1 00:00:04,723 --> 00:00:06,758 OKAY, SO, THANK YOU EVERYONE 2 00:00:06,758 --> 00:00:10,362 WHO WAS BRAVE ENOUGH TO COME 3 00:00:10,362 --> 00:00:12,631 HERE DESPITE THE RAIN AND THANK 4 00:00:12,631 --> 00:00:16,501 YOU TO EVERYONE WHO IS JOINING 5 00:00:16,501 --> 00:00:16,969 US ONLINE. 6 00:00:16,969 --> 00:00:19,004 I GUESS YOU MIGHT BE ABLE TO 7 00:00:19,004 --> 00:00:24,710 HEAR US, WE DEFINITELY CANNOT 8 00:00:24,710 --> 00:00:25,410 HEAR YOU. 9 00:00:25,410 --> 00:00:28,747 AND TODAY IT'S MY PLEASURE TO 10 00:00:28,747 --> 00:00:36,655 INTRODUCE TIM PETROS FROM NIHC-- 11 00:00:36,655 --> 00:00:37,022 NICHD, SORRY. 12 00:00:37,022 --> 00:00:42,461 HE IS OUR NLM GROUP SPEAKER AND 13 00:00:42,461 --> 00:00:45,264 MANY OF US DO KNOW TIM AS AN 14 00:00:45,264 --> 00:00:49,968 ACTIVE CONTRIBUTOR TO THE NLM 15 00:00:49,968 --> 00:00:50,502 [INDISCERNIBLE] PROGRAM. 16 00:00:50,502 --> 00:00:55,007 HE HELP US A LOT WITH HIS WORK 17 00:00:55,007 --> 00:00:57,876 AND INSIGHTS IN THE SUMMER AND 18 00:00:57,876 --> 00:01:01,179 FALL AND GOING TO THE BACKGROUND 19 00:01:01,179 --> 00:01:05,017 TIM EARNED HIS Ph.D. IN 20 00:01:05,017 --> 00:01:05,918 COLUMBIA IN 2009. 21 00:01:05,918 --> 00:01:07,986 AFTER THAT HE WAS A POST 22 00:01:07,986 --> 00:01:14,760 DOCTORAL FELLOW AT CORNELL AND 23 00:01:14,760 --> 00:01:19,097 NYU AND IN 2007 -- IS EVERYTHING 24 00:01:19,097 --> 00:01:19,298 OKAY? 25 00:01:19,298 --> 00:01:23,669 -- IN 2007 NIH IN PARTICULAR, 26 00:01:23,669 --> 00:01:31,310 NICHD HAS BEEN VERY FORTUNATE TO 27 00:01:31,310 --> 00:01:34,846 BRING HIM ON BOARD AS A SENIOR 28 00:01:34,846 --> 00:01:37,082 TRACK INVESTIGATOR AND TIM'S 29 00:01:37,082 --> 00:01:41,887 WORK WHICH HAS STARTED I BELIEVE 30 00:01:41,887 --> 00:01:43,388 IS USING MASS 31 00:01:43,388 --> 00:01:44,656 SPCERTAINLY--CERTAINLY C AND HE 32 00:01:44,656 --> 00:01:47,225 HAS EXPANDED INTO THE FIELD OF 33 00:01:47,225 --> 00:01:49,361 GENETIC AND EPIGENERATEDETEC 34 00:01:49,361 --> 00:01:52,264 MECHANISMS THAT REGULATE EARLY 35 00:01:52,264 --> 00:01:52,965 INTERNEURON DECISIONS DURING 36 00:01:52,965 --> 00:01:56,601 NEUROGENESIS AND THE RESULTS OF 37 00:01:56,601 --> 00:01:59,004 TIM'S GROUP'S WORK HAS BEEN PUB 38 00:01:59,004 --> 00:02:06,211 LESHED IN A LOT OF LEADING 39 00:02:06,211 --> 00:02:07,646 JOURNALS IN NEURON, AND SEVERAL 40 00:02:07,646 --> 00:02:07,879 OTHERS. 41 00:02:07,879 --> 00:02:12,117 IN ADDITION TO THAT, TIM 42 00:02:12,117 --> 00:02:13,285 RECEIVED MULTIPLE AWARDS 43 00:02:13,285 --> 00:02:14,820 INCLUDING THE DIVISION OF 44 00:02:14,820 --> 00:02:18,590 INTRAMURAL RESEARCH OF NICHD 45 00:02:18,590 --> 00:02:19,758 SCIENTIFIC DIRECTOR'S AWARD, 46 00:02:19,758 --> 00:02:21,259 MENTOR AWARDS, ET CETERA, ET 47 00:02:21,259 --> 00:02:21,593 CETERA. 48 00:02:21,593 --> 00:02:28,133 THE LIST IS WAY TO ALONG TO 49 00:02:28,133 --> 00:02:32,070 MENTION JUST ENOUGH, THE TITLE 50 00:02:32,070 --> 00:02:34,139 OF TIM'S WALK IS MECHANISMS 51 00:02:34,139 --> 00:02:37,509 REGULATING FETE AND MATURATION 52 00:02:37,509 --> 00:02:38,310 OF FOREBRAIN INHIBITORY 53 00:02:38,310 --> 00:02:39,144 INTERNEURONS AND BEFORE WE JUMP 54 00:02:39,144 --> 00:02:40,345 INTO THAT I WOULD LIKE TO 55 00:02:40,345 --> 00:02:46,385 BRIEFLY TALK TO THOSE OF US WHO 56 00:02:46,385 --> 00:02:47,085 ARE ATTENDING VIRTUALLY -- I 57 00:02:47,085 --> 00:02:49,287 HOPE CAN YOU HEAR ME OR YOU WILL 58 00:02:49,287 --> 00:02:50,722 BE ABLE TO SEE THE PRESENTATION 59 00:02:50,722 --> 00:02:52,724 EMPLOY IF YOU HAVE ANY QUESTIONS 60 00:02:52,724 --> 00:02:55,127 PLEASE DO SEND THEM TO THE 61 00:02:55,127 --> 00:02:56,995 E-MAIL ADDRESS THAT HOPEFULLY 62 00:02:56,995 --> 00:03:03,835 WILL BE PROVIDED TO YOU BY THE 63 00:03:03,835 --> 00:03:04,936 VIDEO CHAT FUNCTION, BUT IF YOU 64 00:03:04,936 --> 00:03:06,838 SEE THE E-MAIL ADDRESS, PLEASE 65 00:03:06,838 --> 00:03:08,607 FEEL FREE TO SEND YOUR QUESTIONS 66 00:03:08,607 --> 00:03:11,476 THERE, IF NOT YOU CAN ALSO USE 67 00:03:11,476 --> 00:03:14,646 THE FEEDBACK BUTTON ON THE 68 00:03:14,646 --> 00:03:15,247 VIDEOCAST PAGE. 69 00:03:15,247 --> 00:03:16,281 THOSE QUESTIONS WILL BE 70 00:03:16,281 --> 00:03:21,486 DELIVERED TO ME SOMEHOW AND I 71 00:03:21,486 --> 00:03:24,189 WILL ASK TIM THE QUESTIONS AT 72 00:03:24,189 --> 00:03:25,357 END OF HIS PRESENTATION. 73 00:03:25,357 --> 00:03:26,792 ALSO, IF YOU ARE INTERESTED IN 74 00:03:26,792 --> 00:03:28,093 JOINING US FOR LUNCH AT NOON, 75 00:03:28,093 --> 00:03:29,728 YOU ARE MORE THAN WELCOME. 76 00:03:29,728 --> 00:03:32,931 WE WILL GO TO THE CAFETERIA IN 77 00:03:32,931 --> 00:03:38,203 NATCHER WHICH IS FORTUNATELY 78 00:03:38,203 --> 00:03:38,804 OPEN TODAY. 79 00:03:38,804 --> 00:03:40,439 WITH THAT, IT IS MY PLEASURE TO 80 00:03:40,439 --> 00:03:47,712 WELCOME TIM TO OUR NLM GROUP 81 00:03:47,712 --> 00:03:52,017 TALK EMPLOY. 82 00:03:52,017 --> 00:03:53,518 >> OKAY, THANK YOU FOR THAT KIND 83 00:03:53,518 --> 00:03:54,719 INTRODUCTION, IT'S NICE TO SEE 84 00:03:54,719 --> 00:03:56,822 SOME FAMILIAR FACES HERE, I 85 00:03:56,822 --> 00:03:59,324 REALLY ENJOYED MY TIME ON THE 86 00:03:59,324 --> 00:04:00,358 TENURED TRACK SEARCH COMMITTEE. 87 00:04:00,358 --> 00:04:03,095 I LEARNED A LOT ABOUT NLM AND 88 00:04:03,095 --> 00:04:04,529 WHAT YOU GUYS DO AS AN 89 00:04:04,529 --> 00:04:05,931 ORGANIZATION AND AGAIN, I AM 90 00:04:05,931 --> 00:04:08,533 VERY HAPPY TO PRESENT MY WORK 91 00:04:08,533 --> 00:04:08,733 HERE. 92 00:04:08,733 --> 00:04:11,236 SO AS WAS SAID, MY LAB FOCUSES 93 00:04:11,236 --> 00:04:13,538 ON MECHANISMS THAT REGULATE FATE 94 00:04:13,538 --> 00:04:16,274 AND MATURATION OF GABBAERGIC 95 00:04:16,274 --> 00:04:17,676 INHIBITORY NEURONS AND THIS IS 96 00:04:17,676 --> 00:04:20,112 WOB OF MY MANY FAILED COVER 97 00:04:20,112 --> 00:04:21,746 IMAGE SUBMISSIONS BUT BASICALLY, 98 00:04:21,746 --> 00:04:24,950 ON THE LEFT, IS A SECTION 99 00:04:24,950 --> 00:04:25,584 THROUGH THE EMBRYONIC MOUSE 100 00:04:25,584 --> 00:04:27,419 BRAIN, ON THE RIGHT IS A SECTION 101 00:04:27,419 --> 00:04:28,920 THROUGH THE ADULT MOUSE BRAIN 102 00:04:28,920 --> 00:04:30,088 AND SO MY LAB TRIES TO 103 00:04:30,088 --> 00:04:32,290 UNDERSTAND HOW YOU GO FROM THIS 104 00:04:32,290 --> 00:04:34,392 SMALL POPULATION OF CELLS IN THE 105 00:04:34,392 --> 00:04:36,962 EMBRYO, TO GENERATE THIS 106 00:04:36,962 --> 00:04:39,164 INCREDIBLY DIVERSE POPULATION OF 107 00:04:39,164 --> 00:04:40,298 GABBAERGIC CELLS THAT POPULATE A 108 00:04:40,298 --> 00:04:43,268 VARIETY OF BRAIN REGIONS NEAR 109 00:04:43,268 --> 00:04:48,140 CRITICAL AND EVERY ASPECT OF 110 00:04:48,140 --> 00:04:48,807 BRAIN FUNCTION. 111 00:04:48,807 --> 00:04:50,575 SO IT GOES WITHOUT SAYING THAT 112 00:04:50,575 --> 00:04:52,244 BRAIN DEVELOPMENT ACCIDENT IN 113 00:04:52,244 --> 00:04:53,912 PARTICULAR BRAIN DEVELOPMENT IS 114 00:04:53,912 --> 00:04:55,680 AN INCREDIBLY COMPLEX AND NEURAL 115 00:04:55,680 --> 00:04:56,982 PROCESS STARTING WITH THE NEURAL 116 00:04:56,982 --> 00:04:58,717 TUBE STRUCTURE AND THEN OVER 117 00:04:58,717 --> 00:05:00,218 MONTHS AND YEARS FORMING THIS 118 00:05:00,218 --> 00:05:02,821 COMPLEX STRUCTURE THAT WE ALL 119 00:05:02,821 --> 00:05:04,589 TRY TO UNDERSTAND AND HOW YOU 120 00:05:04,589 --> 00:05:05,690 GENERATE THIS COMPLEXITY AND 121 00:05:05,690 --> 00:05:08,126 DIVERSITY, I THINK IS 1 OF THE 122 00:05:08,126 --> 00:05:10,262 MOST EXCITING AND CHALLENGING 123 00:05:10,262 --> 00:05:10,729 QUESTIONS IN BIOLOGY. 124 00:05:10,729 --> 00:05:14,466 NOW IF YOU LOOK AT COMPAREISON 125 00:05:14,466 --> 00:05:16,101 OF SPECIES OF DIFFERENT BRAINS, 126 00:05:16,101 --> 00:05:17,969 YOU CAN SAY UP HERE IS THE MOUSE 127 00:05:17,969 --> 00:05:20,705 BRAIN WE USE TO MODEL SYSTEM AND 128 00:05:20,705 --> 00:05:22,440 OVER TIME YOU EVENTUALLY 129 00:05:22,440 --> 00:05:24,743 EVOLUTION AS GIVEN RISE TO HUMAN 130 00:05:24,743 --> 00:05:26,178 BRAIN, WITH OVER 86 BILLION 131 00:05:26,178 --> 00:05:28,647 NEURONS THAT MAKES TRILLIONS AND 132 00:05:28,647 --> 00:05:29,447 TRILLIANS OF SYNAPSES AND THEN 133 00:05:29,447 --> 00:05:31,850 YOU CAN ALSO LOOK AT CROSS 134 00:05:31,850 --> 00:05:34,953 SECTIONS HERE AND AGAIN, HERE'S 135 00:05:34,953 --> 00:05:35,921 THE MOUSE BRAIN, SO EVEN THOUGH 136 00:05:35,921 --> 00:05:39,357 WE CONSIDER IT A MODEL SYSTEM 137 00:05:39,357 --> 00:05:40,959 AND MORE SIMPLISTIC, IT'S 138 00:05:40,959 --> 00:05:43,028 INCREDIBLY COMPLEX IN TERMS OF 139 00:05:43,028 --> 00:05:44,129 CELLULAR DIVERSITY AND 140 00:05:44,129 --> 00:05:44,462 CONNECTIVITY. 141 00:05:44,462 --> 00:05:45,630 BUT THERE'S A LOT WE CAN LEARN 142 00:05:45,630 --> 00:05:48,300 ABOUT IT AND HOPEFULLY APPLY TO 143 00:05:48,300 --> 00:05:49,100 HUMAN BRAIN DEVELOPMENT. 144 00:05:49,100 --> 00:05:51,836 NOW I SAID THE CELLS, I STUDY 145 00:05:51,836 --> 00:05:54,739 THESE INHIBITORY NEURONS, THE 146 00:05:54,739 --> 00:05:56,708 BRAINS CONTAIN EXCITATORY CELLS 147 00:05:56,708 --> 00:05:58,777 AND INHIBITORY CELLS AND THIS 148 00:05:58,777 --> 00:06:00,679 BALANCE IS CRITICAL FOR 149 00:06:00,679 --> 00:06:01,379 MAINTAINING CIRCUIT FUNCTION. 150 00:06:01,379 --> 00:06:02,814 SO THESE CELLS WERE BORN IN THE 151 00:06:02,814 --> 00:06:03,882 VENTRAL PART OF THE BRAIN, THE 152 00:06:03,882 --> 00:06:08,687 BOTTOM PART OF THE BRAIN FROM 153 00:06:08,687 --> 00:06:09,521 THESE GANGLIONIC EVIDENCES AND 154 00:06:09,521 --> 00:06:12,224 HERE IS A 3D VIEW, THE LATERAL, 155 00:06:12,224 --> 00:06:14,025 MEDIAL AND KAWDAL AND MOST OF MY 156 00:06:14,025 --> 00:06:17,562 WALK IS FOCUSED ON THE MEDIAL 157 00:06:17,562 --> 00:06:20,332 AND CAUDALL, AND THESE CELLS ARE 158 00:06:20,332 --> 00:06:22,200 BORN, THEY BECOME POST MITOTIC 159 00:06:22,200 --> 00:06:23,935 IN THESE REGIONS AND THEN THEY 160 00:06:23,935 --> 00:06:25,737 GO AND POPULATE ALL REGIONS OF 161 00:06:25,737 --> 00:06:26,371 THE FOREBRAIN. 162 00:06:26,371 --> 00:06:28,840 WE KNOW A BIT ABOUT THE GENETIC 163 00:06:28,840 --> 00:06:32,377 MECHANISMS, THE GENES THAT DRIVE 164 00:06:32,377 --> 00:06:34,713 INTERNEURONS FROM THESE REGIONS, 165 00:06:34,713 --> 00:06:36,948 THE CG AND MG CELLS THAT ARISE 166 00:06:36,948 --> 00:06:38,583 FROM THESE REGIONS, THEY FORM 167 00:06:38,583 --> 00:06:39,985 DIFFERENT TYPES OF INTERNEURONS 168 00:06:39,985 --> 00:06:41,953 AND THEY'RE MORE OR LESS 169 00:06:41,953 --> 00:06:43,355 OVERLAPPING, SO DEPENDING ON 170 00:06:43,355 --> 00:06:45,190 WHAT MATURE TYPE OF INTERNEURON 171 00:06:45,190 --> 00:06:46,725 YOU ARE, WE KNOW WHETHER YOU 172 00:06:46,725 --> 00:06:50,762 AROSE FROM THE MG OR CG, THESE 2 173 00:06:50,762 --> 00:06:53,531 REGIONS GIVE COMPLETELY DISTINCT 174 00:06:53,531 --> 00:06:54,232 TYPES OF INTERNEURONS. 175 00:06:54,232 --> 00:06:56,735 AND AGAIN AS I MENTIONED THEY 176 00:06:56,735 --> 00:06:59,037 POPULATE THE ENTIRE BRAIN AND 177 00:06:59,037 --> 00:07:00,438 DIFFERENT DENSITIES AND 178 00:07:00,438 --> 00:07:01,339 DIFFERENT MORPHOLOGIES AND 179 00:07:01,339 --> 00:07:02,841 INVOLVED IN PRETTY MUCH EVERY 180 00:07:02,841 --> 00:07:06,878 ASPECT OF BRAIN CIRCUITRY. 181 00:07:06,878 --> 00:07:08,413 >> [INAUDIBLE QUESTION FROM 182 00:07:08,413 --> 00:07:08,847 AUDIENCE ] 183 00:07:08,847 --> 00:07:10,882 >> IN THE PREVIOUS SLIDE? 184 00:07:10,882 --> 00:07:12,550 LET ME GO BACK. 185 00:07:12,550 --> 00:07:17,722 SO THIS WAS ACTUALLY IN UTERY O 186 00:07:17,722 --> 00:07:20,458 OPERATION, SO THIS IS ME 187 00:07:20,458 --> 00:07:23,094 LABELING CELLS HERE WITH A GFP 188 00:07:23,094 --> 00:07:25,430 PLAZ MIDS MID, THAT EXPRESSES 189 00:07:25,430 --> 00:07:28,566 GFP, SO ABOUT 40 DAYS AFTER THE 190 00:07:28,566 --> 00:07:29,834 ELECTROPORRATION, THIS IS WHERE 191 00:07:29,834 --> 00:07:33,138 THE CELLS MIGRATED AND POPULATE, 192 00:07:33,138 --> 00:07:35,507 SO IT'S JUST A REPORTER OF CELLS 193 00:07:35,507 --> 00:07:38,009 THAT WERE BORN IN THIS BRAIN 194 00:07:38,009 --> 00:07:38,343 REGION. 195 00:07:38,343 --> 00:07:40,345 SO IF WE ZOOM IN ON THESE CELLS, 196 00:07:40,345 --> 00:07:41,780 THESE ARE ACTUAL RECONSTRUCTIONS 197 00:07:41,780 --> 00:07:43,415 OF TYPES OF INTERNEURONS AND IT 198 00:07:43,415 --> 00:07:45,884 CAN GIVE YOU A FIELD FOR THE 199 00:07:45,884 --> 00:07:47,018 STRIKING MORPHOLOGIES, SOME 200 00:07:47,018 --> 00:07:49,321 BASKET CELLS HAVE DENSE 201 00:07:49,321 --> 00:07:50,255 EXCELLENT ARBOR ANDS SYNAPSE 202 00:07:50,255 --> 00:07:53,258 LOCALLY AND THERE ARE OTHER CELL 203 00:07:53,258 --> 00:07:56,127 TYPES LIKE THIS MARNATI CELL 204 00:07:56,127 --> 00:07:57,195 WHICH SENDS CELLS AWAY FROM THE 205 00:07:57,195 --> 00:07:59,297 CELL BODY AND FORMS SYNAPSES ON 206 00:07:59,297 --> 00:07:59,798 DISTAL DENDRITES. 207 00:07:59,798 --> 00:08:00,765 IF YOU LOOK AT THIS CARTOON 208 00:08:00,765 --> 00:08:03,668 ISSUES HAVE YOU THIS VERY BORING 209 00:08:03,668 --> 00:08:04,402 PARAMETRAL EXCITATORY CELL IN 210 00:08:04,402 --> 00:08:05,837 GRAY, AND ALL THE COLORED CELLS 211 00:08:05,837 --> 00:08:08,840 WITH THE DIFFERENT TYPES OF 212 00:08:08,840 --> 00:08:11,142 INTERNEURONS THAT CAN SYNAPSE ON 213 00:08:11,142 --> 00:08:12,210 THESE EXCITATORY CELLS SO THIS 214 00:08:12,210 --> 00:08:14,112 GIVES YOU A FLAVOR FOR THE 215 00:08:14,112 --> 00:08:15,046 DIFFERENT MORPHOLOGIES AND 216 00:08:15,046 --> 00:08:15,880 CONNECTIVITY PATTERNS AND AGAIN 217 00:08:15,880 --> 00:08:18,416 HOW DO YOU GENERATE THIS 218 00:08:18,416 --> 00:08:19,617 INCREDIBLE DIVERSITY OF 219 00:08:19,617 --> 00:08:21,786 INHIBITORY CELLS FROM BASICALLY 220 00:08:21,786 --> 00:08:25,023 2 BRAINS REGIONS IN THE 221 00:08:25,023 --> 00:08:25,790 EMBRYONIC MOUSE BRAIN. 222 00:08:25,790 --> 00:08:27,425 NOW IT'S INTERESTING IS THAT 223 00:08:27,425 --> 00:08:29,994 INTERNEURONS MAKE UP ONLY 20-30% 224 00:08:29,994 --> 00:08:32,897 OF THE NEURONS IN THE BRAIN AND 225 00:08:32,897 --> 00:08:35,133 THE EXCITATORY CELLS MAKE UP 226 00:08:35,133 --> 00:08:35,367 70-80%. 227 00:08:35,367 --> 00:08:37,035 IF YOU LOOK AT THE DIFFERENT 228 00:08:37,035 --> 00:08:39,838 SUBTYPES, THIS IS FROM THAL AN 229 00:08:39,838 --> 00:08:42,273 BRAIN INSTITUTE WHERE THEY 230 00:08:42,273 --> 00:08:43,775 CLASSIFIED SUBTYPE SPACE SOLELY 231 00:08:43,775 --> 00:08:44,442 ON TRANSCRIPT ORDER OF MICRONSS, 232 00:08:44,442 --> 00:08:47,011 AND YOU SEE IN THE MOUSE CORTEX, 233 00:08:47,011 --> 00:08:49,080 50% OF THE DIFFERENT CELL TYPES 234 00:08:49,080 --> 00:08:51,983 IN THE BRAIN DEFINED BY GENE 235 00:08:51,983 --> 00:08:52,684 EXPRESSION ARE INTERNEURONS EVEN 236 00:08:52,684 --> 00:08:54,085 THOUGH THEY MAKE UP A SMALLER 237 00:08:54,085 --> 00:08:55,920 PROPORTION OF THE CELLS. 238 00:08:55,920 --> 00:08:58,256 AND THIS ACTUALLY EXPANDS IN THE 239 00:08:58,256 --> 00:09:02,060 HUMAN CORTEX WHERE ABOUT 60% OF 240 00:09:02,060 --> 00:09:03,361 THE CELLS GENETICALLY DEFINED 241 00:09:03,361 --> 00:09:05,864 CELL TYPES ARE INTERNEURONS, SO 242 00:09:05,864 --> 00:09:06,998 THERE'S INCREDIBLY HIGHER 243 00:09:06,998 --> 00:09:08,466 DIVERSITY COMPARED TO THE 244 00:09:08,466 --> 00:09:10,368 MAJORITY POPULATION OF THE 245 00:09:10,368 --> 00:09:11,536 EXCITATORY CELLS. 246 00:09:11,536 --> 00:09:12,937 NOW DYSFUNCTION OF THESE CELLS 247 00:09:12,937 --> 00:09:15,340 IS ASSOCIATED OR IN SOME CASES 248 00:09:15,340 --> 00:09:19,377 CAUSATIVE WITH THE WIDE VARIETY 249 00:09:19,377 --> 00:09:20,712 OF NEURODEVELOPMENTAL DISEASES, 250 00:09:20,712 --> 00:09:22,814 MOST NOTABLY SOMETHING LIKE 251 00:09:22,814 --> 00:09:24,416 EPILEPSY IF THERE'S TOO LITTLE 252 00:09:24,416 --> 00:09:25,950 INHIBITION IN THE BRAIN, IT CAN 253 00:09:25,950 --> 00:09:27,352 BE EXCITABLE AND THAT CAN LEAD 254 00:09:27,352 --> 00:09:29,454 TO EPILEPS EXPE SEIZURE 255 00:09:29,454 --> 00:09:32,257 DISORDERS, BUT IT'S BEEN NOTE 256 00:09:32,257 --> 00:09:34,225 THE NOTED IN A WIDE VARIETY OF 257 00:09:34,225 --> 00:09:35,960 DISEASES AND IN FACT MANY GENES 258 00:09:35,960 --> 00:09:37,462 ASSOCIATE WIDE THESE DISEASES 259 00:09:37,462 --> 00:09:39,697 ARE ENRICHED WITH NEURAL 260 00:09:39,697 --> 00:09:43,034 PROGENITORS SO CELLS AND OFTEN 261 00:09:43,034 --> 00:09:43,701 TIMES IMMATURE NEURONS. 262 00:09:43,701 --> 00:09:45,570 SO THEN WHAT MY LAB HAS BECOME 263 00:09:45,570 --> 00:09:47,972 MORE INTERESTED IN IS THE 264 00:09:47,972 --> 00:09:49,841 GROWING EVIDENCE THAT MANY 265 00:09:49,841 --> 00:09:53,044 PROTURBATIONS AND MUTATIONS, I'M 266 00:09:53,044 --> 00:09:54,245 ACTUALLY AFFECT EPIGENETIC 267 00:09:54,245 --> 00:09:56,748 MACHINE REGULATORY, RATHER THAN 268 00:09:56,748 --> 00:09:57,582 THE GENE CODING SEQUENCE 269 00:09:57,582 --> 00:09:57,882 THEMSELVES. 