COMMITMENT IS 4441 02:56:31,728 --> 02:56:35,265 HHEX DESCRENT A HUGE IT -- HHEX 4442 02:56:35,265 --> 02:56:36,633 DELETION DOESN'T ARE A HUGE 4443 02:56:36,633 --> 02:56:39,502 EFFECT ON DN2 TO DN3 PROGRESSION 4444 02:56:39,502 --> 02:56:41,704 JUST USING SURFACE MARKERS, IF 4445 02:56:41,704 --> 02:56:47,043 YOU ACTUALLY MONITOR ENDOGENOUS 4446 02:56:47,043 --> 02:56:48,211 BCL11B EXPRESSION, EXPRESSION TO 4447 02:56:48,211 --> 02:56:49,178 COMMITMENT OF THESE CELLS YOU 4448 02:56:49,178 --> 02:56:52,282 CAN SEE THERE'S ACTUALLY QUITE 4449 02:56:52,282 --> 02:56:53,917 ROBUST AND HIGHLY REPRODUCES I 4450 02:56:53,917 --> 02:56:56,986 BELIEVE UPREGULATION OF BCL11B 4451 02:56:56,986 --> 02:56:58,521 AND AT A MUCH EARLIER TIME THAN 4452 02:56:58,521 --> 02:56:59,956 IT WOULD NORMALLY GO UP IF YOU 4453 02:56:59,956 --> 02:57:03,259 TAKE AWAY HHEX SO WE THINK HHEX 4454 02:57:03,259 --> 02:57:06,963 IS ACTUALLY ACTING AS A RECEPTOR 4455 02:57:06,963 --> 02:57:10,066 REPRESSOR OF BCL1 <!0> B IN 4456 02:57:10,066 --> 02:57:10,767 ADDITION TO NEW YORK STOCK 4457 02:57:10,767 --> 02:57:11,801 EXCHANGE CLOSE CHROMATIN AND 4458 02:57:11,801 --> 02:57:15,271 THIS IS HOW MUCH DIFFERENT THE 4459 02:57:15,271 --> 02:57:16,839 BCL11B GEOMETRIC MEAN 4460 02:57:16,839 --> 02:57:19,475 FLUORESCENCE INTENSITIES IS 4461 02:57:19,475 --> 02:57:20,810 BETWEEN THE KNOCK-OUT IN RED AND 4462 02:57:20,810 --> 02:57:23,746 CONTROL IN GRAY HERE DONE BY A 4463 02:57:23,746 --> 02:57:28,518 POST-DOC HERE JIHYUN IRIZARRY. 4464 02:57:28,518 --> 02:57:29,519 HEAVY HITTERS IN THIS SYSTEM, 4465 02:57:29,519 --> 02:57:30,787 THEY WI THIS THE OPPOSITION 4466 02:57:30,787 --> 02:57:32,889 BETWEEN THE ONSET OF THE T CELL 4467 02:57:32,889 --> 02:57:35,625 FACTORS LIKE TCF7 AND GATA3 ON 4468 02:57:35,625 --> 02:57:38,494 THE ONE HAND AND THE MOST 4469 02:57:38,494 --> 02:57:40,029 DOMINANT PROGENITOR FACTOR WHICH 4470 02:57:40,029 --> 02:57:42,432 IS SPE1 IN OUR HANDS, AND I SAY 4471 02:57:42,432 --> 02:57:44,867 MOST DOMINANT BECAUSE SPE1 IS 4472 02:57:44,867 --> 02:57:47,570 REALLY EXPRESSED AND IT'S TBOWND 4473 02:57:47,570 --> 02:57:50,006 THE DNA, SOMETHING OVER 30,000 4474 02:57:50,006 --> 02:57:52,642 SITES IN THE EARLY -- AND IT'S 4475 02:57:52,642 --> 02:57:54,677 TBOWND THE DNA, SOMETHING OVER 4476 02:57:54,677 --> 02:57:59,282 30,000 SOWNDERS, IN THE EARLY -- 4477 02:57:59,282 --> 02:58:00,750 CAN 30,000 SITES, IT ACCOUNTS 4478 02:58:00,750 --> 02:58:02,018 FOR A VERY LARGE FRACTION OF THE 4479 02:58:02,018 --> 02:58:03,753 OPEN CHROMATIN SITES AS CROSS 4480 02:58:03,753 --> 02:58:06,489 THE GENOME, YOU COULD HAVE SEEN 4481 02:58:06,489 --> 02:58:07,690 THAT IN SOME OF THE FIGURES THAT 4482 02:58:07,690 --> 02:58:09,325 ROXANE SHOWED AS WELL BECAUSE IT 4483 02:58:09,325 --> 02:58:10,960 HAS THE SAME MOTIF ALSO 4484 02:58:10,960 --> 02:58:13,162 IDENTIFIED AS SPEB AND IT'S 4485 02:58:13,162 --> 02:58:16,132 ACTUALLY BINDING THERE AND IT'S 4486 02:58:16,132 --> 02:58:16,933 IMPORTANT. 4487 02:58:16,933 --> 02:58:19,535 WE'VE SEEN IN REPEATED CASES 4488 02:58:19,535 --> 02:58:21,704 THAT PO1 KNOCK QULOWT ALSO 4489 02:58:21,704 --> 02:58:23,606 APPEARS -- PU1 KNOCK-OUT ALSO 4490 02:58:23,606 --> 02:58:24,807 MAKE THE CELLS APPEAR TO GO 4491 02:58:24,807 --> 02:58:26,542 FORWARD LOOKING AT THIS MORE 4492 02:58:26,542 --> 02:58:28,511 GLOBALLY AS CROSS THE GENOME 4493 02:58:28,511 --> 02:58:30,580 WHEN JO THE PERSON THIS MY LAB 4494 02:58:30,580 --> 02:58:33,449 WHO DID A LOT OF SINGLE CELL SEQ 4495 02:58:33,449 --> 02:58:35,284 ANALYSES BASICALLY DID A SMALL 4496 02:58:35,284 --> 02:58:37,453 SCALE PERTURBED SEQ EXPERIMENT 4497 02:58:37,453 --> 02:58:38,454 IN WHICH SOME OF THE THINGS WE 4498 02:58:38,454 --> 02:58:42,091 WERE PERTURBING WERE EITHER SPE1 4499 02:58:42,091 --> 02:58:45,094 OR TCF7 OR ERG OR SEVERAL OTHER 4500 02:58:45,094 --> 02:58:48,664 TRANSCRIPTION FACTORS, AND THESE 4501 02:58:48,664 --> 02:58:50,199 WERE ALL HASHTAGS AND THEN 4502 02:58:50,199 --> 02:58:50,867 SEQUENCED TOGETHER. 4503 02:58:50,867 --> 02:58:52,802 WHAT YOU'RE SEEING HERE IN GRAY 4504 02:58:52,802 --> 02:58:55,972 IS THE THE MAN FOLD OF ALL THE 4505 02:58:55,972 --> 02:58:58,174 SAMPLES IN THE EXPERIMENT, THE 4506 02:58:58,174 --> 02:58:59,709 DARK PURPLE DOTS ACTUALLY SHOW 4507 02:58:59,709 --> 02:59:01,778 YOU THE CONTROL CELLS WHICH ARE 4508 02:59:01,778 --> 02:59:04,080 SPREAD IN THIS RANGE OF STAGES, 4509 02:59:04,080 --> 02:59:05,381 THAT'S THE ASYNCHRONY I WAS 4510 02:59:05,381 --> 02:59:08,017 TELLING YOU ABOUT, BECAUSE THE 4511 02:59:08,017 --> 02:59:09,419 GENES ALTHOUGH THIS END, THIS 4512 02:59:09,419 --> 02:59:11,721 LOWER RIGHT END ARE ACTUALLY ALL 4513 02:59:11,721 --> 02:59:12,622 PROGENITOR GENE EXPRESSION 4514 02:59:12,622 --> 02:59:15,324 PATTERNS, THE ONES UP AT THIS 4515 02:59:15,324 --> 02:59:18,961 END ARE EARLY T CELL ENDS, THE 4516 02:59:18,961 --> 02:59:20,163 CONTROLS ARE SPREAD ALL ACROSS 4517 02:59:20,163 --> 02:59:20,363 HERE. 4518 02:59:20,363 --> 02:59:23,866 IF YOU KNOCK OUGHT TCF7 ENCODING 4519 02:59:23,866 --> 02:59:25,401 TCF1 THE CELLS DON'T EVEN GET 4520 02:59:25,401 --> 02:59:27,070 PAST THIS ENTRY GATE, THEY 4521 02:59:27,070 --> 02:59:27,703 BASICALLY DON'T LOOK LIKE 4522 02:59:27,703 --> 02:59:28,671 THEY'VE STARTED THE T CELL 4523 02:59:28,671 --> 02:59:29,472 PROGRAM AT ALL. 4524 02:59:29,472 --> 02:59:32,108 THIS IS CONSISTENT WITH MANY 4525 02:59:32,108 --> 02:59:35,611 OTHERS' WORK INCLUDING AZ 4526 02:59:35,611 --> 02:59:37,380 I NASHA'S BUT WITH P IS YOU GET 4527 02:59:37,380 --> 02:59:38,881 THE EXACT OPPOSITE PHENOTYPE, 4528 02:59:38,881 --> 02:59:42,618 KNOCKING OUT PU1 CELLS 4529 02:59:42,618 --> 02:59:44,353 FAST-FORWARD THROUGH THEIR 4530 02:59:44,353 --> 02:59:46,122 TRANSCRIPTOME TO THIS EXTREME 4531 02:59:46,122 --> 02:59:48,191 ADVANCE END AND SEE THAT PU1 IS 4532 02:59:48,191 --> 02:59:50,493 MOVING CELLS ACROSS THE SAME SET 4533 02:59:50,493 --> 02:59:52,328 OF STAGES BUT SIGNIFICANTLY 4534 02:59:52,328 --> 02:59:54,831 FASTER THAN THE CONTROLS. 4535 02:59:54,831 --> 02:59:57,600 SO THAT INFERS, IMPLIES THAT PU1 4536 02:59:57,600 --> 02:59:59,035 IS ACTUALLY NORMALLY A BREAK ON 4537 02:59:59,035 --> 03:00:00,002 THIS PROCESS. 4538 03:00:00,002 --> 03:00:04,273 AND BECAUSE OF THE SINGLE CELL 4539 03:00:04,273 --> 03:00:05,775 TRIPTDOMICS YOU CAN ACTUALLY SEE 4540 03:00:05,775 --> 03:00:07,009 WHICH GENES INDIVIDUALLY ARE UP 4541 03:00:07,009 --> 03:00:08,744 AND DOWN IN RESPONSE TO EACH 4542 03:00:08,744 --> 03:00:09,745 PERTUBATION SO WE CAN PUT 4543 03:00:09,745 --> 03:00:12,381 TOGETHER A LOT OF NODES IN AN 4544 03:00:12,381 --> 03:00:13,683 ACTUAL FUNCTIONALLY DEFINED T 4545 03:00:13,683 --> 03:00:15,585 CELL GENE REGULATORY NETWORK. 4546 03:00:15,585 --> 03:00:17,954 THIS IS NOT A MOTIF CORRELATED 4547 03:00:17,954 --> 03:00:19,355 WITH GENE EXPRESSION. 4548 03:00:19,355 --> 03:00:20,690 THIS IS ACTUALLY THESE ARE THE 4549 03:00:20,690 --> 03:00:22,859 GENES WHICH ACTUALLY CHANGE 4550 03:00:22,859 --> 03:00:24,527 EXPRESSION, BOOM, WHEN YOU TAKE 4551 03:00:24,527 --> 03:00:26,262 AWAY THIS TRANSCRIPTION FACTOR. 4552 03:00:26,262 --> 03:00:27,797 WHAT I WANT YOU TO TAKE AWAY 4553 03:00:27,797 --> 03:00:29,432 FROM THIS IS NOT ANY OF THE 4554 03:00:29,432 --> 03:00:32,535 DETAILS, BUT THESE COLORED 4555 03:00:32,535 --> 03:00:34,070 REGIONS REPRESENT SIMULTANEOUSLY 4556 03:00:34,070 --> 03:00:36,139 OPERATING NETWORKS IN THE SAME 4557 03:00:36,139 --> 03:00:36,405 CELLS. 4558 03:00:36,405 --> 03:00:38,207 THE BLUE NETWORK IS THE ONE 4559 03:00:38,207 --> 03:00:40,643 DRIVEN BY NOTCH DIRECTLY, AND 4560 03:00:40,643 --> 03:00:42,478 THAT'S INCLUDING THE FEED 4561 03:00:42,478 --> 03:00:43,880 FORWARD CIRCUITRY THAT GIVES YOU 4562 03:00:43,880 --> 03:00:46,849 THE T CELL PROGRAM COMING OUT. 4563 03:00:46,849 --> 03:00:50,353 TCF7, GATA3, TCF12 ALSO DRIVING 4564 03:00:50,353 --> 03:00:52,688 TOWARDS T CELL SPECIFICATION. 4565 03:00:52,688 --> 03:00:56,492 IN THIS ORANGE BOX, THIS IS THIS 4566 03:00:56,492 --> 03:00:58,461 ATTEMPTED SELF-MAINTAINING 4567 03:00:58,461 --> 03:00:59,829 PROGENITOR GENE REGULATORY 4568 03:00:59,829 --> 03:01:02,698 NETWORK WITH A LOT OF 4569 03:01:02,698 --> 03:01:03,299 CROSS-REGULATORY INTERACTIONS, 4570 03:01:03,299 --> 03:01:04,967 EVEN BEYOND THE ONES SHOWN HERE 4571 03:01:04,967 --> 03:01:07,436 IN WHICH SPE1 IS A MAJOR DRIVER 4572 03:01:07,436 --> 03:01:09,272 OF THE EXPRESSION OF OTHER 4573 03:01:09,272 --> 03:01:16,179 FACTORS LIKE LMO2, MEF2C AND 4574 03:01:16,179 --> 03:01:17,480 BCL11A TRYING TO HOLD BACK THE 4575 03:01:17,480 --> 03:01:18,748 CELLS FROM DIFFER HE BE YAITDING 4576 03:01:18,748 --> 03:01:20,216 TOO FAST SO YOU REALLY HAVE THE 4577 03:01:20,216 --> 03:01:22,718 CELLS FOR A WHILE ARE BALANCED 4578 03:01:22,718 --> 03:01:24,487 BETWEEN THE ACTIVITY OF TWO 4579 03:01:24,487 --> 03:01:25,721 DIFFERENT OPPOSING NETWORK ON 4580 03:01:25,721 --> 03:01:28,991 THE SAME CELLS WITH A LOT OF 4581 03:01:28,991 --> 03:01:29,759 CROSSES-REPRESSIVE INTERACTIONS 4582 03:01:29,759 --> 03:01:31,360 AT THE SINGLE GENE LEVEL. 4583 03:01:31,360 --> 03:01:33,029 NOW, HERE ARE THESE GENES IN 4584 03:01:33,029 --> 03:01:36,332 HERE, WHICH DON'T SEEM TO MAKE 4585 03:01:36,332 --> 03:01:37,833 SENSE. 4586 03:01:37,833 --> 03:01:38,701 RUNX1 AND 3. 4587 03:01:38,701 --> 03:01:40,603 THE REASON THEY DO PLAY A VERY 4588 03:01:40,603 --> 03:01:42,138 BIG ROLE, AND IF YOU FOLLOW THE 4589 03:01:42,138 --> 03:01:44,140 ARROWS, YOU CAN SEE THAT THEY'RE 4590 03:01:44,140 --> 03:01:45,308 BIASED, THEY'RE ACTUALLY HELPING 4591 03:01:45,308 --> 03:01:47,710 THE T CELL PROGRAM AND THEY'RE 4592 03:01:47,710 --> 03:01:48,978 ANTAGONIZING THE STEM AND 4593 03:01:48,978 --> 03:01:49,579 PROGENITOR PROGRAM. 4594 03:01:49,579 --> 03:01:58,354 WHAT ARE THEY DOING HERE? 4595 03:01:58,354 --> 03:01:59,689 ESSENTIAL A FLAT LEVEL OF 4596 03:01:59,689 --> 03:02:01,624 EXPRESSION ACROSS THESE STAGES. 4597 03:02:01,624 --> 03:02:03,292 THEY'RE NOT INCREASING WITH T 4598 03:02:03,292 --> 03:02:04,260 CELL DEVELOPMENT. 4599 03:02:04,260 --> 03:02:06,028 IN FACT, IF YOU WERE DOING 4600 03:02:06,028 --> 03:02:06,963 FUNCTIONAL GENOMICS AND YOU WERE 4601 03:02:06,963 --> 03:02:08,497 TRYING TO SAY WHICH 4602 03:02:08,497 --> 03:02:10,066 TRANSCRIPTION FACTORS ARE LIKELY 4603 03:02:10,066 --> 03:02:11,634 TO BE BORING IN THIS PROCESS, 4604 03:02:11,634 --> 03:02:13,336 YOU WOULD LOOK AT THIS FLAT 4605 03:02:13,336 --> 03:02:15,838 EXPRESSION OF RUNX AND SAY IT'S 4606 03:02:15,838 --> 03:02:17,506 OBVIOUSLY NOT DOING ANYTHING 4607 03:02:17,506 --> 03:02:17,807 INTERESTING. 4608 03:02:17,807 --> 03:02:19,775 BUT IN FACT WHAT IT IS DOING IN 4609 03:02:19,775 --> 03:02:22,845 TERMS OF DIRECT REGULATORY 4610 03:02:22,845 --> 03:02:24,780 EFFECT IS IT'S A MAJOR REPRESSOR 4611 03:02:24,780 --> 03:02:26,549 OF THE STEM AND PROGENITOR 4612 03:02:26,549 --> 03:02:28,217 PROGRAM GENES, SEVERAL OF THEM, 4613 03:02:28,217 --> 03:02:31,621 AND IT'S A MAJOR POSITIVE 4614 03:02:31,621 --> 03:02:34,023 REGULATOR OF EACH OF THESE THREE 4615 03:02:34,023 --> 03:02:35,424 MAJOR T CELL PROMOTING 4616 03:02:35,424 --> 03:02:36,959 TRANSCRIPTION FACTORS. 4617 03:02:36,959 --> 03:02:39,528 SO WHAT IS IT DOING THEN P IN 4618 03:02:39,528 --> 03:02:42,031 THE EARLY ETP STAGE WHEN THE 4619 03:02:42,031 --> 03:02:45,167 MAIN BUSINESS OF THIS IS TO 4620 03:02:45,167 --> 03:02:46,936 EXPRESS THESE PROGENITOR GENES? 4621 03:02:46,936 --> 03:02:48,938 WELL, WHAT HAPPENS AND SOME OF 4622 03:02:48,938 --> 03:02:50,406 YOU HAVE HEARD ME TALK ABOUT 4623 03:02:50,406 --> 03:02:52,842 THIS BEFORE, IS THAT RUNX 4624 03:02:52,842 --> 03:02:56,112 BINDING IS VERY, VERY STAGE 4625 03:02:56,112 --> 03:02:56,379 SPECIFIC. 4626 03:02:56,379 --> 03:02:58,214 AND IN THE CELLS THAT ARE STILL 4627 03:02:58,214 --> 03:03:01,584 EXPRESSING A LOT OF PU1, THE 4628 03:03:01,584 --> 03:03:03,886 RUNX IS BINDING TO DIFFERENT 4629 03:03:03,886 --> 03:03:05,855 GENOMIC SITES THAN IT IS AS THE 4630 03:03:05,855 --> 03:03:07,923 CELLS UNDERGO COMMITMENT HERE. 4631 03:03:07,923 --> 03:03:09,825 AND YOU CAN SEE IT'S REALLY, 4632 03:03:09,825 --> 03:03:11,427 THIS REALLY INVOLVES LARGE 4633 03:03:11,427 --> 03:03:12,795 FRACTIONS OF ALL OF THE SITES 4634 03:03:12,795 --> 03:03:15,798 THAT RUNX BINDS IN THE GENOME. 4635 03:03:15,798 --> 03:03:17,266 AND IF YOU'RE INTERESTED AND YOU 4636 03:03:17,266 --> 03:03:19,001 WANT TO LOOK, THIS IS RECENTLY 4637 03:03:19,001 --> 03:03:20,303 PUBLISHED WORK, SO YOU CAN 4638 03:03:20,303 --> 03:03:23,506 ACTUALLY LOOK UP, IT'S NOT JUST 4639 03:03:23,506 --> 03:03:24,940 FOLLOWING TAC ACCESSIBILITY. 4640 03:03:24,940 --> 03:03:27,009 IF ANYTHING SH IT'S ANTICIPATING 4641 03:03:27,009 --> 03:03:28,077 IT. 4642 03:03:28,077 --> 03:03:30,279 THE ATTACK ACCESSIBILITY IS 4643 03:03:30,279 --> 03:03:34,717 GENERALLY ENRICHED AT RUNX SITES 4644 03:03:34,717 --> 03:03:36,686 AND VICE VERSA BUT RUNX CAN GET 4645 03:03:36,686 --> 03:03:39,188 INTO SITES THAT HAVE REMAINED 4646 03:03:39,188 --> 03:03:41,624 TOLLINGLY IN TAC INACCESSIBLE OR 4647 03:03:41,624 --> 03:03:43,059 IT CAN DELAY GETTING INTO SITES 4648 03:03:43,059 --> 03:03:44,460 WHICH ARE JUST TOTALLY OPEN AND 4649 03:03:44,460 --> 03:03:45,995 JUST WAITING, IF THE TIME ISN'T 4650 03:03:45,995 --> 03:03:46,228 RIGHT. 4651 03:03:46,228 --> 03:03:49,065 SO SOMETHING ELSE IS ACTUALLY 4652 03:03:49,065 --> 03:03:51,467 CONTROLLING THE CHANGE IN RUNX 4653 03:03:51,467 --> 03:03:53,436 OCCUPANCY WHICH CORRELATES VERY, 4654 03:03:53,436 --> 03:03:55,971 VERY WELL WITH A SWITCHING OF 4655 03:03:55,971 --> 03:03:58,074 WHOLESALE OF RUNX FUNCTIONAL 4656 03:03:58,074 --> 03:04:00,242 EFFECTS ON THE CELLS. 4657 03:04:00,242 --> 03:04:02,745 SO WHY DO RUNX TBAK TORS BIND 4658 03:04:02,745 --> 03:04:05,281 DIFFERENT SITE AT DIFFERENT 4659 03:04:05,281 --> 03:04:05,548 STAINDLE? 4660 03:04:05,548 --> 03:04:08,551 -- RUNX FACTORS BIND DIFFERENT 4661 03:04:08,551 --> 03:04:10,753 SITES AT DIFFERENT STAGES? 4662 03:04:10,753 --> 03:04:13,489 RMS ACTUALLY IT HAS A SET OF 4663 03:04:13,489 --> 03:04:15,224 ALTERNATIVE INTERACTION PARTNERS 4664 03:04:15,224 --> 03:04:16,092 WHICH ARE IN COMPETITION WITH 4665 03:04:16,092 --> 03:04:17,326 EACH OTHER. 4666 03:04:17,326 --> 03:04:18,961 WE HAD PUBLISHED SOME YEARS AGO 4667 03:04:18,961 --> 03:04:21,997 THAT IF YOU TAKE A CELL, A T 4668 03:04:21,997 --> 03:04:23,332 LINEAGE CELL THAT'S ALREADY 4669 03:04:23,332 --> 03:04:29,772 TURNED OFF PU1, THAT PU1 CAN BE 4670 03:04:29,772 --> 03:04:31,841 ADDED BACK AGAIN AND IT WILL 4671 03:04:31,841 --> 03:04:33,376 TALL -- AND IT WILL ACTUALLY 4672 03:04:33,376 --> 03:04:36,011 CAUSE THE CELL TO LOSE ITS T 4673 03:04:36,011 --> 03:04:37,213 LINEAGE COMMITTED STATUS AND TRY 4674 03:04:37,213 --> 03:04:38,581 TO MAKE MYELOID CELLS AGAIN AND 4675 03:04:38,581 --> 03:04:40,616 THE WAY IT DOES THIS IN PART IS 4676 03:04:40,616 --> 03:04:43,919 BY SHUTTING OFF RUNX'S ABILITY 4677 03:04:43,919 --> 03:04:45,888 TO SERVE APPEARS THE T CELL 4678 03:04:45,888 --> 03:04:47,323 GENES THAT IT WAS SERVICING 4679 03:04:47,323 --> 03:04:47,857 BEFORE. 4680 03:04:47,857 --> 03:04:48,290 HOW? 4681 03:04:48,290 --> 03:04:51,160 BECAUSE THE PU1 THAT YOU ADD 4682 03:04:51,160 --> 03:04:53,129 BINDS TO SITES THAT WERE CLOSED 4683 03:04:53,129 --> 03:04:56,298 BEFORE HAD, PU1'S KNOWN PIONEER 4684 03:04:56,298 --> 03:04:57,833 FAX TORQUES HERE IT IS 4685 03:04:57,833 --> 03:04:59,468 PIONEERING, AND WHAT IT DOES IS 4686 03:04:59,468 --> 03:05:01,637 IT PULLS RUNX ALONG FOR THE 4687 03:05:01,637 --> 03:05:01,837 RIDE. 4688 03:05:01,837 --> 03:05:04,273 SO RUNX WAS NOT BINDING TO THESE 4689 03:05:04,273 --> 03:05:07,343 SITES BEFORE, YOU ADD PU1, RUNX 4690 03:05:07,343 --> 03:05:07,910 GOES THERE. 4691 03:05:07,910 --> 03:05:08,978 BUT THE INTERESTING THING TO US 4692 03:05:08,978 --> 03:05:11,147 IS THAT RUNX DOES THIS AT THE 4693 03:05:11,147 --> 03:05:14,350 EXPENSE OF ITS BINDING TO A 4694 03:05:14,350 --> 03:05:18,721 LARGE FRACTION OF ITS PREVIOUS 4695 03:05:18,721 --> 03:05:19,889 OCCUPANTS, WHAT DOES THIS 4696 03:05:19,889 --> 03:05:20,890 ACTUALLY MEAN? 4697 03:05:20,890 --> 03:05:23,092 COULD PU1 BE PUSHING IT OUT OF 4698 03:05:23,092 --> 03:05:23,726 THESE SITES? 4699 03:05:23,726 --> 03:05:25,361 WELL, NO, IT'S NOT PUSHING 4700 03:05:25,361 --> 03:05:28,664 BECAUSE PU1 ISN'T GOING THERE. 4701 03:05:28,664 --> 03:05:30,633 PU1 IS PULLING IS BUT IT'S NOT 4702 03:05:30,633 --> 03:05:31,734 PUSH IT GO OUT. 4703 03:05:31,734 --> 03:05:33,569 SO DOES THIS MEAN THERE'S NOT 4704 03:05:33,569 --> 03:05:35,237 ENOUGH RU NX TO GO AROUND? 4705 03:05:35,237 --> 03:05:36,639 ONE WAY TO ASK THAT QUESTION 4706 03:05:36,639 --> 03:05:38,274 MORE QUANTITATIVELY IS JUST TO 4707 03:05:38,274 --> 03:05:40,609 ADD MORE RUNX TO THE AND SEE HOW 4708 03:05:40,609 --> 03:05:41,811 THAT CHANGES THE RESULT. 4709 03:05:41,811 --> 03:05:44,847 WHEN YOU DO THAT, YOU GET A VERY 4710 03:05:44,847 --> 03:05:47,016 SATISFYING-LOOKING RESULT IN 4711 03:05:47,016 --> 03:05:51,320 THIS ARTIFICIAL T CELL LINE 4712 03:05:51,320 --> 03:06:00,429 SYSTEM WHERE YOU ADD PU1 NOW, IT 4713 03:06:00,429 --> 03:06:02,364 STILL GOES TO THE PU1 SITES THE 4714 03:06:02,364 --> 03:06:04,533 SAME WAY, BUT NOW IT NO LONGER 4715 03:06:04,533 --> 03:06:07,236 IS DEPLETED FROM THE PREVIOUS 4716 03:06:07,236 --> 03:06:07,470 SITES. 4717 03:06:07,470 --> 03:06:09,805 SO THE QUESTION IS, YOU KNOW, 4718 03:06:09,805 --> 03:06:10,906 AND WE KNOW SOMETHING ABOUT THE 4719 03:06:10,906 --> 03:06:12,541 PHYSICAL NATURE OF THESE SITES, 4720 03:06:12,541 --> 03:06:15,678 WE'VE BEEN VERY -- WE KNOW WHAT 4721 03:06:15,678 --> 03:06:16,612 PART IN OTHER WORDS ARE THERE 4722 03:06:16,612 --> 03:06:18,714 WHAT, MOTIFS ARE THERE, HOW HIGH 4723 03:06:18,714 --> 03:06:21,250 AFFINITY THEY ARE AND SO FORTH. 4724 03:06:21,250 --> 03:06:22,751 BUT THIS IS AN ARTIFICIAL 4725 03:06:22,751 --> 03:06:23,652 SYSTEM, AND THE QUESTION IS, 4726 03:06:23,652 --> 03:06:26,121 DOES THIS REALLY HAVE ANY IN 4727 03:06:26,121 --> 03:06:27,022 VIVO RELEVANCE? 4728 03:06:27,022 --> 03:06:29,959 JUST TO CLARIFY, WE'RE PROPOSING 4729 03:06:29,959 --> 03:06:32,194 THAT IN THE NATURAL STATES WHEN 4730 03:06:32,194 --> 03:06:34,897 THERE'S A LOT OF PU1 AROUND, THE 4731 03:06:34,897 --> 03:06:37,266 PU1 IS NOT JUST CO-BINDING WITH 4732 03:06:37,266 --> 03:06:40,135 THE RUNX BUT THAT IT'S ACTUALLY 4733 03:06:40,135 --> 03:06:42,338 SEQUESTERING THE RUNX AT THOSE 4734 03:06:42,338 --> 03:06:44,740 SITES AND WHEN THE PU1 GOES DOWN 4735 03:06:44,740 --> 03:06:46,609 THAT NOW ALLOWS THE RUNX TO GO 4736 03:06:46,609 --> 03:06:47,142 SOMEWHERE ELSE. 4737 03:06:47,142 --> 03:06:48,477 IF THAT WERE TRUE, THAT WOULD 4738 03:06:48,477 --> 03:06:49,945 MEAN THERE SHOULD BE A 4739 03:06:49,945 --> 03:06:50,679 DEVELOPMENTAL CONSEQUENCE TO 4740 03:06:50,679 --> 03:06:53,649 ADDING MORE RUNX, NOT TOO MUCH 4741 03:06:53,649 --> 03:06:54,283 RUNX BECAUSE TOO MUCH OF 4742 03:06:54,283 --> 03:06:55,818 ANYTHING IS BAD, BUT WHAT ABOUT 4743 03:06:55,818 --> 03:06:56,819 JUST A LITTLE MORE? 4744 03:06:56,819 --> 03:06:59,121 AND WE WERE VERY FORTUNATE TO BE 4745 03:06:59,121 --> 03:07:01,724 ABLE TO HAVE VECTORS WHICH 4746 03:07:01,724 --> 03:07:04,693 RESPONDED TO THEIR OWN USE 4747 03:07:04,693 --> 03:07:06,996 SUPPRESSED RUNX IN A VERY, VERY 4748 03:07:06,996 --> 03:07:08,397 MODERATE MEASURED WAY. SO WE 4749 03:07:08,397 --> 03:07:09,632 ESSENTIALLY JUST DOUBLED THE 4750 03:07:09,632 --> 03:07:11,800 AMOUNT OF RUNX IN THE CELL USING 4751 03:07:11,800 --> 03:07:14,303 THESE VECTORS, WHICH IS NOT 4752 03:07:14,303 --> 03:07:16,605 REALLY VERY MUCH OF AN 4753 03:07:16,605 --> 03:07:16,972 OVEREXPRESSION. 4754 03:07:16,972 --> 03:07:19,008 SO WE COULD THEN TEST THIS AT 4755 03:07:19,008 --> 03:07:23,078 THE PROTEIN LEVEL, VERIFY IT AND 4756 03:07:23,078 --> 03:07:24,947 LOOK AT THE CROSS-REGULATION 4757 03:07:24,947 --> 03:07:26,982 WITH OTHER RUNX FAMILY MEMBERS 4758 03:07:26,982 --> 03:07:28,350 AND WE COULD SEE THAT THEY 4759 03:07:28,350 --> 03:07:29,618 ACTUALLY WENT DOWN WHEN YOU 4760 03:07:29,618 --> 03:07:31,153 OVEREXPRESSED RUNX ONE SO YOU 4761 03:07:31,153 --> 03:07:33,122 END UP WITH NO MORE THAN ABOUT A 4762 03:07:33,122 --> 03:07:34,189 TWOFOLD INCREASE TOTAL. 4763 03:07:34,189 --> 03:07:35,491 THEN YOU COULD ASK, WHAT THOOPS 4764 03:07:35,491 --> 03:07:36,258 THE CELLS? 4765 03:07:36,258 --> 03:07:38,260 WE CAN LOOK IN TERMS OF SINGLY 4766 03:07:38,260 --> 03:07:40,129 CELL RNA SEQ, WE CAN ALSO LOOK 4767 03:07:40,129 --> 03:07:43,966 IN TERMS LONGER TERM T CELL 4768 03:07:43,966 --> 03:07:44,266 DEVELOPMENT. 4769 03:07:44,266 --> 03:07:46,235 THIS IS THE SCHEMA OF HOW WE 4770 03:07:46,235 --> 03:07:49,405 STUDY THIS, THIS WAS A MARVELOUS 4771 03:07:49,405 --> 03:07:55,678 POST-DOC WHO SOME OF YOU KNOW, 4772 03:07:55,678 --> 03:07:57,713 BOYOUNG SHIN WHO SET THIS UP AS 4773 03:07:57,713 --> 03:07:59,114 BOTH A GAIN EXPLOS OF FUNCTION 4774 03:07:59,114 --> 03:08:01,483 OF R YOU NX IN PARALLEL SETS OF 4775 03:08:01,483 --> 03:08:02,685 CELLS AT TWO DIFFERENT TIME 4776 03:08:02,685 --> 03:08:04,553 POINTS FOR EACH, SO TWO DAYS OR 4777 03:08:04,553 --> 03:08:07,189 FOUR DAYS POST ADDITION OF EXTRA 4778 03:08:07,189 --> 03:08:09,124 RUNX, THREE DAYS OR SIX DAYS 4779 03:08:09,124 --> 03:08:12,628 POST DELETION OF BOTH RUNX1 AND 4780 03:08:12,628 --> 03:08:14,730 3 TOGETHER IN THE SAME COHORTS 4781 03:08:14,730 --> 03:08:17,766 OF INPUT CELLS THAT WILL WE'RE 4782 03:08:17,766 --> 03:08:21,570 GETTING FROM EITHER CAS9 B CELL 4783 03:08:21,570 --> 03:08:29,011 2 TRANSGENIC MICE OR JUST B CELL 4784 03:08:29,011 --> 03:08:30,279 2 MICE AND ASKING WHAT THOOPS 4785 03:08:30,279 --> 03:08:31,246 THEIR DEVELOPMENT AND IF YOU 4786 03:08:31,246 --> 03:08:33,682 LOOK AT THE SINGLE CELL RNA SEQ 4787 03:08:33,682 --> 03:08:35,084 EFFECTS OF THESE PERTUBATIONS 4788 03:08:35,084 --> 03:08:37,653 HERE WHAT I'M SHOWING YOU IS THE 4789 03:08:37,653 --> 03:08:39,355 MANIFOLD OF ALL OF THE SAMPLES, 4790 03:08:39,355 --> 03:08:40,990 AGAIN, ALL OF THE TIME POINTS, 4791 03:08:40,990 --> 03:08:42,625 ALL OF THE GENOTYPES, ONE OF THE 4792 03:08:42,625 --> 03:08:43,926 VERY NICE THINGS ABOUT IT IS 4793 03:08:43,926 --> 03:08:47,229 THAT IF YOU MAP, YOU MAP ONE IS 4794 03:08:47,229 --> 03:08:48,864 ALMOST ALWAYS CELL CYCLE SO 4795 03:08:48,864 --> 03:08:51,900 THAT'S REANLTD AN ISSUE HERE. 4796 03:08:51,900 --> 03:08:54,470 BUT U MAP TWO AND U MAP THREE 4797 03:08:54,470 --> 03:08:56,305 ARE INCREDIBLY INFORMATIVE HERE. 4798 03:08:56,305 --> 03:08:58,207 U MAP 2 IS ALMOST PERFECT 4799 03:08:58,207 --> 03:08:59,441 REPRESENTATION OF DEVELOPMENTAL 4800 03:08:59,441 --> 03:09:01,076 PROGRESSION ALONG THE T CELL 4801 03:09:01,076 --> 03:09:04,313 PATHWAY, AND U MAP 3 4802 03:09:04,313 --> 03:09:05,047 MAGNIFICENTLY ENOUGH TURNS ON 4803 03:09:05,047 --> 03:09:07,316 THE TO BE A REFLECTION OF THE 4804 03:09:07,316 --> 03:09:09,084 RUNX GENOTYPE. 4805 03:09:09,084 --> 03:09:11,587 SO WE WERE ABLE TO SEPARATE RUNX 4806 03:09:11,587 --> 03:09:12,588 KNOCK-OUT FROM THE CONTROLS FROM 4807 03:09:12,588 --> 03:09:15,057 THE RUNX OVEREXPRESSION, AND YOU 4808 03:09:15,057 --> 03:09:17,493 CAN ACTUALLY SEE THEN VISUALLY 4809 03:09:17,493 --> 03:09:19,461 WHAT I'M GOING TO SHOW YOU HERE 4810 03:09:19,461 --> 03:09:21,697 CALCULATED FROM THE PSEUDOTIME 4811 03:09:21,697 --> 03:09:24,166 USING MONOCAL WHICH IS AT THAT 4812 03:09:24,166 --> 03:09:26,368 EVERY TIME POINT RUNX 4813 03:09:26,368 --> 03:09:27,469 OVEREXPRESSION MOVES THE CELLS 4814 03:09:27,469 --> 03:09:29,438 FASTER THROUGH THE T CELL 4815 03:09:29,438 --> 03:09:29,672 PATHWAY. 4816 03:09:29,672 --> 03:09:32,875 P AT EVERY TIME POINT THE RUNX 4817 03:09:32,875 --> 03:09:34,543 KNOCK-OUT MOVES THEM SLOWER. 4818 03:09:34,543 --> 03:09:37,179 IS THIS ACTUALLY REALLY RELEVANT 4819 03:09:37,179 --> 03:09:39,148 TO T CELL DEVELOPMENT OR IS IT 4820 03:09:39,148 --> 03:09:41,550 REALLY JUST SINGLE CELL RNA SEQ, 4821 03:09:41,550 --> 03:09:44,520 YOU KNOW, LABELING OF 4822 03:09:44,520 --> 03:09:45,821 TRAJECTORIES AND DOES THIS HAVE 4823 03:09:45,821 --> 03:09:47,790 ANYTHING TO DO WITH REAL T CELL 4824 03:09:47,790 --> 03:09:48,090 DEVELOPMENT? 4825 03:09:48,090 --> 03:09:50,859 WE WERE ABLE TO LOOK AT THIS IN 4826 03:09:50,859 --> 03:09:58,434 AN IN VITRO WAY. 4827 03:09:58,434 --> 03:10:00,302 RETROVIRAL VECTORS, EMPTY VECTOR 4828 03:10:00,302 --> 03:10:03,338 ONE COLOR, THE RUNX 4829 03:10:03,338 --> 03:10:04,106 OVEREXPRESSING ANOTHER COLOR 4830 03:10:04,106 --> 03:10:06,408 MIXED TOGETHER IN THE SAME 4831 03:10:06,408 --> 03:10:07,710 ARTIFICIAL ORGANOID CULTURE 4832 03:10:07,710 --> 03:10:09,578 SYSTEM WHICH IS VERY, VERY GOOD 4833 03:10:09,578 --> 03:10:10,913 FOR PUSH BEING THE CELLS PAST 4834 03:10:10,913 --> 03:10:13,315 THAT DM2 ARREST POINT THAT 4835 03:10:13,315 --> 03:10:15,184 ROXANE TOLD YOU ABOUT. SO THEY 4836 03:10:15,184 --> 03:10:17,119 GO ALL THE WAY INTO BETA 4837 03:10:17,119 --> 03:10:17,386 SELECTION. 4838 03:10:17,386 --> 03:10:18,620 WHAT I THINK YOU CAN SEE HERE IS 4839 03:10:18,620 --> 03:10:22,725 WITH THE RED DOTS BEING THE 4840 03:10:22,725 --> 03:10:24,126 OVEREXPRESSING ONES, YOU CAN SEE 4841 03:10:24,126 --> 03:10:25,994 THAT THE RED CELLS, THE 4842 03:10:25,994 --> 03:10:28,397 OVEREXPRESSING CELLS AT EACH 4843 03:10:28,397 --> 03:10:30,833 TIME POINT GET OUT OF THE 4844 03:10:30,833 --> 03:10:32,668 EARLIER STAGE FASTER AND INTO 4845 03:10:32,668 --> 03:10:34,970 THE LATER STAGE FASTER, OUT OF 4846 03:10:34,970 --> 03:10:37,172 THE EARLY STAGE FASTER, INTO THE 4847 03:10:37,172 --> 03:10:38,807 LATER STAGE FASTER ALL THE WAY 4848 03:10:38,807 --> 03:10:40,342 THROUGH BETA SELECTION, CELLS 4849 03:10:40,342 --> 03:10:42,644 RESPOND TO GO THIS RUNX 4850 03:10:42,644 --> 03:10:43,412 OVEREXPRESSION ACCELERATION 4851 03:10:43,412 --> 03:10:44,613 EFFECT ARE CELLS CAPABLE OF 4852 03:10:44,613 --> 03:10:48,450 GOING ON IN THE T CELL PROGRAM. 4853 03:10:48,450 --> 03:10:49,651 I JUST WANT TO SAY SOMETHING 4854 03:10:49,651 --> 03:10:52,020 ABOUT THE GENE EXPRESSION 4855 03:10:52,020 --> 03:10:54,056 DETAILS OF THIS PHENOTYPE 4856 03:10:54,056 --> 03:10:54,923 BECAUSE THEY'RE REALLY 4857 03:10:54,923 --> 03:10:57,226 INTERESTING FOR THE MECHANISM. 4858 03:10:57,226 --> 03:10:59,094 I ALLUDED TO THE FACT THAT ONE 4859 03:10:59,094 --> 03:11:09,505 OF THE THINGS ASILENCES 4860 03:11:10,305 --> 03:11:11,440 EXPRETION -- THAT SIGH LENS 4861 03:11:11,440 --> 03:11:13,175 EXPRESSION IS GATA3. 4862 03:11:13,175 --> 03:11:15,911 IF GATA3 WAS ALREADY THERE WHICH 4863 03:11:15,911 --> 03:11:17,579 IT WAS WERE WE SIMPLE JUST 4864 03:11:17,579 --> 03:11:19,248 TURNING OFF PU1, IN OTHER WORDS, 4865 03:11:19,248 --> 03:11:21,850 WERE WE SIMPLE REPEATING THE PU1 4866 03:11:21,850 --> 03:11:22,584 KNOCK-OUT EXPERIMENT AGAIN IN 4867 03:11:22,584 --> 03:11:23,986 THESE SPHERMTS? 4868 03:11:23,986 --> 03:11:25,587 OR WERE WE DOING SOMETHING NEW 4869 03:11:25,587 --> 03:11:28,423 THAT HAD TO DO WITH RUNX'S OWN 4870 03:11:28,423 --> 03:11:28,657 EFFECT? 4871 03:11:28,657 --> 03:11:30,259 SO TO ANSWER THAT, WHAT WE DID 4872 03:11:30,259 --> 03:11:33,362 WAS WE TOOK THE SINGLE CELL RNA 4873 03:11:33,362 --> 03:11:35,330 SEQ DATA AND WE BASICALLY SAID, 4874 03:11:35,330 --> 03:11:36,665 WE'RE ONLY INTERESTED IN THE 4875 03:11:36,665 --> 03:11:38,634 SLICES OF CELLS THAT ARE STILL 4876 03:11:38,634 --> 03:11:39,735 EXPRESSING PU1. 4877 03:11:39,735 --> 03:11:42,070 SO THESE ARE THE CELLS THAT ALL 4878 03:11:42,070 --> 03:11:44,907 HAVE STILL PU1 RNA OVER A 4879 03:11:44,907 --> 03:11:46,308 CERTAIN THRESHOLD LEVEL, 4880 03:11:46,308 --> 03:11:48,143 COMPARING THE CONTROLS WITH THE 4881 03:11:48,143 --> 03:11:49,211 RUNX OVEREXPRESSING. 4882 03:11:49,211 --> 03:11:50,379 NOW WE'LL LOOK AT THE OTHER 4883 03:11:50,379 --> 03:11:52,447 GENES THAT THOSE CELLS ARE 4884 03:11:52,447 --> 03:11:52,781 EXPRESSING. 4885 03:11:52,781 --> 03:11:54,983 DO THEY ACTIVATE THE T CELL 4886 03:11:54,983 --> 03:11:56,852 PROGRAM BEFORE THEY DOWN 4887 03:11:56,852 --> 03:11:58,587 REGULATE THE STEM AND PROGENITOR 4888 03:11:58,587 --> 03:12:00,422 GENES, OR DO THEY DO THIS STEM 4889 03:12:00,422 --> 03:12:01,957 AND PROGENITOR GENE DOWN 4890 03:12:01,957 --> 03:12:03,192 REGULATION FIRST? 4891 03:12:03,192 --> 03:12:05,394 AND THE RESULTS CAME OUT 4892 03:12:05,394 --> 03:12:07,796 UNBELIEVABLY CLEARLY, WHICH IS 4893 03:12:07,796 --> 03:12:09,531 THAT IF YOU PICK ON THE CELLS 4894 03:12:09,531 --> 03:12:11,500 THAT INDUSTRIAL PU1 THEY HAVE 4895 03:12:11,500 --> 03:12:12,601 ABSOLUTELY NO SIGNIFICANT 4896 03:12:12,601 --> 03:12:14,236 DIFFERENCE IN THEIR EXPRESSION 4897 03:12:14,236 --> 03:12:15,871 OF ANY OF THE STEM AND 4898 03:12:15,871 --> 03:12:17,306 PROGENITOR FACTORS THAT WE 4899 03:12:17,306 --> 03:12:18,707 LOOKED AT. 4900 03:12:18,707 --> 03:12:20,776 WITH AR WITHOUT THE EXTRA RUNX. 4901 03:12:20,776 --> 03:12:22,311 BUT THOSE SAME INDIVIDUAL CELLS 4902 03:12:22,311 --> 03:12:24,379 HAVE GREATLY UPREGULATED THEIR 4903 03:12:24,379 --> 03:12:30,886 EXPRESSION OF TCF7, LCK, GATA3, 4904 03:12:30,886 --> 03:12:32,654 TCF12 AND ONE THING VERY AMAZE 4905 03:12:32,654 --> 03:12:34,723 SG THEY UPREGULATE SOME ILC 4906 03:12:34,723 --> 03:12:39,194 GENES AND HAVE A TINY BIT OF 4907 03:12:39,194 --> 03:12:41,163 BCL11B VERY PREMATURELY UP 4908 03:12:41,163 --> 03:12:42,030 REGULATED, WHERE WEERVE VERY 4909 03:12:42,030 --> 03:12:43,298 INTERESTED, AGAIN, I'M SURE IF 4910 03:12:43,298 --> 03:12:44,800 WE HAVE ENOUGH TIME SAY THIS B 4911 03:12:44,800 --> 03:12:47,536 THE SITES THAT RUNX IS USING TO 4912 03:12:47,536 --> 03:12:49,338 MEDIATE THESE EFFECTS, IF ANY OF 4913 03:12:49,338 --> 03:12:51,473 YOU ARE INTERESTED, YOU CAN ASK 4914 03:12:51,473 --> 03:12:53,141 ME IN THE QUESTIONS, SO WE'VE 4915 03:12:53,141 --> 03:12:54,209 REALLY LOOKED AT WHAT ARE THE 4916 03:12:54,209 --> 03:12:57,279 SITES THAT ARE OCCUPIED IN THESE 4917 03:12:57,279 --> 03:12:58,347 EXACT EXPERIMENTAL CONDITIONS? 4918 03:12:58,347 --> 03:12:59,314 WE'VE LOOKED AT THE SITES WITH 4919 03:12:59,314 --> 03:13:01,550 REGARD TO THEIR AFFINITY, THEIR 4920 03:13:01,550 --> 03:13:03,518 MOTIF DENSITY, AND THE COFACTORS 4921 03:13:03,518 --> 03:13:04,887 THAT BIND THERE AND THERE ARE 4922 03:13:04,887 --> 03:13:06,688 SOME VERY INTERESTING PATTERNS 4923 03:13:06,688 --> 03:13:08,223 HERE THAT YOU'RE WELCOME TO ASK 4924 03:13:08,223 --> 03:13:08,891 ME ABOUT. 4925 03:13:08,891 --> 03:13:13,262 THE POINT IS THAT THESE ARE NOT 4926 03:13:13,262 --> 03:13:14,930 UNFORTUNATELY PURE RUNX TARGETS 4927 03:13:14,930 --> 03:13:16,999 IN THE SENSE THAT MOST 4928 03:13:16,999 --> 03:13:19,902 TRANSCRIPTION FACTORS WORK AS 4929 03:13:19,902 --> 03:13:21,136 MEMBERS OF ENSEMBLES ON 4930 03:13:21,136 --> 03:13:22,804 INDIVIDUAL ENHANCERS AND 4931 03:13:22,804 --> 03:13:24,106 UNFORTUNATELY A LOT OF THE EE 4932 03:13:24,106 --> 03:13:26,074 NEACTS WE'RE SEEING WITH RUNX 4933 03:13:26,074 --> 03:13:28,810 ARE COMPOSITES OF THE EFFECTS OF 4934 03:13:28,810 --> 03:13:30,545 RUNX ITSELF BINDING TO THESE NEW 4935 03:13:30,545 --> 03:13:31,980 SITES AND THE EFFECTS OF THE 4936 03:13:31,980 --> 03:13:32,948 OTHER TRANSCRIPTION FACTORS THAT 4937 03:13:32,948 --> 03:13:35,717 I JUST TOLD YOU THEY UPREGULATE. 4938 03:13:35,717 --> 03:13:35,918 OKAY. 4939 03:13:35,918 --> 03:13:39,621 SO STANLEY WANTS ME TO STOP. 4940 03:13:39,621 --> 03:13:44,026 SO FOR EXAMPLE, THE BCL11B 4941 03:13:44,026 --> 03:13:46,194 UPREGULATION DEPENDS ALSO ON THE 4942 03:13:46,194 --> 03:13:48,196 TCF7 THAT'S ALSO UP REGULATED. 4943 03:13:48,196 --> 03:13:49,631 SO I JUST WANT YOU TONED WITH 4944 03:13:49,631 --> 03:13:50,799 THIS PICTURE OF WHAT'S ACTUALLY 4945 03:13:50,799 --> 03:13:53,535 HAPPENING IN THESE CELLS. 4946 03:13:53,535 --> 03:13:55,304 THAT DURING ENTRY, TOWARDS THE T 4947 03:13:55,304 --> 03:13:56,838 CELL PROGRAM UNDER THE INFLUENCE 4948 03:13:56,838 --> 03:13:58,807 OF NOTCH SIGNALLING, THE CELLS 4949 03:13:58,807 --> 03:14:01,977 ARE ENTERING THIS PRO T CELL 4950 03:14:01,977 --> 03:14:04,313 STAGE WHICH IS VERY MUCH THE 4951 03:14:04,313 --> 03:14:06,615 STAGE, VERY CLOSE TO THE STAGE 4952 03:14:06,615 --> 03:14:10,118 THAT ROXANE WAS ABLE TO KEEP 4953 03:14:10,118 --> 03:14:12,120 SUSTAINED IN HER CULTURE SYSTEM. 4954 03:14:12,120 --> 03:14:15,057 BUT THAT THESE CELLS ARE REALLY 4955 03:14:15,057 --> 03:14:16,058 BALANCED BETWEEN POSSIBLY TRYING 4956 03:14:16,058 --> 03:14:18,794 TO GO BACK TO STEM AND 4957 03:14:18,794 --> 03:14:19,995 PROGENITOR, THEY HAVEN'T TOTALLY 4958 03:14:19,995 --> 03:14:21,296 LOST ALL OF THAT YET. 4959 03:14:21,296 --> 03:14:23,065 POSSIBLY GOING OFF SIDEWAYS TO 4960 03:14:23,065 --> 03:14:25,033 MYELOID AND DC WHICH IS ONE OF 4961 03:14:25,033 --> 03:14:25,834 THEIR FAVORITE OPTIONS IF YOU 4962 03:14:25,834 --> 03:14:27,669 TAKE AWAY THE NOTCH SIGNALLING, 4963 03:14:27,669 --> 03:14:30,739 OR GOING TO LYMPHOID CELL AND NK 4964 03:14:30,739 --> 03:14:32,908 AND THIS IS BASICALLY AN 4965 03:14:32,908 --> 03:14:35,577 EXPRESSION OF THE COMPLEX GENE 4966 03:14:35,577 --> 03:14:36,979 REGULATORY NETWORK RELATIONSHIPS 4967 03:14:36,979 --> 03:14:39,514 BETWEEN P THE DIFFERENT 4968 03:14:39,514 --> 03:14:41,283 REGULATORS THAT ARE 4969 03:14:41,283 --> 03:14:42,117 COINCIDENTALLY AT WAR IN THE 4970 03:14:42,117 --> 03:14:43,452 SAME CELLS. 4971 03:14:43,452 --> 03:14:45,420 OUR PICTURE OF T CELL 4972 03:14:45,420 --> 03:14:46,755 DEVELOPMENT THROUGH THESE EARLY 4973 03:14:46,755 --> 03:14:48,724 STAGES THEN COMES DOWN TO THESE 4974 03:14:48,724 --> 03:14:49,691 MAJOR POINTS. 4975 03:14:49,691 --> 03:14:51,927 THAT THE ENTRY INTO THE T CELL 4976 03:14:51,927 --> 03:14:54,129 PROGRAM DRIVEN BY NOTCH IS 4977 03:14:54,129 --> 03:14:57,599 REALLY A SLOW SUBVERSION OF AN 4978 03:14:57,599 --> 03:14:59,601 ESTABLISHED MULTITI POE NENLT 4979 03:14:59,601 --> 03:15:00,736 PROGENITOR REGULATORY STATE AND 4980 03:15:00,736 --> 03:15:02,771 THIS IS WHAT THE NOTCH 4981 03:15:02,771 --> 03:15:04,072 SIGNALLING CASCADE CASCADE IS 4982 03:15:04,072 --> 03:15:05,707 BUILT ON BUT THAT'S WHAT THE 4983 03:15:05,707 --> 03:15:07,242 NOTCH SIGNALLING CASCADE 4984 03:15:07,242 --> 03:15:09,978 UNDERMINES AND WE GET 4985 03:15:09,978 --> 03:15:11,179 ACQUISITION OF NEW 4986 03:15:11,179 --> 03:15:12,280 FUNCTIONALITIES OF TRANSCRIPTION 4987 03:15:12,280 --> 03:15:15,217 FACTORS LIKE RUNX AS A RESULT 4988 03:15:15,217 --> 03:15:17,119 BECAUSE OF THE CUMULATIVE 4989 03:15:17,119 --> 03:15:19,855 EXPRESSION OF OTHER PARTNERS, 4990 03:15:19,855 --> 03:15:21,556 WHICH HAS A NUMBER OF BEAUTIFUL 4991 03:15:21,556 --> 03:15:23,125 PIECES OF MOLECULAR BIOLOGY THAT 4992 03:15:23,125 --> 03:15:26,495 YOU CAN STUDY ABOUT HOW THE SITE 4993 03:15:26,495 --> 03:15:27,929 CHOICE WORKS AND HOW THAT 4994 03:15:27,929 --> 03:15:32,200 INTERFACES WITH THE EPIGENOME, 4995 03:15:32,200 --> 03:15:34,503 AND JUST THAT THIS WILL 4996 03:15:34,503 --> 03:15:35,470 ULTIMATELY NARROW OF THE SCOPE 4997 03:15:35,470 --> 03:15:38,240 OF THE RUNX ACTIVATED GENES TO 4998 03:15:38,240 --> 03:15:39,875 FAVOR T OVER INNAITD CELL 4999 03:15:39,875 --> 03:15:41,843 DEVELOPMENT ONCE YOU GET 5000 03:15:41,843 --> 03:15:50,919 DISAMBIGUATING FACTORS LIKE BCL 5001 03:15:50,919 --> 03:15:51,086 11B. 5002 03:15:51,086 --> 03:15:55,057 I WANT TO THANK IT MY LAB, THIS 5003 03:15:55,057 --> 03:15:56,591 IS MY LAB ABOUT A YEAR AK, THESE 5004 03:15:56,591 --> 03:15:57,692 ARE MARVELOUS PEOPLE, THEY 5005 03:15:57,692 --> 03:15:59,027 WORKED VERY, VERY HARD DURING 5006 03:15:59,027 --> 03:16:00,862 THE PANDEMIC EVEN THOUGH IT WAS 5007 03:16:00,862 --> 03:16:02,164 TOUGH TIMES FOR ALL. 5008 03:16:02,164 --> 03:16:03,398 AND I JUST WANT TO THANK THEM 5009 03:16:03,398 --> 03:16:05,267 AND I WANT TO THANK YOU FOR YOUR 5010 03:16:05,267 --> 03:16:05,534 ATTENTION. 5011 03:16:05,534 --> 03:16:06,668 AND IF STANLEY WILL BE KIND 5012 03:16:06,668 --> 03:16:09,504 ENOUGH TO LET ANY OF YOU ASK 5013 03:16:09,504 --> 03:16:11,073 QUESTIONS, EITHER INSIDE OR 5014 03:16:11,073 --> 03:16:13,108 OUTSIDE THE SESSION, I'M 5015 03:16:13,108 --> 03:16:15,210 DELIGHTED. 5016 03:16:15,210 --> 03:16:19,848 [APPLAUSE] 5017 03:16:19,848 --> 03:16:21,349 >> ELLEN, THAT WAS LOVELY. 5018 03:16:21,349 --> 03:16:22,751 CAN YOU HEAR ME? 5019 03:16:22,751 --> 03:16:23,118 >> YES, I CAN. 5020 03:16:23,118 --> 03:16:25,387 >> SO REALLY QUICK QUESTION. 5021 03:16:25,387 --> 03:16:27,022 I'LL ASK YOU ABOUT THIS AFTER, 5022 03:16:27,022 --> 03:16:29,424 BUT IN THE EXPERIMENTS WHERE YOU 5023 03:16:29,424 --> 03:16:33,061 OVEREXPRESSED RUNX SO MAYBE TAKE 5024 03:16:33,061 --> 03:16:35,363 AWAYLY PL ONE -- 5025 03:16:35,363 --> 03:16:37,232 >> CAN YOU COME A LITTLE CLOSER? 5026 03:16:37,232 --> 03:16:40,202 YOU'RE TOO FAR AWAY FROM THE 5027 03:16:40,202 --> 03:16:40,502 MICROPHONE. 5028 03:16:40,502 --> 03:16:42,504 >> THE EXPERIMENTS WHERE THE TWO 5029 03:16:42,504 --> 03:16:48,643 ALLELES OF BCL11B ASYNC NIS 5030 03:16:48,643 --> 03:16:51,246 NUSLY EXPRESSED IN THE SAME 5031 03:16:51,246 --> 03:16:52,614 CELL, IS ITLESS -- 5032 03:16:52,614 --> 03:16:54,182 >> THIS IS A BEAUTIFUL 5033 03:16:54,182 --> 03:16:55,951 EXPERIMENT ASKED ABOUT, I WANT 5034 03:16:55,951 --> 03:16:57,819 DESPERATELY TO DO THAT 5035 03:16:57,819 --> 03:16:59,554 EXPERIMENT H IT JUST HASN'T BEEN 5036 03:16:59,554 --> 03:17:01,223 DONE YET. 5037 03:17:01,223 --> 03:17:03,525 BUT IT IS A GREAT EXPERIMENT. 5038 03:17:03,525 --> 03:17:05,127 WE'RE A LITTLE SHORT OF 5039 03:17:05,127 --> 03:17:06,294 FLUORESCE ENTD PROTEIN COLORS, 5040 03:17:06,294 --> 03:17:09,097 BUT WE SHOULD BE ABLE TO DO IT 5041 03:17:09,097 --> 03:17:09,764 ANYWAY. 5042 03:17:09,764 --> 03:17:10,765 SO I VERY MUCH -- SO THE 5043 03:17:10,765 --> 03:17:13,602 QUESTION IS, DOES THE RUNX 5044 03:17:13,602 --> 03:17:15,604 OVEREXPRESSION COLLAPSE THE 5045 03:17:15,604 --> 03:17:17,005 EPIGENETIC BARRIER? 5046 03:17:17,005 --> 03:17:17,906 AND WE WOULD PREDICT THAT THAT 5047 03:17:17,906 --> 03:17:19,207 MIGHT BE THE CASE, AND I WOULD 5048 03:17:19,207 --> 03:17:20,742 LOVE TO DO THAT EXPERIMENT 5049 03:17:20,742 --> 03:17:24,446 SOONER RATHER THAN LATER. 5050 03:17:24,446 --> 03:17:29,384 ZOOS SO YOU SAID THE UF1 IS 5051 03:17:29,384 --> 03:17:31,153 MOSTLY PROLIFERATION GENES, SO 5052 03:17:31,153 --> 03:17:32,554 HOW MUCH IS THE CELL CYCLE? 5053 03:17:32,554 --> 03:17:35,624 I MEAN, YOU SHOWED IT WITH PU1 A 5054 03:17:35,624 --> 03:17:38,326 LONG TIME AGO, TRANSCRIPTION 5055 03:17:38,326 --> 03:17:42,063 FACTORS, LIKE HOW OFTEN IS THIS 5056 03:17:42,063 --> 03:17:45,367 INFLUENCING THE CONCENTRATION OR 5057 03:17:45,367 --> 03:17:49,604 THE AMOUNT OF PU1 AND FACTORS 5058 03:17:49,604 --> 03:17:51,139 AND INFLUENCING THE CHOICE OF 5059 03:17:51,139 --> 03:17:53,675 THE SITES THAT YOU HAVE? 5060 03:17:53,675 --> 03:17:55,510 >> WHERE THEY ARE IN THE CELL 5061 03:17:55,510 --> 03:17:55,777 CYCLE? 5062 03:17:55,777 --> 03:17:57,179 >> ZEECT WE HAVEN'T DONE THAT 5063 03:17:57,179 --> 03:18:00,215 EXACT SPRMT THE WAY YOU'RE 5064 03:18:00,215 --> 03:18:00,448 ASKING. 5065 03:18:00,448 --> 03:18:02,717 BUT PU1 IS A VERY STABLE 5066 03:18:02,717 --> 03:18:04,386 TRANSCRIPTION FACTOR IN THESE 5067 03:18:04,386 --> 03:18:05,787 CELLS, SO IT GETS CARRIED 5068 03:18:05,787 --> 03:18:07,422 THROUGH THE CELL CYCLE, BUT THAT 5069 03:18:07,422 --> 03:18:09,291 ALSO MEANS THAT ITS LEVEL DROPS 5070 03:18:09,291 --> 03:18:11,393 BY A FOK TORE OF TWO EVERY TIME 5071 03:18:11,393 --> 03:18:12,160 THE CELLS DIVIDE. 5072 03:18:12,160 --> 03:18:12,794 >> EXACT. 5073 03:18:12,794 --> 03:18:13,995 >> AND I REALLY DON'T KNOW THE 5074 03:18:13,995 --> 03:18:16,865 ANSWER TO THAT QUESTION, 5075 03:18:16,865 --> 03:18:17,165 LITERALLY. 5076 03:18:17,165 --> 03:18:20,035 THESE CELLS UNLIKE THE SITUATION 5077 03:18:20,035 --> 03:18:21,903 THAT YOU WERE HEARING ABOUT FROM 5078 03:18:21,903 --> 03:18:23,872 DINAH, THESE CELLS ARE REALLY 5079 03:18:23,872 --> 03:18:25,707 PROLIFERATING ALL THROUGH THIS 5080 03:18:25,707 --> 03:18:28,310 PERIOD, SO EVERY ONE OF THESE 5081 03:18:28,310 --> 03:18:29,644 DEVELOPMENTAL STAGES HAS THE 5082 03:18:29,644 --> 03:18:31,379 FULL RANGE AND THAT'S WHY IT 5083 03:18:31,379 --> 03:18:33,148 REALLY DOESN'T SEPARATE THINGS 5084 03:18:33,148 --> 03:18:36,451 VERY WELL UNTIL YOU GET TO DM3 5085 03:18:36,451 --> 03:18:36,651 STAGE. 5086 03:18:36,651 --> 03:18:38,853 BUT IT'S A REALLY COOL QUESTION, 5087 03:18:38,853 --> 03:18:41,022 AND IT SOUNDS LIKE IT MIGHT BE A 5088 03:18:41,022 --> 03:18:42,657 WINDOW OF OPPORTUNITY FOR GENE 5089 03:18:42,657 --> 03:18:43,992 EXPRESSION CHANGE TO OCCUR. 5090 03:18:43,992 --> 03:18:45,627 SO I REALLY LIKE YOUR QUESTION. 5091 03:18:45,627 --> 03:18:47,829 THANK YOU. 5092 03:18:47,829 --> 03:18:50,365 >> I HAVE A POTENTIALLY NAIVE 5093 03:18:50,365 --> 03:18:50,699 QUESTION. 5094 03:18:50,699 --> 03:18:53,668 SO THE RECRUITMENT OF RUNX TO 5095 03:18:53,668 --> 03:18:54,636 THESE ALTERNATIVE SITES WHEN YOU 5096 03:18:54,636 --> 03:18:57,706 HAVE PU1, HOW IMPORTANT, WHAT IS 5097 03:18:57,706 --> 03:19:00,442 THE CONTRIBUTION OF THAT EFFECT 5098 03:19:00,442 --> 03:19:03,044 BASED ON THE REPRESSIVE COME 5099 03:19:03,044 --> 03:19:05,046 PLEKS THAT RUNX CAN BRING AND 5100 03:19:05,046 --> 03:19:07,382 HOW THAT AFFECTS TRANSCRIPTION 5101 03:19:07,382 --> 03:19:09,784 OF THOSE GENES VERSUS JUST 5102 03:19:09,784 --> 03:19:11,286 DISTRACTING THE RUNX FROM 5103 03:19:11,286 --> 03:19:13,188 BINDING TO THE SITES. 5104 03:19:13,188 --> 03:19:14,422 >> IT'S I'VE GREAT QUESTION. 5105 03:19:14,422 --> 03:19:16,791 IT'S NOT AT ALL NAIVE. 5106 03:19:16,791 --> 03:19:18,860 THE PROBLEM IS RUNX ACTUALLY HAS 5107 03:19:18,860 --> 03:19:20,061 QUITE A LOT OF SITES AND IT 5108 03:19:20,061 --> 03:19:22,364 BINDS TO MULTIPLE KINDS OF SITES 5109 03:19:22,364 --> 03:19:25,433 AROUND EACH OF THE GENES, SO 5110 03:19:25,433 --> 03:19:26,868 WE'VE SPENT AN ENORMOUS AMOUNT 5111 03:19:26,868 --> 03:19:29,371 OF TIME TRYING ON ANALYZE THIS 5112 03:19:29,371 --> 03:19:30,338 GENOMEWIDE AND IT'S REALLY KIND 5113 03:19:30,338 --> 03:19:34,209 OF FRUSTRATING BECAUSE WHATEVER 5114 03:19:34,209 --> 03:19:35,710 THE GENE IS GOING TO DO IN 5115 03:19:35,710 --> 03:19:38,146 RESPONSE TO RUNX, THE MOST 5116 03:19:38,146 --> 03:19:40,215 ABUNDANT KIND OF SITES ARE THE 5117 03:19:40,215 --> 03:19:49,090 SITES THAT ARE ALWAYS THERE. 5118 03:19:49,090 --> 03:19:50,959 DOES IT REALLY EXPLAIN THE 5119 03:19:50,959 --> 03:19:54,029 DIFFERENTIAL ACTIVITY UP AND 5120 03:19:54,029 --> 03:19:54,829 DOWN, WILL BE THERE IN ADDITION 5121 03:19:54,829 --> 03:19:56,197 TO SITES THAT ARE JUST THERE ALL 5122 03:19:56,197 --> 03:19:57,932 THE TIME. 5123 03:19:57,932 --> 03:20:00,702 AND WITH SOMETHING ON THE ORDER 5124 03:20:00,702 --> 03:20:02,137 OF 25,000 SITES OCCUPIED IT'S 5125 03:20:02,137 --> 03:20:05,573 REALLY SCARY TO TRY TO COME UP 5126 03:20:05,573 --> 03:20:08,576 WITH A STRATEGY FOR KNOCKING OUT 5127 03:20:08,576 --> 03:20:11,513 INDIVIDUAL SITES F WE'RE REALLY 5128 03:20:11,513 --> 03:20:12,947 NICE TARGET GENES TO LOOK AT. 5129 03:20:12,947 --> 03:20:14,149 SO WE WOULD LOVE TO KNOW THE 5130 03:20:14,149 --> 03:20:15,784 ANSWER TO YOUR QUESTION. 5131 03:20:15,784 --> 03:20:17,952 WE'VE TRIED TO LOOK FOR MOTIF 5132 03:20:17,952 --> 03:20:20,588 ENRICHMENT THAT MIGHT BE MORE 5133 03:20:20,588 --> 03:20:22,991 ASSOCIATED WITH REPRESSIVE 5134 03:20:22,991 --> 03:20:25,193 VERSUS ACTIVATING INTERACTIONS 5135 03:20:25,193 --> 03:20:28,363 IN A STAGE-SPECIFIC WAY, BUT THE 5136 03:20:28,363 --> 03:20:33,168 SIGNAL YOU GET BACK FROM RUNX X 5137 03:20:33,168 --> 03:20:35,904 WHETHER IT'S RUNX AND PU1 IN 5138 03:20:35,904 --> 03:20:40,842 EARLIER STAGE OR RUNX AND X1 IS 5139 03:20:40,842 --> 03:20:42,377 EARLIER EXRRKS P PROTEIN IN 5140 03:20:42,377 --> 03:20:44,112 LATER STAGE, YOU JUST DON'T SEE 5141 03:20:44,112 --> 03:20:44,879 WHAT MIGHT BE SPECIFIC. 5142 03:20:44,879 --> 03:20:46,281 IT'S A GREAT QUESTION, WE HOPE 5143 03:20:46,281 --> 03:20:47,415 WE'LL COME UP WITH BETTER WAYS 5144 03:20:47,415 --> 03:20:48,016 OF ANSWERING IT. 5145 03:20:48,016 --> 03:20:50,218 >> THANK YOU. 5146 03:20:50,218 --> 03:20:50,819 >> ALL RIGHT. 5147 03:20:50,819 --> 03:20:51,453 THAT WAS GREAT. 5148 03:20:51,453 --> 03:20:53,621 I WAS WONDERING ABOUT THE ROLE 5149 03:20:53,621 --> 03:20:55,290 OF THE GENETIC BACKGROUND OF THE 5150 03:20:55,290 --> 03:20:55,490 MICE. 5151 03:20:55,490 --> 03:20:56,324 >> OH. 5152 03:20:56,324 --> 03:20:58,326 >> AND JUST BECAUSE LIKE WE'VE 5153 03:20:58,326 --> 03:21:01,963 BEEN LOOKING AT MACROPHAGES AND 5154 03:21:01,963 --> 03:21:04,165 SEE VAST DIFFERENCE INSIDE PU1 5155 03:21:04,165 --> 03:21:05,133 ACTIVITY BETWEEN DIFFERENT 5156 03:21:05,133 --> 03:21:06,201 STRAINS OF MICE. 5157 03:21:06,201 --> 03:21:08,470 >> IT IS REALLY A GREAT THING TO 5158 03:21:08,470 --> 03:21:08,636 DO. 5159 03:21:08,636 --> 03:21:10,505 I MEAN, WE DON'T HAVE THE 5160 03:21:10,505 --> 03:21:11,940 RESOURCES TO LOOK AT THE 5161 03:21:11,940 --> 03:21:15,009 DIFFERENT MOUSE STRAINS THE WAY 5162 03:21:15,009 --> 03:21:17,812 WE KNOW SHOULD BE DONE, AND IF 5163 03:21:17,812 --> 03:21:20,014 ANYONE WANTS TO DO THAT WITH A 5164 03:21:20,014 --> 03:21:21,049 GOOD LIBRARY OF MOUSE STRAINS, 5165 03:21:21,049 --> 03:21:22,617 WE WOULD BE HAPPY TO HELP 5166 03:21:22,617 --> 03:21:23,852 BECAUSE I THINK SCIENTIFICALLY 5167 03:21:23,852 --> 03:21:25,987 IT'S SUCH AN IMPORTANT QUESTION. 5168 03:21:25,987 --> 03:21:27,922 YOU KNOW, YOU HAVE 5169 03:21:27,922 --> 03:21:29,357 POLYMORPHISMLESS THAT AFFECT 5170 03:21:29,357 --> 03:21:31,926 THIS, AND IT'S SOMETHING WE WILL 5171 03:21:31,926 --> 03:21:33,528 NEVER BE ABLE TO DO IT AT CAL 5172 03:21:33,528 --> 03:21:35,130 TECH WITH OUR LITTLE MOUSE 5173 03:21:35,130 --> 03:21:36,398 COLONY BUT I WOULD LOVE TO SEE 5174 03:21:36,398 --> 03:21:41,503 SOMEBODY ELSE RUN WITH THAT. 5175 03:21:41,503 --> 03:21:43,638 >> SO IT'S VERY NICE. 5176 03:21:43,638 --> 03:21:45,173 TWO QUICK QUESTIONS. 5177 03:21:45,173 --> 03:21:48,042 FIRST TO GO BACK TO THE 5178 03:21:48,042 --> 03:21:49,010 QUESTION, DO YOU KNOW IF THE 5179 03:21:49,010 --> 03:21:49,244 BRAISH. 5180 03:21:49,244 --> 03:21:51,846 >> CLOSER TO THE MIKE ZEEN THE 5181 03:21:51,846 --> 03:21:55,283 MOTIF THAT BINDS -- AT THE END 5182 03:21:55,283 --> 03:21:57,485 IS NECESSARY TO SEE WHAT YOU 5183 03:21:57,485 --> 03:22:00,188 SEE, AND THAT COULD HELP YOU 5184 03:22:00,188 --> 03:22:01,756 PERHAPS DISTINGUISH BETWEEN 5185 03:22:01,756 --> 03:22:05,794 ACTIVATE AND GO REPRESSING 5186 03:22:05,794 --> 03:22:07,762 FUNCTIONS? 5187 03:22:07,762 --> 03:22:10,932 >> THERE'S VWRPY IS A FAMOUS 5188 03:22:10,932 --> 03:22:12,700 MOTIF AND A LOT OF WORK WAS DONE 5189 03:22:12,700 --> 03:22:15,637 IN JAPAN OVER TEN YEARS AGO 5190 03:22:15,637 --> 03:22:18,940 COMPARING THE EFFECTS OF RUNX1 5191 03:22:18,940 --> 03:22:23,878 WITHOUT VWRPY MOTIF. 5192 03:22:23,878 --> 03:22:25,613 S STILL DOING VERY ELEGANT WORK 5193 03:22:25,613 --> 03:22:27,482 ON THIS AS YOU KNOW BUT OTHERS 5194 03:22:27,482 --> 03:22:30,952 MAY NOT IN THE CD4, THE OLD DATA 5195 03:22:30,952 --> 03:22:34,389 MADE IT SEEM AS THOUGH THAT 5196 03:22:34,389 --> 03:22:35,557 WASN'T IMPORTANT FOR THE EARLY 5197 03:22:35,557 --> 03:22:37,225 STAGES THAT I WORK ON, BUT AT 5198 03:22:37,225 --> 03:22:39,928 THAT TIME IT REALLY WASN'T 5199 03:22:39,928 --> 03:22:41,596 APPRECIATED HOW EXTREMELY 5200 03:22:41,596 --> 03:22:43,998 PERVASIVE RUNX3 IS AT THE SAME 5201 03:22:43,998 --> 03:22:44,766 TIME. 5202 03:22:44,766 --> 03:22:46,468 AND RUNX3 IS BASICALLY EXPRESSED 5203 03:22:46,468 --> 03:22:49,471 THE AAN EVEN HIGHER LEVEL THAN 5204 03:22:49,471 --> 03:22:50,672 RUNX1 IN THE CELLS THAT ARE AT 5205 03:22:50,672 --> 03:22:52,674 THE ETP STAGE THEY'RE ABOUT 5206 03:22:52,674 --> 03:22:56,511 EQUAL IN DM2A AND THEN THE RUNX1 5207 03:22:56,511 --> 03:22:57,912 BECOMES DOMINANT IN THE NEXT 5208 03:22:57,912 --> 03:22:58,146 STAGES. 5209 03:22:58,146 --> 03:22:59,547 SO ESSENTIALLY THEY WOULDN'T 5210 03:22:59,547 --> 03:23:02,617 HAVE SEEN AN EFFECT BECAUSE THEY 5211 03:23:02,617 --> 03:23:05,253 WOULDN'T -- BY SIMPLY NOT IF 5212 03:23:05,253 --> 03:23:06,888 HANGING OUT OF R -- KNOCKING OUT 5213 03:23:06,888 --> 03:23:09,224 OF RUNX 1RBGS IT WOULD HAVE HAD 5214 03:23:09,224 --> 03:23:10,458 TO BE KNOCKED OUT OF BOTH AND I 5215 03:23:10,458 --> 03:23:11,593 DON'T KNOW IF THAT'S BEEN DONE. 5216 03:23:11,593 --> 03:23:12,794 >> YOU COULD DO THAT IS CORRECT 5217 03:23:12,794 --> 03:23:15,663 RIGHT, OVEREXPRESS THE -- 5218 03:23:15,663 --> 03:23:17,832 >> YEAH, BUT YOU WOULD HAVE TO 5219 03:23:17,832 --> 03:23:19,501 WORRY ABOUT, WE MIGHT NOT BE AS 5220 03:23:19,501 --> 03:23:22,871 LUCKY WITH THE REGULATION OF THE 5221 03:23:22,871 --> 03:23:24,072 SELF-REGULATION IF YOU DIDN'T 5222 03:23:24,072 --> 03:23:25,039 HAVE THAT. 5223 03:23:25,039 --> 03:23:27,008 >> THE OTHER QUESTION, WHEN YOU 5224 03:23:27,008 --> 03:23:29,744 KNOCK OUT BOTH 1 AND 3, THE 5225 03:23:29,744 --> 03:23:32,480 CELLS DON'T CRASH? 5226 03:23:32,480 --> 03:23:35,316 THE CELLS DON'T CRASH? 5227 03:23:35,316 --> 03:23:38,920 EVEN DOUBLE POSITIVE THERM 5228 03:23:38,920 --> 03:23:41,122 SIGHTS, THEY DON'T -- 5229 03:23:41,122 --> 03:23:42,223 THYMOCYTES, DHOANT GO VERY FAR? 5230 03:23:42,223 --> 03:23:43,925 >> WE'VE NEVER DONE THAT. 5231 03:23:43,925 --> 03:23:47,362 MAYBE OTHERS HAVE. 5232 03:23:47,362 --> 03:23:50,231 >> THE DOUBLE NEGATIVITY EARLY 5233 03:23:50,231 --> 03:23:51,299 PROGENITORS, THEY SURVIVE? 5234 03:23:51,299 --> 03:23:53,134 >> THEY COMPLETELY -- THEY LIVE. 5235 03:23:53,134 --> 03:23:53,568 >> OKAY. 5236 03:23:53,568 --> 03:23:54,936 >> THEY LIVE. 5237 03:23:54,936 --> 03:23:57,105 BUT THEY DON'T DEVELOP PAST THE 5238 03:23:57,105 --> 03:23:57,639 EARLY STAGE. 5239 03:23:57,639 --> 03:23:58,273 >> OKAY. 5240 03:23:58,273 --> 03:23:59,107 THANK YOU. 5241 03:23:59,107 --> 03:24:00,808 >> THANK YOU. 5242 03:24:00,808 --> 03:24:02,677 >> PERHAPS YOU ALREADY MENTIONED 5243 03:24:02,677 --> 03:24:04,646 THIS, BUT WHEN YOU OVEREXPRESS, 5244 03:24:04,646 --> 03:24:06,481 DO YOU SEE ANY PREFERENTIAL 5245 03:24:06,481 --> 03:24:08,917 BINDING TO SITES LIKE ENHANCERS 5246 03:24:08,917 --> 03:24:09,884 OR PROMOTERS? 5247 03:24:09,884 --> 03:24:12,086 >> SO THE PROMOTERS IN OUR 5248 03:24:12,086 --> 03:24:16,057 SYSTEM ARE ALMOST COMPLETELY 5249 03:24:16,057 --> 03:24:19,961 DEVOID OF USEFUL INFORMATION. 5250 03:24:19,961 --> 03:24:22,797 BASICALLY, YOU SEE IT BINDING TO 5251 03:24:22,797 --> 03:24:24,566 PROMOTERS AT LEAST AS MUCH OF 5252 03:24:24,566 --> 03:24:26,100 GENES THAT IT DOESN'T REGULATE 5253 03:24:26,100 --> 03:24:28,369 AT ALL AS THE GENES THAT IT DOES 5254 03:24:28,369 --> 03:24:28,970 REGULATE. 5255 03:24:28,970 --> 03:24:31,239 SO WE'VE REALLY FOCUSED ON 5256 03:24:31,239 --> 03:24:32,240 NONPROMOTER SITES WHERE MOST OF 5257 03:24:32,240 --> 03:24:34,075 THE BINDING IS AND BY USING CUT 5258 03:24:34,075 --> 03:24:36,377 AND RUN, WE'VE ALSO BIASED OUR 5259 03:24:36,377 --> 03:24:39,547 DETECTION TO NON-PROMOTER SITES. 