1 00:00:06,383 --> 00:00:09,853 >> GOOD AFTERNOON AND WELCOME TO 2 00:00:09,853 --> 00:00:12,789 THE WEDNESDAY AFTERNOON LECTURE 3 00:00:12,789 --> 00:00:13,023 SERIES. 4 00:00:13,023 --> 00:00:16,359 I'M A LAB CHIEF IN NEUROGENETICS 5 00:00:16,359 --> 00:00:21,298 FROM THE NATIONAL INSTITUTE ON 6 00:00:21,298 --> 00:00:23,300 AGING AND HERE AND CO-CHAIR OF 7 00:00:23,300 --> 00:00:25,435 THE SINGLE CELL GENOMICS GROUP 8 00:00:25,435 --> 00:00:30,006 WHO NOMINATED TODAY'S SPEAKER, 9 00:00:30,006 --> 00:00:34,911 DR. JONATHAN WEISSMAN AND I'LL 10 00:00:34,911 --> 00:00:35,946 READ THESE BECAUSE I CAN'T 11 00:00:35,946 --> 00:00:37,581 REMEMBER THIS STUFF. 12 00:00:37,581 --> 00:00:39,850 IF YOU'RE WATCHING REMOTELY AND 13 00:00:39,850 --> 00:00:41,518 I BELIEVE THERE'S A COUPLE 14 00:00:41,518 --> 00:00:42,319 HUNDRED PEOPLE ONLINE WE 15 00:00:42,319 --> 00:00:43,920 ENCOURAGE YOU TO PARTICIPATE IN 16 00:00:43,920 --> 00:00:47,724 THE Q&A AND CLICK ON THE BUTTON 17 00:00:47,724 --> 00:00:50,293 BELOW YOUR VIDEOCAST THAT SAYS 18 00:00:50,293 --> 00:00:56,032 SEND LIVE SPEFEEDBACK AND WE'LL 19 00:00:56,032 --> 00:00:57,400 RELY IT TO HIM AND YOU CAN 20 00:00:57,400 --> 00:00:58,735 SUBMIT AT ANY TIME AND HOPE YOU 21 00:00:58,735 --> 00:00:59,336 DO SO. 22 00:00:59,336 --> 00:01:01,671 FOR THOSE HERE IN PERSON AND WE 23 00:01:01,671 --> 00:01:02,973 HAVE A GREAT CROWD, THANK YOU 24 00:01:02,973 --> 00:01:04,141 ALL FOR BEING HERE. 25 00:01:04,141 --> 00:01:05,876 THERE'LL BE A COUPLE MICROPHONES 26 00:01:05,876 --> 00:01:08,578 LEFT AND RIGHT TO ASK IN PERSON. 27 00:01:08,578 --> 00:01:13,316 THIS IS CRITICAL, WE'RE OFFERING 28 00:01:13,316 --> 00:01:15,752 CONTINUING MEDICAL EDUCATION 29 00:01:15,752 --> 00:01:19,689 CREDIT AND THE CME CODE IS 30 00:01:19,689 --> 00:01:20,090 50128. 31 00:01:20,090 --> 00:01:22,058 PUT THAT AND YOU'RE GET YOUR CME 32 00:01:22,058 --> 00:01:22,292 CREDIT. 33 00:01:22,292 --> 00:01:30,033 SO OUR SPEAKER TODAY, DR 34 00:01:30,033 --> 00:01:35,105 DR. WISEMAN IS A HEAD OF 35 00:01:35,105 --> 00:01:38,275 WHITEHEAD INSTITUTE AND STUDIES 36 00:01:38,275 --> 00:01:42,045 HOW CELLS ENSURE PROTEINS ARE 37 00:01:42,045 --> 00:01:44,080 FOLDED AND HOW IT IMPACTS 38 00:01:44,080 --> 00:01:44,915 DISEASE. 39 00:01:44,915 --> 00:01:48,051 IT'S FAIR TO SAY HE'S FAMOUS FOR 40 00:01:48,051 --> 00:01:50,287 A LOT OF TECHNIQUES DEVELOPED 41 00:01:50,287 --> 00:01:52,022 OVER THE YEARS INCLUDING 42 00:01:52,022 --> 00:01:54,791 RIBOSOME PROFILING AND GENOME 43 00:01:54,791 --> 00:01:56,593 SCALE HIGH CONTENT SCREENS, 44 00:01:56,593 --> 00:01:58,261 CRISPR INTERFERENCE AND 45 00:01:58,261 --> 00:02:00,063 ACTIVATION TO CONTROL EXPRESSION 46 00:02:00,063 --> 00:02:03,400 AND REWIRE THE EPI GENGENOME AN 47 00:02:03,400 --> 00:02:05,235 RECORDING THE HISTORY OF CELLS. 