270 00:09:57,882 --> 00:09:59,551 SO THIS IS KIND OF A DRIVING 271 00:09:59,551 --> 00:10:00,952 THEME OF OUR LAB THAT WE 272 00:10:00,952 --> 00:10:02,487 BELIEVE, WE NEED A BETTER 273 00:10:02,487 --> 00:10:04,122 UNDERSTANDING OF BOTH THE 274 00:10:04,122 --> 00:10:05,390 GENETIC AND GENE REGULATORY 275 00:10:05,390 --> 00:10:07,792 MECHANISMS THAT GUIDE THESE 276 00:10:07,792 --> 00:10:09,627 INITIAL FETE DECISIONS EARLY ON 277 00:10:09,627 --> 00:10:12,397 IN THE EMBRYO AND THAT SHOULD 278 00:10:12,397 --> 00:10:13,898 INCREASE OUR UNDERSTANDING OF 279 00:10:13,898 --> 00:10:16,100 NORMAL DEVELOPMENT AND VARIES 280 00:10:16,100 --> 00:10:16,534 DISEASE ETIOLOGIES. 281 00:10:16,534 --> 00:10:18,303 SO WHEN THERE ARE PROTURBATIONS 282 00:10:18,303 --> 00:10:19,671 IN THESE GENES WHAT GOES ON 283 00:10:19,671 --> 00:10:21,072 EARLY ON THAT EVENTUALLY 284 00:10:21,072 --> 00:10:23,074 MANIFEST INTO A DISEASE 285 00:10:23,074 --> 00:10:23,341 PHONOTYPE? 286 00:10:23,341 --> 00:10:25,443 AS I WILL TELL YOU ABOUT A 287 00:10:25,443 --> 00:10:26,110 COUPLE STORIES TODAY. 288 00:10:26,110 --> 00:10:27,745 THE FIRST IS WE WANT TO DEFINE 289 00:10:27,745 --> 00:10:31,516 WHAT WE CALL AN EPIGENOME ATLAS, 290 00:10:31,516 --> 00:10:32,784 THAT SETS THE GROUND TRUTH FOR 291 00:10:32,784 --> 00:10:34,652 HOW THESE CELLS LOOK FROM THESE 292 00:10:34,652 --> 00:10:36,187 DIFFERENT BRAIN REGIONS IN THE 293 00:10:36,187 --> 00:10:37,255 EMBRYONIC BRAIN AND ONCE WE HAVE 294 00:10:37,255 --> 00:10:39,791 THE BASE LINE, WE'RE ABLE TO SEE 295 00:10:39,791 --> 00:10:40,859 HOW PROTURBATIONS AFFECT THIS 296 00:10:40,859 --> 00:10:41,159 DEVELOPMENT. 297 00:10:41,159 --> 00:10:42,760 SO I WILL TELL YOU ABOUT A 298 00:10:42,760 --> 00:10:45,163 VARIETY OF TECHNIQUES WE USE TO 299 00:10:45,163 --> 00:10:46,564 BASICALLY KIND OF DEFINE THE 300 00:10:46,564 --> 00:10:48,433 GROUND TRUTH OF THESE CELLS, 301 00:10:48,433 --> 00:10:51,202 MANY CELL SEQUENCING ASSAYS AS 302 00:10:51,202 --> 00:10:52,637 WELL AS BULK SEQUENCING ASSAYS 303 00:10:52,637 --> 00:10:55,106 AND ONCE WE ESTABLISHED THIS, 304 00:10:55,106 --> 00:10:57,475 THIS,A LOWSITOUS PERTURB THE 305 00:10:57,475 --> 00:10:59,244 SYSTEM WITH GENETIC MUTATIONS TO 306 00:10:59,244 --> 00:11:01,980 UNDERSTAND HOW ALTERATIONS OF 307 00:11:01,980 --> 00:11:04,282 EITHER GENE REPRESSION OR GENE 308 00:11:04,282 --> 00:11:06,584 ACTIVATION, CAN AFFECT CELL 309 00:11:06,584 --> 00:11:07,986 FATE, CELL FUNCTION AND 310 00:11:07,986 --> 00:11:09,053 ULTIMATELY ANIMAL BEHAVIOR AND 311 00:11:09,053 --> 00:11:10,555 LASTLY IF I HAVE TIME, I WILL 312 00:11:10,555 --> 00:11:14,359 TELL YOU ABOUT NEW DATA LOOKING 313 00:11:14,359 --> 00:11:15,693 AT ONCE TRANSCRIPTION FACTOR 314 00:11:15,693 --> 00:11:17,562 THAT'S HIGHLY CORRELATED WITH 315 00:11:17,562 --> 00:11:18,263 NEURODEVELOPMENTAL DISEASE AND 316 00:11:18,263 --> 00:11:23,067 WHAT INSIGHTS WE HAVE FROM OUR 317 00:11:23,067 --> 00:11:23,535 MOST RECENT STUDIES. 318 00:11:23,535 --> 00:11:25,270 SO A LOT OF OUR EXPERIMENTAL 319 00:11:25,270 --> 00:11:26,838 WORK FLOW FOLLOWS THIS PARADIGM. 320 00:11:26,838 --> 00:11:28,473 WE HAVE OUR EMBRYONIC MOUSE, IN 321 00:11:28,473 --> 00:11:29,674 A PARTICULAR AGE WE'RE 322 00:11:29,674 --> 00:11:32,310 INTERESTED IN, WE CAN THEN OPEN 323 00:11:32,310 --> 00:11:33,678 UP THE BRAIN, HEMISECTIONAL 324 00:11:33,678 --> 00:11:34,712 ANALYSISSED AND LOOK INSIDE. 325 00:11:34,712 --> 00:11:36,581 THIS IS OBVIOUSLY STAINED AND 326 00:11:36,581 --> 00:11:38,449 COLORED BUT YOU CAN ACTUALLY SEE 327 00:11:38,449 --> 00:11:41,319 THE STRUCTURES JUST BY EYE 328 00:11:41,319 --> 00:11:43,154 WITHOUT THE STAINING, THERE ARE 329 00:11:43,154 --> 00:11:44,422 BUMPS INTO THE VENTRICULAR SPACE 330 00:11:44,422 --> 00:11:46,624 SO IT'S VERY EASY TO ACCESS 331 00:11:46,624 --> 00:11:48,493 THESE AND DISSECT OUT THE MG OR 332 00:11:48,493 --> 00:11:50,662 THE CGE AND HERE IT IS IN 333 00:11:50,662 --> 00:11:51,095 CARTOON FORM. 334 00:11:51,095 --> 00:11:53,531 SO WE CAN DISSECT OUT THESE 335 00:11:53,531 --> 00:11:56,534 REGIONS, PREPARE SINGLE CELL OR 336 00:11:56,534 --> 00:11:57,569 SINGLE NUCLEI SUSPENSIONS, OFTEN 337 00:11:57,569 --> 00:11:59,170 FACT SORT THEM, PUT THEM THROUGH 338 00:11:59,170 --> 00:12:01,272 OUR 10 X SYSTEM TO DO SINGLE 339 00:12:01,272 --> 00:12:03,041 CELL SEQUENCING OR ALTERNATIVELY 340 00:12:03,041 --> 00:12:04,876 DO SOME BULK SEQUENCING WITH 341 00:12:04,876 --> 00:12:08,012 OTHER ASSAYS AND THEN USE NICHD 342 00:12:08,012 --> 00:12:10,648 SEQUENCING CORE AND THEN WHEN 343 00:12:10,648 --> 00:12:12,183 MOY LAB SPENDS MOST OF THEIR 344 00:12:12,183 --> 00:12:14,519 TIME DOING IS BIOINFORMATTICS 345 00:12:14,519 --> 00:12:16,754 FIGURING OUT TO YOU MASSAGE THE 346 00:12:16,754 --> 00:12:18,122 DATA AND MANIPULATE IT TO TRY TO 347 00:12:18,122 --> 00:12:19,090 UNDERSTAND BIOLOGY FROM THESE 348 00:12:19,090 --> 00:12:21,125 COMPLEX DATA SETS AND THIS IS A 349 00:12:21,125 --> 00:12:21,759 FAIRLY ACCURATE REPRESENTATION 350 00:12:21,759 --> 00:12:24,028 OF WHAT MY POST DOCS THINK ABOUT 351 00:12:24,028 --> 00:12:26,130 DAY-TO-DAY AS THEY WORK ON THIS 352 00:12:26,130 --> 00:12:27,165 DATA SET. 353 00:12:27,165 --> 00:12:28,366 SO, THERE HAVE BEEN A LOT OF 354 00:12:28,366 --> 00:12:30,935 WORK THAT HAD LOOKED AT 355 00:12:30,935 --> 00:12:36,207 TRANSCRIPT 356 00:12:36,207 --> 00:12:37,375 TRANSCRIPTOMES FROM THESE BRAIN 357 00:12:37,375 --> 00:12:39,444 REGIONS BUT WE WANTED TO LOOK 1 358 00:12:39,444 --> 00:12:40,778 STEP HIGHER AT CHROMATIN 359 00:12:40,778 --> 00:12:43,481 ACCESSIBLE AND BED HOW THE 360 00:12:43,481 --> 00:12:44,882 CHROMATIN LOOKS IN THESE BRAIN 361 00:12:44,882 --> 00:12:46,517 REGIONS THAT GIVE RISE TO 362 00:12:46,517 --> 00:12:47,251 DISTINCT CELL TYPES. 363 00:12:47,251 --> 00:12:49,454 SO COUPLE YEARS AFTER I JOINED 364 00:12:49,454 --> 00:12:50,955 HERE, 10 X DEVELOPED THE 365 00:12:50,955 --> 00:12:53,691 COMMERCIALLY AVAILABLE SINGLE 366 00:12:53,691 --> 00:12:56,894 CELL, ATAQ SEQ PLATFORM, YOU 367 00:12:56,894 --> 00:13:00,331 COULD DO BULK AATTACK SEQ, BUT 368 00:13:00,331 --> 00:13:00,898 IN HETEROGENEOUS ENVIRONMENT 369 00:13:00,898 --> 00:13:02,567 SUCH AS THE DEVELOPING BRAIN 370 00:13:02,567 --> 00:13:08,573 IT'S NOT THAT INFORMATIVE. 371 00:13:08,573 --> 00:13:13,578 SO WHAT ATTACK SEQ IS, THE USE 372 00:13:13,578 --> 00:13:16,014 THE SEQUENCING THAT IDENTIFY 373 00:13:16,014 --> 00:13:18,016 OPEN REGIONS OF CHROMATIN, IT 374 00:13:18,016 --> 00:13:20,118 HAS TAGS, IT CAN CUT THIS DNA SO 375 00:13:20,118 --> 00:13:21,986 THAT YOU'RE LEFT WITH THE PIECES 376 00:13:21,986 --> 00:13:23,821 OF DNA THAT REPRESENT REGIONS OF 377 00:13:23,821 --> 00:13:27,692 CHROMATIN AND THEN YOU CAN 378 00:13:27,692 --> 00:13:30,495 SEQUENCE IT WITH THESE TAGS, SO 379 00:13:30,495 --> 00:13:31,896 WE PERFORM ATAC SINGLE CELL 380 00:13:31,896 --> 00:13:34,432 SEQUENCING ON THESE BRAIN 381 00:13:34,432 --> 00:13:39,704 REGIONS SO WE SECTIONED OUT THE 382 00:13:39,704 --> 00:13:41,706 LG, CG, AND MG, AND THIS GIVES 383 00:13:41,706 --> 00:13:43,041 WAY TO OVERLAPPING NEURONS. 384 00:13:43,041 --> 00:13:45,076 WE PREP THE SAMPLES AND THEN 385 00:13:45,076 --> 00:13:46,244 BASICALLY YOU HAVE THE READ OUT 386 00:13:46,244 --> 00:13:47,679 WHERE YOU HAVE SEQUENCES, 387 00:13:47,679 --> 00:13:48,513 SEQUENCE READS THROUGHOUT THE 388 00:13:48,513 --> 00:13:50,048 GENOME AND WHAT YOU'RE LOOKING 389 00:13:50,048 --> 00:13:52,183 FOR IS LEGIONS WHERE THERE'S A 390 00:13:52,183 --> 00:13:53,618 HIGH,AMOUNT OF SEQUENCE READS IN 391 00:13:53,618 --> 00:13:56,521 A VARIETY OF CELLS AND THEN THAT 392 00:13:56,521 --> 00:13:58,589 WOULD BASICALLY BE ABOVE THE 393 00:13:58,589 --> 00:14:00,625 THRESHOLD YOU WOULD CALL A PEAK 394 00:14:00,625 --> 00:14:02,627 AND THIS IS A REGION THAT IS 395 00:14:02,627 --> 00:14:04,262 ACCESSIBLE IN THE SINGLE CELL OR 396 00:14:04,262 --> 00:14:14,005 THE GROUP OF CELLS THAT WE'RE 397 00:14:14,005 --> 00:14:14,305 LOOKING AT. 398 00:14:14,305 --> 00:14:16,307 -- EACH DOT YOU'RE LOOKING AT 399 00:14:16,307 --> 00:14:17,842 REP REISN'TS CHROMATIN 400 00:14:17,842 --> 00:14:18,543 ACCESSIBLE, RATHER THAN GENE 401 00:14:18,543 --> 00:14:21,012 EXPRESSION AND YOU CAN SEE THAT 402 00:14:21,012 --> 00:14:23,014 THE CELLS FROM EACH REGION KIND 403 00:14:23,014 --> 00:14:24,882 OF CLUSTER TODAY, YOU HAVE 404 00:14:24,882 --> 00:14:26,451 EXCITATORY CELLS IN THE CORTEX, 405 00:14:26,451 --> 00:14:28,419 MG CELLS OVER HERE AND THEN LG 406 00:14:28,419 --> 00:14:31,956 AND CG FOR WHICH WE KNOW LESS, 407 00:14:31,956 --> 00:14:32,557 THERE'S LESS DISTINCT 408 00:14:32,557 --> 00:14:33,925 DIFFERENCES IN GENE EXPRESSION 409 00:14:33,925 --> 00:14:35,593 PROFILES, THEY ACTUALLY KIND OF 410 00:14:35,593 --> 00:14:37,095 CLUSTERED TOGETHER HERE AND YOU 411 00:14:37,095 --> 00:14:38,730 CAN PERFORM DIFFERENT TYPES OF 412 00:14:38,730 --> 00:14:40,131 ANALYSIS JUST LIKE YOU COULD 413 00:14:40,131 --> 00:14:42,700 WITH SINGLE CELL RNA SEQ, AND 414 00:14:42,700 --> 00:14:44,268 YOU CAN HAVE REPEAT CLUSTERS. 415 00:14:44,268 --> 00:14:45,870 IN THIS CASE WE KNOW HOW THESE 416 00:14:45,870 --> 00:14:48,239 CELL DIVISIONS WORK AND WE CAN 417 00:14:48,239 --> 00:14:49,674 KIND OF DEFINE DIFFERENT 418 00:14:49,674 --> 00:14:51,175 PROGENITORS IN HERE AND THEN YOU 419 00:14:51,175 --> 00:14:52,343 CAN ALSO DID PSEUDOTIME TO LOOK 420 00:14:52,343 --> 00:14:54,212 AT THE DEVELOPMENT OF THESE 421 00:14:54,212 --> 00:14:58,516 REGIONS IN THESE CELLS ACROSS 422 00:14:58,516 --> 00:14:58,983 NEUROGENESIS. 423 00:14:58,983 --> 00:15:01,185 NOW IN THIS CASE, WHAT WE'RE 424 00:15:01,185 --> 00:15:04,922 LOOKING AT HERE IS HEAT MAPS 425 00:15:04,922 --> 00:15:07,024 SHOWING REGIONS OF PROMOTERS, OF 426 00:15:07,024 --> 00:15:09,427 VARIOUS GENES THAT ARE 427 00:15:09,427 --> 00:15:11,162 ACCESSIBLE, SO NESTIN IS A GENE 428 00:15:11,162 --> 00:15:14,132 THAT'S EXPRESSED IN RADIAL GLIAL 429 00:15:14,132 --> 00:15:14,899 CELLS, VERY IMMATURE CELLS AND 430 00:15:14,899 --> 00:15:17,001 YOU CAN SEE THAT THE ACCESSIBLE 431 00:15:17,001 --> 00:15:19,403 OF THE NESTIN PROMOTER IS 432 00:15:19,403 --> 00:15:21,339 RESTRICTED TO THESE CELLS THAT 433 00:15:21,339 --> 00:15:22,640 ARE THE RADIAL GLIAL CELLS AND 434 00:15:22,640 --> 00:15:26,410 AS THE CELLS GO FURTHER ALONG TO 435 00:15:26,410 --> 00:15:28,379 DEVELOPMENT, THIS LOCI GETS SHUT 436 00:15:28,379 --> 00:15:29,847 OFF AND BECOMES CLOSED SO CAN 437 00:15:29,847 --> 00:15:31,149 YOU DO THIS WITH VARIOUS GENES 438 00:15:31,149 --> 00:15:32,884 OVER TIME AND YOU CAN LOOK AT 439 00:15:32,884 --> 00:15:37,889 VARIOUS GENES THAT HAVE SPATIAL 440 00:15:37,889 --> 00:15:39,257 EXPRESSION, SO NKX2.1, I WILL 441 00:15:39,257 --> 00:15:40,758 COME BACK TO YOU, THIS IS THE 442 00:15:40,758 --> 00:15:42,260 SPECIFIC GENE, CAN YOU SEE THE 443 00:15:42,260 --> 00:15:43,194 PROMOTER IS ONLY ACCESS CLINICAL 444 00:15:43,194 --> 00:15:44,395 COMMUNITY IN THE MGE AND IT'S 445 00:15:44,395 --> 00:15:46,063 CLOSED OFF IN THE OTHER BRAIN 446 00:15:46,063 --> 00:15:46,430 REGIONS. 447 00:15:46,430 --> 00:15:47,532 AND THEN THERE ARE OTHER WAYS TO 448 00:15:47,532 --> 00:15:49,734 LOOK AT THIS, IF YOU LOOK AT THE 449 00:15:49,734 --> 00:15:50,802 PUTATIVE CLUSTERS AT WHICH THERE 450 00:15:50,802 --> 00:15:53,571 IS ABOUT 25, YOU CAN LOOK AT 451 00:15:53,571 --> 00:15:55,006 CHROMATIN ACCESSIBLE AND 452 00:15:55,006 --> 00:15:56,474 CLUSTERS HERE BASED ON SPATIAL 453 00:15:56,474 --> 00:15:59,110 LOCATION, SO CAN YOU SEE AGAIN 454 00:15:59,110 --> 00:16:01,245 THE NK 2.1 PROMOTER IS MAINLY 455 00:16:01,245 --> 00:16:02,680 ACCESSIBLE IN THE MGE AND THE 456 00:16:02,680 --> 00:16:04,182 ABSENCE OF OTHER CLUSTERS AND 457 00:16:04,182 --> 00:16:06,717 AGAIN, YOU CAN LOOK AT THAT IN 458 00:16:06,717 --> 00:16:08,452 BOTH SPATIAL DIVISIONS AND 459 00:16:08,452 --> 00:16:09,720 DEVELOPMENTAL TIME ACCESS HERE. 460 00:16:09,720 --> 00:16:12,356 NOW THIS IS KIND OF A FAIRLY 461 00:16:12,356 --> 00:16:13,791 COMPLEX IMAGE BUT IT'S PRETTY 462 00:16:13,791 --> 00:16:17,328 GOOD SUMMARY OF HOW THE 463 00:16:17,328 --> 00:16:17,929 DIFREPRESENTIALLY ACCESSIBLE 464 00:16:17,929 --> 00:16:19,797 PEAKS LOOK AT OUR DATA SET, SO 465 00:16:19,797 --> 00:16:21,432 IN THIS CASE, THE TISSUE IS 466 00:16:21,432 --> 00:16:23,067 BROKEN DOWN BY THESE COLOR BARS 467 00:16:23,067 --> 00:16:24,769 UP HERE, THESE ARE DIFFERENT 468 00:16:24,769 --> 00:16:26,370 STAGES OF NEUROGENESIS HERE, AND 469 00:16:26,370 --> 00:16:28,606 WHAT WE DID IS BASICALLY TOOK 470 00:16:28,606 --> 00:16:31,542 THE 25 OR SO CLUSTERS WE HAD, 471 00:16:31,542 --> 00:16:34,645 PUTATIVE CELL CLUSTERS AND DID 472 00:16:34,645 --> 00:16:35,713 UNBIASED HIERARCHICAL CLUSTERING 473 00:16:35,713 --> 00:16:37,348 SO WE CAN IDENTIFY CLUSTERS THAT 474 00:16:37,348 --> 00:16:38,149 WERE MOST CLOSELY ASSOCIATED 475 00:16:38,149 --> 00:16:39,283 WITH EACH OTHER AND SO WHAT YOU 476 00:16:39,283 --> 00:16:43,621 CAN SEE HERE, IF YOU LOOK IN THE 477 00:16:43,621 --> 00:16:45,022 UPPER LEFT, THERE'S 4 CLUSTERS 478 00:16:45,022 --> 00:16:48,025 HERE, THESE ARE ALL CORTICALE 479 00:16:48,025 --> 00:16:49,093 APICAL PROGENITORS SO THEY'RE 480 00:16:49,093 --> 00:16:50,394 ALL FAIRLY SIMILAR AND YOU CAN 481 00:16:50,394 --> 00:16:53,064 SEE THERE'S ACTUALLY FAIR -- 482 00:16:53,064 --> 00:16:55,266 SOME SIMILARITY BETWEEN THE 483 00:16:55,266 --> 00:16:56,000 DIFFERENTIALLY ACCESSIBLE PEAKS 484 00:16:56,000 --> 00:16:57,034 IN THESE DEFINITE CLUSTERS AND 485 00:16:57,034 --> 00:16:58,336 THAT MAKES SENSE BECAUSE THEY'RE 486 00:16:58,336 --> 00:17:00,338 ALL VERY SIMILAR CELL TYPES, 487 00:17:00,338 --> 00:17:01,939 CONVERSELY IF YOU LOOK AT 488 00:17:01,939 --> 00:17:02,807 DIFFERENCES BETWEEN DIFFERENT 489 00:17:02,807 --> 00:17:03,941 BRAIN REGIONS AND DIFFERENT CELL 490 00:17:03,941 --> 00:17:06,277 TYPES, YOU SEE THERE'S A LOT 491 00:17:06,277 --> 00:17:07,445 LESS SIMILARITY HERE SO WHAT WE 492 00:17:07,445 --> 00:17:09,413 SEE IS THAT THE ACCESSIBILITY 493 00:17:09,413 --> 00:17:11,415 PROFILES OF THESE CELLS ACTUALLY 494 00:17:11,415 --> 00:17:13,117 MATCHES PRETTY WELL WITH BOTH 495 00:17:13,117 --> 00:17:15,620 GENE EXPRESSION AND THERE'S 496 00:17:15,620 --> 00:17:16,854 QUITE DISTINCT DIFFERENCES 497 00:17:16,854 --> 00:17:18,689 BETWEEN BOTH RAIN BREEJIONS AND 498 00:17:18,689 --> 00:17:25,963 BETWEEN DIFFERENT CELL TYPES AND 499 00:17:25,963 --> 00:17:26,764 DEVELOPMENT. 500 00:17:26,764 --> 00:17:28,332 SO TO TAKE 1 BRIEF TANGENT, AS 501 00:17:28,332 --> 00:17:31,035 WE'RE DOING THE DATA WE IN THE 502 00:17:31,035 --> 00:17:32,637 LAB GENERATE RNASEQ DAILY BASIS 503 00:17:32,637 --> 00:17:33,838 THEA OF THE 4 BRAIN REGIONS AND 504 00:17:33,838 --> 00:17:35,706 WE DID THIS BECAUSE WE WANTED TO 505 00:17:35,706 --> 00:17:38,042 HAVE BOTH TRANSCRIPTIONAL DATA 506 00:17:38,042 --> 00:17:39,677 AND CHROMATIN ACCESSIBLE IN 507 00:17:39,677 --> 00:17:41,646 THESE BRAIN REGIONS AND SO, I 508 00:17:41,646 --> 00:17:44,515 WON'T GO THROUGH ALL THIS BUT WE 509 00:17:44,515 --> 00:17:49,787 FOUND DISTINCT EXPRESSION 510 00:17:49,787 --> 00:17:52,123 PROFILES IN THESE CELLS BETWEEN 511 00:17:52,123 --> 00:17:54,425 BRAIN REGIONS AND HERE'S 512 00:17:54,425 --> 00:17:57,228 EXAMPLES FOR LG, MG, AND CG, AND 513 00:17:57,228 --> 00:17:58,729 THERE'S ALSO INITANCES WITHIN 514 00:17:58,729 --> 00:18:00,398 THE SINGLE BRAIN REGION, AND YOU 515 00:18:00,398 --> 00:18:01,933 SEE 1 THAT'S ENRICHED IN THE 516 00:18:01,933 --> 00:18:03,367 DORSAL, AND YOU SEE 1 THAT'S 517 00:18:03,367 --> 00:18:04,702 ENRICHED IN THE VENTRAL KRRK GE 518 00:18:04,702 --> 00:18:08,873 AND WE HOPE AND WE THINK THAT 519 00:18:08,873 --> 00:18:10,474 THESE EXPRESSION PROFILES IN 520 00:18:10,474 --> 00:18:11,409 THESE DIFFERENT DOMAINS ARE 521 00:18:11,409 --> 00:18:13,411 CRITICAL FOR GIVING RISE OF 522 00:18:13,411 --> 00:18:14,946 THESE TYPES OF INTERNEURONS BUT 523 00:18:14,946 --> 00:18:17,748 WE HAD THE SINGLE CELL RNA DATA 524 00:18:17,748 --> 00:18:20,418 AND WE GENERATE SINGLE CELL ATAC 525 00:18:20,418 --> 00:18:21,919 DATA WHICH I WILL TOUCH ON LATER 526 00:18:21,919 --> 00:18:32,430 SO WHAT WE DID IS -- THEY COME 527 00:18:36,434 --> 00:18:38,035 FROM THE SAME REGIONS AND TIME 528 00:18:38,035 --> 00:18:39,904 POINTS AND THIS ALLOWED US HOW 529 00:18:39,904 --> 00:18:43,307 TO LOOK AT RNA AND CHROMATIN 530 00:18:43,307 --> 00:18:45,242 ACCESSIBILITY CHANGES OVER TIME 531 00:18:45,242 --> 00:18:47,178 AND IDENTIFY DIFFERENCES IN 532 00:18:47,178 --> 00:18:47,712 BRAIN REGIONS. 