5260 03:24:39,547 --> 03:24:43,151 SO THOSE ARE, IT'S MOSTLY THE 5261 03:24:43,151 --> 03:24:44,919 ADDED SITES, THE ECTOPIC SITES, 5262 03:24:44,919 --> 03:24:49,958 AS IT WERE, ARE, BEING SOME OF 5263 03:24:49,958 --> 03:24:51,559 THEM ARE BASICALLY SITES THAT 5264 03:24:51,559 --> 03:24:53,962 REALLY WERE JUST LOW LEVEL RUNX 5265 03:24:53,962 --> 03:24:55,196 SITES ANYWAY AND THEY WERE DOING 5266 03:24:55,196 --> 03:24:55,663 SOMETHING. 5267 03:24:55,663 --> 03:24:58,232 MANY OF THEM WERE IN CLOSE 5268 03:24:58,232 --> 03:24:59,767 CHROMATIN AND SEEM TO HAVE 5269 03:24:59,767 --> 03:25:03,705 ABSOLUTELY NO SIGNIFICANT 5270 03:25:03,705 --> 03:25:06,774 ASSOCIATION WITH GENES WHICH ARE 5271 03:25:06,774 --> 03:25:09,410 EXPRESSED UNDER RUNX CONTROL AND 5272 03:25:09,410 --> 03:25:11,379 OTHER THINGS FOR EXAMPLE LIKE 5273 03:25:11,379 --> 03:25:13,147 ILC'S, SO I THINK AGAIN THEY'RE 5274 03:25:13,147 --> 03:25:15,216 GOING MAINLY, WE'RE SEEING 5275 03:25:15,216 --> 03:25:18,052 MOSTLY THE INTERGENIC AND 5276 03:25:18,052 --> 03:25:20,688 INTRONIC SITES THAT ARE OCCUPIED 5277 03:25:20,688 --> 03:25:21,055 DIFFERENTIALLY. 5278 03:25:21,055 --> 03:25:23,424 >> AND DOES THE COMPLEX FORM, I 5279 03:25:23,424 --> 03:25:25,360 MEAN, DO YOU FORM A COMPLEX OF 5280 03:25:25,360 --> 03:25:28,129 THE PROTEINS OR IS IT A LARGE 5281 03:25:28,129 --> 03:25:31,332 MACROMOLECULAR COMPLEX WHEN YOU 5282 03:25:31,332 --> 03:25:31,633 OVEREXPRESS? 5283 03:25:31,633 --> 03:25:33,601 >> WE HAVEN'T DONE ANY 5284 03:25:33,601 --> 03:25:34,502 PROTEOMICS ON IT. 5285 03:25:34,502 --> 03:25:36,237 IT'S A TWOFOLD OVEREXPRESSION, 5286 03:25:36,237 --> 03:25:37,905 IT'S BASICALLY GOING TO HIGH 5287 03:25:37,905 --> 03:25:40,608 AFFINITY RUNX SITES THAT WERE 5288 03:25:40,608 --> 03:25:42,377 ENCLOSED CHROMATIN. 5289 03:25:42,377 --> 03:25:42,844 THANKS. 5290 03:25:42,844 --> 03:25:44,345 SORRY TO BE LONG. 5291 03:25:44,345 --> 03:25:46,381 THANKS. 5292 03:25:46,381 --> 03:25:50,918 [APPLAUSE] 5293 03:25:50,918 --> 03:25:52,654 >> I THINK WE ALL AGREE THAT WAS 5294 03:25:52,654 --> 03:25:54,055 A VERY STIMULATING SESSION THIS 5295 03:25:54,055 --> 03:25:54,422 MORNING. 5296 03:25:54,422 --> 03:25:58,159 SO PLEASE ON BEHALF, ANOTHER 5297 03:25:58,159 --> 03:25:59,961 ROUND OF APPLAUSE TO ALL THE 5298 03:25:59,961 --> 03:26:00,103 SPEAKERS THIS MORNING. 5299 03:26:00,103 --> 03:26:02,739 OUR FIRST SPEAKER IS CLAUDIA 5300 03:26:02,739 --> 03:26:06,276 KEMPER FROM NHLBI, TALKING TO US 5301 03:26:06,276 --> 03:26:07,878 ABOUT FUNCTIONS FOR 5302 03:26:07,878 --> 03:26:08,578 INTRACELLULAR COMPLEMENT WHICH 5303 03:26:08,578 --> 03:26:11,081 IS A FIELD THAT SHE DISCOVERED 5304 03:26:11,081 --> 03:26:12,516 AND ELABORATED AND I'M LOOKING 5305 03:26:12,516 --> 03:26:15,652 FORWARD TO HEARING ABOUT IT, 5306 03:26:15,652 --> 03:26:20,690 CLAUDIA. 5307 03:26:20,690 --> 03:26:22,225 >> CAN YOU HEAR ME ALL? 5308 03:26:22,225 --> 03:26:22,492 WONDERFUL. 5309 03:26:22,492 --> 03:26:22,792 THANK YOU. 5310 03:26:22,792 --> 03:26:24,861 I WOULD LIKE TO THANK THE 5311 03:26:24,861 --> 03:26:26,062 ORGANIZERS FOR INVITING ME. 5312 03:26:26,062 --> 03:26:27,330 IT'S A FANTASTIC MEETING. 5313 03:26:27,330 --> 03:26:28,798 I REALLY, REALLY ENJOY IT. 5314 03:26:28,798 --> 03:26:30,634 BUT WHAT I'M GOING TO DO NOW IS 5315 03:26:30,634 --> 03:26:35,205 I'M DOING SOMETHING SLIGHTLY 5316 03:26:35,205 --> 03:26:36,973 DIFFERENT, I'M NOT SHOWCASING 5317 03:26:36,973 --> 03:26:38,208 ONE STORY WE'RE WORKING ON I 5318 03:26:38,208 --> 03:26:39,743 HAVE PUBLISHED RECENTLY BUT I 5319 03:26:39,743 --> 03:26:41,711 WOULD RATHER TAKE YOU ONTO A 5320 03:26:41,711 --> 03:26:43,113 JOURNEY IN MY RESEARCH PROGRAM, 5321 03:26:43,113 --> 03:26:44,848 THE REASON BEING THAT I THINK 5322 03:26:44,848 --> 03:26:46,950 THERE WERE SOME CHANGES IN THE 5323 03:26:46,950 --> 03:26:49,019 COMPLEMENT HERE LATELY THAT ARE 5324 03:26:49,019 --> 03:26:50,220 PRETTY PROFOUND AND THAT MAYBE 5325 03:26:50,220 --> 03:26:54,157 NOT EVERYBODY HAS HEARD ABOUT 5326 03:26:54,157 --> 03:26:55,225 YET AND I THINK SOME OF THOSE 5327 03:26:55,225 --> 03:26:56,893 MAY BE REALLY IMPORTANT FOR WHAT 5328 03:26:56,893 --> 03:26:57,827 YOU'RE WORKING ON. 5329 03:26:57,827 --> 03:26:59,196 ONE OF THE REALLY MOST EXCITING 5330 03:26:59,196 --> 03:27:01,164 PARTS IN SCIENCE IS THAT I THINK 5331 03:27:01,164 --> 03:27:02,699 YOU NEVER TRAVEL ALONE, AND 5332 03:27:02,699 --> 03:27:04,334 NOTHING IS DONE IN ISOLATION. 5333 03:27:04,334 --> 03:27:05,502 SO WHAT I WANT TO DO IS ACTUALLY 5334 03:27:05,502 --> 03:27:07,170 I WANTED TO START FIRST WITH THE 5335 03:27:07,170 --> 03:27:08,471 MOST IMPORTANT SLIDE AND SHOW 5336 03:27:08,471 --> 03:27:10,907 YOU MY TEAM HERE AT THE NIH AND 5337 03:27:10,907 --> 03:27:13,009 ALSO THE COLLABORATORS ACROSS 5338 03:27:13,009 --> 03:27:16,046 THE NIH AND BEYOND THAT HAVE 5339 03:27:16,046 --> 03:27:17,514 HELPED DEVELOPING THESE NEW 5340 03:27:17,514 --> 03:27:18,148 PARADIGMS AND THE COMPLEMENT 5341 03:27:18,148 --> 03:27:22,986 FIELD THAT I'M SHARING WITH YOU 5342 03:27:22,986 --> 03:27:23,653 NOW. 5343 03:27:23,653 --> 03:27:24,621 I. HAD BEEN WHEN YOU HEAR 5344 03:27:24,621 --> 03:27:25,422 COMPLEMENT FIELD OR SOMETHING 5345 03:27:25,422 --> 03:27:28,458 LIKE THIS, OH, MY GOD, CASS 5346 03:27:28,458 --> 03:27:30,427 SCRAID, COMPLEMENT C3 AND C5. 5347 03:27:30,427 --> 03:27:32,162 JUST A BRIEF ONE. 5348 03:27:32,162 --> 03:27:34,431 WE ALL THAN THESE COMPLEMENT 5349 03:27:34,431 --> 03:27:36,299 COMPONENTS ARE SECRETED BY THE 5350 03:27:36,299 --> 03:27:37,500 LIVER, CIRCULATE NG YOUR BLOOD 5351 03:27:37,500 --> 03:27:39,135 AND YOU REALLY REALLY NEED THEM 5352 03:27:39,135 --> 03:27:42,572 BECAUSE C3 AND C5 ACTIVATION 5353 03:27:42,572 --> 03:27:44,808 FRAGMENTS PROTECT YOU AGAINST 5354 03:27:44,808 --> 03:27:46,409 INVADING PATHOGENS. 5355 03:27:46,409 --> 03:27:48,712 SO THAT'S A REALLY, REALLY GOOD 5356 03:27:48,712 --> 03:27:49,613 PART ABOUT THE COMPLEMENT 5357 03:27:49,613 --> 03:27:50,480 SYSTEM. 5358 03:27:50,480 --> 03:27:52,215 HOWEVER, THERE'S UNFORTUNATELY 5359 03:27:52,215 --> 03:27:53,650 ALSO REALLY A QUITE DARK SIDE 5360 03:27:53,650 --> 03:27:55,185 ABOUT THIS SYSTEM. 5361 03:27:55,185 --> 03:27:57,287 WHEN ACCOUNTS DYSREGULATED -- 5362 03:27:57,287 --> 03:28:01,524 WHEN IT IS DYSREGULATED, AND 5363 03:28:01,524 --> 03:28:05,462 THAT UNFORTUNATELY IS BECAUSE 5364 03:28:05,462 --> 03:28:07,230 IT'S TYPICAL 5365 03:28:07,230 --> 03:28:09,633 INFLAMMATION-DRIVING SYSTEM AND 5366 03:28:09,633 --> 03:28:11,735 THESE ACUTE OR, YOU KNOW, 5367 03:28:11,735 --> 03:28:13,570 SUSTAINED CHRONIC COMPLEMENT 5368 03:28:13,570 --> 03:28:15,105 ACTIVATION PATHWAYS CAN THEN 5369 03:28:15,105 --> 03:28:17,307 CONTRIBUTE TO HUMAN DISEASES. 5370 03:28:17,307 --> 03:28:20,877 AND ACTUALLY THE LIST OF HUMAN 5371 03:28:20,877 --> 03:28:22,112 DISEASES, THE COMPLEMENT HAS TO 5372 03:28:22,112 --> 03:28:24,180 BE DEFINED AS A DEFINITIVE 5373 03:28:24,180 --> 03:28:26,483 EITHER INDIERS OR PATHOLOGY 5374 03:28:26,483 --> 03:28:29,653 DRIVER IS REALLY LONG. 5375 03:28:29,653 --> 03:28:31,421 -- EITHER INDIERS OR PATHOLOGY 5376 03:28:31,421 --> 03:28:32,422 DRIVER IS REALLY LONG. 5377 03:28:32,422 --> 03:28:33,456 THIS IS JUST A PART OF IT. 5378 03:28:33,456 --> 03:28:37,327 WE KNOW FOR ALL SIX DECADES THAT 5379 03:28:37,327 --> 03:28:38,528 COMPLEMENT AS SUPREME 5380 03:28:38,528 --> 03:28:39,529 THERAPEUTIC TARGET AND WE HAVE 5381 03:28:39,529 --> 03:28:41,264 WORKED ON TARGETING COMPLEMENT 5382 03:28:41,264 --> 03:28:43,066 IN RARE AND COMMON DISEASES FOR 5383 03:28:43,066 --> 03:28:44,768 A VERY, VERY LONG TIME, AND WHEN 5384 03:28:44,768 --> 03:28:46,970 I MEAN WE, IT'S ACTUALLY 5385 03:28:46,970 --> 03:28:50,707 CLINICIANS, SCIENTISTS AND THE 5386 03:28:50,707 --> 03:28:51,141 INDUSTRY. 5387 03:28:51,141 --> 03:28:53,009 AS YOU CAN SEE JUST THE NIH IN 5388 03:28:53,009 --> 03:29:02,619 THE LAST TEN YEARS SPENT ALMOST 5389 03:29:02,619 --> 03:29:03,820 1,000 MILLION DOLLARS, THE 5390 03:29:03,820 --> 03:29:06,323 OUTCOME SO FAR IS REALLY QUITE 5391 03:29:06,323 --> 03:29:06,656 DISAPPOINTING. 5392 03:29:06,656 --> 03:29:08,625 WE CURRENTLY HAVE ONLY TEN DRUGS 5393 03:29:08,625 --> 03:29:10,694 THAT ARE EITHER FDA APPROVED OR 5394 03:29:10,694 --> 03:29:12,696 APPROVED FOR CLINICAL USAGE OR 5395 03:29:12,696 --> 03:29:14,397 APPLICATION AT THE EUROPEAN OR 5396 03:29:14,397 --> 03:29:15,732 ASIAN MARKET. 5397 03:29:15,732 --> 03:29:18,468 AND WHAT IS REALLY IMPORTANT IS 5398 03:29:18,468 --> 03:29:20,904 ALL OF THESE DRUGS REALLY ONLY 5399 03:29:20,904 --> 03:29:23,139 TACKLE RARE DISEASES, NONE OF 5400 03:29:23,139 --> 03:29:26,209 THESE DRUGS TACKLE 5401 03:29:26,209 --> 03:29:28,044 ATHEROSCHOARSZ OR ARTHRITIS OR 5402 03:29:28,044 --> 03:29:30,280 IBD, WE ALL KNOW COMPLEMENT 5403 03:29:30,280 --> 03:29:31,681 PLAYS A MAJOR ROLE. 5404 03:29:31,681 --> 03:29:33,650 SO WE'RE NOT 100% SURE QULET AS 5405 03:29:33,650 --> 03:29:35,719 A FIELD WHY THAT IS -- YET AS A 5406 03:29:35,719 --> 03:29:36,720 FIELD WHY THAT IS, BUT WHAT I 5407 03:29:36,720 --> 03:29:37,721 WOULD LIKE TO SHARE WITH YOU 5408 03:29:37,721 --> 03:29:39,356 TODAY AND MAKE A CASE FOR IS 5409 03:29:39,356 --> 03:29:40,957 THAT MAYBE ONE OF THE REASONS 5410 03:29:40,957 --> 03:29:42,859 WHY WE HAVEN'T BEEN SO 5411 03:29:42,859 --> 03:29:44,527 SUCCESSFUL IS BECAUSE WE REALLY 5412 03:29:44,527 --> 03:29:46,329 HAVEN'T UNDERSTOOD THE SYSTEM 5413 03:29:46,329 --> 03:29:46,496 YET. 5414 03:29:46,496 --> 03:29:49,833 ALL OF KNEES DRUGS TRADITIONALLY 5415 03:29:49,833 --> 03:29:51,267 TARGET COMPLEMENT IN THE 5416 03:29:51,267 --> 03:29:53,436 EXTRACELLULAR SPACE BECAUSE WE 5417 03:29:53,436 --> 03:29:54,904 WOULD ARGUE MOST OF YOU PROBABLY 5418 03:29:54,904 --> 03:29:57,507 THINK COMPLEMENT IS JUST A SERUM 5419 03:29:57,507 --> 03:30:00,243 BLOOD LIVER SYSTEM. 5420 03:30:00,243 --> 03:30:02,278 NOW, WHAT OUR COLLABORATORS IN 5421 03:30:02,278 --> 03:30:03,413 THE LAB BASICALLY HAVE 5422 03:30:03,413 --> 03:30:04,948 DISCOVERED IS THAT EVERY SINGLE 5423 03:30:04,948 --> 03:30:08,318 CELL IN YOUR BODY HAS ITS OWN 5424 03:30:08,318 --> 03:30:09,652 CELL AUTONOMOUS COMPLEMENT 5425 03:30:09,652 --> 03:30:11,921 SYSTEM, AND THAT COMPLEMENT 5426 03:30:11,921 --> 03:30:14,557 SYSTEM QUITE OFTEN CAN BE EITHER 5427 03:30:14,557 --> 03:30:18,628 SECRETED OR ALSO WORKS IN 5428 03:30:18,628 --> 03:30:19,596 INTRACELLULARLY, AND WITH THIS 5429 03:30:19,596 --> 03:30:23,466 NEW LOCATION OF COMPLEMENT, ITS 5430 03:30:23,466 --> 03:30:24,300 INTRACELLULAR VASCULAR ALSO 5431 03:30:24,300 --> 03:30:27,537 COMES NEW FUNCTIONAL NICHE. 5432 03:30:27,537 --> 03:30:29,706 WHAT CELL AUTONOMOUS OR 5433 03:30:29,706 --> 03:30:30,440 INTRACELLULAR COMPLEMENT IS 5434 03:30:30,440 --> 03:30:34,010 CONTROL A SINGLE CELL METABOLISM 5435 03:30:34,010 --> 03:30:35,512 NORMAL CELL PHYSIOLOGY AND ALSO 5436 03:30:35,512 --> 03:30:37,614 CELL SURVIVAL HOMOEOSTASIS AND 5437 03:30:37,614 --> 03:30:38,982 CELL EFFECTOR FUNCTION. 5438 03:30:38,982 --> 03:30:41,651 STANDOFFS DISCOVERED BY A REALLY 5439 03:30:41,651 --> 03:30:43,620 TALENTED POST-DOC IN MY LAB 5440 03:30:43,620 --> 03:30:46,389 MARTIN COLAR WHO FOUND THAT 5441 03:30:46,389 --> 03:30:48,825 CIRCULATING CD4 POSITIVE T CELLS 5442 03:30:48,825 --> 03:30:51,461 EXPRESS C3 AND ACTIVATE C3 IN 5443 03:30:51,461 --> 03:30:53,396 THE LYSOSOMES. 5444 03:30:53,396 --> 03:30:58,001 THE C38 PORTION OF ACTIVATED C3 5445 03:30:58,001 --> 03:31:00,470 ENGAGES A LYSOSOMAL C3A RECEPTOR 5446 03:31:00,470 --> 03:31:04,441 WHICH IS REALLY IMPORTANT FOR 5447 03:31:04,441 --> 03:31:06,943 CO-ACTIVATION AND REQUIRED FOR 5448 03:31:06,943 --> 03:31:08,411 CD4 POSITIVE T CELL SURVIVAL. 5449 03:31:08,411 --> 03:31:15,051 T CELLS THAT CANNOT PRODUCE C -- 5450 03:31:15,051 --> 03:31:16,586 C3 DIE. 5451 03:31:16,586 --> 03:31:18,254 WHAT MARTIN ALSO DELINEATED THAT 5452 03:31:18,254 --> 03:31:19,689 IN LINE WITH WHAT WE THINK ABOUT 5453 03:31:19,689 --> 03:31:21,191 COMPLEMENT TO BE A PROTECTOR 5454 03:31:21,191 --> 03:31:23,393 AGAINST INFECTIONS, THE SYSTEM 5455 03:31:23,393 --> 03:31:24,961 REALLY ALSO IS IMPORTANT FOR T 5456 03:31:24,961 --> 03:31:27,130 CELL RESPONSES. 5457 03:31:27,130 --> 03:31:29,098 DURING THE PRESENCE OF DANGER 5458 03:31:29,098 --> 03:31:31,067 HERE SIGNIFIED BY T CELL 5459 03:31:31,067 --> 03:31:33,570 RECEPTOR ENGAGEMENT THE C3B 5460 03:31:33,570 --> 03:31:35,505 PORTION GENERATED 5461 03:31:35,505 --> 03:31:37,273 INTRACELLULARLY FLOIPS THE 5462 03:31:37,273 --> 03:31:39,008 OUTSIDE OF THE DISPEL ENGAGES 5463 03:31:39,008 --> 03:31:42,745 ITS RECEPTOR C DP 46 AND SIGNAL 5464 03:31:42,745 --> 03:31:46,683 MEDIATED BY CD46 ABSOLUTELY 5465 03:31:46,683 --> 03:31:50,186 CRITICAL TO DRIVE GAMMA 5466 03:31:50,186 --> 03:31:52,889 PRODUCTION IN T CELLS AND THE UP 5467 03:31:52,889 --> 03:31:55,391 SELL SIGNAL IS NOT IS IMPORTANT 5468 03:31:55,391 --> 03:31:59,662 FOR TH1 OR TH17 RESPONSES. 5469 03:31:59,662 --> 03:32:01,764 MAY BE QUITE CRITICAL FOR 5470 03:32:01,764 --> 03:32:03,766 HEALTH, ALSO IMPORTANT FOR THE 5471 03:32:03,766 --> 03:32:05,602 TH1 SHUT DOWN PROGRAM WHICH IS 5472 03:32:05,602 --> 03:32:07,003 EQUALLY IMPORTANT FOR SURVIVAL 5473 03:32:07,003 --> 03:32:08,972 BECAUSE IF YOU DON'T HAVE TH1 5474 03:32:08,972 --> 03:32:12,375 RESPONSES YOU CAN'T -- 5475 03:32:12,375 --> 03:32:13,476 INFECTIONS BUT IF YOU CAN'T SHUT 5476 03:32:13,476 --> 03:32:15,211 THEM DOWN YOU REALLY DEVELOP 5477 03:32:15,211 --> 03:32:17,747 PROFOUND TISSUE PATH OOLG. 5478 03:32:17,747 --> 03:32:19,749 CD46 ALSO CONTRIBUTES TO THE TH1 5479 03:32:19,749 --> 03:32:21,451 SHUT DOWN PROGRAM BY INDUCING 5480 03:32:21,451 --> 03:32:23,453 THE CORE EXPRESSION OF 5481 03:32:23,453 --> 03:32:25,955 IMMUNOSUPPRESSIVE IL10 AND TH1 5482 03:32:25,955 --> 03:32:28,191 CELLS INTO A TH1 SHUT DOWN AND 5483 03:32:28,191 --> 03:32:29,926 PRERM PROGRAM. 5484 03:32:29,926 --> 03:32:30,960 -- MEMORY PROGRAM. 5485 03:32:30,960 --> 03:32:32,695 WE HAVE SHOWN USING PATIENTS 5486 03:32:32,695 --> 03:32:36,566 THAT IF YOU HAVE -- IF YOU HAVE 5487 03:32:36,566 --> 03:32:40,803 TOO LITTLE OF THE SYSTEM, DON'T 5488 03:32:40,803 --> 03:32:44,807 HAVE SUFFICIENT CD46 YOU SUFFER 5489 03:32:44,807 --> 03:32:47,143 FROM RECURRENT MOST UPPER 5490 03:32:47,143 --> 03:32:47,877 RESPIRATORY TRACT INFECTIONS, 5491 03:32:47,877 --> 03:32:49,812 BUT IF YOUR SYSTEM IS 5492 03:32:49,812 --> 03:32:50,613 DYSREGULATED AND YOU PRODUCE TOO 5493 03:32:50,613 --> 03:32:53,349 MUCH C3 AND YOU HAVE EN 5494 03:32:53,349 --> 03:32:56,185 HADGESSED CD46 OR UNTIMELY CD46 5495 03:32:56,185 --> 03:32:58,688 SIGNALLING, TH1 HYPERACTIVITY 5496 03:32:58,688 --> 03:33:00,557 INDUCED BY THIS PATHWAY 5497 03:33:00,557 --> 03:33:03,826 CONTRIBUTES TO AUTOIMMUNITY AND 5498 03:33:03,826 --> 03:33:06,462 MOSTLY SO FAR IN ARTHRITIC 5499 03:33:06,462 --> 03:33:06,896 DISEASES. 5500 03:33:06,896 --> 03:33:09,399 NOW, WHERE THE REAL EXCITING 5501 03:33:09,399 --> 03:33:10,967 PART FOR MY LAB CAME IN IS WHEN 5502 03:33:10,967 --> 03:33:13,570 WE TRIED TO FIGURE OUT HOW 5503 03:33:13,570 --> 03:33:17,707 EXACTLY IS CD46 INDUCING TH1 5504 03:33:17,707 --> 03:33:17,974 RESPONSES. 5505 03:33:17,974 --> 03:33:20,243 WHAT THEY FOUND IS SO FAR UNTIL 5506 03:33:20,243 --> 03:33:23,413 TODAY THAT CD46 INDUCES THREE 5507 03:33:23,413 --> 03:33:24,514 KEY EVENTS. 5508 03:33:24,514 --> 03:33:27,483 IT INDUCES DIRECTLY AND WE HAVE 5509 03:33:27,483 --> 03:33:30,086 NOW TO THANK THE MECHANISM BUT 5510 03:33:30,086 --> 03:33:31,087 I'M TALKING ABOUT THIS TODAY, 5511 03:33:31,087 --> 03:33:33,389 THE EXPRESSION OF GENES THAT 5512 03:33:33,389 --> 03:33:34,891 ENCODE NUTRIENT TRANSPORTERS 5513 03:33:34,891 --> 03:33:36,559 THAT ALLOW GLUCOSE AND AMINO 5514 03:33:36,559 --> 03:33:37,727 ACIDS TO COME IN BASICALLY FOOD 5515 03:33:37,727 --> 03:33:39,462 FOR TH1 INDUCTION. 5516 03:33:39,462 --> 03:33:44,534 AT THE SAME TIME, CD46 SHUNS THE 5517 03:33:44,534 --> 03:33:45,969 COMPLEX LYSOSOME WHICH NOW 5518 03:33:45,969 --> 03:33:47,503 ALLOWS THE HIGH LEVELS OF 5519 03:33:47,503 --> 03:33:51,007 GLYCOLYSIS AND OXIDATIVE 5520 03:33:51,007 --> 03:33:58,014 PHOSPHORYLATION PLRSZ L AND 5521 03:33:58,014 --> 03:34:00,316 YEAH, THERE'S ALSO INTRACELLULAR 5522 03:34:00,316 --> 03:34:02,719 C5 SYSTEM, IT IS CONTROLLED BY 5523 03:34:02,719 --> 03:34:06,322 CD46 AND IT INDUCES VIA 5524 03:34:06,322 --> 03:34:08,925 MITOCHONDRIAL RECEPTOR ONE 5525 03:34:08,925 --> 03:34:15,164 SIGNALLING ECONOMICAL AND LP3 -- 5526 03:34:15,164 --> 03:34:19,135 ILM LYNN DEDUCTION THAT DOES NOT 5527 03:34:19,135 --> 03:34:20,003 INDUCT -- RATHER SUSTAINS 5528 03:34:20,003 --> 03:34:22,005 RESPONSES AT MUCOSAL SITES. 5529 03:34:22,005 --> 03:34:24,073 SIMILARLY THE TH1 SHUT DOWN 5530 03:34:24,073 --> 03:34:27,343 PROGRAM IS ALSO APEX REGULATED 5531 03:34:27,343 --> 03:34:30,079 BY CD46 BY A METABOLIC 5532 03:34:30,079 --> 03:34:31,014 REPROGRAMING AND HERE INVOLVING 5533 03:34:31,014 --> 03:34:35,118 A LOT OF CHOLESTEROL METABOLISM. 5534 03:34:35,118 --> 03:34:37,186 SO WHEN I MOVED MY LAB WE HAD 5535 03:34:37,186 --> 03:34:39,155 DISCOVERED THIS SYSTEM IN T 5536 03:34:39,155 --> 03:34:41,157 CELLS AND WE STARTED WORKING ON 5537 03:34:41,157 --> 03:34:43,559 THIS SYSTEM MORE BRAWLD HERE AT 5538 03:34:43,559 --> 03:34:43,793 THE NIH. 5539 03:34:43,793 --> 03:34:46,663 ONE OF THE QUESTIONS THEN WAS 5540 03:34:46,663 --> 03:34:47,730 OKAY, WE HAVE FOUND IT IN T 5541 03:34:47,730 --> 03:34:50,500 CELLS H IT'S IMPORTANT FOR MODEL 5542 03:34:50,500 --> 03:34:51,701 T CELL BEHAVIOR. 5543 03:34:51,701 --> 03:34:53,803 IF IT MISBEHAVES IT CONTRIBUTES 5544 03:34:53,803 --> 03:34:56,572 TO TH1 MEDIATED AUTOIMMUNE 5545 03:34:56,572 --> 03:34:59,208 DISEASE OR OTHER DISEASES. 5546 03:34:59,208 --> 03:35:01,177 DIST SYSTEM ALSO EXIST -- DOES 5547 03:35:01,177 --> 03:35:02,612 THE SYSTEM ALSO EXIST IN OTHER 5548 03:35:02,612 --> 03:35:03,312 IMMUNE CELLS? 5549 03:35:03,312 --> 03:35:04,881 I'M JUST GIVING YOU A BROAD 5550 03:35:04,881 --> 03:35:06,716 OVERVIEW HERE, BUT YES, IT DOES, 5551 03:35:06,716 --> 03:35:11,154 IN THE VERY IMPORTANT MONOCYTE 5552 03:35:11,154 --> 03:35:16,392 MARKER NEUTROPHIL LINEAGE NUTD 5553 03:35:16,392 --> 03:35:17,960 NEUTROPHIL LINEAGES, REALLY 5554 03:35:17,960 --> 03:35:20,363 IMPORTANT FOR NORMAL FUNCTION. 5555 03:35:20,363 --> 03:35:22,999 IF THE SYSTEM IS HYPERACTIVE, A 5556 03:35:22,999 --> 03:35:25,935 LOT OF COLLABORATORS HERE AT THE 5557 03:35:25,935 --> 03:35:29,405 NIH HAVE CARVED THIS OUT, 5558 03:35:29,405 --> 03:35:30,306 HYPERACTIVE MACROPHAGES 5559 03:35:30,306 --> 03:35:31,107 CONTRIBUTE TO DISEASES AND WE 5560 03:35:31,107 --> 03:35:34,977 HAVE SHOWN IT CONTRIBUTES TO 5561 03:35:34,977 --> 03:35:36,412 ATHEROSCHER HOSES MR. TORE THE 5562 03:35:36,412 --> 03:35:41,117 INABILITY OF PERCEPTIVE PATIENTS 5563 03:35:41,117 --> 03:35:42,785 TO CLEAR CANDY DAVMENT THIS IS A 5564 03:35:42,785 --> 03:35:44,220 LITTLE BIT OF A TIDBIT HERE 5565 03:35:44,220 --> 03:35:46,255 TODAY FOR UNPUBLISHED DATD. 5566 03:35:46,255 --> 03:35:49,258 WHAT WE HAVE DONE IS ALSO UP 5567 03:35:49,258 --> 03:35:51,561 REGULATED C3 AND C5 IN HUMAN AND 5568 03:35:51,561 --> 03:35:53,730 MOUSE B CELLS AND WHAT WE 5569 03:35:53,730 --> 03:35:57,834 NOTICED UNEXPECTEDLY IS THAT THE 5570 03:35:57,834 --> 03:36:00,002 ANTIBODY PRODUCTION IN THESE 5571 03:36:00,002 --> 03:36:01,971 CELLS THAT CANNOT PRODUCE C5 IN 5572 03:36:01,971 --> 03:36:05,108 AN OTHERWISE C5 SUFFER HOST IS 5573 03:36:05,108 --> 03:36:05,374 INCREASED. 5574 03:36:05,374 --> 03:36:07,410 THAT IS REALLY, REALLY 5575 03:36:07,410 --> 03:36:08,377 UNEXPECTED, AND THEY'RE TRYING 5576 03:36:08,377 --> 03:36:12,014 TO FIGURE OUT NOW WHY THAT IS, 5577 03:36:12,014 --> 03:36:13,616 AND THE ANTIBODY RESPONSE IS 5578 03:36:13,616 --> 03:36:15,284 INCREASED IN T CELL AND 5579 03:36:15,284 --> 03:36:18,354 INDEPENDENT RESPONSES, AND 5580 03:36:18,354 --> 03:36:21,524 CURRENTLY SEEING RNA SEQ TOLL 5581 03:36:21,524 --> 03:36:28,965 FIND OUT HOW CELL AUTONOMOUS 5582 03:36:28,965 --> 03:36:31,701 FLRMSZ DRL C5. 5583 03:36:31,701 --> 03:36:32,802 THE INCOMES BIG QUESTION WE 5584 03:36:32,802 --> 03:36:35,304 TRIED TO TACKLE IN THE LAB WAS, 5585 03:36:35,304 --> 03:36:36,873 IS THE INTRACELLULAR COMPLEMENT 5586 03:36:36,873 --> 03:36:41,544 SYSTEM WHICH WE COINED THE 5587 03:36:41,544 --> 03:36:43,279 COMPLOSOME ALSO IMPORTANT FOR 5588 03:36:43,279 --> 03:36:44,480 NONIMMUNE CELLS, MEANING FOR 5589 03:36:44,480 --> 03:36:45,381 STRUCTURAL CELLS? 5590 03:36:45,381 --> 03:36:49,452 DURING COVID, A LOT OF NAIAD 5591 03:36:49,452 --> 03:36:51,187 PEOPLE HELPED US DEFINE THAT 5592 03:36:51,187 --> 03:36:53,489 CELL AUTONOMOUS C3 IN TYPE TWO 5593 03:36:53,489 --> 03:36:56,526 LUNG EPITHELIAL CELLS IS KICKED 5594 03:36:56,526 --> 03:36:58,928 ON BY SARS-COV-2 AND THEN 5595 03:36:58,928 --> 03:37:01,798 SECRETED AND THAT SECRETED C3A 5596 03:37:01,798 --> 03:37:04,300 LOCALLY REALLY CONTRIBUTES TO 5597 03:37:04,300 --> 03:37:06,202 THE LOCAL PROINFLAMMATORY 5598 03:37:06,202 --> 03:37:07,603 COMPLEMENT DURING RESPONSE THAT 5599 03:37:07,603 --> 03:37:09,338 PATIENTS THAT TRANSITION INTO 5600 03:37:09,338 --> 03:37:11,974 SEVERE COVID ARE EXPERIENCING. 5601 03:37:11,974 --> 03:37:13,409 WE'RE ACTUALLY WORKING RIGHT NOW 5602 03:37:13,409 --> 03:37:15,912 ON TACKLING THIS AXIS. 5603 03:37:15,912 --> 03:37:18,414 NOW, A LOT OF OTHER LABS NOW ARE 5604 03:37:18,414 --> 03:37:20,383 WORKING ON THIS AND WHAT WE NOW 5605 03:37:20,383 --> 03:37:22,485 KNOW IS THAT WHEREVER YOU LOOK, 5606 03:37:22,485 --> 03:37:24,554 BASICALLY EVERY CELL THAT GROUPS 5607 03:37:24,554 --> 03:37:26,756 ACROSS THE WORLD LOOKED AT 5608 03:37:26,756 --> 03:37:30,226 INTRACELLULAR COMPLEMENT IS 5609 03:37:30,226 --> 03:37:31,460 PRESENT. 5610 03:37:31,460 --> 03:37:34,831 HAD BETA CELLS, EPITHELIAL CELLS 5611 03:37:34,831 --> 03:37:37,099 AND HYDROBLASTS AND FOR EACH OF 5612 03:37:37,099 --> 03:37:38,868 THESE CELL TYPES CELL AUTONOMOUS 5613 03:37:38,868 --> 03:37:42,572 CELL TYPES IS REALLY FINANCIAL 5614 03:37:42,572 --> 03:37:44,140 CORRESPONDENT FOR HOMEOSTATIC 5615 03:37:44,140 --> 03:37:46,976 FUNCTION ASK ALSO CONTRIBUTES 5616 03:37:46,976 --> 03:37:48,277 DYSREGULATED LIKE T CELL 5617 03:37:48,277 --> 03:37:57,720 RESPONSES TO DISEASES,, THAT THE 5618 03:37:57,720 --> 03:37:59,121 COMBLOSOME IS PRESENT IN ALL THE 5619 03:37:59,121 --> 03:38:00,456 CELLS, REALLY WE LOOKED AT SO 5620 03:38:00,456 --> 03:38:03,125 FAR, AND THAT CELL AUTONOMOUS 5621 03:38:03,125 --> 03:38:04,994 COMPLEMENT IS ALSO A KEY FEATURE 5622 03:38:04,994 --> 03:38:06,462 WHEN NEW CELLS ENTER THE TISSUE. 5623 03:38:06,462 --> 03:38:07,530 I DON'T HAVE TIME TO SHOW YOU 5624 03:38:07,530 --> 03:38:08,931 THIS DATA, BUT WE HAVE SHOWN 5625 03:38:08,931 --> 03:38:11,567 THAT P THE INDUCTION OF THE CELL 5626 03:38:11,567 --> 03:38:12,468 AUTONOMOUS COMPLEMENT SYSTEM IS 5627 03:38:12,468 --> 03:38:14,570 AN ABSOLUTELY KEY CHARACTERISTIC 5628 03:38:14,570 --> 03:38:17,206 HAD OF TISSUE RESIDENT IMMUNE 5629 03:38:17,206 --> 03:38:18,140 CELLS. 5630 03:38:18,140 --> 03:38:19,709 NOW, JUST A LITTLE BENEFIT A 5631 03:38:19,709 --> 03:38:20,543 SUMMARY SLIDE. 5632 03:38:20,543 --> 03:38:21,944 WHENEVER WE OR OTHER GROUPS 5633 03:38:21,944 --> 03:38:25,414 CARVED AND LOOKED HOW A CELL 5634 03:38:25,414 --> 03:38:26,649 AUTONOMOUS COMPLEMENT CROAMG THE 5635 03:38:26,649 --> 03:38:29,485 FUNCTION ACROSS ALL OF THESE 5636 03:38:29,485 --> 03:38:31,220 CELLS, I'M COMING RIGHT BACK TO 5637 03:38:31,220 --> 03:38:33,756 THE TITLE OF MY TALK. 5638 03:38:33,756 --> 03:38:37,493 IT IS ALL SINGLE CELL 5639 03:38:37,493 --> 03:38:37,793 PHYSIOLOGY. 5640 03:38:37,793 --> 03:38:38,594 SO CONTRIBUTES TO EVERYTHING YOU 5641 03:38:38,594 --> 03:38:42,632 CAN THINK OF, CHOLESTEROL PRRKS 5642 03:38:42,632 --> 03:38:46,135 GENE DIRECT REGULATION SH 5643 03:38:46,135 --> 03:38:47,570 MITOCHONDRIAL ELECTRON TRANSPORT 5644 03:38:47,570 --> 03:38:49,071 CHAIN DIRECTION SO IT'S A REAL 5645 03:38:49,071 --> 03:38:51,607 NEW CONTROLLER OF CELL 5646 03:38:51,607 --> 03:38:51,908 PHYSIOLOGY. 5647 03:38:51,908 --> 03:38:52,942 AND BECAUSE WE'RE YOUR GUEST 5648 03:38:52,942 --> 03:38:56,145 TODAY FOR NCI, I WANTED TO JUST 5649 03:38:56,145 --> 03:38:57,613 GIVE A LITTLE CARROT AND SHARE 5650 03:38:57,613 --> 03:38:58,814 WITH YOU THAT WHAT WE HAVE FOUND 5651 03:38:58,814 --> 03:39:00,583 REALLY, AND THIS IS YOU SHOULD 5652 03:39:00,583 --> 03:39:01,784 UNDER SUBMISSION NOW, WE HAVE 5653 03:39:01,784 --> 03:39:04,820 FOUND THAT TONICALLY ACTIVE C3 5654 03:39:04,820 --> 03:39:07,957 HAVE I AUTO OM MYC SIGNALLING OF 5655 03:39:07,957 --> 03:39:11,093 THE C3A RECEPTOR IN HUMAN 5656 03:39:11,093 --> 03:39:13,829 EPITHELIAL CELLS IS ABSOLUTELY 5657 03:39:13,829 --> 03:39:15,564 CRITICAL TO STATIC TURN OF 5658 03:39:15,564 --> 03:39:16,465 KEEPING THESE CELLS GOING. 5659 03:39:16,465 --> 03:39:18,968 IF YOU TAKE THE C3 SYSTEM OUT, 5660 03:39:18,968 --> 03:39:23,406 THESE CELLS MOVE INTRAMALIGNANT 5661 03:39:23,406 --> 03:39:24,206 TRANCE FOR MAIPTION AND THE 5662 03:39:24,206 --> 03:39:26,075 PATHWAY IS SOME CONTROL OF SOME 5663 03:39:26,075 --> 03:39:28,477 REALLY INTERESTING ONCOGEN AND 5664 03:39:28,477 --> 03:39:30,680 RIBOSOMAL BIOGENESIS PATHWAYS. 5665 03:39:30,680 --> 03:39:32,214 SO I HOPE I HAVE CONVINCED YOU 5666 03:39:32,214 --> 03:39:34,283 THAT INTRACELLULAR COMPLEMENT 5667 03:39:34,283 --> 03:39:36,152 REALLY IS IMPORTANT. 5668 03:39:36,152 --> 03:39:37,887 WHAT I HAVEN'T TALKED WITH YOU 5669 03:39:37,887 --> 03:39:43,526 ABOUT YET IS THAT IT IS ALSO -- 5670 03:39:43,526 --> 03:39:46,095 TO FORMALLY A PHARMACOLOGICAL 5671 03:39:46,095 --> 03:39:46,529 INTERVENTION. 5672 03:39:46,529 --> 03:39:48,197 WHILE WE'VE BEEN SUCCESSFUL IN 5673 03:39:48,197 --> 03:39:50,466 TAKING T CELLS OUT OR MONOCYTES 5674 03:39:50,466 --> 03:39:52,535 AND MACROPHAGES FROM SITES OF 5675 03:39:52,535 --> 03:39:54,637 HUMAN INFLAMMATION LIKE INFLAMED 5676 03:39:54,637 --> 03:39:55,604 JOINT FROM PATIENTS WITH 5677 03:39:55,604 --> 03:39:58,007 ARTHRORIGHTS AND WE HAVE BEEN 5678 03:39:58,007 --> 03:40:01,310 ABLE TO ANALYZE HYPERAK -- 5679 03:40:01,310 --> 03:40:03,279 NORMALIZE RESPONSES DRIVEN BY C3 5680 03:40:03,279 --> 03:40:07,984 OR C5 ACTIVATION, EX-VIVO, WITH 5681 03:40:07,984 --> 03:40:09,118 A CELL COMPLEMENT INHIBITOR 5682 03:40:09,118 --> 03:40:13,789 ACCIDENT DOING THIS TOGETHER. 5683 03:40:13,789 --> 03:40:16,325 ARE WE ARE NOW ALSO WORKING ON 5684 03:40:16,325 --> 03:40:19,795 TESTING WITH APELLIS FIRST 5685 03:40:19,795 --> 03:40:22,732 CLINICAL MOUSE NODLE 5686 03:40:22,732 --> 03:40:23,499 INTRACELLULAR COMPLEMENT IN 5687 03:40:23,499 --> 03:40:24,500 ARTHRITIS AND IN CANCER. 5688 03:40:24,500 --> 03:40:26,002 SO WHAT I WOULD LIKE TO ARGUE 5689 03:40:26,002 --> 03:40:30,840 HERE ALSO IS THAT BECAUSE C3 AND 5690 03:40:30,840 --> 03:40:33,509 C5 HAS A FUNCTION IN SERUM, IN 5691 03:40:33,509 --> 03:40:35,644 THE CIRCULATION, AND IN CELLS, 5692 03:40:35,644 --> 03:40:38,414 AND THE FUNCTION IS DIFFERENT IN 5693 03:40:38,414 --> 03:40:39,648 CELL SPECIFIC, IF YOU ARE 5694 03:40:39,648 --> 03:40:41,150 INTERESTED IN STUDYING THIS 5695 03:40:41,150 --> 03:40:43,919 SYSTEM, I REALLY, REALLY URGE 5696 03:40:43,919 --> 03:40:45,955 YOU TO USE FLOXED AND CELL 5697 03:40:45,955 --> 03:40:47,156 SPECIFIC KNOCK-OUT MICE. 5698 03:40:47,156 --> 03:40:48,858 WE ARE MOVING AWAY IN OUR FIELD 5699 03:40:48,858 --> 03:40:51,127 FROM USE HAD GONE GLOBAL MICE 5700 03:40:51,127 --> 03:40:52,728 BECAUSE THEY MASK PHENOTYPES AND 5701 03:40:52,728 --> 03:40:54,964 DO NOT REALLY GIVE YOU AN ANSWER 5702 03:40:54,964 --> 03:40:56,399 WITH REGARDS TO MECHANISMS THAT 5703 03:40:56,399 --> 03:40:58,367 YOU MIGHT BE INTERESTED IN. 5704 03:40:58,367 --> 03:41:01,237 THE SECOND THING AI WOULD ALSO 5705 03:41:01,237 --> 03:41:05,207 SAY IS, I WOULD BET YOU A VERY 5706 03:41:05,207 --> 03:41:07,710 GOOD BOTTLE THAT IF YOU DIG INTO 5707 03:41:07,710 --> 03:41:09,578 YOUR SINGLE CELL RNA SEQ DATA 5708 03:41:09,578 --> 03:41:11,047 YOU WILL SEE THE COMPLEMENT 5709 03:41:11,047 --> 03:41:12,548 SYSTEM AND I ALSO BET YOU THAT 5710 03:41:12,548 --> 03:41:14,717 YOU PROBABLY IGNORED THAT 5711 03:41:14,717 --> 03:41:15,684 BECAUSE COMPLEMENT SHOULDN'T A, 5712 03:41:15,684 --> 03:41:17,219 BE THERE, AND SECOND, NOBODY 5713 03:41:17,219 --> 03:41:18,521 LIKES THE SYSTEM. 5714 03:41:18,521 --> 03:41:20,356 BUT IF YOU ARE INTERESTED, IT'S 5715 03:41:20,356 --> 03:41:21,924 IN YOUR TOP PATHWAYS, PLEASE 5716 03:41:21,924 --> 03:41:22,892 COME AND TALK TO US. 5717 03:41:22,892 --> 03:41:24,760 WE THINK IT'S A REALLY, REALLY 5718 03:41:24,760 --> 03:41:26,295 COOL SYSTEM, AND WE THINK IT'S 5719 03:41:26,295 --> 03:41:28,464 BY NO MEANS FULLY EXPLORED. 5720 03:41:28,464 --> 03:41:29,899 AND WE ALSO THINK THIS GIVES 5721 03:41:29,899 --> 03:41:32,301 U.S. THE OPPORTUNITY NOW TO 5722 03:41:32,301 --> 03:41:35,137 RETHINK THIS SYSTEM IN HUMAN 5723 03:41:35,137 --> 03:41:38,307 DISEASE BUT ALSO ULTIMATELY FOR 5724 03:41:38,307 --> 03:41:38,974 THERAPEUTIC INTERVENTION BECAUSE 5725 03:41:38,974 --> 03:41:40,443 WE THINK WE NOW NEED TO START 5726 03:41:40,443 --> 03:41:43,245 THINKING ABOUT TACKLING THE 5727 03:41:43,245 --> 03:41:44,013 INTRA AND EXTRACELLULAR 5728 03:41:44,013 --> 03:41:44,780 COMPLEMENT SYSTEM TO REALLY, 5729 03:41:44,780 --> 03:41:50,820 REALLY HAVE A A GO AT ARE 5730 03:41:50,820 --> 03:41:55,057 BROADENING DISEASES. 5731 03:41:55,057 --> 03:42:05,234 [APPLAUSE] 5732 03:42:08,504 --> 03:42:09,939 >> THANK YOU VERY MUCH. 5733 03:42:09,939 --> 03:42:11,373 THE LITTLE I KNOW ABOUT 5734 03:42:11,373 --> 03:42:12,741 COMPLEMENT I THINK THAT C3 HAS 5735 03:42:12,741 --> 03:42:13,776 TO BE CLEAVED BY SOMETHING, I 5736 03:42:13,776 --> 03:42:16,078 KNOW THERE'S SOME SPONTANEOUS 5737 03:42:16,078 --> 03:42:18,380 CLEAVAGE AND THEN C5 HAS TO BE 5738 03:42:18,380 --> 03:42:18,614 CLEAVED. 5739 03:42:18,614 --> 03:42:22,218 WHAT'S UPSTREAM OF C3 AND HOW DO 5740 03:42:22,218 --> 03:42:24,186 ACIDIC COMPARTMENTS IN THE CELLS 5741 03:42:24,186 --> 03:42:25,921 SOMEHOW PERHAPS CONTRIBUTE TO 5742 03:42:25,921 --> 03:42:27,256 SPONTANEOUS CLEAVAGE? 5743 03:42:27,256 --> 03:42:28,791 >> THAT'S A FANTASTIC QUESTION. 5744 03:42:28,791 --> 03:42:30,793 SO IN SERUM IT IS RELATIVELY 5745 03:42:30,793 --> 03:42:33,929 ESTABLISHED THAT YOU NEED IT A 5746 03:42:33,929 --> 03:42:34,864 COMPLEX OF THREE DIFFERENT 5747 03:42:34,864 --> 03:42:37,099 PROTEINS TO ACTIVATE C3. 5748 03:42:37,099 --> 03:42:38,701 WHAT WE HAVE SHOWN DPINLTD HAVE 5749 03:42:38,701 --> 03:42:42,171 TIME TO REALLY DISSECT HERE WAS 5750 03:42:42,171 --> 03:42:46,008 IN CELLS SINGLE AND EVOLUTIONARY 5751 03:42:46,008 --> 03:42:48,611 PROTEASES CAN ACTUALLY DLEEF C3 5752 03:42:48,611 --> 03:42:49,278 DIRECTLY. 5753 03:42:49,278 --> 03:42:50,479 NOW, 24-S REALLY INTERESTING 5754 03:42:50,479 --> 03:42:54,216 BECAUSE SPECIFICALLY KINAPSIN-L 5755 03:42:54,216 --> 03:42:56,519 CAN WORK IN ACIDIC AND IN 5756 03:42:56,519 --> 03:42:57,920 NEUTRAL PH SETTINGS SO THAT 5757 03:42:57,920 --> 03:42:59,255 ACTUALLY WORKS FOR US SO THAT 5758 03:42:59,255 --> 03:43:00,789 YOU HAVE DIFFERENT COMPARTMENTS 5759 03:43:00,789 --> 03:43:04,093 IN A CELL WHERE A PH AND AN OLD 5760 03:43:04,093 --> 03:43:06,162 PROTEASE IS CONDUCIVE TO CLEAVE 5761 03:43:06,162 --> 03:43:11,767 C3 NOW. 5762 03:43:11,767 --> 03:43:14,270 >> GREAT TALK, CLAUDIA. 5763 03:43:14,270 --> 03:43:17,239 SO HEAD TROA TRIMATO-G-PROTEINS 5764 03:43:17,239 --> 03:43:19,175 ARE THEY INVOLVED IN THE 5765 03:43:19,175 --> 03:43:20,943 INTRACELLULAR COMPLEMENT SYSTEM? 5766 03:43:20,943 --> 03:43:23,879 >> WHAT WAS THE WORD BEFORE 5767 03:43:23,879 --> 03:43:24,146 G-PROTEIN? 5768 03:43:24,146 --> 03:43:27,316 GROO HETRO -- 5769 03:43:27,316 --> 03:43:31,420 >> HETRATROMA ATIC. 5770 03:43:31,420 --> 03:43:33,556 >> I THINK YOU'RE TROAMPG THE 5771 03:43:33,556 --> 03:43:35,090 G-PROTEIN SECTORS CAN PAIR AND 5772 03:43:35,090 --> 03:43:39,595 NOT JUST DIMERIC BUT ALSO 5773 03:43:39,595 --> 03:43:39,895 MULTI-MERIC. 5774 03:43:39,895 --> 03:43:42,831 >> ALL YOUR RECEPTORS AS YOU SAY 5775 03:43:42,831 --> 03:43:44,833 ARE RL COMPLEMENT SO INSIDE THE 5776 03:43:44,833 --> 03:43:47,336 RECEPTOR ARE THE BINDING 5777 03:43:47,336 --> 03:43:48,537 G-PROTEINS, BIENGSD SITES FOR 5778 03:43:48,537 --> 03:43:49,071 THOSE G-PROTEINS. 5779 03:43:49,071 --> 03:43:51,140 >> YES, THEY TORQUES THEY ACTUAL 5780 03:43:51,140 --> 03:43:52,675 EVEN BIND THEM INSPECT 5781 03:43:52,675 --> 03:43:54,543 MITOCHONDRIA, AND MITOCHONDRIAL 5782 03:43:54,543 --> 03:43:55,311 SPACE WE HAVE SHOWN AND 5783 03:43:55,311 --> 03:43:56,078 PUBLISHED THAT. 5784 03:43:56,078 --> 03:43:57,479 THEY ALSO PAIR WITH OTHER 5785 03:43:57,479 --> 03:43:59,048 G-PROTEIN COUPLED RECEPTORS 5786 03:43:59,048 --> 03:44:00,449 WHICH MAKES THE WHOLE SITUATION 5787 03:44:00,449 --> 03:44:01,350 REALLY QUITE COMPLEX. 5788 03:44:01,350 --> 03:44:01,951 SO YES AND YES. 5789 03:44:01,951 --> 03:44:04,486 >> SO IT'S THE G-PROTEINS THAT 5790 03:44:04,486 --> 03:44:06,255 ARE GRIEPTING THE SIGNAL THROUGH 5791 03:44:06,255 --> 03:44:07,122 CLASSIC MECHANISMS? 5792 03:44:07,122 --> 03:44:08,357 >> YEAH. 5793 03:44:08,357 --> 03:44:08,557 YES. 5794 03:44:08,557 --> 03:44:10,526 THROUGH ERG SIGNALLING AND 5795 03:44:10,526 --> 03:44:11,660 PHOSPHORYLATION IN THE 5796 03:44:11,660 --> 03:44:15,798 MITOCHONDRIA. 5797 03:44:15,798 --> 03:44:17,032 YEAH. 5798 03:44:17,032 --> 03:44:17,766 >> HI. 5799 03:44:17,766 --> 03:44:20,703 SO I HAVE TWO QUESTIONS THAT ARE 5800 03:44:20,703 --> 03:44:22,238 RELATED <!680> ONE IS WHAT IS AN 5801 03:44:22,238 --> 03:44:26,508 EXAMPLE OF A DISEASE THAT IS 5802 03:44:26,508 --> 03:44:28,143 CAUSED ONLY BY -- COMPLEMENT AND 5803 03:44:28,143 --> 03:44:29,478 THEN SECOND HOW WOULD YOU PLAN 5804 03:44:29,478 --> 03:44:32,748 TO TARGET IT WITHOUT EFFECTIVE 5805 03:44:32,748 --> 03:44:33,782 EXTRACELLULAR COMPLEMENT? 5806 03:44:33,782 --> 03:44:34,917 >> TWO GOOD QUESTIONS. 5807 03:44:34,917 --> 03:44:37,186 ONE DISEASE IS REALLY TRULY, IF 5808 03:44:37,186 --> 03:44:39,455 YOU CANNOT EXPRESS CD46 OR HAVE 5809 03:44:39,455 --> 03:44:41,657 ISSUES WITH C3 SECRETION, THESE 5810 03:44:41,657 --> 03:44:43,626 PATIENTS ARE REALLY PROFOUNDLY 5811 03:44:43,626 --> 03:44:45,594 DEFECTIVE IN TH1 RESPONSES, SO 5812 03:44:45,594 --> 03:44:46,895 YOU REALLY HAVE A CONNECTION 5813 03:44:46,895 --> 03:44:49,632 BETWEEN TH1 BIOLOGY AND THE 5814 03:44:49,632 --> 03:44:51,734 FAULTY CELL AUTONOMOUS 5815 03:44:51,734 --> 03:44:55,004 COMPLEMENT SYSTEM BECAUSE SERUM 5816 03:44:55,004 --> 03:44:56,972 C3 CANNOT COMPENSATE FOR CELL 5817 03:44:56,972 --> 03:44:58,941 AUTONOMOUS C3, ONE. 5818 03:44:58,941 --> 03:45:00,609 B, IT'S REALLY THE PAPER THAT WE 5819 03:45:00,609 --> 03:45:02,578 ARE SUBMITTING RIGHT NOW, VERY 5820 03:45:02,578 --> 03:45:03,679 SURPRISINGLY THAT IF YOU TAKE 5821 03:45:03,679 --> 03:45:07,049 OUT C3 FROM CERTAIN EPITHELIAL 5822 03:45:07,049 --> 03:45:09,218 CELLS, THESE CELLS MOST 5823 03:45:09,218 --> 03:45:10,786 SPONTANEOUSLY ENTER HYPERPLASIA 5824 03:45:10,786 --> 03:45:12,187 AND MA LIG NANLTD 5825 03:45:12,187 --> 03:45:12,554 TRANSFORMATION. 5826 03:45:12,554 --> 03:45:15,491 SO IT'S CANCER AND INFECTION -- 5827 03:45:15,491 --> 03:45:16,125 MALIGNANT TRANSFORMATION. 5828 03:45:16,125 --> 03:45:16,892 SO IT'S CANCER AND INFECTION 5829 03:45:16,892 --> 03:45:17,226 RIGHT NOW. 5830 03:45:17,226 --> 03:45:19,428 >> SO IT'S NOT A DIRECT 5831 03:45:19,428 --> 03:45:20,429 TARGETING OF THE COMPLEMENT 5832 03:45:20,429 --> 03:45:21,697 IFTSZ ITSELF? 5833 03:45:21,697 --> 03:45:22,865 >> THIS VALLEY A BEAST. 5834 03:45:22,865 --> 03:45:25,000 EVERYBODY KNOWS THE STOP 5835 03:45:25,000 --> 03:45:26,135 INHIBITORS IN CANCER IS THE 5836 03:45:26,135 --> 03:45:27,603 PROBLEM, SO WHAT WE'RE TRYING TO 5837 03:45:27,603 --> 03:45:29,405 DO AS A PROGRAM IS TRYING TO 5838 03:45:29,405 --> 03:45:32,041 UNDERSTAND A LITTLE BET HOW THE 5839 03:45:32,041 --> 03:45:33,676 CELL AUTONOMOUS COMPLEMENT 5840 03:45:33,676 --> 03:45:35,210 SYSTEM CONTROLLED FROM THE CELL 5841 03:45:35,210 --> 03:45:36,979 SURVEYS SO WE MAYBE HAVE EVEN 5842 03:45:36,979 --> 03:45:37,613 EASIER TARGET. 5843 03:45:37,613 --> 03:45:38,947 WE'RE COMING HERE WITH A 5844 03:45:38,947 --> 03:45:40,349 HOLISTIC BRAM WE LOOK WHAT ARE 5845 03:45:40,349 --> 03:45:41,450 THE RECEPTORS THAT DRIVE THE 5846 03:45:41,450 --> 03:45:46,121 SYSTEM AND WE KNOW IT'S 5847 03:45:46,121 --> 03:45:46,855 INTEGRANS LFA1 WE HAVE PUBLISHED 5848 03:45:46,855 --> 03:45:47,823 THAT AND ALSO AT THE SAME TIME 5849 03:45:47,823 --> 03:45:57,366 WE ARE TRYING TO UNDERSTAND WHAE 5850 03:45:57,366 --> 03:45:57,633 DIFFICULT. 5851 03:45:57,633 --> 03:45:57,966 >> THANK YOU. 5852 03:45:57,966 --> 03:45:59,234 >> I ACKNOWLEDGE THAT. 5853 03:45:59,234 --> 03:45:59,768 >> THANK YOU. 5854 03:45:59,768 --> 03:46:02,971 >> THANK YOU VERY MUCH. 5855 03:46:02,971 --> 03:46:09,945 [APPLAUSE] 5856 03:46:09,945 --> 03:46:13,148 >> SO OUR NEXT SPEAKER IS THE 5857 03:46:13,148 --> 03:46:15,417 INCREASINGLY FAY FAMOUS SHRUTI 5858 03:46:15,417 --> 03:46:18,287 NAIK FROM NEW YORK UNIVERSITY, 5859 03:46:18,287 --> 03:46:21,390 HE'LL BE TELLING US ABOUT 5860 03:46:21,390 --> 03:46:22,458 IMMUNE-MEDIATED MECHANISMS OF 5861 03:46:22,458 --> 03:46:24,326 ADAPTIVE AND MALADAPTIVE TISSUE 5862 03:46:24,326 --> 03:46:24,593 RESPONSES. 5863 03:46:24,593 --> 03:46:26,395 LET ME ASK THAT WHEN WE ASK 5864 03:46:26,395 --> 03:46:27,529 REQUESTS AT THE END, PERHAPS IF 5865 03:46:27,529 --> 03:46:30,666 THE QUESTIONER WOULD JUST 5866 03:46:30,666 --> 03:46:31,967 INTRODUCE THEMSELVES, IT MIGHT 5867 03:46:31,967 --> 03:46:34,670 BE HELPFUL FOR ALL OF US, THANK 5868 03:46:34,670 --> 03:46:37,206 YOU. 5869 03:46:37,206 --> 03:46:37,439 WELCOME. 5870 03:46:37,439 --> 03:46:40,909 >> THANKS SO MUCH, AL. 5871 03:46:40,909 --> 03:46:41,677 REALLY WE CAN TALK ABOUT 5872 03:46:41,677 --> 03:46:42,644 ANYTHING AND IT'S GOING TO BE 5873 03:46:42,644 --> 03:46:45,514 KIND OF A GRAB BAG, YOU GET A 5874 03:46:45,514 --> 03:46:46,749 CHOCOLATE BOX AND YOU DON'T KNOW 5875 03:46:46,749 --> 03:46:47,850 WHAT YOU'RE GETTING, RIGHT? 5876 03:46:47,850 --> 03:46:48,484 ALL RIGHT. 5877 03:46:48,484 --> 03:46:50,986 SO LET'S SEE WHAT'S HAPPENING 5878 03:46:50,986 --> 03:46:51,186 HERE. 5879 03:46:51,186 --> 03:46:57,359 I HAVE TO FIND MY TALK. 5880 03:46:57,359 --> 03:47:05,134 ALL RIGHT. 5881 03:47:05,134 --> 03:47:07,069 >> FIRST I WANT TO REALLY THANK 5882 03:47:07,069 --> 03:47:09,304 THE ORGANIZER FOR HAVING ME HERE 5883 03:47:09,304 --> 03:47:10,372 HIST A SPECIAL MOMENT TO BE 5884 03:47:10,372 --> 03:47:10,606 HERE. 5885 03:47:10,606 --> 03:47:14,443 I LEFT THE NIH EXACTLY TEN YEARS 5886 03:47:14,443 --> 03:47:15,978 AND 28 DAYS AGO, NOT THAT ANYONE 5887 03:47:15,978 --> 03:47:18,914 IS COUNTING, AND BEING BACK HERE 5888 03:47:18,914 --> 03:47:20,149 IS JUST TRULY SPECIAL. 5889 03:47:20,149 --> 03:47:21,650 THIS IS A COMMUNITY THAT HAS 5890 03:47:21,650 --> 03:47:23,385 RAISED ME, I FEEL LIKE I'M A 5891 03:47:23,385 --> 03:47:25,587 CHILD OF THE NIH AND I'M REALLY 5892 03:47:25,587 --> 03:47:27,189 GRATEFUL FOR THE MENTORS THAT 5893 03:47:27,189 --> 03:47:29,391 I'VE HAD HERE AND THE NOW 5894 03:47:29,391 --> 03:47:31,026 COLLEAGUES AND FRIENDS THAT I 5895 03:47:31,026 --> 03:47:31,593 HAVE. 5896 03:47:31,593 --> 03:47:33,028 SO THANK YOU SO MUCH. 5897 03:47:33,028 --> 03:47:34,563 AS I WAS TELLING AL EARLIER, MY 5898 03:47:34,563 --> 03:47:36,965 TALK IS REALLY IMMUNOLOGY 5899 03:47:36,965 --> 03:47:38,700 ADJACENT SO I'M GOING TO TALK 5900 03:47:38,700 --> 03:47:45,541 ABOUT REALLY IMMUNE CELLS, BUT 5901 03:47:45,541 --> 03:47:47,709 ABOUT THEIR IMPACT ON TISSUES. 5902 03:47:47,709 --> 03:47:48,911 MY LAB TRIES TO BROADLY 5903 03:47:48,911 --> 03:47:50,145 UNDERSTAND HOW ENVIRONMENTAL 5904 03:47:50,145 --> 03:47:52,114 INPUTS ARE INTERPRETED BY OUR 5905 03:47:52,114 --> 03:47:53,415 BODY AT ESSENTIALLY BARRIER 5906 03:47:53,415 --> 03:47:54,616 TISSUES LIKE THE EXPINT GUT, 5907 03:47:54,616 --> 03:47:56,919 THESE ARE OUR WINDOWS TO THE 5908 03:47:56,919 --> 03:47:58,554 EXTERNAL ENVIRONMENT. 5909 03:47:58,554 --> 03:48:00,956 IN OUR ADMITTEDLY BIASED OPINION 5910 03:48:00,956 --> 03:48:02,591 BELIEVE THAT HOW OUR TISSUES 5911 03:48:02,591 --> 03:48:04,193 RESPOND TO THESE ENVIRONMENTAL 5912 03:48:04,193 --> 03:48:05,761 TRIGGERS REALLY DEPEND ON HOW 5913 03:48:05,761 --> 03:48:07,963 IMMUNE CELLS SENSE THESE 5914 03:48:07,963 --> 03:48:09,865 PERTUBATIONS OR ALTERATIONS, AND 5915 03:48:09,865 --> 03:48:11,900 COMMUNICATE WITH THE REST OF THE 5916 03:48:11,900 --> 03:48:12,134 TISSUE. 5917 03:48:12,134 --> 03:48:13,435 SO WE ARE INTERESTED IN 5918 03:48:13,435 --> 03:48:16,271 UNDERSTANDING THE MECHANISMS 5919 03:48:16,271 --> 03:48:17,673 UNDERLYING THESE COMMUNICATIONS 5920 03:48:17,673 --> 03:48:19,341 NOT JUST FROM THE PERSPECTIVE OF 5921 03:48:19,341 --> 03:48:20,742 IMMUNE CELL BUT ALSO FROM THE 5922 03:48:20,742 --> 03:48:22,377 PERSPECTIVE OF THE RECEIVER 5923 03:48:22,377 --> 03:48:24,379 CELLS AND THE CONSEQUENCES OF 5924 03:48:24,379 --> 03:48:25,781 THESE COMMUNICATIONS FOR BOTH 5925 03:48:25,781 --> 03:48:27,082 HEALTH AND DISEASE. 5926 03:48:27,082 --> 03:48:28,617 AS WAS BEAUTIFULLY BROUGHT UP BY 5927 03:48:28,617 --> 03:48:30,385 THE PREVIOUS TALK, THINGS THAT 5928 03:48:30,385 --> 03:48:32,120 ARE REALLY OPERATIVE IN HEALTH 5929 03:48:32,120 --> 03:48:33,655 ARE COOPTED PATHOLOGICALLY IN 5930 03:48:33,655 --> 03:48:34,122 DISEASE. 5931 03:48:34,122 --> 03:48:36,625 SO WE THINK THERE'S REALLY A 5932 03:48:36,625 --> 03:48:38,494 BEEN TIGHTED TO STUDYING HEALTH 5933 03:48:38,494 --> 03:48:40,028 TO UNDERSTAND NOVEL AVENUES OF 5934 03:48:40,028 --> 03:48:40,863 TARGETING DISEASE. 5935 03:48:40,863 --> 03:48:42,664 I'M GOING TO TELL YOU ONE SORT 5936 03:48:42,664 --> 03:48:44,299 OF NARRATIVE THAT'S BEEN EMERGE 5937 03:48:44,299 --> 03:48:46,935 FRG MY LAB OVER THE PAST FEW 5938 03:48:46,935 --> 03:48:49,605 YEARS THAT SORT OF EXEMPLIFIES 5939 03:48:49,605 --> 03:48:49,905 THIS. 5940 03:48:49,905 --> 03:48:53,675 SO WE STUDY IMMUNE TISSUE 5941 03:48:53,675 --> 03:48:55,210 CROSSTALK IN VARIOUS CONTEXT 5942 03:48:55,210 --> 03:48:56,745 TODAY I'LL TALK TO YOU ABOUT THE 5943 03:48:56,745 --> 03:48:58,380 CONTEXT OF TISSUE DAMAGE AND 5944 03:48:58,380 --> 03:49:00,015 REPAIR AND HOW OUR SYSTEMS COPE 5945 03:49:00,015 --> 03:49:01,250 WITH THIS. 5946 03:49:01,250 --> 03:49:02,551 WE'RE ADDRESS THIS QUESTION OF 5947 03:49:02,551 --> 03:49:04,052 THE SKIN WHICH IS OUR OF COURSE 5948 03:49:04,052 --> 03:49:07,222 PRIMARY BARRIER BUT ALSO HAS 5949 03:49:07,222 --> 03:49:09,424 REALLY SOPHISTICATED REPAIR 5950 03:49:09,424 --> 03:49:09,725 MECHANISMS. 5951 03:49:09,725 --> 03:49:11,260 EVERYONE HERE GETS CUTS AND 5952 03:49:11,260 --> 03:49:12,728 SCRAPES ALL THE TIME, AND EVERY 5953 03:49:12,728 --> 03:49:15,898 ONE HERE HAS IMMUNE CELLS AND 5954 03:49:15,898 --> 03:49:18,400 LYMPHOCYTES IN THEIR SKIN, SO WE 5955 03:49:18,400 --> 03:49:20,702 ARE REALLY INTERESTED IN 5956 03:49:20,702 --> 03:49:21,803 UNDERSTANDING WHAT DO THESE LIMB 5957 03:49:21,803 --> 03:49:23,338 FOAF SIGHTS DO WHEN YOU GET A 5958 03:49:23,338 --> 03:49:25,874 CUT OR SCRAPE IN PHYSIOLOGICAL 5959 03:49:25,874 --> 03:49:26,174 CONTEXT? 5960 03:49:26,174 --> 03:49:27,175 WHAT IS THEIR ROLE? 5961 03:49:27,175 --> 03:49:27,943 DO THEY PARTICIPATE? 5962 03:49:27,943 --> 03:49:30,979 AND HOW DO THEY REALLY MEDIATE 5963 03:49:30,979 --> 03:49:32,548 THEIR FUNCTION IN THIS CONTEXT 5964 03:49:32,548 --> 03:49:33,081 OF REPAIR? 5965 03:49:33,081 --> 03:49:34,716 IS IT BEYOND JUST KILLING THE 5966 03:49:34,716 --> 03:49:36,251 BUGS THAT GET IN? 5967 03:49:36,251 --> 03:49:38,320 SO THIS IS JUST AN IMAGE OF 5968 03:49:38,320 --> 03:49:40,088 HEALTHY HUMAN SKIN AND YOU SEE 5969 03:49:40,088 --> 03:49:41,390 THAT THERE ARE REALLY MANY, MANY 5970 03:49:41,390 --> 03:49:42,824 DIFFERENT TYPES OF LYMPHOCYTES 5971 03:49:42,824 --> 03:49:44,459 THAT HANG OUT AROUND THE HAIR 5972 03:49:44,459 --> 03:49:46,995 FOLLICLES AND AROUND THE 5973 03:49:46,995 --> 03:49:48,830 VASCULATURE, AND WE THINK THAT 5974 03:49:48,830 --> 03:49:50,465 UNDERSTANDING THIS QUESTION IS 5975 03:49:50,465 --> 03:49:52,000 REALLY IMPORTANT NOT JUST FOR 5976 03:49:52,000 --> 03:49:53,101 UNDERSTANDING THE FUNDAMENTALS 5977 03:49:53,101 --> 03:49:55,070 OF HOW TISSUES HEAL AND COPE 5978 03:49:55,070 --> 03:49:56,371 WITH DAMAGE BUT BECAUSE THESE 5979 03:49:56,371 --> 03:50:00,108 SAME FACTORS GO AWRY IN A PLET 5980 03:50:00,108 --> 03:50:01,643 AFTER INFLAMMATORY DISEASES. 5981 03:50:01,643 --> 03:50:04,713 DAMAGE PATHOLOGIES ARE COOPTED 5982 03:50:04,713 --> 03:50:06,748 IN MANY, MANY CONDITIONS, IN 5983 03:50:06,748 --> 03:50:08,250 FACT, I THINK THAT OFTEN WE 5984 03:50:08,250 --> 03:50:09,785 SUCCUMB TO DISEASE BECAUSE OF 5985 03:50:09,785 --> 03:50:10,552 TISSUE DAMAGE. 5986 03:50:10,552 --> 03:50:11,954 OUR ORGANS FAIL BECAUSE OF 5987 03:50:11,954 --> 03:50:13,255 TISSUE DAMAGE. 5988 03:50:13,255 --> 03:50:14,256 SO UNDERSTANDING HOW WE COPE 5989 03:50:14,256 --> 03:50:16,658 WITH THIS IS REALLY CENTRAL TO 5990 03:50:16,658 --> 03:50:17,392 ADVANCING HEALTH. 5991 03:50:17,392 --> 03:50:19,962 AND ALSO THESE PATHOLOGIES ARE 5992 03:50:19,962 --> 03:50:21,797 REEMERGED IN THE CONTEXT, SO 5993 03:50:21,797 --> 03:50:24,132 HERE I HAVE TWO EXAMPLES, 5994 03:50:24,132 --> 03:50:25,233 PSORIASIS WHICH IS REALLY A 5995 03:50:25,233 --> 03:50:26,635 PATHOLOGY OF OVERHEALING WHERE 5996 03:50:26,635 --> 03:50:28,704 YOU HAVE EPITHELIAL HYPERPLASIA 5997 03:50:28,704 --> 03:50:30,339 AND I AM UNIHYPERACTIVATION AND 5998 03:50:30,339 --> 03:50:31,373 CROHN'S DISEASE WHICH IS 5999 03:50:31,373 --> 03:50:32,441 PATHOLOGY WHERE YOU HAVE 6000 03:50:32,441 --> 03:50:34,743 REPEATED DAMAGE AND REPAIR 6001 03:50:34,743 --> 03:50:36,178 CYCLES AND INFLAMMATION THAT 6002 03:50:36,178 --> 03:50:38,480 CORRUPTS YOUR ENTIRE INTESTINE. 6003 03:50:38,480 --> 03:50:40,549 AGAIN, INCREASINGLY FOR THIS 6004 03:50:40,549 --> 03:50:42,217 TALK, BOTH OF THESE DISEASES ARE 6005 03:50:42,217 --> 03:50:45,053 IN SOME FORM OR ANOTHER MEDIATED 6006 03:50:45,053 --> 03:50:47,255 BY HYPERACTIVATED IMMUNE CELLS 6007 03:50:47,255 --> 03:50:49,124 AND PATHOLOGICAL SIGNALLING 6008 03:50:49,124 --> 03:50:49,391 EPITHELIA. 6009 03:50:49,391 --> 03:50:50,525 WHAT IS SUSTAINING THIS 6010 03:50:50,525 --> 03:50:52,194 CROSSTALK AND WHAT ARE THE 6011 03:50:52,194 --> 03:50:54,029 MEDIATORS OF THIS CROSSTALK? 6012 03:50:54,029 --> 03:50:55,897 SO WE RECENTLY PUBLISHED THIS 6013 03:50:55,897 --> 03:50:58,400 PAPER, I WANT TO ADVANCE TO 6014 03:50:58,400 --> 03:51:00,936 UNPUBLISHED FINDING, I'M 6015 03:51:00,936 --> 03:51:04,806 SUMMARIZING THIS, TWO FANTASTIC 6016 03:51:04,806 --> 03:51:05,674 POSTDOCTORAL FELL LOAPS IN THE 6017 03:51:05,674 --> 03:51:07,943 LAB UNCOVERED NEW MECHANISMS BY 6018 03:51:07,943 --> 03:51:09,911 WHICH IMMUNE CELLS CROPS THE 6019 03:51:09,911 --> 03:51:11,346 EPITHELIA 6789 SO WHEN YOU HAVE 6020 03:51:11,346 --> 03:51:12,881 A WOUND. 6021 03:51:12,881 --> 03:51:14,650 SO WHEN YOU HAVE A WOUND, AT THE 6022 03:51:14,650 --> 03:51:16,051 EDGE OF THE WOUND YOU HAVE STEM 6023 03:51:16,051 --> 03:51:17,419 CELLS THAT PROLIFERATE AND MIE 6024 03:51:17,419 --> 03:51:19,021 GRAIRT ACCIDENT EXPAND AND GO 6025 03:51:19,021 --> 03:51:23,392 MAKING BRAND-NEW TISSUE. 6026 03:51:23,392 --> 03:51:29,931 WHAT YUE AND PIOTRI FOUND IS LZ 6027 03:51:29,931 --> 03:51:32,467 TH17 WHICH TRIGGERS CELLS TO 6028 03:51:32,467 --> 03:51:34,369 EXPAND -- IL17 WHICH TRIGGERS 6029 03:51:34,369 --> 03:51:36,838 THE CELLS TO EX-BAND AND MOVE. 6030 03:51:36,838 --> 03:51:39,007 LYMPHOID CELLS PLAY CRITICAL 6031 03:51:39,007 --> 03:51:39,608 ROLES? 6032 03:51:39,608 --> 03:51:40,642 CROSSTALKING WITH STEM CELLS AND 6033 03:51:40,642 --> 03:51:42,310 PROMOTING THEIR SORT OF TISSUE 6034 03:51:42,310 --> 03:51:44,479 GROWTH POTENTIAL. 6035 03:51:44,479 --> 03:51:46,248 MORE SURPRISING WAS HOW THIS 6036 03:51:46,248 --> 03:51:46,515 HAPPENED. 6037 03:51:46,515 --> 03:51:48,850 SO WHAT THEY FOUND IS THAT IN 6038 03:51:48,850 --> 03:51:51,687 FACT IL-17A ENABLES THE 6039 03:51:51,687 --> 03:51:53,121 ADAPTATION OF THESE STEM CELLS 6040 03:51:53,121 --> 03:51:56,191 TO THE HARSH HYPOXIC ENVIRONMENT 6041 03:51:56,191 --> 03:51:57,259 OF A WOUND EDGE. 6042 03:51:57,259 --> 03:51:58,560 THIS IS SOME OF THE MECHANISTIC 6043 03:51:58,560 --> 03:51:59,327 EVIDENCE THAT SUPPORTS THIS. 6044 03:51:59,327 --> 03:52:01,463 ON THE TOP YOU HAVE CONTROL 6045 03:52:01,463 --> 03:52:03,065 WOUNDS AND THE RED REALLY MARKS 6046 03:52:03,065 --> 03:52:06,334 THE EPITHELIAL THAT IS RIGHT AT 6047 03:52:06,334 --> 03:52:07,869 THE EDGE OF THE WOUND AND THE 6048 03:52:07,869 --> 03:52:15,644 GREEN DOTS ARE AM AM HIF1 ALPHA, 6049 03:52:15,644 --> 03:52:16,645 MEDIATES HIGH POX YARKS BRIGHTLY 6050 03:52:16,645 --> 03:52:18,313 LIT UP AT THE WOUND'S EDGE 6051 03:52:18,313 --> 03:52:18,513 THERE. 6052 03:52:18,513 --> 03:52:21,583 IF YOU TAKE AWAY TYPE 17 CELLS 6053 03:52:21,583 --> 03:52:22,918 AND FOR THE CONNOISSEURS AS YOU 6054 03:52:22,918 --> 03:52:26,121 GUYS ARE, GAMA DELTA T CELLS, 6055 03:52:26,121 --> 03:52:27,923 DID ALL THE VARIOUS KNOCKOUTS, 6056 03:52:27,923 --> 03:52:30,525 YOU SEE THAT THAT HISTONE ALPHA 6057 03:52:30,525 --> 03:52:33,128 SIGNAL IS GONE, IT'S REMARKABLE 6058 03:52:33,128 --> 03:52:34,162 IT'S GONE EVEN THOUGH THERE IS 6059 03:52:34,162 --> 03:52:35,697 NO IFNLTS DIFFERENCE BETWEEN 6060 03:52:35,697 --> 03:52:36,898 HYPOXIA BETWEEN THESE TWO 6061 03:52:36,898 --> 03:52:38,433 SITUATIONS, THAT TOLD US REALLY 6062 03:52:38,433 --> 03:52:41,069 THAT IL17 WAS CRITICAL FOR THE 6063 03:52:41,069 --> 03:52:43,038 INDUCTION OF THIS TRANSCRIPTION 6064 03:52:43,038 --> 03:52:44,973 FACTOR THEAN ABLED ADAPTATION OF 6065 03:52:44,973 --> 03:52:47,209 THESE CELLS AT THE WOUND'S EDGE 6066 03:52:47,209 --> 03:52:49,144 TO THIS HYPOXIC CONDITION AND 6067 03:52:49,144 --> 03:52:50,645 THAT HYPOXIA WAS NOT SUFFICIENT 6068 03:52:50,645 --> 03:52:51,847 SO WE ESSENTIALLY SORT OF CAME 6069 03:52:51,847 --> 03:52:58,320 TO THIS CONCLUSION THAT, YOU 6070 03:52:58,320 --> 03:53:03,225 KNOW, -- LZ CONSISTENTLY 6071 03:53:03,225 --> 03:53:06,161 EXPRESSED OR DEGRADED, ACUTE 6072 03:53:06,161 --> 03:53:08,029 HYPOXIA, STABILIZED, GO TO 6073 03:53:08,029 --> 03:53:10,899 NUCLEUS AND ENABLE CELL TO 6074 03:53:10,899 --> 03:53:12,634 SURVIVE IN LOW OXYGEN CONDITION. 6075 03:53:12,634 --> 03:53:14,069 CONDITIONS OF CHRONIC HYPOXIA 6076 03:53:14,069 --> 03:53:15,804 YOU REALLY NEED SECONDARY 6077 03:53:15,804 --> 03:53:19,674 SIGNALS LIKE IL17A THAT SIGNAL 6078 03:53:19,674 --> 03:53:23,044 ML TO ENABLE THIS ADAPTATION AND 6079 03:53:23,044 --> 03:53:24,613 EPITHELIAL ORGANOID SYSTEM TO 6080 03:53:24,613 --> 03:53:25,113 DEMONSTRATE THIS. 6081 03:53:25,113 --> 03:53:26,882 SO IF YOU TAKE PRIMARY 6082 03:53:26,882 --> 03:53:28,283 EPITHELIAL ORG NOIDZ, CULTURE 6083 03:53:28,283 --> 03:53:30,919 THEM IN EITHER NORM OXIA OR 6084 03:53:30,919 --> 03:53:32,320 ACUTE OR CHRONIC HOX YARKS WHAT 6085 03:53:32,320 --> 03:53:33,755 YOU SEE IS THAT -- CAN YOU SEE 6086 03:53:33,755 --> 03:53:34,623 MY POINTER? 6087 03:53:34,623 --> 03:53:34,823 YEAH. 6088 03:53:34,823 --> 03:53:37,459 YOU SEE THAT IN ACUTE HYPOXIA 6089 03:53:37,459 --> 03:53:38,560 ALL SYSTEMS ARE G THRINGZ 6090 03:53:38,560 --> 03:53:42,264 WORKING AS THEY THEY SHOULD, 6091 03:53:42,264 --> 03:53:45,233 HIS TURNS ON FURTHER AUGMENTED 6092 03:53:45,233 --> 03:53:46,802 BY IL17. 