48 00:02:05,235 --> 00:02:07,370 VERY IMPRESSIVE TOOL SET AND 49 00:02:07,370 --> 00:02:07,938 THANK YOU. 50 00:02:07,938 --> 00:02:09,506 THESE ARE THINGS MANY US USE IN 51 00:02:09,506 --> 00:02:12,075 OUR LABS. 52 00:02:12,075 --> 00:02:15,178 HE RECEIVED AN AB IN PHYSICS 53 00:02:15,178 --> 00:02:17,581 FROM HARVARD AND PURSUED AN AN 54 00:02:17,581 --> 00:02:20,984 Ph.D. IN PHYSICS AT M.I.T. 55 00:02:20,984 --> 00:02:25,589 HE STUDIED PROTEIN FOLDING BY 56 00:02:25,589 --> 00:02:27,157 PANCREATIC INHIBITOR AND DID A 57 00:02:27,157 --> 00:02:29,759 POSTDOC AT YALE AND HELD 58 00:02:29,759 --> 00:02:32,596 POSITIONS IN DEPENDS OF CELL 59 00:02:32,596 --> 00:02:36,800 MOLECULAR PHARMACOLOGY AT 60 00:02:36,800 --> 00:02:38,268 UNIVERSITY SAN FRANCISCO AND 61 00:02:38,268 --> 00:02:40,904 CO-LEADS THE LABORATORY FOR 62 00:02:40,904 --> 00:02:42,939 GENOMICS RESEARCH TO DRIVE 63 00:02:42,939 --> 00:02:45,575 DEVELOPMENT OF CRISPR-BASED 64 00:02:45,575 --> 00:02:45,909 THERAPEUTICS. 65 00:02:45,909 --> 00:02:49,312 HE CO-FOUNDED THE INNOVATIVE 66 00:02:49,312 --> 00:02:54,117 GENOMICS INSTITUTE IN 2015 AND 67 00:02:54,117 --> 00:02:55,385 REMAINED ON THE ADVISORY BOARD. 68 00:02:55,385 --> 00:02:59,990 HE HAS RECEIVED NUMEROUS AWARDS 69 00:02:59,990 --> 00:03:02,125 TO CITE A FEW ELECTED TO THE 70 00:03:02,125 --> 00:03:02,692 NATIONAL ACADEMIES SCIENCES 71 00:03:02,692 --> 00:03:03,326 ENGINEERING MEDICINE AND WON THE 72 00:03:03,326 --> 00:03:05,896 AWARD FOR SCIENTIFIC DISCOVERY 73 00:03:05,896 --> 00:03:06,329 IN 2015. 74 00:03:06,329 --> 00:03:10,033 SO WITH ALL THAT SAID AND THAT 75 00:03:10,033 --> 00:03:11,434 IS VERY IMPRESSIVE AND THANK YOU 76 00:03:11,434 --> 00:03:14,771 FOR BEING HERE, THE TITLE OF HIS 77 00:03:14,771 --> 00:03:17,440 TALK IS EPIGENOME EDIT BEING AS 78 00:03:17,440 --> 00:03:21,511 A TOOL FOR BASIC DISCOVERY AND 79 00:03:21,511 --> 00:03:25,682 DELIVERY OF CHARM INTERVENTION. 80 00:03:25,682 --> 00:03:29,452 JOIN MIN WELCOMING DR. WEISSMAN. 81 00:03:29,452 --> 00:03:33,456 >> THANK YOU FOR THE KIND 82 00:03:33,456 --> 00:03:35,191 INTRODUCTION AND INVITATION TO 83 00:03:35,191 --> 00:03:35,859 BE HERE. 84 00:03:35,859 --> 00:03:38,395 IT'S BEEN A WONDERFUL DAY SO FAR 85 00:03:38,395 --> 00:03:40,430 AND HAVE VISITED NIH MANY TIMES 86 00:03:40,430 --> 00:03:41,865 AND IT'S WONDERFUL TO BE BACK. 87 00:03:41,865 --> 00:03:43,433 I WISH I HAD MORE TIME BECAUSE 88 00:03:43,433 --> 00:03:44,768 THERE'S SO MANY PEOPLE I DIDN'T 89 00:03:44,768 --> 00:03:46,970 HAVE A CHANCE TO VISIT BUT 90 00:03:46,970 --> 00:03:50,073 HOPEFULLY THERE'LL BE AN 91 00:03:50,073 --> 00:03:50,707 OPPORTUNITY SOME TIME IN THE NOT 92 00:03:50,707 --> 00:03:51,808 TOO DISTANT FUTURE. 