533 00:18:47,712 --> 00:18:50,014 SO WHAT WE SAW WAS IN INSTANCES 534 00:18:50,014 --> 00:18:52,149 WHERE GENES WERE EXPRESSED 535 00:18:52,149 --> 00:18:53,417 EARLIER IN THE NEUROGENIC 536 00:18:53,417 --> 00:18:57,822 TIMELINE AND THEN WEPT DOWN OVER 537 00:18:57,822 --> 00:18:59,724 TIME, WE SAW BLUE HERE IS THE 538 00:18:59,724 --> 00:19:01,759 RNA, RED IS THE GAS AND THAT'S A 539 00:19:01,759 --> 00:19:04,161 FANCY WAY OF SAYING PROMOTER 540 00:19:04,161 --> 00:19:05,429 ACCESSIBILITY AND YOU CAN SEE IF 541 00:19:05,429 --> 00:19:07,531 YOU FOLLOW THE BLUE HERE THE RNA 542 00:19:07,531 --> 00:19:10,434 TURNS OFF PRIOR TO THE PROMOTER 543 00:19:10,434 --> 00:19:11,802 AND THE ENHANCERS SHUTTING DOWN. 544 00:19:11,802 --> 00:19:15,239 SO IT'S AS IF THE RNA IS DOWN 545 00:19:15,239 --> 00:19:17,241 REGULATED BEFORE THE ACTUAL 546 00:19:17,241 --> 00:19:18,976 PROMOTERS AND ENHANCERS TURNED 547 00:19:18,976 --> 00:19:19,176 DOWN. 548 00:19:19,176 --> 00:19:20,945 CONVERSELY IF YOU LOOK AT GENES 549 00:19:20,945 --> 00:19:22,279 THAT ARE EXPRESSED LATER IN 550 00:19:22,279 --> 00:19:24,448 DEVELOP AM, YOU SEE GREEN AND 551 00:19:24,448 --> 00:19:27,118 RED THE PROMOTER AND ENHANCERS 552 00:19:27,118 --> 00:19:29,754 BECOME ACCESSIBLE PRIOR TO THE 553 00:19:29,754 --> 00:19:30,421 RNA BEING EXPRESSED. 554 00:19:30,421 --> 00:19:32,656 AND THAT'S WHAT YOU WOULD 555 00:19:32,656 --> 00:19:35,092 EXPECT, YOU NEED PROMOLT 556 00:19:35,092 --> 00:19:35,793 PROMOTERS AND ENHANCERS OPEN 557 00:19:35,793 --> 00:19:36,861 BEFORE YOU GET THE NEXT 558 00:19:36,861 --> 00:19:37,928 EXPRESSION, SO THIS IS NICE, CAN 559 00:19:37,928 --> 00:19:43,134 YOU DO IT AT MULTIO--METABOLIZD 560 00:19:43,134 --> 00:19:46,370 AND 1 DATA AND 1 SELL, SO WE SEE 561 00:19:46,370 --> 00:19:48,139 THESE 2 PATTERNS WHEN WE SEE 562 00:19:48,139 --> 00:19:49,106 THESE DATA SETS INTEGRATED 563 00:19:49,106 --> 00:19:49,473 TOGETHER. 564 00:19:49,473 --> 00:19:52,710 NOW THIS IS GREAT IN LOOKING AT 565 00:19:52,710 --> 00:19:54,078 RNA EXPRESSION, IT'S INFORMATIVE 566 00:19:54,078 --> 00:19:57,982 BUT IT'S ONLY CORRELATIVE, BUT 567 00:19:57,982 --> 00:19:59,250 WE CAN MAKE PREDICTIONS ABOUT 568 00:19:59,250 --> 00:20:00,384 WHAT'S AN PROMOTER AND ENHANCER 569 00:20:00,384 --> 00:20:02,119 BUT WE WANT TO HAVE GREATER DATA 570 00:20:02,119 --> 00:20:03,788 SO I CAN SAY WITH MIER 571 00:20:03,788 --> 00:20:05,723 CONFIDENCE, THIS IS AN ENHANCER 572 00:20:05,723 --> 00:20:07,992 OF THIS GENE AND IT'S DIRECTLY 573 00:20:07,992 --> 00:20:08,826 REGULATING ITS EXPRESSION EMPLOY 574 00:20:08,826 --> 00:20:12,663 SO 1 WAY TO DO THAT IS LOOK AT 575 00:20:12,663 --> 00:20:14,899 HISTONE MODISKS, SO HISTONES ARE 576 00:20:14,899 --> 00:20:17,001 PROTEINS THAT DNA RAP WRAPS 577 00:20:17,001 --> 00:20:18,702 AROUND THE HISTONES AND THESE 578 00:20:18,702 --> 00:20:20,738 CAN BE MODJUSTIFIED IN A VARIETY 579 00:20:20,738 --> 00:20:23,040 OF WAYS, PHOSPHORYLATED OR, 580 00:20:23,040 --> 00:20:25,543 SETALATED AND DEPENDING ON THE 581 00:20:25,543 --> 00:20:26,710 PATTERN OF MODIFICATIONS, IT CAN 582 00:20:26,710 --> 00:20:28,646 BE INDICATIVE OF THE STATE OF 583 00:20:28,646 --> 00:20:29,146 THE DNA. 584 00:20:29,146 --> 00:20:32,349 SO IF WE LOOK AT 3 OF THESE 585 00:20:32,349 --> 00:20:36,120 MARKS IN PARTICULAR, HK34 IS A 586 00:20:36,120 --> 00:20:41,025 TRI-METHYLATION IS AN ACTIVE 587 00:20:41,025 --> 00:20:47,264 PROMOTER, A3 KME4, AND THEN HK27 588 00:20:47,264 --> 00:20:48,699 TRI METHYL, INDICATOR OF AN 589 00:20:48,699 --> 00:20:49,600 CORRELATED PRESSIVE STATE. 590 00:20:49,600 --> 00:20:53,304 SO WE CAN TAKE THESE AND 591 00:20:53,304 --> 00:20:57,308 PERFORM MODERATIONS AND DO TAG, 592 00:20:57,308 --> 00:20:58,909 SIMILAR TO CHPSEQ, BUT YOU NEED 593 00:20:58,909 --> 00:21:00,778 LESS CELLS TO GET GOOD DATA AND 594 00:21:00,778 --> 00:21:03,447 IT'S MUCH CHEAPER TO LOOK AT IN 595 00:21:03,447 --> 00:21:05,015 BULK THESE METHYLATION PATTERNS 596 00:21:05,015 --> 00:21:07,785 IN THESE DIFFERENT BRAIN 597 00:21:07,785 --> 00:21:08,052 REGIONS. 598 00:21:08,052 --> 00:21:10,788 SO HERE'S A SUMMARY OF HOW THIS 599 00:21:10,788 --> 00:21:12,523 DATA CAN LOOK, SO WHAT WE HAVE 600 00:21:12,523 --> 00:21:14,492 HERE IS AN ANALYSIS CALLED 601 00:21:14,492 --> 00:21:16,360 SISSER O WHERE WE LOOK FOR 602 00:21:16,360 --> 00:21:17,928 INDIVIDUAL CELLS, IF A LOCI IS 603 00:21:17,928 --> 00:21:20,664 OPEN HERE AND THEN YOU GO DOWN, 604 00:21:20,664 --> 00:21:21,966 DOWN STREAM OR UPSTREAM AND 605 00:21:21,966 --> 00:21:22,800 THERE'S ANOTHER REGION HERE, 606 00:21:22,800 --> 00:21:25,136 THAT IS OPEN IN THE SAME CELLS, 607 00:21:25,136 --> 00:21:27,338 YOU KIND OF GET THESE PEAKS SO 608 00:21:27,338 --> 00:21:28,739 THIS TYPE OF ANALYSIS IS 609 00:21:28,739 --> 00:21:30,541 INTEREST, IF YOU'RE LOOKING AT A 610 00:21:30,541 --> 00:21:32,409 PROMOTER AND YOU SEE A REGION 611 00:21:32,409 --> 00:21:35,112 UPSTREAM OR DOWN STREAM THAT IS 612 00:21:35,112 --> 00:21:36,547 OFTEN OPEN OR CO ACCESSIBLE IN 613 00:21:36,547 --> 00:21:37,882 THE SAME CELL, THAT COULD BE 614 00:21:37,882 --> 00:21:39,984 INDICIAIVE IT OF AN ENHANCER, SO 615 00:21:39,984 --> 00:21:42,153 I FOCUS ON 1 GENE HERE CALLED 616 00:21:42,153 --> 00:21:43,854 ASCL 1 AND THIS IS A GENE 617 00:21:43,854 --> 00:21:45,723 EXPRESSIONED IN ALL THE 618 00:21:45,723 --> 00:21:46,657 GANGLIONIC EMINENCES BUT IT'S 619 00:21:46,657 --> 00:21:48,526 NOT EXPRESSED IN CORACLE CELLS 620 00:21:48,526 --> 00:21:51,195 AND IN PREVIOUS WORK HAD ASSAYED 621 00:21:51,195 --> 00:21:54,732 THIS REGION RIGHT HERE AS A 622 00:21:54,732 --> 00:21:56,133 CANDIDATE ENHANCER FOR ASCL1. 623 00:21:56,133 --> 00:21:57,768 NOW WHAT WE SEE IN OUR DATA IS 624 00:21:57,768 --> 00:22:00,437 THAT IF YOU LOOK IN THESE 4 625 00:22:00,437 --> 00:22:02,106 BRAIN REGIONS THIS LOCI IS 626 00:22:02,106 --> 00:22:04,008 CERTAINLY ACCESSIBLE IN ALL THE 627 00:22:04,008 --> 00:22:06,310 BRAIN REGIONS BUT IF YOU LOOK AT 628 00:22:06,310 --> 00:22:08,179 THIS MARKER AND EFFECTIVE 629 00:22:08,179 --> 00:22:09,346 ENHANCER IT'S STRONGER IN THE 630 00:22:09,346 --> 00:22:13,350 CORTEX AND LESSER IN THE 631 00:22:13,350 --> 00:22:14,084 GANGLIONIC INSTANCES AND AGAIN 632 00:22:14,084 --> 00:22:16,921 THE BRAIN IS WHERE THE CORTEX IS 633 00:22:16,921 --> 00:22:18,122 NOT EXPRESSED SO WHEN THEY LOOK 634 00:22:18,122 --> 00:22:20,891 AT THIS, IT DID NOT DO A GOOD 635 00:22:20,891 --> 00:22:22,459 JOB OF REPLICATING GENE 636 00:22:22,459 --> 00:22:23,861 EXPRESSION AND WE WOULD ARGUE 637 00:22:23,861 --> 00:22:25,496 BECAUSE OF DATA LIKE THIS THE 638 00:22:25,496 --> 00:22:27,264 HIGHEST SIGNAL FOR AN ACTIVE 639 00:22:27,264 --> 00:22:28,199 ENHANCER IS ACTUALLY IN CELL 640 00:22:28,199 --> 00:22:30,301 TYPES WHERE THE GENE IS NOT 641 00:22:30,301 --> 00:22:30,634 EXPRESSED. 642 00:22:30,634 --> 00:22:31,936 CONVERSELY IF YOU LOOK DOWN HERE 643 00:22:31,936 --> 00:22:33,237 IN GRAY, WE FOUND 2 OTHER 644 00:22:33,237 --> 00:22:35,306 RENALLIONS WHERE YOU HAVE REALLY 645 00:22:35,306 --> 00:22:37,208 HIGH SIGNAL FOR ACTIVE ENHANCERS 646 00:22:37,208 --> 00:22:39,009 AND MUCH LOWER IN THE CORTEX SO 647 00:22:39,009 --> 00:22:42,146 WE ARGUE THAT BY HAVING THIS 648 00:22:42,146 --> 00:22:44,582 MULTIMODAL DATA SET, CAN YOU DO 649 00:22:44,582 --> 00:22:46,283 A BETTER PREDICTION FOR 650 00:22:46,283 --> 00:22:46,984 CANDIDATE ENHANCERS FOR VARIOUS 651 00:22:46,984 --> 00:22:50,087 YEENS AND SO WE HAVE A WHOLE 652 00:22:50,087 --> 00:22:53,290 LIST OF CANDIDATE ENHANCERS THAT 653 00:22:53,290 --> 00:22:56,093 WE THINK COULD PLAY IMPORTANT 654 00:22:56,093 --> 00:22:57,861 INSIGHT TO GENE REGULATION FOR 655 00:22:57,861 --> 00:23:00,397 VARIOUS YEENS DURING 656 00:23:00,397 --> 00:23:00,864 DEVELOPMENT. 657 00:23:00,864 --> 00:23:03,834 SO THAT'S HIGHLIGHTED THERE. 658 00:23:03,834 --> 00:23:07,638 SO ANOTHER ASSAY WE DID, SO FAR 659 00:23:07,638 --> 00:23:10,474 WE HAVE ACCESSIBILITY, RNA AND 660 00:23:10,474 --> 00:23:11,308 HISTONE MODIFICATIONS BUT IN 661 00:23:11,308 --> 00:23:13,677 ORDER TO HAVE PROMOTER ENHANCER 662 00:23:13,677 --> 00:23:15,446 INTERACTIONS, THE DNA NEEDS TO 663 00:23:15,446 --> 00:23:16,046 COME INTO CONTACT. 664 00:23:16,046 --> 00:23:17,881 AND SO TO REALLY LOOK AT THAT, 665 00:23:17,881 --> 00:23:21,085 WE USED HIGH C OR MORE RECENTLY 666 00:23:21,085 --> 00:23:22,586 MICROC IN COLLABORATION WITH 667 00:23:22,586 --> 00:23:24,922 PEDRO [INDISCERNIBLE]'S LAB AT 668 00:23:24,922 --> 00:23:27,491 NICHD AND SO WHAT HIGH C IS AN 669 00:23:27,491 --> 00:23:30,127 ASSAY TO LOOK AT DIRECT DNA 670 00:23:30,127 --> 00:23:32,029 CONTEXTS BETWEEN 2 DNA REGIONS. 671 00:23:32,029 --> 00:23:34,798 AND SO, IF YOU JUST -- SO THIS 672 00:23:34,798 --> 00:23:36,800 IS KIND OF A SCHEMATIC OF HOW 673 00:23:36,800 --> 00:23:38,602 DNA CAN LOOK, BASIC LYE TO GET 674 00:23:38,602 --> 00:23:41,472 ENHANCER AND PROMOTER TO COME 675 00:23:41,472 --> 00:23:43,807 TOGETHER, THE DNA ACTUALLY HAS 676 00:23:43,807 --> 00:23:44,508 QUITE ORGANIZED STRUCTURE, AND 677 00:23:44,508 --> 00:23:46,944 THERE ARE VARIOUS WAYS TO BRING 678 00:23:46,944 --> 00:23:47,278 THEM TOGETHER. 679 00:23:47,278 --> 00:23:49,747 A LOT OF TIMES THEY INVOLVE THE 680 00:23:49,747 --> 00:23:51,282 CTCF SITES THAT I WILL COME BACK 681 00:23:51,282 --> 00:23:53,717 TO TOWARDS THE END OF MY TALK TO 682 00:23:53,717 --> 00:23:56,654 KIND OF SEGGREGATE THE DNA IN 683 00:23:56,654 --> 00:23:57,187 TOPOGRAPHICALLY ASSOCIATED 684 00:23:57,187 --> 00:23:59,356 DOMAINS OR TABS AND THAT'S THE 685 00:23:59,356 --> 00:24:02,259 WAY TO RESTRICT DNA FROM OTHER 686 00:24:02,259 --> 00:24:05,162 REGIONS, AND ALLOW THESE 687 00:24:05,162 --> 00:24:06,630 PROMOTER ENHANCER INTERACTIONS 688 00:24:06,630 --> 00:24:08,365 TO OCCUR IN SPECIFIC WAYS. 689 00:24:08,365 --> 00:24:12,236 AND SO BY USING THIS HIGH C 690 00:24:12,236 --> 00:24:16,507 ASSAY, YOU CAN IDENTIFY THESE 691 00:24:16,507 --> 00:24:18,375 LOCI WHERE YOU HAVE THE DISTAL 692 00:24:18,375 --> 00:24:21,912 REGIONS COMING INTO CONTACT AND 693 00:24:21,912 --> 00:24:22,980 PROVIDES GREATER INSIGHT TO HOW 694 00:24:22,980 --> 00:24:24,148 THESE GENES ARE REGULATED. 695 00:24:24,148 --> 00:24:29,386 SO THIS IS HOW THE HI-C LOOKS 696 00:24:29,386 --> 00:24:29,620 LIKE. 697 00:24:29,620 --> 00:24:32,356 ARE THESE HOT SPOTS OF VARIOUS 698 00:24:32,356 --> 00:24:33,791 LOCI, AND IF YOU TAKE THE REGION 699 00:24:33,791 --> 00:24:35,526 LIKE THIS, YOU FOLLOW IT DOWN 700 00:24:35,526 --> 00:24:38,495 THE PYRAMID TO HEAR AND COME 701 00:24:38,495 --> 00:24:40,998 COME DOWN THIS SIDE OF THE 702 00:24:40,998 --> 00:24:43,133 PYRAMID AND THIS SIGNAL TELLS 703 00:24:43,133 --> 00:24:44,635 YOU HAVE A DIRECT CONNECTION 704 00:24:44,635 --> 00:24:46,270 BETWEEN THIS REGION HERE IN THE 705 00:24:46,270 --> 00:24:47,938 DNA AND THIS REGION DOWN HERE SO 706 00:24:47,938 --> 00:24:49,273 IF THIS HAPPENS TO BE A PROMOTER 707 00:24:49,273 --> 00:24:51,275 OF A YEEN AND THIS IS A 708 00:24:51,275 --> 00:24:53,043 CABBEDIDATE ENHANCER, THIS 709 00:24:53,043 --> 00:24:53,777 PROVIDES FURTHER EVIDENCE THAT 710 00:24:53,777 --> 00:24:54,945 YOU HAVE THIS GENETIC 711 00:24:54,945 --> 00:24:56,747 INTEGRATION THAT YOU NEED. 712 00:24:56,747 --> 00:25:00,884 SO WHAT WE DID IS WE PERFORM 713 00:25:00,884 --> 00:25:02,720 THESE HI-C ASSAYS ON THESE 4 714 00:25:02,720 --> 00:25:04,788 BRAIN REGIONS AND THEN WE LOOK 715 00:25:04,788 --> 00:25:06,523 AT YEENS OF INTEREST TO FIND 716 00:25:06,523 --> 00:25:09,626 REGIONS WHERE WE SAW 717 00:25:09,626 --> 00:25:12,496 DIFFERENTIAL DNA INTERACTION, SO 718 00:25:12,496 --> 00:25:13,997 HERE'S THE NK2.1 GENE THAT IS 719 00:25:13,997 --> 00:25:15,999 EXPRESSED IN THE MG AND CRITICAL 720 00:25:15,999 --> 00:25:17,968 FOR ALL CELLS DERIVE FRIDAY THE 721 00:25:17,968 --> 00:25:19,336 MG AND HOPEFULLY CAN YOU 722 00:25:19,336 --> 00:25:21,305 APPRECIATE THAT AT THE MGE, HAVE 723 00:25:21,305 --> 00:25:23,173 YOU THE DARK INCREASE INTENSITY 724 00:25:23,173 --> 00:25:24,541 HERE AND THAT'S ABSENT FROM 725 00:25:24,541 --> 00:25:26,210 THESE OTHER BRAIN REGIONS, SO 726 00:25:26,210 --> 00:25:30,881 THIS IS AN MGE SPECIFIC GENOMIC 727 00:25:30,881 --> 00:25:31,248 INTERACTION. 728 00:25:31,248 --> 00:25:34,084 NOW CAPTURE C IS KIND OF A 729 00:25:34,084 --> 00:25:36,053 MODIFICATION OF HI-C AND IT 730 00:25:36,053 --> 00:25:36,820 CONFIRMS THE DIRECT INTERACTION 731 00:25:36,820 --> 00:25:39,123 BETWEEN IN THIS CASE, THE NK 2.1 732 00:25:39,123 --> 00:25:41,625 PROMOTER AND A REGION DOWN 733 00:25:41,625 --> 00:25:41,859 STREAM. 734 00:25:41,859 --> 00:25:44,294 AND SO WHAT YOU SEE IS IN 735 00:25:44,294 --> 00:25:46,196 INCREASED SIGNAL HERE WHICH 736 00:25:46,196 --> 00:25:47,898 INDICATES THIS REAS YOJ HAS 737 00:25:47,898 --> 00:25:50,501 DIRECT CONTACT WITH THE NK2.1 738 00:25:50,501 --> 00:25:51,769 PROMOTER SPECIFIC TO THE MG, YOU 739 00:25:51,769 --> 00:25:54,671 DON'T DO NOT SEE IT IN THE LG, 740 00:25:54,671 --> 00:25:56,840 CG, CORTEX OR LIVERS OF CONTROL. 741 00:25:56,840 --> 00:25:58,442 AND THEN YOU INDGREAT LITTLER 742 00:25:58,442 --> 00:26:01,345 DATA WE HAVE WITH SINGLE CELL 743 00:26:01,345 --> 00:26:02,646 RNASEQ, AND DIFFERENT 744 00:26:02,646 --> 00:26:04,014 REGULATIONS AND REALLY TRULY 745 00:26:04,014 --> 00:26:05,749 UNDERSTAND AT A GENOMIC LEVEL 746 00:26:05,749 --> 00:26:07,418 AND CHROMATIN LEVEL BOTH HOW THE 747 00:26:07,418 --> 00:26:08,752 CHROMATIN LOOKS AS 3D STRUCTURE 748 00:26:08,752 --> 00:26:12,689 WHERE IT'S IRPT ACTIONS ARE, THE 749 00:26:12,689 --> 00:26:13,490 HISTONE MODIFICATIONS TO TELL 750 00:26:13,490 --> 00:26:15,592 YOU WHETHER SOMETHING IS A 751 00:26:15,592 --> 00:26:17,261 LIKELY ENHANCER, PROMOTER OR 752 00:26:17,261 --> 00:26:18,662 REPRESSED AND THEN ACCESSIBILITY 753 00:26:18,662 --> 00:26:20,063 AND RNA AND WHAT I WILL NOTE 754 00:26:20,063 --> 00:26:24,568 HERE IS THAT THIS MG SPECIFIC 755 00:26:24,568 --> 00:26:26,637 INTERACTION OCCURS WITH THIS 756 00:26:26,637 --> 00:26:29,406 GENE CALLED MBIP WHO HAS NOT -- 757 00:26:29,406 --> 00:26:31,275 WHOSE FUNCTION IS NOT KNOWN IN 758 00:26:31,275 --> 00:26:33,143 NEURAL DEVELOPMENT AND IT'S 759 00:26:33,143 --> 00:26:35,179 STUDIED IN CANCER BUT I DID FIND 760 00:26:35,179 --> 00:26:37,714 A PAPER FROM ABOUT 15 YEARS AGO 761 00:26:37,714 --> 00:26:41,385 WHERE THEY DID STAINING FOR MBIP 762 00:26:41,385 --> 00:26:43,320 AND MBIP IS ALSO ONLY EXPRESSION 763 00:26:43,320 --> 00:26:45,355 IN THE MG, IT'S NOT FOUND IN THE 764 00:26:45,355 --> 00:26:45,956 OTHER BRAIN REGIONS. 765 00:26:45,956 --> 00:26:48,125 SO NOW WE KNOW THERE'S A DIRECT 766 00:26:48,125 --> 00:26:48,826 INTERACTION BETWEEN THESE 2 767 00:26:48,826 --> 00:26:50,561 GENES THAT ARE ONLY EXPRESSED IN 768 00:26:50,561 --> 00:26:52,362 THE MG AND NOT FOUND IN OTHER 769 00:26:52,362 --> 00:26:52,830 BRAIN REGIONS. 770 00:26:52,830 --> 00:26:54,231 AND THEN WE WENT AHEAD AND YOU 771 00:26:54,231 --> 00:26:56,333 CAN DO THIS FOR OTHER GENES AND 772 00:26:56,333 --> 00:26:57,534 OTHER BRAIN REGIONS, AND THERE'S 773 00:26:57,534 --> 00:27:00,737 LOTS OF KIND OF EXCITING INSIGHT 774 00:27:00,737 --> 00:27:03,674 YOU HAVE, WE HAVE INTO REGION 775 00:27:03,674 --> 00:27:05,776 SPECIFIC BRAIN REGION SPECIFIC 776 00:27:05,776 --> 00:27:08,145 CHROMATIN INTERACTIONS, IN THE 777 00:27:08,145 --> 00:27:09,413 EMBRYONIC BRAIN AND TO OUR 778 00:27:09,413 --> 00:27:10,714 KNOWLEDGE THIS HASN'T BEEN 779 00:27:10,714 --> 00:27:11,782 EXPLORED IN DETAIL YET. 780 00:27:11,782 --> 00:27:14,618 SO WE REALLY THINK, WE 781 00:27:14,618 --> 00:27:16,119 UNDERSTAND THE GROUND TRUTH OF 782 00:27:16,119 --> 00:27:18,589 HOW THESE CELLS LOOK IN THE 783 00:27:18,589 --> 00:27:21,058 EMBRYO AS WE'RE MAKING THESE 784 00:27:21,058 --> 00:27:22,059 CRITICAL FATE DECISIONS. 785 00:27:22,059 --> 00:27:23,694 NOW I WILL HIGHLIGHT 1 ONGOING 786 00:27:23,694 --> 00:27:26,196 STUDY THAT CAME OUT OF THIS 787 00:27:26,196 --> 00:27:27,498 SPECIFIC NK2.1 INTERACTION. 788 00:27:27,498 --> 00:27:28,932 AS I MENTIONED MASTER REGULATOR 789 00:27:28,932 --> 00:27:30,801 OF CELL FATE AND SO WE WANTED TO 790 00:27:30,801 --> 00:27:33,604 KNOW HOW CRITICAL IS THIS IRPT 791 00:27:33,604 --> 00:27:35,239 ACTION FOR EXPRESS AND FUNCTION 792 00:27:35,239 --> 00:27:35,706 OF THESE YEENS? 