6093 03:53:46,802 --> 03:53:50,806 IN CHRONIC HYPOXIA IT IS NOT THE 6094 03:53:50,806 --> 03:53:51,006 CASE. 6095 03:53:51,006 --> 03:53:54,142 IF YOU GIVE IL17 IT RESCUES 6096 03:53:54,142 --> 03:53:54,342 THIS. 6097 03:53:54,342 --> 03:53:56,011 AND THIS MODE UNTIL THEY GET 6098 03:53:56,011 --> 03:53:59,147 SIGNALS FROM THEIR SORT OF 6099 03:53:59,147 --> 03:54:01,116 IMMUNOLOGICAL BROTHER EN. 6100 03:54:01,116 --> 03:54:02,884 AND BROADLY THE SORT OF BROUGHT 6101 03:54:02,884 --> 03:54:04,186 UP THIS PICTURE THAT WHEN YOU 6102 03:54:04,186 --> 03:54:07,355 HAVE AN INJURY, YOU HAVE 6103 03:54:07,355 --> 03:54:08,790 COOPERATIVITY BETWEEN YOUR 6104 03:54:08,790 --> 03:54:11,526 IMMUNE CELLS AND YOUR WOUND EDGE 6105 03:54:11,526 --> 03:54:13,261 EPITHELIAL PROGENITORS AND THAT 6106 03:54:13,261 --> 03:54:15,163 REALLY THE 17 SIGNAL FROM YOUR 6107 03:54:15,163 --> 03:54:17,566 EU78 UNICELLS ENABLES THIS 6108 03:54:17,566 --> 03:54:18,967 ADAPTATION HYPOXIA WHICH THEN 6109 03:54:18,967 --> 03:54:20,502 ALLOWS THESE CELLS TO CHANGE 6110 03:54:20,502 --> 03:54:23,338 THEIR METABOLISM AND FUEL 6111 03:54:23,338 --> 03:54:23,572 REPAIR. 6112 03:54:23,572 --> 03:54:26,208 SO WE THINK THAT THIS IS IN FACT 6113 03:54:26,208 --> 03:54:28,310 AN EXEMPLAR OF MULTICELLULARITY. 6114 03:54:28,310 --> 03:54:30,545 HOW WE STUDY HYPOXIC RESPONSE 6115 03:54:30,545 --> 03:54:33,281 INSIDE A DISH IS REALLY STUDYING 6116 03:54:33,281 --> 03:54:35,383 ACUTE HYPOXIC RESPONSES WHICH 6117 03:54:35,383 --> 03:54:36,585 REALLY REPRESENT EITHER 6118 03:54:36,585 --> 03:54:37,552 UNICELLULAR ORGANISMS OR WHEN 6119 03:54:37,552 --> 03:54:39,554 YOU ARE FIRST ENCOUNTERING 6120 03:54:39,554 --> 03:54:40,889 HYPOXIA AND WHEN YOU THINK ABOUT 6121 03:54:40,889 --> 03:54:42,157 THE SORT OF PROLONGED TIME SCALE 6122 03:54:42,157 --> 03:54:44,125 OF WHAT IT TAKES TO HEAL A 6123 03:54:44,125 --> 03:54:45,560 TISSUE AND MAKE BRAND-NEW TISSUE 6124 03:54:45,560 --> 03:54:47,095 AND THE ENERGY INVOLVED AND 6125 03:54:47,095 --> 03:54:48,730 COMPLEXITY INVOLVED, WE THINK 6126 03:54:48,730 --> 03:54:50,031 THAT THERE IS ESSENTIALLY 6127 03:54:50,031 --> 03:54:51,099 CROSSTALK BETWEEN IMMUNE CELLS 6128 03:54:51,099 --> 03:54:53,301 AND THE EPITHELIA THAT ENABLE 6129 03:54:53,301 --> 03:54:53,635 THIS. 6130 03:54:53,635 --> 03:54:54,970 SO WHAT HAPPENS WHEN THIS 6131 03:54:54,970 --> 03:54:56,738 CROSSTALK GOES AWRY AND WHEN 6132 03:54:56,738 --> 03:54:58,039 THESE CONVERSATIONS ARE 6133 03:54:58,039 --> 03:54:59,341 UNCHECKED AND WHEN YOU HAVE 6134 03:54:59,341 --> 03:55:02,544 MAYBE TOO MUCH SECONDARY SIGNAL? 6135 03:55:02,544 --> 03:55:05,180 SO THIS IS WHERE I WANT TO COME 6136 03:55:05,180 --> 03:55:06,815 BACK TO PSORIASIS WHICH IS THIS 6137 03:55:06,815 --> 03:55:08,316 DISEASE OF OVERHEALING AND MANY 6138 03:55:08,316 --> 03:55:10,318 OF THE PATHOLOGICAL FEATURES YOU 6139 03:55:10,318 --> 03:55:12,721 SEE IN A WOUND ARE ESSENTIALLY 6140 03:55:12,721 --> 03:55:14,689 EXAGGERATED AND PSORIASIS WHERE 6141 03:55:14,689 --> 03:55:16,625 INSTEAD OF A WOUND HEALING AND 6142 03:55:16,625 --> 03:55:18,293 MAKING NEW TISSUE THIS WAY, IT'S 6143 03:55:18,293 --> 03:55:19,828 NOW 3W45EUBGING PLAQUES UPWARD 6144 03:55:19,828 --> 03:55:21,062 AND WHAT'S REMARKABLE SYNTHESIS 6145 03:55:21,062 --> 03:55:22,297 ONE OF THE DISEASES THAT'S 6146 03:55:22,297 --> 03:55:24,766 RESPONSIVE TO IL17 THERAPY SO IN 6147 03:55:24,766 --> 03:55:28,169 MANY CASES PATIENTS IN FACT THEY 6148 03:55:28,169 --> 03:55:29,371 RESOLVE THEIR DISEASE, THOUGH 6149 03:55:29,371 --> 03:55:30,639 THAT IS NOT CURATIVE. 6150 03:55:30,639 --> 03:55:32,007 SO WE THOUGHT THIS WOULD BE A 6151 03:55:32,007 --> 03:55:33,174 REALLY INTERESTING DISEASE TO 6152 03:55:33,174 --> 03:55:37,012 PROBE IF OUR NOVEL IL17 ALPHA 6153 03:55:37,012 --> 03:55:38,546 AXIS WAS OPERATIVE AND IF IT 6154 03:55:38,546 --> 03:55:40,282 PLAYED ANY ROLE IN DRIVING 6155 03:55:40,282 --> 03:55:41,816 PATHOLOGY OR WAS IT SIMPLY 6156 03:55:41,816 --> 03:55:44,786 FUELING THAT SORT OF EPITHELIAL 6157 03:55:44,786 --> 03:55:45,854 HYPERPROLIFERATION? 6158 03:55:45,854 --> 03:55:48,023 AND SO WE PAIRED UP WITH THE 6159 03:55:48,023 --> 03:55:51,026 DIRECTOR OF ARTHRITIS CENTER AND 6160 03:55:51,026 --> 03:55:56,064 TOGETHER JOSE AND I MENTORED A 6161 03:55:56,064 --> 03:55:58,667 WONDERFUL FELLOW NOW AT BRIGHAM 6162 03:55:58,667 --> 03:56:03,038 CONSTITUTED STUDYING MYOSITIS 6163 03:56:03,038 --> 03:56:06,508 AND LEVERAGED SPATIAL TO 6164 03:56:06,508 --> 03:56:08,109 UNDERSTAND HOW REPAIR MECHANISMS 6165 03:56:08,109 --> 03:56:09,878 MAY BE COOPTED IN THIS DISEASE. 6166 03:56:09,878 --> 03:56:12,047 HERE ICHT TO MAKE A POINT WE 6167 03:56:12,047 --> 03:56:17,519 DIDN'T JUST USE SPATIAL 6168 03:56:17,519 --> 03:56:18,720 SCRIPTOMICS OR PRETTY PICTURES 6169 03:56:18,720 --> 03:56:19,721 OR LOVE SPENDING MONEY WHICH I 6170 03:56:19,721 --> 03:56:21,489 DO NOT, BUT THIS ALLOWS ACCESS 6171 03:56:21,489 --> 03:56:23,224 TO CELL TYPES THAT ARE VERY HARD 6172 03:56:23,224 --> 03:56:24,859 TO GERKTS EITHER CELLS DHEENLTD 6173 03:56:24,859 --> 03:56:27,696 HAVE NUCLEI LIKE DIFFERENTIATED 6174 03:56:27,696 --> 03:56:29,698 EPITHELIAL CELLS OR SITES THAT 6175 03:56:29,698 --> 03:56:31,833 DIE OR EVEN WITH RNA SEQ YOU 6176 03:56:31,833 --> 03:56:34,569 DON'T REALLY GET THEIR 6177 03:56:34,569 --> 03:56:37,505 TRANSCRIPTIONAL LANDSCAPE. 6178 03:56:37,505 --> 03:56:42,477 SO ROCHELLE IS VERY INTERESTED 6179 03:56:42,477 --> 03:56:45,647 IN UNDERSTANDING SORE YAT I CAN 6180 03:56:45,647 --> 03:56:47,849 ARTHRITIS, THE MAN MANUSCRIPT IS 6181 03:56:47,849 --> 03:56:48,116 PUBLISHED. 6182 03:56:48,116 --> 03:56:49,584 WE WERE REALLY INTERESTED IN IS 6183 03:56:49,584 --> 03:56:50,885 OUR AXIS OPERATIVE HERE? 6184 03:56:50,885 --> 03:56:52,187 SO THESE ARE THE KIND OF 6185 03:56:52,187 --> 03:56:55,190 BEAUTIFUL PICTURES YOU GET, FULL 6186 03:56:55,190 --> 03:56:56,391 GENOME EXPRESSION ACROSS THE 6187 03:56:56,391 --> 03:56:58,326 TISH LUND SCAPE, EVERY DOT IS 6188 03:56:58,326 --> 03:57:00,295 YOU GUYS HAVE SEEN LOTS TODAY 6189 03:57:00,295 --> 03:57:02,063 HERE INSTEAD OF EVERY DOT BEING 6190 03:57:02,063 --> 03:57:06,234 A SINGLE CELL, EVERY DOT IS A 5S 6191 03:57:06,234 --> 03:57:07,736 ALREADY OUTDATED BECAUSE NOW 6192 03:57:07,736 --> 03:57:09,738 THERE'S MUCH HIGHER RESOLUTION 6193 03:57:09,738 --> 03:57:11,639 ST OUT THERE WE'VE MOVED TO BUT 6194 03:57:11,639 --> 03:57:14,309 YOU CAN ESSENTIALLY SEE WHAT 6195 03:57:14,309 --> 03:57:15,677 ECOSYSTEMS ARE EXPRESSING AND 6196 03:57:15,677 --> 03:57:16,711 WHAT'S DIFFERENT BETWEEN A 6197 03:57:16,711 --> 03:57:19,481 DISEASE STATE AND NOT AND THEN 6198 03:57:19,481 --> 03:57:21,850 DIG DEEPER INTO WHAT ARE THE 6199 03:57:21,850 --> 03:57:23,518 SPECIFIC GENES THAT ARE 6200 03:57:23,518 --> 03:57:24,285 DYSREGULATED IN THIS. 6201 03:57:24,285 --> 03:57:25,587 SO WE WANT TO DO ASK THIS 6202 03:57:25,587 --> 03:57:26,921 QUESTION, WE SPECIFICALLY WANTED 6203 03:57:26,921 --> 03:57:29,290 TO HONE IN ON THE ECOSYSTEM THAT 6204 03:57:29,290 --> 03:57:31,126 WAS DYSREGULATED IN THE SORE YAT 6205 03:57:31,126 --> 03:57:35,296 I CAN TISSUE -- PSORIATIC 6206 03:57:35,296 --> 03:57:37,098 TISSUE, MARKED IN RED, WITH BIG 6207 03:57:37,098 --> 03:57:39,267 RED ARROWS, LOOKING AT SPECIFIC 6208 03:57:39,267 --> 03:57:41,102 AREAS DYSREGULATED AND REALLY 6209 03:57:41,102 --> 03:57:44,072 EMERGE IN PSORIATIC SKIN AND 6210 03:57:44,072 --> 03:57:46,574 EXCITED TO FIND THAT IN FACT IL7 6211 03:57:46,574 --> 03:57:48,576 WERE UNIQUELY CO-ENRICHED IN 6212 03:57:48,576 --> 03:57:49,878 THESE AREAS, SO THIS IS 6213 03:57:49,878 --> 03:57:51,579 SEQUENCING AND WE MUST VALIDATE 6214 03:57:51,579 --> 03:57:53,048 AT A PROTEIN EXPLEFL A 6215 03:57:53,048 --> 03:57:55,116 FUNCTIONAL LEVEL, SO WE DID JUST 6216 03:57:55,116 --> 03:57:56,317 THAT -- LEVEL. 6217 03:57:56,317 --> 03:57:58,686 SOILY ANOTHER Ph.D. STUDENT IN 6218 03:57:58,686 --> 03:58:01,489 THE LAB AND PITRA TEAMED UP TO 6219 03:58:01,489 --> 03:58:04,059 TAKE THIS ON WITH OUR 6220 03:58:04,059 --> 03:58:05,794 BIOMATHEMATICIAN AND THEY FIRST 6221 03:58:05,794 --> 03:58:08,830 STARTED BY LOOKING AT IS HIS 6222 03:58:08,830 --> 03:58:11,699 OVEREXPRESSED IN -- IS HIF 6223 03:58:11,699 --> 03:58:13,334 OVEREXPRESSED IN INFLAMMATORY 6224 03:58:13,334 --> 03:58:15,203 DISEASES, IN HEALTHY SKIN, NO 6225 03:58:15,203 --> 03:58:17,605 HIF EXPRESSION IN EPIDERMIS OR 6226 03:58:17,605 --> 03:58:18,273 ELSEWHERE, GREEN IN THE 6227 03:58:18,273 --> 03:58:19,240 BACKGROUND IS JUST BACKGROUND. 6228 03:58:19,240 --> 03:58:22,010 THIS GOES UP LIKE GANG BUSTERS 6229 03:58:22,010 --> 03:58:27,582 IN PSORIASIS, ALWAYS TOPIC 6230 03:58:27,582 --> 03:58:29,517 DERMATITIS, AND RS IT CORRELATED 6231 03:58:29,517 --> 03:58:31,519 WITH THE PRESENCE OF IMMUNE 6232 03:58:31,519 --> 03:58:33,388 INFILTRATES AND IN AREAS WHERE 6233 03:58:33,388 --> 03:58:35,457 PATHOLOGY WAS EVEN WORSE AS 6234 03:58:35,457 --> 03:58:37,559 MEASURED BY EPIDERMAL 6235 03:58:37,559 --> 03:58:38,460 HYPERPLASIA AND REALLY 6236 03:58:38,460 --> 03:58:39,394 REMARKABLE ALMOST TO THE POINT 6237 03:58:39,394 --> 03:58:40,628 OF WHERE LIKE I DID NOT BELIEVE 6238 03:58:40,628 --> 03:58:42,263 THIS DARKTS I HAD THE LAB DO IT 6239 03:58:42,263 --> 03:58:43,731 OVER AND OVER AGAIN WAS THAT IF 6240 03:58:43,731 --> 03:58:46,468 YOU LOOKED AT RARE IL17 RECEPTOR 6241 03:58:46,468 --> 03:58:48,503 WAS EXPRESSED IN THE EPITHELIA, 6242 03:58:48,503 --> 03:58:50,038 WHERE, IN RED, AND IF YOU LOOK 6243 03:58:50,038 --> 03:58:53,007 AT WHERE HIF1 ALPHA EXPRESSED IT 6244 03:58:53,007 --> 03:58:54,776 WAS 100% CONCORD GNAT, 6245 03:58:54,776 --> 03:58:56,111 COMPLETELY CO-EXPRESSED IN HUMAN 6246 03:58:56,111 --> 03:58:58,379 TISSUE REALLY SUGGESTING THAT 6247 03:58:58,379 --> 03:59:00,482 HIF1 ACTIVATION IN EPITHELIA WAS 6248 03:59:00,482 --> 03:59:02,383 THIS RHEOSTAT STAT, THIS BROM 6249 03:59:02,383 --> 03:59:04,586 TEFER IL17 SIGNALLING. 6250 03:59:04,586 --> 03:59:05,487 FINALLY WE TOOK ADVANTAGE OF THE 6251 03:59:05,487 --> 03:59:06,821 FACT THAT WE COULD ESSENTIALLY 6252 03:59:06,821 --> 03:59:08,189 HAVE ACCESS TO PATIENTS THAT ARE 6253 03:59:08,189 --> 03:59:11,526 COMPLETELY THERAPY RESPONSIVE 6254 03:59:11,526 --> 03:59:13,661 AND LOOKED ABBOTT THEIR LESIONAL 6255 03:59:13,661 --> 03:59:15,330 SKIN PRE AND POST THERAPY AND 6256 03:59:15,330 --> 03:59:18,032 YOU SEE THAT PRETHERAPY THERE'S 6257 03:59:18,032 --> 03:59:21,102 TONS OF HIF1 ALPHA SIGNAL AND 6258 03:59:21,102 --> 03:59:24,072 POST THERAPY HIF1 ALPHA SIGNAL 6259 03:59:24,072 --> 03:59:26,808 GOES DOWN ON BASE LOON, NICE 6260 03:59:26,808 --> 03:59:27,542 CORRELATING DATA. 6261 03:59:27,542 --> 03:59:29,644 WHAT IS HIF ACTUAL DOOLG AND IS 6262 03:59:29,644 --> 03:59:31,513 IT ACTUALLY PLAYING A ROLE IN 6263 03:59:31,513 --> 03:59:32,147 THIS DISEASE? 6264 03:59:32,147 --> 03:59:33,014 IF SO, HOW? 6265 03:59:33,014 --> 03:59:35,250 TO DO THIS WE ESSENTIALLY SET UP 6266 03:59:35,250 --> 03:59:37,785 A QUOTE UNQUOTE TRIAL IN A DISH 6267 03:59:37,785 --> 03:59:39,220 SYSTEM BECAUSE IT'S A LOT EASIER 6268 03:59:39,220 --> 03:59:47,328 TO GET AN IRB AND THIS WOULD 6269 03:59:47,328 --> 03:59:48,997 REALLY ALLOW US TO REALLY START 6270 03:59:48,997 --> 03:59:51,299 PROBING WHAT HIF'S FUNCTION WAS 6271 03:59:51,299 --> 03:59:52,467 SIDE BY SIDE WITH STANDARD OF 6272 03:59:52,467 --> 03:59:53,835 CARE, WE ESSENTIALLY TOOK 6273 03:59:53,835 --> 03:59:55,003 BIOPSIES FROM PATIENTS WITH 6274 03:59:55,003 --> 03:59:56,137 PSORIASIS AND CULTURED THEM 6275 03:59:56,137 --> 03:59:56,804 EITHER WITH THE STANDARD OF CARE 6276 03:59:56,804 --> 04:00:01,309 OR WITH HIF1 ALPHA INHIBITORS, 6277 04:00:01,309 --> 04:00:04,279 DID ANALYSIS AND YOU SEE HIF1 6278 04:00:04,279 --> 04:00:05,914 AMFA HAS A REALLY DRAMATIC 6279 04:00:05,914 --> 04:00:07,582 EFFECT COMPARED TO STANDARD 6280 04:00:07,582 --> 04:00:08,983 WITHIN 24 HOURS, THESE ARE NOT 6281 04:00:08,983 --> 04:00:10,852 GENES KILLING OFF THE TISSUE OR 6282 04:00:10,852 --> 04:00:11,519 KILLING OFF THE CELLS. 6283 04:00:11,519 --> 04:00:13,021 IF YOU TAKE THESE GENES AND THEN 6284 04:00:13,021 --> 04:00:16,090 PROJECT THEM ONTO CLINICAL TRIAL 6285 04:00:16,090 --> 04:00:18,393 DATA THEY ESSENTIALLY PERFECTLY 6286 04:00:18,393 --> 04:00:21,496 MIRROR THE PRECISE GENES THAT 6287 04:00:21,496 --> 04:00:23,231 REFLECT FOLKS THAT HAVE RECEIVED 6288 04:00:23,231 --> 04:00:25,433 THERAPY FROM PLACEBO AND FURTHER 6289 04:00:25,433 --> 04:00:27,168 DISTINGUISH RESPONDERS FROM 6290 04:00:27,168 --> 04:00:27,669 NONRESPONDERS. 6291 04:00:27,669 --> 04:00:28,403 SO THIS IS REALLY TELLING US 6292 04:00:28,403 --> 04:00:30,471 THAT HIF MAY HAVE A FUNCTIONAL 6293 04:00:30,471 --> 04:00:30,672 ROLE. 6294 04:00:30,672 --> 04:00:32,106 BUT THIS IS PRETTY MUCH AS FAR 6295 04:00:32,106 --> 04:00:33,975 AS WE CAN GO WITH OUR HUMAN 6296 04:00:33,975 --> 04:00:34,309 DATA. 6297 04:00:34,309 --> 04:00:35,743 SO WE HAVE TO GO BACK TO OUR 6298 04:00:35,743 --> 04:00:37,378 MOUSE MODELS, WHICH MADE ME 6299 04:00:37,378 --> 04:00:38,713 VERY, VERY HAPPY BECAUSE THIS IS 6300 04:00:38,713 --> 04:00:41,015 KIND OF MY COMFORT ZONE, I 6301 04:00:41,015 --> 04:00:42,750 ALWAYS CALL MYSELF AN M-D A 6302 04:00:42,750 --> 04:00:45,153 MOUSE DOCTOR. 6303 04:00:45,153 --> 04:00:49,224 SO WE NEUFD A MOUSE MODEL OF 6304 04:00:49,224 --> 04:00:53,628 PSORIASIS, LZ TLR7 LIGAND, WELL 6305 04:00:53,628 --> 04:00:54,295 ESTABLISHED MOUSE MODEL IRVETION 6306 04:00:54,295 --> 04:00:56,431 GO FOR SIX DAYS, GET WELL 6307 04:00:56,431 --> 04:00:57,298 DEFINED COULD YOU TAKEN 6308 04:00:57,298 --> 04:00:58,166 CUTANEOUS PATHOLOGY. 6309 04:00:58,166 --> 04:01:00,268 FOR OUR PURPOSES HIF IS ALSO 6310 04:01:00,268 --> 04:01:01,803 HIGHLY HEIGHTENED IN THIS MODEL 6311 04:01:01,803 --> 04:01:02,570 AND ALSO DEPENDENT ON THE 6312 04:01:02,570 --> 04:01:04,339 EXPRESSION OF IL17 BECAUSE IF 6313 04:01:04,339 --> 04:01:06,007 YOU GOT RID OF THE 17 RECEPTOR 6314 04:01:06,007 --> 04:01:07,909 IN THE EPITHELIA YOU NO LONGER 6315 04:01:07,909 --> 04:01:09,344 HAVE THIS. 6316 04:01:09,344 --> 04:01:13,481 AND WHEN WE GIVE TOPICAL HIF1 6317 04:01:13,481 --> 04:01:15,883 ALPHA INHIBITOR, YOU NOW SEE 6318 04:01:15,883 --> 04:01:19,087 THAT IN FACT THESE MICE WERE 6319 04:01:19,087 --> 04:01:20,521 PROTECTED FROM DEVELOPMENT OF 6320 04:01:20,521 --> 04:01:21,823 DISEASE, SO WE STARTED TO WONDER 6321 04:01:21,823 --> 04:01:25,026 WHAT IS THE ROLE OF HIF IN THIS 6322 04:01:25,026 --> 04:01:26,928 EPITHELIUM AND HOW IS IT 6323 04:01:26,928 --> 04:01:27,562 CONTRIBUTING TO DAMPENING OF 6324 04:01:27,562 --> 04:01:28,296 THIS DISEASE? 6325 04:01:28,296 --> 04:01:30,798 SO WE GENERATED EPITHELIAL 6326 04:01:30,798 --> 04:01:32,000 SPECIFIC KNOCKOUTS AND IN FACT 6327 04:01:32,000 --> 04:01:34,736 THESE MICE JUST GETTING RID OF 6328 04:01:34,736 --> 04:01:36,704 HIF1 ALPHA IN THE EPITHELIA 6329 04:01:36,704 --> 04:01:37,905 COMPLETELY PROTECTED THEM FROM 6330 04:01:37,905 --> 04:01:40,008 DISEASE, ALSO IN THE SECOND 6331 04:01:40,008 --> 04:01:41,309 MODEL PSORIASIS. 6332 04:01:41,309 --> 04:01:43,177 SO WE DUG A LITTLE BIT DEEPER. 6333 04:01:43,177 --> 04:01:44,712 AND OF COURSE HIF IS KNOWN TO 6334 04:01:44,712 --> 04:01:46,314 CONTROL MANY, MANY DIFFERENT 6335 04:01:46,314 --> 04:01:48,716 ASPECTS OF DISEASE PATHOLOGY, 6336 04:01:48,716 --> 04:01:53,921 INCLUDING EXPRESSION OF VEGF 6337 04:01:53,921 --> 04:01:55,857 ALPHA, BUT WE SAW AWE KEY WAY IN 6338 04:01:55,857 --> 04:01:56,858 WHICH THINGS WERE CHANGING IN 6339 04:01:56,858 --> 04:02:00,928 THE HIF1 ALPHA DEFICIENT 6340 04:02:00,928 --> 04:02:02,997 EPITHELIAL DEFICIENT MICE WERE 6341 04:02:02,997 --> 04:02:04,732 GLYCOLYSIS, WONDERED IS TOGGLING 6342 04:02:04,732 --> 04:02:06,167 THIS METABOLIC PROGRAM GOING TO 6343 04:02:06,167 --> 04:02:07,502 ALTER DISEASE PATHOLOGY, IS THIS 6344 04:02:07,502 --> 04:02:10,004 A KEY MEANS BY WHICH THIS FACTOR 6345 04:02:10,004 --> 04:02:12,640 IS FUNCTIONING IN THIS CONTEXT? 6346 04:02:12,640 --> 04:02:14,709 SO FIRST WE NEEDED TO SEE, IS 6347 04:02:14,709 --> 04:02:16,344 THIS ACTUALLY HAPPENING IN VIVO 6348 04:02:16,344 --> 04:02:20,248 AND INDEED IN THE CONTEXT OF 6349 04:02:20,248 --> 04:02:22,950 SORT OF MOUSE MODEL YOU SEW SAW 6350 04:02:22,950 --> 04:02:26,120 THAT EXPRESSION OF GLUT1 HIGHLY 6351 04:02:26,120 --> 04:02:27,555 ENRICHED, HIGH RA INFINITY 6352 04:02:27,555 --> 04:02:28,856 TRANSPORTER FOR GLUCOSE, IF YOU 6353 04:02:28,856 --> 04:02:31,893 GET RID OF HIF1 ALPHA IN THE 6354 04:02:31,893 --> 04:02:33,494 EPITHELIA SPECIFICALLY HERE THE 6355 04:02:33,494 --> 04:02:35,163 ENTIRE IMMUNE COMPONENT IS 6356 04:02:35,163 --> 04:02:37,565 INTACTD YOU LOSE THIS GLUT1 6357 04:02:37,565 --> 04:02:39,334 EXPRESSION, REALLY COMPELLING 6358 04:02:39,334 --> 04:02:40,535 THAT IN FACT IT IS WORKING IN 6359 04:02:40,535 --> 04:02:41,836 THIS WAY. 6360 04:02:41,836 --> 04:02:44,672 SO THEN WE DECIDED TO GET RID OF 6361 04:02:44,672 --> 04:02:46,407 GLUT1 SPECIFICALLY IN THE 6362 04:02:46,407 --> 04:02:48,710 EPITHELIA AND JUST LIMITING THE 6363 04:02:48,710 --> 04:02:50,678 ABILITY OF THE EPITHELIA TO 6364 04:02:50,678 --> 04:02:52,547 UPTAKE GLUCOSE, NOT AT BASELINE, 6365 04:02:52,547 --> 04:02:54,082 BASELINE THESE MICE WERE FINE, 6366 04:02:54,082 --> 04:02:56,050 COMPLETELY PREKED THEM FROM THE 6367 04:02:56,050 --> 04:02:58,419 EPITHELIA PATHOLOGY, AND ALSO 6368 04:02:58,419 --> 04:03:00,855 THE VASCULAR AND NEURONAL 6369 04:03:00,855 --> 04:03:03,057 PATHOLOGY H SEVERAL FACETS OF 6370 04:03:03,057 --> 04:03:05,226 PATHOLOGY DOWNSTREAM OF THE 6371 04:03:05,226 --> 04:03:06,227 EPITHELIA WERE CONTROLLED BY 6372 04:03:06,227 --> 04:03:06,427 THIS. 6373 04:03:06,427 --> 04:03:10,264 AND OF COURSE MOST RELEVANT 6374 04:03:10,264 --> 04:03:12,433 HERE, REALLY CONTROLS THE IMMUNE 6375 04:03:12,433 --> 04:03:13,668 INFILTRATED AND T CELL 6376 04:03:13,668 --> 04:03:14,302 PATHOLOGY. 6377 04:03:14,302 --> 04:03:17,071 SO THIS WAS QUITE EXCITING TO US 6378 04:03:17,071 --> 04:03:18,806 BECAUSE BASICALLY LIMITING THE 6379 04:03:18,806 --> 04:03:20,475 METABOLISM OF ONE CELL TYPE WITH 6380 04:03:20,475 --> 04:03:22,410 NO PERTUBATION IN THE IMMUNE 6381 04:03:22,410 --> 04:03:23,745 SYSTEM COMPLETELY SHUT DOWN THIS 6382 04:03:23,745 --> 04:03:27,315 ENTIRE PATHOLOGICAL CIRCUIT H 6383 04:03:27,315 --> 04:03:28,783 AND REALLY SO WE STARTED TO 6384 04:03:28,783 --> 04:03:29,751 WONDER HOW THIS WAS HAPPENING 6385 04:03:29,751 --> 04:03:32,620 AND WHAT IS THIS METABOLIC 6386 04:03:32,620 --> 04:03:34,889 PROGRAM IN THE EPITHELIA REALLY 6387 04:03:34,889 --> 04:03:35,189 CONTROLLING. 6388 04:03:35,189 --> 04:03:37,859 SO WILL WE WENT BACK TO OUR 6389 04:03:37,859 --> 04:03:46,067 SEQUENCING DATA AND TYPE 17 6390 04:03:46,067 --> 04:03:48,469 IMMUNE CELLS, BOTH ALPHA DAY 6391 04:03:48,469 --> 04:03:51,439 THAT AND GAMMA DELTA INCLUDING 6392 04:03:51,439 --> 04:03:52,640 DJ'S GROUP WHO HAS BEEN A 6393 04:03:52,640 --> 04:03:54,842 PIONEER IN THIS SPACE BUT NONE 6394 04:03:54,842 --> 04:03:56,244 OF THOSE THINGS WERE REALLY 6395 04:03:56,244 --> 04:03:57,211 DIFFERENTIALLY EXPRESSED. 6396 04:03:57,211 --> 04:03:58,413 SO WHAT'S GOING ON? 6397 04:03:58,413 --> 04:03:59,781 WHY ARE WE NOT SEEING ANYTHING 6398 04:03:59,781 --> 04:04:02,683 THAT'S IMPORTANT FOR 6399 04:04:02,683 --> 04:04:04,118 IMMUNE-RELATED CONTROL IN THE 6400 04:04:04,118 --> 04:04:06,287 CONTEXT OF THIS GL YOU T1 6401 04:04:06,287 --> 04:04:08,289 DEFICIENT MOUSE AND WE WONDERED 6402 04:04:08,289 --> 04:04:10,024 MAYBE IT'S INNOCENT OUR GENE 6403 04:04:10,024 --> 04:04:10,858 EXPRESSION, MAYBE GENE 6404 04:04:10,858 --> 04:04:11,659 EXPRESSION DOESN'T CAPTURE WHAT 6405 04:04:11,659 --> 04:04:13,294 WE TBHEED TO SEE AND MAYBE 6406 04:04:13,294 --> 04:04:14,495 IT'S -- WHAT WE NEED TO SEE AND 6407 04:04:14,495 --> 04:04:16,464 MAYBE TS A METABOLITE THAT IS 6408 04:04:16,464 --> 04:04:20,234 REALLY AT THE CRUX OF THIS 6409 04:04:20,234 --> 04:04:20,902 CROSSTALK. 6410 04:04:20,902 --> 04:04:22,503 SO ONE OF THE TERMINAL 6411 04:04:22,503 --> 04:04:24,605 END-PRODUCTS OF GLYCOLYSIS IS 6412 04:04:24,605 --> 04:04:24,839 LACTATE. 6413 04:04:24,839 --> 04:04:25,940 THIS IS REALLY WELL-KNOWN 6414 04:04:25,940 --> 04:04:28,776 WHVMENT WE PUT IL17 INTO OUR 6415 04:04:28,776 --> 04:04:30,845 EPITHELIAL CULTURES WE GET DONS 6416 04:04:30,845 --> 04:04:32,513 TONS OF LACTATE. 6417 04:04:32,513 --> 04:04:37,218 ALSO WHEN YOU LOOK ATLESS IMQ 6418 04:04:37,218 --> 04:04:39,220 MOUSE THE PRESENCE OF THIS 6419 04:04:39,220 --> 04:04:39,754 INFLAMMATORY ENVIRONMENT 6420 04:04:39,754 --> 04:04:41,055 GENERATES A LOT OF LACTATE AND 6421 04:04:41,055 --> 04:04:43,724 DOES SO IN GLYCOLYSIS-DEPENDENT 6422 04:04:43,724 --> 04:04:43,958 MANNER. 6423 04:04:43,958 --> 04:04:46,661 WHEN YOU LOOK AT THE GLUT 1 6424 04:04:46,661 --> 04:04:50,231 DEFICIENT MICE, EPITHELIAL GLUTD 6425 04:04:50,231 --> 04:04:51,466 1 DEFICIENT MICE THEY DON'T MAKE 6426 04:04:51,466 --> 04:04:53,501 AS MUCH LACTATE, TELLING US 6427 04:04:53,501 --> 04:04:54,569 EPITHELIA ARE A PRIMARY SOURCE 6428 04:04:54,569 --> 04:04:56,304 OF LACTATE IN THIS CONTEXT AND 6429 04:04:56,304 --> 04:04:57,505 WONDERED IS LACTATE HERE 6430 04:04:57,505 --> 04:04:59,040 RESPONSIBLE FOR DRIVING THAT 6431 04:04:59,040 --> 04:05:00,808 PATHOLOGICAL IMMUNE RESPONSE? 6432 04:05:00,808 --> 04:05:04,045 SO WE INHIBITED HAD WITH EITHER 6433 04:05:04,045 --> 04:05:08,349 AN LDHLY INHIBITOR INHIBITS 6434 04:05:08,349 --> 04:05:11,285 LACTATE TO PYRUVATE OR INHIBITED 6435 04:05:11,285 --> 04:05:12,820 TRANSPORTERS THAT UPTAKE OR 6436 04:05:12,820 --> 04:05:14,555 RELEASE LACTATE BOTH ON 6437 04:05:14,555 --> 04:05:15,790 EPITHELIAL SIDE AND IMMUNE SIDE 6438 04:05:15,790 --> 04:05:18,493 AND IN BOTH CASES YOU'RE NOW 6439 04:05:18,493 --> 04:05:20,495 CUSHING THE IMMUNE RESPONSE AND 6440 04:05:20,495 --> 04:05:23,231 ESSENTIAL PHENOCOPYING GLUT 1 6441 04:05:23,231 --> 04:05:23,664 DEFICIENCY. 6442 04:05:23,664 --> 04:05:24,966 SO TO SUMMARIZE, I THINK WHAT 6443 04:05:24,966 --> 04:05:28,603 THAT TELLS US IS THAT DISEASE 6444 04:05:28,603 --> 04:05:31,005 STATES ARE MAINTAINED BY 6445 04:05:31,005 --> 04:05:32,940 METABOLIC CIRCUITS, AND WHEN WE 6446 04:05:32,940 --> 04:05:34,408 THINK ABOUT HOW DISEASE IS 6447 04:05:34,408 --> 04:05:36,043 FUELED AND THE ENERGY IT TAKES, 6448 04:05:36,043 --> 04:05:38,145 YOU NEED COOPERATIVITY. 6449 04:05:38,145 --> 04:05:40,081 SO I THINK ABOUT THIS A LOT, HOW 6450 04:05:40,081 --> 04:05:41,949 DO CELLS COMPETE FOR GROWTH 6451 04:05:41,949 --> 04:05:42,183 FACTORS. 6452 04:05:42,183 --> 04:05:44,318 BUT HOW DO THEY COMPETE FOR 6453 04:05:44,318 --> 04:05:45,887 NUTRIENTS AND WHY SHOULD THEY 6454 04:05:45,887 --> 04:05:47,989 COMPETE FOR NUTRIENTS WHEN THEY 6455 04:05:47,989 --> 04:05:50,024 COULD HAVE THIS SORT OF 6456 04:05:50,024 --> 04:05:50,992 COOPERATIVE ADAPTATION? 6457 04:05:50,992 --> 04:05:52,860 WE THINK THAT IN FACT AS THE 6458 04:05:52,860 --> 04:05:53,895 INFLAMMATORY CIRCUIT IS SET UP, 6459 04:05:53,895 --> 04:05:56,063 SO IS THE METABOLIC CIRCUIT AND 6460 04:05:56,063 --> 04:05:59,100 THAT ESSENTIALLY 17 SIGNALS 6461 04:05:59,100 --> 04:06:00,668 PATHOLOGICAL INTO THE EPITHELIA 6462 04:06:00,668 --> 04:06:01,903 WHICH THEN IN TURN PRODUCES A 6463 04:06:01,903 --> 04:06:05,606 NUMBER OF FACTORS THAT DRIVELY 6464 04:06:05,606 --> 04:06:07,575 17 CELLS BUT ALSO THERE'S THIS 6465 04:06:07,575 --> 04:06:08,442 METABOLIC CIRCUIT ESTABLISHED 6466 04:06:08,442 --> 04:06:09,744 AND SUSTAINED THAT SUSTAINS 6467 04:06:09,744 --> 04:06:12,513 THESE TWO CELLS, WHILE GLUCOSE 6468 04:06:12,513 --> 04:06:13,948 PROMOTES GLYCOLYSIS IN 6469 04:06:13,948 --> 04:06:15,383 EPITHELIA, LACTATE IS LIKELY 6470 04:06:15,383 --> 04:06:17,218 FEEDING INTO THE TCA CYCLE 6471 04:06:17,218 --> 04:06:18,386 ALTHOUGH WE'RE STILL WORKING ON 6472 04:06:18,386 --> 04:06:20,121 THOSE STUDIES AND THIS CIRCUIT 6473 04:06:20,121 --> 04:06:21,455 IS IN FACT UNDERPINNING ALL OF 6474 04:06:21,455 --> 04:06:23,124 THE OTHER PATHOLOGICAL FEATURES 6475 04:06:23,124 --> 04:06:25,493 OF THIS DISEASE AND COULD LIKELY 6476 04:06:25,493 --> 04:06:27,562 BE FUNCTIONING IN MANY, MANY 6477 04:06:27,562 --> 04:06:28,696 OTHER CHRONIC CONDITIONS WHERE 6478 04:06:28,696 --> 04:06:30,364 THE CIRCUIT IS LIKELY NOT GOING 6479 04:06:30,364 --> 04:06:32,600 TO BE BETWEEN EPITHELIA AND TYPE 6480 04:06:32,600 --> 04:06:34,368 17 BUT MANY OTHER TYPES OF 6481 04:06:34,368 --> 04:06:36,137 CELLS, FIBROBLASTS, ET CETERA, 6482 04:06:36,137 --> 04:06:37,572 AND IMMUNE CELLS. 6483 04:06:37,572 --> 04:06:38,773 SO I HOPE I'VE CONVINCED YOU AND 6484 04:06:38,773 --> 04:06:40,942 I TRIED TO STAY ON TIME, I CUT A 6485 04:06:40,942 --> 04:06:42,944 LOT OF SLIDES BECAUSE I'M AFRAID 6486 04:06:42,944 --> 04:06:47,748 OF AL, AND SO I HOPE I'VE 6487 04:06:47,748 --> 04:06:49,817 DEFENSED YOU THAT STUDYING 6488 04:06:49,817 --> 04:06:51,152 PHYSIOLOGY HAS A HUGE BENEFITED 6489 04:06:51,152 --> 04:06:52,787 IN UNDERSTANDING PATHOLOGY AND 6490 04:06:52,787 --> 04:06:54,889 CAN ACTUALLY TELL US 6491 04:06:54,889 --> 04:06:56,524 FUNDAMENTALS OF HOW OUR TISSUES 6492 04:06:56,524 --> 04:06:59,160 COPE WITH PERTUBATIONS AND THOSE 6493 04:06:59,160 --> 04:07:01,395 SAME FACTORS ARE COOPTED. 6494 04:07:01,395 --> 04:07:03,230 I DON'T THINK BIOLOGY IS 6495 04:07:03,230 --> 04:07:03,798 PARTICULARLY CREATIVE. 6496 04:07:03,798 --> 04:07:05,166 I THINK THEY JUST HIJACK WHAT'S 6497 04:07:05,166 --> 04:07:05,733 ALREADY THERE. 6498 04:07:05,733 --> 04:07:07,602 SO I'LL STOP THERE. 6499 04:07:07,602 --> 04:07:09,570 HAD AND THANK MY AMAZING LAB, IT 6500 04:07:09,570 --> 04:07:10,905 IS TRULY A PRIVILEGE AND 6501 04:07:10,905 --> 04:07:12,006 PLEASURE TO WORK WITH THEM, AND 6502 04:07:12,006 --> 04:07:13,107 I WILL TAKE ANY QUESTIONS IF 6503 04:07:13,107 --> 04:07:16,444 THERE ARE ANY. 6504 04:07:16,444 --> 04:07:23,384 [APPLAUSE] 6505 04:07:23,384 --> 04:07:25,119 >> VERY NICE TALK. 6506 04:07:25,119 --> 04:07:26,187 CAN YOU HEAR ME? 6507 04:07:26,187 --> 04:07:27,588 >> YES. 6508 04:07:27,588 --> 04:07:31,392 >> SO THE ARE TH17 CELLS DON'T 6509 04:07:31,392 --> 04:07:33,628 COME IN JUST ONE FLAVOR, RIGHT? 6510 04:07:33,628 --> 04:07:35,463 IN THEIR HOMEOSTATIC CONDITIONS 6511 04:07:35,463 --> 04:07:39,400 THAT YOU DESCRIBE, THE IL17 6512 04:07:39,400 --> 04:07:46,540 PRODUCING CELLS -- YOU STILL 6513 04:07:46,540 --> 04:07:48,709 MAINTAIN THE CONTEXT. 6514 04:07:48,709 --> 04:07:50,344 BUT IN THE PATHOLOGICAL 6515 04:07:50,344 --> 04:07:52,146 CONDITION LIKE PSORIASIS YOU 6516 04:07:52,146 --> 04:07:57,118 HAVE IL23 ACTING ON TL17 CELLS, 6517 04:07:57,118 --> 04:07:58,686 IT CHANGES THEIR METABOLIC 6518 04:07:58,686 --> 04:08:00,121 PROFILE AND FUNCTIONAL PROFILE 6519 04:08:00,121 --> 04:08:01,288 AS WELL. 6520 04:08:01,288 --> 04:08:04,492 SO IT MAY NOT BE JUST THAT THE 6521 04:08:04,492 --> 04:08:07,461 SAME CELL IS DOING BOTH BECAUSE 6522 04:08:07,461 --> 04:08:11,766 OF IL17 BECAUSE IF YOU LOOK AT 6523 04:08:11,766 --> 04:08:14,902 THE METABOLIC PROFILE USING 6524 04:08:14,902 --> 04:08:16,537 OTHERS, YOU WILL SEE THAT THE 6525 04:08:16,537 --> 04:08:19,040 METABOLISM OF HOMEOSTATIC TH17 6526 04:08:19,040 --> 04:08:22,076 CELLS IS VERY DIFFERENT FROM THE 6527 04:08:22,076 --> 04:08:24,178 PATHOGENIC TH17 CELLS. 6528 04:08:24,178 --> 04:08:25,646 AND HAVE YOU TRIED TO LOOK 6529 04:08:25,646 --> 04:08:28,249 WHETHER YOUR TH17 CELLS ACTUALLY 6530 04:08:28,249 --> 04:08:30,651 MAKE LACTATE AND IT GOES BACK 6531 04:08:30,651 --> 04:08:32,053 ONTO THE EPITHELIA? 6532 04:08:32,053 --> 04:08:34,155 >> YES. 6533 04:08:34,155 --> 04:08:36,323 SO FIRST, I TOTALLY AGREE WITH 6534 04:08:36,323 --> 04:08:36,490 YOU. 6535 04:08:36,490 --> 04:08:38,092 I THINK THAT WE'RE SO OBSESSED 6536 04:08:38,092 --> 04:08:39,593 WITH SUBSETTING CELLS AND I AM 6537 04:08:39,593 --> 04:08:41,395 MUCH MORE INTERESTED IN SORT OF 6538 04:08:41,395 --> 04:08:43,931 THE END FEEK TORE PROGRAMS LIKE 6539 04:08:43,931 --> 04:08:44,131 IL17. 6540 04:08:44,131 --> 04:08:46,000 IN THE CASE OF THE MOUSE MODELS 6541 04:08:46,000 --> 04:08:48,936 I'VE SHOWN YOU, GAMMA DELTA 17'S 6542 04:08:48,936 --> 04:08:52,473 ARE THE MOST QUOTE, UNQUOTE, 6543 04:08:52,473 --> 04:08:52,773 PREDOMINANT. 6544 04:08:52,773 --> 04:08:54,308 I THINK IN HUMAN TISSUES IT'S A 6545 04:08:54,308 --> 04:08:55,843 VERY DIFFERENT GAME, SO THE 6546 04:08:55,843 --> 04:08:57,478 SUBSET ISSUE IS I THINK MUCH 6547 04:08:57,478 --> 04:08:59,447 MORE AN ABERRATION OF HOW WE 6548 04:08:59,447 --> 04:09:02,883 WRITE PAPERS THAN BIOLOGY. 6549 04:09:02,883 --> 04:09:05,119 SO I 100% AGREE WITH YOU. 6550 04:09:05,119 --> 04:09:07,655 >> CAN I JUST SAY FOR JUST A 6551 04:09:07,655 --> 04:09:08,723 MINUTE -- CAN I JUST DISAGREE 6552 04:09:08,723 --> 04:09:10,057 WITH YOU FOR JUST A MINUTE? 6553 04:09:10,057 --> 04:09:11,592 >> WE'LL DISGREERKS THAT'S. 6554 04:09:11,592 --> 04:09:13,327 >> IF YOU REALLY LOOK AT THE 6555 04:09:13,327 --> 04:09:14,328 HOMEOSTATIC CONDITIONS EVEN IN 6556 04:09:14,328 --> 04:09:18,833 HUMANS IN THE GUT, YOU HAVE 17 6557 04:09:18,833 --> 04:09:20,167 AND TEN PRODUCING CELLS IN THE 6558 04:09:20,167 --> 04:09:21,235 GUT, THOSE ARE THE MAJORITY OF 6559 04:09:21,235 --> 04:09:23,204 THE CELLS IN THE GUT, UNLESS 6560 04:09:23,204 --> 04:09:26,841 THEY GET ACTED BOYNE IL23, THEY 6561 04:09:26,841 --> 04:09:28,876 DON'T BECOME PATHOGENIC. 6562 04:09:28,876 --> 04:09:31,512 THAT'S WHY ENTER IL23 WORKS SO 6563 04:09:31,512 --> 04:09:34,281 WELL IN PSORIASIS AND COLITIS, 6564 04:09:34,281 --> 04:09:36,751 IN FACT, THEY COULDN'T FIND THE 6565 04:09:36,751 --> 04:09:41,689 END DOSE, IL17 WORKS, BUTLY IL23 6566 04:09:41,689 --> 04:09:45,860 SITS OWN DATA, SO IT'S NOT JUST 6567 04:09:45,860 --> 04:09:47,261 ABERRATION OF ACTUALLY 6568 04:09:47,261 --> 04:09:47,862 SUBSETTING. 6569 04:09:47,862 --> 04:09:48,996 IN FACT, IF YOU LOOK AT OUR DATA 6570 04:09:48,996 --> 04:09:52,266 FROM THE MOUSE AND THOSE DATA 6571 04:09:52,266 --> 04:09:54,235 FROM HUMANS, PROFILE IS EXACTLY 6572 04:09:54,235 --> 04:09:56,237 THE SAME. 6573 04:09:56,237 --> 04:09:58,739 IN AND IN CASE OF CAN TI DA 6574 04:09:58,739 --> 04:10:01,142 ALBICANS YOU HAVE PATHOGENIC 17 6575 04:10:01,142 --> 04:10:03,344 CELLS N CASE OF STAPH AUREUS IN 6576 04:10:03,344 --> 04:10:05,813 HUMANS, YOU HAVE HOMEOSTATIC 17. 6577 04:10:05,813 --> 04:10:06,647 >> YES, I UNDERSTAND YOUR 6578 04:10:06,647 --> 04:10:06,914 QUESTION. 6579 04:10:06,914 --> 04:10:08,516 WHAT I'M SAYING IS THE SUBSETS 6580 04:10:08,516 --> 04:10:10,351 ARE VERY DIFFERENT DURING 6581 04:10:10,351 --> 04:10:13,187 BASELINE AND SO IL23 MAY BE 6582 04:10:13,187 --> 04:10:14,188 ENRICHING FOR DIFFERENT SUBSET 6583 04:10:14,188 --> 04:10:14,755 OF CELLS. 6584 04:10:14,755 --> 04:10:16,757 FOR INSTANCE, IN THE SKIN, THE 6585 04:10:16,757 --> 04:10:18,793 DOMINANT PRODUCERS OF IL17 ARE 6586 04:10:18,793 --> 04:10:20,561 NOT TH17 CELLS. 6587 04:10:20,561 --> 04:10:22,129 THEY ARE MATE CELLS, GAMMA 6588 04:10:22,129 --> 04:10:23,964 DELTA, THAT'S WHAT I'M SAYING, 6589 04:10:23,964 --> 04:10:25,699 THERE'S ALSO TISSUE-SPECIFIC 6590 04:10:25,699 --> 04:10:26,000 DIFFERENCES. 6591 04:10:26,000 --> 04:10:28,235 I DO THINK THAT WE HAVEN'T 6592 04:10:28,235 --> 04:10:30,604 TESTED FOR LACTATE PRODUCED FROM 6593 04:10:30,604 --> 04:10:31,906 TH17 CELLS AND AT BASELINE I 6594 04:10:31,906 --> 04:10:32,973 KNOW AND YOU'VE DONE BEAUTIFUL 6595 04:10:32,973 --> 04:10:35,509 WORK IN THIS WHERE THEY ARE 6596 04:10:35,509 --> 04:10:36,911 GLYCOLYTIC AND MAKING 6597 04:10:36,911 --> 04:10:38,679 GLYCOLYSIS, WHEREAS IN THIS 6598 04:10:38,679 --> 04:10:39,780 INFLAMMATORY CONDITION WE THINK 6599 04:10:39,780 --> 04:10:41,649 THEY'RE TURNING ON THE TCA 6600 04:10:41,649 --> 04:10:43,818 CYCLE, THAT MAY BE DEPENDENT ON 6601 04:10:43,818 --> 04:10:44,018 IL23. 6602 04:10:44,018 --> 04:10:45,419 >> I LOVED YOUR WORK. 6603 04:10:45,419 --> 04:10:47,555 >> NO, NO, I THINK WE'RE 6604 04:10:47,555 --> 04:10:48,122 AGREERKS RIGHT? 6605 04:10:48,122 --> 04:10:48,923 I THINK WE'RE AGREEING. 6606 04:10:48,923 --> 04:10:50,291 >> THANK YOU FOR THE ANSWER. 6607 04:10:50,291 --> 04:10:51,592 BUT ACTUALLY VERY NICE WORK. 6608 04:10:51,592 --> 04:10:53,327 >> WE'LL TALK LATER, BECAUSE YOU 6609 04:10:53,327 --> 04:10:56,497 CLEARLY DISAGREE WITH ME STILL. 6610 04:10:56,497 --> 04:10:56,764 [LAUGHTER] 6611 04:10:56,764 --> 04:11:00,134 >> DIDN'T INTRODUCE HIMSELF, 6612 04:11:00,134 --> 04:11:02,369 THAT'S VIJAY. 6613 04:11:02,369 --> 04:11:06,073 >> HI, SHRUTI. 6614 04:11:06,073 --> 04:11:06,907 NICE WORK. 6615 04:11:06,907 --> 04:11:12,046 I HAVE ONE QUESTION. 6616 04:11:12,046 --> 04:11:17,685 I'M R YOU TI, I'M A RESEARCH 6617 04:11:17,685 --> 04:11:20,054 FELLOW IN VANOSH'S LAB AND I 6618 04:11:20,054 --> 04:11:21,856 HAVE TWO QUESTIONS. 6619 04:11:21,856 --> 04:11:26,227 FIRST IS, SAY THAT IL17 ACTS 6620 04:11:26,227 --> 04:11:27,428 SECONDARY SIGNAL IN STABILIZING 6621 04:11:27,428 --> 04:11:29,563 HERE AND THEN DRIVING 6622 04:11:29,563 --> 04:11:32,933 GLYCOLYSIS, BUT DO YOU KNOW HOW 6623 04:11:32,933 --> 04:11:35,870 MUCH IS INDEPENDENT THAT CAN 6624 04:11:35,870 --> 04:11:36,971 IL17 DRIVE. 6625 04:11:36,971 --> 04:11:38,505 >> GLYCOLYSIS INDEPENDENT OF 6626 04:11:38,505 --> 04:11:43,210 HERE, FIRST? 6627 04:11:43,210 --> 04:11:49,149 AND CAN IL17 PRODUCE IT? 6628 04:11:49,149 --> 04:11:51,719 THE ROLE OF IL23'S TO COMPLETELY 6629 04:11:51,719 --> 04:11:52,653 RULE THEM OUT? 6630 04:11:52,653 --> 04:11:55,823 LIKE HAVE YOU TESTED IN RAG 6631 04:11:55,823 --> 04:11:56,824 KNOCK-OUT. 6632 04:11:56,824 --> 04:11:58,659 >> FIRST, I WILL SAY, SO WE 6633 04:11:58,659 --> 04:12:00,394 DON'T THINK THAT IN EPITHELIAL 6634 04:12:00,394 --> 04:12:02,263 CELLS WHEN YOU TREAT WITH IL17 6635 04:12:02,263 --> 04:12:04,098 YOU NEED HIF1 ALPHA. 6636 04:12:04,098 --> 04:12:08,168 THE HIF1 HAD ALPHA KNOCKOUTS 6637 04:12:08,168 --> 04:12:08,802 DON'T INDUCE. 6638 04:12:08,802 --> 04:12:09,904 I KNOW THERE'S OTHER GENES TO 6639 04:12:09,904 --> 04:12:12,206 TURN ON, LIGAND, THAT'S NOT 6640 04:12:12,206 --> 04:12:13,574 OPERATIVE IN THIS CONTEXT. 6641 04:12:13,574 --> 04:12:14,875 DISHEANLT MEAN IT NEVER HAPPENS. 6642 04:12:14,875 --> 04:12:16,010 IN THE ORIGINAL PAPER WE 6643 04:12:16,010 --> 04:12:19,546 ACTUALLY TESTED EVERY POSSIBLE 6644 04:12:19,546 --> 04:12:23,284 GAMMA T DEFICIENT MOUSE, GAMMA 6645 04:12:23,284 --> 04:12:27,755 DELTA'S MATES NTH17'S HAD, ILC'S 6646 04:12:27,755 --> 04:12:29,490 DWE WITH JUST RAG MICE AND SEEMS 6647 04:12:29,490 --> 04:12:31,725 TO COME DOWN TO GAMMA DELTAS IN 6648 04:12:31,725 --> 04:12:31,926 MATES. 6649 04:12:31,926 --> 04:12:33,360 AGAIN THIS COMES BACK TO THIS 6650 04:12:33,360 --> 04:12:34,628 ABERRATION OF MOUSE EXPIN HOW 6651 04:12:34,628 --> 04:12:36,297 IT'S VERY DIFFERENT THAN HUMAN 6652 04:12:36,297 --> 04:12:40,434 SKIN AND JUST THE CELLULAR. 6653 04:12:40,434 --> 04:12:41,068 PLAYER PLAYERS DRIRCHT. 6654 04:12:41,068 --> 04:12:42,870 I WOULD LIKE TO FOCUS -- PLAYERS 6655 04:12:42,870 --> 04:12:43,504 ARE DIFFERENT. 6656 04:12:43,504 --> 04:12:45,005 I WOULD LIKE TO REALLY FOCUS ON 6657 04:12:45,005 --> 04:12:45,706 THE END-PRODUCT. 6658 04:12:45,706 --> 04:12:47,141 >> THANK YOU. 6659 04:12:47,141 --> 04:12:48,509 I'M VALENTINA, ALSO WORKING ON 6660 04:12:48,509 --> 04:12:51,278 THE SKIN, I'M IN THE GROUP CR 6661 04:12:51,278 --> 04:12:54,448 WITH CHEN HERE AT NIH, I'M A 6662 04:12:54,448 --> 04:12:55,149 PH.D. STUDENT. 6663 04:12:55,149 --> 04:12:57,117 I ALSO WORK ON PSORIASIS AND I 6664 04:12:57,117 --> 04:12:59,753 HAVE TO CONFIRM THAT IN MICE 6665 04:12:59,753 --> 04:13:01,588 MODELS IT'S THE MAJOR 17 6666 04:13:01,588 --> 04:13:04,458 PRODUCER ARE GAMMA DELTA T 6667 04:13:04,458 --> 04:13:04,792 CELLS. 6668 04:13:04,792 --> 04:13:07,761 SO MY QUESTION WOULD RATHER BE 6669 04:13:07,761 --> 04:13:10,698 IN STEADY STATE CONDITION, SO 6670 04:13:10,698 --> 04:13:12,633 WHEN YOU WOUND THE MICE, IF ALSO 6671 04:13:12,633 --> 04:13:14,435 THEY ARE THE GAMMA DELTAS ARE 6672 04:13:14,435 --> 04:13:17,938 THE MAJOR IL17 PRODUCER, AND 6673 04:13:17,938 --> 04:13:19,707 THEN MY QUESTION WOULD BE, HOW 6674 04:13:19,707 --> 04:13:22,409 DOES IT COME HAD FOR THEM TO 6675 04:13:22,409 --> 04:13:23,010 PRODUCE 17? 6676 04:13:23,010 --> 04:13:26,447 IS IT BECAUSE BY GENERATING THE 6677 04:13:26,447 --> 04:13:29,316 WOUND YOU ALLOW THE BACTERIA TO 6678 04:13:29,316 --> 04:13:30,851 ENTER THE SYSTEM AND THEN YOU 6679 04:13:30,851 --> 04:13:33,520 HAVE THIS SORT OF REACTION, OR 6680 04:13:33,520 --> 04:13:36,123 DO YOU THINK IT'S MOSTLY BECAUSE 6681 04:13:36,123 --> 04:13:39,159 OF THE EPITHELIAL CELLS DID IN 6682 04:13:39,159 --> 04:13:40,294 INDEED GET DAMAGED AND THEN 6683 04:13:40,294 --> 04:13:42,863 START THIS CASCADE THAT THEN 6684 04:13:42,863 --> 04:13:44,732 PRODUCES OR INDUCES 17 6685 04:13:44,732 --> 04:13:45,032 PRODUCTION? 6686 04:13:45,032 --> 04:13:46,600 >> YEAH, I MEAN, I THINK THAT 6687 04:13:46,600 --> 04:13:48,602 THOSE THINGS ARE NOT MUTE 6688 04:13:48,602 --> 04:13:50,004 YOU'LLLY EXCLUSIVEMENT RIGHT 6689 04:13:50,004 --> 04:13:54,241 SNING THEY'RE PROBABLY ARE NOT 6690 04:13:54,241 --> 04:13:55,142 MUTUALLY EXCLUSIVE. 6691 04:13:55,142 --> 04:13:55,342 RIGHT? 6692 04:13:55,342 --> 04:13:57,211 I THINK THEY'RE PROBABLY 6693 04:13:57,211 --> 04:13:58,178 HAPPENING AT THE SAME TIME. 6694 04:13:58,178 --> 04:14:01,281 I THINK YOU'RE SPLITTING HAIRS. 6695 04:14:01,281 --> 04:14:04,885 IN MICE, GAMMA DELTAS, UNLESS AS 6696 04:14:04,885 --> 04:14:06,620 HAS SHOWN THEY RECEIVE THE RIGHT 6697 04:14:06,620 --> 04:14:09,390 MATE CELL DEVELOPING SIGNAL. 6698 04:14:09,390 --> 04:14:11,458 SO YOU CAN HAVE COMPENSATION 6699 04:14:11,458 --> 04:14:13,627 FROM MATE CELLS IF YOU GIVE MICE 6700 04:14:13,627 --> 04:14:15,496 THE RIGHT SIGNALS TO DEVELOP 6701 04:14:15,496 --> 04:14:17,398 MATE CELLS AS WE DO IN OUR SKIN 6702 04:14:17,398 --> 04:14:18,699 BECAUSE WE GET THE RIGHT SIGNALS 6703 04:14:18,699 --> 04:14:20,334 AT THE RIGHT TIME. 6704 04:14:20,334 --> 04:14:26,140 SO AGAIN SPF MICE JUST DIFFERENT 6705 04:14:26,140 --> 04:14:26,373 BEASTS. 6706 04:14:26,373 --> 04:14:27,241 >> THANK YOU. 6707 04:14:27,241 --> 04:14:28,142 >> THAT WAS GREAT. 6708 04:14:28,142 --> 04:14:30,110 THANK YOU VERY, VERY MUCH. 6709 04:14:30,110 --> 04:14:31,178 SORRY, I THINK IN THE BACK THEY 6710 04:14:31,178 --> 04:14:35,482 DON'T HEAR US IF YOU DON'T TALK 6711 04:14:35,482 --> 04:14:37,151 DIRECTLY INTO THE MIC. 6712 04:14:37,151 --> 04:14:39,453 SPECTACULAR TALK. 6713 04:14:39,453 --> 04:14:42,322 IN TERMS OF YOUR KNOCKOUTS OF 6714 04:14:42,322 --> 04:14:44,224 HIF1 ALPHA AND OF GLUT 1, DO 6715 04:14:44,224 --> 04:14:46,927 THEY HAVE SIMILAR EFFECTS ON THE 6716 04:14:46,927 --> 04:14:47,795 PROLIFERATIVE STATUS OF THE 6717 04:14:47,795 --> 04:14:50,064 CELLS, AND IN THAT CONTEXT, YOU 6718 04:14:50,064 --> 04:14:52,266 REALLY TALK ABOUT THE EFFECT OF 6719 04:14:52,266 --> 04:14:54,301 GLUT 1 IN TERMS OF LACTATE. 6720 04:14:54,301 --> 04:14:54,868 BUT DO YOU KNOW APPROXIMATE 6721 04:14:54,868 --> 04:14:55,869 THERE ARE DIFFERENCES IN TERMS 6722 04:14:55,869 --> 04:14:57,438 OF NUCLEOTIDE POOLS? 6723 04:14:57,438 --> 04:14:59,606 >> OH, WEFNLT LOOKED, WE HAVE 6724 04:14:59,606 --> 04:15:04,078 ACTUALLY NOT ASSESSED THAT THE . 6725 04:15:04,078 --> 04:15:05,279 THAT'S A REALLY IMPORTANT POINT 6726 04:15:05,279 --> 04:15:07,347 BECAUSE THERE IS A HUGE 6727 04:15:07,347 --> 04:15:08,582 PROLIFERATIVE EFFECT AS YOU 6728 04:15:08,582 --> 04:15:09,349 ALLUDED AND I DON'T KNOW HOW 6729 04:15:09,349 --> 04:15:11,318 MUCH OF IT IS BECAUSE IT'S 6730 04:15:11,318 --> 04:15:12,086 DEPLETED NUCLEOTIDE POOLS. 6731 04:15:12,086 --> 04:15:13,387 >> IS IT THE SAME IF YOU KNOCK 6732 04:15:13,387 --> 04:15:16,390 OUT HIF1 FAL OR GLUT 1? 6733 04:15:16,390 --> 04:15:18,826 >> IN TERMS OFLY PROLIFERATION, 6734 04:15:18,826 --> 04:15:18,992 YES. 6735 04:15:18,992 --> 04:15:19,927 >> IT'S EQUIVALENT AND. 6736 04:15:19,927 --> 04:15:22,162 >> IT'S QUIF ZEENT WE CAN TALK 6737 04:15:22,162 --> 04:15:23,230 LATER BUT IT WOULD BE 6738 04:15:23,230 --> 04:15:24,364 INTERESTING TO SEE WHAT THE 6739 04:15:24,364 --> 04:15:25,632 PERCENTAGE IMPACT IS OF LACK 6740 04:15:25,632 --> 04:15:29,269 TAILLIGHT VERSUS NUCLEOTIDE 6741 04:15:29,269 --> 04:15:29,503 POOLSES. 6742 04:15:29,503 --> 04:15:31,572 >> YEAH, WE JUST HAVEN'T EVEN 6743 04:15:31,572 --> 04:15:33,440 CONSIDERED T SO IT'S IT'S A 6744 04:15:33,440 --> 04:15:35,109 REALLY GOOD POINT. 6745 04:15:35,109 --> 04:15:39,379 >> JAY ZABROSKI FROM NCLI, I 6746 04:15:39,379 --> 04:15:40,414 HAVE MAY HAVE MISSED THIS, IS 6747 04:15:40,414 --> 04:15:43,283 THERE A ROLE FOR RGF1 IN THE 6748 04:15:43,283 --> 04:15:45,552 METABOLIC EFFECTS OF HIF1 ALPHA 6749 04:15:45,552 --> 04:15:46,253 THAT YOU DESCRIBE? 6750 04:15:46,253 --> 04:15:47,988 >> THAT'S NOT ONE OF THE FACTORS 6751 04:15:47,988 --> 04:15:50,390 WE SEE DIFFERENTLY EXPRESSED. 6752 04:15:50,390 --> 04:15:52,126 SO I REALLY CAN'T COMMENT ON IF 6753 04:15:52,126 --> 04:15:56,296 THERE IS OR ISN'T AS FAR AS -- 6754 04:15:56,296 --> 04:15:57,698 BUT THAT'S WE DWHROOND BECAUSE 6755 04:15:57,698 --> 04:15:59,032 WE JUST HAVEN'T TESTED IT. 6756 04:15:59,032 --> 04:16:00,701 >> OKAY, BECAUSE IT HAS BEEN 6757 04:16:00,701 --> 04:16:04,271 ASSOCIATED WITH HIF1 EFFECTS SO 6758 04:16:04,271 --> 04:16:06,573 CAN OBVIOUSLY INFLUENCE GLUCOSE 6759 04:16:06,573 --> 04:16:07,141 METABOLISM AND OTHERS. 6760 04:16:07,141 --> 04:16:08,542 >> THAT'S A GOOD POINT. 6761 04:16:08,542 --> 04:16:09,877 ESSENTIALLY THE WAY WE DID THIS 6762 04:16:09,877 --> 04:16:12,146 IS WE TOOK A VERY LET THE DATA 6763 04:16:12,146 --> 04:16:13,914 TELL US WHERE TO GO, AND THE 6764 04:16:13,914 --> 04:16:15,983 GENES THAT WERE THE MOST 6765 04:16:15,983 --> 04:16:17,184 DIFFERENT WERE GLYCOLYSIS, 6766 04:16:17,184 --> 04:16:18,619 THAT'S KIND WHAF WE WENT AFTER. 6767 04:16:18,619 --> 04:16:20,254 WE DIDN'T SEE GENES THAT WERE 6768 04:16:20,254 --> 04:16:22,789 DIFFERENT IN IGF SIGNALLING OR 6769 04:16:22,789 --> 04:16:25,292 THE IGF RECEPTOR, BUT THAT 6770 04:16:25,292 --> 04:16:26,827 DOESN'T MEAN IT'S NOT -- I THINK 6771 04:16:26,827 --> 04:16:28,462 WE JUST HAVEN'T EXPLORED IT. 6772 04:16:28,462 --> 04:16:30,063 THANKS GROO THANK YOU. 6773 04:16:30,063 --> 04:16:33,066 >> THANK YOU. 6774 04:16:33,066 --> 04:16:38,472 [APPLAUSE] 6775 04:16:38,472 --> 04:16:41,542 >> SO I GUESS WE'RE STAYING WITH 6776 04:16:41,542 --> 04:16:43,377 OUR 17 PRODUCING CELLS FOR A B 6777 04:16:43,377 --> 04:16:46,013 OUR NEXT SPEAKER IS VAN 6778 04:16:46,013 --> 04:16:48,815 GENTLEMAN LAZ VICK FROM -- OUR 6779 04:16:48,815 --> 04:16:54,755 NEXT SPEAKER IS VAN GENTLEMAN 6780 04:16:54,755 --> 04:16:55,055 LAZAREVIC,. 6781 04:16:55,055 --> 04:16:56,857 >> THAT'S A PERFECT INTRODUCE 6782 04:16:56,857 --> 04:16:57,824 FOR MY TALK TODAY. 6783 04:16:57,824 --> 04:16:58,892 I'M DELIGHTED TO BE HERE AND 6784 04:16:58,892 --> 04:17:00,427 SHARE WITH YOU OUR MOST RECENT 6785 04:17:00,427 --> 04:17:02,196 STUDY ON THE ROLE OF THE 6786 04:17:02,196 --> 04:17:04,298 TRANSCRIPTION FACTOR ERRING 2 IN 6787 04:17:04,298 --> 04:17:09,102 CONTROLLING -- EGR2 IN 6788 04:17:09,102 --> 04:17:12,406 CONTROLLINGLESS TH17 CELLS IN P 6789 04:17:12,406 --> 04:17:16,443 THE CNS COMPARTMENT. 6790 04:17:16,443 --> 04:17:17,744 THEY SPECIALIZE INTO SUB SETSZ 6791 04:17:17,744 --> 04:17:19,613 UPON CYTOKINE STIMULATION, EACH 6792 04:17:19,613 --> 04:17:21,615 GUIDED BY THEIR LINEAGE DEFINING 6793 04:17:21,615 --> 04:17:23,116 TRANSCRIPTION FACTORS THAT 6794 04:17:23,116 --> 04:17:25,552 CONTROL UNIQUE GENE EXPRESSION 6795 04:17:25,552 --> 04:17:26,753 PROGRAMS. 6796 04:17:26,753 --> 04:17:29,590 WHILE THESE T HELPER CELLS ARE 6797 04:17:29,590 --> 04:17:34,728 INSTRUMENTAL IN CLEARING 6798 04:17:34,728 --> 04:17:38,465 PATHOGENS, SO THIS IS BEST 6799 04:17:38,465 --> 04:17:43,203 EXEMPLIFIED BY TH17 CELLS. 6800 04:17:43,203 --> 04:17:45,339 NOW, IN NORMAL TISSUES NORMAL 6801 04:17:45,339 --> 04:17:46,807 STEADY STATE FOUND HAD BARRIER 6802 04:17:46,807 --> 04:17:47,874 TISSUES WHERE THEY PROTECT US 6803 04:17:47,874 --> 04:17:49,610 FROM FUNGAL AND BACTERIAL 6804 04:17:49,610 --> 04:17:51,378 INFECTIONS, THEY ALSO MEDIATE 6805 04:17:51,378 --> 04:17:53,280 TISSUE HOMOEOSTASIS AND THEY CAN 6806 04:17:53,280 --> 04:17:56,783 ALSO PROMOTE WOUND HEALING. 6807 04:17:56,783 --> 04:17:59,486 THEY ARE -- THEIR PROTOTYPICAL 6808 04:17:59,486 --> 04:18:01,455 HAD FUNCTIONS MEDIATED BY 6809 04:18:01,455 --> 04:18:04,191 CYTOKINES, SUCH AS IL17 AND 22, 6810 04:18:04,191 --> 04:18:07,060 PATIENTS WHO HAVE IMPAIRED TL17 6811 04:18:07,060 --> 04:18:10,631 RESPONSES OFTEN SUFFER FROM 6812 04:18:10,631 --> 04:18:13,600 CHRONIC MUCOSAL CANDIDIASIS CIS 6813 04:18:13,600 --> 04:18:16,570 AND ALSO SKIN INFECTIONS, BUT 6814 04:18:16,570 --> 04:18:17,938 ALSO THEY HAVE BEEN IMPLICATED 6815 04:18:17,938 --> 04:18:19,873 AS KEY DRIVERS OF INFLAMMATORY 6816 04:18:19,873 --> 04:18:21,375 DISEASES AND AUTOIMMUNE DISEASES 6817 04:18:21,375 --> 04:18:25,545 AND DESPITE THIS EXTENSIVE 6818 04:18:25,545 --> 04:18:28,382 RESEARCH INTO TH17 CELL BIOLOGY 6819 04:18:28,382 --> 04:18:29,583 THERE'S SO MUCH TO BE LEARNED 6820 04:18:29,583 --> 04:18:30,917 ABOUT THE TRANSCRIPTIONAL 6821 04:18:30,917 --> 04:18:31,985 REGULATORS OF TH17 CELLS AND 6822 04:18:31,985 --> 04:18:34,288 ALSO THE MECHANISMS THAT 6823 04:18:34,288 --> 04:18:35,956 UNDERLIE TISSUE PATHOLOGY AND 6824 04:18:35,956 --> 04:18:38,025 DIFFERENT TYPES OF AUTOIMMUNE 6825 04:18:38,025 --> 04:18:38,592 DISEASES. 6826 04:18:38,592 --> 04:18:41,795 SO FOR EXAMPLE, IN PSORIASIS, 6827 04:18:41,795 --> 04:18:46,033 TH17 CELLS SECRETE IL22 THAT 6828 04:18:46,033 --> 04:18:51,138 TRIGGERS HYPERPLASIA CRLS BY 6829 04:18:51,138 --> 04:18:53,540 ACTIVATING STAT 3, WHILE IN 6830 04:18:53,540 --> 04:18:57,311 RHEUMATOID ARTHRITIS, INDUCE 6831 04:18:57,311 --> 04:19:00,781 RANK LIGAND AND MIE BRO BLAST 6832 04:19:00,781 --> 04:19:02,516 AND IS AGGRAVATE LOCAL 6833 04:19:02,516 --> 04:19:04,418 INFLAMMATION BY PRODUCING 6834 04:19:04,418 --> 04:19:06,586 INFLAMMATORY CYTOKINES N MOUSE 6835 04:19:06,586 --> 04:19:08,689 MODELS WHERE TH17 CELLS DRIVE 6836 04:19:08,689 --> 04:19:09,589 INFLAMMATORY BOWEL DISEASE 6837 04:19:09,589 --> 04:19:11,091 THERE'S A VERY IMPORTANT 6838 04:19:11,091 --> 04:19:14,828 TRANSITION OF TH17 CELLS INTO 6839 04:19:14,828 --> 04:19:17,831 TH1 LIKE CELLS CHARACTERIZED BY 6840 04:19:17,831 --> 04:19:20,100 EXPRESSION OF GAMMA AND THESE 6841 04:19:20,100 --> 04:19:22,002 CYTOKINES DRIVE PATHOLOGY. 6842 04:19:22,002 --> 04:19:23,870 JUST LIKE INFLAMMATORY BOWEL 6843 04:19:23,870 --> 04:19:25,839 DISEASE, THIS FUNCTIONAL 6844 04:19:25,839 --> 04:19:27,674 PLASTICITY OF TH17 CELLS ALSO 6845 04:19:27,674 --> 04:19:29,910 CONTRIBUTES TO NEUROPATHOLOGY IN 6846 04:19:29,910 --> 04:19:30,143 MS. 6847 04:19:30,143 --> 04:19:32,746 SO IN MY LAB WE TRY TO 6848 04:19:32,746 --> 04:19:36,783 UNDERSTAND HOW TH17 CELLS 6849 04:19:36,783 --> 04:19:37,784 PROMOTE NEUROINFLAMMATION, 6850 04:19:37,784 --> 04:19:39,419 SPECIFICALLY WE WANT TO IDENTIFY 6851 04:19:39,419 --> 04:19:40,954 TRANSCRIPTIONAL REGULATORS THAT 6852 04:19:40,954 --> 04:19:42,689 CONTROL DEVELOPMENT OF 6853 04:19:42,689 --> 04:19:47,194 PATHOGENIC TH17 CELLS IN THE CNS 6854 04:19:47,194 --> 04:19:47,527 COMPARTMENT. 6855 04:19:47,527 --> 04:19:49,396 SO IN LINE WITH THESE INQUIRIES 6856 04:19:49,396 --> 04:19:50,263 THERE ARE SEVERAL IMPORTANT 6857 04:19:50,263 --> 04:19:51,798 QUESTIONS THAT EMERGE. 6858 04:19:51,798 --> 04:19:53,734 FIRST K WE IDENTIFY 6859 04:19:53,734 --> 04:19:56,136 TRANSCRIPTIONAL REGULATORS OF 6860 04:19:56,136 --> 04:19:57,904 PATHOGENIC TH17 CELLS AND CAN WE 6861 04:19:57,904 --> 04:20:00,006 TARG THE PATHOGENIC EFFECTOR 6862 04:20:00,006 --> 04:20:02,409 PROGRAM OF TH17 CELLS WITHOUT 6863 04:20:02,409 --> 04:20:04,478 INTERFERING WITH THEIR 6864 04:20:04,478 --> 04:20:07,214 PROTECTIVE BARRIER FUNCTION IN 6865 04:20:07,214 --> 04:20:07,614 TISSUES? 6866 04:20:07,614 --> 04:20:10,117 SO TO TRY TO UNDERSTAND THE 6867 04:20:10,117 --> 04:20:11,852 TRANSCRIPTIONAL PROGRAM OF 6868 04:20:11,852 --> 04:20:15,155 PATHOGENIC TH17 CELLS, WE 6869 04:20:15,155 --> 04:20:16,556 COMPARED THE TRANSCRIPTIONAL 6870 04:20:16,556 --> 04:20:18,725 PROGRAM OF MEMORY CD4 T CELLS 6871 04:20:18,725 --> 04:20:21,795 THAT WERE CHARACTERIZED AS BEING 6872 04:20:21,795 --> 04:20:23,230 MILDLY REACTIVE OR AT THE TIME 6873 04:20:23,230 --> 04:20:27,501 MEASURE POSITIVE OR MYELIN 6874 04:20:27,501 --> 04:20:29,603 NONREACTIVE OR AT THE TIME TROA 6875 04:20:29,603 --> 04:20:31,872 MERE NEGATIVE, FROM HEALTHY 6876 04:20:31,872 --> 04:20:33,273 CONTROLS AND MS PATIENTS. 6877 04:20:33,273 --> 04:20:36,143 AS YOU CAN SEE HERE MYELIN 6878 04:20:36,143 --> 04:20:38,011 REACTIVE CD4 T CELLS WERE 6879 04:20:38,011 --> 04:20:39,212 CHARACTERIZED BY ELEVATED 6880 04:20:39,212 --> 04:20:40,747 SPREFTION EGR FAMILY OF 6881 04:20:40,747 --> 04:20:43,283 TRANSCRIPTION FACTORS, EGL1, 6882 04:20:43,283 --> 04:20:45,218 2-RBGS APPROXIMATE AND 4 DPIERD 6883 04:20:45,218 --> 04:20:47,154 THE MYELIN NONREACTIVE T CELLS 6884 04:20:47,154 --> 04:20:49,556 FROM THE SAME MS PATIENT. 6885 04:20:49,556 --> 04:20:53,326 WE CAN SEE HERE IF REALLY HIGHLY 6886 04:20:53,326 --> 04:20:53,760 UPREGULATED. 6887 04:20:53,760 --> 04:20:56,062 WHEN WE PERFORMED CLUSTERING 6888 04:20:56,062 --> 04:20:58,565 ANALYSIS INVOLVING PATHOGENICITY 6889 04:20:58,565 --> 04:21:01,401 ASSOCIATED GENES WITH EGR FAMILY 6890 04:21:01,401 --> 04:21:03,203 TRANSCRIPTION FACTORS, AN 6891 04:21:03,203 --> 04:21:04,738 INTERESTING PATTERN EMERGED. 6892 04:21:04,738 --> 04:21:07,441 SO THESE MYELIN REABLGHTTIVE CD4 6893 04:21:07,441 --> 04:21:09,910 T CELLS FROM MS PATIENTS THEY 6894 04:21:09,910 --> 04:21:12,813 CLUSTER SEPARATELY FROM MYELIN 6895 04:21:12,813 --> 04:21:14,748 NONREACTIVE CD4 T CELLS FROM MS 6896 04:21:14,748 --> 04:21:16,283 PATIENTS AND ALSO HEALTHY 6897 04:21:16,283 --> 04:21:17,918 CONTROL, SUGGESTING THAT PERHAPS 6898 04:21:17,918 --> 04:21:20,353 THERE IS ASSOCIATION BETWEEN EGR 6899 04:21:20,353 --> 04:21:21,955 FAMILY TRANSCRIPTION FACTORS AND 6900 04:21:21,955 --> 04:21:24,224 THE PATHOGENIC FUNCTIONS OF CD4 6901 04:21:24,224 --> 04:21:27,194 T CELLS IN MS. 6902 04:21:27,194 --> 04:21:37,170 WE ALSO DECIDED TO LOOK P AT -- 6903 04:21:37,170 --> 04:21:40,106 INDUPSED BY MICROBES IN THE GUT, 6904 04:21:40,106 --> 04:21:42,409 SUCH AS BACTERIUM, SO WE WANTED 6905 04:21:42,409 --> 04:21:44,177 TO EXPLORE THIS DIFFERENTIAL 6906 04:21:44,177 --> 04:21:46,913 EXPRESSION OF EGR FAMILY MEMBERS 6907 04:21:46,913 --> 04:21:49,616 IN HOMEOSTATIC TH17 CELLS 6908 04:21:49,616 --> 04:21:52,285 ISOLATED FROM THE LAMINA PROPRIA 6909 04:21:52,285 --> 04:21:55,155 OF IL17 REPORTER MICE AS STEADY 6910 04:21:55,155 --> 04:21:57,591 STATE TO THAT OF PATHOGENIC CD4 6911 04:21:57,591 --> 04:22:01,294 T CELLS ISOLATED FRAWLS IL17 GFP 6912 04:22:01,294 --> 04:22:03,930 REPORTER MICE AT THE PEAK OF 6913 04:22:03,930 --> 04:22:06,233 NEUROINFLAMMATION USING EAE 6914 04:22:06,233 --> 04:22:06,533 MODEL. 