93 00:03:51,808 --> 00:03:56,046 I'D LIKE TO TELL YOU TODAY ABOUT 94 00:03:56,046 --> 00:03:59,416 THE UNIFYING THEME OF OUR WORK 95 00:03:59,416 --> 00:04:01,251 IS A TOOL OF PROGRAMMABLE 96 00:04:01,251 --> 00:04:02,185 EPIGENETIC EDITORS. 97 00:04:02,185 --> 00:04:06,089 I'D LIKE TO TALK HOW WE USE THIS 98 00:04:06,089 --> 00:04:09,960 AS DISCOVERY TOOL AND PRESENT 99 00:04:09,960 --> 00:04:14,064 FLU WORK ABOUT THE POTENTIAL OF 100 00:04:14,064 --> 00:04:16,866 USING EPIGENETIC SILENCING AS A 101 00:04:16,866 --> 00:04:17,167 THERAPEUTIC. 102 00:04:17,167 --> 00:04:20,837 FIRST, THESE ARE MY DISCLOSURES. 103 00:04:20,837 --> 00:04:23,573 I THINK THE PRESENTATION WILL BE 104 00:04:23,573 --> 00:04:25,041 AVAILABLE FOR SOME TIME TO SEE 105 00:04:25,041 --> 00:04:28,411 THEM IN MORE DETAIL. 106 00:04:28,411 --> 00:04:33,984 SO MY INTRODUCTION TO CRISPR 107 00:04:33,984 --> 00:04:36,086 STARTED THROUGH THREE VERY 108 00:04:36,086 --> 00:04:39,656 FORTUITOUS ENCOUNTERS IN MY 109 00:04:39,656 --> 00:04:41,925 OFFICE IN 2012 AND HAD THE GOOD 110 00:04:41,925 --> 00:04:44,494 FORTUNE OF SITTING IN MY OFFICE 111 00:04:44,494 --> 00:04:45,128 AND SUCCESSIVELY THESE THREE 112 00:04:45,128 --> 00:04:47,831 PEOPLE CAME IN. 113 00:04:47,831 --> 00:04:51,201 THE FIRST WAS JENNIFER DOWDNA I 114 00:04:51,201 --> 00:04:54,070 HAD KNOWN MANY YEARS AND WITH 115 00:04:54,070 --> 00:04:55,572 GREAT EXCITEMENT LIKE OTHER 116 00:04:55,572 --> 00:04:58,208 PEOPLE READ HER PAPER SHOWING 117 00:04:58,208 --> 00:05:08,685 THAT KAS9 AND THE CHIMERIC 118 00:05:13,056 --> 00:05:15,291 POTENTIAL OF CRISPR AND SAID I 119 00:05:15,291 --> 00:05:16,893 HAVE THIS GREAT TOOL, WHAT ARE 120 00:05:16,893 --> 00:05:21,364 THINGS WE CAN DO WITH IT. 121 00:05:21,364 --> 00:05:24,200 THEN STANLEYY WHO WAS THEN A 122 00:05:24,200 --> 00:05:26,770 GRADUATE STUDENT JUST FINISHING 123 00:05:26,770 --> 00:05:30,974 AT BERKELEY AND COMING TO UCSF 124 00:05:30,974 --> 00:05:33,977 AS A FELLOW WAS TALKING ABOUT 125 00:05:33,977 --> 00:05:35,779 WHAT TO DO WITH NEXT STEP AND 126 00:05:35,779 --> 00:05:38,515 DESCRIBED REMARKABLE THING HE 127 00:05:38,515 --> 00:05:41,751 COULD TAKE A DEAD VERSION OF KAS 128 00:05:41,751 --> 00:05:43,119 9 AND EXPRESS SINCE A 129 00:05:43,119 --> 00:05:44,687 PROGRAMMABLE WAY IN BACTERIA AND 130 00:05:44,687 --> 00:05:48,324 USE IT TO BLOCK TRANSCRIPTION. 