793 00:27:35,706 --> 00:27:39,109 IS THERE A WAY TO PERTUSH THIS 794 00:27:39,109 --> 00:27:40,143 INTERACTION TO DISRUPT IT AND 795 00:27:40,143 --> 00:27:42,546 SEE HOW THAT AFFECTS GENE 796 00:27:42,546 --> 00:27:43,680 EXPRESSION CELL FATE AND WHY 797 00:27:43,680 --> 00:27:45,315 THIS IS ALSO RELEVANT IS BECAUSE 798 00:27:45,315 --> 00:27:49,586 IN HUMAN DISEASE, THERE ARE 799 00:27:49,586 --> 00:27:51,221 VARIANTS OF NKX 2.1 YEEN AND IT 800 00:27:51,221 --> 00:27:52,422 GIVES RISE TO COMPLEX SYNDROME 801 00:27:52,422 --> 00:28:00,330 THAT CAN BE CALLED BENIGN 802 00:28:00,330 --> 00:28:02,633 HEREDITARY CHOREA, OR BRAIN LUNG 803 00:28:02,633 --> 00:28:04,034 THYROID SYNDROME, IT'S ALSO 804 00:28:04,034 --> 00:28:06,003 EXPRESSED IN THE LUNG AND 805 00:28:06,003 --> 00:28:06,236 THYROID. 806 00:28:06,236 --> 00:28:08,305 SO PATIENTS THAT HAVE MUTATIONS 807 00:28:08,305 --> 00:28:11,108 AND VARIANTS OF THE NK2.1 CAN 808 00:28:11,108 --> 00:28:12,709 EXPRESS PHENOTYPES THAT SHOW 809 00:28:12,709 --> 00:28:15,145 DYSFUNCTION IN THESE 3 ORGANS 810 00:28:15,145 --> 00:28:17,414 WITH VARIOUS SEVERITYS DEPENDING 811 00:28:17,414 --> 00:28:18,549 ON THE PATIENT. 812 00:28:18,549 --> 00:28:19,383 NOW WHAT'S ALSO REALLY 813 00:28:19,383 --> 00:28:21,251 INTERESTING IS THAT A HANDFUL OF 814 00:28:21,251 --> 00:28:22,886 PATIENTS THAT PRESENT WITH THIS 815 00:28:22,886 --> 00:28:26,390 PHENOTYPE DO NOT ACTUALLY HAVE 816 00:28:26,390 --> 00:28:28,559 MUTATIONS IN NK2.1. 817 00:28:28,559 --> 00:28:30,961 RATHER THEY HAVE DELETIONS 818 00:28:30,961 --> 00:28:33,730 FURTHER UPSTREAM THAT ENCOMPASS 819 00:28:33,730 --> 00:28:36,567 THE MBIP GENE AND THERE ARE NO 820 00:28:36,567 --> 00:28:38,001 KNOWN FUNCTIONS FOR THIS 821 00:28:38,001 --> 00:28:38,402 NEURODEVELOPMENT. 822 00:28:38,402 --> 00:28:39,736 SO NOW WE UNDERSTAND THIS 823 00:28:39,736 --> 00:28:40,904 INTERACTION HERE WE WONDERED IF 824 00:28:40,904 --> 00:28:42,806 IT'S POSSIBLE THAT SOME OF THIS 825 00:28:42,806 --> 00:28:45,309 PHENOTYPE THAT ARISES IN THESE 826 00:28:45,309 --> 00:28:48,345 PATIENTS WERE THE NK2.1 LOCI IS 827 00:28:48,345 --> 00:28:52,282 INTACT BUT THE MBIP IS DISTURBED 828 00:28:52,282 --> 00:28:54,351 OR MUTATED COULD BE DUE TO THIS 829 00:28:54,351 --> 00:28:55,185 CRITICAL INTERACTION HERE, SO 830 00:28:55,185 --> 00:28:56,753 AGAIN, HOW DO WE STUDY THIS, HOW 831 00:28:56,753 --> 00:29:00,257 DO WE BREAK UP THIS 832 00:29:00,257 --> 00:29:00,591 INTERACTION,. 833 00:29:00,591 --> 00:29:02,893 >> MUNICIPAL ADVISERS AND I 834 00:29:02,893 --> 00:29:03,961 ASK'S QUESTION? 835 00:29:03,961 --> 00:29:04,227 >> YES. 836 00:29:04,227 --> 00:29:05,662 >> HOW IS THE GENE PRODUCTION 837 00:29:05,662 --> 00:29:13,337 WITH THIS AND [INDISCERNIBLE]? 838 00:29:13,337 --> 00:29:15,339 >> YEAH, GOOD QUESTION AND KIND 839 00:29:15,339 --> 00:29:15,939 OF COMPLICATED. 840 00:29:15,939 --> 00:29:19,443 SO AT THIS RESOLUTION WITH WE 841 00:29:19,443 --> 00:29:21,345 DID THIS DATA AT HI-C WE DID NOT 842 00:29:21,345 --> 00:29:22,846 HAVE THE RESOLUTION TO KNOW 843 00:29:22,846 --> 00:29:23,780 WHETHER THE INTERACTION, SO THAT 844 00:29:23,780 --> 00:29:26,183 YOU KNOW, ALL I CAN SAY HERE IS 845 00:29:26,183 --> 00:29:28,619 THIS INTERACTION IS SOMEWHERE IN 846 00:29:28,619 --> 00:29:29,586 THIS MBIP LOCUST, YOU CAN SEE 847 00:29:29,586 --> 00:29:32,489 THIS IS A GENE WITH 5 OR 6 AXONS 848 00:29:32,489 --> 00:29:34,691 AND THE RESOLUTION WE HAD IS WE 849 00:29:34,691 --> 00:29:35,692 KNOW THERE'S AN INTERACTION 850 00:29:35,692 --> 00:29:38,328 SOMEWHERE THERE WHETHER IT'S A 851 00:29:38,328 --> 00:29:39,463 PROMOTER-PROMOTER INTERACTION OR 852 00:29:39,463 --> 00:29:41,231 WHETHER THERE'S A PARTICULAR 853 00:29:41,231 --> 00:29:46,003 LOCI WITHIN AN MBIP ENTRON, 854 00:29:46,003 --> 00:29:48,672 THAT'S AN ENHANCER FOR NK2.1 OR 855 00:29:48,672 --> 00:29:49,606 VICE VERSA, WE CAN'T SAY 856 00:29:49,606 --> 00:29:51,241 ANYTHING ABOUT IT AT THIS LEVEL 857 00:29:51,241 --> 00:29:52,643 OF RESOLUTION. 858 00:29:52,643 --> 00:29:55,045 I MY KROSIA I TALK ABOUT A 859 00:29:55,045 --> 00:29:57,147 LITTLE BIT DOES GIVE YOU BETTER 860 00:29:57,147 --> 00:29:59,149 RESOLUTION WHERE YOU CAN LOOK AT 861 00:29:59,149 --> 00:30:00,550 10S OF NUCLEOTIDES AND SO THEN 862 00:30:00,550 --> 00:30:02,519 WE CAN SAY WITH CONFIDENCE THAT 863 00:30:02,519 --> 00:30:06,623 THERE'S INTERACTION BETWEEN THE 864 00:30:06,623 --> 00:30:09,493 NK2.1 PROMOTER AND SOME NEUTRON 865 00:30:09,493 --> 00:30:12,396 WEAN IT AND MBIP BUT THERE'S 866 00:30:12,396 --> 00:30:13,030 PROMOTER-PROMOTER INTERACTIONS 867 00:30:13,030 --> 00:30:14,831 THAT HELP BOTH GENES AND THEN 868 00:30:14,831 --> 00:30:17,134 THERE'S ALSO EVIDENCE FOR 869 00:30:17,134 --> 00:30:18,735 ENHANCERS OR ENTRON NEAR ANOTHER 870 00:30:18,735 --> 00:30:19,703 GENE AFFECTING THE GENE 871 00:30:19,703 --> 00:30:21,304 EXPRESSION OF ANOTHER 1. 872 00:30:21,304 --> 00:30:23,273 SO THAT'S REALLY A QUESTION OF 873 00:30:23,273 --> 00:30:24,007 RESOLUTION, HOW REFINED CAN YOU 874 00:30:24,007 --> 00:30:32,816 GET TO LOOK AT THIS INTERACTION. 875 00:30:32,816 --> 00:30:33,950 >> [INAUDIBLE QUESTION FROM 876 00:30:33,950 --> 00:30:34,451 AUDIENCE ] 877 00:30:34,451 --> 00:30:36,420 >> YEAH, AGAIN ABOUT RESOLUTION. 878 00:30:36,420 --> 00:30:39,489 ALL WE CAN SAY ABOUT THIS IS 879 00:30:39,489 --> 00:30:40,757 THERE'S A DIRECT INTERACTION 880 00:30:40,757 --> 00:30:43,660 BETWEEN SOME REGION HERE IN THE 881 00:30:43,660 --> 00:30:46,496 MBIP YEEN AND THE IN, K2.1 882 00:30:46,496 --> 00:30:47,964 PROMOTER. 883 00:30:47,964 --> 00:30:50,867 IT'S POSSIBLE THERE ARE 2 OR 3 884 00:30:50,867 --> 00:30:51,501 DISTINCT INTERACTION DOMAINS BUT 885 00:30:51,501 --> 00:30:53,070 WE DON'T HAVE THE RESOLUTION 886 00:30:53,070 --> 00:31:01,078 WITH THIS ASSAY TO DIRECTLY 887 00:31:01,078 --> 00:31:01,912 INTERACT WITH THAT. 888 00:31:01,912 --> 00:31:03,080 >> [INAUDIBLE QUESTION FROM 889 00:31:03,080 --> 00:31:03,346 AUDIENCE ] 890 00:31:03,346 --> 00:31:06,683 >> SO CAN YOU LOOK AT 891 00:31:06,683 --> 00:31:07,417 INTERCHROMESOME INTERACTIONS 892 00:31:07,417 --> 00:31:07,684 WITH HI-C. 893 00:31:07,684 --> 00:31:08,952 SO WHAT I'VE DONE HERE IS FOCUS 894 00:31:08,952 --> 00:31:10,454 ON THE SPECIFIC REGION BUT CAN 895 00:31:10,454 --> 00:31:13,757 YOU EXPAND THIS OUT TO ALL 896 00:31:13,757 --> 00:31:14,558 CHROMATIN -- CHROMESOME 897 00:31:14,558 --> 00:31:17,761 INTERACTION SO IF THERE ARE IRPT 898 00:31:17,761 --> 00:31:19,129 ACTIONS BETWEEN CHROMESOMES THAT 899 00:31:19,129 --> 00:31:22,332 OCCUR AT A HIGH ENOUGH PREQUENCY 900 00:31:22,332 --> 00:31:24,067 THAT IT WOULD KIND OF GENERATE 901 00:31:24,067 --> 00:31:25,936 THESE HOT SPOTS YOU COULD 902 00:31:25,936 --> 00:31:27,137 THEORETICALLY DETECT IT IN THIS 903 00:31:27,137 --> 00:31:27,704 TYPE OF DATA. 904 00:31:27,704 --> 00:31:29,706 BUT IN THIS CASE I FOCUSED ON 905 00:31:29,706 --> 00:31:30,774 THIS LOCI OF INTEREST WHERE WE 906 00:31:30,774 --> 00:31:35,078 HAVE THIS GENE AND THEN A STRONG 907 00:31:35,078 --> 00:31:36,580 BRAIN REGION SIGNAL JUST 908 00:31:36,580 --> 00:31:38,849 UPSTREAM, OKAY IN OKAY, SO HOW 909 00:31:38,849 --> 00:31:40,784 CAN WE DISRUPT THIS INTERACTION? 910 00:31:40,784 --> 00:31:42,452 THIS IS WHERE OUR COLLABORATION 911 00:31:42,452 --> 00:31:44,554 WITH PEDRO RO, KREBS CYCLE CO 912 00:31:44,554 --> 00:31:45,989 WAS CRITICAL, SO, IN A STUDY 913 00:31:45,989 --> 00:31:48,024 THAT WE COLLABORATED WITH, HE 914 00:31:48,024 --> 00:31:50,794 WAS LOOKING AT THE SOX 2 LOCUST 915 00:31:50,794 --> 00:31:54,531 WHICH IS A CRITICAL GENE OF 916 00:31:54,531 --> 00:31:56,566 INTERACTION FOR THE EPIBLAST AND 917 00:31:56,566 --> 00:31:59,136 SO BASICALLY YOU HAVE A CRITICAL 918 00:31:59,136 --> 00:32:00,537 ENHANCER OF SOX 2 WHICH IS 919 00:32:00,537 --> 00:32:03,140 CALLED SCR OVER HERE AND THEN 920 00:32:03,140 --> 00:32:04,574 THE SOX 2 GENE IS OVER HERE. 921 00:32:04,574 --> 00:32:07,577 AND YOU HAVE THESE LITTLE ARROWS 922 00:32:07,577 --> 00:32:09,179 ARE CALLED CTCF SITES AND AS I 923 00:32:09,179 --> 00:32:10,680 MENTIONED THAT'S CRITICAL FOR 924 00:32:10,680 --> 00:32:12,182 PARSING THIS GENOME STRUCTURES 925 00:32:12,182 --> 00:32:15,452 THAT YOU HAVE THIS TAD DOMAIN 926 00:32:15,452 --> 00:32:18,655 HERE SO THAT BASICALLY GENOMEEC 927 00:32:18,655 --> 00:32:20,056 LOCI HERE CAN INTERACT WITH EACH 928 00:32:20,056 --> 00:32:22,058 OTHER AND REALLY ARE INHIBITED 929 00:32:22,058 --> 00:32:23,326 FROM INTERACTING OUT HERE. 930 00:32:23,326 --> 00:32:26,663 SO PEDRO'S IDEA IS WHAT IF WE 931 00:32:26,663 --> 00:32:28,565 PUT IN ARTIFICIAL CTF SITES HERE 932 00:32:28,565 --> 00:32:29,966 TO DISRUPT THE INTERACTION AND 933 00:32:29,966 --> 00:32:31,368 THAT'S WHATEE DID IN THIS PAPER, 934 00:32:31,368 --> 00:32:32,769 CAN YOU DO DIFFERENT KIND OF 935 00:32:32,769 --> 00:32:34,905 TYPES OR DIFFERENT ORGANIZATIONS 936 00:32:34,905 --> 00:32:36,206 OF CTCF FACTORS AND I HOPE YOU 937 00:32:36,206 --> 00:32:38,742 CAN APPRECIATE THAT WHEN YOU DO 938 00:32:38,742 --> 00:32:40,443 THESE MODIIVESS WHAT YOU DO IS 939 00:32:40,443 --> 00:32:42,846 DISRUPT OR IN THIS CASE MINIMIZE 940 00:32:42,846 --> 00:32:44,014 THIS INTERACTION BETWEEN THE SOX 941 00:32:44,014 --> 00:32:45,882 2 LOCUST AND THE ENHANCER OVER 942 00:32:45,882 --> 00:32:47,384 HERE AND THEN CAN YOU LOOK AT 943 00:32:47,384 --> 00:32:49,553 HOW THIS AFFECTS GENE EXPRESSION 944 00:32:49,553 --> 00:32:49,820 PHENOTYPE. 945 00:32:49,820 --> 00:32:52,255 SO WE SAID GREAT, CAN WE DO THIS 946 00:32:52,255 --> 00:32:55,425 AT THE LOCI WE CARE ABOUT IN THE 947 00:32:55,425 --> 00:32:56,193 NK2.1 PROMOTER? 948 00:32:56,193 --> 00:32:57,327 AND LUCKILY IT HAD THE SITES SET 949 00:32:57,327 --> 00:32:59,863 UP IN A WAY THAT ALLOWED US TO 950 00:32:59,863 --> 00:33:04,067 DO THIS SO PEDRO DESIGNED 951 00:33:04,067 --> 00:33:06,336 EXOGENOUS CTCF SITES TO INSERT 952 00:33:06,336 --> 00:33:08,505 ON IN THESE 2 YEENS WITH THE 953 00:33:08,505 --> 00:33:09,573 GOAL OF BLOCKING THE 954 00:33:09,573 --> 00:33:09,940 INTERACTION. 955 00:33:09,940 --> 00:33:13,009 THIS IS NEW DATA WITH MY ROUGH 956 00:33:13,009 --> 00:33:15,111 ATOM SEEDS, WITH HIGH 957 00:33:15,111 --> 00:33:17,581 REZONINGUTION AT HI-C AND YOU 958 00:33:17,581 --> 00:33:20,317 HAVE THE NK2.1 YEEN HERE AND 959 00:33:20,317 --> 00:33:21,384 THEN YOU WILD-TYPE HERE IN THE 960 00:33:21,384 --> 00:33:23,720 INTERACTION IN THE MICE WHERE WE 961 00:33:23,720 --> 00:33:25,355 INSERTED THE CTCF SITES, YOU SEE 962 00:33:25,355 --> 00:33:26,623 THE INTERACTION IS GONE AND 963 00:33:26,623 --> 00:33:28,391 INSTEAD YOU HAVE AN ISHT 964 00:33:28,391 --> 00:33:30,026 INTERACTION HERE WHERE YOU HAVE 965 00:33:30,026 --> 00:33:32,429 THE BLUE CTCF SITE, INTERACTING 966 00:33:32,429 --> 00:33:35,632 WITH THESE RED SITES, SO THIS 967 00:33:35,632 --> 00:33:37,434 WAS SUCCESSFUL IN PERTURBING 968 00:33:37,434 --> 00:33:42,572 THIS INTERACTION BETWEEN THESE 2 969 00:33:42,572 --> 00:33:43,373 LOCI. 970 00:33:43,373 --> 00:33:44,007 AGAIN, CAPTURE MICRO-C, ANOTHER 971 00:33:44,007 --> 00:33:46,343 WAY OF LOOKING AT IT, YOU SEE 972 00:33:46,343 --> 00:33:48,044 THE WILD-TYPE, IT'S NOW REDUCED 973 00:33:48,044 --> 00:33:49,779 IN THE MUTANT WITH AN ARTIFICIAL 974 00:33:49,779 --> 00:33:51,748 IRPT ACTION HERE SO WE CONFIRM 975 00:33:51,748 --> 00:33:55,619 THAT BASICALLY WE BLOCK THIS 976 00:33:55,619 --> 00:33:58,922 NK2.1 MBIP INTERACTION IN THESE 977 00:33:58,922 --> 00:33:59,155 MICE. 978 00:33:59,155 --> 00:34:00,490 >> QUESTION [INAUDIBLE QUESTION 979 00:34:00,490 --> 00:34:01,391 FROM AUDIENCE ] 980 00:34:01,391 --> 00:34:03,793 >> SO THIS IS 3, WE HAVE 3 GOING 981 00:34:03,793 --> 00:34:07,731 THIS WAY AND 3 GOING THIS WAY, 982 00:34:07,731 --> 00:34:15,372 AS KIND OF THE MORE CTCFs 983 00:34:15,372 --> 00:34:18,942 SITES YOU IN TERMS OF BLOCKING 984 00:34:18,942 --> 00:34:21,144 THAT, THESE ARE HUMAN SITES SO 985 00:34:21,144 --> 00:34:22,846 THESE MAKES IT EASIER FOR 986 00:34:22,846 --> 00:34:26,283 SEQUENCING AND DOWN STREAMS 987 00:34:26,283 --> 00:34:28,118 OKAY, SO WHAT HAPPENS IN GENE 988 00:34:28,118 --> 00:34:30,687 EXPRESSION, THIS IS INSITUS 989 00:34:30,687 --> 00:34:32,422 LOOKING AT NK2.1 AND MBIP AND 990 00:34:32,422 --> 00:34:35,125 YOU CAN SEE IN THE MUTANT IS 991 00:34:35,125 --> 00:34:37,294 THAT THE MBIP EXPRESSION IS 992 00:34:37,294 --> 00:34:39,829 ALMOST TOTALLY GONE IN THE NK 993 00:34:39,829 --> 00:34:41,264 AND CTCF INSERTION, IT BEING 994 00:34:41,264 --> 00:34:44,634 LOAMACYS LIKE THERE'S DOWN 995 00:34:44,634 --> 00:34:47,270 REGULATION OF NK2.1 PRACTICES 996 00:34:47,270 --> 00:34:48,004 FORMING ADDITIONAL CAN FIRM 997 00:34:48,004 --> 00:34:50,106 THIS, BUT IF YOU LOOK AT THE PC 998 00:34:50,106 --> 00:34:52,175 R, YOU SEE STRONG REDUCTION, 50% 999 00:34:52,175 --> 00:34:55,545 OR SO REDUCTION OF MBIP IN THIS 1000 00:34:55,545 --> 00:34:57,180 MUTANT, IT LOOKS STRONGER BY 1001 00:34:57,180 --> 00:34:59,182 INSITUS AND WE SEE A SLIGHT BUT 1002 00:34:59,182 --> 00:35:01,484 PROBABLY SIGNIFICANT DECREASE IN 1003 00:35:01,484 --> 00:35:01,718 MK2.1. 1004 00:35:01,718 --> 00:35:03,286 AND WE THINK THIS IS EXCITING 1005 00:35:03,286 --> 00:35:04,688 BECAUSE AGAIN GOING BACK TO 1006 00:35:04,688 --> 00:35:07,624 THOSE PATIENTS THIS COULD 1007 00:35:07,624 --> 00:35:08,625 INDICATE THAT IN THESE PATIENTS 1008 00:35:08,625 --> 00:35:10,560 WHERE YOU HAVE THE LOCUST, WHAT 1009 00:35:10,560 --> 00:35:12,429 REALLY COULD BE HAPPENING IS 1010 00:35:12,429 --> 00:35:14,264 THAT YOU'RE DISRUPTING THIS 1011 00:35:14,264 --> 00:35:15,765 PARTICULAR INTERACTION THAT'S 1012 00:35:15,765 --> 00:35:19,069 DECREASING GENE EXPRESSION AND 1013 00:35:19,069 --> 00:35:19,836 PROBABLY PERTURBING NK2.1 1014 00:35:19,836 --> 00:35:21,938 EXPRESSION EVEN THOUGH THE 1015 00:35:21,938 --> 00:35:23,907 NKSUPER 1 LOCUST IS IN TACT AND 1016 00:35:23,907 --> 00:35:25,842 THUS GIVING RISE TO THE 1017 00:35:25,842 --> 00:35:26,142 PHENOTYPE. 1018 00:35:26,142 --> 00:35:27,811 SO WE'RE KIND OF WORKING ON 1019 00:35:27,811 --> 00:35:29,312 THESE MICE, IT CHANGES IN CELL 1020 00:35:29,312 --> 00:35:32,315 FATE AND ANY TYPE OF BEHAVIORIAL 1021 00:35:32,315 --> 00:35:34,718 DEFICITS BUT HERE'S A NICE BRAIN 1022 00:35:34,718 --> 00:35:35,752 REGION SPECIFIC INTERACTION WE 1023 00:35:35,752 --> 00:35:37,387 IDENTIFIED AND WHEN YOU 1024 00:35:37,387 --> 00:35:37,921 MANIPULATE THAT, THIS 1025 00:35:37,921 --> 00:35:39,055 INTERACTION NOT THE GENES 1026 00:35:39,055 --> 00:35:41,024 THEMSELVES, CAN YOU SEE CHANGES 1027 00:35:41,024 --> 00:35:46,262 IN JEN EXPRESSION AND HOPEFULLY 1028 00:35:46,262 --> 00:35:47,497 CELL FATE. 1029 00:35:47,497 --> 00:35:54,104 >> [INAUDIBLE QUESTION FROM 1030 00:35:54,104 --> 00:35:56,639 AUDIENCE ] -- SO IT LOOKS 1031 00:35:56,639 --> 00:35:59,476 REALLY, REALLY STRAIMPLEG TO -- 1032 00:35:59,476 --> 00:36:01,811 STRANGE TO ME, WHAT YOU ARE 1033 00:36:01,811 --> 00:36:04,280 DOING HERE DISRUPTING SUPPOSELY 1034 00:36:04,280 --> 00:36:06,716 EKING LATTERY ELEMENT LOCATED 1035 00:36:06,716 --> 00:36:08,118 INSIDE MBIP, YOU ARE SEARCHING A 1036 00:36:08,118 --> 00:36:12,522 BLOCK BETWEEN THAT ELEMENT AND 1037 00:36:12,522 --> 00:36:12,956 NKX2.1? 1038 00:36:12,956 --> 00:36:18,161 AND YOU DO SEE DECREASE LEVEL OF 1039 00:36:18,161 --> 00:36:19,496 EXPRESSION IN NKX2.1, GREAT, BUT 1040 00:36:19,496 --> 00:36:20,864 YOU'RE NOT REMOVING THAT 1041 00:36:20,864 --> 00:36:24,034 ELMETROPOLITAN AWAY FROM THE 1042 00:36:24,034 --> 00:36:26,469 HOST MBIP GENE AND YET THE LEVEL 1043 00:36:26,469 --> 00:36:27,871 OF EXPRESSION DRIVES DOWN 1044 00:36:27,871 --> 00:36:28,238 SIGNIFICANTLY. 1045 00:36:28,238 --> 00:36:35,678 ARE YOU SAYING THAT NKX2.1 IN 1046 00:36:35,678 --> 00:36:37,247 TURN UPREGULATES MBIP OR -- 1047 00:36:37,247 --> 00:36:38,715 >> THIS GOES BACK TO YOUR 1048 00:36:38,715 --> 00:36:40,683 PREVIOUS QUESTION, IS THIS A 1049 00:36:40,683 --> 00:36:42,752 PROMOTER PROTEIN COMPLEXER 1050 00:36:42,752 --> 00:36:43,353 INTERACTION OR PROMOTER 1051 00:36:43,353 --> 00:36:44,954 INTERACTION WHERE THEY'RE 1052 00:36:44,954 --> 00:36:45,755 HELPING EACH OTHER. 