6915 04:22:06,533 --> 04:22:10,504 UNLIKE EGL1 THERE WAS EXPRESSED 6916 04:22:10,504 --> 04:22:15,942 IN BOTH PROTACTIV AND -- CELLS 6917 04:22:15,942 --> 04:22:17,911 EGR2 SPECIFICALLY UP REGULATED 6918 04:22:17,911 --> 04:22:20,413 IN PATHOGENIC CD4 CELLS FROM THE 6919 04:22:20,413 --> 04:22:24,985 CNS OF EA TEST MICE, UNLIKE 6920 04:22:24,985 --> 04:22:25,886 HUMAN, MOUSE CELLS DO NOT 6921 04:22:25,886 --> 04:22:30,357 EXPRESS EGR3 AND 4 UNDER THESE 6922 04:22:30,357 --> 04:22:30,891 CONDITIONS. 6923 04:22:30,891 --> 04:22:33,627 SO WHAT ARE THE EGR FAMILY 6924 04:22:33,627 --> 04:22:34,494 TRANSCRIPTION FACTORS? 6925 04:22:34,494 --> 04:22:36,329 THEY STAND FOR EARLY GROWTH 6926 04:22:36,329 --> 04:22:37,764 RESPONSE FAMILY, THERE ARE FOUR 6927 04:22:37,764 --> 04:22:40,600 MEMBERS, ONE, TWO, THREE AND 6928 04:22:40,600 --> 04:22:41,701 FOUR. 6929 04:22:41,701 --> 04:22:44,104 IN MOUSE IMMUNE SYSTEM EGR ONE 6930 04:22:44,104 --> 04:22:45,705 TWORKS AND THREE ARE INDUCED BY 6931 04:22:45,705 --> 04:22:48,708 T CELL RECEPTOR SIGNALS. 6932 04:22:48,708 --> 04:22:50,710 THEY SHARE MORE THAN 80% OF HOME 6933 04:22:50,710 --> 04:22:52,212 OMG IN THE DNA BINDING DOMAIN 6934 04:22:52,212 --> 04:22:56,516 AND THEY BIND IDENTICAL DC RICH 6935 04:22:56,516 --> 04:22:57,584 SEQUENCES. 6936 04:22:57,584 --> 04:22:58,785 HOWEVER, IF YOU MAKE MICE THAT 6937 04:22:58,785 --> 04:23:00,620 LACK ONE OF THESE EGR FAMILY 6938 04:23:00,620 --> 04:23:02,155 MEMBERS, THEY HAVE VERY DISTINCT 6939 04:23:02,155 --> 04:23:03,123 FAMILY MEMBERS SUGGESTING 6940 04:23:03,123 --> 04:23:04,558 PERHAPS THEY ALSO HAVE UNIQUE 6941 04:23:04,558 --> 04:23:06,526 GENE REGULATION PROGRAMS. 6942 04:23:06,526 --> 04:23:09,062 IN IMMUNE SYSTEM, AS I MENTIONED 6943 04:23:09,062 --> 04:23:10,597 THEY'RE DOWNSTREAM OF T CELL 6944 04:23:10,597 --> 04:23:12,566 RECEPTOR SIGNALLING, SO NO NOT 6945 04:23:12,566 --> 04:23:13,533 SURPRISINGLY THEY PLAY AN 6946 04:23:13,533 --> 04:23:14,534 IMPORTANT ROLE AT DIFFERENT 6947 04:23:14,534 --> 04:23:15,769 STAGES OF T CELL DEVELOPMENT IN 6948 04:23:15,769 --> 04:23:18,171 THE THYMUS. 6949 04:23:18,171 --> 04:23:20,807 SO WE DECIDED TO LOOK AT THE 6950 04:23:20,807 --> 04:23:23,209 EXPRESSION OF EGR ONE AND EGR 6951 04:23:23,209 --> 04:23:27,814 TWO PROTEIN AT DIFFERENT STAGES 6952 04:23:27,814 --> 04:23:29,015 OF TH17 DIFFERENTIATION AND 6953 04:23:29,015 --> 04:23:31,318 FOUND THAT BOTH EGR ONE AND TWO 6954 04:23:31,318 --> 04:23:32,886 ARE CONSIDERED TO BE EARLY 6955 04:23:32,886 --> 04:23:35,355 INDUCED TRANSCRIPTION FACTORS, 6956 04:23:35,355 --> 04:23:37,357 THEY'RE UP WITHIN FOUR HOURS OF 6957 04:23:37,357 --> 04:23:39,392 ACTIVATION, AND THEN THEY'RE 6958 04:23:39,392 --> 04:23:41,394 CO-EXPRESSED IN ABOUT 30% OF 6959 04:23:41,394 --> 04:23:44,531 TH17 CELLS UP TO 48 HOURS POST 6960 04:23:44,531 --> 04:23:45,999 ACTIVATION AFTER WHICH THEIR 6961 04:23:45,999 --> 04:23:49,502 PROTEIN EXPRESSION IS COMPLETELY 6962 04:23:49,502 --> 04:23:51,871 DECLINED. 6963 04:23:51,871 --> 04:23:55,175 TO LOOK AT THEIR FUNCTION WE USE 6964 04:23:55,175 --> 04:23:56,810 RETRO VIAL ARE TRANS 6965 04:23:56,810 --> 04:23:59,412 TRANSDUCTION SYSTEM, ISOLATED 6966 04:23:59,412 --> 04:24:02,515 CD4 DISPELZ THROUGH RETRO VIAL 6967 04:24:02,515 --> 04:24:04,985 TRANSACTION OVEREXPRESS ERG1 OR 6968 04:24:04,985 --> 04:24:06,119 2 AND LOOK AT THE COMMITMENT 6969 04:24:06,119 --> 04:24:07,120 FIVE DAYS LATER. 6970 04:24:07,120 --> 04:24:10,390 AS YOU CAN SEE HERE, UNLIKE EGR 6971 04:24:10,390 --> 04:24:13,660 ONE, EGR IT TWO WAS ABLE TO 6972 04:24:13,660 --> 04:24:15,629 UPREGULATE EXPRESSION OF QUL RHO 6973 04:24:15,629 --> 04:24:18,031 GAMMA CODING GENE COMPARED TO 6974 04:24:18,031 --> 04:24:19,699 LSZ ANTI-VECTOR CONTROL AND ALSO 6975 04:24:19,699 --> 04:24:20,667 SIGNIFICANTED INCREASED THE 6976 04:24:20,667 --> 04:24:24,404 FREQUENCY OF IF IL17 PRODUCING 6977 04:24:24,404 --> 04:24:26,373 CD4 T CELLS SUGGESTING THAT EGR2 6978 04:24:26,373 --> 04:24:28,908 ACTS AS A POSITIVE REGULATOR OF 6979 04:24:28,908 --> 04:24:31,478 TH17 CELL DIFFERENTIATION 6980 04:24:31,478 --> 04:24:31,778 PROGRAM. 6981 04:24:31,778 --> 04:24:35,682 BUT IS EGR2 REQUIRED FOR TH17 6982 04:24:35,682 --> 04:24:36,349 LINEAGE COMMITMENT? 6983 04:24:36,349 --> 04:24:38,251 SO THIS QUESTION WAS RAISED 6984 04:24:38,251 --> 04:24:40,687 BECAUSE WE HAVE SEEN COMPENSATE 6985 04:24:40,687 --> 04:24:42,922 TO HER INCREASE IN EGR FAMILY 6986 04:24:42,922 --> 04:24:43,957 MEMBERS WHEN ONE OF THE MEMBERS 6987 04:24:43,957 --> 04:24:45,792 WAS DELETED. 6988 04:24:45,792 --> 04:24:49,763 SO EGR1 DEFICIENCY IN 6989 04:24:49,763 --> 04:24:51,698 SIGNIFICANT UPREGULATION OF EGR2 6990 04:24:51,698 --> 04:24:54,401 IN mRNA AND EGR2 DEFICIENCY 6991 04:24:54,401 --> 04:24:57,437 INCREASES EGR1 mRNA EXPRESSION 6992 04:24:57,437 --> 04:25:00,807 WHEREAS EGR1 AND 2 DOUBLE 6993 04:25:00,807 --> 04:25:03,410 INSUFFICIENCY RESULTS IN 6994 04:25:03,410 --> 04:25:08,782 ENHANCED EGR3 mRNA CR 6995 04:25:08,782 --> 04:25:09,983 TRANSCRIPTION, THAT WAS THE 6996 04:25:09,983 --> 04:25:10,183 CASE. 6997 04:25:10,183 --> 04:25:11,518 WE FOUND WE HAVE TO DELETE ALL 6998 04:25:11,518 --> 04:25:14,921 THREE EGR FAMILY MEMBERS TO 6999 04:25:14,921 --> 04:25:19,626 SIGNIFICANTLY INHIBIT CELL 7000 04:25:19,626 --> 04:25:19,959 COMMITMENT. 7001 04:25:19,959 --> 04:25:21,995 BASED ON THESE DATA WE CONCLUDED 7002 04:25:21,995 --> 04:25:24,898 THAT EGR2 IS EXPRESSED IN 7003 04:25:24,898 --> 04:25:25,965 PATHOGENIC TH17 CELLS IN MICE 7004 04:25:25,965 --> 04:25:27,734 AND HUMANS DURING INFLAMMATION 7005 04:25:27,734 --> 04:25:35,975 AND WHEN OVEREXPRESSED EGR -- 7006 04:25:35,975 --> 04:25:37,210 REQUIRED FOR TH17 LINEAGE 7007 04:25:37,210 --> 04:25:39,612 COMMITMENT DUE TO FUNCTIONAL 7008 04:25:39,612 --> 04:25:42,549 REDUNDANCY AMONGST EGR FAMILY 7009 04:25:42,549 --> 04:25:42,782 MEMBERS. 7010 04:25:42,782 --> 04:25:46,252 DOES EGR TWO CONTROL DEVELOPMENT 7011 04:25:46,252 --> 04:25:48,321 HOMEOSTATIC OR INFECTION INDUCED 7012 04:25:48,321 --> 04:25:51,658 TH17 CELLS? 7013 04:25:51,658 --> 04:25:53,126 TO ADDRESS THIS QUESTION, WE 7014 04:25:53,126 --> 04:25:56,529 DECIDED TO LOOK AT EGR2 PROTEIN 7015 04:25:56,529 --> 04:25:58,364 EXPRESSION IN INTESTINAL 7016 04:25:58,364 --> 04:26:00,066 HOMEOSTATIC CD4 T CELLS ISOLATED 7017 04:26:00,066 --> 04:26:04,471 FROM THE LAMINA PROPRIA OF SMALL 7018 04:26:04,471 --> 04:26:06,339 EXPWEFN COLON OF COLONIZED MOUSE 7019 04:26:06,339 --> 04:26:07,774 IN STEADY STATE CONDITION AND 7020 04:26:07,774 --> 04:26:08,975 SURPRISINGLY WE FOUND THAT ONLY 7021 04:26:08,975 --> 04:26:12,278 A SMALL FRACTION OF CD4 T CELLS 7022 04:26:12,278 --> 04:26:15,648 EXPRESSED EGR2 AT STEADY STATE. 7023 04:26:15,648 --> 04:26:16,750 CYTOKINE PRODUCTION IN 7024 04:26:16,750 --> 04:26:19,185 PARTICULAR IL17 WAS COMPARABLE 7025 04:26:19,185 --> 04:26:21,888 BETWEEN EGR2 KNOCK-OUT AND 7026 04:26:21,888 --> 04:26:23,656 WILD-TYPE CD4 T CELLS IN BOTH 7027 04:26:23,656 --> 04:26:26,593 TISSUES, SUGGESTING THAT EGR2 IS 7028 04:26:26,593 --> 04:26:30,663 NOT REQUIRED FOR HOMEOSTATIC 7029 04:26:30,663 --> 04:26:33,199 TH17 CELL DEVELOPMENT VFT BUT TO 7030 04:26:33,199 --> 04:26:36,169 CLEAR INTESTINAL PATH JENS SUCH 7031 04:26:36,169 --> 04:26:38,037 AS REDEMPTION MOUSE AND HUMANS 7032 04:26:38,037 --> 04:26:40,874 DEPEND ON INFECTION INDUCED TH17 7033 04:26:40,874 --> 04:26:41,407 CELLS. 7034 04:26:41,407 --> 04:26:43,376 AND THIS CAN BE ASSOCIATED WITH 7035 04:26:43,376 --> 04:26:46,246 TISSUE INFLAMMATION AND 7036 04:26:46,246 --> 04:26:48,114 PATHOLOGY. 7037 04:26:48,114 --> 04:26:49,949 BUT SURPRISINGLY WHEN WE CHANGED 7038 04:26:49,949 --> 04:26:54,888 THE MICE IT FROMS SIT TROA BAC 7039 04:26:54,888 --> 04:26:57,423 TEST R, THERE WAS NO CHANGE IN 7040 04:26:57,423 --> 04:27:01,461 INTESTINAL CD4 CELLS, MOREOVER, 7041 04:27:01,461 --> 04:27:02,762 KNOCK OUT CELLS EXPRESSED 7042 04:27:02,762 --> 04:27:05,698 SIMILAR AMOUNTS OF GAMMA IN TH17 7043 04:27:05,698 --> 04:27:07,367 COMPARED TO WILD-TYPE CONTROLS. 7044 04:27:07,367 --> 04:27:09,536 ALSO WE FOUND THAT MICE THAT 7045 04:27:09,536 --> 04:27:11,404 LACK EGR2 EXPRESSION 7046 04:27:11,404 --> 04:27:12,705 SPECIFICALLY IN T CELLS 7047 04:27:12,705 --> 04:27:14,574 CONTROLLED BACTERIAL BURDEN AND 7048 04:27:14,574 --> 04:27:16,109 LIKE WILD-TYPE ANIMALS THEY 7049 04:27:16,109 --> 04:27:18,878 DIDN'T SHOW ANY SIGNS OF BODY 7050 04:27:18,878 --> 04:27:19,946 WEIGHT LOSS. 7051 04:27:19,946 --> 04:27:21,981 SO COLLECTIVELY THIS 7052 04:27:21,981 --> 04:27:23,917 DEMONSTRATES THAT EGR2 IS NOT 7053 04:27:23,917 --> 04:27:26,119 REQUIRED FOR EITHER HOMEOSTATIC 7054 04:27:26,119 --> 04:27:30,790 OR INFECTION INDUCED TH17 CELLS. 7055 04:27:30,790 --> 04:27:32,192 BUT THEN WE BECAME INTERESTED 7056 04:27:32,192 --> 04:27:34,828 ABOUT THE ROLE OF EGR2 IN 7057 04:27:34,828 --> 04:27:36,629 CONTROLLING PATHOGENIC CD4 T 7058 04:27:36,629 --> 04:27:39,465 CELLS DURING ITS UP REGULATION 7059 04:27:39,465 --> 04:27:40,733 DURING NEUROINFLAMMATION IN 7060 04:27:40,733 --> 04:27:43,169 HUMANS WITH MS AND ALSO IN MICE 7061 04:27:43,169 --> 04:27:45,238 WITH EAE. 7062 04:27:45,238 --> 04:27:46,806 USING ACTIVE IMMUNIZATION MODEL 7063 04:27:46,806 --> 04:27:48,341 WITH MOCK PROTEIN WE FOUND WITH 7064 04:27:48,341 --> 04:27:51,110 30% OF CNS INFILTRATING CD4 7065 04:27:51,110 --> 04:27:56,115 CELLS EXPRESSED EGR2, AND ALSO 7066 04:27:56,115 --> 04:27:58,418 MICE IN WHICH WE DELETED EGR2 7067 04:27:58,418 --> 04:28:02,222 EITHER ONLY IN T CELLS OR IN KIM 7068 04:28:02,222 --> 04:28:03,756 L17 PRODUCER CELLS WERE 7069 04:28:03,756 --> 04:28:04,824 SIGNIFICANTLY PROTECTED FROM 7070 04:28:04,824 --> 04:28:07,694 DISEASE DPIERTD WILD-TYPE 7071 04:28:07,694 --> 04:28:07,961 CONTROLS. 7072 04:28:07,961 --> 04:28:10,630 -- DPIERD THE WILD-TYPE 7073 04:28:10,630 --> 04:28:11,731 CONTROLS. 7074 04:28:11,731 --> 04:28:13,600 ERRING 1 DEFICIENCY HAD NO 7075 04:28:13,600 --> 04:28:16,102 IMPACT RS CONCLUDE THAT GO EGR2 7076 04:28:16,102 --> 04:28:20,907 BUNLT EGR1 PROMOTES PATHOGENIC 7077 04:28:20,907 --> 04:28:22,942 TH17 CELL DEVELOPMENT OF SOS 7078 04:28:22,942 --> 04:28:25,912 WHAT IS THE FUNCTION OF EGR2 IN 7079 04:28:25,912 --> 04:28:27,480 THIS PATHOGENIC TH17 CELLS? 7080 04:28:27,480 --> 04:28:29,449 WE REALLY WANTED TO GO DEEPER 7081 04:28:29,449 --> 04:28:32,051 INTO THE MOLECULAR MECHANISMS BY 7082 04:28:32,051 --> 04:28:34,220 WHICH EGR2 PROMOTE PATHOGENICITY 7083 04:28:34,220 --> 04:28:36,055 SO TO ANSWER THIS QUESTION WE 7084 04:28:36,055 --> 04:28:38,691 COMPARED THE GENE EXPRESSION 7085 04:28:38,691 --> 04:28:41,561 PROGRAM OF SPLENIC CD4 T CELLS 7086 04:28:41,561 --> 04:28:43,396 OR CNS INFILTRATING CELLS 7087 04:28:43,396 --> 04:28:44,964 BETWEEN THE WILD-TYPE AND 7088 04:28:44,964 --> 04:28:48,268 KNOCKOUTS AND YOU CAN SEE HERE 7089 04:28:48,268 --> 04:28:51,237 THAT EGR2 DEFICIENCY HAD MANY 7090 04:28:51,237 --> 04:28:52,772 MORE IMPACT ON GENE EXPRESSION 7091 04:28:52,772 --> 04:28:54,073 PROGRAM IN THE SPLEEN HOWEVER IN 7092 04:28:54,073 --> 04:28:56,209 THE CNS THERE WAS A HUGE 7093 04:28:56,209 --> 04:28:58,711 DYSREGULATION IN THE GENE 7094 04:28:58,711 --> 04:29:01,214 EXPRESSION IN THE CNS WHERE THEY 7095 04:29:01,214 --> 04:29:05,718 NORMALLY REENCOUNTER THEIR 7096 04:29:05,718 --> 04:29:05,952 ANTIGEN. 7097 04:29:05,952 --> 04:29:08,554 SO WE WANTED TO IDENTIFY 7098 04:29:08,554 --> 04:29:10,523 MOLECULAR PATHWAY THAT IS ARE 7099 04:29:10,523 --> 04:29:13,059 REGULATED BY EGR2 AND WE USED 7100 04:29:13,059 --> 04:29:14,994 EGR2 RETROVIRUS OVEREXPRESSION 7101 04:29:14,994 --> 04:29:18,531 AND WE FOUND THAT EGR2 7102 04:29:18,531 --> 04:29:21,501 OVEREXPRESSION NORMALLY IN 7103 04:29:21,501 --> 04:29:23,136 NONPATHOGENIC TH17 CELLS INDUCED 7104 04:29:23,136 --> 04:29:25,204 THE EXPRESSION OF TEN OUT OF 7105 04:29:25,204 --> 04:29:26,706 SIXTEEN PATHOGENICITY ASSOCIATED 7106 04:29:26,706 --> 04:29:29,042 GENES COMPARED TO THE EMPTY 7107 04:29:29,042 --> 04:29:36,950 VECTOR CONTROL. 7108 04:29:36,950 --> 04:29:37,583 PATHOGENICITY ASSOCIATED GENES 7109 04:29:37,583 --> 04:29:40,486 IN THE CNS DURING EAE. 7110 04:29:40,486 --> 04:29:44,057 AND THIS ALSO INCLUDED THE 7111 04:29:44,057 --> 04:29:48,561 TRANSCRIPTION FACTOR T-BET. 7112 04:29:48,561 --> 04:29:50,930 NOW, BECAUSE T-BET IS SO 7113 04:29:50,930 --> 04:29:53,366 IMPORTANT FOR PATHOGENICITY OF 7114 04:29:53,366 --> 04:29:54,500 TH17 CELLS WE WANTED TO VALIDATE 7115 04:29:54,500 --> 04:29:57,303 THESE RESULTS IN VITRO SO IN 7116 04:29:57,303 --> 04:29:59,906 THIS PARTICULAR EXPERIMENT, WE 7117 04:29:59,906 --> 04:30:01,808 USED CD4 CELLS THAT WERE 7118 04:30:01,808 --> 04:30:03,843 CULTURED UNDER NONPATHOGENIC 7119 04:30:03,843 --> 04:30:05,278 POLARIZING CONDITIONS IN THE 7120 04:30:05,278 --> 04:30:07,847 PRESENCE OF BETA ONE AND EITHER 7121 04:30:07,847 --> 04:30:09,449 TRANSDUCED THEM WITH EMPTY 7122 04:30:09,449 --> 04:30:13,519 VECTOR CONTROL OR EGR1 OR EGR2 7123 04:30:13,519 --> 04:30:14,153 OVEREXPRESSING RETROVIRUS AND 7124 04:30:14,153 --> 04:30:18,157 YOU CAN SEE THAT AS EXPECTED, 7125 04:30:18,157 --> 04:30:19,492 NONPATHOGENIC WILL LSES TH17 7126 04:30:19,492 --> 04:30:21,127 CELLS DO NOT EXPRESS T-BET ASK 7127 04:30:21,127 --> 04:30:24,297 IN CONTRAST TO ARE ERG-1, ERG 7128 04:30:24,297 --> 04:30:26,366 2-6S ABLE TO INDUCE T-BET 7129 04:30:26,366 --> 04:30:27,934 EXPRESSION IN THESE CELLS, 7130 04:30:27,934 --> 04:30:30,636 SUGGEST THAT GO PERHAPS SOME 7131 04:30:30,636 --> 04:30:32,739 PATHOGENICITY IS CONTROLLED BY 7132 04:30:32,739 --> 04:30:35,308 2ERG MEDIATED UPREGULATED 7133 04:30:35,308 --> 04:30:36,409 LAITION OF THE TRANSCRIPTION 7134 04:30:36,409 --> 04:30:37,076 FACTOR T-BET. 7135 04:30:37,076 --> 04:30:39,579 TO TEST THIS WE DID A PASSIVE 7136 04:30:39,579 --> 04:30:43,182 TRANSFER ADOPTED TRANSFER EAE 7137 04:30:43,182 --> 04:30:44,517 EXPERIMENTS IN WHICH THLSZ TH17 7138 04:30:44,517 --> 04:30:47,653 CELLS WERE CULTURED UNDER 7139 04:30:47,653 --> 04:30:48,621 NONPOLARIZING TH17 CONDITIONS IN 7140 04:30:48,621 --> 04:30:55,328 THE PRESENCE OF MS THETA, 7141 04:30:55,328 --> 04:30:55,661 RETROVIRUS. 7142 04:30:55,661 --> 04:30:58,865 ALSO INCLUDED PATHOGENIC IL23 7143 04:30:58,865 --> 04:31:01,167 STIMULATED CELLS AS A POSITIVE 7144 04:31:01,167 --> 04:31:02,035 CONTROL. 7145 04:31:02,035 --> 04:31:04,704 AND UNLIKE NONPATHOGENIC TH17 7146 04:31:04,704 --> 04:31:06,539 CELLS TRANSDUCED WITH THE 7147 04:31:06,539 --> 04:31:08,541 ANTI-VIRUS, THOSE THAT EXPRESS 7148 04:31:08,541 --> 04:31:11,144 EGR2 WERE NOW ABLE TO INDUCE 7149 04:31:11,144 --> 04:31:11,677 NEUROINFLAMMATION. 7150 04:31:11,677 --> 04:31:14,313 SO THESE DATA SUGGEST THAT EGR2 7151 04:31:14,313 --> 04:31:16,182 IS REALLY CONTROLLING THE 7152 04:31:16,182 --> 04:31:18,251 PATHOGENIC PROGRAM OF THESE 7153 04:31:18,251 --> 04:31:21,354 MYELIN REACTIVE PATHOGENIC TH17 7154 04:31:21,354 --> 04:31:21,554 CELLS. 7155 04:31:21,554 --> 04:31:23,523 IN ADDITION TO T-BET THERE WAS 7156 04:31:23,523 --> 04:31:24,624 ONE VERY INTERESTING PATHWAY 7157 04:31:24,624 --> 04:31:27,460 THAT STOOD OUT, AND IT INVOLVED 7158 04:31:27,460 --> 04:31:29,862 GENES INVOLVING T CELL MIGRATION 7159 04:31:29,862 --> 04:31:31,164 INCLUDING CHEMOKINE AND IS 7160 04:31:31,164 --> 04:31:32,632 CHEMOKINE RECEPTORS, SAME SUBSET 7161 04:31:32,632 --> 04:31:34,901 OF CHEMOKINES AND CHEMOKINE 7162 04:31:34,901 --> 04:31:36,769 RECEPTORS WAS ALSO UP REGULATED 7163 04:31:36,769 --> 04:31:39,572 IN PATHOGENIC IL23 STIMULATED 7164 04:31:39,572 --> 04:31:41,941 TH17 CELLS SO WE POSTULATED THAT 7165 04:31:41,941 --> 04:31:43,242 PERHAPS 2ERG CONTROLS IMMUNE 7166 04:31:43,242 --> 04:31:47,947 CELL MIGRATION TO THIS -- CNS, 7167 04:31:47,947 --> 04:31:50,383 INDEED THE CASE, COMPARTMENT WAS 7168 04:31:50,383 --> 04:31:51,918 NORMAL BETWEEN WILD-TYPE AND 7169 04:31:51,918 --> 04:31:53,886 EGR2 KNOCK-OUT MICE BUT IN THE 7170 04:31:53,886 --> 04:31:55,421 CNS THERE WAS A HUGE DECREASE IN 7171 04:31:55,421 --> 04:31:58,257 THE ACCUMULATION OF PATHOGENIC 7172 04:31:58,257 --> 04:31:59,926 CD4 T CELLS. 7173 04:31:59,926 --> 04:32:03,429 IN ADDITION, WE FOUND THAT THE 7174 04:32:03,429 --> 04:32:09,836 EXPRESSION OF MYELOCYTE KINES 7175 04:32:09,836 --> 04:32:13,406 L1, 3 AND 4 EXPRESSION, SINCE 7176 04:32:13,406 --> 04:32:15,408 THESE CHEMOKINES RECRUIT, WE 7177 04:32:15,408 --> 04:32:18,211 FOUND SIGNIFICANT REDUCTION IN 7178 04:32:18,211 --> 04:32:19,312 INFLAMMATORY MONOCYTES 7179 04:32:19,312 --> 04:32:23,149 SUMMARIZED HERE AND ALSO CAN 7180 04:32:23,149 --> 04:32:24,784 EXPRESSING DENDRITIC CELLS. 7181 04:32:24,784 --> 04:32:25,885 SO COLLECTIVELY WHAT I HAVE 7182 04:32:25,885 --> 04:32:28,754 SHOWN YOU HERE IS THAT WE HAVE 7183 04:32:28,754 --> 04:32:29,922 IDENTIFIED A NOVEL FUNCTION FOR 7184 04:32:29,922 --> 04:32:33,893 THE TRANSCRIPTION FACTOR EGR2 IN 7185 04:32:33,893 --> 04:32:35,962 DIFFERENTIATING PATHOGENIC TH17 7186 04:32:35,962 --> 04:32:37,730 CELLS THAT PROMOTE INFLAMMATORY 7187 04:32:37,730 --> 04:32:39,565 DISEASES SUCH AS MS, AND THOSE 7188 04:32:39,565 --> 04:32:42,568 THAT ARE PROTECTIVE AND 7189 04:32:42,568 --> 04:32:44,036 BENEFICIAL TH17 CELLS IN THE 7190 04:32:44,036 --> 04:32:44,303 GUT. 7191 04:32:44,303 --> 04:32:46,105 I HAVEN'T SHOWN YOU BUT WE ALSO 7192 04:32:46,105 --> 04:32:50,710 LOOKED AT CANDIDA ALBICANS AND 7193 04:32:50,710 --> 04:32:52,578 BACTERIUM AND RESPONSES, THESE 7194 04:32:52,578 --> 04:32:53,779 MUCOSAL TH17 CELLS ARE 7195 04:32:53,779 --> 04:32:55,414 COMPLETELY NORMAL WHEN WE KNOCK 7196 04:32:55,414 --> 04:32:56,415 OUT EGR2. 7197 04:32:56,415 --> 04:33:00,219 BUT IN THE CONTEXT OF THE CNS 7198 04:33:00,219 --> 04:33:05,691 AND AUTOIMMUNE INFLAM ARE 7199 04:33:05,691 --> 04:33:07,460 INFLAMMATION, THIS -- 7200 04:33:07,460 --> 04:33:08,361 PATHOGENICITY ASSOCIATED GENES 7201 04:33:08,361 --> 04:33:10,596 AND ALSO CHEMOKINES AND CELL 7202 04:33:10,596 --> 04:33:12,131 RECRUITMENT TO THE CNS. 7203 04:33:12,131 --> 04:33:13,499 I SLEANLT A CHANCE TO SHOW YOU 7204 04:33:13,499 --> 04:33:15,701 THE DATA, BUT THIS CONTEXTUAL 7205 04:33:15,701 --> 04:33:17,570 REQUIREMENT FOR EGR2 WAS 7206 04:33:17,570 --> 04:33:19,205 REGULATED BY THE AFFINITY AND 7207 04:33:19,205 --> 04:33:21,374 THE STRENGTH OF T CELL RECEPTOR 7208 04:33:21,374 --> 04:33:22,909 SIGNAL SIGNALLING AS WELL AS 7209 04:33:22,909 --> 04:33:24,577 TISSUE SPECIFIC CUES. 7210 04:33:24,577 --> 04:33:26,646 AND FINALLY, I WOULD LIKE TO 7211 04:33:26,646 --> 04:33:29,382 THANK THE MEBLES OF MY TEAMLY. 7212 04:33:29,382 --> 04:33:31,350 -- THE MEKS OF MY TEAM. 7213 04:33:31,350 --> 04:33:33,352 -- AND FINAL HL, I WOULD LIKE TO 7214 04:33:33,352 --> 04:33:43,462 THANK THE MEMBERS OF MY TEAM. 7215 04:33:43,462 --> 04:33:46,732 SO I WOULD LIKE TO THANK MY NIH 7216 04:33:46,732 --> 04:33:48,501 COLLABORATORS AND MY COLLEAGUES 7217 04:33:48,501 --> 04:33:49,669 AT EXPERIMENTAL IMMUNOLOGY 7218 04:33:49,669 --> 04:33:49,902 BRANCH. 7219 04:33:49,902 --> 04:33:51,003 THANK YOU VERY MUCH FOR YOUR 7220 04:33:51,003 --> 04:33:51,270 ATTENTION. 7221 04:33:51,270 --> 04:33:52,972 I'M HAPPY TO ANSWER ANY 7222 04:33:52,972 --> 04:33:55,041 QUESTIONS. 7223 04:33:55,041 --> 04:34:01,714 [APPLAUSE] 7224 04:34:01,714 --> 04:34:05,785 SO THIS IS DR. NICKI -- 7225 04:34:05,785 --> 04:34:07,186 >> YES, A WONDERFUL TALK. 7226 04:34:07,186 --> 04:34:08,020 CAN YOU HEAR ME? 7227 04:34:08,020 --> 04:34:08,421 >> YES. 7228 04:34:08,421 --> 04:34:10,089 >> SO THE WAY YOU PRESENTED 7229 04:34:10,089 --> 04:34:14,760 THIS, IT SEEMS LIKE EGR2 IS VERY 7230 04:34:14,760 --> 04:34:16,596 SPECIFIC TO YOUR INFLAMMATION 7231 04:34:16,596 --> 04:34:17,797 FROM WHAT BEUF SEEN SO FAR. 7232 04:34:17,797 --> 04:34:19,232 CAN YOU ELABORATE A LITTLE MORE 7233 04:34:19,232 --> 04:34:21,767 ON THE TISSUE SPECIFIC CUES 7234 04:34:21,767 --> 04:34:22,735 REGULATED IN THE BRAIN OR 7235 04:34:22,735 --> 04:34:24,303 CONTEXT OF INFLAMMATION AND 7236 04:34:24,303 --> 04:34:26,038 BECAUSE SINCE YOU'RE SHOWING IT 7237 04:34:26,038 --> 04:34:27,840 MIGRATION, SOME OF IT MIGHT BE 7238 04:34:27,840 --> 04:34:29,742 UPREGULATED BEFORE THEY GET 7239 04:34:29,742 --> 04:34:31,377 THERE OR HOW ARE YOU THINKING OF 7240 04:34:31,377 --> 04:34:31,811 THAT? 7241 04:34:31,811 --> 04:34:35,114 >> SO THE SIGNAL THAT DEFINITELY 7242 04:34:35,114 --> 04:34:37,183 CROAMS THE LEVEL OF EGR2 7243 04:34:37,183 --> 04:34:39,485 EXPRESSION EVEN IN MONOCLONAL 7244 04:34:39,485 --> 04:34:41,087 POPULATION CELLS IS THE STRENGTH 7245 04:34:41,087 --> 04:34:43,222 AND THE AFFINITY OF T CELL 7246 04:34:43,222 --> 04:34:43,956 RECEPTOR SIGNALLING. 7247 04:34:43,956 --> 04:34:46,459 ONE POSSIBILITY IS THIS AUTO 7248 04:34:46,459 --> 04:34:47,860 REACTIVE T CELLS THAT EXPRESS 7249 04:34:47,860 --> 04:34:50,630 HIGH AFFINITY TCR'S MAY HAVE 7250 04:34:50,630 --> 04:34:53,799 HIGH LEVELS OF EGR2 THAN 7251 04:34:53,799 --> 04:34:55,534 POLYFUNCTIONAL MICROBIAL 7252 04:34:55,534 --> 04:34:58,304 SPECIFIC T CELLS. 7253 04:34:58,304 --> 04:34:59,839 SO ONE OF THE THINGS THAT CAN BE 7254 04:34:59,839 --> 04:35:01,674 MAKING THE DIFFERENCES ARE THOSE 7255 04:35:01,674 --> 04:35:01,941 TISSUE -- 7256 04:35:01,941 --> 04:35:04,210 >> IMMUNITY AREA, ARE THERE 7257 04:35:04,210 --> 04:35:05,645 SETTINGS BETWEEN IMMUNITY YOU 7258 04:35:05,645 --> 04:35:07,280 COULD HAVE THAT AS WELL 7259 04:35:07,280 --> 04:35:07,613 POTENTIALLY? 7260 04:35:07,613 --> 04:35:10,883 >> SO ONE THING WE LOOKED AT WAS 7261 04:35:10,883 --> 04:35:11,984 COLITIS FOR EXAMPLE, T CELL 7262 04:35:11,984 --> 04:35:13,753 DRIVEN COA LIGHTS MODEL. 7263 04:35:13,753 --> 04:35:17,356 THERE JUST LIKE WITH CITRO 7264 04:35:17,356 --> 04:35:20,526 BACTER RETENTION WE FOUND NO 7265 04:35:20,526 --> 04:35:22,695 CHANGE IN DR.-- EXPRESSION, DOWN 7266 04:35:22,695 --> 04:35:22,962 REGULATED. 7267 04:35:22,962 --> 04:35:23,863 THERE'S SOMETHING ABOUT THE GUT 7268 04:35:23,863 --> 04:35:27,400 THAT IS SUPPRESSING EGR2. 7269 04:35:27,400 --> 04:35:29,268 NOW, TGF BETA SOMEBODY 7270 04:35:29,268 --> 04:35:31,570 MENTIONED, YES, WE LOOKED AT IT, 7271 04:35:31,570 --> 04:35:34,507 DEFINITELY HAS A DOSE DEPENDENT 7272 04:35:34,507 --> 04:35:36,942 INHIBITION OF ALL EGR FAMILY 7273 04:35:36,942 --> 04:35:38,678 MEMBERS BUT THE CNS ALSO HAS 7274 04:35:38,678 --> 04:35:41,080 HIGH LEVELS OF TGF BETA AS WELL 7275 04:35:41,080 --> 04:35:43,816 SO WE'RE NOT QUITE SURE WHETHER 7276 04:35:43,816 --> 04:35:46,352 TGF BETA IS THE ONLY SIGHT DHIEN 7277 04:35:46,352 --> 04:35:46,719 SUPPRESSES IT. 7278 04:35:46,719 --> 04:35:49,155 >> SECOND SIDE OF TISSUE 7279 04:35:49,155 --> 04:35:51,457 COMPARTMENT WITH THE HUMAN ATLAS 7280 04:35:51,457 --> 04:35:52,591 DATA OUT I DON'T KNOW IF YOU HAD 7281 04:35:52,591 --> 04:35:54,460 AN OPPORTUNITY TO LOOK THE ADATA 7282 04:35:54,460 --> 04:35:55,461 PUBLISHED IN HUMANS AND MICE AT 7283 04:35:55,461 --> 04:35:57,096 DIFFERENT SETTINGS TO SEE 7284 04:35:57,096 --> 04:36:00,066 WHETHER T CELLS IN DIFFERENT 7285 04:36:00,066 --> 04:36:01,033 PATHOLOGIES HAVE 2ERG OR NOT. 7286 04:36:01,033 --> 04:36:03,269 >> WE DID LOOK AT, YEAH, DIDN'T 7287 04:36:03,269 --> 04:36:05,204 LOOK AT OTHER PSORIASIS FOR 7288 04:36:05,204 --> 04:36:06,305 EXAMPLE, WE HAVEN'T LOOKED IN 7289 04:36:06,305 --> 04:36:07,406 THAT PARTICULAR CASE. 7290 04:36:07,406 --> 04:36:08,808 BUT WE WILL DEFINITELY LOOK AT 7291 04:36:08,808 --> 04:36:09,375 THAT IN THE FUTURE. 7292 04:36:09,375 --> 04:36:10,209 >> WONDERFUL TALK. 7293 04:36:10,209 --> 04:36:15,181 >> THANK YOU. 7294 04:36:15,181 --> 04:36:16,082 THANKS. 7295 04:36:16,082 --> 04:36:16,949 >> HELLO. 7296 04:36:16,949 --> 04:36:17,983 I CAN JUST SPEAK LOUD. 7297 04:36:17,983 --> 04:36:20,653 CAN YOU HEAR ME? 7298 04:36:20,653 --> 04:36:21,053 >> YES. 7299 04:36:21,053 --> 04:36:23,789 >> SO IT WAS A VERY NICE TALK. 7300 04:36:23,789 --> 04:36:25,558 I HAD A QUESTION ON THE 7301 04:36:25,558 --> 04:36:26,992 CHEMOKINE PART. 7302 04:36:26,992 --> 04:36:29,729 SO KIND OF THE OTHER SIDE OF THE 7303 04:36:29,729 --> 04:36:30,596 COIN, ON THE RESIDENTIAL PART, 7304 04:36:30,596 --> 04:36:32,231 DO YOU THINK THERE'S A LOSS OF 7305 04:36:32,231 --> 04:36:34,133 RESIDENCY AND MORE MIGRATION AND 7306 04:36:34,133 --> 04:36:38,270 CIRCULATION OF THOSE CELLS? 7307 04:36:38,270 --> 04:36:38,437 ARE. 7308 04:36:38,437 --> 04:36:40,406 >> IN THE ABSENCE OF -- 7309 04:36:40,406 --> 04:36:40,973 >> YEAH. 7310 04:36:40,973 --> 04:36:42,308 >> THAT'S ACTUALLY A GREAT 7311 04:36:42,308 --> 04:36:42,575 QUESTION. 7312 04:36:42,575 --> 04:36:45,478 WE DO NOT LOOK, FOR EXAMPLE, AT 7313 04:36:45,478 --> 04:36:46,846 S1P1 RECEPTOR EXPRESSION BECAUSE 7314 04:36:46,846 --> 04:36:48,547 YOU NEED TO DOWN REGULATED IT TO 7315 04:36:48,547 --> 04:36:50,750 STAY IN THE CNS COMPARTMENT. 7316 04:36:50,750 --> 04:36:51,050 >> EXACTLY. 7317 04:36:51,050 --> 04:36:52,385 >> NORMALLY EVEN IN THE 7318 04:36:52,385 --> 04:36:54,453 WILD-TYPE SETTING WHEN WE LOOK 7319 04:36:54,453 --> 04:36:56,122 AT THE TH17 CELLS THAT GOT TO 7320 04:36:56,122 --> 04:37:01,460 THE CNS WE DIDN'T SEE MUCH AS 7321 04:37:01,460 --> 04:37:02,561 RECEPTOR SO WE CAN DEFINITELY 7322 04:37:02,561 --> 04:37:04,530 LOOK AT LEAST AT THE mRNA 7323 04:37:04,530 --> 04:37:06,065 LEVEL WHETHER WE SEE ANY DIMPS 7324 04:37:06,065 --> 04:37:06,499 THERE. 7325 04:37:06,499 --> 04:37:08,467 IT DIDN'T POP OUT AS ONE OF THE 7326 04:37:08,467 --> 04:37:09,668 PATHWAYS THAT IS DIFFERENTIALLY 7327 04:37:09,668 --> 04:37:11,003 REGULATED GROO BECAUSE EVEN THE 7328 04:37:11,003 --> 04:37:14,240 OTHER MEMBERS, ALL THE RETENTION 7329 04:37:14,240 --> 04:37:15,908 MODULES REGULATED BY THE T CELLS 7330 04:37:15,908 --> 04:37:17,676 BECAUSE MAYBE THEY'RE JUST, I 7331 04:37:17,676 --> 04:37:20,112 DON'T KNOW, MIGRATING MORE 7332 04:37:20,112 --> 04:37:22,581 BETWEEN THE TISSUE AND HAVING A 7333 04:37:22,581 --> 04:37:22,815 MORE -- 7334 04:37:22,815 --> 04:37:25,785 >> OTHER TISSUE RETENTION ARE WE 7335 04:37:25,785 --> 04:37:28,888 LOOKED AT WERE CD69 AND 103 AND 7336 04:37:28,888 --> 04:37:30,689 THERE WERE NO DIFFERENCES IN 7337 04:37:30,689 --> 04:37:31,123 PROTEIN EXPRESSION. 7338 04:37:31,123 --> 04:37:31,524 >> OKAY. 7339 04:37:31,524 --> 04:37:33,626 THANK YOU. 7340 04:37:33,626 --> 04:37:34,960 >> THANK YOU VERY MUCH FOR YOUR 7341 04:37:34,960 --> 04:37:35,161 TALK. 7342 04:37:35,161 --> 04:37:41,734 MY NAME IS -- I WORK IN NIAD ON 7343 04:37:41,734 --> 04:37:43,602 VIRUSLESS RECEPTION, SO SINCE 7344 04:37:43,602 --> 04:37:45,337 THOSE EGR TRANSCRIPTION FACTORS 7345 04:37:45,337 --> 04:37:47,106 ARE INDUCERS OF TH17 CELLS HAVE 7346 04:37:47,106 --> 04:37:49,275 YOU LOOKED TO SEE WHETHER THEY 7347 04:37:49,275 --> 04:37:52,378 AFFECT EITHER DIRECTLY OR 7348 04:37:52,378 --> 04:37:54,213 INDIRECTLY ROR OR STAT OF THE 7349 04:37:54,213 --> 04:37:55,414 TRANSCRIPTION FACTORS? 7350 04:37:55,414 --> 04:37:58,951 >> THEY DEFINITELY WELL EGR2 NOT 7351 04:37:58,951 --> 04:38:01,887 OTHER EGR FAMILY MEMBERS, 7352 04:38:01,887 --> 04:38:03,422 REGULATE GAMMA T EXPRESSION, WE 7353 04:38:03,422 --> 04:38:05,524 DIDN'T SEE ANY DIFFERENCES IN 7354 04:38:05,524 --> 04:38:07,259 HIF1 OR STAT 3 EXPRESSION WHEN 7355 04:38:07,259 --> 04:38:11,297 WE KNOCK OUT EGR2 BUT DEFINITELY 7356 04:38:11,297 --> 04:38:15,367 EGR2 UP REGULAR LAILTS FOR GAMMA 7357 04:38:15,367 --> 04:38:17,903 T SO PROMOTES TH17 7358 04:38:17,903 --> 04:38:19,738 PROLIFERATION, LOSES EFFECT WITH 7359 04:38:19,738 --> 04:38:21,907 IF WE INCOME OUT GAMMA T. 7360 04:38:21,907 --> 04:38:23,876 IT'S DEFINITELY GAMMA T 7361 04:38:23,876 --> 04:38:24,143 DEPENDENT. 7362 04:38:24,143 --> 04:38:24,477 >> THANK YOU. 7363 04:38:24,477 --> 04:38:26,045 >> THANK YOU FOR THE NICE TALK. 7364 04:38:26,045 --> 04:38:29,815 I HAVE A BASIC QUESTION. 7365 04:38:29,815 --> 04:38:32,451 -- TH17 ARE LIKE RESIDENT CELLS, 7366 04:38:32,451 --> 04:38:35,621 BUT EAE MODEL ARE THEY COMING 7367 04:38:35,621 --> 04:38:39,124 FROM LIKE NOT THE RNTD OR THEIR 7368 04:38:39,124 --> 04:38:41,760 RECRUIT, GET RECRUITED TO THE 7369 04:38:41,760 --> 04:38:41,961 SITE? 7370 04:38:41,961 --> 04:38:43,596 -- ARE NOT OR THEIR RECRUIT? 7371 04:38:43,596 --> 04:38:44,563 IS THAT POSSIBLE THAT THAT 7372 04:38:44,563 --> 04:38:47,566 SITUATION MAKES A DIFFERENCE? 7373 04:38:47,566 --> 04:38:49,001 >> THANK YOU FOR THE QUESTION. 7374 04:38:49,001 --> 04:38:52,938 DEFINITELY EAE IS DRIVEN BY THE 7375 04:38:52,938 --> 04:38:54,473 RECRUITMENT OF THESE PATHOGENIC 7376 04:38:54,473 --> 04:38:55,841 TH17 CELLS SO RETENTION WOULD BE 7377 04:38:55,841 --> 04:38:58,010 ONE OF POSSIBLE MECHANISM THAT 7378 04:38:58,010 --> 04:38:59,745 IF IT'S DYSFUNCTIONAL COULD LEAD 7379 04:38:59,745 --> 04:39:01,180 IN THEIR CIRCULATION BACK OUT OF 7380 04:39:01,180 --> 04:39:03,048 THE CNS COMPARTMENT BUT THAT'S 7381 04:39:03,048 --> 04:39:04,049 WHATNOT WE SEE. 7382 04:39:04,049 --> 04:39:05,451 USUALLY WE DON'T SEE ENOUGH 7383 04:39:05,451 --> 04:39:07,319 RECRUITMENT TO THE CNS SO THE 7384 04:39:07,319 --> 04:39:08,687 MICE REMAIN PROTECTED. 7385 04:39:08,687 --> 04:39:09,522 THANK YOU. 7386 04:39:09,522 --> 04:39:10,756 THANKS A LOT FOR YOUR QUESTION. 7387 04:39:10,756 --> 04:39:13,125 THANK YOU. 7388 04:39:13,125 --> 04:39:23,302 [APPLAUSE] 7389 04:39:25,771 --> 04:39:29,441 >> OUR NEXT SPEAKER AND FINAL 7390 04:39:29,441 --> 04:39:31,844 SPEAKER BEFORE THE BREAK IS 7391 04:39:31,844 --> 04:39:36,282 VIJAY KUCHROO, FROM HARVARD 7392 04:39:36,282 --> 04:39:39,218 MEDICAL SCHOOL, NEURO IMMUNE 7393 04:39:39,218 --> 04:39:40,753 INTERACTIONS IN ENTERIC NERVOUS 7394 04:39:40,753 --> 04:39:41,320 SYSTEM. 7395 04:39:41,320 --> 04:39:45,090 THANK YOU FOR COMING, VIJAY. 7396 04:39:45,090 --> 04:39:48,527 >> AFTER SHRUTI AND VAN 7397 04:39:48,527 --> 04:39:49,962 GENTLEMAN'S TALK, I THOUGHT I 7398 04:39:49,962 --> 04:39:54,099 SHOULD ALSO GIVE A TH17 TALK. 7399 04:39:54,099 --> 04:39:57,269 -- AND VANJA'S TALK, I THOUGHT I 7400 04:39:57,269 --> 04:40:04,810 SHOULD ALSO GIVE A TH17 TALK. 7401 04:40:04,810 --> 04:40:08,080 S ARE. 7402 04:40:08,080 --> 04:40:17,089 [PAUSE] THIS IS NOT THE MAIN TOM 7403 04:40:17,089 --> 04:40:19,491 TOOP -- THIS IS NOT THE MAIN -- 7404 04:40:19,491 --> 04:40:21,360 THIS IS NOT THE MAIN TOPIC OF MY 7405 04:40:21,360 --> 04:40:23,329 LAB BUT I THOUGHT I WILL TALK 7406 04:40:23,329 --> 04:40:24,863 ABOUT SOMETHING I NEVER TALK B 7407 04:40:24,863 --> 04:40:27,633 I'LL TALK BTD NEURO IMMUNE 7408 04:40:27,633 --> 04:40:28,801 INTERACTIONS AND AND HOW 7409 04:40:28,801 --> 04:40:31,036 IMPORTANT THEY MAY BE IN HOW THE 7410 04:40:31,036 --> 04:40:32,237 IMMUNE SYSTEM SEES THE NERVOUS 7411 04:40:32,237 --> 04:40:33,739 SYSTEM AND NERVOUS SYSTEM SEES 7412 04:40:33,739 --> 04:40:34,974 IMMUNE SYSTEM AND HOW THEY 7413 04:40:34,974 --> 04:40:35,841 CROSSTALK WITH EACH OTHER. 7414 04:40:35,841 --> 04:40:37,343 CAN YOU GUYS HEAR ME AT THE 7415 04:40:37,343 --> 04:40:37,576 BACK? 7416 04:40:37,576 --> 04:40:37,876 >> YES. 7417 04:40:37,876 --> 04:40:38,978 >> OKAY. 7418 04:40:38,978 --> 04:40:41,313 >> SOY OFTEN SEEM TO RUSH AT THE 7419 04:40:41,313 --> 04:40:43,382 END, AND IN FACT, THE PEOPLE 7420 04:40:43,382 --> 04:40:44,683 FROM IT WAS A COLLABORATION 7421 04:40:44,683 --> 04:40:53,125 BETWEEN THREE LABS, MY LAB AND 7422 04:40:53,125 --> 04:40:58,931 THESE OTHER LABS, SAM AND (?) 7423 04:40:58,931 --> 04:41:01,100 DID MOST OF THE WORK AND HELPED 7424 04:41:01,100 --> 04:41:04,403 US WITH SOME OF THE DATASETS. 7425 04:41:04,403 --> 04:41:10,376 AND WE GOT LOTS OF -- FROM THE 7426 04:41:10,376 --> 04:41:14,079 CHILDREN'S HOSPITAL AND GAVE US 7427 04:41:14,079 --> 04:41:19,551 R13 RECEPTOR, AND WITH ISAAC 7428 04:41:19,551 --> 04:41:21,387 CHLU WE DID SOME OF THE WORK AND 7429 04:41:21,387 --> 04:41:23,889 DAVID ARTIS I DON'T KNOW IF IS 7430 04:41:23,889 --> 04:41:26,025 IN THE AUDIENCE HERE OR NOT BUT 7431 04:41:26,025 --> 04:41:28,060 ALSO PROVIDED US WITH SOME HELP. 7432 04:41:28,060 --> 04:41:29,995 SO MY TALK I'LL DO IN TWO PARTS. 7433 04:41:29,995 --> 04:41:31,630 THE VERY FIRST PART OF THE TALK 7434 04:41:31,630 --> 04:41:36,435 IS SOME HISTORIC DATA ON ON HOW 7435 04:41:36,435 --> 04:41:37,970 WE IDENTIFIED NEUROPEPTIDE 7436 04:41:37,970 --> 04:41:39,705 RECEPTORS ON INNATE LYMPHOID 7437 04:41:39,705 --> 04:41:42,608 CELLS AND HOW THEY MAY FB 7438 04:41:42,608 --> 04:41:45,411 CROSSTALK INNATE LYMPH CELLS ARE 7439 04:41:45,411 --> 04:41:46,745 GETTING CUES FROM THE NERVOUS 7440 04:41:46,745 --> 04:41:47,046 SYSTEM. 7441 04:41:47,046 --> 04:41:49,948 THE SECOND PART OF THE TALK, I 7442 04:41:49,948 --> 04:41:52,217 EVER NOT SPOKEN ABOUT IT BEFORE, 7443 04:41:52,217 --> 04:41:54,720 SO YOU ARE MY FIRST GUINEA PIGS. 7444 04:41:54,720 --> 04:41:56,288 SO LET ME GET THE FIRST PART OF 7445 04:41:56,288 --> 04:41:56,789 THE TALK. 7446 04:41:56,789 --> 04:41:59,425 IN FACT, THE STORY STARTS ABOUT 7447 04:41:59,425 --> 04:42:03,462 30 YEARS AGO WHEN MY SON WAS 7448 04:42:03,462 --> 04:42:06,098 LITTLE, AND EFS FOUND TO BE 7449 04:42:06,098 --> 04:42:08,067 ALLERGIC TO ALMOST EVERYTHING. 7450 04:42:08,067 --> 04:42:13,305 HAD TO MILK, TO EKSZ HR TO NUTS, 7451 04:42:13,305 --> 04:42:14,740 AND -- TO EGGS, TO NUTS, THIS 7452 04:42:14,740 --> 04:42:17,710 WAS A TIME WHEN THERE WERE NO 7453 04:42:17,710 --> 04:42:19,178 PEANUT-FREE ZONES IN KINDER GARR 7454 04:42:19,178 --> 04:42:20,379 TENSE OR WHATEVER AND IT OFTEN 7455 04:42:20,379 --> 04:42:22,414 SO HAPPENED THAT ONCE HE HAD A 7456 04:42:22,414 --> 04:42:24,283 BOUT OF FOOD ALLERGIES, HE WILL 7457 04:42:24,283 --> 04:42:26,919 ACTUALLY WE CAN GIVE HIM 7458 04:42:26,919 --> 04:42:30,522 EPI-PEN, WE CAN GIVE HIM 7459 04:42:30,522 --> 04:42:32,257 ANTIHISTAMINES, EVERYTHING WAS 7460 04:42:32,257 --> 04:42:33,659 NORMAL OUT OF DANGER BUT IF HE 7461 04:42:33,659 --> 04:42:35,194 STARTED THROWING UP, IT WOULD 7462 04:42:35,194 --> 04:42:37,596 NOT STOP. 7463 04:42:37,596 --> 04:42:39,665 IN FACT, WE HAD TO TAKE HIM TO 7464 04:42:39,665 --> 04:42:40,699 THE BOSTON CHILDREN'S HOSPITAL 7465 04:42:40,699 --> 04:42:41,767 TO GET SEDATED. 7466 04:42:41,767 --> 04:42:44,403 THIS APPARENTLY HAPPENS IN ABOUT 7467 04:42:44,403 --> 04:42:47,673 20% OF THE YOUNG KIDS THAT HAVE 7468 04:42:47,673 --> 04:42:50,275 VERY SERIOUS FOOD ALLERGIES. 7469 04:42:50,275 --> 04:42:54,246 SO AT THAT TIME, IN FACT, I CAME 7470 04:42:54,246 --> 04:42:56,582 WITH THIS IDEA THAT MAYBE THEIR 7471 04:42:56,582 --> 04:42:58,217 IMMUNE SYSTEM AND NERVOUS SYSTEM 7472 04:42:58,217 --> 04:43:02,087 ARE INTIMATELY COMMUNICATING 7473 04:43:02,087 --> 04:43:05,124 WITH EACH OTHER, AND IT'S 7474 04:43:05,124 --> 04:43:06,992 POSSIBLE THAT THE IMMUNE SYSTEM 7475 04:43:06,992 --> 04:43:09,194 IS MAKING CYTOKINES AND 7476 04:43:09,194 --> 04:43:11,363 CYTOKINES ACTIVATE THE NERVOUS 7477 04:43:11,363 --> 04:43:14,433 SYSTEM WITH NEUROPEPTIDES AND 7478 04:43:14,433 --> 04:43:15,968 THAT ACT ON THEIR IMMUNE CELLS 7479 04:43:15,968 --> 04:43:17,970 AND THIS IS LIKE A FEED-FORWARD 7480 04:43:17,970 --> 04:43:19,471 LOOP AMPLIFYING AND YOU GET 7481 04:43:19,471 --> 04:43:21,206 ACTIVATION AND IRRITATION OF 7482 04:43:21,206 --> 04:43:23,942 NERVES HAD. 7483 04:43:23,942 --> 04:43:26,812 AND IRRITATION MUCH NERVES. 7484 04:43:26,812 --> 04:43:30,749 HAD AND THE IDEA THAT YOU SEE. 7485 04:43:30,749 --> 04:43:33,452 SO ONE OF THE EXPERIMENTS WE DID 7486 04:43:33,452 --> 04:43:35,687 MANY QULIERTZ IS THAT WE 7487 04:43:35,687 --> 04:43:39,191 LOOKED -- MANY YEARS LATER IS WE 7488 04:43:39,191 --> 04:43:41,794 DID SIMILAR ILC2'S IN THE GUT 7489 04:43:41,794 --> 04:43:43,295 AND LUNG, THIS IS FROM THE LUNG, 7490 04:43:43,295 --> 04:43:51,003 YOU CAN SEE, IL 25RBGS IL 7491 04:43:51,003 --> 04:43:52,971 23-RBGS ABOUT 80,000 SINGLE 7492 04:43:52,971 --> 04:43:55,040 CELLS YOU CAN SEE THEY CLUSTER 7493 04:43:55,040 --> 04:43:56,041 DIFFERENT AND WILL WE IDENTIFIED 7494 04:43:56,041 --> 04:43:59,211 THIS RECEPTOR NEUROPEPTIDE 7495 04:43:59,211 --> 04:44:02,381 RECEPTOR FOR NMUR1 WHICH IS 7496 04:44:02,381 --> 04:44:04,049 GENERALLY PRESENT AND HAD BEEN 7497 04:44:04,049 --> 04:44:06,218 IDENTIFIED BEFORE ON SMOOTH 7498 04:44:06,218 --> 04:44:09,721 MUSCLES AND WHEN AK TI VASE 7499 04:44:09,721 --> 04:44:13,325 NMUR1 SMOOTH MUSCLE IT CAUSES 7500 04:44:13,325 --> 04:44:14,459 BRONCHIAL SPASM AND IS DIARRHEA 7501 04:44:14,459 --> 04:44:17,262 AND VOMITING, INTERESTING THAT 7502 04:44:17,262 --> 04:44:18,797 THIS NEUROPEPTIDE RECEPTOR 7503 04:44:18,797 --> 04:44:20,766 EXPRESSED ON INNATE LYMPHOID 7504 04:44:20,766 --> 04:44:23,836 CELL CLEARLY INVOLVED IN TYPE 2 7505 04:44:23,836 --> 04:44:26,004 IMMUNITY AND ALLERGIES AND 7506 04:44:26,004 --> 04:44:27,773 ASTHMAS, AND THE QUESTION WAS 7507 04:44:27,773 --> 04:44:30,843 HAD, WHAT'S THE ROLE OF THIS 7508 04:44:30,843 --> 04:44:32,377 NEUROPEPTIDE COMING FROM NEURONS 7509 04:44:32,377 --> 04:44:34,780 ACTING ON INNATE LYMPHOID CELLS? 7510 04:44:34,780 --> 04:44:36,548 AND WHAT HAPPENS TO INNATE 7511 04:44:36,548 --> 04:44:37,082 LYMPHOID CELLS? 7512 04:44:37,082 --> 04:44:39,384 AND THE DATA IS PUBLISHED. 7513 04:44:39,384 --> 04:44:44,857 AND IT WAS VERY CLEAR THAT -- 7514 04:44:44,857 --> 04:44:46,458 WILL EXPAND INNATE LYMPHOID 7515 04:44:46,458 --> 04:44:48,961 CELLS UNDER MUCOSAL SURFACE BUT 7516 04:44:48,961 --> 04:44:50,762 WILL NOT EN DO YOU SAY DISEASE 7517 04:44:50,762 --> 04:44:52,831 OR ALLERGIES UNLESS THEY GET A 7518 04:44:52,831 --> 04:44:55,667 SECOND SIGNAL DPROMG NEURONS AND 7519 04:44:55,667 --> 04:44:58,403 FALL OFF NEUROPEPTIDE CALLED NMU 7520 04:44:58,403 --> 04:45:00,372 AND THE TWO TOGETHER WILL GIVE 7521 04:45:00,372 --> 04:45:02,107 YOU CHANGE THIS INNATD LYMPHOID 7522 04:45:02,107 --> 04:45:04,776 CELL FROM BEING HOMEOSTATIC INTO 7523 04:45:04,776 --> 04:45:08,480 INFLAMMATORY ILC2'S AND IF YOU 7524 04:45:08,480 --> 04:45:12,017 INTERRUPTED THIS NMU, NMUR1 7525 04:45:12,017 --> 04:45:13,018 PATHWAY YOU CAN ESSENTIALLY 7526 04:45:13,018 --> 04:45:16,054 INHIBIT THE DEVELOPMENT OF THIS 7527 04:45:16,054 --> 04:45:21,226 ALLERGY BOUT AT LEAST IN THE 7528 04:45:21,226 --> 04:45:28,000 MOUSE. 7529 04:45:28,000 --> 04:45:29,601 SO EVERY TIME I PUBLISH A GOOD 7530 04:45:29,601 --> 04:45:30,836 PAPER THERE ARE MANY OTHERS 7531 04:45:30,836 --> 04:45:32,738 RIGHT WITH T THREE PAPERS COMING 7532 04:45:32,738 --> 04:45:35,807 UP TOGETHER SHOWING NMU AND 7533 04:45:35,807 --> 04:45:37,643 NMUR1 IN TRACTIONS ARE CRITICAL 7534 04:45:37,643 --> 04:45:39,711 FOR ACTIVATE INNATE LYMPHOID 7535 04:45:39,711 --> 04:45:41,580 CELLS AND IN FACT THE YES IS, 7536 04:45:41,580 --> 04:45:44,650 WHO PRODUCES NMU AND WHEN? 7537 04:45:44,650 --> 04:45:47,886 AND WE BELIEVE THAT IT IS 7538 04:45:47,886 --> 04:45:53,659 ACTUALLY IN THE GUT, THE PSM2 7539 04:45:53,659 --> 04:45:55,928 NEURONS, SENSORY NEURONS THAT 7540 04:45:55,928 --> 04:46:04,603 ARE COMING FROM MI ONTR -- 7541 04:46:04,603 --> 04:46:05,370 MYENTERIC PLEXUS, MUSCLES THAT 7542 04:46:05,370 --> 04:46:08,774 GO AROUND THE GUT, AND THEY FORM 7543 04:46:08,774 --> 04:46:11,076 THESE GANGLIA AND WILL YOU CAN 7544 04:46:11,076 --> 04:46:13,145 SEE THE SUB MUCOSAL PLEXUS AND 7545 04:46:13,145 --> 04:46:17,716 THESE ARE YOUR VILII AND WE 7546 04:46:17,716 --> 04:46:20,786 THINK ESSENTIALLY WE HAVE WE CAN 7547 04:46:20,786 --> 04:46:23,855 IDENTIFY THESE NEURONS THAT 7548 04:46:23,855 --> 04:46:24,723 PRODUCE NMU. 7549 04:46:24,723 --> 04:46:28,894 IN FACT, THEY START FROM THIS 7550 04:46:28,894 --> 04:46:30,295 MYENTERIC PLEXUS AND GO ALL THE 7551 04:46:30,295 --> 04:46:32,197 WAY THROUGH INTO THEVILLE LIE 7552 04:46:32,197 --> 04:46:34,800 AND THAT'S -- THE VILLI, AND 7553 04:46:34,800 --> 04:46:36,335 THEY INTERACT WITH IMMUNE CELLS 7554 04:46:36,335 --> 04:46:37,536 AND REGULATE THEIR FUNCTION BOTH 7555 04:46:37,536 --> 04:46:39,271 IN TERMS OF IMMUNE CELLS 7556 04:46:39,271 --> 04:46:40,605 REGULATING THE NERVOUS SYSTEM 7557 04:46:40,605 --> 04:46:43,342 FUNCTIONS AND NEURONS AND 7558 04:46:43,342 --> 04:46:44,843 NEURONS ACTUALLY REGULATING 7559 04:46:44,843 --> 04:46:45,510 IMMUNE FUNCTION. 7560 04:46:45,510 --> 04:46:46,945 THEY ACTUALLY HAVE MANY OTHER 7561 04:46:46,945 --> 04:46:47,980 FUNCTIONS IN WHICH YOU WILL SEE 7562 04:46:47,980 --> 04:46:50,916 IN A MINUTE. 7563 04:46:50,916 --> 04:46:54,052 SO IT'S VERY CLEAR THAT THESE 7564 04:46:54,052 --> 04:46:57,222 HAD LONG DISTANCE RUNNERS COMING 7565 04:46:57,222 --> 04:47:01,493 FROM THE MYENTERIC PLEXUS ARE 7566 04:47:01,493 --> 04:47:04,429 COMING FROM THE VILLI AND 7567 04:47:04,429 --> 04:47:06,631 INTERACTING WITH IMMUNE CELLS. 7568 04:47:06,631 --> 04:47:07,933 S THE FIRST PART OF THE TALK 7569 04:47:07,933 --> 04:47:11,269 WHAT I TOLD YOU, TO SUM UP, BY 7570 04:47:11,269 --> 04:47:14,306 DOING SINGLE CELL RNA SEQ OF 7571 04:47:14,306 --> 04:47:15,741 ILC'S AND WE IDENTIFIED 7572 04:47:15,741 --> 04:47:21,113 RECEPTORS FOR NEUROPEPTIDES AND 7573 04:47:21,113 --> 04:47:26,685 NMUR1. 7574 04:47:26,685 --> 04:47:28,887 AND ILC2'S. 7575 04:47:28,887 --> 04:47:31,757 WHAT I ALSO TODAY YOU IS THAT 7576 04:47:31,757 --> 04:47:34,593 THE NMU NEUROPEPTIDE WAS 7577 04:47:34,593 --> 04:47:36,795 SYNERGIZED WITH THESE ALARMENS 7578 04:47:36,795 --> 04:47:38,930 TO MAKE THESE ILC'S 7579 04:47:38,930 --> 04:47:40,465 INFLAMMATORY, AND TOGETHER THEY 7580 04:47:40,465 --> 04:47:42,234 WILL IN FACT PROMOTE P THE 7581 04:47:42,234 --> 04:47:43,101 ALLERGIC REACTION AND THIS IS 7582 04:47:43,101 --> 04:47:47,472 WHAT WE SAW AND I THINK DAVID 7583 04:47:47,472 --> 04:47:51,676 ART I S' LAB AND HENRIK'S LAB IN 7584 04:47:51,676 --> 04:47:53,745 FACT IDENTIFIED THE SAME 7585 04:47:53,745 --> 04:47:55,280 INTERACTION IN THE GUT. 7586 04:47:55,280 --> 04:47:57,249 SO I TOLD YOU THE FIRST PART OF 7587 04:47:57,249 --> 04:47:59,618 THE TALK IS THAIN DEED 7588 04:47:59,618 --> 04:48:03,121 NEUROPEPTIDE RECEPTORS EXPRESSED 7589 04:48:03,121 --> 04:48:05,090 ON THEIR INNATE LYMPHOID CELLS 7590 04:48:05,090 --> 04:48:07,392 THAT INDUCE ALLERGIC REACTION, 7591 04:48:07,392 --> 04:48:08,460 AMPLIFY ALLERGIC REACTION. 7592 04:48:08,460 --> 04:48:12,864 BUT THE QUESTION IS THAT WE 7593 04:48:12,864 --> 04:48:17,669 SHOWED YOU THAT THE NEURONS OR 7594 04:48:17,669 --> 04:48:19,771 NEUROENDOCRINE CELL WILL MAKE 7595 04:48:19,771 --> 04:48:22,741 NMU AND ACT ON INNATE LYMPHOID 7596 04:48:22,741 --> 04:48:24,776 CELLS BUT WHAT ARE THE CYTOKINE 7597 04:48:24,776 --> 04:48:27,612 RECEPTORS ON THE NEURONS THAT 7598 04:48:27,612 --> 04:48:30,615 WILL RESPOND TO THESE CYTOKINE 7599 04:48:30,615 --> 04:48:32,084 SIGNALS COMING FROM IMMUNE 7600 04:48:32,084 --> 04:48:32,317 SYSTEM? 7601 04:48:32,317 --> 04:48:36,088 AND IF SO, WHAT DO THEY DO? 7602 04:48:36,088 --> 04:48:37,989 AND THE QUESTION IS WHAT 7603 04:48:37,989 --> 04:48:39,958 CYTOKINE RECEPTORS ARE EXPRESSED 7604 04:48:39,958 --> 04:48:43,795 ON THESE NEURONS? 7605 04:48:43,795 --> 04:48:47,966 AND FORTUNATELY FOR US,S THERE 7606 04:48:47,966 --> 04:48:50,602 WAS A BIG DATASET BOTH FROM 7607 04:48:50,602 --> 04:48:52,671 HUMAN AND MOUSE, WHERE THEY HAD 7608 04:48:52,671 --> 04:48:55,307 LOOKED AT ALL THE RECEPTORS 7609 04:48:55,307 --> 04:48:56,875 EXPRESSED ON NEURONS. 7610 04:48:56,875 --> 04:48:58,810 IN FACT, WE TOOK THREE DIFFERENT 7611 04:48:58,810 --> 04:49:00,879 DATASETS THAT WERE PUBLISHED AND 7612 04:49:00,879 --> 04:49:03,482 TRIED TO MAKE AN ATLAS OR A 7613 04:49:03,482 --> 04:49:05,150 CATALOG OF ALL THE RECEPTORS 7614 04:49:05,150 --> 04:49:09,087 THAT ARE EXPRESSED ON THESE 7615 04:49:09,087 --> 04:49:11,923 ENTERIC NERVOUS SYSTEM, ENTERIC 7616 04:49:11,923 --> 04:49:12,924 NEURONS. 7617 04:49:12,924 --> 04:49:14,993 AND HERE IT IS ACTUALLY I HAVE 7618 04:49:14,993 --> 04:49:16,761 CUT A LOT OF DATA DOWN, AND 7619 04:49:16,761 --> 04:49:18,063 ANYBODY WHO HAS QUESTIONS CAN 7620 04:49:18,063 --> 04:49:23,101 COME AND TALK TO ME A LITTLE 7621 04:49:23,101 --> 04:49:23,401 LATER. 7622 04:49:23,401 --> 04:49:25,837 THESE ARE YOUR NMU POSITIVE 7623 04:49:25,837 --> 04:49:29,307 NEURONS, ALSO CALLED PSM2, 7624 04:49:29,307 --> 04:49:32,777 SENSORY NEURO NEURONS, IPAN'S T 7625 04:49:32,777 --> 04:49:34,846 THESE ARE ALL THE CYTOKINE 7626 04:49:34,846 --> 04:49:36,148 RECEPTORS WE COULD IDENTIFY IN 7627 04:49:36,148 --> 04:49:38,483 THESE THREE DATASETS AND WE 7628 04:49:38,483 --> 04:49:39,985 CATALOGED THEM TOGETHER. 7629 04:49:39,985 --> 04:49:42,654 AND THEN YOU CAN FIND IF YOU 7630 04:49:42,654 --> 04:49:46,525 LOOK AT TYPE 2 CYTOKINES IL4 AND 7631 04:49:46,525 --> 04:49:48,193 IL13 CYTOKINE RECEPTORS YOU CAN 7632 04:49:48,193 --> 04:49:51,263 CLEARLY SEE THEM ON THESE NMU 7633 04:49:51,263 --> 04:49:52,030 NEURONS, YOU CAN SEE THAT 7634 04:49:52,030 --> 04:49:55,100 THERE'S IL4 RECEPTOR AND IL13 7635 04:49:55,100 --> 04:49:56,234 RECEPTOR EXPRESSED. 7636 04:49:56,234 --> 04:49:59,638 SO IT'S NOT JUST IMAGINATION. 7637 04:49:59,638 --> 04:50:02,807 THAT INDEED, THE NEUROPEPTIDE 7638 04:50:02,807 --> 04:50:05,076 RECEPTORS ARE PRESENT ON INNATE 7639 04:50:05,076 --> 04:50:07,512 LYMPHOID CELLS, AND INNATE 7640 04:50:07,512 --> 04:50:08,914 LYMPHOID CELLS WOULD MAKE 7641 04:50:08,914 --> 04:50:11,750 CYTOKINES LIKE IL4, 5 AND IL13 7642 04:50:11,750 --> 04:50:13,618 AND NOW WE ARE FINDING FROM THIS 7643 04:50:13,618 --> 04:50:16,454 DATASET THAT IL13 AND IL4 7644 04:50:16,454 --> 04:50:18,190 RECEPTORS ARE ACTUALLY PRESENT 7645 04:50:18,190 --> 04:50:22,460 ON ENTERIC NEURONS, IPAN'S OR 7646 04:50:22,460 --> 04:50:28,800 PSM2 CLUSTER OF SENSORY NEURONS. 7647 04:50:28,800 --> 04:50:30,769 YOU GUYS ARE ALL IN IMMUNOLOGY, 7648 04:50:30,769 --> 04:50:33,638 YOU KNOW THAT IL13 AND IL4 FORM 7649 04:50:33,638 --> 04:50:35,240 DIFFERENT TYPES OF RECEPTORS, IN 7650 04:50:35,240 --> 04:50:37,809 FACT, IL4 RECEPTOR IMMUNE SYSTEM 7651 04:50:37,809 --> 04:50:40,312 IS MADE BY IL4 RECEPTOR TOGETHER 7652 04:50:40,312 --> 04:50:46,551 WITH COMMON GAMMA CHAIN AND THE 7653 04:50:46,551 --> 04:50:47,986 PATHWAY, AND THE JAK PATHWAY. 7654 04:50:47,986 --> 04:50:49,454 IN OTHER CELL TYPES INCLUDING 7655 04:50:49,454 --> 04:50:50,455 SMOOTH MUSCLES IT'S A DIFFERENT 7656 04:50:50,455 --> 04:50:55,360 TYPE OF RECEPTOR AND TRACK TIL4 7657 04:50:55,360 --> 04:51:00,098 AND IL13 FORM THESE HETERODIMER 7658 04:51:00,098 --> 04:51:01,700 RICK RECEPTORS ACCIDENT I TOOK A 7659 04:51:01,700 --> 04:51:04,236 BUNCH OF SLIDES OUT, THAT'S THE 7660 04:51:04,236 --> 04:51:06,838 RECEPTOR WE ARE FIENG ON THE 7661 04:51:06,838 --> 04:51:07,706 NEURONS THAT IT IS A COMBINATION 7662 04:51:07,706 --> 04:51:11,042 OF IL4 RECEPTOR WITH IL13 7663 04:51:11,042 --> 04:51:13,411 RECEPTOR ALPHA CHAIN. 7664 04:51:13,411 --> 04:51:16,348 SO WE ACTUALLY STRUGGLE A LOT TO 7665 04:51:16,348 --> 04:51:19,851 REALLY FIND WHAT ROLE DOES 7666 04:51:19,851 --> 04:51:28,293 CYTOKINES HAVE, AND A MASTER 7667 04:51:28,293 --> 04:51:29,227 STUDENT IN FACT STRUCK HE WOULD 7668 04:51:29,227 --> 04:51:32,230 TO FORM THESE IN VITRO 7669 04:51:32,230 --> 04:51:33,698 NEUROSPEAR CULL CULTURES AND 7670 04:51:33,698 --> 04:51:34,633 FINALLY WE FOUND THE CONDITIONS 7671 04:51:34,633 --> 04:51:37,168 WHERE WE CAN ACTUALLY GENERATE 7672 04:51:37,168 --> 04:51:38,703 THESE SENSORY NEURONS IN THE 7673 04:51:38,703 --> 04:51:39,471 CULTURE DISH. 7674 04:51:39,471 --> 04:51:42,107 WHAT THEY DID IS THAT THEY PUT 7675 04:51:42,107 --> 04:51:46,278 ON IL13 OR IL4 ON THEM, AND WE 7676 04:51:46,278 --> 04:51:47,912 CAN IMMEDIATELY SEE THAT WHEN 7677 04:51:47,912 --> 04:51:52,417 YOU PUT THE IL13 RECOMBINANT 7678 04:51:52,417 --> 04:51:53,151 CYTOKINE IN VITRO CULTURE YOU 7679 04:51:53,151 --> 04:51:54,719 CAN SEE AN INCREASE IN THE 7680 04:51:54,719 --> 04:51:57,422 EXPRESSION OF THESE NEURO 7681 04:51:57,422 --> 04:52:03,995 NEUROPEPTIDES NMUU THIS CGRP AND 7682 04:52:03,995 --> 04:52:04,829 GRP. 7683 04:52:04,829 --> 04:52:06,731 AND YOU CAN LOOK AT THIS HERE 7684 04:52:06,731 --> 04:52:10,101 AGAIN IS THAT YOU CAN SEE THAT 7685 04:52:10,101 --> 04:52:11,636 IF YOU HUS A COMBINATION OF 7686 04:52:11,636 --> 04:52:13,438 CYTOKINES AS YOU KNOW THAT ENTER 7687 04:52:13,438 --> 04:52:15,273 ON GAMMA GENERALLY INHIBITS TH 7688 04:52:15,273 --> 04:52:17,008 TO CYTOKINE PRODUCTION AND THE 7689 04:52:17,008 --> 04:52:18,643 FUNCTIONS, AND SO WE STUDY USING 7690 04:52:18,643 --> 04:52:21,813 THE COMBINATIONS OF IL4 AND IL13 7691 04:52:21,813 --> 04:52:25,850 AND GAMMA ON IL4 AND IL13 7692 04:52:25,850 --> 04:52:29,387 TOGETHER AND LOOKING AT THE 7693 04:52:29,387 --> 04:52:30,889 NEUROPEPTIDE FLUKS IN VITRO 7694 04:52:30,889 --> 04:52:32,324 CULTURE AND IT'S CLEAR THAT THE 7695 04:52:32,324 --> 04:52:37,495 IL4 AND IL13 WILL BOTH INDUCE 7696 04:52:37,495 --> 04:52:43,902 NMU, CGR ARE P DATA AND 7697 04:52:43,902 --> 04:52:45,337 NEUROPEPTIDES, INHAVE I TROAT 7698 04:52:45,337 --> 04:52:46,671 NEUROSPEAR CULTURES. 7699 04:52:46,671 --> 04:52:47,405 IT'S NOT FOR EVERYTHING. 7700 04:52:47,405 --> 04:52:51,376 IN FACT, THERE ARE NEUROPEPTIDES 7701 04:52:51,376 --> 04:52:53,678 LIKE VIP THAT THE CYTOKINE 7702 04:52:53,678 --> 04:52:55,013 RECEPTOR TRIGGERING WOONTD 7703 04:52:55,013 --> 04:52:55,547 INDUCE THEM. 7704 04:52:55,547 --> 04:52:57,982 SO IT LOOKS LIKE IT'S SOMEHOW 7705 04:52:57,982 --> 04:52:58,917 SPECIFIC HOW WE ARE DOING WITH 7706 04:52:58,917 --> 04:53:01,019 THE TIME? 7707 04:53:01,019 --> 04:53:01,653 YEAH? 7708 04:53:01,653 --> 04:53:01,886 OKAY. 7709 04:53:01,886 --> 04:53:04,155 SO AT LEAST IN VITRO, WHEN YOU 7710 04:53:04,155 --> 04:53:07,559 USE A NEUROSPHERE AND ACTIVATE 7711 04:53:07,559 --> 04:53:08,893 THE RECOMBINANT CYTOKINES YOU 7712 04:53:08,893 --> 04:53:10,729 CAN SEE THAT YOU ARE FULFILLING 7713 04:53:10,729 --> 04:53:13,264 THIS POSTULATE THAT THE CYTOKINE 7714 04:53:13,264 --> 04:53:15,200 RECEPTOR SIGNALLING IS ACTUALLY 7715 04:53:15,200 --> 04:53:16,668 INDUCING THESE NEUROPEPTIDES, 7716 04:53:16,668 --> 04:53:18,403 IT'S NOT TRUE FOR ALL OF THEM. 7717 04:53:18,403 --> 04:53:22,807 IN FACT, NMU AND CGRP ARE THE 7718 04:53:22,807 --> 04:53:24,275 TWO NEUROPEPTIDES THAT WE AND 7719 04:53:24,275 --> 04:53:27,946 OTHERS HAVE PUBLISHED TO HELP 7720 04:53:27,946 --> 04:53:29,180 IMMUNOMODULATORY OR 7721 04:53:29,180 --> 04:53:30,915 IMMUNOREGULATORY PROPERTIES. 7722 04:53:30,915 --> 04:53:33,918 BUT ET IN VITRO IS WELL AND GOOD 7723 04:53:33,918 --> 04:53:35,687 BUT WE WANTED TO SEE WHAT 7724 04:53:35,687 --> 04:53:37,956 HAPPENS IN IN VIVO, BECAUSE 7725 04:53:37,956 --> 04:53:39,824 CYTOKINES HAVE SUCH SHORT 7726 04:53:39,824 --> 04:53:40,158 HALF-LIFE. 7727 04:53:40,158 --> 04:53:42,594 WE USED IL4 COME PLEKS AND GAVE 7728 04:53:42,594 --> 04:53:44,462 THEM IN VIVO INTO THE MICE AND 7729 04:53:44,462 --> 04:53:46,197 THEN LOOKED AT THE ENTERIC 7730 04:53:46,197 --> 04:53:48,733 NERVOUS SYSTEM TO SEE WHETHER 7731 04:53:48,733 --> 04:53:51,336 THIS RECOME BY NANTD IL4 GRIFN 7732 04:53:51,336 --> 04:53:54,406 IN VIVO FOR ANTIBODY CYTOKINE 7733 04:53:54,406 --> 04:53:55,774 COMPLEX WILL INDUPS THE SIGHT 7734 04:53:55,774 --> 04:53:57,742 EXIENS IS THIS IS DIFFERENT 7735 04:53:57,742 --> 04:54:01,780 PARTS OF THE GUT, DUODENUM ILEUM 7736 04:54:01,780 --> 04:54:03,648 AND COLON AND YOU CAN SEE THAT 7737 04:54:03,648 --> 04:54:05,617 IL4 COMPLEX WILL INDUCE NMU FROM 7738 04:54:05,617 --> 04:54:08,920 ALL THESE REGIONS, ALSO INDUCE 7739 04:54:08,920 --> 04:54:11,423 CGRP AND IT WILL ALSO INDUCE GRP 7740 04:54:11,423 --> 04:54:13,625 FROM THESE NEURONS. 7741 04:54:13,625 --> 04:54:17,028 AND AGAIN, AND IT HAD NO EFFECT 7742 04:54:17,028 --> 04:54:18,129 ON VIP. 7743 04:54:18,129 --> 04:54:21,833 SO WHAT I TOLD YOU IS THAT THE 7744 04:54:21,833 --> 04:54:23,601 CYTOKINE RECEPTORS INDEED ARE 7745 04:54:23,601 --> 04:54:25,703 PRESENT ON ENTERIC NEURONS. 