1053 00:36:45,755 --> 00:36:47,457 I CAN'T SAY DIRECTLY WHAT THE 1054 00:36:47,457 --> 00:36:48,058 WORRY IS. 1055 00:36:48,058 --> 00:36:49,659 BASED ON THIS, THESE ARE 2 1056 00:36:49,659 --> 00:36:52,729 PROMOTERS COMING TOGETHER THAT 1057 00:36:52,729 --> 00:36:53,997 ARE HELPING EACH OTHER 1058 00:36:53,997 --> 00:36:55,165 UPREGULATE THEIR EXPRESSION AND 1059 00:36:55,165 --> 00:36:58,068 NOW WE FORCE SEPARATION OF THIS 1060 00:36:58,068 --> 00:37:02,405 AND FOR WHATEVER REASON MBIP IS 1061 00:37:02,405 --> 00:37:04,474 STRONGLY REDUCING NKX2 PBT 1, 1062 00:37:04,474 --> 00:37:09,679 AND AS I MENTIONED, NKX2.1 IS A 1063 00:37:09,679 --> 00:37:11,581 CRITICAL REGULATORS FROM ALL 1064 00:37:11,581 --> 00:37:14,017 CELLS FROM THE MBIP, SO MY GUESS 1065 00:37:14,017 --> 00:37:15,752 IS THERE'S A MORE EVOLUTIONARY 1066 00:37:15,752 --> 00:37:17,287 CONSERVATION THERE THAT IS 1067 00:37:17,287 --> 00:37:19,589 CRITICAL FOR KEEPING NK2.1 ON SO 1068 00:37:19,589 --> 00:37:21,524 I DIDN'T MENTION IT, I'LL HOP 1069 00:37:21,524 --> 00:37:23,827 BACK REAL QUICK BUT IN THAT SOX 1070 00:37:23,827 --> 00:37:25,462 2 LOCUST STUDY, YOU SEE HERE IS 1071 00:37:25,462 --> 00:37:27,097 THAT DESPITE THE BEST EFFORTS 1072 00:37:27,097 --> 00:37:28,598 THERE'S STILL ACTUALLY AN 1073 00:37:28,598 --> 00:37:31,568 INTERACTION HERE DESPITE PUTTING 1074 00:37:31,568 --> 00:37:32,135 IN THESE SITES. 1075 00:37:32,135 --> 00:37:35,038 WHAT THIS PAPER SHOWED IS THAT 1076 00:37:35,038 --> 00:37:38,408 IT SEEMS LIKE EVOLUTION HAS 1077 00:37:38,408 --> 00:37:41,578 FOUND A WAY AROUND VARIOUS 1078 00:37:41,578 --> 00:37:42,946 PROTURBATIONS LIKE THIS BECAUSE 1079 00:37:42,946 --> 00:37:44,881 SOX 2 IS SO CRITICAL FOR 1080 00:37:44,881 --> 00:37:45,949 DEVELOPMENT THAT ANY LITTLE 1081 00:37:45,949 --> 00:37:49,586 MUTATION HERE OR THERE, THERE'S 1082 00:37:49,586 --> 00:37:51,187 MULTIPLE MECHANISMS TO HELP 1083 00:37:51,187 --> 00:37:52,689 THESE INTERACTIONS COME TOGETHER 1084 00:37:52,689 --> 00:37:54,390 SO THAT SOX 2 EXPRESSION IS 1085 00:37:54,390 --> 00:37:55,458 FOCUSED AND I WOULD ARGUE THAT 1086 00:37:55,458 --> 00:37:57,794 MAYBE THE SAME THING IS 1087 00:37:57,794 --> 00:37:59,262 HAPPENING AT NK2.1 BECAUSE IT'S 1088 00:37:59,262 --> 00:38:01,131 SO CRITICAL FOR BRAIN 1089 00:38:01,131 --> 00:38:02,365 DEVELOPMENT, LUNG DEVELOPMENT, 1090 00:38:02,365 --> 00:38:02,999 THYROID DEPRIVATIONMENT THAT 1091 00:38:02,999 --> 00:38:06,336 THERE ARE YEENS THAT HAVE THIS 1092 00:38:06,336 --> 00:38:07,270 EVOLUTIONARY CONSERVED MECHANISM 1093 00:38:07,270 --> 00:38:09,439 THAT IF EVEN WE TRY TO PERSH 1094 00:38:09,439 --> 00:38:10,607 TURB THEM WITH OTHER 1095 00:38:10,607 --> 00:38:11,708 INTERACTIONS THEY HAVE OTHER 1096 00:38:11,708 --> 00:38:13,643 ENHANCERS OR OTHER WAYS TO TRY 1097 00:38:13,643 --> 00:38:17,113 TO KEEP EXPRESSION OF THE GENE 1098 00:38:17,113 --> 00:38:18,014 AT CRITICAL TIMES. 1099 00:38:18,014 --> 00:38:19,182 THAT'S THE BEST ANSWER I HAVE. 1100 00:38:19,182 --> 00:38:22,318 BUT I THINK THAT IS A PROMOTER, 1101 00:38:22,318 --> 00:38:23,386 PROMOTER INTERACTION RATHER THAN 1102 00:38:23,386 --> 00:38:24,320 AN EN--STRATEGIES HANSER 1103 00:38:24,320 --> 00:38:25,488 PROMOTER BUT AGAIN, WE DON'T 1104 00:38:25,488 --> 00:38:26,756 HAVE GREAT EVIDENT FOR THAT 1 1105 00:38:26,756 --> 00:38:30,059 WAY OR THE OTHER. 1106 00:38:30,059 --> 00:38:31,561 >> OKAY, SO, THIS KIND OF WRAPS 1107 00:38:31,561 --> 00:38:33,429 UP THE FIRST PART OF THE STORY 1108 00:38:33,429 --> 00:38:35,732 WHERE I'VE SHOWN YOU HOW WE 1109 00:38:35,732 --> 00:38:37,267 GENERATE AN EPIGENOME ATLAS IN 1110 00:38:37,267 --> 00:38:38,468 THE EMBRYONIC MOUSE BRAIN WITH A 1111 00:38:38,468 --> 00:38:40,203 VARIETY OF DATA SO THAT WE TRULY 1112 00:38:40,203 --> 00:38:42,038 UNDERSTAND HOW THESE CELLS LOOK 1113 00:38:42,038 --> 00:38:43,306 BOTH AT AN INDIVIDUAL CELL 1114 00:38:43,306 --> 00:38:47,310 LEVEL, AND A GLOBAL CHROME TIN 1115 00:38:47,310 --> 00:38:47,644 LEVEL. 1116 00:38:47,644 --> 00:38:48,912 WE'VE IDENTIFIED MANY, WHAT I 1117 00:38:48,912 --> 00:38:51,247 WOULD CALL HIGH CONFIDENCE 1118 00:38:51,247 --> 00:38:52,515 ENHANCERS FOR THAT QUITE 1119 00:38:52,515 --> 00:38:53,449 POSSIBLY OR CRITICAL FOR 1120 00:38:53,449 --> 00:38:57,620 REGULATION OF GENES THAT ARE 1121 00:38:57,620 --> 00:38:59,022 IMPORTANT FOR DEVELOPMENT OF 1122 00:38:59,022 --> 00:39:00,557 CELLS IN THESE BRAIN REGIONS. 1123 00:39:00,557 --> 00:39:02,825 IT'S IMPORTANT FOR ME TO MAKE 1124 00:39:02,825 --> 00:39:04,260 THIS DAT AS PUBLICLY AVAILABLE 1125 00:39:04,260 --> 00:39:06,062 AS POSSIBLE, SO IN ADDITION TO 1126 00:39:06,062 --> 00:39:08,398 ALL THE SEQUENCING BEING 1127 00:39:08,398 --> 00:39:10,567 DEPOSITED WE USED UCSD BROWSER 1128 00:39:10,567 --> 00:39:12,936 SO ANYBODY CAN GO TO THE WEBSITE 1129 00:39:12,936 --> 00:39:14,871 AND SEARCH FOR THEIR FAVORITE 1130 00:39:14,871 --> 00:39:19,042 GENE AND LOOK FOR MODIFICATION, 1131 00:39:19,042 --> 00:39:20,443 HISTONE DEVELOPMENT AND WE THINK 1132 00:39:20,443 --> 00:39:24,314 IT'S A VERY USEFUL TOOL FOR 1133 00:39:24,314 --> 00:39:26,583 STUDYING BRAIN DEVELOPMENT AND I 1134 00:39:26,583 --> 00:39:28,117 GAVE EXAMPLES OF HOW PERTURBING 1135 00:39:28,117 --> 00:39:29,786 THEM CAN LEAD TO INSIGHTS AND 1136 00:39:29,786 --> 00:39:32,155 MECHANISMS THAT POSSIBLY HAVE A 1137 00:39:32,155 --> 00:39:33,323 ROLE IN HUMAN DISEASE. 1138 00:39:33,323 --> 00:39:36,626 OKAY, SO NOW THAT WE'VE KIND OF 1139 00:39:36,626 --> 00:39:38,161 DEVELOPED THIS KNOWLEDGE AND 1140 00:39:38,161 --> 00:39:40,430 THESE STRATEGIES, WE'RE PRETTY 1141 00:39:40,430 --> 00:39:41,331 WELL POSITIONED TO UNDERSTAND 1142 00:39:41,331 --> 00:39:43,600 HOW THESE ASSPEBTS OF GENES OR 1143 00:39:43,600 --> 00:39:45,735 GENE REGULATORY MECH NIMS CAN 1144 00:39:45,735 --> 00:39:49,138 AFFECT CELL FATE AND DISEASE, SO 1145 00:39:49,138 --> 00:39:50,740 WE'VE BASICALLY KIND OF BEEN 1146 00:39:50,740 --> 00:39:51,808 GOING THROUGH THIS TYPE OF 1147 00:39:51,808 --> 00:39:53,176 PIPELINE TO ARK ASSESSES THIS 1148 00:39:53,176 --> 00:39:55,278 REWRILIENCE WE HAVE A VERY 1149 00:39:55,278 --> 00:39:56,179 MUTATION, PROTURBATION, WE CAN 1150 00:39:56,179 --> 00:39:58,548 LOOK AT CHROME TIN ORGANIZATION 1151 00:39:58,548 --> 00:39:59,215 AND TRANSCRIPT ORDER OF 1152 00:39:59,215 --> 00:40:00,049 MICRONSAL WILL SINGLE CELL 1153 00:40:00,049 --> 00:40:02,018 LEVEL, WE CAN ASSAY CELL FATE, 1154 00:40:02,018 --> 00:40:03,886 WE CAN LOOK AT CELL FUNCTION 1155 00:40:03,886 --> 00:40:07,557 WITH THE COLLABORATORS 1156 00:40:07,557 --> 00:40:08,324 PERFORMING ELECTROPHYSIOLOGICAL 1157 00:40:08,324 --> 00:40:09,692 EXPERIMENTS AND THEN ULTIMATELY 1158 00:40:09,692 --> 00:40:11,694 ANIMAL BEHAVIOR AND I WILL SHOW 1159 00:40:11,694 --> 00:40:12,862 YOU HOW THESE AFFECT ANIMAL 1160 00:40:12,862 --> 00:40:14,397 BEHAVIOR AND HOW IT MIGHT RELATE 1161 00:40:14,397 --> 00:40:16,432 TO HUMAN TEASE AND SO THERE'S 3 1162 00:40:16,432 --> 00:40:17,467 STORIES DEPENDING ON TIME, I 1163 00:40:17,467 --> 00:40:19,469 WILL SEE HOW MUCH I CAN GET 1164 00:40:19,469 --> 00:40:19,702 THROUGH. 1165 00:40:19,702 --> 00:40:21,571 THE FIRST 1 IS WHERE WE LOOK 1166 00:40:21,571 --> 00:40:23,940 THEA PROTURBATION OF YEEN 1167 00:40:23,940 --> 00:40:27,377 REPRESSION USING LOSS OF ACH2 1168 00:40:27,377 --> 00:40:29,012 AND THEN ACTIVATION AND THEN 1169 00:40:29,012 --> 00:40:32,081 DISEASE, SO WE'RE FESTER TAKING 1170 00:40:32,081 --> 00:40:33,516 A SLEDGE HAMMER TO THE WHOLE 1171 00:40:33,516 --> 00:40:33,750 SYSTEM. 1172 00:40:33,750 --> 00:40:35,051 WE KNOW WHAT'S UPON HAING NOW IF 1173 00:40:35,051 --> 00:40:37,353 WE TRY TO DISRUPT GENERAL GENE 1174 00:40:37,353 --> 00:40:38,521 REPRESSION OR GENERAL GENE 1175 00:40:38,521 --> 00:40:39,789 ACTIVATION SPECIFICALLY IN THE 1176 00:40:39,789 --> 00:40:42,158 MG OF THESE MICE. 1177 00:40:42,158 --> 00:40:45,228 AND I WILL GIVE BACKGROUND ON 1178 00:40:45,228 --> 00:40:47,797 ECH 2, THIS IS A CRITICAL 1179 00:40:47,797 --> 00:40:49,065 METHYLTRANSFERASE, THIS IS A BIG 1180 00:40:49,065 --> 00:40:50,133 COMPLEX BUT WHAT'S IMPORTANT TO 1181 00:40:50,133 --> 00:40:55,004 KNOW IS THAT IT'S CRITICAL FOR 1182 00:40:55,004 --> 00:40:57,407 METHALATING K3 H3 K27 AND HAD IS 1183 00:40:57,407 --> 00:41:01,444 IMPORTANT FOR GENE REPRESSION. 1184 00:41:01,444 --> 00:41:03,346 SO BY BLOCKING OR PERTURBING ANY 1185 00:41:03,346 --> 00:41:05,181 COMPLEX, YOU WILL DISRUPT THE 1186 00:41:05,181 --> 00:41:08,017 NORMAL GENE REPRESSION THAT WILL 1187 00:41:08,017 --> 00:41:09,552 LEAD TO ABERRANT GENE 1188 00:41:09,552 --> 00:41:10,320 PROTURBATION. 1189 00:41:10,320 --> 00:41:12,155 NOW WHY THIS IS IMPORTANT 1190 00:41:12,155 --> 00:41:14,123 BECAUSE OF A NUMBER OF STUDIES 1191 00:41:14,123 --> 00:41:19,762 THAT LOOK AT EZH2 IN THE BRAIN 1192 00:41:19,762 --> 00:41:22,732 DEVELOPMENT AND WE SEE THAT LOSS 1193 00:41:22,732 --> 00:41:26,836 OF EZH2 CAN CHANGE THE LOSS OF 1194 00:41:26,836 --> 00:41:28,004 FUNCTION, MIGRATION PATTERNS, 1195 00:41:28,004 --> 00:41:30,073 AND THE ROLE THAT WAS STUDIED IN 1196 00:41:30,073 --> 00:41:32,742 BRAIN DEVELOPMENT BUT NOT 1197 00:41:32,742 --> 00:41:33,810 PARTICULARLY IN THE FOREBRAIN 1198 00:41:33,810 --> 00:41:37,246 NUROONS AND BECAUSE THESE CELLS 1199 00:41:37,246 --> 00:41:38,581 UNDERGO COMPLEX BEHAVIOR, 1200 00:41:38,581 --> 00:41:39,549 CRITICAL FOR CELL FATE WE 1201 00:41:39,549 --> 00:41:42,185 THOUGHT THIS WAS A NICE TARGET 1202 00:41:42,185 --> 00:41:43,920 TO DISRUPT GENE REPRESSION IN 1203 00:41:43,920 --> 00:41:45,988 THE MGE AND LOOK AT HOW THAT 1204 00:41:45,988 --> 00:41:47,757 SLEDGE HAMMER APPROACH AFFECTS 1205 00:41:47,757 --> 00:41:49,192 CELL FATE. 1206 00:41:49,192 --> 00:41:52,228 SO WE USE AN NPXCREE TARGET AND 1207 00:41:52,228 --> 00:41:57,266 THIS IS TARGETED TO THE MGE WITH 1208 00:41:57,266 --> 00:41:58,468 THE FLOX EZH2, AND YOU SEE THE 1209 00:41:58,468 --> 00:41:59,535 NORMAL EXPRESSION AND THEN THE 1210 00:41:59,535 --> 00:42:01,838 BEING IMO OUT THAT EXPRESSION IS 1211 00:42:01,838 --> 00:42:04,807 STRONGLY DOWN REGULATED 1212 00:42:04,807 --> 00:42:06,576 SPECIFICALLY IN THE MG BUT 1213 00:42:06,576 --> 00:42:08,077 INTACT IN THE LG AND CORTEX. 1214 00:42:08,077 --> 00:42:10,880 THE READ OUT FOR THIS YEEN IS 1215 00:42:10,880 --> 00:42:15,618 CRITICAL FOR METHALATING THE H3 1216 00:42:15,618 --> 00:42:18,054 K RESIDUE AND WE SEE A DOWN 1217 00:42:18,054 --> 00:42:19,889 REGULATION OF THIS HISTONE 1218 00:42:19,889 --> 00:42:20,223 MODIFICATION. 1219 00:42:20,223 --> 00:42:21,924 SO WHAT WE DID IS WE TOOK THE 1220 00:42:21,924 --> 00:42:22,892 MICE, HARVESTED THE BRAINS IN 1221 00:42:22,892 --> 00:42:25,928 THE ADULT AND WE COULD PERFORM 1222 00:42:25,928 --> 00:42:30,967 AMINO STAINING SO SST AND PV ARE 1223 00:42:30,967 --> 00:42:32,268 MARKERS FOR 2 SUBTYPES THAT,A 1224 00:42:32,268 --> 00:42:34,470 RISE FROM THE NG, BASICALLY 95% 1225 00:42:34,470 --> 00:42:35,972 OF THE CELLS IN THE CORTEX THAT 1226 00:42:35,972 --> 00:42:39,642 ARE BORN IN THE MG EXPRESS 1227 00:42:39,642 --> 00:42:42,645 EITHER SST OR PV, SO THESE ARE 1228 00:42:42,645 --> 00:42:43,946 NONOVERLAPPING SUBGROUPS THAT 1229 00:42:43,946 --> 00:42:44,981 ENCOMPASS ALL CELLS IN THE 1230 00:42:44,981 --> 00:42:46,082 CORTEX, SO WHAT YOU SEE IN THE 1231 00:42:46,082 --> 00:42:48,050 KNOCK OUT HERE IS WE SEE AN 1232 00:42:48,050 --> 00:42:49,852 OVERALL DECREASE IN THE NUMBER 1233 00:42:49,852 --> 00:42:51,587 OF TOMATO CELLS, TOMATO IS A 1234 00:42:51,587 --> 00:42:53,389 MARKER FOR ALL MG CELLS, THIS IS 1235 00:42:53,389 --> 00:42:54,557 JUST A REPORTER SO MAYBE CAN YOU 1236 00:42:54,557 --> 00:42:56,559 TELL HERE BUT THERE'S ABOUT A 1237 00:42:56,559 --> 00:42:59,295 20% REDUCTION OR SO OF ALL MG 1238 00:42:59,295 --> 00:43:01,164 CELLS IN THE CORTEX BUT WE 1239 00:43:01,164 --> 00:43:02,899 ACTUALLY SAW A DIFFERENT AFFECT 1240 00:43:02,899 --> 00:43:03,466 IN CELL FATE. 1241 00:43:03,466 --> 00:43:06,269 IF YOU LOOK AT SST IN GREEN HERE 1242 00:43:06,269 --> 00:43:08,438 THERE'S AN INCREASE IN THE 1243 00:43:08,438 --> 00:43:12,208 AMOUNT OF SST INTERNONS AND 1244 00:43:12,208 --> 00:43:14,377 CORRESPONDING DECREASE IN THE PV 1245 00:43:14,377 --> 00:43:15,611 CELLS. 1246 00:43:15,611 --> 00:43:17,847 SO THIS FEWER CELLS DUE TO THIS 1247 00:43:17,847 --> 00:43:19,682 MUTATION SPECIFICALLY IN THE 1248 00:43:19,682 --> 00:43:19,916 MGE. 1249 00:43:19,916 --> 00:43:22,385 I WON'T SHOW YOU THE DATA. 1250 00:43:22,385 --> 00:43:24,654 THERE'S ALSO INTERNEURONS THAT 1251 00:43:24,654 --> 00:43:27,089 ARE IN THE STRIATUM AND 1252 00:43:27,089 --> 00:43:28,691 HIPPOCAMPUS AND WE SAW MORE FATE 1253 00:43:28,691 --> 00:43:32,094 SWITCH WHERE WE SAW MORE PV AND 1254 00:43:32,094 --> 00:43:32,862 SST CELLS. 1255 00:43:32,862 --> 00:43:35,164 I WILL NOTE THESE ARE CYCLING 1256 00:43:35,164 --> 00:43:36,966 CELLS, WE USE A DIFFERENT CREE 1257 00:43:36,966 --> 00:43:39,068 LINE TO KNOCK OUT POST MITOTIC 1258 00:43:39,068 --> 00:43:40,136 CELLS SO AFTER THESE CELLS ARE 1259 00:43:40,136 --> 00:43:41,971 BORN AND WE DO NOT SEE THESE 1260 00:43:41,971 --> 00:43:42,839 CHANGES IN CELL FATE. 1261 00:43:42,839 --> 00:43:46,742 SO THIS TELLS US THE FUNCTION OF 1262 00:43:46,742 --> 00:43:49,245 ECH2 IS CRITICAL, AND ONCE THEY 1263 00:43:49,245 --> 00:43:50,346 BECOME POST MITOTIC AND REMOVING 1264 00:43:50,346 --> 00:43:53,316 THOSE IN THE CELLS DOES NOT 1265 00:43:53,316 --> 00:43:54,884 CHANGE CELL FATE. 1266 00:43:54,884 --> 00:43:56,552 SO WITH OUR COLLEAGUES 1267 00:43:56,552 --> 00:43:59,222 [INDISCERNIBLE] AT NIMH, THEY 1268 00:43:59,222 --> 00:43:59,922 PERFORMED ELECTROPHYSIOLOGY 1269 00:43:59,922 --> 00:44:03,693 RECORDINGS TO SEE IF IN THESE 1270 00:44:03,693 --> 00:44:05,061 MUTANTS IS THE PHYSIOLOGY 1271 00:44:05,061 --> 00:44:05,795 AFFAIRS TEAM LEADERRERRED? 1272 00:44:05,795 --> 00:44:07,196 SO THEY LOOKED AT ABOUT 20 OF 1273 00:44:07,196 --> 00:44:08,731 THESE CELL ANDS THERE WAS VERY, 1274 00:44:08,731 --> 00:44:09,899 VERY LITTLE DIFFERENCES BUT WHAT 1275 00:44:09,899 --> 00:44:11,300 THEY DID NOTICE, STRIKING 1276 00:44:11,300 --> 00:44:19,475 CHANGES IN THE MORPHOLOGY, SO 1277 00:44:19,475 --> 00:44:21,911 THESE ARE BARK LBUMIN, AND 1278 00:44:21,911 --> 00:44:23,346 THERE'S MORE BRANCH XG THERE'S 1279 00:44:23,346 --> 00:44:24,614 QUANTIFIED OVER HERE SO WHAT 1280 00:44:24,614 --> 00:44:26,349 THIS MEANS THAT IN THIS MOUSE 1281 00:44:26,349 --> 00:44:28,451 LINE WHERE YOU HAVE FEWER PV 1282 00:44:28,451 --> 00:44:31,354 CELLS, THE PV CELLS REMAIN HAVE 1283 00:44:31,354 --> 00:44:32,655 GREATER AXONAL PROJECTIONS AND 1284 00:44:32,655 --> 00:44:34,624 GREATER TARGETS AND PRESUMABLY 1285 00:44:34,624 --> 00:44:36,325 GREATER SYNAPSES, SO WE'VE 1286 00:44:36,325 --> 00:44:38,194 EITHER SINCE THE COMPENSATION 1287 00:44:38,194 --> 00:44:39,962 MECHANISM, HOW DOES THE BRAIN 1288 00:44:39,962 --> 00:44:42,365 COMPENSATE FOR FUHRER INHIBITORY 1289 00:44:42,365 --> 00:44:44,100 NEURONS AND THE WAY TO DO IS 1290 00:44:44,100 --> 00:44:46,035 THAT THE 1S THAT SURVIVE, THEY 1291 00:44:46,035 --> 00:44:47,103 HAVE MORE PROCESSES, CONTACT 1292 00:44:47,103 --> 00:44:50,540 MORE CELLS TO TRY TO INCREASE 1293 00:44:50,540 --> 00:44:50,840 INHIBITION. 1294 00:44:50,840 --> 00:44:55,378 AND THEN WHAT WE WANT TO DO IS 1295 00:44:55,378 --> 00:44:56,546 PERFORM MULTIOHM ANALYSIS SO IN 1296 00:44:56,546 --> 00:44:57,914 ADDITION TO LOOKING AT CHANGES 1297 00:44:57,914 --> 00:44:59,115 IN CELL FATE WE CAN UNDERSTAND 1298 00:44:59,115 --> 00:45:02,251 WHAT'S GOING ON GENETICALLY AND 1299 00:45:02,251 --> 00:45:03,219 GENE REGULATORY MECHANISMS IN 1300 00:45:03,219 --> 00:45:04,754 THESE CELLS, AT THIS POINT 1301 00:45:04,754 --> 00:45:08,424 MULTIOHM WAS AVAILABLE, SO THIS 1302 00:45:08,424 --> 00:45:10,893 IS SINGLE CELL ATAC AND RNA IN 1303 00:45:10,893 --> 00:45:14,030 THE SAME CELL SO WE PERFORM 1304 00:45:14,030 --> 00:45:15,631 MULTIOHM ANALYSIS, KNOCKOUT MICE 1305 00:45:15,631 --> 00:45:16,766 ON 2 DIFFERENT EMBRYONIC AGES. 1306 00:45:16,766 --> 00:45:18,467 AGAIN WE CAN KIND OF LOOK AT 1307 00:45:18,467 --> 00:45:20,303 GENE EXPRESSION PROFILES SO WE 1308 00:45:20,303 --> 00:45:26,809 KNOW THE TRANSLATIONAL -- TRAJO 1309 00:45:26,809 --> 00:45:27,643 MOST MITOTIC CELLS ARE NEAR THE 1310 00:45:27,643 --> 00:45:29,445 BOTTOM AND WE WANTED TO SEE, ARE 1311 00:45:29,445 --> 00:45:31,347 THERE SIGNATURES IN THE EMBRYO 1312 00:45:31,347 --> 00:45:33,182 THAT ARE INDICATIVE OR CAUSATIVE 1313 00:45:33,182 --> 00:45:34,483 FOR THESE CHANGES IN CELL FATE. 1314 00:45:34,483 --> 00:45:37,286 SO IF WE LOOK AT SST HERE, AND 1315 00:45:37,286 --> 00:45:38,955 AGAIN, THIS IS -- THERE'S MORE 1316 00:45:38,955 --> 00:45:41,057 SST CELLS IN THE ADULT MOUSE, WE 1317 00:45:41,057 --> 00:45:42,892 SEE AN INCREASE IN SST 1318 00:45:42,892 --> 00:45:44,894 EXPRESSION HERE IN THE KNOCK OUT 1319 00:45:44,894 --> 00:45:49,098 MOUSE, AT E15, COMPARED TO THE 1320 00:45:49,098 --> 00:45:49,365 WILD-TYPE. 1321 00:45:49,365 --> 00:45:51,200 CONVERSELY, PARVE O BIEWMIN IS 1322 00:45:51,200 --> 00:45:53,369 NOT EXPRESSED, IT DOESN'T COME 1323 00:45:53,369 --> 00:45:54,203 ON UNTIL POSTINATEALLY, BUT 1324 00:45:54,203 --> 00:45:57,840 THERE ARE A COUPLE GENES MAP 2 C 1325 00:45:57,840 --> 00:45:59,942 AND MAP B, THAT ARE PREDICTIVE 1326 00:45:59,942 --> 00:46:00,876 OF THE WILD-TYPE CELLS. 