7746 04:54:25,703 --> 04:54:31,176 AND I GAVE YOU EXAMPLE OF PSN2 7747 04:54:31,176 --> 04:54:35,980 OR IPAN THEY'RE CALLED AND THEY 7748 04:54:35,980 --> 04:54:39,451 EXPRESS HETEROIL4 AND IL13 7749 04:54:39,451 --> 04:54:45,356 RECEPTOR, CYTOKINES COMING FROM 7750 04:54:45,356 --> 04:54:46,791 LY THE CELLS WILL INDUCE 7751 04:54:46,791 --> 04:54:49,194 EXPRESSION OF THESE 7752 04:54:49,194 --> 04:54:51,629 NEUROPEPTIDES NMU AND CPRG FROM 7753 04:54:51,629 --> 04:54:53,164 THEM. 7754 04:54:53,164 --> 04:54:55,867 SO THE QUESTION IS THAT IN 7755 04:54:55,867 --> 04:55:00,738 ACTUALLY I TEACH THIS COURSE AT 7756 04:55:00,738 --> 04:55:03,007 HARVARD ABOUT THE BEFORE T CELL 7757 04:55:03,007 --> 04:55:05,076 SUBSETS AND THE FAVORITE LINE IS 7758 04:55:05,076 --> 04:55:08,379 THAT P THE TH2 CELLS MAKE THE 7759 04:55:08,379 --> 04:55:12,317 CYTOKINES IL2, 5 AND IL13 AND 7760 04:55:12,317 --> 04:55:14,285 IL13 ACTS ON SMOOTH MUSCLES AND 7761 04:55:14,285 --> 04:55:16,588 SMOOTH MUSCLES THEN CONTRACT AND 7762 04:55:16,588 --> 04:55:20,525 THAT GIVES YOU BRONCHOSPASMS, 7763 04:55:20,525 --> 04:55:30,935 THAT GIVES YOU -- WE HAVE A QUAN 7764 04:55:30,935 --> 04:55:33,471 DAIRY FINDING IL13 RECEPTORS, 7765 04:55:33,471 --> 04:55:36,140 THEY TOOK THE SMOOTH MUSCLES AND 7766 04:55:36,140 --> 04:55:38,309 IN FACT FROM ORGAN THEY PUT IL13 7767 04:55:38,309 --> 04:55:40,345 ON AND THEY SHOWED THAT INDUCES 7768 04:55:40,345 --> 04:55:41,813 SMOOTH MUSCLE CONTRACTION. 7769 04:55:41,813 --> 04:55:44,649 SO THE QUESTION, IS IT DIRECTLY 7770 04:55:44,649 --> 04:55:45,750 ACTING ON SMOOTH MUSCLES OR WHAT 7771 04:55:45,750 --> 04:55:48,353 IS THE ROLE OF THE IL13 RECEPTOR 7772 04:55:48,353 --> 04:55:50,889 THAT WE ARE FINDING ON THE 7773 04:55:50,889 --> 04:55:52,323 ENTERIC NERVOUS SYSTEM? 7774 04:55:52,323 --> 04:55:55,894 SO THIS IS WHAT WE TELL 7775 04:55:55,894 --> 04:55:59,297 STUDENTS, THAT THE ELEMENTS AND 7776 04:55:59,297 --> 04:56:01,699 ALLERGENS WILL ACT WITH 7777 04:56:01,699 --> 04:56:02,800 EPITHELIA AT THE BARRIER SITE 7778 04:56:02,800 --> 04:56:05,436 AND THEY WILL PRODUCE THESE 7779 04:56:05,436 --> 04:56:10,041 ALARMING, IL 25RBGS TSLP, IL33 7780 04:56:10,041 --> 04:56:13,444 WHICH WILL INDUCE ILC 2Z WHICH 7781 04:56:13,444 --> 04:56:15,613 WILL AMPLIFY TH2 RESPONSES AND 7782 04:56:15,613 --> 04:56:18,249 THEN IL13 WILL ACT ON THE SMOOTH 7783 04:56:18,249 --> 04:56:20,451 MUSCLES AND MAST CELLS AND 7784 04:56:20,451 --> 04:56:23,087 GOBLET CELLS AND EOSINOPHILS AND 7785 04:56:23,087 --> 04:56:25,356 INDUCE ALLERGIC REACTION, WHAT 7786 04:56:25,356 --> 04:56:28,092 IS CALLED WEEP AND SWEEP 7787 04:56:28,092 --> 04:56:28,393 PHENOMENON. 7788 04:56:28,393 --> 04:56:31,829 WHICH IS WHAT YOU SEE THERE'S A 7789 04:56:31,829 --> 04:56:37,669 LOT OF MUCOUS SECRETION AND 7790 04:56:37,669 --> 04:56:39,571 BRONCHOSPASMS AND DIARRHEA AND 7791 04:56:39,571 --> 04:56:40,638 VOMITING, ALL OF IT ASSOCIATED 7792 04:56:40,638 --> 04:56:44,943 WITH THE IL4, IL5, TH2 7793 04:56:44,943 --> 04:56:45,276 RESPONSES. 7794 04:56:45,276 --> 04:56:46,711 BUT THEN HERE IS THE PROBLEM. 7795 04:56:46,711 --> 04:56:48,279 THE PROBLEM IS THAT WE ARE 7796 04:56:48,279 --> 04:56:50,114 FINDING THAT THE IL13 RECEPTORS 7797 04:56:50,114 --> 04:56:53,217 AND IL4 RECEPTOR INDEED PRESENT 7798 04:56:53,217 --> 04:56:54,719 ON THE ENTERIC NERVOUS SYSTEM 7799 04:56:54,719 --> 04:56:56,254 AND OTHER NERVOUS SYSTEMS, 7800 04:56:56,254 --> 04:56:57,922 NEURONS AS WELL, AND WHAT'S THE 7801 04:56:57,922 --> 04:57:01,392 ROLE OF THESE CYTOKINE RECEPTORS 7802 04:57:01,392 --> 04:57:04,696 ON NERVOUS SYSTEM? 7803 04:57:04,696 --> 04:57:07,432 AND WE GOT THIS IL13 RECEPTOR 7804 04:57:07,432 --> 04:57:12,470 KNOCK-OUT MOUSE AND IN FACT THE 7805 04:57:12,470 --> 04:57:14,739 BLACK C I S MICE RNLT VERY GOOD 7806 04:57:14,739 --> 04:57:16,507 AT RESPONDING AND MOUNTING 7807 04:57:16,507 --> 04:57:18,176 ALLERGIC REACTION, SO WE HAVE TO 7808 04:57:18,176 --> 04:57:21,346 ACTUALLY USE THIS MODEL WHERE 7809 04:57:21,346 --> 04:57:23,848 THE PARASITE GOES IN AND BURROWS 7810 04:57:23,848 --> 04:57:26,784 IN THE GUT INTO THE MUSCULARIS, 7811 04:57:26,784 --> 04:57:30,054 I HAD A VERY NICE PICTURE, IT 7812 04:57:30,054 --> 04:57:34,892 GOES VERY CLOSE TO THE MYENTERIC 7813 04:57:34,892 --> 04:57:36,094 PLEXUS IN THE MUSCLE AND COMES? 7814 04:57:36,094 --> 04:57:38,830 VERY CLOSE TO THESE NEURONS THAT 7815 04:57:38,830 --> 04:57:41,499 PRODUCE MMU -- PRODUCE NMU AND 7816 04:57:41,499 --> 04:57:46,237 MAKES A BUNDLE. 7817 04:57:46,237 --> 04:57:49,073 AND WE WANTED TO DIG THIS IL13 7818 04:57:49,073 --> 04:57:51,142 RECEPTOR FLOCKS ALLELE AND 7819 04:57:51,142 --> 04:57:54,545 WANTED TO GET RID OF ONLY ON THE 7820 04:57:54,545 --> 04:57:56,214 ENTERIC NEURONS. 7821 04:57:56,214 --> 04:58:00,485 BUT THE PROBLEM IS THERE IS NOT 7822 04:58:00,485 --> 04:58:04,255 A SPECIFIC CRE DRIVER THAT ONLY 7823 04:58:04,255 --> 04:58:06,624 DELETES IT FROM ENTERIC NERVOUS 7824 04:58:06,624 --> 04:58:06,858 SYSTEM. 7825 04:58:06,858 --> 04:58:10,328 IN FACT, ROCKY TOOK THE IL13 7826 04:58:10,328 --> 04:58:11,863 FLOCKS FLOCKS ALLELE AND STARTED 7827 04:58:11,863 --> 04:58:13,431 CROSSING TO EVERY CRE HE COULD 7828 04:58:13,431 --> 04:58:15,166 FIND AND THERE WAS A WHOLE 7829 04:58:15,166 --> 04:58:17,802 ANIMAL HOUSE BREEDING WITH ONE 7830 04:58:17,802 --> 04:58:21,706 JUST FLOCKS ONE FLOX ALLELE. 7831 04:58:21,706 --> 04:58:23,908 HE COULD FIND A COUPLE OF CRE 7832 04:58:23,908 --> 04:58:26,844 DRIVERS THAT ARE DELETING IL13 7833 04:58:26,844 --> 04:58:29,147 REACCEPT NOR A CELL INTRINSIC 7834 04:58:29,147 --> 04:58:32,116 MANNER IN THE ENTERIC NERVES. 7835 04:58:32,116 --> 04:58:34,852 AND WHAT HE DID THEN IS THAT HE 7836 04:58:34,852 --> 04:58:37,822 USED A COUPLE OF THESE DRIVERS, 7837 04:58:37,822 --> 04:58:41,526 THIS IS CRE CROSSED ON IL 7838 04:58:41,526 --> 04:58:45,263 RECEPTOR FLOX-FLOX AEMPLOY LEGAL 7839 04:58:45,263 --> 04:58:47,532 AND INJECTED THEM AND FIRST 7840 04:58:47,532 --> 04:58:49,200 LOOKED WHAT THOOPS THE EGG 7841 04:58:49,200 --> 04:58:50,635 BURDEN AND IT WAS VERY CLEAR 7842 04:58:50,635 --> 04:58:54,872 THAT IF YOU DELETE THE IL13 7843 04:58:54,872 --> 04:58:56,507 RECEPTOR JUST FROM ENTERIC 7844 04:58:56,507 --> 04:58:58,710 NERVOUS SYSTEM AND ENTERIC 7845 04:58:58,710 --> 04:59:03,748 NEUROHONS, YOU HAVE INCREASED 7846 04:59:03,748 --> 04:59:05,049 FOCUNDITY, TECHNICAL WORD FOR 7847 04:59:05,049 --> 04:59:05,717 INCREASED EGG BURDEN. 7848 04:59:05,717 --> 04:59:09,554 I HAD TO LOOK IT UP. 7849 04:59:09,554 --> 04:59:13,257 AND THEN IF YOU CROSS IT ONTO 7850 04:59:13,257 --> 04:59:15,593 INDUCIBLE I BELIEVE CGRP CRE 7851 04:59:15,593 --> 04:59:18,629 DRIVER AND YOU CAN'T EXPEL WORMS 7852 04:59:18,629 --> 04:59:24,135 AS WELL AS THE WILD-TYPE MICE. 7853 04:59:24,135 --> 04:59:26,104 BUT IT IS NOT THAT EVERY CRE 7854 04:59:26,104 --> 04:59:31,709 DRIVER DOES IT, IN FACT -- WHICH 7855 04:59:31,709 --> 04:59:33,745 DELET'S FROMS VAIOUS NERVE ASK 7856 04:59:33,745 --> 04:59:35,947 OTHER NERVES AS WELL, NO EFFECT 7857 04:59:35,947 --> 04:59:38,883 ON FOCUNDITY OR ACTUALLY 7858 04:59:38,883 --> 04:59:40,118 EXPELLING WORMTZ, SO IT LOOKS 7859 04:59:40,118 --> 04:59:41,185 LIKE ONE OF THE FUNCTIONS OF 7860 04:59:41,185 --> 04:59:44,455 THESE NEURONS, CYTOKINE 7861 04:59:44,455 --> 04:59:46,858 RECEPTORS OR NEURONS IS IN FACT 7862 04:59:46,858 --> 04:59:48,493 PROMOTE THE GUT MOTILITY, IN 7863 04:59:48,493 --> 04:59:51,362 FACT, THAT'S WHAT THE MYENTERIC 7864 04:59:51,362 --> 04:59:53,998 PLEXUS DOES, IN FACT, THIS PSM2 7865 04:59:53,998 --> 04:59:55,399 GOES ALL THE WAY AND IN FACT 7866 04:59:55,399 --> 04:59:57,135 INCREASE THE MUSCULAR 7867 04:59:57,135 --> 04:59:58,035 CONTRACTIONS, THAT'S WHAT THEIR 7868 04:59:58,035 --> 04:59:59,370 JOB IS. 7869 04:59:59,370 --> 05:00:01,038 AND WILL WHAT YOU HAVE IS THAT 7870 05:00:01,038 --> 05:00:03,141 IT CAN'T HAVE AS MUCH, THEY 7871 05:00:03,141 --> 05:00:06,077 CAN'T EXPEL WORMS AND THERE'S AN 7872 05:00:06,077 --> 05:00:09,247 INCREASE FOCUNDITY IN THOSE 7873 05:00:09,247 --> 05:00:09,714 MICE. 7874 05:00:09,714 --> 05:00:13,851 WHAT'S ALSO INTERESTING IS THAT 7875 05:00:13,851 --> 05:00:17,588 THESE LARVAES BURROW INSIDE AND 7876 05:00:17,588 --> 05:00:18,990 FORM THESE LARGE WOUNDS AND YOU 7877 05:00:18,990 --> 05:00:21,993 CAN SEE HEMORRHAGIC WOUNDS, IF 7878 05:00:21,993 --> 05:00:24,595 YOU DELETE THE IL13 RECEPTOR 7879 05:00:24,595 --> 05:00:26,497 FROM THE NEURONS. 7880 05:00:26,497 --> 05:00:28,099 AND IT'S ACTUALLY VERY 7881 05:00:28,099 --> 05:00:29,400 SIGNIFICANT, ONLY A COUPLE OF 7882 05:00:29,400 --> 05:00:30,368 EXPERIMENTS YOU CAN SEE THERE 7883 05:00:30,368 --> 05:00:31,602 ARE LARGER WOUNDS, THEY DON'T 7884 05:00:31,602 --> 05:00:35,173 HEAL AND THEY BECOME VERY 7885 05:00:35,173 --> 05:00:35,473 HEMORRHAGIC. 7886 05:00:35,473 --> 05:00:37,375 THEN THE QUESTION IS THAT IF 7887 05:00:37,375 --> 05:00:39,977 IL13 RECEPTOR, IL4 RECEPTOR IS 7888 05:00:39,977 --> 05:00:43,247 EXPRESSED ON ENTERIC NEUROTONS, 7889 05:00:43,247 --> 05:00:44,682 SENSORY NEURONS, WHAT IS ITS 7890 05:00:44,682 --> 05:00:46,384 NORMAL JOB? 7891 05:00:46,384 --> 05:00:48,786 BECAUSE AS YOU REMEMBER THAT WE 7892 05:00:48,786 --> 05:00:52,490 HAVE HOMEOSTATIC ILC 2Z UNDER 7893 05:00:52,490 --> 05:00:54,559 THE MUCOSA THAT ARE PRODUCING 7894 05:00:54,559 --> 05:00:57,962 IL4, IL13 AT LOW LEVELS ALL THE 7895 05:00:57,962 --> 05:00:58,296 TIME. 7896 05:00:58,296 --> 05:00:59,831 AND WE STARTED LOOKING AT WHAT 7897 05:00:59,831 --> 05:01:01,966 HAPPENS TO NERVES THEMSELVES AT 7898 05:01:01,966 --> 05:01:05,536 THE HOMEOSTATIC LEVEL AND 7899 05:01:05,536 --> 05:01:10,041 FOLLOWING INFECTION WITH -- AND 7900 05:01:10,041 --> 05:01:11,175 ONE OF THE VERY FIRST THINGS 7901 05:01:11,175 --> 05:01:21,719 THAT YOU SEE -- AND ACTUALLY IT 7902 05:01:21,953 --> 05:01:24,956 DROPS HAD AND YOU ACTUALLY START 7903 05:01:24,956 --> 05:01:27,024 LOSING NUMBER OF NEURONS FROM 7904 05:01:27,024 --> 05:01:31,295 THE MYENTERIC PLEXUS INNERVATING 7905 05:01:31,295 --> 05:01:33,497 THE V I LLI. 7906 05:01:33,497 --> 05:01:35,032 THAT'S THE FIRST THING YOU SEE. 7907 05:01:35,032 --> 05:01:36,234 SECOND THING YOU SEE IS THAT 7908 05:01:36,234 --> 05:01:37,869 THEY START PRODUCING THE 7909 05:01:37,869 --> 05:01:38,636 NEUROPEPTIDES THAT I TALKED TO 7910 05:01:38,636 --> 05:01:40,538 YOU ABOUT THAT MODULATE OR 7911 05:01:40,538 --> 05:01:43,608 REGULATE IMMUNE SYSTEM. 7912 05:01:43,608 --> 05:01:45,243 SO WHAT DO THEY TELL YOU IN THE 7913 05:01:45,243 --> 05:01:51,816 SUMMARY NUMBER TWO IS THAT IF 7914 05:01:51,816 --> 05:01:58,756 YOU INFECT THESE MICE WITH 7915 05:01:58,756 --> 05:02:01,259 H POLYGYRUS AND NMU PRODUCING 7916 05:02:01,259 --> 05:02:03,694 NEURONS IN THE WILD-TYPE MICE 7917 05:02:03,694 --> 05:02:05,730 YOU EXPEL WORMS AND YOU ALSO 7918 05:02:05,730 --> 05:02:07,698 DON'T HAVE AS MUCH EGG BURDEN, 7919 05:02:07,698 --> 05:02:10,701 THE MOMENT YOU GET RID OF IL13 7920 05:02:10,701 --> 05:02:12,203 RECEPTOR ON ENTERIC NERVOUS 7921 05:02:12,203 --> 05:02:17,575 SYSTEM YOU DON'T HAVE AS MANY 7922 05:02:17,575 --> 05:02:19,143 PSM2 NEURONS GOING THROUGH INTO 7923 05:02:19,143 --> 05:02:23,547 THE VILLI, THE NEURON ACTUALLY 7924 05:02:23,547 --> 05:02:26,150 COLLAPSES AND YOU MAINTAIN A 7925 05:02:26,150 --> 05:02:30,755 LARGER NUMBER OF THESE WORMS AND 7926 05:02:30,755 --> 05:02:31,956 FOCUNDITY IN THE GUT AND YOU 7927 05:02:31,956 --> 05:02:35,593 CAN'T EXPEL WORMS AS MUCH. 7928 05:02:35,593 --> 05:02:36,027 OKAY? 7929 05:02:36,027 --> 05:02:40,298 NOW, THE QUESTION IS, AND HOW 7930 05:02:40,298 --> 05:02:43,334 DOES IT DO IT? 7931 05:02:43,334 --> 05:02:47,638 AND IN FACT, WE HAVE STHEEN DATA 7932 05:02:47,638 --> 05:02:48,739 FROM EVERY WHICH WAY YOU CAN 7933 05:02:48,739 --> 05:02:50,241 THINK OF AND I'M GOING ON BORE 7934 05:02:50,241 --> 05:02:51,342 YOU WITH ALL OF THAT. 7935 05:02:51,342 --> 05:03:01,319 SO WE TOOK ALL THE RECALL CELLS 7936 05:03:01,319 --> 05:03:03,587 ANDLY SEQ AND LOOKED AT 7937 05:03:03,587 --> 05:03:05,056 MUSCULARIS CELLS IN MUSCULARIS 7938 05:03:05,056 --> 05:03:09,994 AND NEURONS IN AND LOOKED AT 7939 05:03:09,994 --> 05:03:11,062 EPITHELIAL CELLS AND I'LL SHOW 7940 05:03:11,062 --> 05:03:12,296 YOU ONE SLIDE AND ANYBODY WHO 7941 05:03:12,296 --> 05:03:14,432 HAS QUESTIONS CAN TALK TO ME. 7942 05:03:14,432 --> 05:03:16,400 SO WE TOOK THIS WILD-TYPE AND 7943 05:03:16,400 --> 05:03:18,602 KNOCK-OUT MOUSE AND INFECTED 7944 05:03:18,602 --> 05:03:23,441 THEM WITH THE H PHYSICAL POLYAND 7945 05:03:23,441 --> 05:03:26,143 YOU CAN MAKE ALL THE SEQ YOU 7946 05:03:26,143 --> 05:03:28,546 WANT, MAKE ALL THESE NEW MAPS 7947 05:03:28,546 --> 05:03:29,547 AND ONE OF THE FIRST THING YOU 7948 05:03:29,547 --> 05:03:31,082 SEE IS THESE MICE IN WHICH YOU 7949 05:03:31,082 --> 05:03:33,351 HAVE DELETED IL13 RECEPTOR FROM 7950 05:03:33,351 --> 05:03:36,320 THE NEURONS, THERE'S A DEFECT IN 7951 05:03:36,320 --> 05:03:38,389 BOTH THE FREQUENCY AND THE 7952 05:03:38,389 --> 05:03:42,093 AMOUNT OF ILC2'S IN SUBMUCOSA. 7953 05:03:42,093 --> 05:03:43,227 THAT'S THE FIRST THING THAT YOU 7954 05:03:43,227 --> 05:03:45,296 SEE. 7955 05:03:45,296 --> 05:03:45,563 OKAY? 7956 05:03:45,563 --> 05:03:49,266 WHAT YOU ALSO SEE IS MAXIMUM 7957 05:03:49,266 --> 05:03:52,036 NUMBER OF INFECT CHANGED IN THE 7958 05:03:52,036 --> 05:03:53,771 MYELOID CELLS -- CHANGES IN THE 7959 05:03:53,771 --> 05:03:55,539 MYELOID CELLS, AND LOOKING AT 7960 05:03:55,539 --> 05:03:58,009 WHAT ARE THE CHANGES AND THE 7961 05:03:58,009 --> 05:03:59,577 CHANGES ARE ASSOCIATED WITH I 7962 05:03:59,577 --> 05:04:02,146 CAN'T READ IT FROM HERE, IS THAT 7963 05:04:02,146 --> 05:04:04,548 THERE'S A DEFECT IN INTERACTION 7964 05:04:04,548 --> 05:04:06,450 BETWEEN SPECIES, THAT MEANS THE 7965 05:04:06,450 --> 05:04:08,419 WORM IS BURROWING INTO A MAMMAL 7966 05:04:08,419 --> 05:04:10,488 AND THAT PATHWAY IS DEFECTIVE. 7967 05:04:10,488 --> 05:04:14,225 THERE'S A DEFECT IN MOUNTING 7968 05:04:14,225 --> 05:04:15,226 INFLAMMATION, AND SOME OF THE 7969 05:04:15,226 --> 05:04:18,763 HEALING PROCESSES ARE LOST IN 7970 05:04:18,763 --> 05:04:22,900 THE IL13 RECEPTOR DELETION ON 7971 05:04:22,900 --> 05:04:24,235 ENTERIC NEURONS. 7972 05:04:24,235 --> 05:04:25,302 THERE'S ALSO SOMETHING THAT I 7973 05:04:25,302 --> 05:04:27,705 HAVE TAKEN OUT OF THOSE SLIDES 7974 05:04:27,705 --> 05:04:29,273 COMPLETELY, THERE'S A CHANGE IN 7975 05:04:29,273 --> 05:04:33,177 THE NUMBER OF PANA CELLS ARE 7976 05:04:33,177 --> 05:04:34,612 DECREASING, THERE'S A REMODELING 7977 05:04:34,612 --> 05:04:37,181 OF GUT EPITHELIALIUM. 7978 05:04:37,181 --> 05:04:40,551 IMAGINE YOU ONLY DELETED IL 7979 05:04:40,551 --> 05:04:41,619 RECEPTOR FROM THE NEUROHON, YOU 7980 05:04:41,619 --> 05:04:44,021 ARE CHANGING THE GUT BARRIER HAS 7981 05:04:44,021 --> 05:04:46,757 CHANGED AND THERE'S A CHANGE IN 7982 05:04:46,757 --> 05:04:48,726 EPITHELIA IN THE GUT, YOU CAN'T 7983 05:04:48,726 --> 05:04:56,133 MAKE AS MANICALLY PEPTIDES TRKTS 7984 05:04:56,133 --> 05:04:57,368 ENTEROMICROPEPTIDES GO DOWN AND 7985 05:04:57,368 --> 05:04:59,637 THE HEALING, WHAT YOU SEE 7986 05:04:59,637 --> 05:05:01,138 NORMALLY UPON INFECTION IS 7987 05:05:01,138 --> 05:05:04,141 ACTUALLY REGULATED BY THESE IL13 7988 05:05:04,141 --> 05:05:05,376 RECEPTOR OR NEURONS SOMETHING, 7989 05:05:05,376 --> 05:05:08,979 THAT I AM NO GISTS IT'S HARD TO 7990 05:05:08,979 --> 05:05:10,881 BELIEVE BUT THAT'S WHAT YOU SEE. 7991 05:05:10,881 --> 05:05:18,189 SO LET'S MAKE SURE WE SUM IT UP. 7992 05:05:18,189 --> 05:05:20,124 RECEPTORS ON ENTERIC NEURONS, 7993 05:05:20,124 --> 05:05:23,661 AND I GAVE YOU EXAMPLE OF IL4, 7994 05:05:23,661 --> 05:05:27,498 IL13 HEAD TROA DIMER AND PSM2 7995 05:05:27,498 --> 05:05:29,867 SENSORY NEURONS, AND WHEN YOU 7996 05:05:29,867 --> 05:05:31,769 DELETE IT AND YOU ARE NOT ABLE 7997 05:05:31,769 --> 05:05:36,807 TO EXPEL WORMS AND THIS INCREASE 7998 05:05:36,807 --> 05:05:39,310 FOCUNDITY, AND THEN WHAT YOU DO 7999 05:05:39,310 --> 05:05:41,312 ALSO IS THERE'S ACTIVATION 8000 05:05:41,312 --> 05:05:43,380 PATHWAY BUT IL13 ACTING ON THE 8001 05:05:43,380 --> 05:05:45,583 NEURONS MAKING NEUROPEPTIDES AND 8002 05:05:45,583 --> 05:05:46,784 ALSO WE'RE FINDING A WHOLE 8003 05:05:46,784 --> 05:05:50,054 SERIES OF NEW NEUROPEPTIDES THAT 8004 05:05:50,054 --> 05:05:51,455 THE NEUROSCIENTISTS HAVE NOT 8005 05:05:51,455 --> 05:05:52,790 ACTUALLY SEEN BEFORE. 8006 05:05:52,790 --> 05:05:54,658 AND BECAUSE THEY ARE ALWAYS 8007 05:05:54,658 --> 05:05:56,193 USING NEURONAL TRIGGERS, HERE WE 8008 05:05:56,193 --> 05:05:59,196 ARE USING CYTOKINE AS A TRIGGER, 8009 05:05:59,196 --> 05:06:01,999 AND THESE NEUROPEPTIDES ARE 8010 05:06:01,999 --> 05:06:06,237 ACTIVATING THE MYELOID CELLS. 8011 05:06:06,237 --> 05:06:08,472 AND ALSO, SOME INTERACTION WE 8012 05:06:08,472 --> 05:06:11,075 DON'T KNOW WHAT THAT IS 8013 05:06:11,075 --> 05:06:13,711 GENERATED BY NEURONS TO REGULATE 8014 05:06:13,711 --> 05:06:15,579 THE EPITHELIAL BARRIER WHEN YOU 8015 05:06:15,579 --> 05:06:18,516 LOSE IT, YOUR EPITHELIAL BARRIER 8016 05:06:18,516 --> 05:06:21,819 IS AFFECTED BY JUST CHANGING THE 8017 05:06:21,819 --> 05:06:22,820 EFFECT ON NEURONS. 8018 05:06:22,820 --> 05:06:23,721 LET ME SUM IT UP. 8019 05:06:23,721 --> 05:06:25,422 THIS IS THE LAST PART OF MY 8020 05:06:25,422 --> 05:06:25,623 TALK. 8021 05:06:25,623 --> 05:06:27,424 SO THAT I SHOWED YOU THAT THE 8022 05:06:27,424 --> 05:06:29,593 CYTOKINE RECEPTORS ARE ON 8023 05:06:29,593 --> 05:06:32,129 ENTERIC NEURONS, AND ACTIVATION 8024 05:06:32,129 --> 05:06:37,134 WITH IL4 AND IL13 OF THIS 8025 05:06:37,134 --> 05:06:40,004 HETERODIMER WILL INDUCE 8026 05:06:40,004 --> 05:06:43,374 NEUROPEPTIDE AND IF YOU DELETE 8027 05:06:43,374 --> 05:06:45,442 IL13 RECEPTORS YOU CAN'T IN FACT 8028 05:06:45,442 --> 05:06:46,577 EXPEL WORMS AND YOU HAVE 8029 05:06:46,577 --> 05:06:49,647 INCREASED FOCUNDITY AND LOSS OF 8030 05:06:49,647 --> 05:06:51,582 IL13 RE RECEPTOR AND NEURONS 8031 05:06:51,582 --> 05:06:53,017 WILL ALSO DECREASE THE NUMBER 8032 05:06:53,017 --> 05:06:55,653 AND FREQUENCY AND THE 8033 05:06:55,653 --> 05:06:59,890 INNERVATION OF THE NM -- NMU 8034 05:06:59,890 --> 05:07:00,791 POSITIVE NEURONS. 8035 05:07:00,791 --> 05:07:02,760 I SHOWED YOU THAT NEURONS 8036 05:07:02,760 --> 05:07:04,628 REMODEL THE GUT EPITHELIUM AND 8037 05:07:04,628 --> 05:07:06,463 PRODUCTION OF MANY OF THE 8038 05:07:06,463 --> 05:07:09,099 ANTIMICROBIAL PEPTIDES. 8039 05:07:09,099 --> 05:07:10,534 JUST WHAT I TOLD YOU FROM THE 8040 05:07:10,534 --> 05:07:11,735 VERY BEGINNING, 30 YEARS AGO 8041 05:07:11,735 --> 05:07:12,836 WHEN I SAID THAT MAYBE THE 8042 05:07:12,836 --> 05:07:13,904 NERVOUS SYSTEM HAS SOMETHING TO 8043 05:07:13,904 --> 05:07:15,873 DO WITH THE IMMUNE SYSTEM AND IN 8044 05:07:15,873 --> 05:07:18,075 FACT FROM MY SON'S ALLERGY 8045 05:07:18,075 --> 05:07:19,310 BOUTS, AND THIS IS PROBABLY ONE 8046 05:07:19,310 --> 05:07:20,277 OF THE THINGS AT THAT CAME OUT 8047 05:07:20,277 --> 05:07:24,848 OF IT, IS THAT YOU INDEED 8048 05:07:24,848 --> 05:07:25,983 THERE'S INDEED INTERACTION 8049 05:07:25,983 --> 05:07:29,119 BETWEEN THE TWO SYSTEMS, WE 8050 05:07:29,119 --> 05:07:31,322 DON'T LIVE IN SILOS, AND 8051 05:07:31,322 --> 05:07:32,623 IMMUNOLOGY IS NOT WORKING ON 8052 05:07:32,623 --> 05:07:33,624 IMMUNE SYSTEM ALONE. 8053 05:07:33,624 --> 05:07:35,259 IT'S WORKING WITH EVERYBODY ELSE 8054 05:07:35,259 --> 05:07:36,360 THAT'S IN THE SYSTEM. 8055 05:07:36,360 --> 05:07:38,329 SO I SHOWED YOU THE CYTOKINES 8056 05:07:38,329 --> 05:07:39,863 PRODUCED BY AT LOW LEVELS 8057 05:07:39,863 --> 05:07:41,298 GENERALLY ARE PRODUCED BY THE 8058 05:07:41,298 --> 05:07:43,400 INNATE LYMPHOID CELLS AND THE 8059 05:07:43,400 --> 05:07:45,102 CYTOKINE RECEPTORS ARE ON THE 8060 05:07:45,102 --> 05:07:48,005 NEURONS AND THE NEURONS PRODUCE 8061 05:07:48,005 --> 05:07:49,640 NEUROPEPTIDES AND THAT WILL 8062 05:07:49,640 --> 05:07:52,409 BECOME A SELF-AMPLIFYING LOOP. 8063 05:07:52,409 --> 05:07:55,846 THIS MUST BE ESSENTIAL, IN FACT, 8064 05:07:55,846 --> 05:07:57,014 THE NEUROPEPTIDES THAT WE ARE 8065 05:07:57,014 --> 05:08:00,818 SEEING BY CYTOKINE ACTIVATION OF 8066 05:08:00,818 --> 05:08:02,219 NEURONS, THESE NOVEL 8067 05:08:02,219 --> 05:08:05,489 NEUROPEPTIDES HAVE VERY POTENT 8068 05:08:05,489 --> 05:08:06,790 ANTIMICROBIAL ACTIVITY AND MAYBE 8069 05:08:06,790 --> 05:08:08,325 THE NERVOUS SYSTEM IS PROBABLY 8070 05:08:08,325 --> 05:08:10,160 THE FIRST INNATE IMMUNE SYSTEM 8071 05:08:10,160 --> 05:08:12,730 THAT EVER EXISTED. 8072 05:08:12,730 --> 05:08:16,300 AND IN FACT, THEY DIRECTLY 8073 05:08:16,300 --> 05:08:19,069 AFFECT THE MICROBIOME AND 8074 05:08:19,069 --> 05:08:21,171 MICROBIAL SPECIES, IN FACT, IT 8075 05:08:21,171 --> 05:08:23,540 CHANGES THE MICROBIOTA. 8076 05:08:23,540 --> 05:08:24,975 ACTUALLY I DON'T WANT TO BLABBER 8077 05:08:24,975 --> 05:08:26,644 TOO MUCH AND I WANT TO INTRODUCE 8078 05:08:26,644 --> 05:08:30,481 YOU TO MY LAB, AND WE HAVE A 8079 05:08:30,481 --> 05:08:36,053 SOCCER TEAM, IT'S CALLED CT 8080 05:08:36,053 --> 05:08:38,455 KUCHROO IT LIONS, WE HAVE LOST 8081 05:08:38,455 --> 05:08:39,890 EVERY GAME WE HAVE PLAYED. 8082 05:08:39,890 --> 05:08:40,457 [LAUGHTER] 8083 05:08:40,457 --> 05:08:42,092 FINALLY BEFORE COVID WE DID WIN 8084 05:08:42,092 --> 05:08:44,061 A GAME, AND THEN WE HAD ALL 8085 05:08:44,061 --> 05:08:45,829 THESE POSTERS AND MAGNETS THAT 8086 05:08:45,829 --> 05:08:48,632 WE BECAME THE CHAMPIONS OF 8087 05:08:48,632 --> 05:08:52,169 HARVARD MEDICAL SCHOOL, AND THIS 8088 05:08:52,169 --> 05:08:54,872 IS ROCKY BARELLA WHO DID MOST OF 8089 05:08:54,872 --> 05:08:56,106 THE WORK. 8090 05:08:56,106 --> 05:08:57,341 I'M HAPPY TO TAKE ANY QUESTIONS 8091 05:08:57,341 --> 05:08:58,008 YOU MAY HAVE. 8092 05:08:58,008 --> 05:09:02,046 THANK YOU FOR YOUR ATTENTION. 8093 05:09:02,046 --> 05:09:06,216 [APPLAUSE] 8094 05:09:06,216 --> 05:09:08,352 >> THANK YOU. 8095 05:09:08,352 --> 05:09:11,322 MITCHELL CORN BELLE BERG -- 8096 05:09:11,322 --> 05:09:11,588 INSTITUTE. 8097 05:09:11,588 --> 05:09:18,128 I'M SUPPOSED TO INTRODUCE -- 8098 05:09:18,128 --> 05:09:20,564 ILC2 CAN MAKE OTHER THINGS, CAN 8099 05:09:20,564 --> 05:09:24,802 MAKE REGULAR LYNN, MAYBE IL9 AND 8100 05:09:24,802 --> 05:09:25,069 CAPULENS. 8101 05:09:25,069 --> 05:09:31,075 DO YOU SEE RECEPTORS FOR OTHER 8102 05:09:31,075 --> 05:09:33,811 ILC2 PRODUCTS ON THE NERVES? 8103 05:09:33,811 --> 05:09:38,615 >> I THINK THIS IS GOING TO BE A 8104 05:09:38,615 --> 05:09:40,984 BE -- AND WE ARE FINDING THINGS 8105 05:09:40,984 --> 05:09:42,820 THAT WE NEVER THOUGHT EXISTED. 8106 05:09:42,820 --> 05:09:44,888 AND THERE ARE LOTS OF CYTOKINE 8107 05:09:44,888 --> 05:09:47,057 RECEPTORS AND OTHER MOLECULAR 8108 05:09:47,057 --> 05:09:48,359 RECEPTORS ON THESE NEURONS AS 8109 05:09:48,359 --> 05:09:50,327 YOU GO THROUGH THE ATLAS. 8110 05:09:50,327 --> 05:09:55,866 AND WE THINK THE EXPRESSION OF 8111 05:09:55,866 --> 05:09:57,835 AMPHOREGLAN BY THE ILC2'S IS 8112 05:09:57,835 --> 05:09:59,269 VERY IMPORTANT FOR MAINTAINING 8113 05:09:59,269 --> 05:10:02,139 THE NEURONS AND NEURON BARRIER 8114 05:10:02,139 --> 05:10:04,174 FUNCTIONS AND ALSO MAINTAINING 8115 05:10:04,174 --> 05:10:04,808 HOMOEOSTASIS. 8116 05:10:04,808 --> 05:10:06,243 SO ACTUALLY ONCE WE STARTED 8117 05:10:06,243 --> 05:10:07,644 HAVING THE CROSSTALK BETWEEN THE 8118 05:10:07,644 --> 05:10:09,613 TWO SYSTEMS, ACTUALLY IT'S A 8119 05:10:09,613 --> 05:10:13,784 WHOLE NEW WORLD. 8120 05:10:13,784 --> 05:10:14,985 >> CL (AWAY FROM MICROPHONE). 8121 05:10:14,985 --> 05:10:17,488 >> CORRECT. 8122 05:10:17,488 --> 05:10:18,722 CORRECT. 8123 05:10:18,722 --> 05:10:22,626 >> JAY -- 8124 05:10:22,626 --> 05:10:25,729 >> JAY FROM NCI. 8125 05:10:25,729 --> 05:10:27,498 VIJAY, THAT'S A FASCINATING 8126 05:10:27,498 --> 05:10:27,698 TALK. 8127 05:10:27,698 --> 05:10:30,567 SINCE BOTH IL4 AND IL13 CAN ACT 8128 05:10:30,567 --> 05:10:32,669 ON THAT SAME RECEPTOR THAT 8129 05:10:32,669 --> 05:10:39,109 INVOLVES IL4 ALPHA AND IL13 8130 05:10:39,109 --> 05:10:40,277 ALPHA IS THERE SOME PREFERENCE 8131 05:10:40,277 --> 05:10:43,046 FOR IL13 AS YOU WERE DESCRIBING 8132 05:10:43,046 --> 05:10:45,215 IT, IS IT BECAUSE IL13 IS MORE 8133 05:10:45,215 --> 05:10:49,820 PREVALENT IN THAT ENVIRONMENT? 8134 05:10:49,820 --> 05:10:52,055 THEY BOTH INDUCE PHOSPHORYLATION 8135 05:10:52,055 --> 05:10:53,390 DOWNSTREAM OF THE RECEPTOR. 8136 05:10:53,390 --> 05:10:55,993 >> SO I TOOK ALL THAT DATA OUT 8137 05:10:55,993 --> 05:10:58,929 AND IN FACT, WE CAN CLEARLY SEE 8138 05:10:58,929 --> 05:11:01,698 THAT UPON ACTIVATION WITH EITHER 8139 05:11:01,698 --> 05:11:05,602 IL4 OR IL13 YOU GET 8140 05:11:05,602 --> 05:11:07,237 PHOSPHORYLATION OF THE RECEPTOR 8141 05:11:07,237 --> 05:11:10,607 TAIL, AND YOU ALSO INDUCE STAT 6 8142 05:11:10,607 --> 05:11:13,410 IN THESE NEURONS. 8143 05:11:13,410 --> 05:11:15,813 AND EVERYTHING LOOKS NORMAL. 8144 05:11:15,813 --> 05:11:17,681 THE REASON WE WENT THROUGH IL13 8145 05:11:17,681 --> 05:11:21,718 IS SIMPLY BECAUSE OF I THOUGHT 8146 05:11:21,718 --> 05:11:23,253 IL13 ACTS ON SMOOTH MUSCLES TO 8147 05:11:23,253 --> 05:11:25,889 INDUCE THE VOMITING AND DIARRHEA 8148 05:11:25,889 --> 05:11:27,991 AND BRONCHOSPOOSM WENT AFTER 8149 05:11:27,991 --> 05:11:29,493 IL13 RECEPTOR DELETION. 8150 05:11:29,493 --> 05:11:30,928 BUT WE COULD HAVE EASILY DONE IT 8151 05:11:30,928 --> 05:11:34,331 WITH IL4 RECEPTOR BECAUSE BOTH 8152 05:11:34,331 --> 05:11:36,500 IL4 RECEPTOR AND IL13 RECEPTOR 8153 05:11:36,500 --> 05:11:39,703 ARE FORM RNG THE HETERODIMER AND 8154 05:11:39,703 --> 05:11:44,508 IL4, IL13 ACTIVATION WILL INDUCE 8155 05:11:44,508 --> 05:11:46,076 BOTH THOSE NEUROPEPTIDES. 8156 05:11:46,076 --> 05:11:49,680 >> IN 234 AL WE STUDIED ONE OF 8157 05:11:49,680 --> 05:11:51,715 THE REASONS WHY IN THAT CASE 8158 05:11:51,715 --> 05:11:56,653 IL13 WORKED AND IL BE 4 DIDN'T 8159 05:11:56,653 --> 05:11:58,722 WAS THAT YOU ALSO NEEDED TO 8160 05:11:58,722 --> 05:12:02,392 ACTIVATE THE IL13 OR ALPHA 2. 8161 05:12:02,392 --> 05:12:03,660 >> YEAH. 8162 05:12:03,660 --> 05:12:06,196 >> THAT IT RESPONDS ONLY TO IL13 8163 05:12:06,196 --> 05:12:08,365 AND THAT UP REGULATE THE OTHER 8164 05:12:08,365 --> 05:12:08,632 RECEPTORS. 8165 05:12:08,632 --> 05:12:11,001 >> CORRECT. 8166 05:12:11,001 --> 05:12:13,737 BUT IN THE NEURONS IT'S 8167 05:12:13,737 --> 05:12:16,573 SPECIFICALLY THESE TWO RECEPTORS 8168 05:12:16,573 --> 05:12:19,977 FORM THE HETERODIMER THAT ARE 8169 05:12:19,977 --> 05:12:20,277 RESPONDING. 8170 05:12:20,277 --> 05:12:21,178 >> THANK YOU. 8171 05:12:21,178 --> 05:12:26,216 >> HI. 8172 05:12:26,216 --> 05:12:27,985 IN. 8173 05:12:27,985 --> 05:12:30,387 REALLY BEAUTIFUL WORK YOU D I 8174 05:12:30,387 --> 05:12:31,922 WAS REALLY STRUCK BY THE LOSS OF 8175 05:12:31,922 --> 05:12:33,757 INNER VAIPTION IN THE IL13 8176 05:12:33,757 --> 05:12:35,425 DEFICIENT AND HOW THAT RELATED 8177 05:12:35,425 --> 05:12:39,129 TO THE FAILURE TO SORT OF 8178 05:12:39,129 --> 05:12:40,130 REEPITHELIAL EYES. 8179 05:12:40,130 --> 05:12:41,565 WHAT DO YOU THINK THE NEURONS 8180 05:12:41,565 --> 05:12:43,100 ARE FUNCTIONING IN THAT SYSTEM? 8181 05:12:43,100 --> 05:12:44,368 THEY DON'T SEEM TO BE AT THE 8182 05:12:44,368 --> 05:12:45,469 BOTTOM, UNLESS I JUST COULDN'T 8183 05:12:45,469 --> 05:12:47,671 SEE IN THE BACK R THEY AFFECTING 8184 05:12:47,671 --> 05:12:49,439 STEM CELLS OR DIRECTLY AFFECTING 8185 05:12:49,439 --> 05:12:50,440 THE DIFFERENTIATED CELLS? 8186 05:12:50,440 --> 05:12:52,543 >> IT'S A GOOD QUESTION. 8187 05:12:52,543 --> 05:12:54,344 I DON'T HAVE A DIRECT ANSWER FOR 8188 05:12:54,344 --> 05:12:54,511 YOU. 8189 05:12:54,511 --> 05:13:01,318 WE SEE THE FREQUENCY OF NEURONS, 8190 05:13:01,318 --> 05:13:03,387 NMU POSITIVE NEURONS ACTUALLY 8191 05:13:03,387 --> 05:13:05,322 NUMBERS GO DOWN AND INNERVATION 8192 05:13:05,322 --> 05:13:08,158 GOES DOWN AND THEY ARE NOT GOING 8193 05:13:08,158 --> 05:13:11,228 DOWN FROM THE MYENTERIC PLEXUS 8194 05:13:11,228 --> 05:13:20,404 INTO THE VILLI. 8195 05:13:20,404 --> 05:13:22,573 SOILY ARE NEURONAL THAT GOES 8196 05:13:22,573 --> 05:13:24,441 INTO THE VILLI. 8197 05:13:24,441 --> 05:13:27,177 ALTHOUGH IF YOU TUK A 8198 05:13:27,177 --> 05:13:27,945 NEUROSCIENTIST, THEY -- IF YOU 8199 05:13:27,945 --> 05:13:29,913 TALK ON A NEUROSCIENTIST THEY 8200 05:13:29,913 --> 05:13:30,914 WILL ALWAYS SAY THAT ONCE YOU 8201 05:13:30,914 --> 05:13:32,416 HAVE THE NEURONS, NOTHING ELSE 8202 05:13:32,416 --> 05:13:33,850 CHANGES, WHICH IS PROBABLY NOT 8203 05:13:33,850 --> 05:13:34,084 TRUE. 8204 05:13:34,084 --> 05:13:36,019 BUT WE DO FIND THAT IT'S 8205 05:13:36,019 --> 05:13:38,689 ESSENTIAL FOR MAINTAINING THAT 8206 05:13:38,689 --> 05:13:42,826 CLUSTER OF NEURONS, THE PSM2 AND 8207 05:13:42,826 --> 05:13:44,695 NMU POSITIVE, CRG POSITIVE 8208 05:13:44,695 --> 05:13:46,229 NEURONS, BUT WE DO NOT KNOW AT 8209 05:13:46,229 --> 05:13:48,265 WHAT STAGE IS IT NECESSARY, IS 8210 05:13:48,265 --> 05:13:52,002 IT STEM CELLS MAKING THEM GROW 8211 05:13:52,002 --> 05:13:53,637 OR IS IT MAKING NEW NEURONS OR 8212 05:13:53,637 --> 05:13:54,972 IS IT MAINTAINING THE NEURONS 8213 05:13:54,972 --> 05:13:56,306 THAT YOU ALREADY HAVE, WE DO NOT 8214 05:13:56,306 --> 05:13:56,506 KNOW. 8215 05:13:56,506 --> 05:13:57,074 >> COOL. 8216 05:13:57,074 --> 05:13:58,375 THANK YOU. 8217 05:13:58,375 --> 05:14:01,011 >> SO I HAVE TWO QUICK SLIGHTLY 8218 05:14:01,011 --> 05:14:04,748 NAIVE QUESTIONS. 8219 05:14:04,748 --> 05:14:10,087 >> HOLD ON -- AM OKAY. 8220 05:14:10,087 --> 05:14:14,024 SO SAME STORY, SO TOUGH CELLS 8221 05:14:14,024 --> 05:14:16,860 AND BECAUSE AS IL33 PRODUCERS 8222 05:14:16,860 --> 05:14:19,096 AND IS THIS THE SAME PATHWAY 8223 05:14:19,096 --> 05:14:24,167 THAT WORKS IN IT CUTANEOUS 8224 05:14:24,167 --> 05:14:25,836 ALLERGIES WITH MAST CELL 8225 05:14:25,836 --> 05:14:26,803 PERIPHERAL NEURON CIRCUITS? 8226 05:14:26,803 --> 05:14:29,406 >> DO NOT KNOW BUT WE SUSPECT 8227 05:14:29,406 --> 05:14:29,906 SO. 8228 05:14:29,906 --> 05:14:31,208 >> TOUGH CELLS DO THEY HAVE A 8229 05:14:31,208 --> 05:14:32,976 ROLE IN THIS GUT SITUATION? 8230 05:14:32,976 --> 05:14:34,011 ARE THEY ACTUALLY -- 8231 05:14:34,011 --> 05:14:36,613 >> IN FACT, WE DO SEE WHEN YOU 8232 05:14:36,613 --> 05:14:38,415 DELETE ALPHA 13 RECEPTORS FROM 8233 05:14:38,415 --> 05:14:41,852 THE NEURONS YOU DO SEE CHANGE IN 8234 05:14:41,852 --> 05:14:43,186 BOTH DEPENDENT CELLS AND TOUGH 8235 05:14:43,186 --> 05:14:43,920 CELL IN THE GUT. 8236 05:14:43,920 --> 05:14:44,588 >> THANK YOU. 8237 05:14:44,588 --> 05:14:48,992 >> OKAY. 8238 05:14:48,992 --> 05:14:49,326 DAVID? 8239 05:14:49,326 --> 05:14:53,397 >> HELLO. 8240 05:14:53,397 --> 05:14:55,232 DAVID DID -- HELLO, VIJAY. 8241 05:14:55,232 --> 05:14:55,666 GREAT TALK. 8242 05:14:55,666 --> 05:14:57,868 I CAME IN A TINY BIT LATE. 8243 05:14:57,868 --> 05:14:59,302 I JUST WANTED TO REMIND MYSELF 8244 05:14:59,302 --> 05:15:01,038 AND THE AWD YEN, YOU KNOW, NOT 8245 05:15:01,038 --> 05:15:05,075 ALL WRORMS THE SAME, AS YOU KNOE 8246 05:15:05,075 --> 05:15:10,881 EXPAIM HLY POLIGYRUS IS UNUSUAL 8247 05:15:10,881 --> 05:15:12,282 BECAUSE IT LIVES IN THE 8248 05:15:12,282 --> 05:15:15,919 MUSCULARIS, IT'S NOT A MODEL FOR 8249 05:15:15,919 --> 05:15:18,455 STUDYING NEURAL HE THEY'LL YOLG 8250 05:15:18,455 --> 05:15:19,790 BIOLOGY NECESSARILY, IT'S A 8251 05:15:19,790 --> 05:15:21,625 BRILLIANT MODEL IT STUDY NEURO 8252 05:15:21,625 --> 05:15:23,160 IMMUNE INTERACTIONS WITH THE 8253 05:15:23,160 --> 05:15:24,795 MUSCULARIS PERHAPS. 8254 05:15:24,795 --> 05:15:28,498 BUT YOUR CONCLUSIONS ABOUT NEURO 8255 05:15:28,498 --> 05:15:30,033 EPITHELIAL STUFF MIGHT BE AN 8256 05:15:30,033 --> 05:15:30,901 EXAGGERATION S THAT FAIR? 8257 05:15:30,901 --> 05:15:34,938 >> FOR THIS ZOODLE OR HAVE YOU 8258 05:15:34,938 --> 05:15:37,240 DONE ALL THELY ALL THE OTHER 8259 05:15:37,240 --> 05:15:37,474 SYSTEMS? 8260 05:15:37,474 --> 05:15:39,476 SO WE DON'T LEAVE THE AUDIENCE 8261 05:15:39,476 --> 05:15:44,815 THINKING EVERY WORM IS THE SAME. 8262 05:15:44,815 --> 05:15:49,319 H POLYGYRUS IS THE LEAST USED GI 8263 05:15:49,319 --> 05:15:50,954 NEMATODE SYSTEM AND ITS UNUSUAL 8264 05:15:50,954 --> 05:15:52,355 BECAUSE ITS BIOLOGY LIVES IN THE 8265 05:15:52,355 --> 05:15:53,323 MUSCLE S THAT FAIR? 8266 05:15:53,323 --> 05:15:54,991 >> SO I THINK I DIDN'T QUITE GET 8267 05:15:54,991 --> 05:15:56,693 THE FULL QUESTION. 8268 05:15:56,693 --> 05:15:58,128 >> THE QUESTION IS. 8269 05:15:58,128 --> 05:15:59,396 >> BUT -- 8270 05:15:59,396 --> 05:16:01,598 >> IS IT THE BEST MODEL SYSTEM 8271 05:16:01,598 --> 05:16:03,800 WHEN IT LIVES IN THE MUSCULARIS 8272 05:16:03,800 --> 05:16:08,705 TO TALK ABOUT NEURO EPITHELIAL 8273 05:16:08,705 --> 05:16:08,939 BIOLOGY? 8274 05:16:08,939 --> 05:16:11,408 BECAUSE PROBABLY NIPO OR 8275 05:16:11,408 --> 05:16:15,045 TRICHINELLA OR TRICURUS ARE MORE 8276 05:16:15,045 --> 05:16:16,813 APPROPRIATE SYSTEMS TO 8277 05:16:16,813 --> 05:16:17,481 INTERROGATE BIOLOGY. 8278 05:16:17,481 --> 05:16:27,724 THAT'S WHAT -- 8279 05:16:38,001 --> 05:16:42,672 WE LOOK AT IMMUNITY, THERE IS 8280 05:16:42,672 --> 05:16:43,673 CHANGE IN THE FUNCTIONS AND HOW 8281 05:16:43,673 --> 05:16:45,642 MUCH OF THAT IS DIRECT OR 8282 05:16:45,642 --> 05:16:47,277 INDIRECT WE DO NOT KNOW. 8283 05:16:47,277 --> 05:16:49,212 BUT WHAT HE SAID IS ARE THERE 8284 05:16:49,212 --> 05:16:50,647 OTHER MODELS WE KNOW COULD 8285 05:16:50,647 --> 05:16:53,283 ACTUALLY ADDRESS THIS BETTER. 8286 05:16:53,283 --> 05:16:55,952 >> YEAH I'M JUST ASKING IF IT 8287 05:16:55,952 --> 05:16:57,754 TRAVERSE, IF THE BIOLOGY YOU'RE 8288 05:16:57,754 --> 05:17:01,091 PROPOSING TRAVERSES OTHER MODEL 8289 05:17:01,091 --> 05:17:01,458 SYSTEMS. 8290 05:17:01,458 --> 05:17:05,128 >> I THINK YOU AND I SHOULD TALK 8291 05:17:05,128 --> 05:17:05,762 OVER A BEER. 8292 05:17:05,762 --> 05:17:07,364 >> TONIGHT, DINNER, SEE YOU 8293 05:17:07,364 --> 05:17:07,864 THEN. 8294 05:17:07,864 --> 05:17:08,565 >> DINNER TION TONIGHT. 8295 05:17:08,565 --> 05:17:11,301 >> YOU OWE ME A BEER. 8296 05:17:11,301 --> 05:17:13,670 >> RACHEL? 8297 05:17:13,670 --> 05:17:17,741 >> SO THANK YOU FOR A GREAT 8298 05:17:17,741 --> 05:17:17,974 TALK. 8299 05:17:17,974 --> 05:17:19,509 THIS MAY BE A LITTLE BET RELATED 8300 05:17:19,509 --> 05:17:21,711 TO WHAT DAVID WAS ASKING. 8301 05:17:21,711 --> 05:17:23,313 THIS MAY BE A LITTLE BIT RELATED 8302 05:17:23,313 --> 05:17:26,049 TO WHAT DAVID WAS ASKING. 8303 05:17:26,049 --> 05:17:30,887 YOU TALK A LOT ABOUT EFFECT OF 8304 05:17:30,887 --> 05:17:31,988 EPITHELIAL CELLS BUT I WAS 8305 05:17:31,988 --> 05:17:34,357 WONDERING DO YOU SEE EFFECTS ON 8306 05:17:34,357 --> 05:17:34,925 PERISTALSIS, FOR EXAMPLE SH. 8307 05:17:34,925 --> 05:17:36,993 >> OH, YEAH. 8308 05:17:36,993 --> 05:17:37,427 YEAH. 8309 05:17:37,427 --> 05:17:42,399 SO I SPENT $40,000 TO GET THE 8310 05:17:42,399 --> 05:17:47,871 MACHINE, AND TO DO EXACTLY THAT. 8311 05:17:47,871 --> 05:17:51,341 AND THE DATA IS NOT QUITE -- 8312 05:17:51,341 --> 05:17:53,210 SOME DAYS IT WORKSZ AND SOME 8313 05:17:53,210 --> 05:17:54,611 DAYS IT LOOKS LIKE IT AFFECTS 8314 05:17:54,611 --> 05:17:56,513 AND SOME DAYS IT DOESN'T. 8315 05:17:56,513 --> 05:17:58,114 SO SWREANT COME TO ANY 8316 05:17:58,114 --> 05:17:59,349 PARTICULAR CONCLUSION THAT BY 8317 05:17:59,349 --> 05:18:02,552 LOSS OF IL IF CU RECEPTORS ARE 8318 05:18:02,552 --> 05:18:04,821 AUTO EFFECT EFFECTING THE NORMAL 8319 05:18:04,821 --> 05:18:06,156 PERISTALSIS MOMENT AND 8320 05:18:06,156 --> 05:18:06,423 EXPULSION. 8321 05:18:06,423 --> 05:18:07,357 THE JURY IS STILL OUT. 8322 05:18:07,357 --> 05:18:09,192 IT MAY BE ASKED TOO BECAUSE WE 8323 05:18:09,192 --> 05:18:13,830 DO NOT KNOW HOW TO DO THESE 8324 05:18:13,830 --> 05:18:14,064 THINGS. 8325 05:18:14,064 --> 05:18:20,637 >> JIM DID HE SANTO -- JIM DE 8326 05:18:20,637 --> 05:18:21,004 SANTO. 8327 05:18:21,004 --> 05:18:21,972 WHAT DO YOU THINK TURNS HAD OFF? 8328 05:18:21,972 --> 05:18:24,708 YOU MENTIONED IT'S AN AMPLIFYING 8329 05:18:24,708 --> 05:18:26,243 LOOP AND WE ALL PROBABLY 8330 05:18:26,243 --> 05:18:27,110 EXPERIENCE ACTIVATION OF THESE 8331 05:18:27,110 --> 05:18:28,678 CELLS AT SOME POINT IN OUR LIVES 8332 05:18:28,678 --> 05:18:30,513 BUT DON'T END UP ALL MAYBE 8333 05:18:30,513 --> 05:18:31,581 HAVING THE KIND OF CONSEQUENCE 8334 05:18:31,581 --> 05:18:32,782 THAT IS YOUR SON HAD. 8335 05:18:32,782 --> 05:18:34,084 WHAT DO YOU THINK IS REGULATING 8336 05:18:34,084 --> 05:18:34,284 THIS? 8337 05:18:34,284 --> 05:18:36,419 >> AND HOW TO INTERRUPT IT. 8338 05:18:36,419 --> 05:18:39,890 IN FACT, I WAS TALKING TO -- AT 8339 05:18:39,890 --> 05:18:42,192 MASS GENERAL AND APPARENTLY THIS 8340 05:18:42,192 --> 05:18:43,960 IRRITATION AND VOMITING 8341 05:18:43,960 --> 05:18:45,695 CONTINUOUS VOMITING ONCE IT 8342 05:18:45,695 --> 05:18:48,999 STARTS IS THERE IN 20% OF THE 8343 05:18:48,999 --> 05:18:51,134 PATIENTS THAT GET THE FOOD 8344 05:18:51,134 --> 05:18:53,169 ALLERGIES, AND ALL THEY DO IS TO 8345 05:18:53,169 --> 05:18:56,139 TAKE THEM IN AND SEDATE THEM. 8346 05:18:56,139 --> 05:18:58,842 SO I DO NOT KNOW WHAT WOULD BE 8347 05:18:58,842 --> 05:19:00,510 NORMAL WAY TO INTERRUPT IT, IS 8348 05:19:00,510 --> 05:19:02,012 THERE A NORMAL BIOLOGICAL 8349 05:19:02,012 --> 05:19:04,214 PROCESS BY WHICH TO TURN IT OFF. 8350 05:19:04,214 --> 05:19:05,115 I DO NOT KNOW. 8351 05:19:05,115 --> 05:19:05,315 OKAY? 8352 05:19:05,315 --> 05:19:07,884 THANK YOU VERY MUCH. 8353 05:19:07,884 --> 05:19:08,151 [APPLAUSE] 8354 05:19:08,151 --> 05:19:10,887 >> I HAVE A QUESTION FOR YOU. 8355 05:19:10,887 --> 05:19:11,154 [APPLAUSE] 8356 05:19:11,154 --> 05:19:14,457 >> SO REMODELING THAT YOU SEE, 8357 05:19:14,457 --> 05:19:16,393 IS THAT WORM DEPENDENT? 8358 05:19:16,393 --> 05:19:18,561 DOES THIS HAPPEN IN THE IL13 8359 05:19:18,561 --> 05:19:19,963 RECEPTOR KNOCK-OUT, FOR EXAMPLE, 8360 05:19:19,963 --> 05:19:20,997 IF YOU DON'T HAVE THE WORMS? 8361 05:19:20,997 --> 05:19:21,865 >> YEAH, IT DOES. 8362 05:19:21,865 --> 05:19:23,934 >> IT DOES, OKAY, GOOD. 8363 05:19:23,934 --> 05:19:25,902 SO THE NEXT SESSION, SESSION WE 8364 05:19:25,902 --> 05:19:28,538 HAVE A COFFEE BREAK UNTIL 8365 05:19:28,538 --> 05:19:32,943 3:15 SO LET'S REASSEMBLE AT 34 8366 05:19:32,943 --> 05:19:34,611 1R5 AND WE'LL FINISH IT OFF. 8367 05:19:34,611 --> 05:19:36,723 >> THANK YOU ACTUAL THE SPEAKERS 8368 05:19:36,723 --> 05:19:40,960 HELLO, I THINK GET STARTED. 8369 05:19:40,960 --> 05:19:47,033 I'M PAM SCHWARTZBERG FROM N I 8370 05:19:47,033 --> 05:19:52,472 AD, OUR FIRST SPEAKER. 8371 05:19:52,472 --> 05:19:54,107 >> I WOULD LIKE TO THANK OF 8372 05:19:54,107 --> 05:19:55,375 COURSE THE ORGANIZER AND I HAVE 8373 05:19:55,375 --> 05:19:56,609 TO IMMEDIATELY CORRECT YOU 8374 05:19:56,609 --> 05:19:58,445 BECAUSE WITHIN A FEW WEEKS I 8375 05:19:58,445 --> 05:20:00,847 WILL BE MOVING FROM EAST COAST 8376 05:20:00,847 --> 05:20:02,182 TO WEST COAST. 8377 05:20:02,182 --> 05:20:04,184 BUT FOR NOW, IT'S STILL NEW 8378 05:20:04,184 --> 05:20:04,384 YORK. 8379 05:20:04,384 --> 05:20:05,785 AND THAT IS WHERE ALL OF THIS 8380 05:20:05,785 --> 05:20:06,720 WORK WAS DONE. 8381 05:20:06,720 --> 05:20:10,790 I DO HAVE A CONFLICT OF 8382 05:20:10,790 --> 05:20:11,791 INTEREST, SOME OF THE WORK I 8383 05:20:11,791 --> 05:20:13,460 WILL BE SHOWING YOU WAS ACTUALLY 8384 05:20:13,460 --> 05:20:16,363 SPONSORED BY A CCH SIRRUS AND 8385 05:20:16,363 --> 05:20:17,797 THEY'VE ALSO HECHED US TO DO A 8386 05:20:17,797 --> 05:20:18,998 TRIAL WHICH I WILL GET TO 8387 05:20:18,998 --> 05:20:19,766 TOWARDS THE END. 8388 05:20:19,766 --> 05:20:25,705 SO ANYWAY TO GET MICROPHONE 8389 05:20:25,705 --> 05:20:27,874 HIGHER? 8390 05:20:27,874 --> 05:20:29,976 BECAUSE I'M SLUMPING OVER HERE. 8391 05:20:29,976 --> 05:20:40,286 I'M NOT THAT TALL. 8392 05:20:40,653 --> 05:20:42,422 CAN YOU HEAR ME? 8393 05:20:42,422 --> 05:20:43,289 OKAY, GOOD. 8394 05:20:43,289 --> 05:20:45,358 SO FOR THIS PURPOSE, SINCE I'M 8395 05:20:45,358 --> 05:20:47,594 GOING TO TELL YOU MOSTLY ABOUT 8396 05:20:47,594 --> 05:20:48,795 GRAFT VERSUS HOST VERY QUICKLY 8397 05:20:48,795 --> 05:20:50,397 FOR THOSE OF YOU WHO DOAN KNOW 8398 05:20:50,397 --> 05:20:57,737 OR DON'T DEAL OFTEN WITH AN 8399 05:20:57,737 --> 05:20:59,506 ALLOGENEIC TRANCE PLNT, DOING 8400 05:20:59,506 --> 05:21:01,608 ABOUT 40 THOWCH TRANSPLANTS DONE 8401 05:21:01,608 --> 05:21:04,110 SPECIFICALLY FOR -- 40,000 8402 05:21:04,110 --> 05:21:05,078 TRANSPLANTS DONE SPECIFICALLY 8403 05:21:05,078 --> 05:21:07,714 FOR PATIENTS WITH BLOOD 8404 05:21:07,714 --> 05:21:09,249 TRANSPLANTS, ONE OF THE FEW 8405 05:21:09,249 --> 05:21:11,351 CURATIVE THERAPIES WE HAVE, 8406 05:21:11,351 --> 05:21:13,753 MEANING WE'RE WILLING TO TAKE 8407 05:21:13,753 --> 05:21:15,355 ENORMOUS RISKS, ONE OF THE MAJOR 8408 05:21:15,355 --> 05:21:25,832 RISKS IS GRAFT VERSUS HOST. 8409 05:21:35,074 --> 05:21:36,509 TESTING ONE TWORKS, THREE, GOOD, 8410 05:21:36,509 --> 05:21:37,444 THANK YOU VERY MUCH. 8411 05:21:37,444 --> 05:21:39,612 SORRY, GUYS. 8412 05:21:39,612 --> 05:21:42,782 SO GRAFT VERSUS HOST IS A 8413 05:21:42,782 --> 05:21:46,719 DISEASE THAT LEADS TO OFTEN 8414 05:21:46,719 --> 05:21:47,754 MAJOR COMPLICATIONS. 8415 05:21:47,754 --> 05:21:50,457 SOME OF THEM CAN ACTUALLY BE 8416 05:21:50,457 --> 05:21:50,690 LETHAL. 8417 05:21:50,690 --> 05:21:51,858 AND ALL YOU NEED TO KNOW FOR 8418 05:21:51,858 --> 05:21:53,526 THIS TALK IS THAT WHAT IS 8419 05:21:53,526 --> 05:21:56,663 CRITICAL THERE IS THAT IT'S ALL 8420 05:21:56,663 --> 05:22:03,603 DEPENDENT ON IT TELLS WITHIN THE 8421 05:22:03,603 --> 05:22:08,441 ALLO, WE'LL SEE VARIOUS ALLO, 8422 05:22:08,441 --> 05:22:09,976 MOSTLY IN GUT, LIVER AND SKIN 8423 05:22:09,976 --> 05:22:12,178 AND IT'S REALLY GULT GRAFT 8424 05:22:12,178 --> 05:22:13,513 VERSUS HOST WHICH LEADS TO MOST 8425 05:22:13,513 --> 05:22:18,384 OF THE PROBLEMS. 8426 05:22:18,384 --> 05:22:22,622 SO TRANSPLANT INTERESTED IN GUT 8427 05:22:22,622 --> 05:22:24,390 FLORA MICROBIOME? 8428 05:22:24,390 --> 05:22:26,626 STUDIES IN THE 1970 INDICATED 8429 05:22:26,626 --> 05:22:28,528 THAT GERM FREE MICE HAD LESS 8430 05:22:28,528 --> 05:22:30,063 GRAFT THAN HOST AND THAT 8431 05:22:30,063 --> 05:22:32,365 ACTUALLY LED FOR DECADES TO TO 8432 05:22:32,365 --> 05:22:33,700 EVERY CENTER DOING THEIR OWN 8433 05:22:33,700 --> 05:22:34,033 THING. 8434 05:22:34,033 --> 05:22:35,468 PATIENTS BEING PUT ON DIFFERENT 8435 05:22:35,468 --> 05:22:37,103 TYPES OF ANTIBIOTICS, SOMETIMES 8436 05:22:37,103 --> 05:22:39,205 INTO A BUBBLE SETTING REALLY AND 8437 05:22:39,205 --> 05:22:41,407 DOING ALL KINDS OF THINGS TO DO 8438 05:22:41,407 --> 05:22:43,443 SOMETHING WITH THE GUT FLORA. 8439 05:22:43,443 --> 05:22:45,078 BUT IT WAS NEVER COMPLETELY 8440 05:22:45,078 --> 05:22:46,513 CLEAR WHAT WAS ACTUALLY 8441 05:22:46,513 --> 05:22:48,615 NECESSARY AND WHAT ACTUALLY 8442 05:22:48,615 --> 05:22:48,948 WORKED. 8443 05:22:48,948 --> 05:22:50,717 SO THAT WAS ONE OF THE REASONS 8444 05:22:50,717 --> 05:22:53,453 WHY IN 2009 OR SO WE TOOK A 8445 05:22:53,453 --> 05:22:56,122 FRESH LOOK AT THAT AND OVER THE 8446 05:22:56,122 --> 05:22:58,558 YEARS WE'VE LEARNED SOMETHING 8447 05:22:58,558 --> 05:22:59,359 ABOUT THIS. 8448 05:22:59,359 --> 05:23:00,994 I'M TRYING TO SUMMARIZE A LOT OF 8449 05:23:00,994 --> 05:23:03,263 WORK HERE NOW FROM SPECIFICALLY 8450 05:23:03,263 --> 05:23:08,334 MY COLLEAGUE ERYK BA AMER AND 8451 05:23:08,334 --> 05:23:10,603 WORK FROM MY GROUP FROM THIS 8452 05:23:10,603 --> 05:23:12,605 GRAM WHICH INDICATES IN BLUE THE 8453 05:23:12,605 --> 05:23:13,973 VARIOUS TEXTS THAT WE HAVE SHOWN 8454 05:23:13,973 --> 05:23:17,510 BOTH IN MOUSE AND MEN MEN TO BE 8455 05:23:17,510 --> 05:23:21,881 OF SOME GA FAVORABLE OUTCOME AND 8456 05:23:21,881 --> 05:23:25,018 THOSE THAT HAVE BEEN IN RED 8457 05:23:25,018 --> 05:23:26,486 LINKED TO WORSE OUTCOME, I'M 8458 05:23:26,486 --> 05:23:28,955 SHOWING YOU HERE HOW THESE ARE 8459 05:23:28,955 --> 05:23:31,424 LINKED TO GRAFT VERSUS HOST. 8460 05:23:31,424 --> 05:23:32,525 BASICALLY ALL OF THE STUDIES WE 8461 05:23:32,525 --> 05:23:33,927 HAVE DONE OVER THE LAST DECADE 8462 05:23:33,927 --> 05:23:36,763 OR SO WE HAVE SHOWN THAT THESE 8463 05:23:36,763 --> 05:23:38,865 VARIOUS TECHS CAN BE IMPLICATED 8464 05:23:38,865 --> 05:23:40,166 IN ALMOST EVERY SERIOUS 8465 05:23:40,166 --> 05:23:44,537 COMPLICATION OR OUTCOME AFTER 8466 05:23:44,537 --> 05:23:46,039 ALLOGENIC TRANSPLANT. 8467 05:23:46,039 --> 05:23:47,507 SOME OF THE LESSONS WE HAVE 8468 05:23:47,507 --> 05:23:49,108 LEARNED IN BODIES RELEVANT FOR 8469 05:23:49,108 --> 05:23:50,777 THE ONE STORY THAT I WOULD LIKE 8470 05:23:50,777 --> 05:23:52,545 TO FOCUS ON IS THAT THE LOSS OF 8471 05:23:52,545 --> 05:23:54,280 THE DIVERSITY WITHIN THE GUT 8472 05:23:54,280 --> 05:23:56,382 FLORA HAPPENS IN ALMOST ALL OF 8473 05:23:56,382 --> 05:23:58,351 THESE PATIENTS AND IF IT'S MORE 8474 05:23:58,351 --> 05:23:59,886 SERIOUS THEN IT'S ACTUALLY ALSO 8475 05:23:59,886 --> 05:24:03,923 LINKED TO LETHAL GRAFT VERSUS 8476 05:24:03,923 --> 05:24:09,529 HOST, COMMENSAL ANAEROBES 8477 05:24:09,529 --> 05:24:11,364 TYPICALLY LOST IN THE FIRST DAY 8478 05:24:11,364 --> 05:24:13,766 OR WEEK AFTER TRANSPLANT AND KEY 8479 05:24:13,766 --> 05:24:18,137 FOCUS BACTERIA WE'VE FOCUSESSED 8480 05:24:18,137 --> 05:24:21,774 ON IS BLAUTIA, ALMOST ALL 8481 05:24:21,774 --> 05:24:23,509 SPRAISHTS A COURSE OF DOMINANCE 8482 05:24:23,509 --> 05:24:25,378 DURING THE COURSE OF THEIR 8483 05:24:25,378 --> 05:24:26,779 TRANSPLANT IN SOME CASES MEANING 8484 05:24:26,779 --> 05:24:30,617 THEIR FLORA IS TAKING OVER BY A 8485 05:24:30,617 --> 05:24:32,719 SINGLE REMEMBER, MORE FREQUENTLY 8486 05:24:32,719 --> 05:24:34,621 THAN TR ANY OTHERS IS 8487 05:24:34,621 --> 05:24:39,392 ENTEROCOCCUS WITHIN THE MOUSE 8488 05:24:39,392 --> 05:24:42,128 ENTEROCOCCUSLY -- AND WITHIN THE 8489 05:24:42,128 --> 05:24:45,031 HUMAN ENTEROCOCCUS SOME OF THE 8490 05:24:45,031 --> 05:24:46,966 THINGS IT DOES AND HOW IT MAKES 8491 05:24:46,966 --> 05:24:48,868 GRAFT VERSUS HOST WORSE. 8492 05:24:48,868 --> 05:24:50,303 LESS ANOTHER STRIKING FEATURE IS 8493 05:24:50,303 --> 05:24:52,205 OF COURSE ALL OF THESE PATIENT 8494 05:24:52,205 --> 05:24:54,207 GOING THROUGH ANNAL LOWGENIC 8495 05:24:54,207 --> 05:24:54,974 TRANSPLANT, GOING THROUGH 8496 05:24:54,974 --> 05:24:56,809 PERIODS OF NEUTROPENA WILL BE 8497 05:24:56,809 --> 05:24:58,578 TREATED AT ONE POINT WITH 8498 05:24:58,578 --> 05:25:00,647 ANTIBIOTICS, SPECIFICALLY THE 8499 05:25:00,647 --> 05:25:02,682 EXPOSURE TO BROAD-SPECTRUM 8500 05:25:02,682 --> 05:25:03,416 ANTIBIOTICS HAS BEEN LINKED TO 8501 05:25:03,416 --> 05:25:06,386 AN INCREASED RISK FOR GRAFT 8502 05:25:06,386 --> 05:25:08,421 VERSUS HOST. 8503 05:25:08,421 --> 05:25:10,523 BUT THERE ARE MANY FACTORS THAT 8504 05:25:10,523 --> 05:25:12,258 CAN HAVE IMPACT OF COURSE ON 8505 05:25:12,258 --> 05:25:14,260 CHANGES WITHIN THE GUT FLORA, 8506 05:25:14,260 --> 05:25:16,129 BUT WE'RE LOOKING AT CURRENTLY 8507 05:25:16,129 --> 05:25:19,365 AND I'M ONLY SHOWING ONE VIN NET 8508 05:25:19,365 --> 05:25:23,970 HERE IS DIET -- VIGNETTE HERE IS 8509 05:25:23,970 --> 05:25:26,773 DIET VERSUS MOUSE AND MAN, 8510 05:25:26,773 --> 05:25:29,642 PATIENT CAN TAKE PHOTOS OF THEIR 8511 05:25:29,642 --> 05:25:30,610 MEALS AND USE THAT FOR 8512 05:25:30,610 --> 05:25:32,612 DOCUMENTATION OF THEIR DIET, AN 8513 05:25:32,612 --> 05:25:34,514 APP, WHICH IS WORKING VERY, VERY 8514 05:25:34,514 --> 05:25:34,747 NICELY. 8515 05:25:34,747 --> 05:25:36,883 WHEEF LEARNED SO FAR IS THAT 8516 05:25:36,883 --> 05:25:38,618 CALORIE INTAKE AND FIBER INTAKE 8517 05:25:38,618 --> 05:25:41,387 IS POSITIVELY CORRELATED WITH A 8518 05:25:41,387 --> 05:25:45,525 DIVERSITY AND WITH BLAUTIA SO 8519 05:25:45,525 --> 05:25:48,695 THAT ONE KEYSTONE AND NEGLECT 8520 05:25:48,695 --> 05:25:50,897 TWIFL ENTEROCOCCUS WHICH IS THE 8521 05:25:50,897 --> 05:25:54,167 ULTIMATE BAD BUG IN ALL OF OUR 8522 05:25:54,167 --> 05:25:57,336 STUDIES. 8523 05:25:57,336 --> 05:25:57,770 HAD. 8524 05:25:57,770 --> 05:25:59,439 STUDY WE RECENTLY PUBLISHED HAS 8525 05:25:59,439 --> 05:26:00,840 BEEN LOOKING ACTUAL AT ALL OF 8526 05:26:00,840 --> 05:26:03,910 THE OTHER DRUGS THAN THESE 8527 05:26:03,910 --> 05:26:05,011 TYPICAL ANTIBIOTICS THAT HAVE 8528 05:26:05,011 --> 05:26:07,080 BEEN IMPLICATED BY MANY AND MANY 8529 05:26:07,080 --> 05:26:09,482 DIFFERENT SETTINGS TO LEAD TO 8530 05:26:09,482 --> 05:26:10,783 CHANGES IN THE GUT FLORA. 8531 05:26:10,783 --> 05:26:13,319 SO WHAT WE DID IS WE TOOK ALL OF 8532 05:26:13,319 --> 05:26:14,854 THE PATIENTS THAT WE HAD AT THAT 8533 05:26:14,854 --> 05:26:18,191 POINT AND ALL OF THE SAMPLES, SO 8534 05:26:18,191 --> 05:26:20,526 THERE WAS MORE THAN 1,000 8535 05:26:20,526 --> 05:26:23,196 PATIENTS AND ALMOST 9,000 8536 05:26:23,196 --> 05:26:23,496 SAMPLES. 8537 05:26:23,496 --> 05:26:27,867 WE PUT THEM TOGETHER INTO A U 8538 05:26:27,867 --> 05:26:30,603 MAP CLUSTERED THEM IN TEN 8539 05:26:30,603 --> 05:26:31,804 DIFFERENT CLUSTERS AND SINCE WE 8540 05:26:31,804 --> 05:26:34,207 HAD IN MANY CASES SEQUENTIAL 8541 05:26:34,207 --> 05:26:35,441 SAMPLES AND VERY DETAILED DATA 8542 05:26:35,441 --> 05:26:36,609 OF ALL OF THE DRUGS THAT THEY 8543 05:26:36,609 --> 05:26:38,377 WERE ON, ALL OF THESE PATIENTS 8544 05:26:38,377 --> 05:26:40,146 AT ANY MOMENT ARE ON DIFFERENT 8545 05:26:40,146 --> 05:26:42,749 TYPES OF DRUGS FOR PAIN, FOR 8546 05:26:42,749 --> 05:26:46,452 NAUSEA, FOR WHATEVER, AND SO 8547 05:26:46,452 --> 05:26:48,087 LOOK P AT THESE CLUSTERS AND 8548 05:26:48,087 --> 05:26:50,189 THEN LOOK NG A SEQUENTIAL WAY AT 8549 05:26:50,189 --> 05:26:51,824 WHAT DRUGS MIGHT DO TO HAVE 8550 05:26:51,824 --> 05:26:53,359 PATIENTS MOVE FROM ONE CLUSTER 8551 05:26:53,359 --> 05:26:55,528 TO THE OTHER OR STAY WHERE THEY 8552 05:26:55,528 --> 05:26:57,063 ARE, GAVE US SOME INDICATORS 8553 05:26:57,063 --> 05:26:58,598 ABOUT WHAT ALL OF THESE OTHER 8554 05:26:58,598 --> 05:27:02,034 DRUGS CAN ACTUALLY DO IN TERMS 8555 05:27:02,034 --> 05:27:04,003 OF CHANGES IN THE LEVELS OF THE 8556 05:27:04,003 --> 05:27:11,477 FREQUENCY OF THESE BACTERIA. 8557 05:27:11,477 --> 05:27:13,079 AGAIN, FOR THESE TWO ULTIMATE 8558 05:27:13,079 --> 05:27:17,016 GOOD AND BAD BUGS, BLAUITY A AND 8559 05:27:17,016 --> 05:27:18,885 ENTEROCOCCUS THAT ALL KINDS OF 8560 05:27:18,885 --> 05:27:21,420 DRUGS LIKE NAUSEA MEDICATIONS, 8561 05:27:21,420 --> 05:27:23,890 PAIN MEDICATIONS CAN HAVE IMPACT 8562 05:27:23,890 --> 05:27:25,224 ALSO, SO THIS OACHES THE DOOR IT 8563 05:27:25,224 --> 05:27:26,726 MAYBE START IN A DIFFERENT WAY 8564 05:27:26,726 --> 05:27:28,594 ABOUT THE CHOICES FOR NAUSEA 8565 05:27:28,594 --> 05:27:30,830 MEDICATIONS, PAIN MEDICATIONS, 8566 05:27:30,830 --> 05:27:32,899 CHOICES THAT WE MAKE DAILY FOR 8567 05:27:32,899 --> 05:27:34,867 ALL OF OUR PATIENTS. 8568 05:27:34,867 --> 05:27:36,068 ONE THING THAT I WANT TO 8569 05:27:36,068 --> 05:27:37,170 HIGHLIGHT ALSO SPECIFICALLY 8570 05:27:37,170 --> 05:27:39,806 DRUGS THAT CAN MAYBE LEAD TO 8571 05:27:39,806 --> 05:27:43,009 LOWER LEVELS OF ENTEROCOCCUS, 8572 05:27:43,009 --> 05:27:45,845 ONE DRUG I WANT TO HIGHLIGHT IS 8573 05:27:45,845 --> 05:27:47,480 OTCA BECAUSE THAT IS SORT OF A 8574 05:27:47,480 --> 05:27:48,381 BRIDGE TO THE NEXT STORY THAT I 8575 05:27:48,381 --> 05:27:49,348 WOULD LIKE TO TELL YOU. 8576 05:27:49,348 --> 05:27:51,517 THIS IS A SECONDARY BOWEL ACID 8577 05:27:51,517 --> 05:27:54,587 WHICH IS COMMONLY USED IN MANY 8578 05:27:54,587 --> 05:27:55,822 TRANSPLANT CENTERS AT THE MOMENT 8579 05:27:55,822 --> 05:27:57,123 FOR THREE MONTHS OR SO DURING 8580 05:27:57,123 --> 05:28:00,393 THE WHOLE COURSE OF A 8581 05:28:00,393 --> 05:28:01,060 TRANSPLANT. 8582 05:28:01,060 --> 05:28:02,161 AND PERFORMANCE THOUGHT ALWAYS 8583 05:28:02,161 --> 05:28:03,830 TO BE BENEFICIAL FOR LIVER 8584 05:28:03,830 --> 05:28:07,533 PROBLEMS, VOD, SOS SYNDROME AND 8585 05:28:07,533 --> 05:28:09,168 WE WON'T GO INTO ALL OF THE 8586 05:28:09,168 --> 05:28:11,304 DETAILS NOW ABOUT THAT. 8587 05:28:11,304 --> 05:28:13,005 SO I TOLD YOU ABOUT THIS 8588 05:28:13,005 --> 05:28:14,106 SECONDARY BOWEL ACID BECAUSE 8589 05:28:14,106 --> 05:28:16,075 THAT'S THE ONE STORY THAT IS 8590 05:28:16,075 --> 05:28:20,012 GOING TO COME OUT SOON, AND I 8591 05:28:20,012 --> 05:28:21,981 APOLOGIZE TO ALAN AND TO ELLA 8592 05:28:21,981 --> 05:28:23,382 BECAUSE THEY'VE ACTUALLY HEARD 8593 05:28:23,382 --> 05:28:26,786 PART OF THAT STORY ALREADY 8594 05:28:26,786 --> 05:28:29,222 ALREADY ON BEAUTIFUL ISLANDS IN 8595 05:28:29,222 --> 05:28:30,189 THE GREAT BARRIER REEF, RIGHT? 8596 05:28:30,189 --> 05:28:32,491 BUT NOW YOU'RE GOING TO HEAR THE 8597 05:28:32,491 --> 05:28:33,359 FULLER STORY BECAUSE WE ANSWER 8598 05:28:33,359 --> 05:28:35,328 TO ALL OF THE REVIEWERS' 8599 05:28:35,328 --> 05:28:35,895 QUESTIONS. 8600 05:28:35,895 --> 05:28:38,397 SO WHAT WE WERE LOOKING AT WAS 8601 05:28:38,397 --> 05:28:40,399 THE IMPACT OF CHANGES WITHIN THE 8602 05:28:40,399 --> 05:28:42,535 BOWEL ACIDS ON OUTCOMES BOTH IN 8603 05:28:42,535 --> 05:28:45,538 MOUSE AND MEN AFTER ALLOGENIC 8604 05:28:45,538 --> 05:28:46,839 TRANSPLANT. SO FOR THOSE OF YOU 8605 05:28:46,839 --> 05:28:48,374 WHO DON'T OFTEN THINK ABOUT 8606 05:28:48,374 --> 05:28:50,109 BOWEL ACIDS BUT MAYBE YOU STILL 8607 05:28:50,109 --> 05:28:52,111 KNOW IT FROM HIGH SCHOOL OR 8608 05:28:52,111 --> 05:28:54,447 SOMETHING, BOWEL ACIDS ARE OF 8609 05:28:54,447 --> 05:28:56,382 COURSE MADE WITHIN THE LIVER, 8610 05:28:56,382 --> 05:28:59,218 PRIMARY BOWEL ACIDS SPECIFICALLY 8611 05:28:59,218 --> 05:29:02,855 ARE CONJUGATED TO GLYCINE AND TO 8612 05:29:02,855 --> 05:29:05,358 TOUR IN-- TAURINE, GHET INTO THE 8613 05:29:05,358 --> 05:29:09,528 GUT THROUGH THE BLADDER AND OF 8614 05:29:09,528 --> 05:29:11,931 COURSE THEIR PRIMARY FUNCTION IS 8615 05:29:11,931 --> 05:29:13,900 FOR LIPID UPTAKE AND A FEW OTHER 8616 05:29:13,900 --> 05:29:14,133 THINGS. 8617 05:29:14,133 --> 05:29:15,501 BUT ONCE THEY GET FURTHER INTO 8618 05:29:15,501 --> 05:29:18,804 IT AND GET INTO THE TERMINAL 8619 05:29:18,804 --> 05:29:20,206 ILEUM OR WITHIN THE LARGE BOWEL, 8620 05:29:20,206 --> 05:29:21,974 THAT IS WHERE THEY COME INTO 8621 05:29:21,974 --> 05:29:25,244 CONTACT WITH ALL THE VARIOUS 8622 05:29:25,244 --> 05:29:26,679 BACTERIA AND BACTERIA ACTUALLY 8623 05:29:26,679 --> 05:29:30,283 CARRY A PLETHORA OF ENZYMES TO 8624 05:29:30,283 --> 05:29:32,785 TURN PRIMARY BOWEL ACIDS INTO 8625 05:29:32,785 --> 05:29:34,587 SECONDARY BOWEL ACIDS. 8626 05:29:34,587 --> 05:29:36,455 WE ARE VERY FORTUNATE THAT ONE 8627 05:29:36,455 --> 05:29:38,057 OF THE PEOPLE WHO WORKED WITH 8628 05:29:38,057 --> 05:29:39,025 JUSTIN CROSS TOOK A SPECIAL 8629 05:29:39,025 --> 05:29:41,327 INTEREST AND MADE ALL OF THESE 8630 05:29:41,327 --> 05:29:42,662 ASSAYS TO LOOK AT ALL THESE 8631 05:29:42,662 --> 05:29:44,830 DIFFERENT PRIMARY AND ALSO 8632 05:29:44,830 --> 05:29:45,932 SECONDARY BOWEL ACIDS THAT WE 8633 05:29:45,932 --> 05:29:47,566 COULD TAKE A MORE DETAILED LOOK 8634 05:29:47,566 --> 05:29:49,135 AT WHAT WAS HAPPENING BOTH IN 8635 05:29:49,135 --> 05:29:50,536 MOUSE AND MEN. 8636 05:29:50,536 --> 05:29:51,404 THE OTHER THING THAT YOU NEED TO 8637 05:29:51,404 --> 05:29:54,340 KNOW IS THAT THESE SECONDARY 8638 05:29:54,340 --> 05:29:56,008 BOWEL ACIDS SPECIFICALLY BUT 8639 05:29:56,008 --> 05:29:57,777 ALSO SOME OF THE PRIMARY BOWEL 8640 05:29:57,777 --> 05:30:01,480 ACIDS CAN BIND TO AGAIN A WHOLE 8641 05:30:01,480 --> 05:30:03,549 SERIES OF RECEPTORS AND THE ONE 8642 05:30:03,549 --> 05:30:09,455 THAT I'M GOING TO FOCUS ON IS 8643 05:30:09,455 --> 05:30:11,090 FXR, A FINAL THING ON HIGHLIGHT 8644 05:30:11,090 --> 05:30:13,059 ON THIS SLIDE IS THAT ONE OF THE 8645 05:30:13,059 --> 05:30:14,927 FIRST THINGS THAT ALL OF THESE 8646 05:30:14,927 --> 05:30:18,731 BACTERIA DO IS USE AN ENZYME 8647 05:30:18,731 --> 05:30:22,601 BOWEL SALT HYDROLASE TO DEACON 8648 05:30:22,601 --> 05:30:29,909 JEW GAIRKTS TAKE OFF THE TAURINE 8649 05:30:29,909 --> 05:30:32,878 FROM BOWEL ACIDS. 8650 05:30:32,878 --> 05:30:34,547 SECONDARY ARE ACIDS HAVE BEEN 8651 05:30:34,547 --> 05:30:36,182 STUDIED BY MANY PEOPLE IN THIS 8652 05:30:36,182 --> 05:30:38,351 AWD EXPWRENS AUTHORS IN -- AND 8653 05:30:38,351 --> 05:30:39,652 AWRD YENS AND AUTHOR INSIDE SOME 8654 05:30:39,652 --> 05:30:41,420 OF THESE PAPERS, JUST 8655 05:30:41,420 --> 05:30:42,555 SUMMARIZING, SOME OF THESE 8656 05:30:42,555 --> 05:30:44,090 PAPERS INDICATING THAT SECONDARY 8657 05:30:44,090 --> 05:30:46,559 BOWEL ACIDS CAN BE RELEVANT FOR 8658 05:30:46,559 --> 05:30:48,861 TH17 AND FOR TREX SPECIFICALLY 8659 05:30:48,861 --> 05:30:51,597 LOWER LEVELS OF TH17 AND 8660 05:30:51,597 --> 05:30:54,467 INCREASED LEVELS OF TREGS AND 8661 05:30:54,467 --> 05:30:55,434 FOR REASONS THAT I DON'T KNOW 8662 05:30:55,434 --> 05:30:56,969 ALL OF THESE PAPERS ARE 8663 05:30:56,969 --> 05:30:58,404 PUBLISHED IN NATURE, I GUESS 8664 05:30:58,404 --> 05:31:02,041 NATURE LIKES BOWEL ACIDS. 8665 05:31:02,041 --> 05:31:03,342 WHAT WAS KNOWN WITHIN THE 8666 05:31:03,342 --> 05:31:10,549 CONTEXT OF THE GRAFT VERSUS HOST 8667 05:31:10,549 --> 05:31:15,521 NRKSZ IF YOU GIVE THIS STEKD RI 8668 05:31:15,521 --> 05:31:17,923 BOWEL ACID THAT THAT COULD LEAD 8669 05:31:17,923 --> 05:31:21,060 TO LOWER LEVELS OF GRAFT VERSUS 8670 05:31:21,060 --> 05:31:23,329 HOST, OLDER STUDIES TWO YEARS 8671 05:31:23,329 --> 05:31:24,997 PLUS OLD FROM SWEDEN ACTUALLY 8672 05:31:24,997 --> 05:31:27,433 THAT INDICATED THAT IF YOU GIVE 8673 05:31:27,433 --> 05:31:29,035 UDCA TO PATIENTS THAT THAT COULD 8674 05:31:29,035 --> 05:31:30,936 LEAD TO OVERALL BETTER OUTCOMES 8675 05:31:30,936 --> 05:31:32,371 AND LOWER LEVELS OF GRAFT VERSUS 8676 05:31:32,371 --> 05:31:33,672 HOST, BUT THERE NEVER HAD BEEN 8677 05:31:33,672 --> 05:31:35,508 ANY FURTHER STUDIES OR ANY 8678 05:31:35,508 --> 05:31:37,943 FURTHER FOWP ON THOSE EARLY 8679 05:31:37,943 --> 05:31:39,779 CLINICAL DATASETS. 8680 05:31:39,779 --> 05:31:42,081 SO WE TOOK A FRESH LOOK AND WE 8681 05:31:42,081 --> 05:31:44,050 STARTED WITH MOUSE MODEL SO WE 8682 05:31:44,050 --> 05:31:45,851 OFTEN USED MOUSE MODELS FOR 8683 05:31:45,851 --> 05:31:46,886 GRAFT VERSUS HOST. 8684 05:31:46,886 --> 05:31:48,220 FIRST THANK WE NOTICED WAS THAT 8685 05:31:48,220 --> 05:31:50,790 IF WE LOOKED ABOUT A WEEK OR SO 8686 05:31:50,790 --> 05:31:53,492 OUT FROM TRANSPLANT, THAT IN 8687 05:31:53,492 --> 05:31:54,560 THOSE MICE DEVELOPING GRAFT 8688 05:31:54,560 --> 05:31:57,630 VERSUS HOST THE LEVELS TOTAL 8689 05:31:57,630 --> 05:31:58,831 BOWEL ACIDS POOL WITHIN THE GUT 8690 05:31:58,831 --> 05:32:01,500 AND ALSO WITHIN THE SERUM WITHIN 8691 05:32:01,500 --> 05:32:05,738 THE PLASMA ACTUALLY WAS LOWER. 8692 05:32:05,738 --> 05:32:06,872 SO THINKING THAT MIGHT BE 8693 05:32:06,872 --> 05:32:08,274 BECAUSE THERE'S LIVER DAMAGE, 8694 05:32:08,274 --> 05:32:09,809 RIGHT, MENTIONED THAT THERE'S 8695 05:32:09,809 --> 05:32:11,677 LIVER GRAFT VERSUS HOST, SO 8696 05:32:11,677 --> 05:32:12,845 MAYBE THAT'S WHY. 8697 05:32:12,845 --> 05:32:15,347 SO WE DID SOME SEQUENCING OF THE 8698 05:32:15,347 --> 05:32:17,983 LIVER OF THESE MICE WITH A GRAFT 8699 05:32:17,983 --> 05:32:19,185 VERSUS HOST AND INDEED NOTICED 8700 05:32:19,185 --> 05:32:21,454 THAT ALL OF THE PATHWAYS, ALL OF 8701 05:32:21,454 --> 05:32:22,955 THE ENZYMES THAT WERE RELEVANT 8702 05:32:22,955 --> 05:32:25,091 FOR THE SYNTHESIS OF BOWEL ACIDS 8703 05:32:25,091 --> 05:32:26,792 WERE PRESENT AT LOWER LEVELS AND 8704 05:32:26,792 --> 05:32:28,394 OF COURSE YOU SEE EVIDENCE OF 8705 05:32:28,394 --> 05:32:29,261 THE INFLAMMATION. 8706 05:32:29,261 --> 05:32:31,964 BUT WHEN WE LOOKED INTO MORE 8707 05:32:31,964 --> 05:32:33,866 DETAIL AT THE DROP IN THE BOWEL 8708 05:32:33,866 --> 05:32:36,469 ACID WE SAW -- POOL WE SAW THAT 8709 05:32:36,469 --> 05:32:37,903 MOST OF THE LOSS WAS ACTUALLY 8710 05:32:37,903 --> 05:32:40,840 WITHIN THE SECONDARY BOWEL ACIDS 8711 05:32:40,840 --> 05:32:46,879 AND DEACON JEW GATED BOWEL -- 8712 05:32:46,879 --> 05:32:48,514 DECONJUGATEB BOWEL ACIDS, COULD 8713 05:32:48,514 --> 05:32:48,948 BE RELEVANT. 8714 05:32:48,948 --> 05:32:52,218 WHEN WE LOOKED AT MORE DETAIL AT 8715 05:32:52,218 --> 05:32:53,786 STIEPS OF DOWEL ACIDS AND SAW 8716 05:32:53,786 --> 05:32:55,621 ALL THE ONES IMPLICATED TO 8717 05:32:55,621 --> 05:32:58,791 REGULAR LATE TH17 AND TREG ALL 8718 05:32:58,791 --> 05:33:00,092 OF THOSE WERE PRESENT IN MUCH 8719 05:33:00,092 --> 05:33:02,328 LOWER LEVELS IN THOSE MICE 8720 05:33:02,328 --> 05:33:06,565 DEVELOPING GRAFT VERSUS HOST. 8721 05:33:06,565 --> 05:33:10,302 WE THEN DID SOME SHUT BEGUN 8722 05:33:10,302 --> 05:33:11,837 SEQUENCES -- SHUT GUN SEQUENCING 8723 05:33:11,837 --> 05:33:13,139 SO WE COULD BETTER ANALYZE GENES 8724 05:33:13,139 --> 05:33:14,473 OF INTEREST AND ONLY THING TO 8725 05:33:14,473 --> 05:33:15,774 HIGHLIGHT HERE IS THE LOWER 8726 05:33:15,774 --> 05:33:19,245 LEVEL OF THIS DISIEM THAT I 8727 05:33:19,245 --> 05:33:20,646 MENTIONED EARLIER WAS RELEVANT 8728 05:33:20,646 --> 05:33:25,918 IN THE FIRST STEP BY TBOORKT 8729 05:33:25,918 --> 05:33:27,553 DECONJUGATE PRIMARY BOWEL ACIDS 8730 05:33:27,553 --> 05:33:30,089 AND SPECIFICALLY BMH IN GRAFT 8731 05:33:30,089 --> 05:33:32,925 VERSUS HOST WAS PRESENT AT A 8732 05:33:32,925 --> 05:33:34,560 SIGNIFICANTLY LOWER LEVEL -- 8733 05:33:34,560 --> 05:33:34,727 BSH. 8734 05:33:34,727 --> 05:33:36,529 I TOLD YOU ALREADY THAT WE HAD A 8735 05:33:36,529 --> 05:33:38,497 SPECIAL INTEREST IN FXR, ONE OF 8736 05:33:38,497 --> 05:33:40,699 THE MOST RELEVANT PATHWAYS HERE, 8737 05:33:40,699 --> 05:33:43,002 SPECIFICALLY FOR BOWEL ACID 8738 05:33:43,002 --> 05:33:43,302 SYNTHESIS. 8739 05:33:43,302 --> 05:33:47,139 BUT FXR CAN BE FOUND ON MANY 8740 05:33:47,139 --> 05:33:49,008 OTHER CELL TYPES AND HAVE BEEN 8741 05:33:49,008 --> 05:33:50,409 IMPLICATED IN A NUMBER OF OTHER 8742 05:33:50,409 --> 05:33:51,544 PROCESSES AND OF COURSE OUR 8743 05:33:51,544 --> 05:33:54,146 INTEREST WAS SPECIFICALLY IF IT 8744 05:33:54,146 --> 05:33:56,348 COULD REGULATE IMMUNE CELLS. 8745 05:33:56,348 --> 05:33:58,317 WHEN WE STARTED OUT WITH WAS 8746 05:33:58,317 --> 05:33:59,552 DEVELOPING AN ASSAY SO THAT WE 8747 05:33:59,552 --> 05:34:02,888 COULD MEASURE THE ACTIVITY ON 8748 05:34:02,888 --> 05:34:06,158 FXR OR THROUGH FXR OF A PANEL OF 8749 05:34:06,158 --> 05:34:08,494 BOWEL ACIDS, BOTH PRIMARY AND 8750 05:34:08,494 --> 05:34:09,795 SECOND BY BOWEL ACIDS AND WHAT 8751 05:34:09,795 --> 05:34:11,564 WAS VERY STRIKELY IN THIS ASSAY 8752 05:34:11,564 --> 05:34:14,533 WAS THAT ONLY ONE PRIMARY BOWEL 8753 05:34:14,533 --> 05:34:16,735 ACIDS CDCA WAS ABLE TO FUNCTION 8754 05:34:16,735 --> 05:34:18,938 AS AN AGONIST, ALL OF THE OTHERS 8755 05:34:18,938 --> 05:34:20,906 WERE VERY WEAK OR PARTIAL 8756 05:34:20,906 --> 05:34:23,008 AGONIST AND ACTUALLY IF YOU MIX 8757 05:34:23,008 --> 05:34:25,911 IT, THEY COULD FUNCTION SOMEWHAT 8758 05:34:25,911 --> 05:34:29,014 AS AN ANTAGONIST ALSO. 8759 05:34:29,014 --> 05:34:31,050 WE THEN ZOOMED IN SPECIFICALLY 8760 05:34:31,050 --> 05:34:32,585 ON T CELLS FOR TWO REASONS. 8761 05:34:32,585 --> 05:34:34,954 FIRST OF ALL, BECAUSE I TOLD YOU 8762 05:34:34,954 --> 05:34:36,422 ALREADY THAT T CELLS WITHIN THE 8763 05:34:36,422 --> 05:34:38,290 CON ESKS IT A GRAFT VERSUS HOST 8764 05:34:38,290 --> 05:34:40,426 ARE THE MOST CRITICAL CELLS THAT 8765 05:34:40,426 --> 05:34:42,761 WILL GET YOU TO A GRAFT VERSUS 8766 05:34:42,761 --> 05:34:44,630 HOST, AND THE SECOND THING WAS 8767 05:34:44,630 --> 05:34:45,764 THAT ONE OF THE FOLKS WHO WERE 8768 05:34:45,764 --> 05:34:50,269 WORKING WITH US ON THIS CLARISSA 8769 05:34:50,269 --> 05:34:52,538 CAMPBELL WHO NOW HAS HER OWN LAB 8770 05:34:52,538 --> 05:34:55,307 IN AUSTRIA, THAT SHE HAD ALREADY 8771 05:34:55,307 --> 05:34:56,742 DEMONSTRATED THAT IN ACTIVATED T 8772 05:34:56,742 --> 05:35:00,579 CELLS YOU CAN SEE IN TIME AN 8773 05:35:00,579 --> 05:35:01,347 UP-REGULATION OF THE EXPRESSION 8774 05:35:01,347 --> 05:35:07,987 OF FXR. 8775 05:35:07,987 --> 05:35:13,792 SO AFTER THEY HAD BEEN 8776 05:35:13,792 --> 05:35:15,327 ACTIVATED, I'M ONLY SHOWING YOU 8777 05:35:15,327 --> 05:35:17,963 HERE THE CD4 DATA AND SHOWING 8778 05:35:17,963 --> 05:35:22,334 INDEED THAT CDCA, PRIME PRIMARY 8779 05:35:22,334 --> 05:35:23,769 BOWEL ACID THAT COULD FUNCTION 8780 05:35:23,769 --> 05:35:26,171 AS AN AGONIST COULD READ TO 8781 05:35:26,171 --> 05:35:27,606 UPREGULATION OF INFLAMMATORY 8782 05:35:27,606 --> 05:35:30,009 GENES WHERE UDCA SEKSD RI BOWEL 8783 05:35:30,009 --> 05:35:31,744 ACIDS THAT CAN FUNCTION AS A 8784 05:35:31,744 --> 05:35:34,613 VERY WEAK AGONIST OR MAYBE EVEN 8785 05:35:34,613 --> 05:35:36,482 AN ANTAGONIST DOESN'T DO THAT. 8786 05:35:36,482 --> 05:35:38,984 AND THESE ARE THE CONTROLS BOTH 8787 05:35:38,984 --> 05:35:41,720 DRUG THAT CAN FUNCTION AS AN FXR 8788 05:35:41,720 --> 05:35:43,455 AGONIST OR AN ANTAGONIST. 8789 05:35:43,455 --> 05:35:46,091 SO ACTUALLY PRETTY STRONG 8790 05:35:46,091 --> 05:35:49,962 AGNOSTIC FUNCTION FOR CDCA. 8791 05:35:49,962 --> 05:35:51,897 WE THEN TOOK THAT TO SOME 8792 05:35:51,897 --> 05:35:56,368 FUNCTIONAL ASSAYS AND SHOWED IN 8793 05:35:56,368 --> 05:35:58,604 AN ASSAY THAT IF YOU LOOK AT THE 8794 05:35:58,604 --> 05:36:00,773 BASELINE PROLIFERATION OF 8795 05:36:00,773 --> 05:36:03,609 ACTIVATED T CELLS IF YOU ADD 8796 05:36:03,609 --> 05:36:05,277 AGAIN THAT PRIMARY BOWEL ACID 8797 05:36:05,277 --> 05:36:07,346 THAT CAN FUNCTION AS AN AGONIST 8798 05:36:07,346 --> 05:36:10,516 TO CDCA THAT YOU CAN BETTER 8799 05:36:10,516 --> 05:36:12,384 PROLIFERATION WHEREAS UDCA LEADS 8800 05:36:12,384 --> 05:36:14,253 TO WORSE PROLIFERATION. 8801 05:36:14,253 --> 05:36:15,988 FINALLY LOOKING AT ACTIVATION 8802 05:36:15,988 --> 05:36:18,057 MEASURED HERE BY CD25 8803 05:36:18,057 --> 05:36:19,925 EXPRESSION, WE SAW AGAIN THE 8804 05:36:19,925 --> 05:36:22,995 SAME THING, HERE ARE FXR 8805 05:36:22,995 --> 05:36:23,862 WILD-TYPE T CELLS SHOWING THAT 8806 05:36:23,862 --> 05:36:26,732 THE PRIMARY BOWEL ACID CDCA CAN 8807 05:36:26,732 --> 05:36:29,468 LEAD TO HIGHER LEVELS IN UDCA TO 8808 05:36:29,468 --> 05:36:31,303 LOWER LEVELS, WE ALSO DID THIS 8809 05:36:31,303 --> 05:36:33,505 WITH AN FXR KNOCK-OUT EXPHOWS 8810 05:36:33,505 --> 05:36:36,442 THERE WE SAW NO ACTIVITY AT ALL 8811 05:36:36,442 --> 05:36:40,612 OF CDCA AND YOU DCA. 8812 05:36:40,612 --> 05:36:42,881 SO THEN WE WENT BACK TO MOUSE 8813 05:36:42,881 --> 05:36:44,216 MODELS FOR GRAFT VERSUS HOST AND 8814 05:36:44,216 --> 05:36:45,984 STARTED BY SIMPLY GIVING THESE 8815 05:36:45,984 --> 05:36:49,154 MICE AN AGONIST AND SO 8816 05:36:49,154 --> 05:36:51,757 OVERALL -- AND SAW OVERALL WORST 8817 05:36:51,757 --> 05:36:53,192 OUTCOMES, WORST GRAFT VERSUS 8818 05:36:53,192 --> 05:36:55,694 HOST, THAT DOESN'T TELL YOU YET 8819 05:36:55,694 --> 05:36:57,363 THROUGH WHICH PATHWAY, WHICH 8820 05:36:57,363 --> 05:36:58,864 CELL TYPE. 8821 05:36:58,864 --> 05:37:00,466 WE GENERATED CONDITIONAL 8822 05:37:00,466 --> 05:37:02,234 KNOCK-OUT MOUSE AND IN THESE 8823 05:37:02,234 --> 05:37:03,435 MODELS YOU CAN SPECIFICALLY TAKE 8824 05:37:03,435 --> 05:37:06,839 ONLY T CELLS THAT ARE DEFICIENT 8825 05:37:06,839 --> 05:37:07,906 FOR FXR. 8826 05:37:07,906 --> 05:37:10,209 AND IN TWO DIFFERENT MODELS WE 8827 05:37:10,209 --> 05:37:12,311 SAW LESS GRAFT VERSUS HOST IF WE 8828 05:37:12,311 --> 05:37:17,116 USE FXR DEFICIENT DONOR T CELLS. 8829 05:37:17,116 --> 05:37:18,917 WE ANALYZED THAT ALSO UPSIDE 8830 05:37:18,917 --> 05:37:20,719 DOWN AS WE ALWAYS DO OF ALL OF 8831 05:37:20,719 --> 05:37:23,655 THE TISSUES, THE CYTOKINES, THE 8832 05:37:23,655 --> 05:37:25,090 INFILTRATION OF T CELLS AND SO 8833 05:37:25,090 --> 05:37:27,426 ON AND SO ON, WILL SPARE YOU 8834 05:37:27,426 --> 05:37:29,061 THAT AND JUST SAY THAT OVERALL 8835 05:37:29,061 --> 05:37:31,196 WE SAW LESS GRAFT VERSUS HOST 8836 05:37:31,196 --> 05:37:33,065 SPECIFICALLY WE SAW LESS GRAFT 8837 05:37:33,065 --> 05:37:37,336 VERSUS HOST WITHIN THE LARGE 8838 05:37:37,336 --> 05:37:37,569 BOWEL. 8839 05:37:37,569 --> 05:37:41,407 SO THEN WE TOOK ALL OF THIS TO A 8840 05:37:41,407 --> 05:37:43,909 SETTING AND SELECTED 60 PLUS 8841 05:37:43,909 --> 05:37:47,246 PATIENTS THAT WERE MATCHED, ONE 8842 05:37:47,246 --> 05:37:49,314 GROUP WITH WITH, ONE GROUP 8843 05:37:49,314 --> 05:37:50,516 WITHOUT GRAFT VERSUS HOST AND 8844 05:37:50,516 --> 05:37:52,618 I'LL TELL YOU NOW THAT THAT 8845 05:37:52,618 --> 05:37:53,685 BASICALLY MIRRORED WHAT WE FOWN 8846 05:37:53,685 --> 05:37:56,188 IN THESE MOUSE MODEL -- FOUND IN 8847 05:37:56,188 --> 05:37:57,156 THESE MOUSE MODELS. 8848 05:37:57,156 --> 05:37:59,057 SO STARTING WITH HAVING INDEED 8849 05:37:59,057 --> 05:38:00,993 IN PATIENTS WITH GRAFT VERSUS 8850 05:38:00,993 --> 05:38:04,062 HOST, LOWER LEVELS OF BOWEL 8851 05:38:04,062 --> 05:38:06,899 ACIDS OVERALL. 8852 05:38:06,899 --> 05:38:09,201 AGAIN, SMESKLY LOWER LEVELS OF 8853 05:38:09,201 --> 05:38:13,338 THESE SECONDARY BOWEL ACIDS AND 8854 05:38:13,338 --> 05:38:16,642 DECONJUGAT HE D BOWEL ACIDS AND 8855 05:38:16,642 --> 05:38:18,710 SHOTGUN SEQUENCING WE SAW AGAIN 8856 05:38:18,710 --> 05:38:20,345 SPECIFICALLY A LOWER EXPRESSION 8857 05:38:20,345 --> 05:38:23,282 OF BOWEL SALTS, HYDROLASE IN 8858 05:38:23,282 --> 05:38:25,484 BACTERIA FROM PATIENTS WITH 8859 05:38:25,484 --> 05:38:29,121 GRAFT VERSUS HOST. 8860 05:38:29,121 --> 05:38:31,623 AND AGAIN, THESE SECONDARY BOWEL 8861 05:38:31,623 --> 05:38:33,125 ACIDSES THAT HAD BEEN IMPLICATED 8862 05:38:33,125 --> 05:38:36,695 TO BE RELEVANT FOR TH167 AND FOR 8863 05:38:36,695 --> 05:38:38,997 REGULATORY -- TH17 AND FOR 8864 05:38:38,997 --> 05:38:40,299 REGULATORY T CELLS WERE FOUND IN 8865 05:38:40,299 --> 05:38:41,834 MUCH LOWER LEVELS IN PATIENTS 8866 05:38:41,834 --> 05:38:44,336 WITH GRAFT VERSUS HOST. 8867 05:38:44,336 --> 05:38:47,439 NOW, INTERESTING AND I GUESS 8868 05:38:47,439 --> 05:38:48,407 FORTUNATE FOR US, THERE WAS A 8869 05:38:48,407 --> 05:38:50,709 TIME HAD OUR CENTER WAS NOT 8870 05:38:50,709 --> 05:38:55,247 GIVING PATIENTS IRSODIOL, 8871 05:38:55,247 --> 05:38:57,015 SECONDARY BOWEL ACIDS I 8872 05:38:57,015 --> 05:38:58,383 MENTIONED ALREADY SO WE LOOKED 8873 05:38:58,383 --> 05:38:59,985 BACK AT PATIENTS THAT WERE NOT 8874 05:38:59,985 --> 05:39:02,754 GETTING URS DIOL AND AFTERWARDS 8875 05:39:02,754 --> 05:39:04,122 PATIENTS WHO DID AND SAW INDEED 8876 05:39:04,122 --> 05:39:10,229 A STRIKING DIFFERENCE IN THE 8877 05:39:10,229 --> 05:39:13,198 INCIDENCE HF ENTEROCOCCUS, I 8878 05:39:13,198 --> 05:39:15,067 TOLD YOU EARLY ERB IN THAT NAIRP 8879 05:39:15,067 --> 05:39:18,337 WE RECENTLY PUBLISHED THAT 8880 05:39:18,337 --> 05:39:23,575 SPECIFICALLYILY OTICA COULD BE 8881 05:39:23,575 --> 05:39:25,677 ONE OF THE DRUGS ASSOCIATED WITH 8882 05:39:25,677 --> 05:39:27,779 LOWER LEVELS OF ENTEROCOCCUS, 8883 05:39:27,779 --> 05:39:29,481 SEEMED TO BE HOLDING UP ALSO IN 8884 05:39:29,481 --> 05:39:30,215 THESE PATIENTS. 8885 05:39:30,215 --> 05:39:32,150 SO I DON'T KNOW HOW I'M DOING ON 8886 05:39:32,150 --> 05:39:32,518 TIME. 8887 05:39:32,518 --> 05:39:34,453 IT SEEMS LIKE I VINYL FEW 8888 05:39:34,453 --> 05:39:36,088 MINUTE, SO I'M GOING TO READ THE 8889 05:39:36,088 --> 05:39:37,089 WHOLE SUMMARY HERE. 8890 05:39:37,089 --> 05:39:38,957 JUST TO TELL YOU THAT WHERE WE 8891 05:39:38,957 --> 05:39:39,925 ARE AT THE MOMENT IS OF COURSE 8892 05:39:39,925 --> 05:39:42,227 TRYING TO TAKE THIS INTO THE 8893 05:39:42,227 --> 05:39:44,296 CLINIC, AS YOU KNOW, MANY 8894 05:39:44,296 --> 05:39:45,831 SETTINGS AT THE MOMENT IS WHERE 8895 05:39:45,831 --> 05:39:48,467 PEOPLE ARE TESTING FECAL 8896 05:39:48,467 --> 05:39:49,668 TRANSPLANTS, BUT WE ARE 8897 05:39:49,668 --> 05:39:50,969 INTERESTED IN AND PARTNERING 8898 05:39:50,969 --> 05:39:54,907 WITH THIS COMPANY TO GIVE REALLY 8899 05:39:54,907 --> 05:39:59,411 DEFINED CON SORE -- CONSORTIA. 8900 05:39:59,411 --> 05:40:00,546 WE ARE GIVING 16 STRAINS 8901 05:40:00,546 --> 05:40:02,948 SELECTED FOR VARIOUS FUNCTIONS 8902 05:40:02,948 --> 05:40:04,416 THAT WE THINK WOULD BE RELEVANT 8903 05:40:04,416 --> 05:40:07,553 FOR THE OUTCOME AFTER ALLOGENIC 8904 05:40:07,553 --> 05:40:08,854 TRANSPLANT FROM GRAFT VERSUS 8905 05:40:08,854 --> 05:40:11,290 HOST TO IMPROVED BARRIER TO 8906 05:40:11,290 --> 05:40:15,460 REMEMBER COLONY REZIS SENTENCE 8907 05:40:15,460 --> 05:40:17,095 AGAINST PATHOGEN -- RESISTANCE 8908 05:40:17,095 --> 05:40:19,064 AGAINST PATHOGENS, THIS TRIAL IS 8909 05:40:19,064 --> 05:40:20,165 ONGOING AT THE MOMENT AND 8910 05:40:20,165 --> 05:40:21,800 ENTERING PRESS RELEASE ACTUALLY 8911 05:40:21,800 --> 05:40:24,369 SHOWS THAT SULFUR IT -- THAT SO 8912 05:40:24,369 --> 05:40:25,771 FAR IT SEEMS TO BE WORKING 8913 05:40:25,771 --> 05:40:26,872 ACCORDING TO PLAN AND THAT 8914 05:40:26,872 --> 05:40:27,973 SPECIFIC WHAT WILL WE'RE SEEING 8915 05:40:27,973 --> 05:40:30,842 IS LESS DOMINATION WITH 8916 05:40:30,842 --> 05:40:33,045 PATHOGENS IN PATIENTS GETTING 8917 05:40:33,045 --> 05:40:34,546 BEFORE AND AT A CERTAIN TIME 8918 05:40:34,546 --> 05:40:36,615 POINT AFTER THE ALLOGENIC 8919 05:40:36,615 --> 05:40:39,017 TRANSPLANT A COUPLE OF DAYS OF 8920 05:40:39,017 --> 05:40:42,654 THIS DEFINED CONSORTIUM. 8921 05:40:42,654 --> 05:40:44,289 SO THAT'S ALL THAT -- I MEAN, I 8922 05:40:44,289 --> 05:40:46,558 HAVE A SECOND STORY, BUT I KNEW 8923 05:40:46,558 --> 05:40:48,293 ALREADY THAT I WOULDN'T GET TO 8924 05:40:48,293 --> 05:40:48,727 THAT. 8925 05:40:48,727 --> 05:40:48,994 [LAUGHTER] 8926 05:40:48,994 --> 05:40:50,696 THAT'S WHY I PUT MY END SLIDE 8927 05:40:50,696 --> 05:40:50,896 HERE. 8928 05:40:50,896 --> 05:40:52,664 OF COURSE I'M GOING TO THANK MY 8929 05:40:52,664 --> 05:40:53,765 FUNDING AND OF COURSE HERE IN 8930 05:40:53,765 --> 05:40:55,334 THIS ROOM I SPECIFICALLY WANT TO 8931 05:40:55,334 --> 05:40:59,471 THANK THE NIH THAT HAS BEEN VERY 8932 05:40:59,471 --> 05:41:01,273 GENEROUS TO US. 8933 05:41:01,273 --> 05:41:02,541 AND THE VARIOUS GRANTS TO THE 8934 05:41:02,541 --> 05:41:03,942 PEOPLE WITHIN MY LAB AND OF 8935 05:41:03,942 --> 05:41:05,010 COURSE FIRST AND MOST OF AWFUL 8936 05:41:05,010 --> 05:41:06,478 THE PEOPLE IN MY LAB WHO WERE 8937 05:41:06,478 --> 05:41:07,446 DOING ALL OF THIS WORK. 8938 05:41:07,446 --> 05:41:10,349 THANK YOU VERY MUCH. 8939 05:41:10,349 --> 05:41:18,023 [APPLAUSE] 8940 05:41:18,023 --> 05:41:22,828 >> THANK YOU. 8941 05:41:22,828 --> 05:41:25,263 QUESTIONS? 8942 05:41:25,263 --> 05:41:28,200 >> I HAVE ONE L BEAUTIFUL TALK, 8943 05:41:28,200 --> 05:41:30,836 AS ALWAYS. 8944 05:41:30,836 --> 05:41:33,538 THERE WILL NCI, FOR THOSE THAT 8945 05:41:33,538 --> 05:41:36,174 ME. 8946 05:41:36,174 --> 05:41:38,477 ACTUALLY, I WAS CAUGHT WITH THE 8947 05:41:38,477 --> 05:41:39,911 LIST OF DRUGS YOU HAD THERE THAT 8948 05:41:39,911 --> 05:41:40,979 THE PATIENTS RECEIVE AND THEY 8949 05:41:40,979 --> 05:41:42,447 ACTUALLY HAVE A REALLY HARD TIME 8950 05:41:42,447 --> 05:41:43,615 WITHOUT THEM, SO IT'S HARD TO 8951 05:41:43,615 --> 05:41:44,483 TAKE THOSE AWAY. 8952 05:41:44,483 --> 05:41:49,655 SO MY QUESTION WAS A BIT BROADER 8953 05:41:49,655 --> 05:41:51,089 AND WHAT IS THERE THAT ONE CAN 8954 05:41:51,089 --> 05:41:51,957 DO ABOUT IT? 8955 05:41:51,957 --> 05:41:53,392 BECAUSE THIS IS A QUESTION THAT 8956 05:41:53,392 --> 05:41:55,093 I KEEP ASKING, YOU KNOW, THE 8957 05:41:55,093 --> 05:41:57,529 ANSWER WE USUALLY GET FROM 8958 05:41:57,529 --> 05:41:59,831 ONCOLOGISTS IS THAT'S THE 8959 05:41:59,831 --> 05:42:00,098 PROTOCOL. 8960 05:42:00,098 --> 05:42:02,467 DO YOU WANT TO TAKE YOUR 8961 05:42:02,467 --> 05:42:02,701 CHANCES? 8962 05:42:02,701 --> 05:42:04,269 YOU KNOW, NOT THE ANSWER WE LIKE 8963 05:42:04,269 --> 05:42:05,003 TO HEAR. 8964 05:42:05,003 --> 05:42:07,739 BUT DO YOU SEE ANY POSSIBILITY 8965 05:42:07,739 --> 05:42:09,408 OF REALLY DOING, YOU KNOW, 8966 05:42:09,408 --> 05:42:10,842 MEANINGFUL STUDIES TO ADDRESS 8967 05:42:10,842 --> 05:42:15,814 THE IMPACT OF ALL THESE DRUGS IN 8968 05:42:15,814 --> 05:42:19,951 MIKE BOAM AND THEREFORE -- 8969 05:42:19,951 --> 05:42:20,619 MICROBIOME AND THEREFORE 8970 05:42:20,619 --> 05:42:21,620 RESPONSE TO SAY THERAPIES? 8971 05:42:21,620 --> 05:42:23,055 >> YES, FOR SURE. 8972 05:42:23,055 --> 05:42:24,790 A STUDY IS OPEN NOW THAT I 8973 05:42:24,790 --> 05:42:25,757 DIDN'T MENTION STARTED WITH OF 8974 05:42:25,757 --> 05:42:28,293 COURSE THE FIRST CATEGORY OF 8975 05:42:28,293 --> 05:42:30,128 DRUGS THAT WE'RE INTERESTED IN 8976 05:42:30,128 --> 05:42:31,797 WHICH IS BROAD-SPECTRUM 8977 05:42:31,797 --> 05:42:32,597 ANTIBIOTICS. 8978 05:42:32,597 --> 05:42:33,432 EVERY CENTER FOR REASONS THAT 8979 05:42:33,432 --> 05:42:34,866 ARE OFTEN NOT COMPLETELY CLEAR 8980 05:42:34,866 --> 05:42:36,935 HAVE THEIR OWN FAVORITE, BUT 8981 05:42:36,935 --> 05:42:39,004 THERE ARE PHASE III STUDIES THAT 8982 05:42:39,004 --> 05:42:40,305 HAVE DEMONSTRATED THAT ABOUT 8983 05:42:40,305 --> 05:42:42,774 FIVE, SIX DIFFERENT ONES HAVE 8984 05:42:42,774 --> 05:42:43,442 EQUAL OUTCOMES. 8985 05:42:43,442 --> 05:42:44,176 SO YOU HAVE A CHOICE. 8986 05:42:44,176 --> 05:42:45,510 YOU CAN MAKE A CHOICE DL. 8987 05:42:45,510 --> 05:42:50,315 AND JUST TO BE VERY SPECIFIC, 8988 05:42:50,315 --> 05:42:54,586 BACTIN, BANT, CEPHAPIME MUCH 8989 05:42:54,586 --> 05:42:56,321 BETTER SO WE ARE DOING A 8990 05:42:56,321 --> 05:42:57,322 RANDOMIZED STUDY AT THE MOMENT 8991 05:42:57,322 --> 05:43:00,158 WHERE PATIENTS WHO AFTER ANNAL 8992 05:43:00,158 --> 05:43:01,893 LOWGENIC TRANSPLANT HAVE FEVER 8993 05:43:01,893 --> 05:43:04,062 AND NEUTROPOENA ARE BEING 8994 05:43:04,062 --> 05:43:11,403 RANDOMIZED TO EITHER GETTING -- 8995 05:43:11,403 --> 05:43:13,605 FOR NAUSEA MEDICATIONS WHERE WE 8996 05:43:13,605 --> 05:43:15,240 CHOOSE FROM AT LEAST FIVE OR SIX 8997 05:43:15,240 --> 05:43:18,710 DIFFERENT ONES, OFTEN THEY'RE 8998 05:43:18,710 --> 05:43:20,612 EQUALLY BENEFICIAL, AND PAIN 8999 05:43:20,612 --> 05:43:20,879 MEDICINES. 9000 05:43:20,879 --> 05:43:23,448 SO CERTAINLY IN THOSE TWO 9001 05:43:23,448 --> 05:43:24,516 CATEGORIES, WE HAVE CHOICES, SO 9002 05:43:24,516 --> 05:43:29,855 WE CAN TEST THAT. 9003 05:43:29,855 --> 05:43:31,790 >> HEY, MARCEL. 9004 05:43:31,790 --> 05:43:32,591 >> HEY, DAVID. 9005 05:43:32,591 --> 05:43:33,458 >> HELLO. 9006 05:43:33,458 --> 05:43:35,327 >> HI THERE NEIGHBOR. 9007 05:43:35,327 --> 05:43:40,432 >> WE MUST STOP DATING LIKE THIS 9008 05:43:40,432 --> 05:43:41,199 IN BETHESDA. 9009 05:43:41,199 --> 05:43:43,301 WE DO LIVE NEXT-DOOR TO EACH 9010 05:43:43,301 --> 05:43:43,502 OTHER. 9011 05:43:43,502 --> 05:43:44,603 GREAT PRESENTATION, AS ALWAYS. 9012 05:43:44,603 --> 05:43:46,271 I'LL ASK YOU A SLIGHTLY BROADER 9013 05:43:46,271 --> 05:43:47,539 QUESTION THAT RELATES TO A 9014 05:43:47,539 --> 05:43:48,774 CONVERSATION I HAD EARLIER THIS 9015 05:43:48,774 --> 05:43:50,408 WEEK ABOUT THE NIH STRATEGIC 9016 05:43:50,408 --> 05:43:56,314 MISSION ON NUTRITION AND HUMAN 9017 05:43:56,314 --> 05:43:58,316 HEALTH, 2030 DOCUMENT I'M SURE 9018 05:43:58,316 --> 05:43:59,618 EVERYONE IS AWARE OF. 9019 05:43:59,618 --> 05:44:00,786 SO BEAUTIFUL WHAT YOU'RE TALKING 9020 05:44:00,786 --> 05:44:04,456 ABOUT IN TERMS OF FIBER AND CDCA 9021 05:44:04,456 --> 05:44:07,826 IN PARTICULAR AND REGULATING 9022 05:44:07,826 --> 05:44:08,827 INFLAMMATORY T CELL RESPONSES. 9023 05:44:08,827 --> 05:44:11,663 HOW DOES IT RELATE TO PRECISION 9024 05:44:11,663 --> 05:44:11,930 NUTRITION? 9025 05:44:11,930 --> 05:44:13,932 WHAT DO WE BRING THIS TO REAL 9026 05:44:13,932 --> 05:44:14,132 LIFE? 9027 05:44:14,132 --> 05:44:16,468 BECAUSE YOU DO THE GREAT BASIC 9028 05:44:16,468 --> 05:44:17,469 SCIENCE COOL EXPERIMENTS IN 9029 05:44:17,469 --> 05:44:19,638 MICE, RIGHT, ALL THE COOL STUFF. 9030 05:44:19,638 --> 05:44:21,940 BUT YOU ALSO DO THE HARD STUFF 9031 05:44:21,940 --> 05:44:23,875 IN REAL-LIFE CLINICAL TRIALS. 9032 05:44:23,875 --> 05:44:27,412 SO HOW CAN WE BRING THE 9033 05:44:27,412 --> 05:44:32,217 NUTRITION PIECE PAST, YOU KNOW, 9034 05:44:32,217 --> 05:44:34,219 AM BUY BAD PRODUCTS OR BADLY 9035 05:44:34,219 --> 05:44:35,387 LABELED IN THE SUPER MARKETED? 9036 05:44:35,387 --> 05:44:37,789 HOW DO WE BRING IT TO REAL LIFE? 9037 05:44:37,789 --> 05:44:40,625 >> RECALL SO TELL ME C WITH THE 9038 05:44:40,625 --> 05:44:43,495 HEALTH FOOD STORIES FINALLY HAVE 9039 05:44:43,495 --> 05:44:45,497 TO END, AND FOR THOSE OF YOU WHO 9040 05:44:45,497 --> 05:44:47,799 ARE NOT CLINICALLY ACTIVE BUT 9041 05:44:47,799 --> 05:44:50,101 EVERYBODY WHO IS CLINICALLY 9042 05:44:50,101 --> 05:44:51,503 ACTIVE AND IS TREATING A CANCER 9043 05:44:51,503 --> 05:44:53,338 PATIENT AT ONE POINT WILL GET 9044 05:44:53,338 --> 05:44:56,441 THE QUESTION, WHAT SHOULD I EAT, 9045 05:44:56,441 --> 05:44:56,975 DOCTOR? 9046 05:44:56,975 --> 05:44:59,377 IT'S ON EVERY PATIENT'S MIND 9047 05:44:59,377 --> 05:44:59,811 ALWAYS. 9048 05:44:59,811 --> 05:45:02,047 SO DIET STUDIES CAN BE DONE 9049 05:45:02,047 --> 05:45:03,415 BETTER, I'M SURE YOU KNOW ABOUT 9050 05:45:03,415 --> 05:45:04,950 SOME COMPANIES THAT ARE ACTUALLY 9051 05:45:04,950 --> 05:45:09,120 DOING THAT BASED UPON MICROBIOME 9052 05:45:09,120 --> 05:45:10,155 PROFILING, WHAT WE ARE DOING AT 9053 05:45:10,155 --> 05:45:12,090 THE MOMENT IS IN TWO WAYS 9054 05:45:12,090 --> 05:45:14,192 ANALYZING IN GREAT DETAIL WHAT 9055 05:45:14,192 --> 05:45:15,594 PEOPLE ARE EATING. 9056 05:45:15,594 --> 05:45:18,897 FIRST OF ALL, WHEN THEY ARE 9057 05:45:18,897 --> 05:45:19,965 INPATIENTS WE KNOW EXACTLY WHAT 9058 05:45:19,965 --> 05:45:21,900 FOOD WE ARE SERVING AND WE KNOW 9059 05:45:21,900 --> 05:45:23,702 EXACTLY ALL OF THE DETAILS OF 9060 05:45:23,702 --> 05:45:25,637 EVERY FIBER CONTENT AND 9061 05:45:25,637 --> 05:45:26,638 WHATEVER, AND I DIDN'T WANT TO 9062 05:45:26,638 --> 05:45:28,406 GO, I MEAN, THAT IS A WHOLE TALK 9063 05:45:28,406 --> 05:45:30,609 THAT I DIDN'T WANT TO GIVE NOW. 9064 05:45:30,609 --> 05:45:33,211 SO THERE WE CAN GET IN MUCH MORE 9065 05:45:33,211 --> 05:45:36,281 DETAIL ABOUT WHAT TYPE OF 9066 05:45:36,281 --> 05:45:36,815 NUTRIENTS. 9067 05:45:36,815 --> 05:45:38,116 BUT THERE'S A SECOND WAY OF 9068 05:45:38,116 --> 05:45:41,953 DOING IT WHICH IS A TAXONOMY OF 9069 05:45:41,953 --> 05:45:42,153 FOOD. 9070 05:45:42,153 --> 05:45:44,723 YOU ARE MIGHT HAVE SEEN SOME OF 9071 05:45:44,723 --> 05:45:47,392 THE TALKS BY FORMER POST-DOC WHO 9072 05:45:47,392 --> 05:45:52,864 NOW HAS HIS OWN LAB, SONY PALLET 9073 05:45:52,864 --> 05:45:54,432 WHERE HE FOR INSTANCE SHOWS HA 9074 05:45:54,432 --> 05:45:57,269 CERTAIN FRUITS AND JUICES 9075 05:45:57,269 --> 05:45:58,436 SPECIFICALLY PROVIDE SUGAR THAT 9076 05:45:58,436 --> 05:46:00,438 IS CAN BE BAD FOR THE OUTGROWTH 9077 05:46:00,438 --> 05:46:01,840 OF THESE PATD GENERALS THAT CAN 9078 05:46:01,840 --> 05:46:02,707 LEAD TO DOMINANCE. 9079 05:46:02,707 --> 05:46:05,810 SO THERE ARE TWO WAYS -- 9080 05:46:05,810 --> 05:46:06,077 PATHOGENS. 9081 05:46:06,077 --> 05:46:08,213 TWO WAYS OF ANALYZING, SIMPLY 9082 05:46:08,213 --> 05:46:09,681 THE CONTENTS OF THE NUTRIENTS 9083 05:46:09,681 --> 05:46:11,716 AND SECOND THE FOOD GROUPS. 9084 05:46:11,716 --> 05:46:12,884 BASED UPON THAT, WE SHOULD BE 9085 05:46:12,884 --> 05:46:15,086 ABLE TO DEFINE STUDIES. 9086 05:46:15,086 --> 05:46:17,689 WE'RE DOING SOME OF THAT ALREADY 9087 05:46:17,689 --> 05:46:19,925 WITHIN MYELOMA, DIFFERENT 9088 05:46:19,925 --> 05:46:21,426 SEGHTD, WHERE WE HAVE POSITIVE 9089 05:46:21,426 --> 05:46:25,196 OUTCOMES WHEN WE JUST INCREASE 9090 05:46:25,196 --> 05:46:26,398 FIBER, WHICH IS A VERY OBVIOUS 9091 05:46:26,398 --> 05:46:27,966 ONE AND I DON'T HAVE TO TELL YOU 9092 05:46:27,966 --> 05:46:29,267 WHY, RIGHT? 9093 05:46:29,267 --> 05:46:31,002 SHORT CHAIN FATTY ACID, HEALING 9094 05:46:31,002 --> 05:46:33,705 OF THE GUT, ET CETERA. 9095 05:46:33,705 --> 05:46:35,607 >> MY QUESTION IS A COMPLEMENT, 9096 05:46:35,607 --> 05:46:36,908 NOT A CRITICISM I THINK WHERE 9097 05:46:36,908 --> 05:46:40,312 CANCER IS IN TERMS OF 9098 05:46:40,312 --> 05:46:41,813 UNDERSTANDING NEW THRITION AL 9099 05:46:41,813 --> 05:46:43,581 INTERVENTION INSIDE -- 9100 05:46:43,581 --> 05:46:45,684 NUTRITIONAL INTERVENTIONS IN 9101 05:46:45,684 --> 05:46:48,119 HUMAN HEALTH AND DISEASE,LESS 9102 05:46:48,119 --> 05:46:49,921 SOME OF THE BEST STUDIES BEING 9103 05:46:49,921 --> 05:46:52,557 DONE IN HUMAN AND MOUSE RELATE 9104 05:46:52,557 --> 05:46:54,626 AROUND HOW NUTRITION IS 9105 05:46:54,626 --> 05:46:54,893 AFFECTING. 9106 05:46:54,893 --> 05:46:55,961 ONE OF THE STUDIES YOU WERE 9107 05:46:55,961 --> 05:46:57,162 INVOLVED IN I THINK HIGHLIGHTS 9108 05:46:57,162 --> 05:46:59,431 IT THAT, YOU KNOW, HIGH FIBER 9109 05:46:59,431 --> 05:47:01,533 DIETS CAN BE BENEFICIAL, 9110 05:47:01,533 --> 05:47:04,135 PROBIOTICS CAN BE DETRIMENTAL IN 9111 05:47:04,135 --> 05:47:04,836 CERTAIN CIRCUMSTANCES. 9112 05:47:04,836 --> 05:47:05,537 >> I THINK -- 9113 05:47:05,537 --> 05:47:08,406 >> AND I HAVE TO STOP TALKING. 9114 05:47:08,406 --> 05:47:09,507 BUT I THINK YOU WERE PROVIDING 9115 05:47:09,507 --> 05:47:13,545 US WITH -- SO THANK YOU. 9116 05:47:13,545 --> 05:47:15,747 >> THANK YOU RK DAVID WE'LL TALK 9117 05:47:15,747 --> 05:47:15,947 LATER. 9118 05:47:15,947 --> 05:47:17,816 >> SHORT QUESTIONS ZEES SURE. 9119 05:47:17,816 --> 05:47:19,451 I WILL MAKE OUT WITH THIS 9120 05:47:19,451 --> 05:47:20,785 MICROPHONE LIKE NOBODY'S 9121 05:47:20,785 --> 05:47:21,052 BUSINESS. 9122 05:47:21,052 --> 05:47:22,287 >> THANK YOU. 9123 05:47:22,287 --> 05:47:24,089 >> DAVID ALREADY ASKED A BIG 9124 05:47:24,089 --> 05:47:25,757 PART OF MY QUESTION. 9125 05:47:25,757 --> 05:47:28,126 YOU KNOW, RELATED TO PRECISION 9126 05:47:28,126 --> 05:47:31,229 MEDICINE AND HOW YOUR BEAUTIFUL 9127 05:47:31,229 --> 05:47:33,398 PHASE I THRIEL YOU'RE -- THAT 9128 05:47:33,398 --> 05:47:35,000 YOU SPOKE ABOUT AT THE END, 9129 05:47:35,000 --> 05:47:36,634 YOU'RE ACTUALLY TAKING CONSORTIA 9130 05:47:36,634 --> 05:47:38,169 AND APPLYING IT TO EVERYONE BUT 9131 05:47:38,169 --> 05:47:40,472 IT'S THE SAME CONSORTIA SHA 9132 05:47:40,472 --> 05:47:42,774 WHICH I'M WONDERING HOW DO 9133 05:47:42,774 --> 05:47:44,609 DIFFERENT PEOPLE REACT TO THAT 9134 05:47:44,609 --> 05:47:46,378 SAME CONSORTIA AND I WAS REALLY 9135 05:47:46,378 --> 05:47:47,712 IMPRESSED THAT THERE WERE NO 9136 05:47:47,712 --> 05:47:49,781 ADVERSE EVENTS IN CONTRAST TO 9137 05:47:49,781 --> 05:47:53,518 THE LAST TRIAL, SO I'M CURIOUS 9138 05:47:53,518 --> 05:47:55,253 WHAT LESSONS HAVE YOU LEARNED 9139 05:47:55,253 --> 05:47:56,688 AND IMPROVED UPON THAT HAVE 9140 05:47:56,688 --> 05:47:58,123 ALLOWED YOU TO REACH THIS 9141 05:47:58,123 --> 05:47:59,190 REMARKABLE MILESTONE? 9142 05:47:59,190 --> 05:48:05,463 >> YEAH 6789. 9143 05:48:05,463 --> 05:48:08,099 SO YOU'RE HINTING, PART OF AN 9144 05:48:08,099 --> 05:48:10,668 NIH PROJECT AND THAT IS TO 9145 05:48:10,668 --> 05:48:12,871 UPFRONT MEASURE WHAT'S THE 9146 05:48:12,871 --> 05:48:15,807 DEFICIENT SIZ THAT MIGHT BE IN 9147 05:48:15,807 --> 05:48:18,243 SOMEBODY'S MICROBIOME. 9148 05:48:18,243 --> 05:48:20,945 BASED UPON THAT PROFILE, DEFINE 9149 05:48:20,945 --> 05:48:23,281 A SPECIFIC CONSORTIUM. 9150 05:48:23,281 --> 05:48:24,883 SO PRECISION MEDICINE IN TERMS 9151 05:48:24,883 --> 05:48:26,418 OF THE CONSORTIA THAT WE WOULD 9152 05:48:26,418 --> 05:48:31,356 WANT TO GIVE. 9153 05:48:31,356 --> 05:48:32,690 THINK ONE OF THE REASONS WHY OUR 9154 05:48:32,690 --> 05:48:34,426 STRATEGY WAS RELATIVELY SAFE WAS 9155 05:48:34,426 --> 05:48:35,527 BECAUSE OF THE BACTERIA ONLY 9156 05:48:35,527 --> 05:48:36,628 BEING GIVEN BEFORE THE 9157 05:48:36,628 --> 05:48:37,562 CONDITIONING REGIMEN. 9158 05:48:37,562 --> 05:48:40,665 AT THAT POINT WE DIDN'T GIVE 9159 05:48:40,665 --> 05:48:42,534 ANYTHING ANYMORE UNTIL THEY WERE 9160 05:48:42,534 --> 05:48:44,169 OFF ANTIBIOTICS AND HAD THEIR 9161 05:48:44,169 --> 05:48:45,203 NEUTROPHILS BACK, SO THEY WERE 9162 05:48:45,203 --> 05:48:48,873 ABOUT THREE WEEKS OUT FROM 9163 05:48:48,873 --> 05:48:50,408 ALLOGENIC TRANSPLANT, THEN THEY 9164 05:48:50,408 --> 05:48:51,709 GOT A SECOND COURSE, MAYBE 9165 05:48:51,709 --> 05:48:53,144 THAT'S WHY IT WAS RELATIVELY 9166 05:48:53,144 --> 05:48:54,112 ZIEF VERY IMPRESSIVE. 9167 05:48:54,112 --> 05:48:54,579 >> THANK YOU. 9168 05:48:54,579 --> 05:49:04,823 >> THANK YOU. 9169 05:49:11,096 --> 05:49:14,833 >> IS IT OKAY IF I USE THIS? 9170 05:49:14,833 --> 05:49:25,143 >> THE OTHER ONE. 9171 05:49:29,280 --> 05:49:29,747 HAD. 9172 05:49:29,747 --> 05:49:30,849 >> DOES THIS WORK? 9173 05:49:30,849 --> 05:49:31,816 OKAY, GOOD. 9174 05:49:31,816 --> 05:49:37,088 OH, THAT'S NOT MY TALK. 9175 05:49:37,088 --> 05:49:39,057 OKAY. 9176 05:49:39,057 --> 05:49:44,762 R MY TALK WITHOUT GLASSES, NOT 9177 05:49:44,762 --> 05:49:44,963 EASY. 9178 05:49:44,963 --> 05:49:47,465 I WANT TO THANK THE ORGANIZER 9179 05:49:47,465 --> 05:49:51,870 FOR INVITING ME TO PRESENT HERE 9180 05:49:51,870 --> 05:49:52,337 TODAY. 9181 05:49:52,337 --> 05:49:52,537 OKAY. 9182 05:49:52,537 --> 05:49:53,071 THERE WE ARE. 9183 05:49:53,071 --> 05:49:55,273 I'M REALLY HAPPY AND ACTUALLY 9184 05:49:55,273 --> 05:49:57,408 HAD, THIS COULD HAVE NOT BEEN 9185 05:49:57,408 --> 05:49:59,677 BETTER BACK TO BACK 9186 05:49:59,677 --> 05:50:01,746 PRESENTATIONS AND ALL THE 9187 05:50:01,746 --> 05:50:05,016 QUESTIONS HAVE REALLY KIND OF 9188 05:50:05,016 --> 05:50:06,751 INTRODUCED WHAT I'M GOING TO 9189 05:50:06,751 --> 05:50:10,155 TALK ABOUT, AND YOU'LL SEE WHY 9190 05:50:10,155 --> 05:50:11,856 I'M SAYING THIS IN A MOMENT, BUT 9191 05:50:11,856 --> 05:50:13,324 I'M GOING TO FOCUS ON WHAT WE'RE 9192 05:50:13,324 --> 05:50:17,428 DOING ON TRY TO UNDERSTAND HOW 9193 05:50:17,428 --> 05:50:19,297 MICROBIOTA ACTUALLY REGULATE 9194 05:50:19,297 --> 05:50:21,432 THESE ANTI-TUMOR RESPONSES. 9195 05:50:21,432 --> 05:50:22,934 I DON'T NEED TO TELL YOU THAT 9196 05:50:22,934 --> 05:50:25,103 TUMORS DEVELOP ALONGSIDE A 9197 05:50:25,103 --> 05:50:29,374 HIGHLY COMPLEX AND DYNAMIC TUMOR 9198 05:50:29,374 --> 05:50:30,508 MICROENVIRONMENT AND ALL THE 9199 05:50:30,508 --> 05:50:32,377 RESULTS WE CAN FIND IN IMMUNE 9200 05:50:32,377 --> 05:50:36,114 CELLS PRESENT IN THESE TUMOR 9201 05:50:36,114 --> 05:50:39,484 MICROENVIRONMENT, THOSE FAMILIAR 9202 05:50:39,484 --> 05:50:42,320 WITH MACK FAJS, MONOCYTES AND 9203 05:50:42,320 --> 05:50:43,288 DENDRITIC CELLS MAJOR COMPONENTS 9204 05:50:43,288 --> 05:50:44,589 OF THESE TME. 9205 05:50:44,589 --> 05:50:46,758 THESE CELLS ARE CRITICAL 9206 05:50:46,758 --> 05:50:49,494 INITIATORS AND REGULATORS OF 9207 05:50:49,494 --> 05:50:50,929 IMMUNITY AND PLAY A VERY 9208 05:50:50,929 --> 05:50:55,833 IMPORTANT ROLE BOTH IN 9209 05:50:55,833 --> 05:50:56,935 PATROLMANNING OR CONTROLLING 9210 05:50:56,935 --> 05:50:58,002 TUMOR PROGRESSION AS WELL AS 9211 05:50:58,002 --> 05:50:59,137 RESPONSE TO THERAPY. 9212 05:50:59,137 --> 05:51:00,238 PROBABLY FOR THE MOST PART WHEN 9213 05:51:00,238 --> 05:51:01,839 PEOPLE THINK ABOUT MACROFADGES 9214 05:51:01,839 --> 05:51:03,841 IN THE CONTEXT OF THE TME THEY 9215 05:51:03,841 --> 05:51:06,144 TONED THINK ABOUT A PROTEOGENIC 9216 05:51:06,144 --> 05:51:07,011 CELL TYPE WHILE ON THE OTHER 9217 05:51:07,011 --> 05:51:09,948 HAND WHEN THEY THINK ABOUT 9218 05:51:09,948 --> 05:51:12,050 DENDRITIC CELLS, THEIR UNIQUE 9219 05:51:12,050 --> 05:51:14,519 ABILITY TO PRIME OR ACTIVATE 19 9220 05:51:14,519 --> 05:51:17,288 CELLS WE THINK ABOUT THEM AS 9221 05:51:17,288 --> 05:51:18,923 ANTI-TUMORGENIC, BUT THE 9222 05:51:18,923 --> 05:51:20,258 REALITIES THAT NOTHING IS SO 9223 05:51:20,258 --> 05:51:22,227 BLACK AND WHITE AND THESE CELLS 9224 05:51:22,227 --> 05:51:27,699 ARE VERY PLASTIC AND THEY CAN 9225 05:51:27,699 --> 05:51:32,537 ALLLY OBSERVE -- AND 9226 05:51:32,537 --> 05:51:33,304 ANTI-TUMORGENIC. 9227 05:51:33,304 --> 05:51:36,474 WHAT ARE THE CUE FOR THESE -- 9228 05:51:36,474 --> 05:51:38,109 THAT REGULATE IN THE TME? 9229 05:51:38,109 --> 05:51:39,577 I DON'T NEED TOLL YOU NOW WE 9230 05:51:39,577 --> 05:51:42,046 KNOW THAT MICROBUY ON THE 9231 05:51:42,046 --> 05:51:44,983 MICROBIOTA PLAYS A IMPORTANT 9232 05:51:44,983 --> 05:51:46,317 ROLE IN ANTICANCER TREATMENT. 9233 05:51:46,317 --> 05:51:48,286 FOR THE MOST PART THE STUDIES 9234 05:51:48,286 --> 05:51:50,355 THAT NOW ARE MANY THAT HAVE BEEN 9235 05:51:50,355 --> 05:51:52,657 TAKING PLACE IN THE PAST DECADE 9236 05:51:52,657 --> 05:51:56,728 OR SO HAVE MOSTLY FOCUSED ON THE 9237 05:51:56,728 --> 05:51:58,229 ADAPTING IMMUNE RESPONSE. 9238 05:51:58,229 --> 05:52:00,365 SO WHAT THESE STUDIES HAVE SHOWN 9239 05:52:00,365 --> 05:52:03,401 IS THAT MICROBIOTA INFLUENCES 9240 05:52:03,401 --> 05:52:05,670 RESPONSE TO ANTICANCER THERAPIES 9241 05:52:05,670 --> 05:52:07,405 AND THIS IS EITHER BY DIRECTLY 9242 05:52:07,405 --> 05:52:10,041 OR INDIRECTLY REGULATING 9243 05:52:10,041 --> 05:52:12,043 ANTI-TUMOR T CELL FUNCTION. 9244 05:52:12,043 --> 05:52:14,879 HOWEVER, EVEN THE STUDIES THAT 9245 05:52:14,879 --> 05:52:20,485 IDENTIFY SPECIFIC MICROBIOTA 9246 05:52:20,485 --> 05:52:21,886 METABOLITES, T CELL INFECTION, 9247 05:52:21,886 --> 05:52:23,321 THEY SHOW THESE BENEFICIAL 9248 05:52:23,321 --> 05:52:25,156 EFFECTS OF MICROBOT AT THAT 9249 05:52:25,156 --> 05:52:28,993 STILL RELIES ON THE PRESENCE OF 9250 05:52:28,993 --> 05:52:31,763 MONO THUK LAR. 9251 05:52:31,763 --> 05:52:33,197 OUR PARTICULAR INTEREST IN THESE 9252 05:52:33,197 --> 05:52:36,234 CELLS TO ASK WHETHER MICROBIOTA 9253 05:52:36,234 --> 05:52:40,905 REGULATE THE FUNCTIONAL STATE OF 9254 05:52:40,905 --> 05:52:42,573 MONONUCLEAR PHAGOCYTES AND IF SO 9255 05:52:42,573 --> 05:52:42,740 HOW. 9256 05:52:42,740 --> 05:52:45,710 I WANT TO MAKE A VERY LONG STORY 9257 05:52:45,710 --> 05:52:48,012 VERY SHORT AND GIVE YOU THE 9258 05:52:48,012 --> 05:52:49,113 SUMMARY BUT BASICALLY THIS IS 9259 05:52:49,113 --> 05:52:51,082 THE WORK, THIS IS PUBLISHED SO 9260 05:52:51,082 --> 05:52:52,583 I'M GOING TO GO THROUGH ALL THE 9261 05:52:52,583 --> 05:52:56,020 DETAILS, THIS WORK WAS LED BY 9262 05:52:56,020 --> 05:52:59,190 TWO FELLOWS IN THE LAB, AND WHAT 9263 05:52:59,190 --> 05:53:05,229 THEY SHOWED WAS THAT MICROBIOTA 9264 05:53:05,229 --> 05:53:10,301 AGO -- STING AGONISTS, ANTI-TOMB 9265 05:53:10,301 --> 05:53:14,572 MONO SIETLE IN TERM ACTIVATING 9266 05:53:14,572 --> 05:53:20,278 NK CELLS THAT PRODUCE XCL1 AND 9267 05:53:20,278 --> 05:53:22,880 CCL5 AND AT THE SAME TIME TYPE 1 9268 05:53:22,880 --> 05:53:26,417 INTERFERON HAS A MONOCYTE THAT 9269 05:53:26,417 --> 05:53:27,952 DIFFERENTIATES INTO MACROPHAGES 9270 05:53:27,952 --> 05:53:30,755 WITH A MORE ANTI-TUMORGENIC 9271 05:53:30,755 --> 05:53:31,222 PROFILE. 9272 05:53:31,222 --> 05:53:35,126 IN THE ABSENCE OF MICROBIOTA, 9273 05:53:35,126 --> 05:53:37,528 THIS CASCADE IS ALTERED AND 9274 05:53:37,528 --> 05:53:41,799 MONOCYTES INSTEAD DEFAULT ON A 9275 05:53:41,799 --> 05:53:44,302 MORE PROTEINGENIC MACROPHAGE 9276 05:53:44,302 --> 05:53:44,535 PROFILE. 9277 05:53:44,535 --> 05:53:46,838 WE WERE ABLE TO SHOW THAT 9278 05:53:46,838 --> 05:53:52,744 ACTUALLY HAD SIMPLY 9279 05:53:52,744 --> 05:53:55,113 ADMINISTERING CDR, STING, WE 9280 05:53:55,113 --> 05:53:57,014 COULD COMPLETELY RESTORE THE 9281 05:53:57,014 --> 05:53:59,751 SYSTEM INDUCE TYPE I IN K AXIS 9282 05:53:59,751 --> 05:54:02,520 AND RESTORE THE REPERTOIRE OF 9283 05:54:02,520 --> 05:54:06,124 MONONUCLEAR PHAGOCYTES. 9284 05:54:06,124 --> 05:54:07,592 SO WE BASICALLY ESTABLISHED THAT 9285 05:54:07,592 --> 05:54:12,063 ACTUALLY INDEED MICROBIOTA 9286 05:54:12,063 --> 05:54:14,031 REGULATE ANTI-TUMOR IMMUNITY VIA 9287 05:54:14,031 --> 05:54:16,367 THE REGULATION OF MONONUCLEAR 9288 05:54:16,367 --> 05:54:19,070 PHAGOCYTES IN TME AND THEIR VERY 9289 05:54:19,070 --> 05:54:20,204 COMPLEX INTERPLAY WITH OTHER 9290 05:54:20,204 --> 05:54:23,641 CELLS AND SHOWED YOU CDM COULD 9291 05:54:23,641 --> 05:54:25,743 RESCUE FUNCTIONING IN TME BUT 9292 05:54:25,743 --> 05:54:26,978 THE QUESTION WAS IS THERE 9293 05:54:26,978 --> 05:54:29,247 ANYTHING WE COULD DO TO 9294 05:54:29,247 --> 05:54:30,448 MANIPULATE MICROBIOTA PRIOR SO 9295 05:54:30,448 --> 05:54:32,517 THAT WE CAN SET THE STAGE FOR A 9296 05:54:32,517 --> 05:54:34,585 MORE PERMISSIVE ANTI-TUMOR 9297 05:54:34,585 --> 05:54:37,455 RESPONSE AND BETTER RESPONSES TO 9298 05:54:37,455 --> 05:54:37,889 THERAPY? 9299 05:54:37,889 --> 05:54:40,291 AND AS MENTIONED AMONG THE MANY 9300 05:54:40,291 --> 05:54:42,794 APPROACHES THAT WE CAN USE TO 9301 05:54:42,794 --> 05:54:46,197 CHANGE OR ALTER MICROBIOTA 9302 05:54:46,197 --> 05:54:48,499 COMPOSITION, WE HAVE THE FMT'S, 9303 05:54:48,499 --> 05:54:51,202 WE HAVE DIET INTERVENTIONS, WE 9304 05:54:51,202 --> 05:54:52,737 KNOW THE USE OF ANTIBIOTICS OR 9305 05:54:52,737 --> 05:54:55,373 SINGLE AGENT OR COMBINED 9306 05:54:55,373 --> 05:54:55,673 PROBIOTICS. 9307 05:54:55,673 --> 05:54:57,975 SO WE DECIDED TO START WITH DIET 9308 05:54:57,975 --> 05:55:00,344 BECAUSE WE FIGURE IT'S PRETTY 9309 05:55:00,344 --> 05:55:02,380 EASY TO TRANSLATE CLINICAL LI 9310 05:55:02,380 --> 05:55:03,948 AND PROBABLY A LITTLE BIT MORE 9311 05:55:03,948 --> 05:55:07,418 PLEASANT THAN SOME OF THE OTHER 9312 05:55:07,418 --> 05:55:08,586 APPROACHES. 9313 05:55:08,586 --> 05:55:10,788 AND INDEED IN THIS CASE WAS WE 9314 05:55:10,788 --> 05:55:12,523 PUT ANIMALS ON DIFFERENT DIETS 9315 05:55:12,523 --> 05:55:14,826 AND THEN HE LOOKED AT THE EFFECT 9316 05:55:14,826 --> 05:55:17,261 OF THESE DIETS ON TUMOR GROWTH. 9317 05:55:17,261 --> 05:55:18,896 WHAT YOU'RE LOOKING AT IN THIS 9318 05:55:18,896 --> 05:55:20,865 GRAPH HERE IS THE EFFECT OF THE 9319 05:55:20,865 --> 05:55:23,367 DIET OF EACH OF THESE DIET 9320 05:55:23,367 --> 05:55:26,237 COMPARED TO THE STANDARD CHOW 9321 05:55:26,237 --> 05:55:28,072 THAT WE FEED THE MICE AND THE 9322 05:55:28,072 --> 05:55:28,873 OVERALL TUMOR GROWTH. 9323 05:55:28,873 --> 05:55:31,809 SO ANYTHING THAT IS TO THE 9324 05:55:31,809 --> 05:55:35,079 RIGHT, SO ABOVE C MEANS IT'S A 9325 05:55:35,079 --> 05:55:36,948 DETRIMENTAL EFFECT THE TUMORS 9326 05:55:36,948 --> 05:55:38,716 WERE BIGGER AND ANYTHING BELOW 9327 05:55:38,716 --> 05:55:40,051 ZERO SHOWS A BENEFICIAL EFFECT 9328 05:55:40,051 --> 05:55:41,118 OF THE DIET. 9329 05:55:41,118 --> 05:55:43,521 AND YOU CAN SEE THAT DIFFERENT 9330 05:55:43,521 --> 05:55:44,722 DIETS MAY HAVE DIFFERENT 9331 05:55:44,722 --> 05:55:44,956 EFFECTS. 9332 05:55:44,956 --> 05:55:47,124 SOME ARE ACTUALLY WORSE, SOME 9333 05:55:47,124 --> 05:55:48,326 DON'T MAKE ANY DIFFERENCE AND 9334 05:55:48,326 --> 05:55:50,728 SOME CAN BE VERY POWERFUL AT 9335 05:55:50,728 --> 05:55:51,896 IMPROVING TUMOR GROWTH CONTROL 9336 05:55:51,896 --> 05:55:54,665 AS IS THE CASE OF HIGH FIBER 9337 05:55:54,665 --> 05:55:54,999 DIETS. 9338 05:55:54,999 --> 05:55:56,701 NOW, I JUST WANT TO POINT OUT 9339 05:55:56,701 --> 05:55:57,535 HERE BECAUSE WE WERE TALKING AND 9340 05:55:57,535 --> 05:55:59,270 THE QUESTION CAME UP ABOUT, YOU 9341 05:55:59,270 --> 05:56:01,138 KNOW, WHAT DO WE EAT, WHAT 9342 05:56:01,138 --> 05:56:04,308 FIBERS, ET CETERA, NOT EVERY 9343 05:56:04,308 --> 05:56:06,277 FIBER HAS THE SAME EFFECT. SO 9344 05:56:06,277 --> 05:56:07,912 THAT'S SOMETHING TO ADD TO THE 9345 05:56:07,912 --> 05:56:09,313 DISCUSSION THAT WAS HAPPENING 9346 05:56:09,313 --> 05:56:09,547 EARLIER. 9347 05:56:09,547 --> 05:56:11,716 BUT WE DECIDED TO FOCUS ON THESE 9348 05:56:11,716 --> 05:56:13,384 FIBER IN PARTICULAR THAT HAD THE 9349 05:56:13,384 --> 05:56:16,254 MOST STRIKING EFFECT AND WE DID 9350 05:56:16,254 --> 05:56:17,688 ALL OUR WORK DOWNSTREAM ANALYSIS 9351 05:56:17,688 --> 05:56:19,657 WITH THAT AND WE SHOWED THAT 9352 05:56:19,657 --> 05:56:21,592 INDEED NOT ONLY IT IMPROVES 9353 05:56:21,592 --> 05:56:23,394 TUMOR GROWTH CONTROL, IT 9354 05:56:23,394 --> 05:56:25,296 ACTUALLY HAS A SYNERGISTIC 9355 05:56:25,296 --> 05:56:30,134 EFFECT WHEN WE TREAT THE ANIMALS 9356 05:56:30,134 --> 05:56:34,805 WITH ANTI-P PDL1. 9357 05:56:34,805 --> 05:56:36,374 DIS YOU MAY BE WONDERING DOES 9358 05:56:36,374 --> 05:56:37,241 THIS HAVE ANYTHING TO DO WITH 9359 05:56:37,241 --> 05:56:40,077 THE MECHANISM I SHOWED YOU THERE 9360 05:56:40,077 --> 05:56:41,512 WITH AXIS AND THE SHORT ANSWER 9361 05:56:41,512 --> 05:56:42,713 IS YES, IT DOES. 9362 05:56:42,713 --> 05:56:44,882 ACTUALLY WHAT WE FOUND WAS THAT 9363 05:56:44,882 --> 05:56:46,317 HAD THE ANIMALS THAT WERE ON 9364 05:56:46,317 --> 05:56:48,052 HIGH FIBER DIET WHEN WE LOOKED 9365 05:56:48,052 --> 05:56:54,759 AT THE TUMOR IN THE MIKE 9366 05:56:54,759 --> 05:56:58,229 MICROBIOTA ENVIRONMENT, FOUND IT 9367 05:56:58,229 --> 05:57:03,467 INCREASED PRODUCTION -- COMPARED 9368 05:57:03,467 --> 05:57:04,368 TO CONTROL. 9369 05:57:04,368 --> 05:57:05,803 FURTHERMORE AND IF WE THINK 9370 05:57:05,803 --> 05:57:09,974 ABOUT A PROBIOTICS APPROACH, WE 9371 05:57:09,974 --> 05:57:11,809 LOOKED AT THE MICROBIOME CHANGES 9372 05:57:11,809 --> 05:57:15,246 IN THESE ANIMALS AND IDENTIFY. 9373 05:57:15,246 --> 05:57:16,948 A BACTERIUM THAT WAS ACTUALLY 9374 05:57:16,948 --> 05:57:18,749 ENRICHED BY THE HIGH FIBER DIET 9375 05:57:18,749 --> 05:57:21,752 THAT WAS CAPABLE OF COMPLETELY 9376 05:57:21,752 --> 05:57:23,588 RECAPITULATING THE FIBER DIET 9377 05:57:23,588 --> 05:57:25,256 BENEFICIAL EFFECT. 9378 05:57:25,256 --> 05:57:28,593 TRIGGERING EXACTLY ALL THE SAME 9379 05:57:28,593 --> 05:57:30,094 INNATE MECHANISMS. 9380 05:57:30,094 --> 05:57:32,863 SO HOW IS THIS RELEVANT TO 9381 05:57:32,863 --> 05:57:33,164 PATIENTS? 