1327 00:46:00,876 --> 00:46:02,678 YOU CAN SEE HERE WE HAVE STRONG 1328 00:46:02,678 --> 00:46:04,146 EXPRESSION OF THE CELLS AND IN 1329 00:46:04,146 --> 00:46:05,781 THE KNOCK OUT YOU SEE DOWN 1330 00:46:05,781 --> 00:46:06,582 REGULATION OF THESE. 1331 00:46:06,582 --> 00:46:08,684 IN ADDITION BECAUSE IT'S 1332 00:46:08,684 --> 00:46:10,753 MULTIOHM, WE CAN LOOK AT THE 1333 00:46:10,753 --> 00:46:12,421 MOTIF FOR THE BINDING SITE OF 1334 00:46:12,421 --> 00:46:14,123 THESE FACTORS IN OUR DNA, SORRY 1335 00:46:14,123 --> 00:46:16,959 IN OUR CELLS AND YOU CAN SEE 1336 00:46:16,959 --> 00:46:18,761 AGAIN, A DECREASE IN THE MOTIF 1337 00:46:18,761 --> 00:46:19,962 ACCESSIBILITY IN THE KNOCK OUTS 1338 00:46:19,962 --> 00:46:21,664 COMPARED TO THE WILD-TYPE. 1339 00:46:21,664 --> 00:46:25,501 SO NOW WE CAN IDENTIFY IN THE 1340 00:46:25,501 --> 00:46:28,037 EMBRYO, SOME GENETIC LOGIC AS TO 1341 00:46:28,037 --> 00:46:30,573 WHY WE SEE THE CELL FATE CHANGES 1342 00:46:30,573 --> 00:46:33,142 INCREASED IN MET STATIN IN D15 1343 00:46:33,142 --> 00:46:35,645 AND DECREASED MARKERS OF PV 1344 00:46:35,645 --> 00:46:39,382 CELLS ALREADY PRESENT ALSO AT 1345 00:46:39,382 --> 00:46:39,548 E15. 1346 00:46:39,548 --> 00:46:41,617 WE CAN DO KIND OF UNBIASED 1347 00:46:41,617 --> 00:46:43,819 COMPARISON TO LOOK AT GENES THAT 1348 00:46:43,819 --> 00:46:44,854 ARE UPREGULATED AT THESE 1349 00:46:44,854 --> 00:46:46,589 DIFFERENT AGES IN THESE VOLCANO 1350 00:46:46,589 --> 00:46:48,691 PLOTS AND AGAIN YOU SEE 2 GENES 1351 00:46:48,691 --> 00:46:50,559 HERE PREDICTED OF PV CELLS THAT 1352 00:46:50,559 --> 00:46:51,861 ARE WILD-TYPE ENRICHED MEANING 1353 00:46:51,861 --> 00:46:54,063 DOWN REGULATED OF THE KNOCK OUT 1354 00:46:54,063 --> 00:46:56,632 AND SST OVER HERE IS STRONGLY 1355 00:46:56,632 --> 00:46:58,801 ENRICHED IN THE KNOCK OUT. 1356 00:46:58,801 --> 00:47:00,803 THERE'S A FAIR AMOUNT OF OTHER 1357 00:47:00,803 --> 00:47:01,637 GENETIC ANALYSIS WE'VE DONE WITH 1358 00:47:01,637 --> 00:47:03,506 THIS BUT I DON'T WANT TO BELABOR 1359 00:47:03,506 --> 00:47:05,241 THE POINT TOO MUCH IS BASICALLY 1360 00:47:05,241 --> 00:47:06,776 THE TAKE HOME MESSAGE IS THAT WE 1361 00:47:06,776 --> 00:47:09,178 SEE A FATE SWITCH IN THIS MOUSE, 1362 00:47:09,178 --> 00:47:12,882 AND ALSO, WE SEE CHANGES IN GENE 1363 00:47:12,882 --> 00:47:14,417 EXPRESSION IN THE EMBRYO THAT 1364 00:47:14,417 --> 00:47:16,252 ARE CERTAINLY CAUSATIVE FOR THAT 1365 00:47:16,252 --> 00:47:16,786 FATE SWITCH. 1366 00:47:16,786 --> 00:47:20,356 OKAY, NOW I'M GOING TO JUMP TO 1367 00:47:20,356 --> 00:47:21,657 ANOTHER STORY, SO THAT WAS 1368 00:47:21,657 --> 00:47:23,092 LOOKING AT GENE REPRESSION, AND 1369 00:47:23,092 --> 00:47:25,127 SO THIS IS -- SORRY, I FORGOT I 1370 00:47:25,127 --> 00:47:25,695 HAD THIS SLIDE. 1371 00:47:25,695 --> 00:47:30,199 SO IN ARK DITION TO THE MULTIOHM 1372 00:47:30,199 --> 00:47:31,300 DATA WE ALSO PERFORM TAG TO 1373 00:47:31,300 --> 00:47:34,170 LOOK THEA DIFFERENCES OF THIS 1374 00:47:34,170 --> 00:47:36,906 KTWEIVE METHYLATION, AGAIN THIS 1375 00:47:36,906 --> 00:47:40,009 IS THE TARGET OF ECH2, AND THE 1376 00:47:40,009 --> 00:47:41,711 TAKE HOME MESSAGE WE FOUND THAT 1377 00:47:41,711 --> 00:47:45,047 IN THIS KNOCK OUT THE CHANGES IN 1378 00:47:45,047 --> 00:47:46,449 THE K27 METHYLATION WAS NOT 1379 00:47:46,449 --> 00:47:48,084 EQUIVALENT ACROSS THE GENOME. 1380 00:47:48,084 --> 00:47:50,619 WE SAW SOME LOCI THAT WERE MORE 1381 00:47:50,619 --> 00:47:54,290 SUSCEPTIBLE SUCH AS THE FOX 1382 00:47:54,290 --> 00:47:55,925 PPAYS LOCI HERE, YOU SEE THE 1383 00:47:55,925 --> 00:47:57,460 WILD-TYPE AND YOU SEE A REALLY 1384 00:47:57,460 --> 00:47:59,328 STRONG REDUCTION IN MULTIPLE 1385 00:47:59,328 --> 00:48:00,696 REPLICATES IN THE KNOCK OUT. 1386 00:48:00,696 --> 00:48:04,100 SO THIS IS A LOCI THAT SEEMS TO 1387 00:48:04,100 --> 00:48:11,140 BE EXTREMELY SUSCEPTIBLE TO THIS 1388 00:48:11,140 --> 00:48:11,474 MANIPULATION. 1389 00:48:11,474 --> 00:48:12,575 CONVERSELY, NKX2.1, YOU CAN SEE 1390 00:48:12,575 --> 00:48:13,976 A RELATIVE INCREASE IN THE LEVEL 1391 00:48:13,976 --> 00:48:15,277 OF METHYLATION, SO THIS DOESN'T 1392 00:48:15,277 --> 00:48:17,246 MEAN IT'S MORE METHALATED THAN 1393 00:48:17,246 --> 00:48:18,647 THE WILD-TYPE, IT JUST MEANS 1394 00:48:18,647 --> 00:48:20,282 IT'S MORE RESISTANT, THE 1395 00:48:20,282 --> 00:48:20,850 METHYLATION WAS NOT REMOVED 1396 00:48:20,850 --> 00:48:22,017 BECAUSE ALL OF THIS IS RELATIVE. 1397 00:48:22,017 --> 00:48:23,652 SO THIS KIND OF GOES BACK TO 1398 00:48:23,652 --> 00:48:25,488 YOUR EARLIER POINT, I THINK 1399 00:48:25,488 --> 00:48:28,624 THERE'S EVIDENCE THAT NK2.1 HAS 1400 00:48:28,624 --> 00:48:30,926 SOME SORT OF -- BECAUSE OF ITS 1401 00:48:30,926 --> 00:48:34,063 CRITICAL ROLE IN FATE, I THINK 1402 00:48:34,063 --> 00:48:35,297 THERE'S EVOLUTIONARY 1403 00:48:35,297 --> 00:48:36,866 CONSERVATION THAT HAS CAUSED 1404 00:48:36,866 --> 00:48:40,369 THIS LOCI TO BE MORE RESISTANT 1405 00:48:40,369 --> 00:48:42,138 TO VARIOUS CHANGES BECAUSE 1406 00:48:42,138 --> 00:48:45,841 DESPITE KNOCKING OUT A GENE FRM 1407 00:48:45,841 --> 00:48:46,709 RETIRED KRAL METHYLATION, IT 1408 00:48:46,709 --> 00:48:49,345 TRIES AS BEST IT CAN TO REFUTE 1409 00:48:49,345 --> 00:48:50,746 THIS PROTURBATION AND KEEP HIGH 1410 00:48:50,746 --> 00:48:52,148 LEVELS OF METHYLATION HERE, SO 1411 00:48:52,148 --> 00:48:54,116 THAT IT CAN BE REPRESSED OR 1412 00:48:54,116 --> 00:48:58,320 ACTIVATED IN THE PROPER WAY. 1413 00:48:58,320 --> 00:48:59,388 >> OKAY, SO THAT WAS YEEN 1414 00:48:59,388 --> 00:49:00,890 REPRESSION AND NOW I WANT TO 1415 00:49:00,890 --> 00:49:02,725 TALK A LITTLE BIT ABOUT GENE 1416 00:49:02,725 --> 00:49:04,894 ACTIVATION BUT ALSO WHY THIS 1417 00:49:04,894 --> 00:49:06,395 PARTICULAR MODE OF LOOKING AT 1418 00:49:06,395 --> 00:49:09,732 GENE ACTIVATION IS CRITICAL. 1419 00:49:09,732 --> 00:49:12,468 SO METHYLATION AT H3 K4, IS 1420 00:49:12,468 --> 00:49:13,602 CRITICAL FOR GENE ACTIVATION, 1421 00:49:13,602 --> 00:49:15,538 BUT THERE'S A LOT OF EVIDENCE 1422 00:49:15,538 --> 00:49:17,673 THROUGH YEARS OF WORK THAT HAVE 1423 00:49:17,673 --> 00:49:19,975 SHOWN THAT THIS REGULATION OF 1424 00:49:19,975 --> 00:49:21,377 THIS LOCI AND THIS METHYLATION 1425 00:49:21,377 --> 00:49:24,713 IS ASSOCIATED WITH A VARIETY OF 1426 00:49:24,713 --> 00:49:27,449 NEURAL DEVELOPMENTAL DECS, 1427 00:49:27,449 --> 00:49:32,154 SCHIZOPHRENIA, ALSO, DRUG 1428 00:49:32,154 --> 00:49:33,322 DEPENDENCY, THIS IS A NICE 1429 00:49:33,322 --> 00:49:35,357 SUMMARY OF A HOST OF YEENS THAT 1430 00:49:35,357 --> 00:49:37,493 ARE EPIGENETIC MACHINERY THAT 1431 00:49:37,493 --> 00:49:39,728 CAN EITHER WRITE OR REMOVE 1432 00:49:39,728 --> 00:49:41,564 EPIGENETIC MARKS AND I WILL 1433 00:49:41,564 --> 00:49:43,966 HIGHLIGHT 3 GENES HERE, ALL 3 OF 1434 00:49:43,966 --> 00:49:45,034 THESE YEENS TARGET METHYLATION 1435 00:49:45,034 --> 00:49:46,435 OF THIS PARTICULAR RESIDUE AND 1436 00:49:46,435 --> 00:49:49,505 ALL HAVE VARIOUS SYMPTOMS AND 1437 00:49:49,505 --> 00:49:49,939 DRIVE HUMAN DISEASE. 1438 00:49:49,939 --> 00:49:51,473 SO WE THINK A BETTER 1439 00:49:51,473 --> 00:49:54,009 UNDERSTANDING OF HOW THIS 1440 00:49:54,009 --> 00:49:55,945 PARTICULAR LOCI, GETS METHALATED 1441 00:49:55,945 --> 00:49:57,213 IT'S CRITICAL FOR UNDERSTANDING 1442 00:49:57,213 --> 00:49:59,415 GENE ACTIVATION AS WELL AS MAYBE 1443 00:49:59,415 --> 00:50:02,885 KIND OF THE UNDERLYING CAUSES OF 1444 00:50:02,885 --> 00:50:03,552 VARIOUS HUMAN DISEASES. 1445 00:50:03,552 --> 00:50:05,187 NOW WHAT MAKES STUDYING THIS 1446 00:50:05,187 --> 00:50:07,690 COMPLICATE SIDE THAT UNLIKE THE 1447 00:50:07,690 --> 00:50:10,025 PREVIOUS EXAMPLE WHERE IT'S THE 1448 00:50:10,025 --> 00:50:11,861 PRIMARY METHYLTRANSFERASE THAT 1449 00:50:11,861 --> 00:50:13,696 TARGETS THAT HISTONE, IN THIS 1450 00:50:13,696 --> 00:50:15,431 CASE YOU HAVE 6 DIFFERENT 1451 00:50:15,431 --> 00:50:18,400 ENZYMES THAT CAN METHALATE K4, 1452 00:50:18,400 --> 00:50:20,236 THAT CAN METHALATE THE K4 1453 00:50:20,236 --> 00:50:22,872 RESIDUE AND THESE ARE EXPRESSED 1454 00:50:22,872 --> 00:50:25,074 OFTEN TIMES IN OVERLAPPING CELLS 1455 00:50:25,074 --> 00:50:27,476 AND CAN OVERLAPPING COMPENSATORY 1456 00:50:27,476 --> 00:50:28,844 FUNCTION SO SIMPLY KNOCKING OUT 1457 00:50:28,844 --> 00:50:30,846 1 OF THESE IS SOMETIMES NOT 1458 00:50:30,846 --> 00:50:32,248 SUFFICIENT TO HAVE A ROBUST 1459 00:50:32,248 --> 00:50:33,182 EFFECT. 1460 00:50:33,182 --> 00:50:34,783 NOW WE WERE LUCKY THAT A 1461 00:50:34,783 --> 00:50:37,686 COLLEAGUE HERE AT NIDDK, DR. GUY 1462 00:50:37,686 --> 00:50:39,655 HAD DEVELOPED A MOUSE WHERE HE 1463 00:50:39,655 --> 00:50:43,926 MUTATED THE SLICE IN 4 RESIDUE 1464 00:50:43,926 --> 00:50:48,297 TO A AN M METHYL, I FORGET, K4 M 1465 00:50:48,297 --> 00:50:51,967 AND BASICALLY WHAT THIS DOES IS 1466 00:50:51,967 --> 00:50:54,103 BLOCKS THE LIESING, BECAUSE IT 1467 00:50:54,103 --> 00:50:55,838 CAN NEVER BE LIESINE BECAUSE 1468 00:50:55,838 --> 00:50:57,673 IT'S NEVER THERE ANYMORE AND 1469 00:50:57,673 --> 00:50:59,575 IT'S CREE DEPENDENT WHICH,A 1470 00:50:59,575 --> 00:51:00,643 LOWSITOUS EXPRESS IN MUTATED 1471 00:51:00,643 --> 00:51:02,077 HISTONE IN THE CELLS WE WANT AND 1472 00:51:02,077 --> 00:51:03,846 WHAT'S NICE IS THAT THIS IS 1473 00:51:03,846 --> 00:51:08,150 TRANSGENIC SO BASICALLY YOU HAVE 1474 00:51:08,150 --> 00:51:10,586 IN A K-FORM HOMOZYGOUS MOUSE, 1475 00:51:10,586 --> 00:51:13,055 YOU HAVE 2 WILD-TYPE ARK LEGALS, 1476 00:51:13,055 --> 00:51:14,690 AND ALSO ELSEWHERE WHERE THIS 1477 00:51:14,690 --> 00:51:17,026 WAS INSERTED YOU HAVE 2 MUTANT 1478 00:51:17,026 --> 00:51:17,393 ALLELES. 1479 00:51:17,393 --> 00:51:20,029 SO THIS IS MIMICS HUMAN DISEASE 1480 00:51:20,029 --> 00:51:22,197 WHERE YOU HAVE A KNOCK DOWN OR 1481 00:51:22,197 --> 00:51:24,166 MUTATION IN AN ENZYME THAT 1482 00:51:24,166 --> 00:51:25,801 AFFECTS THIS AND YOU'RE LOOKING 1483 00:51:25,801 --> 00:51:27,903 AT A LOSS, NOT A COMPLETE LOSS 1484 00:51:27,903 --> 00:51:28,971 OF FUNCTION BUT A IMK DOWN. 1485 00:51:28,971 --> 00:51:31,507 SO WE THINK THIS IS A COOL TOOL 1486 00:51:31,507 --> 00:51:34,343 TO BASICALLY HAVE A HOMOZYGOUS 1487 00:51:34,343 --> 00:51:36,111 MOUSE AS A FUNCTIONAL HAT TO 1488 00:51:36,111 --> 00:51:40,316 STUDY MANIPULATION OF THIS 1489 00:51:40,316 --> 00:51:40,549 RESIDUE. 1490 00:51:40,549 --> 00:51:43,619 SO WHAT WE SEE OUT OF THESE 1491 00:51:43,619 --> 00:51:44,720 HOMOZYGOUS MOUSE, THEY'RE 1492 00:51:44,720 --> 00:51:45,788 ACTUALLY UNDERWEIGHT EARLY ON, A 1493 00:51:45,788 --> 00:51:47,957 NUMBER OF THEM DIE, 50% OF THEM 1494 00:51:47,957 --> 00:51:49,992 BY BY 20 WEEKS, THEIR BRAINS ARE 1495 00:51:49,992 --> 00:51:51,527 ALSO SMALLER WHICH YOU CAN SEE 1496 00:51:51,527 --> 00:51:53,729 HERE, THE CORTEX HAS DECREASED 1497 00:51:53,729 --> 00:51:54,196 THICKNESS, BUT WHAT'S 1498 00:51:54,196 --> 00:51:56,065 INTERESTING IS THAT THE 1S THAT 1499 00:51:56,065 --> 00:51:57,566 SURVIVE AFTER ABOUT 12 MONTHS 1500 00:51:57,566 --> 00:51:58,834 BECOME QUITE OBESE, THEY CAN BE 1501 00:51:58,834 --> 00:52:02,271 DOUBLE THE WEIGHT OF THEIR 1502 00:52:02,271 --> 00:52:04,773 COUNTERPARTS AND AGAIN, I WON'T 1503 00:52:04,773 --> 00:52:06,508 GO THROUGH THIS IN MUCH DETAIL 1504 00:52:06,508 --> 00:52:09,044 BUT WE ARE FAIRLY CONFIDENT THIS 1505 00:52:09,044 --> 00:52:11,513 IS PROTURBATIONS IN THE 1506 00:52:11,513 --> 00:52:12,715 HYPOTHALAMUS, AND HYPOTHAT 1507 00:52:12,715 --> 00:52:13,882 WILLUS IS CRITICAL FOR FEEDING 1508 00:52:13,882 --> 00:52:14,516 BEHOUSTON AND RICE FOOTBALL AND 1509 00:52:14,516 --> 00:52:16,485 VARIETY OF OTHER HORMONAL 1510 00:52:16,485 --> 00:52:16,752 BEHAVIORS. 1511 00:52:16,752 --> 00:52:18,921 NOW IF YOU LOOK AT CHANGES IN 1512 00:52:18,921 --> 00:52:22,358 THE INTERNEURON COUNTS, WE SEE 1513 00:52:22,358 --> 00:52:23,993 THERE'S A DECREASE IN MEAS MICE 1514 00:52:23,993 --> 00:52:27,529 IN THE NEURONS AND A DEC -- 1515 00:52:27,529 --> 00:52:29,932 STRONG DECREASE IN P A LBALBUT 1516 00:52:29,932 --> 00:52:33,002 MIN AND IN THE ECH 2 MOUSE WE 1517 00:52:33,002 --> 00:52:34,737 SAW FATE SWITCH WHERE 1 WENT UP 1518 00:52:34,737 --> 00:52:36,005 AND 1 WENT DOWN. 1519 00:52:36,005 --> 00:52:38,474 IN THIS MOUSE WE SEE A DECREASE 1520 00:52:38,474 --> 00:52:39,775 OF BOTH CELL TYPES AND THIS IS 1521 00:52:39,775 --> 00:52:41,076 MORE STRIKING IN THE HIPPOCAMPUS 1522 00:52:41,076 --> 00:52:43,312 WHERE IF YOU LOOK IN THE 1523 00:52:43,312 --> 00:52:44,413 WILD-TYPE, THERE'S LOTS OF 1524 00:52:44,413 --> 00:52:45,047 TOMATO CELLS HERE AND YOU SLEEP 1525 00:52:45,047 --> 00:52:45,881 APNEA AND OBESITYY THE HUGE 1526 00:52:45,881 --> 00:52:49,284 POCKET WHERE IS THERE'S FEW 1527 00:52:49,284 --> 00:52:50,486 INTERNEURONS IN THESE MICE AND 1528 00:52:50,486 --> 00:52:52,121 WE QUANTIFIED ALL THAT AS WELL. 1529 00:52:52,121 --> 00:52:53,789 I WANT TO SHOW YOU A VIDEO OF 1530 00:52:53,789 --> 00:52:55,424 THESE MICE, SO THIS IS A KNOCK 1531 00:52:55,424 --> 00:52:58,460 OUT MOUSE HERE, THESE MICE HAVE 1532 00:52:58,460 --> 00:52:59,728 SPONTANEOUS EPILEPSY, YOU SEE 1533 00:52:59,728 --> 00:53:00,662 THESE PRONT PAWS HAVING THIS 1534 00:53:00,662 --> 00:53:03,866 BACK AND FORTH MOVEMENT AND THEN 1535 00:53:03,866 --> 00:53:06,869 KIND OF STARTS TO ARE HAVE THESE 1536 00:53:06,869 --> 00:53:08,470 RAPID MOVEMENTS, THIS IS UPDATER 1537 00:53:08,470 --> 00:53:10,339 GOING A SPONTANEOUS SEIZURE, 1538 00:53:10,339 --> 00:53:11,974 HERE'S ANOTHER EXAMPLE, THIS 1'S 1539 00:53:11,974 --> 00:53:14,076 KIND OF SEVERE, KIND OF A 1540 00:53:14,076 --> 00:53:15,010 CATATONIC SEIZURE, IT WILL 1541 00:53:15,010 --> 00:53:17,012 FREEZE UP AND FALL OVER, SO 1542 00:53:17,012 --> 00:53:18,047 THESE HOMOZYGOUS MOUSE ARE VERY 1543 00:53:18,047 --> 00:53:23,986 SICK IN TERMS OF THE FACT THEY 1544 00:53:23,986 --> 00:53:24,620 DISPLAY SPONTANEOUS DISEASURES 1545 00:53:24,620 --> 00:53:27,022 AND AGAIN THIS CORRELATES WITH 1546 00:53:27,022 --> 00:53:28,991 FEWER INTERNEURONS, SO YOU WE 1547 00:53:28,991 --> 00:53:31,326 CAN DO TESTS TO LOOK AT SEVERITY 1548 00:53:31,326 --> 00:53:32,995 OF THE SEIZURE, IT INDUCES 1549 00:53:32,995 --> 00:53:34,530 SEIZURES AND YOU SEE THAT THE 1550 00:53:34,530 --> 00:53:38,400 HOMOZYGOUS HERE IN RED, HAVE A 1551 00:53:38,400 --> 00:53:40,269 MUCH MORE SUSCEPTIBLE TO THIS 1552 00:53:40,269 --> 00:53:41,737 TASK, THEY UPDATER GO SEIZURES 1553 00:53:41,737 --> 00:53:43,338 OF MUCH MORE SEVERE LEVEL 1554 00:53:43,338 --> 00:53:45,574 COMPARED TO THE BLUE IN THE 1555 00:53:45,574 --> 00:53:49,478 HETEROZYGOUS WILD-TYPE MICE BE 1556 00:53:49,478 --> 00:53:50,946 AND WE'VE ALSO DONE 1557 00:53:50,946 --> 00:53:51,580 CERTAINLY--CERTAINLY LECT ROUGH 1558 00:53:51,580 --> 00:53:53,949 ATOM PHYSIOLOGY ON THESE, ON THE 1559 00:53:53,949 --> 00:53:54,917 LEFT THERE ARE 3 DIFFERENT 1560 00:53:54,917 --> 00:53:56,652 SUBTYPES AND YOU CAN SEE THE RED 1561 00:53:56,652 --> 00:53:58,720 DOTS AND GREEN DOTS AND BLUE 1562 00:53:58,720 --> 00:54:00,789 DOTS ARE FAIRLY WELL CLUSTERED 1563 00:54:00,789 --> 00:54:02,858 TOGETHER IF YOU LOOK IN THE 1564 00:54:02,858 --> 00:54:04,059 MUTANT THERE MUCH MORE SPARSE, 1565 00:54:04,059 --> 00:54:07,362 SO THIS INDICATES THE ACTUAL 1566 00:54:07,362 --> 00:54:09,531 PHYSIOLOGICAL CELLS IN THE 1567 00:54:09,531 --> 00:54:12,067 MUTANT THERE SEEM TO BE MUCH 1568 00:54:12,067 --> 00:54:14,470 MORE LIKE PROPERTIES OF MATURE 1569 00:54:14,470 --> 00:54:16,438 INTERNEURONS, BUT NOT ONLY DO 1570 00:54:16,438 --> 00:54:19,875 YOU HAVE FEWER INTERNEURONS BUT 1571 00:54:19,875 --> 00:54:21,743 THEY'RE ALL ELECTROPHYSIOLOGICAL 1572 00:54:21,743 --> 00:54:23,145 PROPERTIES ARE COMPROMISED AND 1573 00:54:23,145 --> 00:54:25,881 MAKE MORE AN IMMATURE STATE 1574 00:54:25,881 --> 00:54:27,950 RATHER THAN A MATURE 1575 00:54:27,950 --> 00:54:28,250 INTERNEURON. 1576 00:54:28,250 --> 00:54:30,519 WE HAD A LOT OF BEHAVIOR ASSAYS 1577 00:54:30,519 --> 00:54:31,854 ON THIS, I WILL TOUCH ON A 1578 00:54:31,854 --> 00:54:32,421 COUPLE. 1579 00:54:32,421 --> 00:54:34,523 THIS IS I FIELD TEST AND YOU PUT 1580 00:54:34,523 --> 00:54:38,160 THE MICE IN THE BIG 4 BY 4 1581 00:54:38,160 --> 00:54:39,461 BASICALLY PLEXIGLOSS BOX AND AT 1582 00:54:39,461 --> 00:54:41,029 THE CAMERA YOU CAN MONITOR 1583 00:54:41,029 --> 00:54:42,731 MOVEMENT AND QUANTIFY IT, THIS 1584 00:54:42,731 --> 00:54:44,900 IS A WILD-TYPE MICE, THEY KIND 1585 00:54:44,900 --> 00:54:46,101 OF PERFORM THE MITSD BUT 1586 00:54:46,101 --> 00:54:48,737 THEY'RE HAPPY TO EXPLORE A 1587 00:54:48,737 --> 00:54:49,171 ROUND. 