9382 05:57:33,164 --> 05:57:34,498 WE JUST HEARD A LOT OF THESE 9383 05:57:34,498 --> 05:57:36,167 TYPE OF APPROACHES ARE ACTUALLY 9384 05:57:36,167 --> 05:57:37,668 BEING USE IN THE CLINIC. SO WE 9385 05:57:37,668 --> 05:57:39,637 ASKED THE SAME QUESTION, AND HAD 9386 05:57:39,637 --> 05:57:44,809 WHAT WE FOUND WAS THAT ACTUALLY 9387 05:57:44,809 --> 05:57:47,979 FMT, SO MICROBIOTA FROM IMMUNE 9388 05:57:47,979 --> 05:57:50,915 BLOCKADE WAS IN A PATIENT, 9389 05:57:50,915 --> 05:57:53,751 TRANSFERRED INTO ANIMALS, 9390 05:57:53,751 --> 05:57:56,554 CAPABLE OF COMPLETELY 9391 05:57:56,554 --> 05:57:57,855 RERECAPITULATE THE SAME 9392 05:57:57,855 --> 05:58:00,491 PHENOTYPE, TRIGGEREDLY TYPE 1 9393 05:58:00,491 --> 05:58:02,293 AND K PATHWAY, REMODEL THE 9394 05:58:02,293 --> 05:58:03,628 REPERTOIRE AND LED TO IMPROVED 9395 05:58:03,628 --> 05:58:06,030 TUMOR GROWTH CONTROL, NOT ONLY 9396 05:58:06,030 --> 05:58:07,665 THAT, WHEN WE ANALYZED DATA, 9397 05:58:07,665 --> 05:58:09,734 WHEN WE ANALYZED TUMORS, RNA 9398 05:58:09,734 --> 05:58:12,770 SEQUENCING FROM TUMOR SAMPLES OF 9399 05:58:12,770 --> 05:58:14,438 THE HUMAN TRIALS THAT HAVE BEEN 9400 05:58:14,438 --> 05:58:16,741 RECENTLY PUBLISHED, WE FOUND 9401 05:58:16,741 --> 05:58:19,343 THAT ACTUALLY THE HUMAN TO HUMAN 9402 05:58:19,343 --> 05:58:21,412 FMT RESULTED IN THE SAME, 9403 05:58:21,412 --> 05:58:23,180 INCREASING ALL THESE INGNAT 9404 05:58:23,180 --> 05:58:25,049 IMMUNE SIGNATURES THAT WERE 9405 05:58:25,049 --> 05:58:27,818 ASSOCIATED WITH BETTER RESPONSE 9406 05:58:27,818 --> 05:58:28,819 TO IMMUNE BLOCKADE. 9407 05:58:28,819 --> 05:58:31,255 SO ALL OF THESE IS CURRENTLY 9408 05:58:31,255 --> 05:58:33,290 ACTUALLY BEING DONE IN THE 9409 05:58:33,290 --> 05:58:33,958 CLINIC. 9410 05:58:33,958 --> 05:58:36,460 NOW, THE REALITY IS THAT AND 9411 05:58:36,460 --> 05:58:38,262 THAT WAS RELATED TO ONE OF THE 9412 05:58:38,262 --> 05:58:40,331 QUESTIONS, NOT ALL PATIENTS ARE 9413 05:58:40,331 --> 05:58:41,432 RESPONDING THE SAME WAY TO THE 9414 05:58:41,432 --> 05:58:45,169 SAME TYPE OF MICROBIOTA TARGETED 9415 05:58:45,169 --> 05:58:46,337 APPROACH, THAT WAS VERY CLEAR IN 9416 05:58:46,337 --> 05:58:49,140 THE CASE OF FMT INITIAL TRIALS 9417 05:58:49,140 --> 05:58:52,076 THAT MAISHTS IT WE SEE, SAME 9418 05:58:52,076 --> 05:58:53,944 DONOR, MICROBIOTA, SOME 9419 05:58:53,944 --> 05:58:56,781 RESPONDED BUT SOME DIDN'T. 9420 05:58:56,781 --> 05:58:58,115 THE PATIENTS THAT WE SEE. 9421 05:58:58,115 --> 05:58:59,950 THAT REALLY TRIGGERED US TO 9422 05:58:59,950 --> 05:59:02,486 WONDER, DO YOU ATUMORS RESPOND 9423 05:59:02,486 --> 05:59:06,424 EQUALLY TO THE SAME MICROBIOTA 9424 05:59:06,424 --> 05:59:07,525 TARGETED THERAPY? 9425 05:59:07,525 --> 05:59:09,560 TO ADDRESS THIS QUESTION, WHAT 9426 05:59:09,560 --> 05:59:10,695 KIM IS CURRENTLY DOING IN THE 9427 05:59:10,695 --> 05:59:13,731 LAB IS HE WENT BACK TO THESE 9428 05:59:13,731 --> 05:59:15,800 FIBER DIET THAT HAD SUCH A 9429 05:59:15,800 --> 05:59:17,768 STRONG RESPONSE IN THE MODELS WE 9430 05:59:17,768 --> 05:59:20,938 HAD LOOKED AT AT THE TIME AND 9431 05:59:20,938 --> 05:59:23,774 TESTED 16 DIFFERENT MODELS, 9432 05:59:23,774 --> 05:59:25,976 INCLUDING IMPLANTED 9433 05:59:25,976 --> 05:59:27,845 SUBCUTANEOUSLY IMPLANTED AND 9434 05:59:27,845 --> 05:59:28,946 SPONTANEOUS TUMOR MODELS AND YOU 9435 05:59:28,946 --> 05:59:33,984 CAN SEE IN THE GRAPHS AGAIN IF 9436 05:59:33,984 --> 05:59:36,620 WE GO ABOVE ZERO, DETRIMENTAL 9437 05:59:36,620 --> 05:59:41,325 EFFECT, BELOW ZERO -- CERTAIN 9438 05:59:41,325 --> 05:59:45,062 TUMORS MAMMARY CARCINOMA SRKSZ 9439 05:59:45,062 --> 05:59:46,363 HAVE ACTUALLY NO RESPONSE TO THE 9440 05:59:46,363 --> 05:59:47,998 DIET OR THEY ACTUALLY MAKE IT 9441 05:59:47,998 --> 05:59:50,267 WORSE, WHILE THE VAST MAJORITY 9442 05:59:50,267 --> 05:59:53,671 OF MELANOMAS ACTUALLY DO 9443 05:59:53,671 --> 05:59:54,071 RESPOND. 9444 05:59:54,071 --> 05:59:57,508 BUT THEY DO SO AT VARYING 9445 05:59:57,508 --> 05:59:57,742 DEGREES. 9446 05:59:57,742 --> 06:00:02,780 SO WE CAN STRATIFY THESE TUMOR 9447 06:00:02,780 --> 06:00:04,515 AS NONRESPONDERS, PARTIAL 9448 06:00:04,515 --> 06:00:08,986 RESPONDERS OR EXCEPTIONAL 9449 06:00:08,986 --> 06:00:09,286 RESPONDERS. 9450 06:00:09,286 --> 06:00:10,855 WHAT IS THANK OR WHAT IS 9451 06:00:10,855 --> 06:00:11,722 AFFECTING ALL OF THESE AND ONE 9452 06:00:11,722 --> 06:00:13,691 OF THE MAIN THINGS THAT COME TO 9453 06:00:13,691 --> 06:00:16,527 MIND ARE HOST FACTORS, BUT WHAT 9454 06:00:16,527 --> 06:00:17,795 KIM DID TO ADDRESS THAT QUESTION 9455 06:00:17,795 --> 06:00:20,564 WAS HE TOOK ANIMALS THAT FED THE 9456 06:00:20,564 --> 06:00:22,666 DIET AND THEN HE IMPLANTED IN 9457 06:00:22,666 --> 06:00:25,402 THE SAME ANIMAL CONTRALATERALLY 9458 06:00:25,402 --> 06:00:28,038 EITHER AN EXCEPTIONAL RESPONDER 9459 06:00:28,038 --> 06:00:30,207 MELANOMA OR IN THE OPPOSITE SIDE 9460 06:00:30,207 --> 06:00:34,044 A MAMMARY CARCINOMA IN THAT 9461 06:00:34,044 --> 06:00:35,279 DOESN'T RESPOND. 9462 06:00:35,279 --> 06:00:36,247 WHAT YOU'RE LOOKING AT IN THIS 9463 06:00:36,247 --> 06:00:38,315 GRAPH HERE IS THAT EVEN WHEN 9464 06:00:38,315 --> 06:00:41,519 IMPLANTED INTO THE SAME HOST, 9465 06:00:41,519 --> 06:00:43,921 THE EFFECT OF FIBER IS 9466 06:00:43,921 --> 06:00:45,689 MAINTAINED, RESPONDERS ARE STILL 9467 06:00:45,689 --> 06:00:48,392 RESPONDER AND NONRESPONDERS ARE 9468 06:00:48,392 --> 06:00:50,127 NONRESPONDERS, WHICH MEANS THERE 9469 06:00:50,127 --> 06:00:52,763 IS CLEARLY A TUMOR INTRINSIC 9470 06:00:52,763 --> 06:00:58,235 EFFECT THAT SOMEHOW IS 9471 06:00:58,235 --> 06:01:01,872 CONTROLLING -- SO WHAT ARE THESE 9472 06:01:01,872 --> 06:01:02,106 FACTORS? 9473 06:01:02,106 --> 06:01:03,941 THESE ARE THE THINGS WE ARE 9474 06:01:03,941 --> 06:01:04,241 ADDRESSING. 9475 06:01:04,241 --> 06:01:05,676 I DON'T HAVE THE ANSWERS YET. 9476 06:01:05,676 --> 06:01:06,944 THIS IS WHAT WE'RE TRYING TO 9477 06:01:06,944 --> 06:01:08,212 FIGURE OUT. 9478 06:01:08,212 --> 06:01:10,714 BUT WE KNOW TUMOR TYPE MAKES A 9479 06:01:10,714 --> 06:01:11,015 DIFFERENCE. 9480 06:01:11,015 --> 06:01:14,318 WE KNOW THAT TUMOR IN 9481 06:01:14,318 --> 06:01:15,653 MICROENVIRONMENT IS VERY 9482 06:01:15,653 --> 06:01:17,521 IMPORTANT, WE SEE CHANGES IN 9483 06:01:17,521 --> 06:01:18,489 RESPONDER TUMORS THAT DO NOT 9484 06:01:18,489 --> 06:01:20,891 HAPPEN IN THE NONRESPONDER 9485 06:01:20,891 --> 06:01:21,125 TUMORS. 9486 06:01:21,125 --> 06:01:23,327 WE HAVE NOW PRIMARILY DATA THAT 9487 06:01:23,327 --> 06:01:24,728 SUGGESTS TUMOR METABOLISM MAY BE 9488 06:01:24,728 --> 06:01:28,532 A FACTOR AS WELL AND TUMORLY 9489 06:01:28,532 --> 06:01:28,899 IMMUNOGENICITY. 9490 06:01:28,899 --> 06:01:31,969 I'LL SHOW YOU A LITTLE BIT OF 9491 06:01:31,969 --> 06:01:35,773 WHAT WE HAVE WITH TUMOR 9492 06:01:35,773 --> 06:01:36,574 IMMUNOGENICITY, BECAUSE I THINK 9493 06:01:36,574 --> 06:01:37,174 IT'S IMPORTANT. 9494 06:01:37,174 --> 06:01:39,944 IT FAVORS THE RESPONSE, SO IT 9495 06:01:39,944 --> 06:01:42,546 ENCOMPASSES THE RESPONSE OF NOT 9496 06:01:42,546 --> 06:01:43,214 ALREADY RESPONDER TUMOR WHAVMENT 9497 06:01:43,214 --> 06:01:44,815 I MEAN WI THAT IS IF WE TAKE ONE 9498 06:01:44,815 --> 06:01:46,383 OF THE MELANOMAS THAT WE 9499 06:01:46,383 --> 06:01:48,252 CLASSIFY AS A PARTIAL RESPONDER 9500 06:01:48,252 --> 06:01:51,522 AND NOW WE ARTIFICIALLY INCREASE 9501 06:01:51,522 --> 06:01:53,557 THE IMMUNOGENICITY BY FOR 9502 06:01:53,557 --> 06:01:59,997 EXAMPLE OVEREXPRESSING VITAMIN 9503 06:01:59,997 --> 06:02:01,498 HIGHLILY -- IMMUNOGENIC TO MICE, 9504 06:02:01,498 --> 06:02:04,034 IF WE DO THAT WITH MAMMARY 9505 06:02:04,034 --> 06:02:05,269 CARCINOMAS THAT DON'TS 9506 06:02:05,269 --> 06:02:05,803 RERESPONSIBILITY, NOTHING 9507 06:02:05,803 --> 06:02:06,136 CHANGES. 9508 06:02:06,136 --> 06:02:08,472 THIS MAKES SENSE THAT INCREASING 9509 06:02:08,472 --> 06:02:09,006 IMMUNOGENICITY CONTRIBUTES 9510 06:02:09,006 --> 06:02:10,074 BECAUSE SOMETHING I DIDN'T SHOW 9511 06:02:10,074 --> 06:02:12,276 YOU IS THAT -- SORRY, THE 9512 06:02:12,276 --> 06:02:15,379 CHANGES THAT WE SEE IN THE TUMOR 9513 06:02:15,379 --> 06:02:16,447 MICROENVIRONMENT OF THOSE TUMORS 9514 06:02:16,447 --> 06:02:19,283 THAT BENEFIT OF THE FIBER EE 9515 06:02:19,283 --> 06:02:21,252 EFFECT ARE THAT DOWNSTREAM ALL 9516 06:02:21,252 --> 06:02:22,786 THESE INNATE PATHWAY THAT I 9517 06:02:22,786 --> 06:02:24,989 SHOWED YOU EARLIER AND OF COURSE 9518 06:02:24,989 --> 06:02:27,057 RELATED TO THE DENDRITIC CELLS 9519 06:02:27,057 --> 06:02:29,059 THAT WE SEE, WE SEE CHANGE IN 9520 06:02:29,059 --> 06:02:30,794 THE T CELL COMPARTMENT PARTMENT 9521 06:02:30,794 --> 06:02:36,934 AS WELL WHICH ARE A REDUCTION IN 9522 06:02:36,934 --> 06:02:38,836 RECALL RM T CELLS AND UP 9523 06:02:38,836 --> 06:02:41,105 REGULATION AND ENHANCED, SEE 9524 06:02:41,105 --> 06:02:42,740 REDUCTION IN T REGULATORY CELLS 9525 06:02:42,740 --> 06:02:45,409 AND MORE CD4 ACTIVATED T CELLS 9526 06:02:45,409 --> 06:02:47,244 WHICH SETS REALLY A VERY NICE 9527 06:02:47,244 --> 06:02:49,346 STAGE FOR A GOOD RESPONSE TO AN 9528 06:02:49,346 --> 06:02:52,383 IMMUNE CHECK BLOCKADE SUCH AS 9529 06:02:52,383 --> 06:02:53,484 EPITHELIAL 1 THAT I SHOWED YOU 9530 06:02:53,484 --> 06:02:55,452 EARLY HAD SYNERGISTIC EFFECT AND 9531 06:02:55,452 --> 06:02:57,521 WHAT'S BEING SHOWN IN THE CLINIC 9532 06:02:57,521 --> 06:02:59,823 IN OBSERVATION STUDIES WITH 9533 06:02:59,823 --> 06:03:00,824 FIBER AND BLOCKADE. 9534 06:03:00,824 --> 06:03:01,992 WHAT ABOUT OTHER THERAPIES? 9535 06:03:01,992 --> 06:03:03,794 NOW WE GO BACK TO THE PREVIOUS 9536 06:03:03,794 --> 06:03:04,128 CONVERSATION. 9537 06:03:04,128 --> 06:03:06,931 WHAT HAPPENS TO PATIENTS THAT 9538 06:03:06,931 --> 06:03:09,566 ARE RECEIVING CHEMOTHERAPY? 9539 06:03:09,566 --> 06:03:11,302 THESE PATIENTS I ALWAYS ALMOST 9540 06:03:11,302 --> 06:03:15,873 INEVITABLY TREATED WITH -- 9541 06:03:15,873 --> 06:03:17,608 THESE PATIENTS ARE ALMOST 9542 06:03:17,608 --> 06:03:20,177 INEVITABLY TREATED WITH 9543 06:03:20,177 --> 06:03:20,945 ANTIBIOTICS. 9544 06:03:20,945 --> 06:03:22,579 CHECKPOINT, I APOLOGIZE, IT'S A 9545 06:03:22,579 --> 06:03:23,681 REALITY WHAVMENT WE SHOWED IN 9546 06:03:23,681 --> 06:03:24,815 OUR ORIGINAL STUDIES TEN YEARS 9547 06:03:24,815 --> 06:03:28,085 AGO WAS THAT MICROBIOTA ALSO 9548 06:03:28,085 --> 06:03:29,553 INFLUENCES RESPONSE TO CHEMO, 9549 06:03:29,553 --> 06:03:31,789 FOR EXAMPLE, PLATINUM BASED I AM 9550 06:03:31,789 --> 06:03:33,657 FLOW THERAPIES, SO FOR EXAMPLE 9551 06:03:33,657 --> 06:03:36,193 ANIMALS THAT LACK MICROBIOTA DO 9552 06:03:36,193 --> 06:03:41,198 NOT RESPOND TO OXALIPLATIN, 9553 06:03:41,198 --> 06:03:43,634 THESE DRUGS AND MANY OTHER 9554 06:03:43,634 --> 06:03:46,070 CHEMOS NEED IS HIGH ACTIVATION 9555 06:03:46,070 --> 06:03:48,238 OF REACTIVATION SPECIES AND WE 9556 06:03:48,238 --> 06:03:49,740 SHOWED AT THE TIME THAT IN 9557 06:03:49,740 --> 06:03:52,476 ABSENCE OF MICROBIOTA THERE WAS 9558 06:03:52,476 --> 06:03:54,611 AN IMPAIRED PRODUCTION OF ROS IN 9559 06:03:54,611 --> 06:03:56,647 THE TUMOR AND HOST IMMUNE CELLS 9560 06:03:56,647 --> 06:03:58,148 WERE NEEDED FOR THE RESPONSE F 9561 06:03:58,148 --> 06:04:00,818 WE GO BACK TO THE TME AND THE 9562 06:04:00,818 --> 06:04:02,653 MINOR COMPARTMENT WHO ARE 9563 06:04:02,653 --> 06:04:04,188 AMAZING HOST PRODUCERS WE CAN 9564 06:04:04,188 --> 06:04:07,358 SEE IN THE TME NEUTROPHILS. 9565 06:04:07,358 --> 06:04:08,459 ROS PRODUCERS. 9566 06:04:08,459 --> 06:04:10,427 SO THE ROLE OF NEUTROPHILS IN 9567 06:04:10,427 --> 06:04:12,963 RESPONSE TO CHEMOTHERAPY, WORK 9568 06:04:12,963 --> 06:04:15,332 LED BY ROMEO R RUSSIA IN THE LAB 9569 06:04:15,332 --> 06:04:17,768 AND SHOWED THAT THE NEUTROPHIL 9570 06:04:17,768 --> 06:04:20,704 IS INDEED THE MAIN SOURCE OF ROS 9571 06:04:20,704 --> 06:04:22,139 IN RESPONSE AND THEY'RE ACTUALLY 9572 06:04:22,139 --> 06:04:24,441 NECESSARY FOR THERAPEUTIC 9573 06:04:24,441 --> 06:04:24,808 EFFICACY. 9574 06:04:24,808 --> 06:04:25,976 BUT FURTHERMORE, WHAT WE FOUND 9575 06:04:25,976 --> 06:04:28,379 WAS THAT MICROBIOTA REGULATES 9576 06:04:28,379 --> 06:04:30,147 THEIR QUANTITY AND THEIR QUALITY 9577 06:04:30,147 --> 06:04:32,549 IN THE TME AND THAT THE CHANGES 9578 06:04:32,549 --> 06:04:35,085 THAT ARE INDUCED BY CHEMO, WHICH 9579 06:04:35,085 --> 06:04:37,488 IS A REPROGRAMING OF THE 9580 06:04:37,488 --> 06:04:39,890 NEUTROPHILS THAT BECOME MORE 9581 06:04:39,890 --> 06:04:40,758 ANTI-TUMORGENIC AND PRODUCE 9582 06:04:40,758 --> 06:04:42,960 BUCKETS OF ROS LEADING TO AN 9583 06:04:42,960 --> 06:04:44,528 EFFECTIVE RESPONSE IS IMPAIRED 9584 06:04:44,528 --> 06:04:49,933 WHEN WE DON'T HAVE MICROBIOTA. 9585 06:04:49,933 --> 06:04:51,135 SO WE'RE GOING BACK TO THE 9586 06:04:51,135 --> 06:04:52,870 BIGGEST QUESTIONS, HOW, AND IS 9587 06:04:52,870 --> 06:04:54,271 THAT THE SAME MECHANISM AT THAT 9588 06:04:54,271 --> 06:04:57,808 I SHOWED BEFORE, THE OTHER ONE 9589 06:04:57,808 --> 06:05:03,280 WAS VERY -- AND AGAIN, I'M GOING 9590 06:05:03,280 --> 06:05:06,583 TO MAKE IT SHORT, BUT WE KNEW 9591 06:05:06,583 --> 06:05:11,422 THAT MICROBIOTA REGULATES 9592 06:05:11,422 --> 06:05:12,389 NEUTROPHIL ANTIMICROBIAL 9593 06:05:12,389 --> 06:05:12,656 FUNCTIONS. 9594 06:05:12,656 --> 06:05:15,325 WE ALSO KNEW THAT IT REGULATES 9595 06:05:15,325 --> 06:05:17,394 ROS PRODUCTION BY MACROPHAGES 9596 06:05:17,394 --> 06:05:22,199 AND THIS WAS THE ACTIVATION OF 9597 06:05:22,199 --> 06:05:25,169 THESE NOD1 RECEPTOR, WE HAD A 9598 06:05:25,169 --> 06:05:28,238 SIMPLE HYPOTHESIS, DIST SAME 9599 06:05:28,238 --> 06:05:31,075 PATHWAY REGULATE ROS PRODUCTION 9600 06:05:31,075 --> 06:05:34,278 IN NEUTROPHILS IN TUMOR 9601 06:05:34,278 --> 06:05:35,045 MICROENVIRONMENT, WE TESTED 9602 06:05:35,045 --> 06:05:37,114 THAT, AS I TOLD YOU EARLIER IF 9603 06:05:37,114 --> 06:05:38,982 THERE'S NO MICROBIOTA THE ROS 9604 06:05:38,982 --> 06:05:41,618 PRODUCTION STLINLT, WE GAVE 9605 06:05:41,618 --> 06:05:43,587 LIGAND AND THERE WAS SLEURTLY NO 9606 06:05:43,587 --> 06:05:43,921 EFFECT. 9607 06:05:43,921 --> 06:05:48,625 HOWEVER, WHEN WE GAVE THEM A 9608 06:05:48,625 --> 06:05:50,494 DIFFERENT NDP THAT BINDS TO A 9609 06:05:50,494 --> 06:05:52,329 DIFFERENT RECEPTOR NOT TO 9610 06:05:52,329 --> 06:05:53,363 COMPLETELY RESTORE NEUTROPHIL 9611 06:05:53,363 --> 06:05:55,199 FUNCTION IT WASN'T JUST A 9612 06:05:55,199 --> 06:05:56,500 FUNCTION, IT'S RESTORED THE 9613 06:05:56,500 --> 06:05:59,570 WHOLE DYNAMICS OF NEUTROPHILS 9614 06:05:59,570 --> 06:06:02,706 AND WAS ABSENCE OF MICROBIAL BUT 9615 06:06:02,706 --> 06:06:04,942 MOST IMPORTANTLY IT RESTORED THE 9616 06:06:04,942 --> 06:06:06,343 ABILITY OF THESE ANIMALS TO 9617 06:06:06,343 --> 06:06:09,546 RESPOND TO TREATMENT. 9618 06:06:09,546 --> 06:06:11,381 SO BASICALLY WE HAVE A SCENARIO 9619 06:06:11,381 --> 06:06:13,784 WHERE NOW WE HAVE CHEMOTHERAPY, 9620 06:06:13,784 --> 06:06:15,752 IT ENGAGES ALTHOUGH VERY 9621 06:06:15,752 --> 06:06:18,288 DIFFERENT CELLULAR SEQUENCE, 9622 06:06:18,288 --> 06:06:20,357 NEUTROPHILS ARE IMPORTANT AND IF 9623 06:06:20,357 --> 06:06:22,459 WE HAVE MICROBIAL SIGNALS THESE 9624 06:06:22,459 --> 06:06:24,228 NEUTROPHILS ARE GOING TO RESPOND 9625 06:06:24,228 --> 06:06:26,930 PRODUCE ROS AND LEAD TO TUMOR 9626 06:06:26,930 --> 06:06:27,231 ERADICATION. 9627 06:06:27,231 --> 06:06:31,902 IF NOT, THEY WILL BECOME -- 9628 06:06:31,902 --> 06:06:33,704 FNLT, WE'LL FAIL TO TREAT THESE 9629 06:06:33,704 --> 06:06:35,038 TUMORS, BUT I JUST SHOWED YOU 9630 06:06:35,038 --> 06:06:37,541 THAT WE IDENTIFIED MDP THAT CAN 9631 06:06:37,541 --> 06:06:39,610 REVERT THIS SCENARIO. 9632 06:06:39,610 --> 06:06:41,478 AND THIS IS VERY IMPORTANT IN 9633 06:06:41,478 --> 06:06:43,780 THE CLINIC FOR PATIENTS 9634 06:06:43,780 --> 06:06:44,882 RECEIVING CHEMO BECAUSE AS 9635 06:06:44,882 --> 06:06:48,118 MARCEL JUST MENTIONED, THESE 9636 06:06:48,118 --> 06:06:49,887 PATIENTS BECOME NEUTROPOENIC, 9637 06:06:49,887 --> 06:06:51,455 DEVELOP FEVERS AND THERE THEY GO 9638 06:06:51,455 --> 06:06:54,625 TO RECEIVE THEIR ANTIBIOTIC 9639 06:06:54,625 --> 06:06:55,058 COCKTAIL. 9640 06:06:55,058 --> 06:06:56,927 SO AS NEUTROPHILS ARE COMING 9641 06:06:56,927 --> 06:07:00,330 BACK, THEY ARE DOING SO IN AN 9642 06:07:00,330 --> 06:07:01,965 ENVIRONMENT WITH AN ALTERED 9643 06:07:01,965 --> 06:07:03,066 MICROBIOTA BECAUSE OF THE 9644 06:07:03,066 --> 06:07:04,034 ANTIBIOTIC TREATMENT. 9645 06:07:04,034 --> 06:07:06,069 SO IF WE HAVE A MOLECULE THAT WE 9646 06:07:06,069 --> 06:07:08,972 CAN USE TO REEDUCATE THESE 9647 06:07:08,972 --> 06:07:10,140 NEUTROPHILS AS THEY COME BACK 9648 06:07:10,140 --> 06:07:11,808 AND GET READY FOR THE NEXT DOSE 9649 06:07:11,808 --> 06:07:14,411 OF CHEMO, THAT CAN MAKE A BIG 9650 06:07:14,411 --> 06:07:15,746 DIFFERENCE IN TERMS OF 9651 06:07:15,746 --> 06:07:17,047 THERAPEUTIC RESPONSE. 9652 06:07:17,047 --> 06:07:18,348 AND WHAT IS EVEN BETTER ABOUT 9653 06:07:18,348 --> 06:07:22,586 THIS IS THAT THERE ARE MANY MVP 9654 06:07:22,586 --> 06:07:24,154 SYNTHETIC AN ANALOGS THAT HAVE 9655 06:07:24,154 --> 06:07:25,622 ALREADY BEEN DEEMED SAFE IN THE 9656 06:07:25,622 --> 06:07:28,659 CLINIC AND ARE USING DIFFERENT 9657 06:07:28,659 --> 06:07:29,760 SETTINGS, SOME INCLUDING 9658 06:07:29,760 --> 06:07:30,727 ACTUALLY CANCER. 9659 06:07:30,727 --> 06:07:32,629 SO I THINK THAT IT SETS REALLY 9660 06:07:32,629 --> 06:07:34,031 GIVES THE RATIONALE TO START 9661 06:07:34,031 --> 06:07:35,566 THINKING ABOUT CLINICAL TRIALS 9662 06:07:35,566 --> 06:07:37,901 OR SOMETHING LIKE THIS WHERE IT 9663 06:07:37,901 --> 06:07:39,503 COULD BE DONE FOR CHEMO. 9664 06:07:39,503 --> 06:07:42,239 I'M FINISHING WITH THE LAST 9665 06:07:42,239 --> 06:07:42,773 SLIDE. 9666 06:07:42,773 --> 06:07:44,007 WHAT I WANT TO YOU TAKE HOME AS 9667 06:07:44,007 --> 06:07:47,477 A BIG MESSAGE FROM THIS IS, YES, 9668 06:07:47,477 --> 06:07:48,879 MICROBIOTA PROGRAMS THE TME FOR 9669 06:07:48,879 --> 06:07:51,215 A GOOD RESPONSE, BUT AND IT'S A 9670 06:07:51,215 --> 06:07:53,617 BIG BUT, NOT ALL TUMORS RESPOND 9671 06:07:53,617 --> 06:07:56,420 EQUALLY TO THE SAME MICROBIOTA 9672 06:07:56,420 --> 06:07:58,855 TARGETED THERAPY, BOTH HOST AND 9673 06:07:58,855 --> 06:08:00,324 TUMOR INTRINSIC FACTORS 9674 06:08:00,324 --> 06:08:01,525 INFLUENCE THIS RESPONSE, AND 9675 06:08:01,525 --> 06:08:04,795 DIFFERENT THERAPIES, DIFFERENT 9676 06:08:04,795 --> 06:08:06,997 CELLULAR CIRCUITS AND DIFFERENT 9677 06:08:06,997 --> 06:08:09,266 MICROBIOTA MEDIATED MECHANISMS. 9678 06:08:09,266 --> 06:08:12,869 THE ONLY WAY WE CAN COME UP WITH 9679 06:08:12,869 --> 06:08:14,071 MEANINGFUL ACTIONABLE APPROACHES 9680 06:08:14,071 --> 06:08:16,340 IS IF WE REALLY UNDERSTAND HOW 9681 06:08:16,340 --> 06:08:19,243 ALL OF THIS IS HAPPENING, SO 9682 06:08:19,243 --> 06:08:20,844 THAT WE CAN BYPASS A LOT OF WHAT 9683 06:08:20,844 --> 06:08:23,247 IS UPSTREAM AND HAVE SOMETHING 9684 06:08:23,247 --> 06:08:25,849 WE CAN USE AS A DRIVER DIRECTLY 9685 06:08:25,849 --> 06:08:27,551 AND I CAN'T EMPHASIZE THAT 9686 06:08:27,551 --> 06:08:27,851 ENOUGH. 9687 06:08:27,851 --> 06:08:29,553 AND I'M GOING TO FINISH, I 9688 06:08:29,553 --> 06:08:34,124 ALREADY MENTIONED KIM, I WANT TO 9689 06:08:34,124 --> 06:08:35,058 THANK EVERYBODY IN THE LAB, 9690 06:08:35,058 --> 06:08:35,959 SMALL GROUP, EVERYBODY 9691 06:08:35,959 --> 06:08:37,594 CONTRIBUTES RG ALL THE 9692 06:08:37,594 --> 06:08:39,263 COLLABORATORS WITHIN AND OUTSIDE 9693 06:08:39,263 --> 06:08:40,230 NIH AND MANY COURSE AND 9694 06:08:40,230 --> 06:08:41,798 FACILITIES THAT WE HAVE ACCESS 9695 06:08:41,798 --> 06:08:43,634 HERE THAT MAKE ALL OF THIS 9696 06:08:43,634 --> 06:08:43,900 POSSIBLE. 9697 06:08:43,900 --> 06:08:44,768 I THANK YOU FOR YOUR ATTENTION 9698 06:08:44,768 --> 06:08:46,603 AND I LOOK FORWARD TO YOUR 9699 06:08:46,603 --> 06:08:48,672 QUESTIONS. 9700 06:08:48,672 --> 06:08:53,610 [APPLAUSE] 9701 06:08:53,610 --> 06:08:55,445 >> VERY NICE TALK. 9702 06:08:55,445 --> 06:08:58,181 SO YOUR HIGH FIBER DIET MODEL, 9703 06:08:58,181 --> 06:09:04,321 SO YOU MENTIONED THAT XL IS 1 IS 9704 06:09:04,321 --> 06:09:05,088 INCREASED, RIGHT? 9705 06:09:05,088 --> 06:09:07,591 SO JUST WONDERING, THE OTHER 9706 06:09:07,591 --> 06:09:09,993 CHEMOKINE FUNCTION YOUR MODELS 9707 06:09:09,993 --> 06:09:12,729 CCR5 HAVE YOU ALSO CHECKED THAT? 9708 06:09:12,729 --> 06:09:14,164 AND I'M JUST WONDERING IF THAT 9709 06:09:14,164 --> 06:09:15,032 IS SPECIFICALLY. 9710 06:09:15,032 --> 06:09:16,867 >> YES, ALSO INCREASED AND IT'S 9711 06:09:16,867 --> 06:09:18,869 ALSO PRIMARILY PRODUCED BY 9712 06:09:18,869 --> 06:09:20,837 ENCASING THE TME ALSO T CELLS 9713 06:09:20,837 --> 06:09:22,439 ALSO PRODUCE IT BUT IT'S 9714 06:09:22,439 --> 06:09:23,106 INCREASED IN ALL. 9715 06:09:23,106 --> 06:09:29,846 >> OKAY, THANK YOU. 9716 06:09:29,846 --> 06:09:31,381 >> YOUNG PEOPLE FIRST. 9717 06:09:31,381 --> 06:09:36,286 >> GREAT TALK. 9718 06:09:36,286 --> 06:09:37,587 I APPRECIATE THAT. 9719 06:09:37,587 --> 06:09:42,959 I'M A POST-DOC IN -- LAB. 9720 06:09:42,959 --> 06:09:45,362 MICROBIOME AFFECTING NEUTROPHILS 9721 06:09:45,362 --> 06:09:46,997 IN TUMOR ENVIRONMENT, WHY IS IT 9722 06:09:46,997 --> 06:09:48,332 HAPPENING ONLY IN THE TUMOR 9723 06:09:48,332 --> 06:09:48,632 ENVIRONMENT? 9724 06:09:48,632 --> 06:09:49,633 GROO THAT'S A VERY GOOD 9725 06:09:49,633 --> 06:09:50,500 QUESTION, AND ONE THAT I DON'T 9726 06:09:50,500 --> 06:09:52,903 HAVE AN ANSWER TO YES. 9727 06:09:52,903 --> 06:09:55,439 BUT WE DON'T KNOW WHERE EXACTLY 9728 06:09:55,439 --> 06:09:58,508 P THE EDUCATION ACTIVATION 9729 06:09:58,508 --> 06:09:59,576 WHATEVER NAME YOU'RE GOING TO 9730 06:09:59,576 --> 06:10:01,011 GIVE IT OF COURSE WE'RE LOOK 9731 06:10:01,011 --> 06:10:03,747 INTUG T I'M HOPING ONE DAY WE'LL 9732 06:10:03,747 --> 06:10:05,515 KNOW. 9733 06:10:05,515 --> 06:10:06,550 BUT I DON'T THINK, IT MIGHT BE 9734 06:10:06,550 --> 06:10:08,552 MORE THAN ONE PLACE. 9735 06:10:08,552 --> 06:10:09,653 LET'S LEAVE IT LIKE THAT. 9736 06:10:09,653 --> 06:10:11,421 >> IN YOUR STUDIES EVER SEE 9737 06:10:11,421 --> 06:10:14,157 AFTER ADDING THE DRUG THAT 9738 06:10:14,157 --> 06:10:17,327 ACTIVATES, LIKE MDV IS THERE 9739 06:10:17,327 --> 06:10:18,829 ANNUITY FILLS THAT COULD 9740 06:10:18,829 --> 06:10:20,364 POTENTIALLY CAUSE DAMAGE TO THE 9741 06:10:20,364 --> 06:10:20,564 BODY? 9742 06:10:20,564 --> 06:10:22,366 >> I DON'T THINK THAT WE LOOKED 9743 06:10:22,366 --> 06:10:23,166 IN PARTICULAR. 9744 06:10:23,166 --> 06:10:25,302 I BELIEVE WITH MDP IT WAS MOSTLY 9745 06:10:25,302 --> 06:10:27,637 LOOKED AT IN THE TUMOR, ROMEO, I 9746 06:10:27,637 --> 06:10:29,005 DON'T KNOW IF EUROPE IN THE 9747 06:10:29,005 --> 06:10:30,974 AUDIENCE, IF YOU LOOKED IN 9748 06:10:30,974 --> 06:10:31,274 CIRCULATION. 9749 06:10:31,274 --> 06:10:32,909 I THINK TELL YOU SHE'S SAYING NO 9750 06:10:32,909 --> 06:10:35,145 FROM THE BACK, I CAN TELL YOU 9751 06:10:35,145 --> 06:10:36,880 THAT THE CHANGES WE SEE IN THE 9752 06:10:36,880 --> 06:10:38,749 TUMOR IN THE NEUTROPHILS AFTER 9753 06:10:38,749 --> 06:10:39,950 TREATMENT FOR EXAMPLE WE DO NOT 9754 06:10:39,950 --> 06:10:41,885 SEE THOSE IN BLOOD. 9755 06:10:41,885 --> 06:10:43,019 WHAT HAPPENS IN THE TUMOR 9756 06:10:43,019 --> 06:10:44,855 HAPPENS IN THE TUMOR, IS NOT 9757 06:10:44,855 --> 06:10:47,624 REFLECTED IN CIRCULATION. 9758 06:10:47,624 --> 06:10:53,230 THAT'S ANOTHER -- 9759 06:10:53,230 --> 06:10:54,164 >> GREAT TALK. 9760 06:10:54,164 --> 06:10:55,332 MY NAME SUSPECT NATALIE. 9761 06:10:55,332 --> 06:10:56,266 CAN YOU HEAR ME? 9762 06:10:56,266 --> 06:10:56,600 >> YEAH. 9763 06:10:56,600 --> 06:10:59,136 >> GREAT. 9764 06:10:59,136 --> 06:11:01,037 >> MY NAME IS NATALIE. 9765 06:11:01,037 --> 06:11:02,272 HE ASKED MY FIRST QUESTION WHICH 9766 06:11:02,272 --> 06:11:03,774 IS ABOUT WHERE THAT PRIMING IS 9767 06:11:03,774 --> 06:11:05,075 ACTUALLY TAKING PLACE. 9768 06:11:05,075 --> 06:11:08,578 BUT THEN THITION SORT -- BUT 9769 06:11:08,578 --> 06:11:09,780 THEN I GUESS SORT OF SIMILARLY 9770 06:11:09,780 --> 06:11:11,081 TO THAT QUERKS I WANTED TO ASK, 9771 06:11:11,081 --> 06:11:13,850 YOU LOOKED AT MAMMARY GLAND AND 9772 06:11:13,850 --> 06:11:15,385 MELANOMA AND OTHER MODELS AS 9773 06:11:15,385 --> 06:11:16,553 WELL BUT I WANTED TO ASK IF 9774 06:11:16,553 --> 06:11:18,188 YOU'VE LOOKED AT OTHER MODEL 9775 06:11:18,188 --> 06:11:20,257 SYSTEMS THAT HAD A LARGER SORT 9776 06:11:20,257 --> 06:11:23,593 OF LIKE INTERFACE WITH 9777 06:11:23,593 --> 06:11:26,663 MICROBIOTA IN GENERAL, LIKE 9778 06:11:26,663 --> 06:11:28,932 INTESTINAL OR LIVER FOR EXAMPLE. 9779 06:11:28,932 --> 06:11:33,203 >> YEAH, NO, WE IT USED 9780 06:11:33,203 --> 06:11:36,873 CANCER -- CANCER BUT THAT WAS 9781 06:11:36,873 --> 06:11:38,074 SUBCUTANEOUSLY IMPLANTED, WE 9782 06:11:38,074 --> 06:11:39,576 HAVE NOT LOOKED IN THE GUT, MAIN 9783 06:11:39,576 --> 06:11:41,344 FOCUS IS LOOKING AT SYSTEM 9784 06:11:41,344 --> 06:11:42,446 SITES, WE'RE NOT REALLY LOOKING 9785 06:11:42,446 --> 06:11:43,947 AT LOCAL EFFECT. 9786 06:11:43,947 --> 06:11:45,248 BUT SOMETHING TO BE DONE. 9787 06:11:45,248 --> 06:11:46,783 >> THANK YOU. 9788 06:11:46,783 --> 06:11:53,590 >> IN THE BACK. 9789 06:11:53,590 --> 06:11:56,326 >> YOUNG PEOPLE FIRST. 9790 06:11:56,326 --> 06:11:58,295 >> YOUNG PEOPLE FIRST? 9791 06:11:58,295 --> 06:12:01,131 >> YEAH. 9792 06:12:01,131 --> 06:12:04,634 [LAUGHTER] 9793 06:12:04,634 --> 06:12:07,471 >> ONE QUESTION APIECE. 9794 06:12:07,471 --> 06:12:08,138 SOMEBODY START. 9795 06:12:08,138 --> 06:12:10,841 >> IN THE BACK. 9796 06:12:10,841 --> 06:12:12,309 >> GREAT TALK, 9797 06:12:12,309 --> 06:12:12,709 SUPER-INTERESTING. 9798 06:12:12,709 --> 06:12:16,012 I WAS WONDERING ABOUT GERM-FREE 9799 06:12:16,012 --> 06:12:17,814 EXPERIMENTS THAT WERE DONE AND 9800 06:12:17,814 --> 06:12:19,716 THE RESPONSE THAT YOU GOT. 9801 06:12:19,716 --> 06:12:23,587 HOW EARLY WOULD YOU NEED TO 9802 06:12:23,587 --> 06:12:27,524 TRANSFER THE -- OR TRANSFER THE 9803 06:12:27,524 --> 06:12:28,959 MICROBIOME TO THE GERM FREE MICE 9804 06:12:28,959 --> 06:12:31,261 IN ORDER TO GET THE ANTI-TUMOR 9805 06:12:31,261 --> 06:12:31,528 RESPONSE? 9806 06:12:31,528 --> 06:12:33,363 >> SORRY, I MISSED THE FIRST 9807 06:12:33,363 --> 06:12:33,663 PART. 9808 06:12:33,663 --> 06:12:34,231 HOW WHAT? 9809 06:12:34,231 --> 06:12:35,632 >> HOW EARLY WOULD YOU ZAISH ON 9810 06:12:35,632 --> 06:12:35,999 HOW EARLY. 9811 06:12:35,999 --> 06:12:36,733 >> YEAH. 9812 06:12:36,733 --> 06:12:39,803 >> I BELIEVE WE DID TWO WEEKS 9813 06:12:39,803 --> 06:12:40,003 PRIOR. 9814 06:12:40,003 --> 06:12:41,972 >> SO TWO WEEKS IS ENOUGH FOR 9815 06:12:41,972 --> 06:12:44,508 THE MICE TO THEN DEVELOP THE 9816 06:12:44,508 --> 06:12:45,909 ANTI-TUMOR RESPONSE? 9817 06:12:45,909 --> 06:12:47,711 >> YEAH, AND PROBABLY LESS. 9818 06:12:47,711 --> 06:12:49,312 GERM FREE MICE GET COL NIELTZED 9819 06:12:49,312 --> 06:12:52,616 VERY EASILY. 9820 06:12:52,616 --> 06:12:55,151 DON'T HAVE ANYBODY TO COMPETE 9821 06:12:55,151 --> 06:12:55,352 WITH. 9822 06:12:55,352 --> 06:12:56,820 >> I'M SORRY, I'M BEING TOLD 9823 06:12:56,820 --> 06:12:58,989 THAT WE NEED TO CUT OFF JUST 9824 06:12:58,989 --> 06:13:01,491 BECAUSE WE'RE LOSING THE ROOM 9825 06:13:01,491 --> 06:13:01,825 SOON. 9826 06:13:01,825 --> 06:13:02,726 I'M REALLY SORRY. 9827 06:13:02,726 --> 06:13:08,164 >> I'LL BE AROUND FOR MORE 9828 06:13:08,164 --> 06:13:18,341 QUESTIONS. 9829 06:13:22,279 --> 06:13:27,350 >> I'M FROM NCI, I'LL CHAIR THE 9830 06:13:27,350 --> 06:13:28,685 REMAINING SESSION, MY IMPORTANT 9831 06:13:28,685 --> 06:13:29,986 JOB THAT WE'LL GET OUT OF THE 9832 06:13:29,986 --> 06:13:31,588 ROOM BY 5, OTHERWISE WE'LL GET 9833 06:13:31,588 --> 06:13:34,224 KICKED OUT. 9834 06:13:34,224 --> 06:13:37,627 AND YOU CAN THINK THE SCREEN IS 9835 06:13:37,627 --> 06:13:38,995 FLICKERING. 9836 06:13:38,995 --> 06:13:47,504 IT IS NOT YOUR EYES. 9837 06:13:47,504 --> 06:13:51,875 (AWAY FROM MICROPHONE). 9838 06:13:51,875 --> 06:13:53,143 FROM NCI. 9839 06:13:53,143 --> 06:13:55,812 THE LONGEST THIETLE OF THE 9840 06:13:55,812 --> 06:13:59,950 CONFERENCE FOR THE TALK WILL -- 9841 06:13:59,950 --> 06:14:00,717 TITLE. 9842 06:14:00,717 --> 06:14:04,588 EXCELLENT SPEAKER. 9843 06:14:04,588 --> 06:14:04,754 OF. 9844 06:14:04,754 --> 06:14:05,422 >> ALL RIGHT. 9845 06:14:05,422 --> 06:14:06,656 CAN EVERYONE HEAR ME? 9846 06:14:06,656 --> 06:14:06,957 >> YEAH. 9847 06:14:06,957 --> 06:14:07,958 >> ALL RIGHT. 9848 06:14:07,958 --> 06:14:09,492 SO I WOULD REALLY LIKE TO THANK 9849 06:14:09,492 --> 06:14:11,895 THE ORGANIZERS THE OPPORTUNITY 9850 06:14:11,895 --> 06:14:13,563 TO PRESENTED VERY NEW FLERCH MY 9851 06:14:13,563 --> 06:14:13,730 LAB. 9852 06:14:13,730 --> 06:14:15,465 I AM SOMEONE PASSIONATE ABOUT 9853 06:14:15,465 --> 06:14:17,067 UNDERSTANDING HOW HERPESVIRUS 9854 06:14:17,067 --> 06:14:18,835 SYN EFFECT FOR LIFE AND I'M VERY 9855 06:14:18,835 --> 06:14:20,770 INTERESTED IN CANCER CAUSING 9856 06:14:20,770 --> 06:14:21,104 HERPESVIRUSES. 9857 06:14:21,104 --> 06:14:22,539 SO HERE I'M TAKING KNOWLEDGE IN 9858 06:14:22,539 --> 06:14:24,374 THE FIELD ON HOW THESE VIRUSES 9859 06:14:24,374 --> 06:14:26,476 REPLICATE AND HOW THEY ESTABLISH 9860 06:14:26,476 --> 06:14:28,011 LATENCY TO NOW MAKE MUTANT 9861 06:14:28,011 --> 06:14:29,512 VIRUSES THAT AREN'T ABLE TO DO 9862 06:14:29,512 --> 06:14:31,481 SO AND SEE IF THOSE ARE 9863 06:14:31,481 --> 06:14:34,351 EFFECTIVE VACCINES IN VIVO. 9864 06:14:34,351 --> 06:14:36,286 SO MANY OF YOU ARE AWARE THAT 9865 06:14:36,286 --> 06:14:37,954 THERE ARE MANY VIRUSES THAT ARE 9866 06:14:37,954 --> 06:14:40,023 ASSOCIATED WITH HUMAN CANCERS, 9867 06:14:40,023 --> 06:14:44,828 IN TWO OF THESE ARE CAPIC THY 9868 06:14:44,828 --> 06:14:48,131 HERPESVIRUS AND EPSTEIN-BARR 9869 06:14:48,131 --> 06:14:58,308 VIRUS, GAMMA HERPESVIRUS. 9870 06:14:58,308 --> 06:14:59,709 CAPISARCOMA, NEOPLASIA TYPICALLY 9871 06:14:59,709 --> 06:15:01,745 IN SKIN, CAN BE IN THE LUNG AND 9872 06:15:01,745 --> 06:15:03,480 GI TRACT AND EPSTEIN-BARR VIRUS 9873 06:15:03,480 --> 06:15:06,883 ASSOCIATED WITH CAUSATIVE AGENT 9874 06:15:06,883 --> 06:15:07,984 PNEUMATIC CLEEOSIS ALSO 9875 06:15:07,984 --> 06:15:10,754 ASSOCIATED WITH MANY LYMPHOMAS 9876 06:15:10,754 --> 06:15:13,256 IN ADDITION TO CANCERS OF THE 9877 06:15:13,256 --> 06:15:13,590 EPITHELIUM. 9878 06:15:13,590 --> 06:15:16,393 SO THESE VIRUSES ARE ESPECIALLY 9879 06:15:16,393 --> 06:15:18,461 IMPORTANT IN PEOPLE LIVING WITH 9880 06:15:18,461 --> 06:15:20,530 HIV WHICH WE HAVE A LARGE 9881 06:15:20,530 --> 06:15:21,531 POPULATION WORLDWIDE AND PEOPLE 9882 06:15:21,531 --> 06:15:23,366 WHO ARE NEWLY INFECTED EVERY 9883 06:15:23,366 --> 06:15:26,002 DAY, EVEN IN THOSE THAT HAVE 9884 06:15:26,002 --> 06:15:27,971 GOOD CD4 SCROWNTS A HIGHER 9885 06:15:27,971 --> 06:15:32,809 PREVALENCE OF GAMMA HERPESVIRUS 9886 06:15:32,809 --> 06:15:33,043 CANCERS. 9887 06:15:33,043 --> 06:15:34,210 A LITTLE BIT OF INFORMATION 9888 06:15:34,210 --> 06:15:35,412 ABOUT HERPESVIRUSES, THERE ARE 9889 06:15:35,412 --> 06:15:36,980 NINE OF THEM, THEY'RE BIG 9890 06:15:36,980 --> 06:15:38,448 VIRUSES, DNA VIRUSES AND PART OF 9891 06:15:38,448 --> 06:15:39,382 THE COMPLEXITY IS THEY HAVE A 9892 06:15:39,382 --> 06:15:43,319 LOT OF GENES THAT MANIPULATE THE 9893 06:15:43,319 --> 06:15:44,954 HOST, THEY ALSO HAVE NONCODING 9894 06:15:44,954 --> 06:15:46,723 RNA AND IS THEY ACTUALLY BOTH 9895 06:15:46,723 --> 06:15:48,992 REPLICATE TO MAKE NEW INFECTIOUS 9896 06:15:48,992 --> 06:15:49,893 PARTICLES, GETTING THIS STRAIGHT 9897 06:15:49,893 --> 06:15:51,361 HERE, AND THEY CAN ESTABLISH 9898 06:15:51,361 --> 06:15:52,395 LATENCY WHERE THEY'RE ACTUALLY 9899 06:15:52,395 --> 06:15:54,264 NOT EXPRESSING MANY GENES AND 9900 06:15:54,264 --> 06:15:56,132 THAT'S A GREAT WAY TO EVADE 9901 06:15:56,132 --> 06:15:57,500 IMMUNE CONTROL. 9902 06:15:57,500 --> 06:15:59,069 WE HAVE ANTI-VIRAL DRUGS BUT 9903 06:15:59,069 --> 06:16:02,238 THOSE ARE ONLY EFFECTIVE AGAINST 9904 06:16:02,238 --> 06:16:03,907 THE LYTIC PHASE NOT THE LATE 9905 06:16:03,907 --> 06:16:05,208 LATENT FADES OF HERPESVIRUS 9906 06:16:05,208 --> 06:16:06,743 INFECTIONS ASK WE HAVE VACCINES 9907 06:16:06,743 --> 06:16:08,178 AVAILABLE ONLY AGAINST ONE OF 9908 06:16:08,178 --> 06:16:16,252 THE HERPESVIRUSES, VARI TELL LA 9909 06:16:16,252 --> 06:16:16,720 ZOSTER. 9910 06:16:16,720 --> 06:16:18,354 I'VE UNDERLINED THE ONES 9911 06:16:18,354 --> 06:16:19,389 ACTUALLY BEING USED IN PEOPLE 9912 06:16:19,389 --> 06:16:22,092 RIGHT NOW, SHIN GREKS USED IN 9913 06:16:22,092 --> 06:16:23,693 OLDER INDIVIDUALS TO PREVENT 9914 06:16:23,693 --> 06:16:25,495 SHINGLES, REACTIVATION AND WE 9915 06:16:25,495 --> 06:16:28,398 HAVE ATTENUATED VIRUS ZOSTIVAX 9916 06:16:28,398 --> 06:16:30,467 USE #-D IN CHILDREN FREERCHT 9917 06:16:30,467 --> 06:16:32,836 CHICKENPOX, PRIMARY INFECTION. 9918 06:16:32,836 --> 06:16:34,170 ATTENUATED VIRUS THAT DOES 9919 06:16:34,170 --> 06:16:35,572 ESTABLISH LATENCY, DOES NOT 9920 06:16:35,572 --> 06:16:37,207 PREVENTED STERILIZING IMMUNITY 9921 06:16:37,207 --> 06:16:39,743 BUT IT DOES PREVENT REALLY 9922 06:16:39,743 --> 06:16:44,647 DETRIMENTAL CHICKENPOX 9923 06:16:44,647 --> 06:16:51,755 INFECTIONS. 9924 06:16:51,755 --> 06:16:54,557 I'M VERY EXCITED WITH A mRNA 9925 06:16:54,557 --> 06:16:58,394 BASED ARE VACCINE H ITD TO 9926 06:16:58,394 --> 06:17:03,199 MENTION THAT JEFF COHEN AT 9927 06:17:03,199 --> 06:17:06,636 NIANIAD AT NIH, INSTRUMENTAL IN 9928 06:17:06,636 --> 06:17:08,605 VACCINES IN COLLABORATION ALSO 9929 06:17:08,605 --> 06:17:11,508 IN N I AD, THOSE ARE ALSO IN 9930 06:17:11,508 --> 06:17:12,208 CLINICAL TRIALS. 9931 06:17:12,208 --> 06:17:14,377 WHAT YOU DO NOT SEE ON THIS LIST 9932 06:17:14,377 --> 06:17:18,815 IS KSHV, NOILY VACCINE IN 9933 06:17:18,815 --> 06:17:19,883 PROGRESS FOR HERPESVIRUS. 9934 06:17:19,883 --> 06:17:20,617 WHY IS THAT? 9935 06:17:20,617 --> 06:17:22,185 IT IS A LITTLE BIT BEHIND, IT'S 9936 06:17:22,185 --> 06:17:23,620 A MORE NEWLY DISCOVERED VIRUS, 9937 06:17:23,620 --> 06:17:26,890 MORE RECENTLY DISCOVERED THAN 9938 06:17:26,890 --> 06:17:28,958 EBV, WE HAVE THE PROBLEM WAWFLT 9939 06:17:28,958 --> 06:17:30,727 HERPESVIRUSES IS THERE'S NOT 9940 06:17:30,727 --> 06:17:32,595 JUST ONE RECEPTOR, THERE'S NOT 9941 06:17:32,595 --> 06:17:34,998 JUST ONE GLYCOPROTEIN THERE ARE 9942 06:17:34,998 --> 06:17:36,633 MULTIPLE GLYCOPROTEINS THAT FORM 9943 06:17:36,633 --> 06:17:38,401 COME PLEKS AND THESE 9944 06:17:38,401 --> 06:17:40,136 GLYCOPROTEINS CAN ENGAGE 9945 06:17:40,136 --> 06:17:41,437 MULTIPLE RECEPTORS FOR BOTH 9946 06:17:41,437 --> 06:17:42,338 ATTACHMENT AND ENTRY. 9947 06:17:42,338 --> 06:17:45,074 SO IT IS A COMPLICATED SCENARIO 9948 06:17:45,074 --> 06:17:47,377 TO DEVELOP NEUTRALIZING 9949 06:17:47,377 --> 06:17:49,579 ANTIBODIES AGAINST. 9950 06:17:49,579 --> 06:17:53,316 KSHD AND THE -- KSHV AND THE 9951 06:17:53,316 --> 06:17:56,319 MOUSE PATH I'LL TELL YOU ABOUT 9952 06:17:56,319 --> 06:17:59,389 TODAY SHARELY KINASES FOR ENTRY, 9953 06:17:59,389 --> 06:18:02,625 A POST DMOK MY LAB HAS DONE 9954 06:18:02,625 --> 06:18:03,660 AMAZING WORK THAT, THERE IS 9955 06:18:03,660 --> 06:18:05,895 OVERLAP THERE. 9956 06:18:05,895 --> 06:18:10,667 SO ANOTHER ASPECT OF KSHV IS UN 9957 06:18:10,667 --> 06:18:12,101 -- IS IT DOES NOT TAPE A 9958 06:18:12,101 --> 06:18:13,636 LARGE PARTED OF OUR T CELL 9959 06:18:13,636 --> 06:18:14,838 REPERTOIRE, THOSE INFECTED WE 9960 06:18:14,838 --> 06:18:16,606 DON'T SEE ANY CLEAR PATTERNS FOR 9961 06:18:16,606 --> 06:18:18,441 IMMUNE DOMINANT EPITOPES AND 9962 06:18:18,441 --> 06:18:20,143 DON'T HAVE DEFINED IMMUNOCAR 9963 06:18:20,143 --> 06:18:20,677 LATS OF PROTECTION. 9964 06:18:20,677 --> 06:18:22,478 HOW CAN WE GENERATE A SAFE 9965 06:18:22,478 --> 06:18:24,013 VACCINE WITH BROAD 9966 06:18:24,013 --> 06:18:25,582 IMMUNOGENICITY THAT CAN BLOCK OR 9967 06:18:25,582 --> 06:18:27,450 CONTROL KSHV INFECTION? 9968 06:18:27,450 --> 06:18:30,720 I USE A HERPESVIRUS PATHOGEN, 9969 06:18:30,720 --> 06:18:33,022 NATURAL PATHOGEN OF RODENTS AND 9970 06:18:33,022 --> 06:18:35,859 GENOME COLINEAR WITH KSHV, VIRUS 9971 06:18:35,859 --> 06:18:37,227 STUDIED FOR ABOUT 30 YEARS NOW 9972 06:18:37,227 --> 06:18:39,028 AND WE HAVE A NICE GENETICALLY 9973 06:18:39,028 --> 06:18:40,997 TRACKABLE SYSTEM WHERE WE'VE 9974 06:18:40,997 --> 06:18:42,665 USED MUTANT VIRUSES AND MUTANT 9975 06:18:42,665 --> 06:18:44,968 MICE AND LEARNED A LOT ABOUT THE 9976 06:18:44,968 --> 06:18:53,843 SPATIAL TEMPORAL KINETICS. 9977 06:18:53,843 --> 06:18:55,378 LAB ACTUALLY WHEN HE WAS AT 9978 06:18:55,378 --> 06:18:57,580 SAINT JUDZ A LOT OF THE 9979 06:18:57,580 --> 06:18:59,949 FOUNDATION -- SAINT JUD HE'S, WE 9980 06:18:59,949 --> 06:19:02,151 KNOW A LOT ABOUT TYPES OF IMMUNE 9981 06:19:02,151 --> 06:19:04,087 CELLS THAT RESPOND TO CONTROL, 9982 06:19:04,087 --> 06:19:05,655 REPLICATION AND I LA TENDENCY. 9983 06:19:05,655 --> 06:19:06,589 TAKEN TOGETHER THIS IS A GREAT 9984 06:19:06,589 --> 06:19:09,292 SYSTEM TO EXPLORE POSSIBLE 9985 06:19:09,292 --> 06:19:10,927 VACCINE PLATFORMS AND BLEARNT 9986 06:19:10,927 --> 06:19:12,929 IMMUNE CORRELATES OF PROTECTION. 9987 06:19:12,929 --> 06:19:15,331 SO MY COLLABORATOR CRAIG FOREST, 9988 06:19:15,331 --> 06:19:17,367 UNIVERSITY OF ARKANSAS MEDICAL 9989 06:19:17,367 --> 06:19:18,368 SIENLSES I FORGOT TO MENTION ON 9990 06:19:18,368 --> 06:19:20,136 THE FIRST SLIDE, WE'VE 9991 06:19:20,136 --> 06:19:21,337 COLLABORATED AND DECIDED TO DO A 9992 06:19:21,337 --> 06:19:22,639 RATIONAL APPROACH WHERE WE'VE 9993 06:19:22,639 --> 06:19:25,041 KNOCKED OUT DETERMINANTS OFLYTIC 9994 06:19:25,041 --> 06:19:27,477 REPLICATION HERE I'M FOCUSED ON 9995 06:19:27,477 --> 06:19:29,979 RTA ENCODE BY OR50, NECESSARY 9996 06:19:29,979 --> 06:19:32,615 AND SUFFICIENT TO TRIGGERLYTIC 9997 06:19:32,615 --> 06:19:33,917 GENE EXPRESSION AND THEN WE'VE 9998 06:19:33,917 --> 06:19:35,985 ALSO TARGETED LATENCY 9999 06:19:35,985 --> 06:19:38,087 DETERMINIST LEFT IN THE VIRUS 10000 06:19:38,087 --> 06:19:39,822 THAT PROMOTE LATENCY AND AGAIN 10001 06:19:39,822 --> 06:19:45,161 THIS IS MHV68 COLINEAR WITH KHSV 10002 06:19:45,161 --> 06:19:47,497 ON THE -- KSHV ON THE BOTTOM. 10003 06:19:47,497 --> 06:19:49,332 DECIDE TO DO MAKE A REPLICATION 10004 06:19:49,332 --> 06:19:49,933 DEAD VIRUS. 10005 06:19:49,933 --> 06:19:51,534 HOW DO YOU GROW SUCH A VIRUS IN 10006 06:19:51,534 --> 06:19:54,037 WE CAME WUP A STRATEGY WHERE WE 10007 06:19:54,037 --> 06:19:55,471 CAN PRODUCE IT IN A CELL LINE 10008 06:19:55,471 --> 06:19:56,673 WITH RESPECT TO THE 10009 06:19:56,673 --> 06:19:58,708 COMPLIMENTING GENE IS CODON 10010 06:19:58,708 --> 06:19:59,676 SHUFFLED, PREVENTS RECOMBINATION 10011 06:19:59,676 --> 06:20:01,844 SO WE DON'T GET REVERT ENS. 10012 06:20:01,844 --> 06:20:03,713 WE END UP WITH A VIRUS THAT 10013 06:20:03,713 --> 06:20:05,381 BASICALLY ENTERS A CELL AND 10014 06:20:05,381 --> 06:20:06,916 UNDERGOES AN ABORTIVE INFECTION, 10015 06:20:06,916 --> 06:20:08,151 SINGLE ROUND OF INFECTION WITH 10016 06:20:08,151 --> 06:20:09,819 NO INFECTIOUS PARTICLES ON THE 10017 06:20:09,819 --> 06:20:10,787 BACK END. 10018 06:20:10,787 --> 06:20:12,188 WE'VE TAKEN THESE VIRUS STOCKS 10019 06:20:12,188 --> 06:20:13,990 AND INFECTED IMMUNODEFICIENT 10020 06:20:13,990 --> 06:20:16,926 MICE AND IN THIS SYSTEM WE CAN 10021 06:20:16,926 --> 06:20:20,296 ONE PFU A WILD-TYPE IMAGE V68 10022 06:20:20,296 --> 06:20:22,832 CAN LEAD TO 100% MORTALITY, WE 10023 06:20:22,832 --> 06:20:24,901 PUT IN A MILLION PFU OF OUR 10024 06:20:24,901 --> 06:20:26,869 VIRUS WHICH I'LL REFER TO AAS 10025 06:20:26,869 --> 06:20:32,976 THE RDV ORF50 STOP CODON, 100% 10026 06:20:32,976 --> 06:20:33,242 SURVIVAL. 10027 06:20:33,242 --> 06:20:35,345 SO WE ENVISION THIS VIRUS AS 10028 06:20:35,345 --> 06:20:36,980 ENTERING A CELL, IT'S BASICALLY 10029 06:20:36,980 --> 06:20:38,748 SORT OF LIKE A VLP BECAUSE THE 10030 06:20:38,748 --> 06:20:39,949 VIRUS IS INTACT. 10031 06:20:39,949 --> 06:20:41,851 BUT IT HAS A GENOME, IT DELIVERS 10032 06:20:41,851 --> 06:20:43,987 THAT TO THE NUCLEUS AND YOU HAVE 10033 06:20:43,987 --> 06:20:45,455 NEW GENE EXPRESSION AND 10034 06:20:45,455 --> 06:20:46,789 DEPENDING ON THE CELL TYPE YOU 10035 06:20:46,789 --> 06:20:50,226 CAN GET MHC1 AND 2 PRESENTATION. 10036 06:20:50,226 --> 06:20:53,296 WE LOOKED AT THE RNA SEQ OF 10037 06:20:53,296 --> 06:20:55,198 INFECTED CELLS WITH THE 50 STOP 10038 06:20:55,198 --> 06:20:56,933 VIRUS HERE OR THE WILD-TYPE 10039 06:20:56,933 --> 06:20:58,134 VIRUS ON THE TOP AND YOU CAN SEE 10040 06:20:58,134 --> 06:21:00,236 THERE'S GREATLY DIMINISHED GENE 10041 06:21:00,236 --> 06:21:02,739 EXPRESSION, THESE ARE THE ORF'S 10042 06:21:02,739 --> 06:21:07,443 OF MHV68 MUCH DEPRESSED GENE 10043 06:21:07,443 --> 06:21:08,945 EXPRESSION WRVMENT WE SEE GENE 10044 06:21:08,945 --> 06:21:10,046 EXPRESSION IT'S INTERESTING 10045 06:21:10,046 --> 06:21:11,414 BECAUSE SOME OF THESE ARE 10046 06:21:11,414 --> 06:21:12,115 LATENCY DETERMINE ABILITY, 10047 06:21:12,115 --> 06:21:13,449 SOMETHING TO KEEP IN MIND AND WE 10048 06:21:13,449 --> 06:21:15,718 ALSO HAVE WHAT WE USE AS 10049 06:21:15,718 --> 06:21:17,153 IMMUNODOMINANT EPITOPES TO 10050 06:21:17,153 --> 06:21:19,222 FOLLOW VIRUS SPECIFIC T CELL 10051 06:21:19,222 --> 06:21:21,324 RESPONSES IN BLACK SIX MICE, 10052 06:21:21,324 --> 06:21:23,826 EXPRESSED INDEPENDENTLY OF RTA. 10053 06:21:23,826 --> 06:21:24,027 OKAY. 10054 06:21:24,027 --> 06:21:26,229 SO HE WITH TAKE THIS VIRUS AND 10055 06:21:26,229 --> 06:21:28,431 NOW WE'RE INFECTING BLACK SIX 10056 06:21:28,431 --> 06:21:28,631 MICE. 10057 06:21:28,631 --> 06:21:32,168 WE DO A PRIME WITH A MOIL YON 10058 06:21:32,168 --> 06:21:34,370 PFU IP, BOOST ONCE OR TWICE, AND 10059 06:21:34,370 --> 06:21:36,239 THEN FOLLOW ANTIBODY AND VIRUS 10060 06:21:36,239 --> 06:21:37,640 SPECIFIC T CELL RESPONSES. 10061 06:21:37,640 --> 06:21:39,409 AND THIS IS A BLOT WHERE WE'VE 10062 06:21:39,409 --> 06:21:41,277 TAKEN SERUM FROM MICE EITHER 10063 06:21:41,277 --> 06:21:42,712 INFECTED LIKE THE NORMAL 10064 06:21:42,712 --> 06:21:44,313 WILD-TYPE VIRUS OR VACCINATED 10065 06:21:44,313 --> 06:21:46,416 AND IT'S PRETTY OBVIOUS WE DON'T 10066 06:21:46,416 --> 06:21:49,118 SEE MUCH SIGNAL REACTING AGAINST 10067 06:21:49,118 --> 06:21:51,954 VIRALLY INFECTED LYSATES WHERE 10068 06:21:51,954 --> 06:21:53,723 50 STOP AS COMPARED TO WILD-TYPE 10069 06:21:53,723 --> 06:21:55,825 BY SECOND BOOST RER SEEING SOME 10070 06:21:55,825 --> 06:21:56,125 REACTIVITY. 10071 06:21:56,125 --> 06:21:57,860 THIS WAS ALSO CONFIRMED BY OUR 10072 06:21:57,860 --> 06:22:01,297 VIRUS SPECIFIC IGG ELISA, WE 10073 06:22:01,297 --> 06:22:03,032 HAVE WILD-TYPE PREDOMINANT, LOTS 10074 06:22:03,032 --> 06:22:04,667 OF GOOD ANTIBODIES BEING 10075 06:22:04,667 --> 06:22:05,735 GENERATED, WE HAVE LESS SO FOR 10076 06:22:05,735 --> 06:22:08,071 THE RTA50 STOP. 10077 06:22:08,071 --> 06:22:10,273 INTERESTINGLY THOUGH, WE DO SEE 10078 06:22:10,273 --> 06:22:12,041 THAT NEUTRALIZING ANTIBODIES BY 10079 06:22:12,041 --> 06:22:13,476 THIS PLAQUE REDUCTION 10080 06:22:13,476 --> 06:22:16,512 NEUTRALIZATION ASSAY WE DO SEE 10081 06:22:16,512 --> 06:22:17,613 NEUTRALIZING ANTIBODIES THAT 10082 06:22:17,613 --> 06:22:19,148 APPROACH WILD-TYPE INFECTION. 10083 06:22:19,148 --> 06:22:20,683 SO WE ARE GENERATING 10084 06:22:20,683 --> 06:22:21,984 NEUTRALIZING ANTIBODIES WITH 10085 06:22:21,984 --> 06:22:23,419 THIS VACCINE. 10086 06:22:23,419 --> 06:22:25,555 SO I'LL SHOW YOU DATA NOW, WE 10087 06:22:25,555 --> 06:22:26,956 ALSO DID T CELL WORK-UP BUT I'LL 10088 06:22:26,956 --> 06:22:28,724 SHOW YOU THAT T CELL DATA NOW IN 10089 06:22:28,724 --> 06:22:30,193 THE NEXT SET OF DATA. 10090 06:22:30,193 --> 06:22:31,794 NOW WE'RE PRIME BOOSTING THESE 10091 06:22:31,794 --> 06:22:33,429 MICE AND THEN WE'RE CHALLENGING 10092 06:22:33,429 --> 06:22:37,467 WITH WILD-TYPE MHD68 BOOSTING IP 10093 06:22:37,467 --> 06:22:39,335 CHALLENGING INTRANASALLY. 10094 06:22:39,335 --> 06:22:41,270 WHAT WE GET HERE IS REPLICATION 10095 06:22:41,270 --> 06:22:42,605 IN THE LUNGS FOR THE ACUTE PHASE 10096 06:22:42,605 --> 06:22:44,674 AND THEN WE FOLLOW LATENCY 10097 06:22:44,674 --> 06:22:46,776 ESTABLISHMENT IN THE SPLEEN FOR 10098 06:22:46,776 --> 06:22:52,582 LATENCY. 10099 06:22:52,582 --> 06:22:54,984 LUNGS OF THESE VAC NAITD MICE, 10100 06:22:54,984 --> 06:22:57,753 HERE WE HAVE SHA AM VACCINATOR 10101 06:22:57,753 --> 06:23:01,891 IN RED AND VACCINATED WITH RDV50 10102 06:23:01,891 --> 06:23:02,959 STOP VIRUS IN BLUE. 10103 06:23:02,959 --> 06:23:04,327 YOU CAN APPRECIATE WE HAVE A 10104 06:23:04,327 --> 06:23:06,062 FOUR LOG REDUCTION IN THE LEVELS 10105 06:23:06,062 --> 06:23:08,064 OF VIRUS REPLICATION IN THE 10106 06:23:08,064 --> 06:23:09,665 LUNGS OF THESE MICE AND IT LOOKS 10107 06:23:09,665 --> 06:23:10,867 A LITTLE BIT BETTER WITH TWO 10108 06:23:10,867 --> 06:23:12,068 BOOST HERE ALTHOUGH THE MICE 10109 06:23:12,068 --> 06:23:13,302 NUMBERS ARE LIMITED. 10110 06:23:13,302 --> 06:23:15,505 I'M SHOWING YOU WE ALSO SAW 10111 06:23:15,505 --> 06:23:16,572 NEUTRALIZING ANTIBODIES AS I 10112 06:23:16,572 --> 06:23:18,574 SHOWED YOU BEFORE THAT ARE 10113 06:23:18,574 --> 06:23:20,810 COMPARABLE TO WILD-TYPE. 10114 06:23:20,810 --> 06:23:22,278 WHAT ABOUT THE T CELL RESPONSE? 10115 06:23:22,278 --> 06:23:23,913 SO WHAT ARE THOSE IMMUNE 10116 06:23:23,913 --> 06:23:24,680 CORRELATES OF PROTECTION? 10117 06:23:24,680 --> 06:23:25,982 HERE WE'RE LOOKING AT THE LUNGS 10118 06:23:25,982 --> 06:23:28,151 AT THE SAME DAY, DAY SEVEN POST 10119 06:23:28,151 --> 06:23:30,920 CHALLENGE AND I HAVE FOUR 10120 06:23:30,920 --> 06:23:33,422 GROUPS, WE HAVE -- SORRY -- WE 10121 06:23:33,422 --> 06:23:36,259 HAVE THE SHAM VACCINATED, NOT 10122 06:23:36,259 --> 06:23:37,360 CHALLENGED OR WILD-TYPE 10123 06:23:37,360 --> 06:23:38,895 CHALLENGE, WE HAVE OUR 10124 06:23:38,895 --> 06:23:40,763 VACCINATED ANIMALS NOT 10125 06:23:40,763 --> 06:23:42,532 CHALLENGED AND VACCINATED MICE 10126 06:23:42,532 --> 06:23:44,800 CHAL HE COULD WITH WILD-TYPE 10127 06:23:44,800 --> 06:23:46,235 VIRUS -- CHALLENGED WITH 10128 06:23:46,235 --> 06:23:53,376 WILD-TYPE VIRUS, LOOKING AT P79 10129 06:23:53,376 --> 06:23:54,744 AND 56, LOOKING BIND TO 10130 06:23:54,744 --> 06:23:56,379 RECOGNIZE THESE TWO EPITOPES. 10131 06:23:56,379 --> 06:23:58,581 AND WE FIND THAT IN THE 10132 06:23:58,581 --> 06:24:00,116 VACCINATED ANIMALS UPON 10133 06:24:00,116 --> 06:24:01,551 WILD-TYPE CHALLENGE WE SEE A 10134 06:24:01,551 --> 06:24:03,186 STRONG VIRUS SPECIFIC T CELL 10135 06:24:03,186 --> 06:24:05,354 RESPONSE IN THE LUNGS, AND THAT 10136 06:24:05,354 --> 06:24:08,057 IS WITH BOTH P 1K3-79 P56 AT THE 10137 06:24:08,057 --> 06:24:10,393 TIME MEASURES. 10138 06:24:10,393 --> 06:24:11,827 -- TETRAMERS. 10139 06:24:11,827 --> 06:24:14,897 IN THOSE VACCINATED ANIMALS 10140 06:24:14,897 --> 06:24:16,966 SEEING T CELL PRESENT WHICH 10141 06:24:16,966 --> 06:24:18,034 CORRELATES WITH REPLICATION. 10142 06:24:18,034 --> 06:24:19,202 WE ALL LOOKED IN THE SPLEEN AND 10143 06:24:19,202 --> 06:24:20,570 WE CAN ACTUALLY SEE A POPULATION 10144 06:24:20,570 --> 06:24:22,305 OF VIRUS SPECIFIC T CELLS IN THE 10145 06:24:22,305 --> 06:24:24,840 VACCINATED ANIMALS WITHOUT 10146 06:24:24,840 --> 06:24:25,107 CHALLENGE. 10147 06:24:25,107 --> 06:24:26,709 SHOWN HERE. 10148 06:24:26,709 --> 06:24:28,678 WE SEE SIMILAR LEVELS WITH AND 10149 06:24:28,678 --> 06:24:29,645 WITHOUT CHALLENGE. 10150 06:24:29,645 --> 06:24:33,015 THOSE VACCINATED MICE THAT THE 10151 06:24:33,015 --> 06:24:34,350 VIRUS SPECIFIC T CELLS HAVE A 10152 06:24:34,350 --> 06:24:36,419 SHORT LIVED EFFECTOR PHENOTYPE 10153 06:24:36,419 --> 06:24:38,154 AND OF THE IMPACTS WE ALSO SEE 10154 06:24:38,154 --> 06:24:41,023 THAT THEY ARE CD62L LOW 10155 06:24:41,023 --> 06:24:42,458 INDICATING THEY HAVE EFFECTOR 10156 06:24:42,458 --> 06:24:43,993 SERVICE PHENOTYPES CONSISTENT 10157 06:24:43,993 --> 06:24:46,596 WITH EFFECTOR T CELLS. 10158 06:24:46,596 --> 06:24:48,030 THEN WE LOOKED AT THEIR ABILITY 10159 06:24:48,030 --> 06:24:49,865 TO PRODUCE EFFECTOR CYTOKINES. 10160 06:24:49,865 --> 06:24:51,434 SO WE STIMULATED THE T CELLS 10161 06:24:51,434 --> 06:24:53,069 AGAIN FROM THESE SAME SETS OF 10162 06:24:53,069 --> 06:24:56,505 ANIMALS AND THIS COMPARISON WITH 10163 06:24:56,505 --> 06:24:58,541 EITHER P79 OR P56 AND WE'RE 10164 06:24:58,541 --> 06:25:01,811 LOOKING FOR TNF ALPHA AND 10165 06:25:01,811 --> 06:25:02,912 INTERFERON GAMMA PRODUCTION 10166 06:25:02,912 --> 06:25:04,247 UPPER RIGHT QUADRANT AND SEE 10167 06:25:04,247 --> 06:25:07,516 GHEAN THE VACCINATED ANIMALS 10168 06:25:07,516 --> 06:25:08,618 DOING LESS VIRUS SPECIFIC T 10169 06:25:08,618 --> 06:25:10,486 CELLS THAT CAN RAPIDLY RESPOND 10170 06:25:10,486 --> 06:25:17,293 TO PEPTIDE STIMULATION. 10171 06:25:17,293 --> 06:25:18,761 WHAT ABOUT LATENCY? 10172 06:25:18,761 --> 06:25:20,796 IMPORTANT PART OF GAMMA 10173 06:25:20,796 --> 06:25:21,998 HERPESVIRUS AND IS WE SEE THAT 10174 06:25:21,998 --> 06:25:24,734 ONE INDICATOR FOR MHD68 10175 06:25:24,734 --> 06:25:26,035 COLONIZATION OF THE SPLEEN, 10176 06:25:26,035 --> 06:25:27,737 SPLEN NOTICEMEGALY, SIMILAR TO 10177 06:25:27,737 --> 06:25:29,238 LYMPH NODE ENLARGEMENT ONE MIGHT 10178 06:25:29,238 --> 06:25:33,276 SEE WITH PATIENTS WITH EBD IM, 10179 06:25:33,276 --> 06:25:35,111 INFECTIOUS MONO NUCLEOSIS, THESE 10180 06:25:35,111 --> 06:25:37,413 MICE DEVELOP THREEFOLD 10181 06:25:37,413 --> 06:25:39,181 SPLENOMEGALY IN THE SHAM 10182 06:25:39,181 --> 06:25:40,916 VACCINATED CONDITIONS, IF 10183 06:25:40,916 --> 06:25:43,019 VACCINATED, WE SEE NO SPLEN 10184 06:25:43,019 --> 06:25:43,352 NOTICEMEGALY. 10185 06:25:43,352 --> 06:25:44,854 TWO LAW REDUCTION IN THE 10186 06:25:44,854 --> 06:25:47,623 FREQUENCY OF SITES THAT HARBOR 10187 06:25:47,623 --> 06:25:50,326 THE GENOME AND WE DO THREE WEEK 10188 06:25:50,326 --> 06:25:52,762 EX-PLANT REACTIVATION ASSAY, 10189 06:25:52,762 --> 06:25:53,763 SENSITIVE AND QUANTITATIVE 10190 06:25:53,763 --> 06:25:55,931 MEASUREMENT OF REACTIVATION FROM 10191 06:25:55,931 --> 06:25:58,434 LATENCY, WE SEE NEAR COMPLETE 10192 06:25:58,434 --> 06:26:00,002 ABLAIFTION REACTIVATION IN THOSE 10193 06:26:00,002 --> 06:26:02,438 VACCINATED ANIMALS. 10194 06:26:02,438 --> 06:26:04,173 NOT GOING ON SHOW YOU THE DATA. 10195 06:26:04,173 --> 06:26:05,708 BUT WE WAITED UNTIL THREE MONTHS 10196 06:26:05,708 --> 06:26:08,010 POST BOOST AND WE SEE SIMILAR 10197 06:26:08,010 --> 06:26:09,178 PROTECTION, SIMILAR IMMUNE 10198 06:26:09,178 --> 06:26:09,512 RESPONSES. 10199 06:26:09,512 --> 06:26:12,815 SO IT IS DURABLE PROTECTION. 10200 06:26:12,815 --> 06:26:14,450 SO THEN WE MOVED TO KIND OF HOW 10201 06:26:14,450 --> 06:26:16,886 COULD WE PUSH THE SYSTEM TO 10202 06:26:16,886 --> 06:26:18,621 REALLY CHECK EFFICACY. 10203 06:26:18,621 --> 06:26:21,123 HERE WE'RE LOOKING AT TYPE ONE 10204 06:26:21,123 --> 06:26:22,558 INTERFERON RECEPTOR KNOCK-OUT 10205 06:26:22,558 --> 06:26:24,527 MICE, WE'RE USING A DOSE ABOUT 10206 06:26:24,527 --> 06:26:27,296 TWO MILLION PFEYM WITH WILD-TYPE 10207 06:26:27,296 --> 06:26:29,465 LEADING TO AREA OF ROBUST 10208 06:26:29,465 --> 06:26:33,069 LETHALITY IN THESE ANIMALS, 10209 06:26:33,069 --> 06:26:35,738 THESE ANIMALS WERE PRIME BOOSTED 10210 06:26:35,738 --> 06:26:38,274 WITH R ARE BT STOP VIRUS AND 10211 06:26:38,274 --> 06:26:39,942 LOOKING AT THE RED LINE 10212 06:26:39,942 --> 06:26:41,410 UNVACCINATED ANIMALS DPIERD THE 10213 06:26:41,410 --> 06:26:43,145 BLACK LINES THAT, INDICATES THE 10214 06:26:43,145 --> 06:26:44,547 VACCINATED ANIMALS, WE SEE NO 10215 06:26:44,547 --> 06:26:48,517 WEIGHT LOSS OR SEE VERY LIMITED 10216 06:26:48,517 --> 06:26:54,757 WEIGHT LOSS IN THE -- NOW, ONE 10217 06:26:54,757 --> 06:26:57,259 ASPECT OF THIS IS WE LOOKED OF 10218 06:26:57,259 --> 06:26:59,662 THE MICE THAT SURVIVED, WE 10219 06:26:59,662 --> 06:27:01,731 LOOKED AT WHAT TYPE OF VIRUS IS 10220 06:27:01,731 --> 06:27:04,934 PRESENT IN THOSE SPLEN NOTICE 10221 06:27:04,934 --> 06:27:08,571 SIGHTS, WHAT IS THE VIRAL GENOME 10222 06:27:08,571 --> 06:27:09,638 GONE LATE ENTD. 10223 06:27:09,638 --> 06:27:12,708 INTERESTING TO FIND, WE DO SEE 10224 06:27:12,708 --> 06:27:14,243 LATEN GENE EXPRESSION IN 50 STOP 10225 06:27:14,243 --> 06:27:15,578 VIRUS AND WE DO SEE EVIDENCE 10226 06:27:15,578 --> 06:27:19,281 THAT THIS IS THE GENOTYPING 10227 06:27:19,281 --> 06:27:20,716 ASSAY LOOKING FOR THE 50 STOP 10228 06:27:20,716 --> 06:27:23,552 VIRUS USING GENOME TYPE SPECIFIC 10229 06:27:23,552 --> 06:27:25,955 PRIMERS AND THEN JUST A PAN 10230 06:27:25,955 --> 06:27:29,024 DEFECTION FOR ALL OF MRV68 AND 10231 06:27:29,024 --> 06:27:31,961 GET A COUNT OF ABOUT 16% OF THE 10232 06:27:31,961 --> 06:27:34,497 VIRUS OUR VACCINE VIRUS, CAIGHTD 10233 06:27:34,497 --> 06:27:35,931 THAT VACCINE VIRUS CAN ESTABLISH 10234 06:27:35,931 --> 06:27:37,266 LATENCY AND ALSO INDICATES WE'RE 10235 06:27:37,266 --> 06:27:39,301 NOT BEING STERILIZING IMMUNITY 10236 06:27:39,301 --> 06:27:41,070 BECAUSE WE'RE DETECHING SOME 10237 06:27:41,070 --> 06:27:45,908 WILD-TYPE VIRUS IN THOSE 10238 06:27:45,908 --> 06:27:46,242 SPLENOCYTES. 10239 06:27:46,242 --> 06:27:49,612 RECENTLY WE DECIDED TO TOWARDS 10240 06:27:49,612 --> 06:27:51,647 KNOCKING OUT LATENCY, HERE 10241 06:27:51,647 --> 06:27:54,750 ATTACKING LEFT MH68 WHICH HAS M 10242 06:27:54,750 --> 06:27:56,786 GENES UNIQUE TO 68 THAT PROMOTE 10243 06:27:56,786 --> 06:27:58,421 LATENCY, USED SIMPLE COMPARISON, 10244 06:27:58,421 --> 06:27:59,955 THIS REPLICATION DEAD VIRUS TO 10245 06:27:59,955 --> 06:28:01,690 NOW REPLICATION DEAD VIRUS 10246 06:28:01,690 --> 06:28:02,792 LAYERED ON THAT WE'VE KNOCKED 10247 06:28:02,792 --> 06:28:05,194 OUT GENES NEEDED FOR LATENCY. 10248 06:28:05,194 --> 06:28:06,962 AND WE FOUND THAT IF YOU COMPARE 10249 06:28:06,962 --> 06:28:09,365 THE BLACK TO THE BLUE SQUARES IN 10250 06:28:09,365 --> 06:28:11,534 ALL OF OUR COMPARISONS, BOTH ARE 10251 06:28:11,534 --> 06:28:11,834 PROTECTIVE. 10252 06:28:11,834 --> 06:28:15,404 WE HAVE LOSS OF SPLENO 10253 06:28:15,404 --> 06:28:17,339 MEGMEGALY, WE DON'T SEE LATENCY 10254 06:28:17,339 --> 06:28:19,975 AND EVEN LESS WITH DELTA 4 10255 06:28:19,975 --> 06:28:22,278 MUTANT VIRUS AND DON'T SEE 10256 06:28:22,278 --> 06:28:22,611 REACTIVATION. 10257 06:28:22,611 --> 06:28:25,347 WE ALSO SEE COMPARABLE LEVELS OF 10258 06:28:25,347 --> 06:28:27,116 VIRUS SPECIFIC T CELLS IN THE 10259 06:28:27,116 --> 06:28:28,451 LUNGSZ, THE SPLEENS AND THEY 10260 06:28:28,451 --> 06:28:32,021 HAVE AN EFFECTOR PHENOTYPE AND 10261 06:28:32,021 --> 06:28:36,325 PRODUCE ANTI-VIRAL CYTOKINES. 10262 06:28:36,325 --> 06:28:38,027 SO I'VE SHOWN YOU WE CAN PRIME 10263 06:28:38,027 --> 06:28:39,261 BOOST WITH REPLICATION DEAD 10264 06:28:39,261 --> 06:28:41,564 VIRUS AND A VIRUS THAT'S 10265 06:28:41,564 --> 06:28:43,098 DEFECTIVE FOR LATENCY AND 10266 06:28:43,098 --> 06:28:44,166 PROTECT AGAINST WILD-TYPE 10267 06:28:44,166 --> 06:28:46,469 CHALLENGE USING MANY ASPECTS OF 10268 06:28:46,469 --> 06:28:48,337 PATHOGENESIS IN ANIMAL MODEL, 10269 06:28:48,337 --> 06:28:50,306 REDUCE REPLICATIONS, SPLEN 10270 06:28:50,306 --> 06:28:51,740 NOTICEMEGALY AND PROTECT FROM 10271 06:28:51,740 --> 06:28:53,075 LETHAL DISEASE, AND I'VE SHOWN 10272 06:28:53,075 --> 06:28:54,710 YOU SOME IMMUNE CORRELATES OF 10273 06:28:54,710 --> 06:28:55,578 PROTECTION WHICH ARE STARTING TO 10274 06:28:55,578 --> 06:28:57,213 GIVE US SOME IN ROADS INTO HOW 10275 06:28:57,213 --> 06:28:58,414 THIS MAY BE WORKING. 10276 06:28:58,414 --> 06:29:00,082 THERE'S A LOT MORE WORK TO DO, 10277 06:29:00,082 --> 06:29:01,283 WE'RE VERY EXCITED BECAUSE NOW 10278 06:29:01,283 --> 06:29:03,285 WE'RE ACTUALLY GOING TO TRY TO 10279 06:29:03,285 --> 06:29:04,920 COMPLETELY AN LATE ALL LATENCY 10280 06:29:04,920 --> 06:29:08,190 BY LOCKING OUT LANA NEED TO DO 10281 06:29:08,190 --> 06:29:10,259 TETHER THE VIRAL EPISOME TO THE 10282 06:29:10,259 --> 06:29:12,328 HOST AND PUSH BACK THAT BLOCK OF 10283 06:29:12,328 --> 06:29:13,229 REPLICATION FROM EARLY TO 10284 06:29:13,229 --> 06:29:15,397 ACTUALLY VERY LATE IN THE LYTIC 10285 06:29:15,397 --> 06:29:17,032 LIFE CYCLE BY TARGETING 10286 06:29:17,032 --> 06:29:17,933 DIFFERENT VIRAL PROTEIN AND WE 10287 06:29:17,933 --> 06:29:19,969 LIKE TO COMPARE IT THIS TO HEAT 10288 06:29:19,969 --> 06:29:21,871 AND ACTIVATION S THERE BENEFIT 10289 06:29:21,871 --> 06:29:23,439 FROM ADJUVANT AND I'M SETTING UP 10290 06:29:23,439 --> 06:29:26,041 A COLLABORATION WITH JEFF COHEN 10291 06:29:26,041 --> 06:29:28,777 AND NASARO TO USE THAT APPROACH 10292 06:29:28,777 --> 06:29:32,381 WITH EBV USING SERATIN NANO 10293 06:29:32,381 --> 06:29:33,415 PARTICLES THAT WE CAN SEE IF WE 10294 06:29:33,415 --> 06:29:35,784 CAN DO A SUBUNIT BOOST TO 10295 06:29:35,784 --> 06:29:37,786 FURTHER INCREASE PROTECTION. 10296 06:29:37,786 --> 06:29:39,355 AND I APPRECIATE ANY FEEDBACK 10297 06:29:39,355 --> 06:29:40,756 FROM THE I AM NO GISTS IN THE 10298 06:29:40,756 --> 06:29:42,057 ROOM BECAUSE WHAT WE NEED TO DO 10299 06:29:42,057 --> 06:29:43,259 IS ACTUALLY TAKE THIS A STEP 10300 06:29:43,259 --> 06:29:43,692 FURTHER. 10301 06:29:43,692 --> 06:29:47,630 ARE WE GETTING SECRETORY IGA, IS 10302 06:29:47,630 --> 06:29:51,066 THIS GENERATING NEUTRALIZATION 10303 06:29:51,066 --> 06:29:52,034 AGAINST SPECIFIC PROTEINS AND 10304 06:29:52,034 --> 06:29:54,236 HOW ARE THE ANTIBODIES WORKING 10305 06:29:54,236 --> 06:29:56,405 BEYOND NEUTRALIZING ANTIBODIES, 10306 06:29:56,405 --> 06:29:58,240 WHAT IS THE ROLE OF CD4 T CELLS 10307 06:29:58,240 --> 06:30:01,110 AND CAN WE USE THIS TO A MODEL 10308 06:30:01,110 --> 06:30:04,813 THAT PSES KSHV DISEASE AND 10309 06:30:04,813 --> 06:30:06,815 EFFECTIVE VACCINE THERE. 10310 06:30:06,815 --> 06:30:08,584 COLLABORATORS CRAIG FOREST, I'VE 10311 06:30:08,584 --> 06:30:10,452 GREAT INSIGHT FROM BRIAN 10312 06:30:10,452 --> 06:30:11,987 SHERIDAN T CELL I AM NO GIST 10313 06:30:11,987 --> 06:30:15,691 FROM STONY BROOK UNIVERSITY 10314 06:30:15,691 --> 06:30:19,161 AND -- IN MY GROUP ANNA GROSCOFF 10315 06:30:19,161 --> 06:30:22,665 AND DIPA HAVE GENERATED DATA TO 10316 06:30:22,665 --> 06:30:28,938 THIS PROJECT AND PREVIOUS HADLY 10317 06:30:28,938 --> 06:30:30,773 DPAISH IS I'LL STOP THERE AND 10318 06:30:30,773 --> 06:30:33,742 TAKE ANY QUESTIONS. 10319 06:30:33,742 --> 06:30:41,750 [APPLAUSE] 10320 06:30:41,750 --> 06:30:44,587 >> I DON'T KNOW WHO IS OLDER, 10321 06:30:44,587 --> 06:30:46,655 REMY, YOU OR ME, SO I'LL GO 10322 06:30:46,655 --> 06:30:46,855 FIRST. 10323 06:30:46,855 --> 06:30:48,624 THAT WAS GREARKTS LAURIE, THANK 10324 06:30:48,624 --> 06:30:48,791 YOU. 10325 06:30:48,791 --> 06:30:50,159 -- THAT WAS GREAT, LAURIE, THANK 10326 06:30:50,159 --> 06:30:50,926 YOU. 10327 06:30:50,926 --> 06:30:51,627 A QUICK QUESTION, BECAUSE IT 10328 06:30:51,627 --> 06:30:53,495 LOOKED TO ME, MAYBE I MISSED IT, 10329 06:30:53,495 --> 06:30:54,797 THAT THE REDUCTION OF YOUR 10330 06:30:54,797 --> 06:30:56,999 LATENCY WAS LESS THAN YOUR 10331 06:30:56,999 --> 06:30:59,068 REACTIVATION FROM LATENCY. 10332 06:30:59,068 --> 06:31:01,537 SO IN THE CONTEXT OF SOME OF THE 10333 06:31:01,537 --> 06:31:03,672 TRANSFORMATIONS WHICH ARE LATENT 10334 06:31:03,672 --> 06:31:06,542 WHAT DO YOU THINK WOULD HAPPEN 10335 06:31:06,542 --> 06:31:06,909 THEN? 10336 06:31:06,909 --> 06:31:09,411 >> SO IN GENERAL, WILD-TYPE 10337 06:31:09,411 --> 06:31:10,813 VIRUS ABOUT TEN PERCENT OF THE 10338 06:31:10,813 --> 06:31:14,183 VIRUS THAT ESTABLISHES LATENCY 10339 06:31:14,183 --> 06:31:15,184 REACTIVATES, SO THAT'S ONE 10340 06:31:15,184 --> 06:31:16,251 DIFFERENCE THAT MAY HAVE LOOKED 10341 06:31:16,251 --> 06:31:17,553 DIFFERENT IN THOSE GRAPHS. 10342 06:31:17,553 --> 06:31:19,088 WE ACTUAL HAVE A BETTER BLOCK 10343 06:31:19,088 --> 06:31:20,789 WITH REACTIVATION THAN LATENCY. 10344 06:31:20,789 --> 06:31:23,592 >> THAT'S WHAT I WAS SAYING. 10345 06:31:23,592 --> 06:31:23,759 YES. 10346 06:31:23,759 --> 06:31:25,694 >> SO AT THIS POINT WE DO NOT 10347 06:31:25,694 --> 06:31:27,763 HAVE STERILIZING IMMUNITY, VIRUS 10348 06:31:27,763 --> 06:31:30,366 IS STILL ESTABLISHING LATENCY, I 10349 06:31:30,366 --> 06:31:31,834 THINK WHAT I FORESEE SYNC WE 10350 06:31:31,834 --> 06:31:33,902 WOULD BE REDUCING, WE CAN TEST 10351 06:31:33,902 --> 06:31:35,537 THIS VIRAL LOAD OVERALL BUT I 10352 06:31:35,537 --> 06:31:36,872 THINK THE IMPORTANT PART IS 10353 06:31:36,872 --> 06:31:38,207 BLOCKERRING REACTIVATION BECAUSE 10354 06:31:38,207 --> 06:31:40,809 A LOT OF GAMMA HERPESVIRUS 10355 06:31:40,809 --> 06:31:42,011 DISEASES ARE WHEN YOU LOSE 10356 06:31:42,011 --> 06:31:43,746 IMMUNE CONTROL, THESE VIRUSES 10357 06:31:43,746 --> 06:31:45,948 COULD REACTIVATE, INFECT NEW 10358 06:31:45,948 --> 06:31:48,751 CELLS AND DRIVE 10359 06:31:48,751 --> 06:31:50,085 LYMPHOPROLIFERATIVE EXPANSION OF 10360 06:31:50,085 --> 06:31:50,786 THOSE TARGETED CELLS. 10361 06:31:50,786 --> 06:31:53,155 >> I GUESS YOU COULD ARGUE BOTH. 10362 06:31:53,155 --> 06:31:54,289 I AGREE WITH YOU WE WOULD 10363 06:31:54,289 --> 06:31:55,658 PREDICT IT WOULD BE HARDER TO 10364 06:31:55,658 --> 06:31:58,093 CONTROL LATENCY BECAUSE THERE'S 10365 06:31:58,093 --> 06:31:59,728 SO MANY FEWER OPEN READING FRAME 10366 06:31:59,728 --> 06:32:00,896 EXPRESSED IN TERMS OF THE 10367 06:32:00,896 --> 06:32:01,430 IMMUNITY AS WELL. 10368 06:32:01,430 --> 06:32:01,930 >> RIGHT. 10369 06:32:01,930 --> 06:32:03,866 SO AGAIN, IT'S ACTUALLY ALSO NOT 10370 06:32:03,866 --> 06:32:05,401 WELL DEFINED IN HERPESVIRUSES 10371 06:32:05,401 --> 06:32:07,036 WHEN YOU HAVE A LATENT 10372 06:32:07,036 --> 06:32:08,337 INFECTION, THERE IS A 10373 06:32:08,337 --> 06:32:09,872 REANIMATION, THE VIRUS IS 10374 06:32:09,872 --> 06:32:11,940 CONSTANTLY REACTIVATING TO SOME 10375 06:32:11,940 --> 06:32:13,709 DEGREE SO ANTIGENS ARE BEING 10376 06:32:13,709 --> 06:32:14,743 EXPRESSED SO WE'RE HOPING WE 10377 06:32:14,743 --> 06:32:15,911 WOULD GENERATE MORE OF A 10378 06:32:15,911 --> 06:32:18,647 RESPONSE TO THOSE LYTIC ANTIGENS 10379 06:32:18,647 --> 06:32:21,717 THAT MAY CLEAR INFECTED CELLS. 10380 06:32:21,717 --> 06:32:23,986 >> SO THIS IS A VERY NAIVE AND 10381 06:32:23,986 --> 06:32:24,753 SIMPLISTIC QUESTION, I DON'T 10382 06:32:24,753 --> 06:32:28,290 KNOW IF YOU CAN HEAR ME. 10383 06:32:28,290 --> 06:32:29,892 WE'RE HEARING MANY GREAT THINGS 10384 06:32:29,892 --> 06:32:32,961 ON BACTERIA, FUNGI AND THE LIKE. 10385 06:32:32,961 --> 06:32:34,496 >> YOU NEED TO KISS THE MIKE 10386 06:32:34,496 --> 06:32:35,764 ZEEN I NEED TO WHAT? 10387 06:32:35,764 --> 06:32:36,632 TO DO THIS? 10388 06:32:36,632 --> 06:32:36,965 OKAY. 10389 06:32:36,965 --> 06:32:44,073 SO WE ARE HEARING MANY GREAT 10390 06:32:44,073 --> 06:32:47,276 THINGS -- IS THERE -- IS IT 10391 06:32:47,276 --> 06:32:49,144 POSSIBLE THAT THEY ACT AS 10392 06:32:49,144 --> 06:32:53,082 COMMENSURATE FOR US TOO SO THAT 10393 06:32:53,082 --> 06:32:56,819 WE USE THE RESPONSES, DID 10394 06:32:56,819 --> 06:32:58,821 ANYBODY EVER LOOK AT THIS? 10395 06:32:58,821 --> 06:33:02,624 >> I'M NOT SURE I CAUGHT THAT. 10396 06:33:02,624 --> 06:33:04,026 COULD YOU REWORD THAT AGAIN? 10397 06:33:04,026 --> 06:33:05,894 I'M SORRY, IT KEPT ZIESH YOU 10398 06:33:05,894 --> 06:33:06,562 DON'T UNDERSTAND FRENCH. 10399 06:33:06,562 --> 06:33:09,398 >> I'M SORRY. 10400 06:33:09,398 --> 06:33:10,866 [LAUGHTER] 10401 06:33:10,866 --> 06:33:14,002 >> THE QUESTION IS, IS IT 10402 06:33:14,002 --> 06:33:24,446 POSSIBLE, I SUPPOSE THESE 10403 06:33:35,424 --> 06:33:37,659 >> I THINK THERE IS A PAPER A 10404 06:33:37,659 --> 06:33:41,230 FEW YEARS AGO FROM SKIP BURGES 10405 06:33:41,230 --> 06:33:42,498 LAB THAT HERPES IS GOOD FOR YOU. 10406 06:33:42,498 --> 06:33:44,533 >> I GINT THAT FAR SCWHRO THAT 10407 06:33:44,533 --> 06:33:46,168 IF YOU HAD A HERPESVIRUS 10408 06:33:46,168 --> 06:33:47,603 INFECTION IT COULD PROVIDE SOME 10409 06:33:47,603 --> 06:33:49,104 PROTECTION AGAINST SOME OTHER, 10410 06:33:49,104 --> 06:33:50,539 IT COULD ENHANCE CYTOKINE 10411 06:33:50,539 --> 06:33:52,941 RESPONSES AND SOME OTHER 10412 06:33:52,941 --> 06:33:53,175 EFFECTS. 10413 06:33:53,175 --> 06:33:54,810 >> ACTIVITY WITH SOME OTHER 10414 06:33:54,810 --> 06:33:55,377 ANTIGEN OR -- 10415 06:33:55,377 --> 06:33:57,980 >> I THINK THERE WAS A GOOD TALK 10416 06:33:57,980 --> 06:33:59,715 ON VCN THAT I WASN'T REALLY 10417 06:33:59,715 --> 06:34:00,415 AWARE OF. 10418 06:34:00,415 --> 06:34:02,918 SO IT WAS PRETTY SURPRISING. 10419 06:34:02,918 --> 06:34:06,655 KSHV IS UNLIKE EBV IT'S NOT 10420 06:34:06,655 --> 06:34:08,390 SEROPREVALENT IN THIS COUNTRY, 10421 06:34:08,390 --> 06:34:10,259 IT'S ENDEMIC IN CERTAIN AREAS 10422 06:34:10,259 --> 06:34:12,995 SUCH AS SUB-SAHARAN AFRICA AND 10423 06:34:12,995 --> 06:34:13,996 THOSE POPULATIONS CERTAINLY 10424 06:34:13,996 --> 06:34:18,901 SUFFER FROM MORE KSHV ASSOCIATED 10425 06:34:18,901 --> 06:34:21,503 DISEASE -- DISEASE DISEASES THAN 10426 06:34:21,503 --> 06:34:21,737 CANCER. 10427 06:34:21,737 --> 06:34:26,241 I THINK THE JURY OUT ON EBV AND 10428 06:34:26,241 --> 06:34:26,742 CANCER AND THAT. 10429 06:34:26,742 --> 06:34:27,176 >> THANK YOU. 10430 06:34:27,176 --> 06:34:37,419 >> THANK YOU. 10431 06:34:38,587 --> 06:34:41,356 >> CAN YOU HEAR ME? 10432 06:34:41,356 --> 06:34:44,960 SO MY JOB IS TO GET EVERYBODY 10433 06:34:44,960 --> 06:34:46,161 OUT BY 5:00. 10434 06:34:46,161 --> 06:34:48,997 SO I WILL TRY TO DO THAT, 10435 06:34:48,997 --> 06:34:50,766 BECAUSE THIS WORK, SO I'M GOING 10436 06:34:50,766 --> 06:34:54,169 TO TELL YOU ABOUT, SWITCH GEARS 10437 06:34:54,169 --> 06:34:54,703 A BIT. 10438 06:34:54,703 --> 06:35:00,342 BY THE WAY, I'M -- IN CASE YOU 10439 06:35:00,342 --> 06:35:00,843 NOTICED. 10440 06:35:00,843 --> 06:35:03,011 SO I'M GOING TO TALK ABOUT A NEW 10441 06:35:03,011 --> 06:35:04,913 APPROACH FOR CANCER 10442 06:35:04,913 --> 06:35:06,415 IMMUNOTHERAPY THAT INVOLVES WHAT 10443 06:35:06,415 --> 06:35:08,817 WE THINK IS AN INTERESTING 10444 06:35:08,817 --> 06:35:11,486 EPIGENETIC ENZYME CALLED COM1. 10445 06:35:11,486 --> 06:35:14,089 SO WE DISCOVERED COM1 IN A T 10446 06:35:14,089 --> 06:35:18,026 CELL CRISPR SCREEN SO WE WANTED 10447 06:35:18,026 --> 06:35:22,197 TO IDENTIFY NOVEL REGULATORS OF 10448 06:35:22,197 --> 06:35:23,732 T CELL FUNCTION, MAYBE I'LL USE 10449 06:35:23,732 --> 06:35:25,167 THE POINTER HERE. 10450 06:35:25,167 --> 06:35:29,571 SO THE WAY THE SCREEN WORKED WAS 10451 06:35:29,571 --> 06:35:34,209 THAT WE INJECTED EDITED T CELLS 10452 06:35:34,209 --> 06:35:36,511 INTO MICE WITH SUBCUTANEOUS 10453 06:35:36,511 --> 06:35:38,680 MELANOMAS THAT EXPRESSED THE 10454 06:35:38,680 --> 06:35:40,449 OVER TUMOR ANTIGEN AND THE IDEA 10455 06:35:40,449 --> 06:35:43,518 WAS TO IDENTIFY GUIDE RNAS THAT 10456 06:35:43,518 --> 06:35:46,655 WOULD ENHANCE T CELL EXPANSION 10457 06:35:46,655 --> 06:35:48,023 SELECTIVELY IN THE TUMOR BUT NOT 10458 06:35:48,023 --> 06:35:52,928 IN THE CONTROL ORGAN LIKE THE 10459 06:35:52,928 --> 06:35:53,362 SPLEEN. 10460 06:35:53,362 --> 06:35:56,231 SO THE TOP HINT IN THE SCREEN 10461 06:35:56,231 --> 06:35:58,734 WAS COM1 WHICH IS AN EPIGENETIC 10462 06:35:58,734 --> 06:36:00,402 ENZYME, THE NUMBER OF POSITIVE 10463 06:36:00,402 --> 06:36:03,038 CONTROLS HERE FOR EXAMPLE PDC1 10464 06:36:03,038 --> 06:36:06,875 IS PD1, SMAT2 IS A KEY 10465 06:36:06,875 --> 06:36:09,811 TRANSCRIPTION FACTOR IN TGF BETA 10466 06:36:09,811 --> 06:36:10,579 PATHWAY SO WE ARE REALLY 10467 06:36:10,579 --> 06:36:14,016 INTERESTED IN WHAT THIS ENZYME 10468 06:36:14,016 --> 06:36:14,216 DOES. 10469 06:36:14,216 --> 06:36:18,020 SO CARM1 IS A CO-TRANSCRIPTIONAL 10470 06:36:18,020 --> 06:36:18,287 ACTIVATOR. 10471 06:36:18,287 --> 06:36:20,322 IT IS ACTUALLY INITIALLY 10472 06:36:20,322 --> 06:36:23,525 DISCOVERED AS A CO-ACTIVATOR OF 10473 06:36:23,525 --> 06:36:24,960 NUCLEAR HORMONE RECEPTORS AND 10474 06:36:24,960 --> 06:36:26,361 THEREFORE PLAYS A VERY IMPORTANT 10475 06:36:26,361 --> 06:36:29,197 ROLE IN THE PATHOGENESIS OF BOTH 10476 06:36:29,197 --> 06:36:31,867 BREAST AND PROS STATED CANCERS 10477 06:36:31,867 --> 06:36:33,769 BUT IT -- PROSTATE CANCERS BUT 10478 06:36:33,769 --> 06:36:36,571 IT ALSO METH LATES, SO WHAT 10479 06:36:36,571 --> 06:36:47,082 CARM1 DOES IS IT METH LATES -- 10480 06:36:48,550 --> 06:36:50,585 METHYLATES AND SPECIFICALLY THE 10481 06:36:50,585 --> 06:36:54,389 BATH ARE 55 SUBUNIT. 10482 06:36:54,389 --> 06:36:56,692 BAF155 SUB UBT. 10483 06:36:56,692 --> 06:36:59,528 WE DID TO VALIDATE THAT CARM1 10484 06:36:59,528 --> 06:37:02,164 HAS A ROLE IN T CELLS AND KNOCK 10485 06:37:02,164 --> 06:37:04,232 OUT CARM1 GENE IN T CELLS IN 10486 06:37:04,232 --> 06:37:08,603 VITRO AND THEN TRANSLATE THESE 10487 06:37:08,603 --> 06:37:10,973 CELLS TO MICE WITH OVER TUMORS 10488 06:37:10,973 --> 06:37:12,441 WE FIND THAT THE T CELL NUMBERS 10489 06:37:12,441 --> 06:37:14,343 IN THE TUMORS ARE GREATLY 10490 06:37:14,343 --> 06:37:15,344 INCREASED, THE T CELLS HAVE 10491 06:37:15,344 --> 06:37:17,746 GREATLY INCREASED FUNCTIONALITY, 10492 06:37:17,746 --> 06:37:20,248 AS READ OUT HERE BY GAMMA 10493 06:37:20,248 --> 06:37:22,317 INTERFERON EXPRESSION, AND 10494 06:37:22,317 --> 06:37:23,919 THERE'S ALSO RETENTION OF A 10495 06:37:23,919 --> 06:37:27,322 GREATER POPULATION OF MEMORY T 10496 06:37:27,322 --> 06:37:32,160 CELLS THAT EXPRESS TCF7. 10497 06:37:32,160 --> 06:37:34,696 AND INDUCTIVE TRANSFER APPROACH 10498 06:37:34,696 --> 06:37:37,632 WE SHOWED THAT CARM1 DEFICIENCY 10499 06:37:37,632 --> 06:37:40,102 CELLS HAVE ENHANCED INTERTUMOR 10500 06:37:40,102 --> 06:37:43,638 IMMUNITY, SHOWING HERE AGAIN THE 10501 06:37:43,638 --> 06:37:45,507 B16 MELANOMA MODEL WITHOUT T 10502 06:37:45,507 --> 06:37:47,442 CELL TRANSFER THE TUMORS GROW 10503 06:37:47,442 --> 06:37:48,543 VERY RAPIDLY. 10504 06:37:48,543 --> 06:37:50,846 THE OT1 TLZ HAVE SOME ACTIVITY 10505 06:37:50,846 --> 06:37:52,948 IN SLOWING TUMOR GROWTH BUT THEN 10506 06:37:52,948 --> 06:37:55,884 KNOCKING OUT CARM1 SIGNIFICANTLY 10507 06:37:55,884 --> 06:37:58,820 SLOWS TUMOR GROWTH AND PROLONGS 10508 06:37:58,820 --> 06:38:01,023 SURVIVAL IN THESE MICE. 10509 06:38:01,023 --> 06:38:03,425 NOW, THE MOST INTERESTING 10510 06:38:03,425 --> 06:38:07,362 FINDING HERE WAS THAT IN 10511 06:38:07,362 --> 06:38:08,897 ACTIVATING CARM1 SUBSTANTIALLY 10512 06:38:08,897 --> 06:38:11,099 REDUCED THE EXPRESSION OF 10513 06:38:11,099 --> 06:38:13,769 INHIBITORY RECEPTORS, SO BOTH 10514 06:38:13,769 --> 06:38:16,171 PD1 AND TIM3, COMPARING HERE THE 10515 06:38:16,171 --> 06:38:18,140 CONTROL KNOCK-OUT MICE THAT HAVE 10516 06:38:18,140 --> 06:38:20,642 A SUBSTANTIAL POPULATION OF BOTH 10517 06:38:20,642 --> 06:38:25,147 PD1 AND TM3 POSITIVELY T CELLS 10518 06:38:25,147 --> 06:38:26,982 COMPARED TO CARM1 KNOCK-OUT T 10519 06:38:26,982 --> 06:38:27,983 CELLS WHERE EXPRESSION OF BOTH 10520 06:38:27,983 --> 06:38:30,218 OF THESE INHIBITORY RECEPTORS 10521 06:38:30,218 --> 06:38:32,020 WAS SUBSTANTIALLY DIMINISHED, 10522 06:38:32,020 --> 06:38:35,190 ALSO EXTENDED TO EXPRESSION OF 10523 06:38:35,190 --> 06:38:37,793 CD39 INHIBITORY RECEPTOR. 10524 06:38:37,793 --> 06:38:41,096 SO AS I MENTIONED EARLIER, IT 10525 06:38:41,096 --> 06:38:44,132 CARM1 KNOCK-OUT ACTUALLY 10526 06:38:44,132 --> 06:38:45,567 ENHANCED MAINTENANCE OF 10527 06:38:45,567 --> 06:38:48,503 RESERVOIR OF MEMORY-LIKE T CELLS 10528 06:38:48,503 --> 06:38:50,038 WITHIN TUMORS THAT IS SHOWN HERE 10529 06:38:50,038 --> 06:38:54,176 BASED ON STAINING WITH TCF7 AND 10530 06:38:54,176 --> 06:38:56,611 BCL2 DPIERD THE CONTROL 10531 06:38:56,611 --> 06:38:58,580 KNOCK-OUT T CELLS AND QUANTIFIED 10532 06:38:58,580 --> 06:38:59,681 HERE ON THE RIGHT-HAND SIDE. 10533 06:38:59,681 --> 06:39:02,284 SO WHEN WE LOOK AT GENE 10534 06:39:02,284 --> 06:39:04,486 EXPRESSION DATA, SO THIS IS 10535 06:39:04,486 --> 06:39:06,788 BLOCK RNA SEQ DATA, YOU CAN SEE 10536 06:39:06,788 --> 06:39:09,091 OF COURSE THAT CARM1 IS DOWN AND 10537 06:39:09,091 --> 06:39:11,726 THE CARM1 KNOCK-OUT T CELLS AS 10538 06:39:11,726 --> 06:39:14,362 EXPECTED, BUT THEN THERE'S UP 10539 06:39:14,362 --> 06:39:16,064 REGULATION OF MANY GENES THAT 10540 06:39:16,064 --> 06:39:20,202 ARE INVOLVED IN T CELL MEMORY, 10541 06:39:20,202 --> 06:39:27,576 FOR EXAMPLE TCF7, MYC, CD27 AND 10542 06:39:27,576 --> 06:39:32,814 IGE AND ALSO VALIDATED THESE BY 10543 06:39:32,814 --> 06:39:33,148 QPCR. 10544 06:39:33,148 --> 06:39:34,983 NOW, WHAT WE'RE DOING RIGHT NOW 10545 06:39:34,983 --> 06:39:37,018 IS SO THIS IS ALL ONGOING. 10546 06:39:37,018 --> 06:39:40,789 SO WE'RE RIGHT NOW PERFORMING 10547 06:39:40,789 --> 06:39:43,959 ATAXIC AND CUT AND WRUN THE 10548 06:39:43,959 --> 06:39:45,393 CARM1 ANTIBODY TO DEFINE THE 10549 06:39:45,393 --> 06:39:47,162 PRECISE MECHANISM OF ACTION IN 10550 06:39:47,162 --> 06:39:51,266 THE T CELLS. 10551 06:39:51,266 --> 06:39:52,834 HOWEVER, WHAT WE ALSO NOTICED IS 10552 06:39:52,834 --> 06:39:57,305 THAT CARM1 IS BROADLY EXPRESSED 10553 06:39:57,305 --> 06:39:58,507 IN MANY MANY DIFFERENT HUMAN 10554 06:39:58,507 --> 06:39:59,941 CANCERS AND IT'S ALSO AMPLIFIED 10555 06:39:59,941 --> 06:40:04,079 IN SOME CANCERS AND 10556 06:40:04,079 --> 06:40:05,213 OVEREXPRESSED. 10557 06:40:05,213 --> 06:40:06,748 FURTHERMORE, HIGH LEVELS OF 10558 06:40:06,748 --> 06:40:08,817 CARM1 NEGATIVELY CORRELATE WITH 10559 06:40:08,817 --> 06:40:09,918 SURVIVAL IN A NUMBER OF HUMAN 10560 06:40:09,918 --> 06:40:12,220 CANCER TYPES, FOR EXAMPLE, IN 10561 06:40:12,220 --> 06:40:15,490 MELANOMA, BLADDER CANCER, 10562 06:40:15,490 --> 06:40:18,994 SARCOMAS, KIDNEY CANCER AND 10563 06:40:18,994 --> 06:40:19,327 MESOTHELIOMA. 10564 06:40:19,327 --> 06:40:21,596 WHEN YOU LOOK AT THE KEY 10565 06:40:21,596 --> 06:40:23,932 PATHWAYS THAT CORRELATE WITH 10566 06:40:23,932 --> 06:40:28,303 CARM1 EXPRESSION PATTERNS, THE 10567 06:40:28,303 --> 06:40:31,239 PATHWAYS ARE MYC TARKTS AND 10568 06:40:31,239 --> 06:40:33,308 PATHWAYS RELATED TO 10569 06:40:33,308 --> 06:40:36,278 PROLIFERATION AND GLYCOLYSIS AND 10570 06:40:36,278 --> 06:40:44,386 IMPORTANTLY THE PATHWAYS THAT -- 10571 06:40:44,386 --> 06:40:46,988 THE INTERFERON RESPONSE, 10572 06:40:46,988 --> 06:40:49,291 ALLOGRAPH WHICH ACTUAL SL A TERM 10573 06:40:49,291 --> 06:40:52,794 FOR T CELL RESPONSE AND THEN 10574 06:40:52,794 --> 06:40:54,663 TNRF SIGNALLING AND THIS IS TRUE 10575 06:40:54,663 --> 06:40:56,698 FOR MULTIPLE CANCER TYPES. 10576 06:40:56,698 --> 06:40:58,800 SO WHAT WE DID THEN WAS ACTUALLY 10577 06:40:58,800 --> 06:41:02,637 KNOCK OUT CARM1 IN THE TUMOR 10578 06:41:02,637 --> 06:41:04,206 CELLS AND WE GOT A VERY 10579 06:41:04,206 --> 06:41:04,639 SURPRISING RESULT. 10580 06:41:04,639 --> 06:41:06,942 SO THE CONTROL KNOCK YOWTD 10581 06:41:06,942 --> 06:41:08,910 TUMORS ARE OF COURSE -- 10582 06:41:08,910 --> 06:41:10,779 KNOCK-OUT TUMORS OF COURSE GROW 10583 06:41:10,779 --> 06:41:15,684 RAPIDLY IN THESE MICE BUT CARM1 10584 06:41:15,684 --> 06:41:16,885 KNOCK-OUT TUMORS ACTUALLY GROW 10585 06:41:16,885 --> 06:41:18,887 VERY SLOWLY, WE ASKED IF THIS T 10586 06:41:18,887 --> 06:41:21,423 CELL TYPE IS -- CANCER TYPE IS T 10587 06:41:21,423 --> 06:41:23,992 CELL MEDIATED, WE DEPLETE IN 10588 06:41:23,992 --> 06:41:25,560 CARM1 KNOCK-OUT DISPURMS YOU CAN 10589 06:41:25,560 --> 06:41:27,395 SEE THEY GROW AGAIN VERY RAPIDLY 10590 06:41:27,395 --> 06:41:30,432 SO CARM1 KNOCK-OUT IN THE TUMOR 10591 06:41:30,432 --> 06:41:33,935 CELLS ACTUALLY IT ELICIT A T 10592 06:41:33,935 --> 06:41:36,037 CELL MEDIATED RESPONSE. 10593 06:41:36,037 --> 06:41:37,906 IN VITRO WE SEE SIMILAR 10594 06:41:37,906 --> 06:41:40,308 PHENOTYPE, THIS IS A T CELL 10595 06:41:40,308 --> 06:41:42,611 CYTOTOXICITY ASSAY FROM LEFT TO 10596 06:41:42,611 --> 06:41:44,346 RIGHT, WE ARE INCREASING THE 10597 06:41:44,346 --> 06:41:46,214 RATIO OF T CELLS TO TUMOR CELLS, 10598 06:41:46,214 --> 06:41:48,416 YOU CAN SEE THAT THE CONTROL 10599 06:41:48,416 --> 06:41:50,018 KNOCK-OUT CELLS CAN BE KILLED BY 10600 06:41:50,018 --> 06:41:52,087 T CELLS BUT YOU NEED SUBSTANTIAL 10601 06:41:52,087 --> 06:41:53,622 NUMBER OF T CELLS FOR EFFICIENT 10602 06:41:53,622 --> 06:41:56,057 KILLING, WHILE THE CARM1 10603 06:41:56,057 --> 06:41:57,158 KNOCK-OUT TUMOR CELLS ARE 10604 06:41:57,158 --> 06:42:01,029 ACTUALLY VERY SENSITIVE TO 10605 06:42:01,029 --> 06:42:02,631 CYTOTOXIC T CELLS. 10606 06:42:02,631 --> 06:42:05,066 NOW, IN 41 BREAST CANCER MODEL 10607 06:42:05,066 --> 06:42:06,501 WE FOUND THAT NOT ONLY GROWTH OF 10608 06:42:06,501 --> 06:42:09,804 THE PRIMARY TUMORS WAS REDUCED, 10609 06:42:09,804 --> 06:42:14,309 BUT ALSO THE NUMBER OF LUNG MET 10610 06:42:14,309 --> 06:42:16,444 AS CITY SEES WAS 10611 06:42:16,444 --> 06:42:20,415 SIGNIFICANTLY -- METASTESES WAS 10612 06:42:20,415 --> 06:42:21,383 SIGNIFICANTLY DIMINISHED. 10613 06:42:21,383 --> 06:42:25,420 WE OF COURSE ASKED WHETHER CARM1 10614 06:42:25,420 --> 06:42:29,691 KNOCK-OUT WOULD IMMUNIZE, THIS 10615 06:42:29,691 --> 06:42:33,828 IS RESISTANT TO ANTI-CTLA4, 10616 06:42:33,828 --> 06:42:36,131 ANTI-PD1 OR EVEN THE COMBINATION 10617 06:42:36,131 --> 06:42:41,269 OF PD1 ANDLY CTLA4 ANTIBODIES 10618 06:42:41,269 --> 06:42:43,905 AND WE FOUND THERE WAS PROFOUND 10619 06:42:43,905 --> 06:42:47,108 SYNERGY WITH ANTI-CTLA4 SO THESE 10620 06:42:47,108 --> 06:42:48,076 TUMORS GREW VERY SLOWLY AND 10621 06:42:48,076 --> 06:42:49,611 THERE WAS A VERY SUBSTANTIAL 10622 06:42:49,611 --> 06:42:52,714 SURVIVAL BENEFIT WHEN THE CARM1 10623 06:42:52,714 --> 06:42:55,650 KNOCK-OUT WAS COMBINED WITH 10624 06:42:55,650 --> 06:42:57,085 ANTI-CTLA4 COMPARED WITH THE 10625 06:42:57,085 --> 06:42:59,387 CARM1 KNOCK-OUT WITH ISOTOPE 10626 06:42:59,387 --> 06:43:00,889 CONTROL ANTIBODY. 10627 06:43:00,889 --> 06:43:03,425 WE OF COURSE WANTED TO KNOW HOW 10628 06:43:03,425 --> 06:43:05,593 THIS WAS WORKING AT A 10629 06:43:05,593 --> 06:43:07,562 MECHANISTIC LEVEL, AND SO THERE 10630 06:43:07,562 --> 06:43:10,298 WAS A VERY STRIKING PHENOTYPE IN 10631 06:43:10,298 --> 06:43:12,801 THIS RNA SEQ ANALYSIS WHERE WE 10632 06:43:12,801 --> 06:43:15,704 GREW CARM1 KNOCK-OUT OR CONTROL 10633 06:43:15,704 --> 06:43:18,306 TUMOR CELLS IN VITRO. SO AS A 10634 06:43:18,306 --> 06:43:21,376 STRIKING UP REGULATION OF MANY 10635 06:43:21,376 --> 06:43:24,112 GENES IN INTERFERON SIGNALLING 10636 06:43:24,112 --> 06:43:26,214 IN THE CARM1 KNOCK-OUT CELLS 10637 06:43:26,214 --> 06:43:28,183 DPIERTD CONTROL KNOCK-OUT AND 10638 06:43:28,183 --> 06:43:32,554 ALSO UP REGULATION OF TNF 10639 06:43:32,554 --> 06:43:32,887 SIGNALLING. 10640 06:43:32,887 --> 06:43:36,057 SO I SHOWED YOU THE LEFT PART OF 10641 06:43:36,057 --> 06:43:38,259 THE CYTOTOXICITY ASSAY EARLIER, 10642 06:43:38,259 --> 06:43:42,063 BUT THEN WHAT WE DID HERE WAS TO 10643 06:43:42,063 --> 06:43:44,699 KNOCK OUT CGAS BECAUSE WE 10644 06:43:44,699 --> 06:43:47,769 THOUGHT THAT THE CGAS THINK 10645 06:43:47,769 --> 06:43:48,870 PATHWAY COULD BE IMPORTANT IN 10646 06:43:48,870 --> 06:43:50,505 THE TYPE ONE PATHWAY INTERFERON 10647 06:43:50,505 --> 06:43:52,340 RESPONSE I JUST ILLUSTRATED, SO 10648 06:43:52,340 --> 06:43:54,676 WHEN WE KNOCK OUT CGAS, THESE 10649 06:43:54,676 --> 06:43:56,611 TUMOR CELLS ACTUALLY BECOME 10650 06:43:56,611 --> 06:43:58,813 HIGHLY, THE CARM1 KNOCK-OUT 10651 06:43:58,813 --> 06:44:00,415 TUMOR CELLS BECOME HIGHLY 10652 06:44:00,415 --> 06:44:04,119 RESISTANT TO CYTOTOXIC T CELLS. 10653 06:44:04,119 --> 06:44:08,790 NOW, FORTUNATELY, THERE IS A 10654 06:44:08,790 --> 06:44:11,092 VERY SELECTIVE IMPORTANT CARM1 10655 06:44:11,092 --> 06:44:12,193 INHIBITOR AVAILABLE THAT WAS 10656 06:44:12,193 --> 06:44:16,131 DEVELOPED BY EPIZYME, STILL IN 10657 06:44:16,131 --> 06:44:16,898 PRECLINICAL DEVELOPMENT BUT 10658 06:44:16,898 --> 06:44:19,067 EFFORT TO ADVANCE THIS TO THE 10659 06:44:19,067 --> 06:44:19,300 CLINIC. 10660 06:44:19,300 --> 06:44:23,071 SO THIS INHIBITOR HAS AN IC50 OF 10661 06:44:23,071 --> 06:44:26,408 6 UNANIMOUS MOW MOLAR AND IS 10662 06:44:26,408 --> 06:44:28,042 SELECT -- NANOMOLAR AND 10663 06:44:28,042 --> 06:44:29,477 SELECTIVE AS COMPARED TO RELATED 10664 06:44:29,477 --> 06:44:29,878 ENZYMES. 10665 06:44:29,878 --> 06:44:31,980 THIS INHIBITOR ACTUALLY INDUCES 10666 06:44:31,980 --> 06:44:33,715 SIGNIFICANT UP REGULATION OF 10667 06:44:33,715 --> 06:44:36,551 MULL PELL INTERFERONS BOTH 10668 06:44:36,551 --> 06:44:44,092 INTERFERON ALPHAS AND -- THIS 10669 06:44:44,092 --> 06:44:48,930 INHIBITOR WOULD ACTUALLY SHOW 10670 06:44:48,930 --> 06:44:49,798 IMMUNOCHECKPOINT AND I'M SHOWING 10671 06:44:49,798 --> 06:44:54,969 THIS HERE WITH EF4, SIMILAR DATA 10672 06:44:54,969 --> 06:44:57,372 ALSO WITH ANTI-PD1 AND WE FIND 10673 06:44:57,372 --> 06:45:00,341 THAT THE INHIBITOR ALONE 10674 06:45:00,341 --> 06:45:02,177 ACTUALLY HAS SIGNIFICANT 10675 06:45:02,177 --> 06:45:03,311 THERAPEUTIC ACTIVITY, BUT THERE 10676 06:45:03,311 --> 06:45:05,747 IS AGAIN PROFOUND ACTIVITY WHEN 10677 06:45:05,747 --> 06:45:09,217 WE COMBINE THIS INHIBITOR WITH 10678 06:45:09,217 --> 06:45:10,318 ANTI-CDLA4 WITH A VERY 10679 06:45:10,318 --> 06:45:14,489 SUBSTANTIAL SURVIVAL BENEFIT. 10680 06:45:14,489 --> 06:45:20,395 AGAIN, THIS IS A MODEL THAT IS 10681 06:45:20,395 --> 06:45:21,796 RESISTANT IN CHECKPOINT. 10682 06:45:21,796 --> 06:45:23,364 WE WANTED TO KNOW WHETHER THIS 10683 06:45:23,364 --> 06:45:25,200 INHIBITOR HAD TOXICITY, SO AT 10684 06:45:25,200 --> 06:45:26,968 THE SAME DOSE, AT THE SAME 10685 06:45:26,968 --> 06:45:28,903 DURATION OF TREATMENT, WE 10686 06:45:28,903 --> 06:45:30,438 TREATED MICE WITHOUT TUMORS AND 10687 06:45:30,438 --> 06:45:33,608 THEN DID A COMPLETE PATH 10688 06:45:33,608 --> 06:45:35,577 ANALYSIS AND I'M SHOWING YOU A 10689 06:45:35,577 --> 06:45:38,446 SELECTED ORGANS, THERE WERE NO 10690 06:45:38,446 --> 06:45:39,848 INFLAMMATORY CHANGES, AND THERE 10691 06:45:39,848 --> 06:45:42,784 WAS ALSO NO CHANGE IN THE BODY 10692 06:45:42,784 --> 06:45:47,188 WEIGHT REGARDLESS OF WHETHER THE 10693 06:45:47,188 --> 06:45:49,924 MICE GOT VEHICLE OR CARM1 10694 06:45:49,924 --> 06:45:50,191 INHIBITOR. 10695 06:45:50,191 --> 06:45:51,226 WHAT WAS PARTICULARLY STRIKING 10696 06:45:51,226 --> 06:45:54,863 WAS THE DEGREE OF T CELL 10697 06:45:54,863 --> 06:45:57,932 INFILTRATION. 10698 06:45:57,932 --> 06:46:02,303 TEN IS RESISTANT TO THE BLOCKADE 10699 06:46:02,303 --> 06:46:04,906 BECAUSE AT BASELINE VERY LIMITED 10700 06:46:04,906 --> 06:46:08,643 INFILTRATION BY CDA T CELLS, AS 10701 06:46:08,643 --> 06:46:12,580 YOU'RE PROBABLY AWARE, 10702 06:46:12,580 --> 06:46:14,649 ANTI-CTLA4 AND ANTI-PD1 DOES NOT 10703 06:46:14,649 --> 06:46:16,251 REALLY INCREASE INFILTRATION 10704 06:46:16,251 --> 06:46:17,352 SUBSTANTIALLY HOWEVER WHEN WE 10705 06:46:17,352 --> 06:46:19,888 LOOK AT THE CARM1 MONOTHERAPY 10706 06:46:19,888 --> 06:46:21,956 TREATED MICE YOU CAN SEE THAT 10707 06:46:21,956 --> 06:46:24,592 MORE THAN HALF ALL OF THE IMMUNE 10708 06:46:24,592 --> 06:46:25,693 CELLS IN THE TUMOR SO THIS 10709 06:46:25,693 --> 06:46:29,063 INCLUDES ALL OF THE MYELOID 10710 06:46:29,063 --> 06:46:31,266 CELLS ACTUALLY BECOME CD8 T 10711 06:46:31,266 --> 06:46:33,801 CELLS AND THIS INCREASES FURTHER 10712 06:46:33,801 --> 06:46:37,839 WHEN WE TREAT THE MICE ALSO WITH 10713 06:46:37,839 --> 06:46:41,342 ANTI-CTLA4 AND ANTI-PD1 AND THIS 10714 06:46:41,342 --> 06:46:42,243 IS QUANTIFIED HERE ON THE 10715 06:46:42,243 --> 06:46:43,411 RIGHT-HAND SIDE. 10716 06:46:43,411 --> 06:46:44,279 FURTHERMORE, THERE'S SUBSTANTIAL 10717 06:46:44,279 --> 06:46:48,783 UP REGULATION OF GRAHAM ENZYME B 10718 06:46:48,783 --> 06:46:50,451 WHICH AGAIN IN THE ISOTOPE 10719 06:46:50,451 --> 06:46:53,521 CONTROL MICE IS VERY LOW IN THE 10720 06:46:53,521 --> 06:46:55,390 CD8 T CELLS BUT BECOME 10721 06:46:55,390 --> 06:46:59,327 SUBSTANTIAL IN THE CARM1 10722 06:46:59,327 --> 06:47:01,062 INHIBITOR MONOTHERAPY OR IN THE 10723 06:47:01,062 --> 06:47:02,196 COMBINATION THERAPY. 10724 06:47:02,196 --> 06:47:05,199 WHAT IS ALSO INTERESTING IS THAT 10725 06:47:05,199 --> 06:47:06,834 CARM1 INHIBITOR TREATMENT 10726 06:47:06,834 --> 06:47:10,104 INDUCES INFLUX OF NK CELLS WHICH 10727 06:47:10,104 --> 06:47:13,107 ARE RARE IN THE BASELINE TUMORS 10728 06:47:13,107 --> 06:47:16,477 AND INDUCES RECRUITMENT OF 10729 06:47:16,477 --> 06:47:18,980 DENDRITIC CELLS AND SO DEN DRITD 10730 06:47:18,980 --> 06:47:21,683 I CAN CELL ACTUALLY PURSUING 10731 06:47:21,683 --> 06:47:23,718 RIGHT NOW AND AS A CONSEQUENCE 10732 06:47:23,718 --> 06:47:26,988 OF THESE CHANGES IN THE IMMUNE 10733 06:47:26,988 --> 06:47:29,057 MICROENVIRONMENT, THE STRIKING 10734 06:47:29,057 --> 06:47:34,195 CHANGE IN THE CD8 TO FOX P RATIO 10735 06:47:34,195 --> 06:47:36,831 WHICH USUALLY IS LOW BUT IN 10736 06:47:36,831 --> 06:47:40,001 THESE INHIBITOR TREATED MICE 10737 06:47:40,001 --> 06:47:44,606 IT'S AT APPROXIMATELY 200 OR 10738 06:47:44,606 --> 06:47:45,273 HIGHER. 10739 06:47:45,273 --> 06:47:47,976 SO THE CD8 EFFECTOR CELLS VASTLY 10740 06:47:47,976 --> 06:47:50,511 OUTNUMBER THE REGULATORY T 10741 06:47:50,511 --> 06:47:50,712 CELLS. 10742 06:47:50,712 --> 06:47:53,114 SO WHAT I'VE SHOWN YOU, I'VE 10743 06:47:53,114 --> 06:47:57,285 TRIED TO BE BRIEF, IS THAT CARM1 10744 06:47:57,285 --> 06:48:00,154 AND ACTIVATION AND CD8 CELLS 10745 06:48:00,154 --> 06:48:05,193 GREATLY ENHANCES TUMOR -- AND 10746 06:48:05,193 --> 06:48:09,731 INHIBITORY RECEPTORS AND IT 10747 06:48:09,731 --> 06:48:11,799 HELPS INTRATUMORAL RESERVOIR OF 10748 06:48:11,799 --> 06:48:12,567 MEMORY-LIKE CELLS. 10749 06:48:12,567 --> 06:48:16,304 WE'RE CURRENTLY WORKING ON A 10750 06:48:16,304 --> 06:48:18,506 MECHANISM, AND THAT IN THE TUMOR 10751 06:48:18,506 --> 06:48:20,808 CELLS, WE ALSO HAVE A STRIKING 10752 06:48:20,808 --> 06:48:23,411 EFFECT AS AN INDUCTION OF A TYPE 10753 06:48:23,411 --> 06:48:27,248 1 INTERFERON RESPONSE THAT IS 10754 06:48:27,248 --> 06:48:29,550 INDUCED BY CGAS SIGNALLING AND 10755 06:48:29,550 --> 06:48:32,420 THAT SENSITIZES THE TUMOR CELLS 10756 06:48:32,420 --> 06:48:35,657 TO T CELLS AND CARM1 SELECTIVE 10757 06:48:35,657 --> 06:48:38,960 CARM1 INHIBITOR WE SEE A 10758 06:48:38,960 --> 06:48:44,966 STRIKELY INFILTRATION OF TUMORS 10759 06:48:44,966 --> 06:48:47,635 LY BY CD8 DISPELZ NK 10760 06:48:47,635 --> 06:48:49,904 EXPRELZ CDC1 DENDRITIC CELLS. 10761 06:48:49,904 --> 06:48:52,006 THE POINT HERE IS THAT THIS 10762 06:48:52,006 --> 06:48:52,974 INHIBITOR ACTUALLY HAS DUAL 10763 06:48:52,974 --> 06:48:57,278 ACTION ON BOTH TUMOR CELLS AND T 10764 06:48:57,278 --> 06:48:59,347 CELLS, AND ONE OF THE MAJOR 10765 06:48:59,347 --> 06:49:00,882 ACTIVITIES IN THE LAB RIGHT NOW 10766 06:49:00,882 --> 06:49:03,918 IS TO ACTUALLY ALSO LOOK AT THE 10767 06:49:03,918 --> 06:49:04,986 MOLECULAR PATHWAYS THAT ARE 10768 06:49:04,986 --> 06:49:06,888 INDUCED IN TUMOR CELLS. 10769 06:49:06,888 --> 06:49:13,861 I CAN TELL YOU THAT THE R 10770 06:49:13,861 --> 06:49:17,031 RNA SEQ DATA IN TUMOR CELLS AND 10771 06:49:17,031 --> 06:49:18,966 T CELLS ACTUALLY VERY DIFFERENT 10772 06:49:18,966 --> 06:49:21,302 SO IT'S NOT THE SAME MOLECULAR 10773 06:49:21,302 --> 06:49:23,237 PATHWAY THAT'S RESPONSIBLE FOR 10774 06:49:23,237 --> 06:49:25,740 THIS DUAL EFFECT. 10775 06:49:25,740 --> 06:49:30,545 AND THE CGAS THINK PATHWAYS ALSO 10776 06:49:30,545 --> 06:49:32,847 NOT RELEVANT FOR THE IMPROVED T 10777 06:49:32,847 --> 06:49:33,548 CELL RESPONSE. 10778 06:49:33,548 --> 06:49:34,849 SO THERE ARE MANY PEOPLE I NEED 10779 06:49:34,849 --> 06:49:41,689 TO THANK HERE, SOILY SUSHUA MOST 10780 06:49:41,689 --> 06:49:44,892 OF THE WORK TOGETHER WITH CC 10781 06:49:44,892 --> 06:49:48,696 ANDLY WAHAN, DID INITIAL GENETIC 10782 06:49:48,696 --> 06:49:51,666 SCREENING, WE HAD A GREAT 10783 06:49:51,666 --> 06:49:56,471 COLLABORATION WITH SHIRLEY LIU'S 10784 06:49:56,471 --> 06:49:58,339 LAB AND COMPUTATIONAL ANALYSIS I 10785 06:49:58,339 --> 06:50:00,708 SHOWED YOU, THE GENETIC 10786 06:50:00,708 --> 06:50:02,243 LIBRARIES AND THE T CELL SCREEN 10787 06:50:02,243 --> 06:50:07,215 DONE IN COLLABORATION WITH JOHN 10788 06:50:07,215 --> 06:50:10,918 DE -- WITH JOHN D DOENCH AND 10789 06:50:10,918 --> 06:50:14,622 STEVE ELLEDGE AND ALSO A GREAT 10790 06:50:14,622 --> 06:50:17,558 COLLABORATION WITH DAVID MOONEY. 10791 06:50:17,558 --> 06:50:18,993 I THANK YOU FOR YOUR ATTENTION. 10792 06:50:18,993 --> 06:50:19,260 [APPLAUSE] 10793 06:50:19,260 --> 06:50:19,761 >> HI. 10794 06:50:19,761 --> 06:50:20,194 BACK HERE. 10795 06:50:20,194 --> 06:50:21,062 >> OKAY. 10796 06:50:21,062 --> 06:50:23,698 >> I WILL KISS THE MIC AGAIN. 10797 06:50:23,698 --> 06:50:24,999 REALLY BEAUTIFUL WORK. 10798 06:50:24,999 --> 06:50:27,101 I'M CURIOUS ABOUT YOUR CARM1 10799 06:50:27,101 --> 06:50:28,736 DATA IN THE TUMOR CELLS 10800 06:50:28,736 --> 06:50:29,036 THEMSELVES. 10801 06:50:29,036 --> 06:50:29,370 >> YES. 10802 06:50:29,370 --> 06:50:32,140 >> AND ESSENTIALLY ACTIVATING 10803 06:50:32,140 --> 06:50:33,107 THE C GAS PATHWAY. 10804 06:50:33,107 --> 06:50:36,444 DOES IT INDUCE CHROMOSOME 10805 06:50:36,444 --> 06:50:37,779 INSTABILITY IN MICRONUCLEI IF 10806 06:50:37,779 --> 06:50:40,481 YOU HAVE THE INHIBITOR IS THAT 10807 06:50:40,481 --> 06:50:42,316 WHAT'S GOING OR WHAT IS 10808 06:50:42,316 --> 06:50:43,251 MECHANISTICALLY GOING ON? 10809 06:50:43,251 --> 06:50:45,887 >> LIKE VIJAY I CUT OUT A LOT OF 10810 06:50:45,887 --> 06:50:46,120 SLIDES. 10811 06:50:46,120 --> 06:50:47,188 I COULD HAVE TALKED FOR A LONG 10812 06:50:47,188 --> 06:50:48,289 TIME. 10813 06:50:48,289 --> 06:50:48,756 YEAH. 10814 06:50:48,756 --> 06:50:51,592 SO THERE IS EXACTLY, THERE IS 10815 06:50:51,592 --> 06:50:55,630 DNA DAMAGE AND YOU KNOW WITH 10816 06:50:55,630 --> 06:51:00,334 GAMMA H2X WE CAN SEE DNA DAMAGE 10817 06:51:00,334 --> 06:51:03,404 AND WE ALSO CAN VISUALIZE 10818 06:51:03,404 --> 06:51:05,573 CYTOPLASMIC MIE YOA NUCLEI AND 10819 06:51:05,573 --> 06:51:07,341 THAT ACTIVATES THE C GAS 10820 06:51:07,341 --> 06:51:09,310 PATHWAY. 10821 06:51:09,310 --> 06:51:10,511 THAT DOES NOT HAPPEN IN THE T 10822 06:51:10,511 --> 06:51:11,946 CELLS, WE LOOKED AT IT VERY 10823 06:51:11,946 --> 06:51:12,213 CAREFULLY. 10824 06:51:12,213 --> 06:51:14,115 THERE IS NO DNA DAMAGE MT T 10825 06:51:14,115 --> 06:51:14,315 CELLS. 10826 06:51:14,315 --> 06:51:15,917 SO THE MECHANISM OF ACTION IN T 10827 06:51:15,917 --> 06:51:20,721 CELLS AND TUMOR CELLS IS THE 10828 06:51:20,721 --> 06:51:20,922 SAME. 10829 06:51:20,922 --> 06:51:24,759 >> I'M ELLEN ROTHENBURG. 10830 06:51:24,759 --> 06:51:25,426 ONE OF THE THINGS INTERESTING 10831 06:51:25,426 --> 06:51:27,862 ABOUT THIS IS WHICH NORMAL CELLS 10832 06:51:27,862 --> 06:51:29,397 EXPRESS CARM1 AT REASONABLE 10833 06:51:29,397 --> 06:51:31,032 LEVELS, I WAS CHECK MGEN AND 10834 06:51:31,032 --> 06:51:32,366 IT'S PRETTY LOW IN T CELLS 10835 06:51:32,366 --> 06:51:33,568 ACCORDING TO THE RNA. 10836 06:51:33,568 --> 06:51:35,136 SO WHERE ELM IN THE NORMAL BODY 10837 06:51:35,136 --> 06:51:37,438 IS THIS INHIBITOR GOING TO BE 10838 06:51:37,438 --> 06:51:41,008 HAVING AN UNSCHEDULED EFFECT? 10839 06:51:41,008 --> 06:51:43,644 >> SO THAT'S PRETTY BROADLY 10840 06:51:43,644 --> 06:51:45,746 EXPRESSED BECAUSE IT'S A 10841 06:51:45,746 --> 06:51:47,148 TRANSCRIPTIONAL ACTIVATOR FOR A 10842 06:51:47,148 --> 06:51:51,118 NUMBER OF TRANSCRIPTION FACTORS. 10843 06:51:51,118 --> 06:51:53,721 IN T CELLS IT'S ACTUALLY 10844 06:51:53,721 --> 06:51:55,056 STRONGLY UP REGULATED, DIDN'T 10845 06:51:55,056 --> 06:51:57,592 SHOW YOU THAT, IT'S STRONGLY UP 10846 06:51:57,592 --> 06:51:59,093 REGULATED FOLLOWING ACTIVATION, 10847 06:51:59,093 --> 06:52:01,295 ACTUALLY VERY EARLY FOLLOWING 10848 06:52:01,295 --> 06:52:02,897 ACTIVATION. 10849 06:52:02,897 --> 06:52:04,765 AND THEN WE REACTIVATE THE T 10850 06:52:04,765 --> 06:52:06,300 CELLS IT GOES UP FURTHER SO WE 10851 06:52:06,300 --> 06:52:09,704 THINK IT ACCUMULATES SORT OF 10852 06:52:09,704 --> 06:52:11,439 WITH REPETITIVE T CELL 10853 06:52:11,439 --> 06:52:11,739 STIMULATION. 10854 06:52:11,739 --> 06:52:13,207 >> I GUESS I'M WONDERING ABOUT 10855 06:52:13,207 --> 06:52:14,942 GETTING RID OF SOME OF THESE 10856 06:52:14,942 --> 06:52:18,646 PROTECTIVE MECHANISMS THAT ARE 10857 06:52:18,646 --> 06:52:19,981 NORMALLY WORK AGAINST 10858 06:52:19,981 --> 06:52:20,314 AUTOIMMUNITY. 10859 06:52:20,314 --> 06:52:22,250 IS THERE ANY POSSIBILITY THAT 10860 06:52:22,250 --> 06:52:25,119 IT'S GOING TO SENSITIZE TO 10861 06:52:25,119 --> 06:52:26,087 AUTOIMMUNE ATTACK? 10862 06:52:26,087 --> 06:52:29,857 >> IN THE MICE, AS I SAID, THE 10863 06:52:29,857 --> 06:52:31,792 TOXICITY EXPERIMENT WAS DONE FOR 10864 06:52:31,792 --> 06:52:33,661 THE SAME DURATION AND AT THE 10865 06:52:33,661 --> 06:52:36,264 SAME DOSE, AND WE DIDN'T SEE ANY 10866 06:52:36,264 --> 06:52:38,799 INFLAMMATORY LESIONS ANYWAY. 10867 06:52:38,799 --> 06:52:40,768 SO BUT YOU BRING UP A GREAT 10868 06:52:40,768 --> 06:52:41,202 POINT. 10869 06:52:41,202 --> 06:52:42,837 SO, YOU KNOW, HOW DO YOU -- I 10870 06:52:42,837 --> 06:52:46,440 MEAN, WITH ALL IMMUNOTHERAPY 10871 06:52:46,440 --> 06:52:48,843 DRUGS THERE IS IMMUNE RELATED 10872 06:52:48,843 --> 06:52:50,645 TOXICITY HAD, AND THE QUESTION S 10873 06:52:50,645 --> 06:52:52,046 HOW TO MANAGE IT. 10874 06:52:52,046 --> 06:52:54,815 SO WITH THIS TYPE OF DRUG, WE 10875 06:52:54,815 --> 06:52:56,751 MAY ACTUALLY NOT WANT TO GIVE IT 10876 06:52:56,751 --> 06:52:57,618 CONTINUOUSLY FOR A LONG PERIOD 10877 06:52:57,618 --> 06:52:59,053 OF TIME, BUT YOU MAY WANT TO 10878 06:52:59,053 --> 06:53:01,589 GIVE IT AT ENTER VAMS BECAUSE 10879 06:53:01,589 --> 06:53:03,424 ACTUALLY IN ACUTE INTERFERON 10880 06:53:03,424 --> 06:53:05,526 RESPONSE MAY BE MORE BENEFICIAL 10881 06:53:05,526 --> 06:53:06,928 THAN SOME OF LIKE A CHRONIC 10882 06:53:06,928 --> 06:53:09,230 RESPONSE WHICH CAN INDUCE 10883 06:53:09,230 --> 06:53:09,497 NEGATIVE. 10884 06:53:09,497 --> 06:53:13,801 >> THANKS VERY MUCH. 10885 06:53:13,801 --> 06:53:15,503 >> GREAT TALK. 10886 06:53:15,503 --> 06:53:16,704 I HAVE TWO QUESTIONS. 10887 06:53:16,704 --> 06:53:18,339 ONE WAS A LITTLE BIT OF A 10888 06:53:18,339 --> 06:53:19,407 FOLLOW-UP FROM WHAT YOUR 10889 06:53:19,407 --> 06:53:20,541 RESPONSE WAS TO ALAN. 10890 06:53:20,541 --> 06:53:23,311 SO IS CARM1 UP REGULATED IN 10891 06:53:23,311 --> 06:53:24,145 EXHAUSTED CELLS AND THEREFORE 10892 06:53:24,145 --> 06:53:26,547 IT'S A BARRIER? 10893 06:53:26,547 --> 06:53:28,883 IS THAT PART OF THE EXHAUSTION 10894 06:53:28,883 --> 06:53:30,184 PROCESS AND IF SO, WHAT'S 10895 06:53:30,184 --> 06:53:31,252 DRIVING THAT? 10896 06:53:31,252 --> 06:53:33,421 SECONDLY, WHAT IS CARM1 ACTUALLY 10897 06:53:33,421 --> 06:53:36,090 DOING THAT MOD LAITSZ T CELL 10898 06:53:36,090 --> 06:53:37,358 FUNCTION -- MODULATES T CELL 10899 06:53:37,358 --> 06:53:38,559 FUNCTION ASK MIGRATION? 10900 06:53:38,559 --> 06:53:39,927 IS IT SORT OF GENERAL RELEASING 10901 06:53:39,927 --> 06:53:41,562 THE BRAKES SO EVERYTHING GOES 10902 06:53:41,562 --> 06:53:43,331 UP, METABOLISM, ACTIVATION, ET 10903 06:53:43,331 --> 06:53:44,532 CETERA, OR IS IT MORE SELECTIVE 10904 06:53:44,532 --> 06:53:47,935 IN TERMS OF WHAT IT'S TARGETING? 10905 06:53:47,935 --> 06:53:50,237 >> SO THE MOLECULAR MECHANISM 10906 06:53:50,237 --> 06:53:53,174 WITHIN THE ACTIVE INVESTIGATION 10907 06:53:53,174 --> 06:53:58,145 SO WE HAVE SOME ATAXIC DATA. 10908 06:53:58,145 --> 06:54:02,783 SO IN P THE KNOCK-OUT CELLS WE 10909 06:54:02,783 --> 06:54:05,586 SEE GREATER ACCESSIBILITY AT 10910 06:54:05,586 --> 06:54:10,091 GENES THAT ARE -- AT SITES WHERE 10911 06:54:10,091 --> 06:54:11,625 AP1 TRANSCRIPTION FACTORS BIND. 10912 06:54:11,625 --> 06:54:16,063 SO WE THINK THE KNOCK-OUT 10913 06:54:16,063 --> 06:54:19,767 ENHANCES ACTIVATION. 10914 06:54:19,767 --> 06:54:23,504 BUT SO, YOU KNOW, WHAT WE'RE 10915 06:54:23,504 --> 06:54:26,007 STILL WORKING ON, THE CUT AND 10916 06:54:26,007 --> 06:54:33,347 RUN DATA, I MEAN WE HAVE SOME 10917 06:54:33,347 --> 06:54:36,851 INITIAL -- WE HAVE SOME SPECIFIC 10918 06:54:36,851 --> 06:54:39,453 HYPOTHESES AS TO WHAT THE TARGET 10919 06:54:39,453 --> 06:54:42,089 GENES ARE, BUT THOSE EXPERIMENTS 10920 06:54:42,089 --> 06:54:43,190 STILL NEED TO BE DONE. 10921 06:54:43,190 --> 06:54:45,493 SO I REALLY DON'T KNOW. 10922 06:54:45,493 --> 06:54:45,693 YEAH. 10923 06:54:45,693 --> 06:54:46,394 AT THIS POINTED. 10924 06:54:46,394 --> 06:54:48,662 >> FIRST QUESTION IS IS THIS 10925 06:54:48,662 --> 06:54:50,664 SPECIFICALLY UP REGULATED IN 10926 06:54:50,664 --> 06:54:51,032 EXHAUSTION? 10927 06:54:51,032 --> 06:54:52,600 >> BUT THE SPREKS IS CLEARLY 10928 06:54:52,600 --> 06:54:55,036 RELATED TO T CELL ACTIVATION, 10929 06:54:55,036 --> 06:54:57,038 AND AS I MENTIONED IN MY 10930 06:54:57,038 --> 06:54:58,973 RESPONSE TO ELLEN, SO WITH 10931 06:54:58,973 --> 06:55:01,275 REPETITIVE STIMULATION, THE 10932 06:55:01,275 --> 06:55:02,576 EXPRESSION GOES UP. 10933 06:55:02,576 --> 06:55:06,113 SO WE THINK IT'S ONE OF THE 10934 06:55:06,113 --> 06:55:08,616 ENZYMES THAT'S RELATED TO T CELL 10935 06:55:08,616 --> 06:55:10,451 EXHAUSTION, SO IN A BROADER 10936 06:55:10,451 --> 06:55:10,651 SENSE. 10937 06:55:10,651 --> 06:55:11,252 >> THANKS. 10938 06:55:11,252 --> 06:55:12,887 >> OKAY. 10939 06:55:12,887 --> 06:55:14,622 >> HI. 10940 06:55:14,622 --> 06:55:17,258 IS THIS ON? 10941 06:55:17,258 --> 06:55:17,458 HELLO? 10942 06:55:17,458 --> 06:55:18,559 CAN YOU HEAR ME? 10943 06:55:18,559 --> 06:55:19,193 >> YES. 10944 06:55:19,193 --> 06:55:19,994 >> OKAY. 10945 06:55:19,994 --> 06:55:21,195 >> GO AHEAD. 10946 06:55:21,195 --> 06:55:23,164 >> FROM NAID. 10947 06:55:23,164 --> 06:55:24,598 WONDERFUL TALK. 10948 06:55:24,598 --> 06:55:26,333 THINKING ABOUT MAYBE THERE IS A 10949 06:55:26,333 --> 06:55:29,403 COMMON MECHANISM IN TUMOR VERSUS 10950 06:55:29,403 --> 06:55:32,273 IN CD8'S, HAVE YOU CAREFULLY 10951 06:55:32,273 --> 06:55:37,511 CHECKED THE CARM1 KNOCKOUTS CD8S 10952 06:55:37,511 --> 06:55:38,813 WHETHER THEY HAVE ANY 10953 06:55:38,813 --> 06:55:39,914 PROLIFERATION, I WAS WONDERING 10954 06:55:39,914 --> 06:55:41,215 IF SLOWING DOWN THE 10955 06:55:41,215 --> 06:55:43,084 PROLIFERATION OF CD8 THAT THEY 10956 06:55:43,084 --> 06:55:44,952 MAY RE RETAIN SOME STEM LIKE 10957 06:55:44,952 --> 06:55:46,353 FEATURE SO THEY CAN ACTUALLY 10958 06:55:46,353 --> 06:55:48,989 RESIST TO EXHAUSTION AND 10959 06:55:48,989 --> 06:55:50,791 EX-ZOOND I CAN TELL YOU THAT -- 10960 06:55:50,791 --> 06:55:53,594 >> I CAN TELL YOU THAT IN VIVO 10961 06:55:53,594 --> 06:55:54,929 TUMOR CELLS AND T CELLS ISN'T 10962 06:55:54,929 --> 06:55:56,230 RELATED TO PROLIFERATION. 10963 06:55:56,230 --> 06:56:00,501 SO IF YOU SORT OF RECALL HOW WE 10964 06:56:00,501 --> 06:56:03,671 INITIALLY FOUND IT WAS ACTUALLY 10965 06:56:03,671 --> 06:56:05,940 BY LOOKING AT T CELLS THAT 10966 06:56:05,940 --> 06:56:07,374 ACCUMULATED TUMORS, RIGHT? 10967 06:56:07,374 --> 06:56:10,244 SO IT'S ACTUALLY THE T CELLS 10968 06:56:10,244 --> 06:56:13,848 ACCUMULATE BETTER IN TUMORS WHEN 10969 06:56:13,848 --> 06:56:15,916 WE INACTIVATE CARM1 AND WE'VE 10970 06:56:15,916 --> 06:56:17,718 LOOKED VERY CAREFUL AT TUMOR 10971 06:56:17,718 --> 06:56:20,654 CELL PROLIFERATION IN VITRO AND 10972 06:56:20,654 --> 06:56:22,523 THERE'S ABSOLUTELY NO DIFFERENCE 10973 06:56:22,523 --> 06:56:24,258 IN TUMOR CELL PROLIFERATION, SO 10974 06:56:24,258 --> 06:56:25,259 THEANT THE MECHANISM. 10975 06:56:25,259 --> 06:56:26,927 SO WE THINK THAT -- SO THAT'S 10976 06:56:26,927 --> 06:56:30,331 NOT THE MECHANISM. SO WE THINK 10977 06:56:30,331 --> 06:56:31,932 THAT TUMOR CELLS HAVE DEFECTS IN 10978 06:56:31,932 --> 06:56:34,835 DNA REPAIR PATHWAYS, AND WE 10979 06:56:34,835 --> 06:56:37,705 THINK THAT CARM1 IS INVOLVED AS 10980 06:56:37,705 --> 06:56:40,107 A CO-TRANSCRIPTIONAL ACTIVATOR 10981 06:56:40,107 --> 06:56:42,610 OF DNA REPAIR PATHWAYS, BUT THE 10982 06:56:42,610 --> 06:56:46,013 DETAILS NEED TO BE WORKED OUT. 10983 06:56:46,013 --> 06:56:47,581 >> LAST QUESTION. 10984 06:56:47,581 --> 06:56:52,153 >> SO YOU MENTIONED MOST LIKELY 10985 06:56:52,153 --> 06:56:53,854 LTS RELATED IN THE TUMORS -- IT 10986 06:56:53,854 --> 06:56:55,656 IS RELATED, IN THE TUMORS IT IS 10987 06:56:55,656 --> 06:56:58,859 RELATED TO DNA DAMAGE REPAIR, 10988 06:56:58,859 --> 06:57:02,029 BUT HAVE YOU EXPLORED THE 10989 06:57:02,029 --> 06:57:06,500 POSSIBILITY OF A CHANGE IN THE 10990 06:57:06,500 --> 06:57:09,603 EXPRESSION OF MHC ONE BECAUSE I 10991 06:57:09,603 --> 06:57:11,338 KNOW MANY TIMES FREQUENTLY IN 10992 06:57:11,338 --> 06:57:13,307 BASIC CELLS THERE IS DOWN 10993 06:57:13,307 --> 06:57:16,443 REGULATION OF MHC ONE SO I'M 10994 06:57:16,443 --> 06:57:18,279 WONDERING IF IN THE KNOCK-OUT 10995 06:57:18,279 --> 06:57:23,584 YOU CAUSE THE BE -- THERE ARE 10996 06:57:23,584 --> 06:57:25,553 MORE DETECTABLE BY THE T CELLS? 10997 06:57:25,553 --> 06:57:26,754 >> ACTUALLY WHAT HAPPENS SUSPECT 10998 06:57:26,754 --> 06:57:29,490 THAT THE CARM1 KNOCK-OUT CELLS 10999 06:57:29,490 --> 06:57:33,194 HAVE THIS ENDOGENOUS TYPE 1LY 11000 06:57:33,194 --> 06:57:34,862 INTERFERON RESPONSE AND WE'VE 11001 06:57:34,862 --> 06:57:37,031 SHOWN THAT THE KNOCK-OUT CELLS 11002 06:57:37,031 --> 06:57:39,433 ARE VERY SENSITIVE TO GAMG MA 11003 06:57:39,433 --> 06:57:40,968 INTERFERON DUE TO THIS 11004 06:57:40,968 --> 06:57:43,404 ENDOGENOUS TYPE OF INTERFERON 11005 06:57:43,404 --> 06:57:44,138 SIGNALLING. 11006 06:57:44,138 --> 06:57:47,641 SO ENDOGENOUS TYPE 1 INTERFERON 11007 06:57:47,641 --> 06:57:49,810 SIGNALLING SENSITIZES THEM TO 11008 06:57:49,810 --> 06:57:51,445 GAMMA INTERFERON AND UP 11009 06:57:51,445 --> 06:57:53,647 REGULATED EVEN WITH LOW LEVEL OF 11010 06:57:53,647 --> 06:57:55,082 GAM GAMMA INTERFERON, THAT'S HOW 11011 06:57:55,082 --> 06:57:56,050 IT WORKS. 11012 06:57:56,050 --> 06:57:57,585 THAT EXPLAINS SORT OF THE 11013 06:57:57,585 --> 06:58:01,522 HEIGHTENED SENSITIVITY IN T CELL 11014 06:58:01,522 --> 06:58:02,489 KILLING HAD. 11015 06:58:02,489 --> 06:58:03,490 >> GREAT. 11016 06:58:03,490 --> 06:58:04,825 THAT'S REALLY COOL. 11017 06:58:04,825 --> 06:58:05,059 THANKS. 11018 06:58:05,059 --> 06:58:07,661 >> OKAY. 11019 06:58:07,661 --> 06:58:08,362 [APPLAUSE] 11020 06:58:08,362 --> 06:58:12,333 >> THANK YOU. 11021 06:58:12,333 --> 06:58:14,435 >> SO THANK YOU ALL FOR STAYING 11022 06:58:14,435 --> 06:58:17,071 UNTIL THE VERY END, ALSO FOR THE 11023 06:58:17,071 --> 06:58:18,505 SPEEKS, JUST SOME CLOSING 11024 06:58:18,505 --> 06:58:18,839 REMARKS. 11025 06:58:18,839 --> 06:58:20,808 TOMORROW WE ARE GOING TO START 11026 06:58:20,808 --> 06:58:23,644 AT 8:30 WITH A BANG, JIM DE 11027 06:58:23,644 --> 06:58:25,512 SANTO WILL BE OUR FIRST SPEAKER, 11028 06:58:25,512 --> 06:58:26,780 AMAZING SPEAKER, PLEASE COME 11029 06:58:26,780 --> 06:58:33,387 BIVMENT OUR SESSION CHAIRS FOR 11030 06:58:33,387 --> 06:58:35,155 THE FIRST SESSION, WE START AT 11031 06:58:35,155 --> 06:58:35,456 8:30. 11032 06:58:35,456 --> 06:58:36,657 IF YOU ARE LATE, YOU ARE MIRING 11033 06:58:36,657 --> 06:58:38,092 A LOT OF THINGS, SO PLEASE BE 11034 06:58:38,092 --> 06:58:40,227 HERE AT 8:30 AND WE'LL SEE YOU. 11035 06:58:40,227 --> 06:58:40,961 THANK YOU VERY MUCH. 11036 06:58:40,961 --> 06:58:50,961 SEE YOU TOMORROW.