1588 00:54:49,171 --> 00:54:50,939 THIS HOMOZYGOUS MICE, YOU CAN 1589 00:54:50,939 --> 00:54:52,875 SEE THEY AVOID THE MITSD, THEY 1590 00:54:52,875 --> 00:54:54,409 YOU CAN SEE IT'S A SIGN OF 1591 00:54:54,409 --> 00:54:55,711 ANXIETY, THEY DON'T WANT TO GO 1592 00:54:55,711 --> 00:54:57,779 IN MIDDLE, THEY WANT TO HUG THE 1593 00:54:57,779 --> 00:54:59,748 EDGE WHERE THEY FEEL SAFER AND 1594 00:54:59,748 --> 00:55:00,816 MORE SECURE, NOT ONLY DO THEY 1595 00:55:00,816 --> 00:55:02,818 SPEND TIME IN THE CENTER, BUT IN 1596 00:55:02,818 --> 00:55:03,986 PARTICULAR THE FEMALES MOVE A 1597 00:55:03,986 --> 00:55:05,721 LOT MORE, THEY TRAVEL A GREATER 1598 00:55:05,721 --> 00:55:08,056 DISTANCE AND ALSO A LOT FASTER. 1599 00:55:08,056 --> 00:55:10,726 SO 1 WAY TO THINK OF THIS IS IN 1600 00:55:10,726 --> 00:55:13,362 PARTICULARLY THE FEMALE MICE ARE 1601 00:55:13,362 --> 00:55:14,730 ZOOMING AROUND THIS BOX WHICH IS 1602 00:55:14,730 --> 00:55:15,898 A SIGN OF HIGH ANXIETY. 1603 00:55:15,898 --> 00:55:17,432 ANOTHER WAY TO LOOK AT IT, TO 1604 00:55:17,432 --> 00:55:19,501 TEST ANXIETY IS TO PUT MICE IN 1605 00:55:19,501 --> 00:55:22,004 THIS LIGHT DARK BOX, WITH KIND 1606 00:55:22,004 --> 00:55:28,443 OF A 6 C DECIBEL MICE WILL 1607 00:55:28,443 --> 00:55:29,711 PREFER IT IN THE DARK IN 1608 00:55:29,711 --> 00:55:33,448 GENERAL, BUT IF YOU SEE THESE 1609 00:55:33,448 --> 00:55:34,283 PREFERENCES OF THE LIGHT 1610 00:55:34,283 --> 00:55:40,055 CONTROLLER BOX 1611 00:55:40,055 --> 00:55:42,791 COMPARED TO THE DARK BOX. 1612 00:55:42,791 --> 00:55:46,862 WE HAVE DATA ON SOCIAL 1613 00:55:46,862 --> 00:55:48,830 INTERACTION, SOUNDS, MEMORY 1614 00:55:48,830 --> 00:55:50,365 DEFICITS, AND SEE THE FEMALES 1615 00:55:50,365 --> 00:55:51,433 HAVE MORE DEFICITS THAN THE MALE 1616 00:55:51,433 --> 00:55:52,834 MICE AND I WON'T GO INTO TOO 1617 00:55:52,834 --> 00:55:54,002 MUCH MORE ABOUT THAT. 1618 00:55:54,002 --> 00:55:57,439 NOW, AGAIN WE WANTED TO 1619 00:55:57,439 --> 00:55:58,574 PERFORM MULTIOHM ANALYSIS IN 1620 00:55:58,574 --> 00:55:59,875 THIS CASE BECAUSE THE MUTATION 1621 00:55:59,875 --> 00:56:01,410 IS DOING THINGS WE THINK ARE 1622 00:56:01,410 --> 00:56:03,378 CRITICAL IN BOTH THE 1623 00:56:03,378 --> 00:56:06,014 INTERNEURONS AND THE HYPOTHAM 1624 00:56:06,014 --> 00:56:07,249 MUSAND WE HOUSTON AND RICE 1625 00:56:07,249 --> 00:56:09,284 FOOTBALLESTED EMBRYONIC MGE AND 1626 00:56:09,284 --> 00:56:10,986 HEM BRIANAONIC HYPOTHALAMUS AND 1627 00:56:10,986 --> 00:56:13,155 AS WELL ADULT HYPOTHALAMUS AND 1628 00:56:13,155 --> 00:56:14,223 ADULT INTERNEURONS IN THE CORTEX 1629 00:56:14,223 --> 00:56:16,658 AND THIS IS HOW THE MAP LOOKS. 1630 00:56:16,658 --> 00:56:18,260 WE CAN TAKE OUT THE DIFFERENT 1631 00:56:18,260 --> 00:56:21,296 SUBTYPES AND DO ALL KINDS OF 1632 00:56:21,296 --> 00:56:23,765 ANALYSIS AND EMBRYONIC 1S WE 1633 00:56:23,765 --> 00:56:26,301 SEE, INDICATORS OF ALL THESE 1634 00:56:26,301 --> 00:56:27,269 HYPOTHALAMIC NUCLEI THAT EXIST, 1635 00:56:27,269 --> 00:56:28,937 WE SEE THE 2 TYPES OF 1636 00:56:28,937 --> 00:56:30,439 INTERNEURONS AND THEN THE 1637 00:56:30,439 --> 00:56:31,707 HYPOTHALAMUS AND I WILL 1638 00:56:31,707 --> 00:56:32,674 HIGHLIGHT 2 THINGS WE'VE LEARNED 1639 00:56:32,674 --> 00:56:33,809 FROM THIS DATA THAT'S 1640 00:56:33,809 --> 00:56:36,411 INTERESTING, 1 IS THAT IF WE 1641 00:56:36,411 --> 00:56:38,247 FOCUSED ON P60 CORTEX SO THESE 1642 00:56:38,247 --> 00:56:39,781 ARE THE MATURE INTERNEURONS WE 1643 00:56:39,781 --> 00:56:42,217 CAN AGAIN LOOK AT DEFERENTIALLY 1644 00:56:42,217 --> 00:56:45,621 EXPRESSED GENES, WE CAN LOOK AT 1645 00:56:45,621 --> 00:56:46,288 THE DIFFERENT TERMINOLOGY 1646 00:56:46,288 --> 00:56:47,422 FACTORS THAT I HAVE BUT WHAT'S 1647 00:56:47,422 --> 00:56:51,159 INTERESTING TO US IS IF YOU TAKE 1648 00:56:51,159 --> 00:56:52,728 THESE MOST DIFFERENTLY EXPRESSED 1649 00:56:52,728 --> 00:56:54,830 GENES HERE AND MAP THEM ON TO 1650 00:56:54,830 --> 00:56:57,232 VARIOUS HUMAN DISEASES, WHAT YOU 1651 00:56:57,232 --> 00:57:00,736 SEE IS A LOT OF HITS ON 1652 00:57:00,736 --> 00:57:01,336 EPILEPSY, EPILECTIC SYNDROME, 1653 00:57:01,336 --> 00:57:02,037 THIS IS INTERESTING BECAUSE A 1654 00:57:02,037 --> 00:57:04,673 LOT OF THE YEENS THAT ARE MOST 1655 00:57:04,673 --> 00:57:06,742 DIFFERENTLY EXPRESSED IN THESE 1656 00:57:06,742 --> 00:57:09,478 MUTANT MICE ARE ASSOCIATED WITH 1657 00:57:09,478 --> 00:57:11,947 EPILEPSY WHICH WE SEE THESE MICE 1658 00:57:11,947 --> 00:57:13,282 HAVE, THAT'S 1 WAY WE CAN USE 1659 00:57:13,282 --> 00:57:15,284 THIS DATA TO KIND OF CORRELATE 1660 00:57:15,284 --> 00:57:16,952 CHANGES IN GENE EXPRESSION WITH 1661 00:57:16,952 --> 00:57:17,185 DISEASE. 1662 00:57:17,185 --> 00:57:19,821 IF WE LOOK IN THE ADULT 1663 00:57:19,821 --> 00:57:21,189 HYPOTHALAMUS AND WE BREAK IT 1664 00:57:21,189 --> 00:57:23,158 DOWN IN BETWEEN CELLS THAT COME 1665 00:57:23,158 --> 00:57:24,926 FROM THE WILD-TYPE AND 1666 00:57:24,926 --> 00:57:28,463 HOMOZYGOUS MICE SO WE IDENTIFY 1667 00:57:28,463 --> 00:57:30,165 NEURONS, OLIGIOS ASTRO SIGHTS 1668 00:57:30,165 --> 00:57:32,534 AND 10 SAITS AND YOU CAN SEE A 1669 00:57:32,534 --> 00:57:33,702 DIFFERENCE HERE IN THE ASTRO 1670 00:57:33,702 --> 00:57:36,338 SLIGHT CLUSTER WHERE THE MUTANT 1671 00:57:36,338 --> 00:57:37,739 AND WILD-TYPE MICE ARE CLUSTERED 1672 00:57:37,739 --> 00:57:40,909 DIFFERENTLY AND WE CAN AGAIN 1673 00:57:40,909 --> 00:57:41,610 EXTRACT SPECIFICALLY THESE 1674 00:57:41,610 --> 00:57:44,046 SUBSETS AND KIND OF LOOK AT THE 1675 00:57:44,046 --> 00:57:45,647 DIFREBTIAL GENE EXPRESSION AND 1676 00:57:45,647 --> 00:57:46,948 THAT THERE'S A WHOLE POPULATION 1677 00:57:46,948 --> 00:57:48,350 OF ASTRO SIGHTS THAT ARE 1678 00:57:48,350 --> 00:57:49,651 BASICALLY MISSING IN THE 1679 00:57:49,651 --> 00:57:52,721 HOMOZYGOUS MICE IN TERMS OF GENE 1680 00:57:52,721 --> 00:57:53,221 EXPRESSION AND FUNCTION. 1681 00:57:53,221 --> 00:57:53,722 ANOTHER THING THAT WAS 1682 00:57:53,722 --> 00:57:54,856 INTERESTING IS THAT IF YOU LOOK 1683 00:57:54,856 --> 00:57:57,426 AT THE PROPORTION OF CELLS, 1684 00:57:57,426 --> 00:57:58,994 AGAIN, ASTRO SIGHTS ARE BLUE 1685 00:57:58,994 --> 00:58:00,829 HERE, BASED ON THIS, IT LOOKS 1686 00:58:00,829 --> 00:58:02,664 LIKE THERE'S MORE ASTRO SIGHTS 1687 00:58:02,664 --> 00:58:04,299 IN THE MUTANT COMPARED TO 1688 00:58:04,299 --> 00:58:05,133 WILD-TYPE. 1689 00:58:05,133 --> 00:58:06,001 IS THAT TRUE? 1690 00:58:06,001 --> 00:58:06,735 IT IS. 1691 00:58:06,735 --> 00:58:08,570 WHEN WE UMPIRESUNE O STAINING 1692 00:58:08,570 --> 00:58:09,871 FOR THIS MARKER OF ASTRO SIGHTS 1693 00:58:09,871 --> 00:58:12,207 WE SEE A LOT MORE ASTRO SIGHTS 1694 00:58:12,207 --> 00:58:13,642 IN THE MUTANT COMPARED TO THE 1695 00:58:13,642 --> 00:58:14,509 WILD-TYPE, SO IT'S NOT SOMETHING 1696 00:58:14,509 --> 00:58:16,011 WE COULD HAVE PREDICTED, NOT 1697 00:58:16,011 --> 00:58:17,312 SOMETHING WE HAD AN IDEA ABOUT, 1698 00:58:17,312 --> 00:58:19,848 BUT IT LOOKS LIKE THIS MUTATION 1699 00:58:19,848 --> 00:58:20,916 CAUSES SIGNIFICANT INCREASE IN 1700 00:58:20,916 --> 00:58:22,851 ASTRO SIGHTS AND PABLLY A LOSS 1701 00:58:22,851 --> 00:58:29,257 OF A PARTICULAR SUBSET OF ASTRO 1702 00:58:29,257 --> 00:58:37,933 CITES IN THE WILD-TYPE. 1703 00:58:37,933 --> 00:58:40,635 NOW TAM, IS A REGION WHERE CURES 1704 00:58:40,635 --> 00:58:42,604 CAN ENTER THE BLOOD STREAM AND 1705 00:58:42,604 --> 00:58:43,472 HAVE CROSS TALK IN GENERAL IN 1706 00:58:43,472 --> 00:58:46,007 THE REST OF BRAIN, THERE'S 1707 00:58:46,007 --> 00:58:46,775 SIGNIFICANT RESTRICTION FOR 1708 00:58:46,775 --> 00:58:48,377 ANYTHING TO CROSS THE BLOOD 1709 00:58:48,377 --> 00:58:50,579 BRAIN BARRIER BUT THIS REGION IN 1710 00:58:50,579 --> 00:58:52,314 PARTICULAR, THESE TAN O SIGHTS 1711 00:58:52,314 --> 00:58:55,717 ARE CRITICAL WHERE THE BRAIN CAN 1712 00:58:55,717 --> 00:58:56,284 COMMUNICATE WITH THE BLOOD 1713 00:58:56,284 --> 00:58:58,920 STREAM AND THEY HAVE A VARIETY 1714 00:58:58,920 --> 00:59:00,222 OF FUNCTION IN AND ROLES AND 1715 00:59:00,222 --> 00:59:01,723 VARIETY OF FUNCTIONS WE THINK 1716 00:59:01,723 --> 00:59:03,892 ARE AFFECTED IN THESE MICE, SO 1717 00:59:03,892 --> 00:59:07,095 WE TOOK CROSS SECTIONS TO 1718 00:59:07,095 --> 00:59:09,664 AMENOSTAIN THESE REGIONS WITH 1719 00:59:09,664 --> 00:59:13,001 GFAP, SO THE ASTROCYTES MARKER 1720 00:59:13,001 --> 00:59:14,536 AND THESE ORGANIZATION IN TERMS 1721 00:59:14,536 --> 00:59:18,407 OF THE CELLS, IF YOU LOOK IN THE 1722 00:59:18,407 --> 00:59:20,442 MUTANT MOUSE WHICH YOU SEE A 1723 00:59:20,442 --> 00:59:22,010 STRIKING DISRUPTION OF THIS 1724 00:59:22,010 --> 00:59:23,378 ORGANIZE EGG, THERE'S WAY MORE 1725 00:59:23,378 --> 00:59:27,282 NK CELLS LIAISONS RUPTED MANNER 1726 00:59:27,282 --> 00:59:28,583 HERE, ASTROCYTES ARE ON THE 1727 00:59:28,583 --> 00:59:29,418 EDGE, SWITCHED AND NOT IN THE 1728 00:59:29,418 --> 00:59:31,820 MIDDLE AND YOU CAN SEE STRIKING 1729 00:59:31,820 --> 00:59:35,190 DIFFERENTS IN THESE TANYCYTES 1730 00:59:35,190 --> 00:59:35,524 MORPHOLOGY. 1731 00:59:35,524 --> 00:59:43,298 SO THERE'S SOMETHING GOING ON 1732 00:59:43,298 --> 00:59:45,167 THAT'S PERTURBING THESE CELLS, 1733 00:59:45,167 --> 00:59:46,234 BETWEEN THE BLOOD AND HORMONES 1734 00:59:46,234 --> 00:59:48,170 IN THE BRAIN AND THIS ALL FITS 1735 00:59:48,170 --> 00:59:50,872 INTO WHY THESE MICE POSSIBLY 1736 00:59:50,872 --> 00:59:52,941 HAVE OBESITY AND PERTURBATION OF 1737 00:59:52,941 --> 00:59:57,078 BEHAVIORIAL DEFICITS AND FEEDING 1738 00:59:57,078 --> 00:59:57,412 DEFICITS. 1739 00:59:57,412 --> 01:00:00,048 OKAY, SO, I WILL RUN THIS LAST 1740 01:00:00,048 --> 01:00:00,615 PART VERY QUICKLY. 1741 01:00:00,615 --> 01:00:03,151 SO THE LAST 2 THINGS AGAIN WE'RE 1742 01:00:03,151 --> 01:00:04,586 KIND OF TAKING A SLEDGE HAMMER 1743 01:00:04,586 --> 01:00:05,821 APPROACH SO NOW WE WANT TO LOOK 1744 01:00:05,821 --> 01:00:08,557 AT GENES THAT ARE REALLY MORE 1745 01:00:08,557 --> 01:00:09,624 AFFECTED, MORE CORRELATED WITH 1746 01:00:09,624 --> 01:00:13,161 HUMAN DEC AND 1 IS CALLED THIS 1747 01:00:13,161 --> 01:00:14,429 GENE ARC'S TRANSCRIPTION FACTOR. 1748 01:00:14,429 --> 01:00:16,198 THIS HAS BEEN WELL STUDIED IN 1749 01:00:16,198 --> 01:00:17,699 THE LITERATURE, ON THE X 1750 01:00:17,699 --> 01:00:19,634 ROAMSOME AND IS RESPONSIBLE FOR 1751 01:00:19,634 --> 01:00:23,438 ABOUT 10% OF X-LINKED 1752 01:00:23,438 --> 01:00:23,939 INTELLECTUAL DISABILITY 1753 01:00:23,939 --> 01:00:24,206 DISEASES. 1754 01:00:24,206 --> 01:00:26,208 YOU CAN SEE THERE'S A TON OF 1755 01:00:26,208 --> 01:00:28,276 POINT MUTATIONS IN THIS GENE, 1756 01:00:28,276 --> 01:00:29,678 AND MORE RECENTLY SINCE AT 1757 01:00:29,678 --> 01:00:31,646 X-LINKED IT WAS THOUGHT TO 1758 01:00:31,646 --> 01:00:33,915 PRIMARILY OR ONLY AFFECT MAILS 1759 01:00:33,915 --> 01:00:36,451 BUT THERE'S RECENT EVIDENCE NOW 1760 01:00:36,451 --> 01:00:38,753 THAT FEMALE CARRIERS, HETZ HAVE 1761 01:00:38,753 --> 01:00:39,921 MILD AND SEVERE PHENOTYPES. 1762 01:00:39,921 --> 01:00:41,122 WHAT'S INTERESTING FOR US IS 1763 01:00:41,122 --> 01:00:43,959 THIS GENE IS KNOWN TO BE 1764 01:00:43,959 --> 01:00:46,528 EXPRESSED IN THE GANGLIONIC 1765 01:00:46,528 --> 01:00:47,696 EMINENCES AND STUDY VS KNOCKED 1766 01:00:47,696 --> 01:00:49,865 OUT THIS YEEN IN POST MITOTIC 1767 01:00:49,865 --> 01:00:53,068 CELLS BUT THERE HASN'T BEEN MUCH 1768 01:00:53,068 --> 01:00:55,036 STUDIED WHEN YOU IMK IT OUT IN 1769 01:00:55,036 --> 01:00:56,204 CYCLING CELLS. 1770 01:00:56,204 --> 01:01:01,443 SO WE USE OUR NKX2.1 CRE TO SEE 1771 01:01:01,443 --> 01:01:05,380 HOW IT AFFECTS THE CELLS FATE 1772 01:01:05,380 --> 01:01:07,582 AND ALLOWS US TO LOOK AT HOW 1773 01:01:07,582 --> 01:01:09,317 ETICSERY ZYGOUS MOUSE AFFECTS A 1774 01:01:09,317 --> 01:01:10,785 FEMALE BECAUSE THAT HASN'T 1775 01:01:10,785 --> 01:01:11,219 REALLY BEEN TODAYED. 1776 01:01:11,219 --> 01:01:12,888 SO JUST TO RUN THROUGH THIS DATA 1777 01:01:12,888 --> 01:01:15,891 PRETTY QUICK, THE MALES ARE VERY 1778 01:01:15,891 --> 01:01:17,392 SICK, THEY'RE UNDERWEIGHT, THEY 1779 01:01:17,392 --> 01:01:20,061 ALL DIE BY 8 WEEKS. 1780 01:01:20,061 --> 01:01:21,997 WHERE PHENOTYPICALLY THE FEMALES 1781 01:01:21,997 --> 01:01:24,966 HESES ARE NORMAL SIZE, THE MALES 1782 01:01:24,966 --> 01:01:27,068 ARE KNOCKED OUT, THE FEMALES 1783 01:01:27,068 --> 01:01:31,973 AGAIN BECAUSE IT'S X-LINKED. 1784 01:01:31,973 --> 01:01:33,775 AND HERE'S VIDEOS, SO THE MALES 1785 01:01:33,775 --> 01:01:36,444 ARE VERY SICK, HERE'S INFANNITE 1786 01:01:36,444 --> 01:01:37,746 SEIZURES, SPASMS YOU SEE EVEN 1787 01:01:37,746 --> 01:01:40,282 WHEN THE PUPS ARE ABOUT 8 WEEKS 1788 01:01:40,282 --> 01:01:43,218 OLD, THIS SHOULD LOOK FAMILIAR 1789 01:01:43,218 --> 01:01:45,887 FROM THE K4 MMOUSE AND THESE 1790 01:01:45,887 --> 01:01:47,722 MICE ARE ALSO HYPEY ACTIVE. 1791 01:01:47,722 --> 01:01:50,325 WE PUT THEM THROUGH THE SAME 1792 01:01:50,325 --> 01:01:51,293 SEIZURE SCORE, AGAIN ONLY 1793 01:01:51,293 --> 01:01:52,928 FEMALES BECAUSE THE MAILS DIE 1794 01:01:52,928 --> 01:01:54,095 INJECTING THE DRUG CAUSES THEM 1795 01:01:54,095 --> 01:01:56,431 TO DIE, BUT WE SEE THE FEMALES 1796 01:01:56,431 --> 01:01:58,266 HAVE A HIGHER SUSCEPTIBILITY 1797 01:01:58,266 --> 01:01:59,568 SEIZURES AND AGAIN WE'RE 1798 01:01:59,568 --> 01:02:02,337 CURRENTLY PUTTING THEM THROUGH A 1799 01:02:02,337 --> 01:02:03,471 BATTERY OF ASSAYS. 1800 01:02:03,471 --> 01:02:05,774 WHAT YOU CAN SEE IS REALLY 1801 01:02:05,774 --> 01:02:07,208 STRIKING LOSS OF INTERNEURONS SO 1802 01:02:07,208 --> 01:02:09,311 THIS IS THE CORTEX HIPPOCAMPUS, 1803 01:02:09,311 --> 01:02:12,881 THIS IS THE MORMAL AMOUNT OF MG 1804 01:02:12,881 --> 01:02:15,016 DRIVING THE NEURONS. 1805 01:02:15,016 --> 01:02:17,319 THE FEMALES AGAIN HAVE DECREASE 1806 01:02:17,319 --> 01:02:19,387 AND THE MALE KNOCK OUTS HAVE A 1807 01:02:19,387 --> 01:02:20,555 STRIKING LOSS OF INTERNEURONS 1808 01:02:20,555 --> 01:02:22,190 AND THAT CAN BE SEEN HERE AND 1809 01:02:22,190 --> 01:02:23,258 AGAIN QUANTIFIED WHERE YOU HAVE 1810 01:02:23,258 --> 01:02:26,828 JUST A STRIKING LOSS, 80-90% 1811 01:02:26,828 --> 01:02:27,963 LOSS OF INTERNEURONS IN THIS 1812 01:02:27,963 --> 01:02:28,330 MOUSE. 1813 01:02:28,330 --> 01:02:30,398 THIS IS MORE SEVERE THAN THE 1814 01:02:30,398 --> 01:02:34,502 OTHER STUDIES THAT KNOCKED OUT 1815 01:02:34,502 --> 01:02:37,305 ARX POST MITOTICALLY, THEY 1816 01:02:37,305 --> 01:02:38,440 DIDN'T HAVE NEARLY THIS AMOUNT 1817 01:02:38,440 --> 01:02:40,308 OF CELL LOSS OF 1 COOL THING 1818 01:02:40,308 --> 01:02:41,509 WE'RE TRYING TO INDGREAT INTO 1819 01:02:41,509 --> 01:02:44,479 THE LAB, IS CELL 1820 01:02:44,479 --> 01:02:45,747 TRANSCRIPTOMICS, BUT WITH 1821 01:02:45,747 --> 01:02:46,548 SPATIAL TRANSKRPT ORDER OF 1822 01:02:46,548 --> 01:02:49,417 MICRONSICS YOU CAN LOOK AT 1823 01:02:49,417 --> 01:02:50,685 SPATIAL INFORMATION IN DIFFERENT 1824 01:02:50,685 --> 01:02:52,988 YALE GENE EXPRESSION AS WELL AS 1825 01:02:52,988 --> 01:02:54,155 LOOK AT NONCELL AUTONOMOUS 1826 01:02:54,155 --> 01:02:54,389 EFFECTS. 1827 01:02:54,389 --> 01:02:56,257 SO THIS IS JUST KIND OF INITIAL 1828 01:02:56,257 --> 01:02:58,059 DATA WE'VE HAD FOR A COUPLE 1829 01:02:58,059 --> 01:03:00,095 WEEKS, BEING LOAMACYING AT THE 1830 01:03:00,095 --> 01:03:01,396 WILD-TYPE IN THE HETEROZYGOUS 1831 01:03:01,396 --> 01:03:02,697 MOUSE WHERE WHAT YOU'RE DOING IS 1832 01:03:02,697 --> 01:03:05,000 RATHER THAN HAVING A SINGLE CELL 1833 01:03:05,000 --> 01:03:06,401 REAS LIEWGZ, LOOK AT A CELL 1834 01:03:06,401 --> 01:03:07,569 REGION AND LOOK AT GENE 1835 01:03:07,569 --> 01:03:09,404 EXPRESSION BOTH IN TERPS OF THE 1836 01:03:09,404 --> 01:03:12,741 SPATIAL MAP, AS WELL AS IN A 1837 01:03:12,741 --> 01:03:13,274 UMAP PLOT. 1838 01:03:13,274 --> 01:03:15,043 YOU CAN LOOK AT GENE EXPRESSION 1839 01:03:15,043 --> 01:03:15,410 PROFILE. 1840 01:03:15,410 --> 01:03:17,278 SO AGAIN, HERE'S THE WILD-TYPE 1841 01:03:17,278 --> 01:03:19,781 AND THEN IN THIS CASE, THE HAT 1842 01:03:19,781 --> 01:03:21,416 FEMALE, CAN YOU SEE THIS IN THE 1843 01:03:21,416 --> 01:03:23,718 MALE, BUT YOU CAN SEE STRONG 1844 01:03:23,718 --> 01:03:26,921 REGULATION OF P A LB-ALBUT MIN, 1845 01:03:26,921 --> 01:03:30,558 AND WE'RE TRYING TO SINGLE CELL 1846 01:03:30,558 --> 01:03:32,093 DATA AND TRANSCRIPTIONAL DAT TO 1847 01:03:32,093 --> 01:03:33,828 HAVE A COMREHENSIVE WAI TO LOOK 1848 01:03:33,828 --> 01:03:35,664 AT SPATIAL INFORMATION AND CELL 1849 01:03:35,664 --> 01:03:37,399 AUTONOMOUS EFFECTS AND CELL 1850 01:03:37,399 --> 01:03:38,333 NONAUTONOMOUS EFFECTS LIKE THIS. 1851 01:03:38,333 --> 01:03:40,669 OKAY, SO JUST TO SUMMARIZE, I'VE 1852 01:03:40,669 --> 01:03:43,505 SHOWED YOU HOW WE DEVELOPED A 1853 01:03:43,505 --> 01:03:45,373 EPIGENOME AT LIAISONS WITH A 1854 01:03:45,373 --> 01:03:46,141 MULTIMODAL APPROACH TO 1855 01:03:46,141 --> 01:03:48,510 CHARACTERIZE THE CELLS IN THE 1856 01:03:48,510 --> 01:03:50,011 EMBRYONIC BRAIN AS THEY'RE 1857 01:03:50,011 --> 01:03:51,246 MAKING CRITICAL DECISIONS IN 1858 01:03:51,246 --> 01:03:52,213 CELL FATE. 1859 01:03:52,213 --> 01:03:53,515 I'VE SHOWN 2 APPROACHES WE USE 1860 01:03:53,515 --> 01:03:56,051 TO TAKE A SLEDGE HAMMER TO YEEN 1861 01:03:56,051 --> 01:03:57,218 REPRESSION AND ACTIVATION AND 1862 01:03:57,218 --> 01:03:59,754 SHOW HOW THAT AFFECTS BOTH THE 1863 01:03:59,754 --> 01:04:01,723 FATE AND THE PERCENTAGES OF 1864 01:04:01,723 --> 01:04:03,258 CELLS AND CELL LOSS. 1865 01:04:03,258 --> 01:04:04,859 IN SPL CASES REALLY STRIKING 1866 01:04:04,859 --> 01:04:06,027 SEVERE MITRAL SITS IN ANIMAL 1867 01:04:06,027 --> 01:04:07,095 BEHAVIOR AND NOW WE'RE WELL 1868 01:04:07,095 --> 01:04:08,830 POSITIONED TO KIND OF LOOK MORE 1869 01:04:08,830 --> 01:04:10,999 SPECIFICALLY AT YEENS WE KNOW 1870 01:04:10,999 --> 01:04:12,400 ARE CRITICAL BOTH IN INTERNEURON 1871 01:04:12,400 --> 01:04:14,035 DEVELOPMENT AS WELL AS HUMAN 1872 01:04:14,035 --> 01:04:16,971 DISEASE, AND KIND OF TAKE THIS 1873 01:04:16,971 --> 01:04:18,940 PIPELINE TO REALLY HAMMER HOME 1874 01:04:18,940 --> 01:04:20,508 AND HOPEFULLY PROVIDE IMPORTANT 1875 01:04:20,508 --> 01:04:22,210 INSIGHTS INTO SOME OF THESE 1876 01:04:22,210 --> 01:04:22,477 DISEASES. 1877 01:04:22,477 --> 01:04:24,112 SO I HOPE YOU KIND OF CONVINCED 1878 01:04:24,112 --> 01:04:25,814 YOU HOW WE'RE TRYING TO USE A 1879 01:04:25,814 --> 01:04:27,315 VARIETY OF APPROACHES TO REALLY 1880 01:04:27,315 --> 01:04:30,852 UNDERSTAND HOW YOU GO FROM THESE 1881 01:04:30,852 --> 01:04:32,454 KIND OF HETEROGENEOUS POPULATION 1882 01:04:32,454 --> 01:04:34,322 IN THE EMBRYO TO THIS INCREDIBLE 1883 01:04:34,322 --> 01:04:35,890 DIVERSITY IN THE ADULT BRAIN, 1884 01:04:35,890 --> 01:04:37,892 HOW GENETIC FACTORS AND 1885 01:04:37,892 --> 01:04:39,127 EPIGENETIC AND GENE REGULATORY 1886 01:04:39,127 --> 01:04:41,362 FACTORS COMBINE WITH THE BRAIN 1887 01:04:41,362 --> 01:04:42,430 ENVIRONMENT TO INFLUENCE FATE 1888 01:04:42,430 --> 01:04:43,631 AND MATURATION OF THESE CELLS 1889 01:04:43,631 --> 01:04:45,834 BOTH IN NORMAL DEVELOPMENT AND 1890 01:04:45,834 --> 01:04:47,635 MOST CRITICALLY, WHAT GOES WRONG 1891 01:04:47,635 --> 01:04:50,438 IN DISEASE VARIOUS DISEASES. 1892 01:04:50,438 --> 01:04:51,773 SO I JUST WANT TO MAKE EVERYBODY 1893 01:04:51,773 --> 01:04:55,376 IN MY LAB IN PARTICULAR, 3 POST 1894 01:04:55,376 --> 01:04:56,811 DOCS HIGHLIGHTED HERE IN THE 1895 01:04:56,811 --> 01:04:58,313 VARIOUS PROJECTS I TALKED ABOUT. 1896 01:04:58,313 --> 01:05:01,716 A VARIETY OF POST BACKS, 1897 01:05:01,716 --> 01:05:04,252 IMPORTANT COLLABORATOR, 1898 01:05:04,252 --> 01:05:07,388 PARTICULARLY PEDRO ROCCA, THE 1899 01:05:07,388 --> 01:05:10,425 Mc BRAIN LAB FOR 1900 01:05:10,425 --> 01:05:13,128 [INDISCERNIBLE] LAB, AND RYAN 1901 01:05:13,128 --> 01:05:13,795 [INDISCERNIBLE] RUNS THE 1902 01:05:13,795 --> 01:05:15,163 BIO-INFORMATICS CORE THAT WE 1903 01:05:15,163 --> 01:05:16,531 COULDN'T DO A LOT WITHOUT. 1904 01:05:16,531 --> 01:05:16,831 THAT'S IT. 1905 01:05:16,831 --> 01:05:26,241 HAPPY TO TAKE QUESTIONS. 1906 01:05:26,241 --> 01:05:36,584 >> ANY QUESTIONS? 1907 01:05:41,222 --> 01:05:45,160 >> CAN I ASK A QUESTION ABOUT 1908 01:05:45,160 --> 01:05:51,766 YOUR MARK ON DISRUPTION OF H3 K4 1909 01:05:51,766 --> 01:05:52,133 TRI-METHYLATION? 1910 01:05:52,133 --> 01:05:55,270 DID YOU DO IT SPECIFICALLY FOR 1911 01:05:55,270 --> 01:05:58,773 THE NK2.1 LOCUST OR IT WAS DONE 1912 01:05:58,773 --> 01:06:04,546 ACROSS THE SPECTRUM OF ALL 1913 01:06:04,546 --> 01:06:04,746 GENES? 1914 01:06:04,746 --> 01:06:13,221 >> IT'S A FLOCKS ALLELE, SO WE 1915 01:06:13,221 --> 01:06:15,590 USE NKX2.1 CRE, SO THE MUTATED 1916 01:06:15,590 --> 01:06:16,858 ALLELE WAS ONLY EXPRESSED IN 1917 01:06:16,858 --> 01:06:18,426 THOSE REGIONS AND WE DID CONFIRM 1918 01:06:18,426 --> 01:06:20,295 THAT, THE MUTANT ALLELE IS 1919 01:06:20,295 --> 01:06:22,230 HA-TAG SO WE CONFIRMED BOTH 1920 01:06:22,230 --> 01:06:27,202 REDUCTION IN THE K4 METHYLATION 1921 01:06:27,202 --> 01:06:29,904 -- IN THE MG AND HYPOTHALAMUS 1922 01:06:29,904 --> 01:06:31,506 AND ALSO CONFIRMED UPREGULATION 1923 01:06:31,506 --> 01:06:33,474 OF HA SPECIFICALLY IN THOSE 1924 01:06:33,474 --> 01:06:33,741 REGIONS. 1925 01:06:33,741 --> 01:06:35,210 SO MY PREDECLARATION IS IF WE 1926 01:06:35,210 --> 01:06:36,177 DID THIS GLOBALLY AND THE PEOPLE 1927 01:06:36,177 --> 01:06:38,847 THAT MADE THE MOUSE DID IT, 1928 01:06:38,847 --> 01:06:41,249 THESE ARE DIFFERENT CRE DRIVER 1929 01:06:41,249 --> 01:06:43,218 BUT THE MICE DIED EARLY, SO I 1930 01:06:43,218 --> 01:06:45,620 THINK THE WILD-TYPE ALLELE IS 1931 01:06:45,620 --> 01:06:47,155 INTACT BUT I THINK UPREGULATION 1932 01:06:47,155 --> 01:06:49,424 OF THIS MUTANT ALLELE, ACROSS 1933 01:06:49,424 --> 01:06:52,160 THE BOARD, LIKE WITH A PAN CRE 1934 01:06:52,160 --> 01:06:54,128 DRIVER WOULD ALMOST CERTAINLY 1935 01:06:54,128 --> 01:06:58,433 RESULT IN ANIMAL DEATH WHICH THE 1936 01:06:58,433 --> 01:06:59,934 GROUP THAT DEVELOPED THIS MOUSE 1937 01:06:59,934 --> 01:07:04,706 LINE SAW USING A DIFFERENT CRE 1938 01:07:04,706 --> 01:07:05,073 DRIVER. 1939 01:07:05,073 --> 01:07:07,775 >> AND I ASSUME THAT BY DOING 1940 01:07:07,775 --> 01:07:09,510 THAT, YOU EFFECTIVELY DOWN 1941 01:07:09,510 --> 01:07:13,081 REGULATE THE ACTIVITY OF ALL THE 1942 01:07:13,081 --> 01:07:14,148 ENHANCERS OF NKX2.1 AND IF YOU 1943 01:07:14,148 --> 01:07:16,784 WOULD LIKE TO GO INTO THE 1944 01:07:16,784 --> 01:07:18,753 GRANULARITY OF THE REGULATORY 1945 01:07:18,753 --> 01:07:20,588 PROGRAM, YOU CAN PROBABLY 1946 01:07:20,588 --> 01:07:24,726 SOMEHOW GO 1 BY 1 AND SOMETHING 1947 01:07:24,726 --> 01:07:27,896 LIKE THAT AND SEE HOW YOU WERE 1948 01:07:27,896 --> 01:07:33,601 TALKING ABOUT THIS EVOLUTIONARY 1949 01:07:33,601 --> 01:07:34,736 WAYS OF STABILIZING GENE 1950 01:07:34,736 --> 01:07:36,738 EXPRESSION SO THAT PROBLEM IS 1951 01:07:36,738 --> 01:07:38,239 THE [INDISCERNIBLE] ENHANCERS 1952 01:07:38,239 --> 01:07:40,575 GENETIC ACTIVITY OF MULTIPLE 1953 01:07:40,575 --> 01:07:44,145 SHOULD KNOWN TO DECIPHER THAT 1954 01:07:44,145 --> 01:07:46,614 INSIDE OF NKX2.1 LOCUST, I'M 1955 01:07:46,614 --> 01:07:47,782 GUESSING LIKE CRSPR WOULD BE 1956 01:07:47,782 --> 01:07:49,751 NEEDED TO BREAK THE PIECES? 1957 01:07:49,751 --> 01:07:51,286 >> YEAH, THE LAST PIECE OF DATA 1958 01:07:51,286 --> 01:07:53,388 ON THAT STORY IS CUT AND TAG 1959 01:07:53,388 --> 01:07:54,989 WITH THE K4 TRI-METHYLATION, SO 1960 01:07:54,989 --> 01:07:57,525 THAT WE CAN LOOK AT DIFFERENCES 1961 01:07:57,525 --> 01:07:58,326 THROUGHOUT THE GENOME. 1962 01:07:58,326 --> 01:08:01,629 BECAUSE AGAIN, IF IT'S NKX2.1 1963 01:08:01,629 --> 01:08:03,865 WERE MORE RESIST ABT AS YOU WERE 1964 01:08:03,865 --> 01:08:05,967 HINTING AT, MAYBE THE 2.1 LOCUST 1965 01:08:05,967 --> 01:08:07,702 WILL SHOW GREATER RESISTANCE TO 1966 01:08:07,702 --> 01:08:09,237 THE PROTURBATION THAN OTHER LOCI 1967 01:08:09,237 --> 01:08:10,571 SO THAT'S THE LAST PIECE OF DATA 1968 01:08:10,571 --> 01:08:12,707 THAT CAN SPEAK TO THAT THAT 1969 01:08:12,707 --> 01:08:15,343 WE'RE KIND OF WAITING ON RIGHT 1970 01:08:15,343 --> 01:08:15,710 NOW. 1971 01:08:15,710 --> 01:08:18,579 BUT THERE ARE OTHER WAYS YOU CAN 1972 01:08:18,579 --> 01:08:20,348 MANIPULATE THAT WITH CRSPR, AND 1973 01:08:20,348 --> 01:08:21,849 OTHER WAYS TO LOOK AT THAT 1974 01:08:21,849 --> 01:08:22,317 EFFECT. 1975 01:08:22,317 --> 01:08:24,986 IT'S TRICKY, TOO, BECAUSE 50% OF 1976 01:08:24,986 --> 01:08:26,254 THESE HOMOZYGOUS MICE DIE, SO 1977 01:08:26,254 --> 01:08:27,422 WE'RE LOOKING AT THE HEALTHIEST 1978 01:08:27,422 --> 01:08:30,959 OF THE SICK MICE THAT SURVIVE 1979 01:08:30,959 --> 01:08:33,127 AND BECAUSE THEORETICALLY IT'S A 1980 01:08:33,127 --> 01:08:34,429 50/50 RATIO OF WHETHER THE 1981 01:08:34,429 --> 01:08:35,964 MUTANT ARK LEGAL IS THERE, THE 1982 01:08:35,964 --> 01:08:37,799 WILD-TYPE ALLELE IS THERE, IT 1983 01:08:37,799 --> 01:08:39,901 MIGHT NOT BE 50/50, WE HAVEN'T 1984 01:08:39,901 --> 01:08:41,970 -- IT'S VERY COMPLICATED TO LOOK 1985 01:08:41,970 --> 01:08:43,638 AT THAT, BUT IT'S A DIRTY MOUSE 1986 01:08:43,638 --> 01:08:46,007 IN THE SENSE THAT THERE'S GOING 1987 01:08:46,007 --> 01:08:47,542 TO BE VARIABILITY JUST 1988 01:08:47,542 --> 01:08:49,243 THEORETICALLY AND WHICH ALLELE 1989 01:08:49,243 --> 01:08:50,645 IS INTEGRATED INTO WHICH SITE 1990 01:08:50,645 --> 01:08:52,513 AND SOME SITES MIGHT HAVE THE 1991 01:08:52,513 --> 01:08:53,915 MUTANT ALLELE AND THOSE SITES 1992 01:08:53,915 --> 01:08:55,650 MIGHT BE MORE CRITICAL FOR 1993 01:08:55,650 --> 01:08:56,584 PARTICULAR GENE ACTIVATION AND 1994 01:08:56,584 --> 01:08:59,187 THUS GIVE A HIGHER PHENOTYPE. 1995 01:08:59,187 --> 01:09:01,122 SO, MOUSE MODELS REALLY NICE FOR 1996 01:09:01,122 --> 01:09:02,757 FOR THINGS BUT IT IS SORT OF 1997 01:09:02,757 --> 01:09:13,301 DIRTY IN A SENSE OF HAVING THAT 1998 01:09:16,904 --> 01:09:24,278 LEVEL OF GRANULARITY. 1999 01:09:24,278 --> 01:09:24,712 >> [INDISCERNIBLE] 2000 01:09:24,712 --> 01:09:25,913 >> WE DON'T HAVE ANYMORE 2001 01:09:25,913 --> 01:09:27,515 QUESTIONS ONLINE AND I WOULD 2002 01:09:27,515 --> 01:09:31,586 LIKE -- 1 MORE? 2003 01:09:31,586 --> 01:09:34,188 >> I WAS WONDERING, YOU KNOW I 2004 01:09:34,188 --> 01:09:37,825 MAY HAVE MISSED SOME OF YOUR 2005 01:09:37,825 --> 01:09:40,962 OBSERVATIONS BUT YOU SAID YOU DO 2006 01:09:40,962 --> 01:09:42,663 A KNOCK OUT IN THE EXPERIMENT IN 2007 01:09:42,663 --> 01:09:45,533 WHICH YOU DO THE KNOCK OUT OF 2008 01:09:45,533 --> 01:09:50,605 THE ENHANCER OF THE STCH2. 2009 01:09:50,605 --> 01:09:53,541 IT SEEMS LIKE AT THE END YOU GET 2010 01:09:53,541 --> 01:09:57,378 A DIFFERENT PROPORTION OF THE 3 2011 01:09:57,378 --> 01:10:01,682 TYPES OF 3 TYPES OF CELLS AND I 2012 01:10:01,682 --> 01:10:04,252 WAS WONDERING WHERE IS THIS 2013 01:10:04,252 --> 01:10:05,887 AFFECT COMING FROM? 2014 01:10:05,887 --> 01:10:09,290 IS IT COMING FROM A CHANGE IN 2015 01:10:09,290 --> 01:10:10,725 FATE OF YOUR CELLS? 2016 01:10:10,725 --> 01:10:15,096 IS THIS COMING FROM A DIFFERENT 2017 01:10:15,096 --> 01:10:18,266 CELL MIGRATION AND 2018 01:10:18,266 --> 01:10:19,067 DIFFERENTIATION? 2019 01:10:19,067 --> 01:10:21,569 IT COMING FROM CELL DIVISION, 2020 01:10:21,569 --> 01:10:23,037 THE CHANGE OR PROPORTION OF 2021 01:10:23,037 --> 01:10:23,671 THESE CELLS? 2022 01:10:23,671 --> 01:10:27,508 SORE IS THIS COMING FROM CELL 2023 01:10:27,508 --> 01:10:30,411 DEATH THAT AFFECTED, YOU KNOW, 2024 01:10:30,411 --> 01:10:31,112 THESE CELLS DIFFERENTIALLY SO 2025 01:10:31,112 --> 01:10:35,817 THAT YOU END UP WITH THE 2026 01:10:35,817 --> 01:10:36,951 DIFFERENTIAL -- 2027 01:10:36,951 --> 01:10:38,219 >> THAT'S AN EXCELLENT QUESTION 2028 01:10:38,219 --> 01:10:39,921 AND YOU HIT ON ALMOST ALL THE 2029 01:10:39,921 --> 01:10:40,588 POSSIBILITIES, I THINK. 2030 01:10:40,588 --> 01:10:41,789 SO WE HAVEN'T NAILED THAT DOWN. 2031 01:10:41,789 --> 01:10:43,658 I WILL SAY WE LOOK AT CELL 2032 01:10:43,658 --> 01:10:45,726 DEATH, WE DON'T SEE SIGNIFICANT 2033 01:10:45,726 --> 01:10:45,960 CHANGES. 2034 01:10:45,960 --> 01:10:48,262 ONE NUANCE AND DEVELOPMENT OF 2035 01:10:48,262 --> 01:10:49,697 THE PBS ST CELLS THAT I 2036 01:10:49,697 --> 01:10:51,732 MENTIONED IS THAT THESE CELLS 2037 01:10:51,732 --> 01:10:55,369 START TO BE BORN AROUND E11ISH 2038 01:10:55,369 --> 01:10:56,537 AND THEY'RE BASICALLY CONTINUED 2039 01:10:56,537 --> 01:10:58,940 TO BE BORN UNTIL ABOUT THE 2040 01:10:58,940 --> 01:11:01,476 ANIMAL'S -- UNTIL THE ANIMAL IS 2041 01:11:01,476 --> 01:11:02,610 BORN AT ABOUT E18. 2042 01:11:02,610 --> 01:11:05,513 SO YOU HAVE A RANGE OF 7 DAYS 2043 01:11:05,513 --> 01:11:07,482 THAT'S PRODUCING THESE 2044 01:11:07,482 --> 01:11:09,584 INTERNEURONS NOW THE PB AND SST 2045 01:11:09,584 --> 01:11:10,885 CELLS ARE NOT PRODUCED OVER THAT 2046 01:11:10,885 --> 01:11:12,120 TIME, SO IN THE BEGINNING 2047 01:11:12,120 --> 01:11:14,188 THERE'S A BURST OF SOPHISTICATED 2048 01:11:14,188 --> 01:11:15,590 MATOSTATIN CELLS AND THEN THE 2049 01:11:15,590 --> 01:11:17,225 PRODUCTION GOES DOWN OVER TIME 2050 01:11:17,225 --> 01:11:19,627 ON DURING EMBRYO GENESIS, 2051 01:11:19,627 --> 01:11:20,995 [INDISCERNIBLE] CELLS ARE MORE 2052 01:11:20,995 --> 01:11:21,262 CONSTANT. 2053 01:11:21,262 --> 01:11:24,599 SO IF YOU LOOK AT E12, E13,URE 2054 01:11:24,599 --> 01:11:27,201 MAKING A 50/50 MIX OF PB AND SST 2055 01:11:27,201 --> 01:11:29,871 CELLS BEING BORN, IF YOU LOOK AT 2056 01:11:29,871 --> 01:11:33,741 E16, IT'S AN 80/20 MIX OF 2057 01:11:33,741 --> 01:11:34,408 [INDISCERNIBLE] CELLS OVER 2058 01:11:34,408 --> 01:11:35,176 SOPHISTICATED MAT O CELLS. 2059 01:11:35,176 --> 01:11:37,145 SO IF YOU HAVE A DEVELOPMENT OF 2060 01:11:37,145 --> 01:11:38,212 MUTATION LIKE I THINK IS 2061 01:11:38,212 --> 01:11:39,647 HAPPENING IN THE ECH 2, WHERE 2062 01:11:39,647 --> 01:11:42,049 YOU ARE KICKING CELLS OUT OF THE 2063 01:11:42,049 --> 01:11:44,252 CELL CYCLE TOO EARLY, WHAT WILL 2064 01:11:44,252 --> 01:11:46,020 HAPPEN IF YOU'RE MAKING TOO MANY 2065 01:11:46,020 --> 01:11:47,221 TOO EARLY, THERE'S A BIAS FOR 2066 01:11:47,221 --> 01:11:48,723 THOSE TO BE SOPHISTICATED 2067 01:11:48,723 --> 01:11:50,691 MATOSTATIN CELLS AND IF YOU'RE 2068 01:11:50,691 --> 01:11:51,459 DEPLETING THE PROGENITOR POOL 2069 01:11:51,459 --> 01:11:54,529 LATER ON SO YOU HAVE FEWER CELLS 2070 01:11:54,529 --> 01:11:56,631 BEING BORN LATER ON WHERE PB 2071 01:11:56,631 --> 01:11:59,167 CELLS DOMINATE, YOU WILL HAVE 2072 01:11:59,167 --> 01:11:59,734 FEWER PB CELLS. 2073 01:11:59,734 --> 01:12:03,437 SO WE LOOK AT THAT A LITTLE BIT. 2074 01:12:03,437 --> 01:12:04,639 THE DATA WASN'T AS CONVINCING AS 2075 01:12:04,639 --> 01:12:06,807 I HOPED BUT MY BEST GUESS 2076 01:12:06,807 --> 01:12:08,709 BECAUSE A LITTLE BIT OF WHAT WE 2077 01:12:08,709 --> 01:12:10,478 KNOW ABOUT ECH2 AND 3 INNESS 2078 01:12:10,478 --> 01:12:11,779 KACCTER IS THAT TOO MANY CELLS 2079 01:12:11,779 --> 01:12:13,881 ARE BORN TOO EARLY, DEPLETING 2080 01:12:13,881 --> 01:12:15,049 THE PROIENITTOR POOL AND BECAUSE 2081 01:12:15,049 --> 01:12:16,751 OF THE TIME COURSE OF 2082 01:12:16,751 --> 01:12:20,655 SOPHISTICATED MAT O 7 PB-CELLS 2083 01:12:20,655 --> 01:12:22,356 ARISE, THAT EXPLANATION OF AND 2084 01:12:22,356 --> 01:12:23,991 OF ITSELF IS SUFFICIENT TO HAVE 2085 01:12:23,991 --> 01:12:25,560 A DECREASE OVERALL IN THE NUMBER 2086 01:12:25,560 --> 01:12:27,695 OF INTERNEURONS THAT WE SEE AND 2087 01:12:27,695 --> 01:12:29,197 ALSO HAVE THIS FATE SWITCH. 2088 01:12:29,197 --> 01:12:31,065 SO MAYBE, YOU KNOW FATE SWITCH 2089 01:12:31,065 --> 01:12:32,099 IS A DANGEROUS WORD. 2090 01:12:32,099 --> 01:12:35,303 I THINK WHAT'S MORE ACCURATE IS 2091 01:12:35,303 --> 01:12:36,604 THAT INCREASED PRODUCTION OF 2092 01:12:36,604 --> 01:12:38,906 INTERNEURONS EARLY AT A TIME 2093 01:12:38,906 --> 01:12:40,875 WHEN YOU'RE BIASED FOR 2094 01:12:40,875 --> 01:12:42,043 SOPHISTICATED MATOSTATIN CELLS 2095 01:12:42,043 --> 01:12:43,744 AND DECREASE PRODUCTION LATER AT 2096 01:12:43,744 --> 01:12:47,815 A TIME WHEN YOU'RE BIASED FOR PB 2097 01:12:47,815 --> 01:12:49,350 CELLS, THAT'S HOW YOU GET THE 2098 01:12:49,350 --> 01:12:50,184 FATE SWITCH APPEARANCE BUT I 2099 01:12:50,184 --> 01:12:52,787 DON'T THINK THE INDIVIDUAL CELLS 2100 01:12:52,787 --> 01:12:54,155 ARE SWITCHING FATE ALTHOUGH WE 2101 01:12:54,155 --> 01:12:55,156 DON'T HAVE DIRECT EVIDENCE AGAIN 2102 01:12:55,156 --> 01:12:56,958 THAT I THINK THIS OTHER SCENARIO 2103 01:12:56,958 --> 01:13:04,599 IS MUCH MORE LIKELY FOR THE 2104 01:13:04,599 --> 01:13:04,899 EXPLANATION. 2105 01:13:04,899 --> 01:13:08,302 >> [INDISCERNIBLE]. 2106 01:13:08,302 --> 01:13:09,237 2107 01:13:09,237 --> 01:13:09,704 2108 01:13:09,704 --> 01:13:09,971 >> OKAY. 2109 01:13:09,971 --> 01:13:10,771 >> [ APPLAUSE ] 2110 01:13:10,771 --> 01:13:21,048 >> THANK YOU.