131 00:05:48,324 --> 00:05:51,394 THEN FINALLY LUKE GILBERT JUST 132 00:05:51,394 --> 00:05:58,301 JOINED MY LAB AS A POSTD-DOCTORL 133 00:05:58,301 --> 00:06:01,604 FELLOW AND HAD EXPERIENCE IN 134 00:06:01,604 --> 00:06:03,440 VECTOROLOGY AND MAMMALIAN STUDY 135 00:06:03,440 --> 00:06:06,109 AND WE HIT ON WHAT WE COULD DO 136 00:06:06,109 --> 00:06:09,412 WITH JENNIFER'S PROGRAMMABLE 137 00:06:09,412 --> 00:06:09,946 NUCLE 138 00:06:09,946 --> 00:06:10,213 NUCLEASE. 139 00:06:10,213 --> 00:06:13,416 WE WERE GOING TO BREAK IT SO IT 140 00:06:13,416 --> 00:06:14,317 COULDN'T CUT ANYTHING. 141 00:06:14,317 --> 00:06:15,351 THE REASON WE DID THIS IS 142 00:06:15,351 --> 00:06:18,121 BECAUSE NOW INSTEAD OF HAVING A 143 00:06:18,121 --> 00:06:22,459 PROGRAMMABLE NUCLEASE YOU HAD A 144 00:06:22,459 --> 00:06:25,228 READILY PROGRAMMABLE DNA BINDING 145 00:06:25,228 --> 00:06:25,528 PROTEIN. 146 00:06:25,528 --> 00:06:26,396 WE KNEW FROM SEMINOLE WORK THIS 147 00:06:26,396 --> 00:06:32,268 COULD BE USED AS A PLATFORM WHEN 148 00:06:32,268 --> 00:06:36,106 FUSED TO VECTOR DOMAINS AND THE 149 00:06:36,106 --> 00:06:38,808 FIRST WERE TRANSCRIPTIONAL 150 00:06:38,808 --> 00:06:41,578 REPRESSOR TO TURN OFF THE 151 00:06:41,578 --> 00:06:43,847 NEIGHBORING GENE OR 152 00:06:43,847 --> 00:06:49,185 TRANSCRIPTIONAL ACTIVATOR TO 153 00:06:49,185 --> 00:06:50,286 TURN ON. 154 00:06:50,286 --> 00:06:54,290 IN ESSENCE WE NOW HAD A DIAL TO 155 00:06:54,290 --> 00:06:57,160 TUNE ANY GENE EXPRESSION DOWN OR 156 00:06:57,160 --> 00:06:59,762 UP OF ANY GENE OR SET OF GENES 157 00:06:59,762 --> 00:07:01,431 IN THIS PROGRAMMABLE WAY. 158 00:07:01,431 --> 00:07:04,434 SINCE THIS TIME WE AND OTHER 159 00:07:04,434 --> 00:07:07,770 LABS HAVE PUT DIFFERENT EFFECTER 160 00:07:07,770 --> 00:07:08,004 DOMAINS. 161 00:07:08,004 --> 00:07:13,143 PERHAPS THE MOST SPECTACULAR OF 162 00:07:13,143 --> 00:07:17,514 WHICH ARE THE MANY REVERSE 163 00:07:17,514 --> 00:07:20,617 TRANSCRIPTASES TO GENERATE THE 164 00:07:20,617 --> 00:07:22,418 BASE EDITORS AND PRIME EDITOR 165 00:07:22,418 --> 00:07:32,095 STRATEGIES. 166 00:07:32,095 --> 00:07:34,297 THANKS TO THE THE ADVANCES YOU 167 00:07:34,297 --> 00:07:36,799 CAN MAKE GUIDE RNAs TO TARGET 168 00:07:36,799 --> 00:07:39,936 THE FULL SET OF GENES IN THE 169 00:07:39,936 --> 00:07:40,170 GENOME. 170 00:07:40,170 --> 00:07:43,039 NOW YOU CAN TRANSDUCE IN 171 00:07:43,039 --> 00:07:44,340 INDIVIDUAL CELLS THE LIBRARIES 172 00:07:44,340 --> 00:07:50,280 SO THAT EACH CELL GOT ONE OR A 173 00:07:50,280 --> 00:07:53,349 COMBINATION OF DIFFERENT RNA AND 174 00:07:53,349 --> 00:07:54,184 IT WOULD PROGRAM THE CHANGING 175 00:07:54,184 --> 00:07:56,786 EXPRESSION OF THE TARGETED GENE 176 00:07:56,786 --> 00:07:58,788 AND ACT AS A BAR CODE THAT TOLD 177 00:07:58,788 --> 00:08:00,657 YOU UP THROUGH SEQUENCING WHICH 178 00:08:00,657 --> 00:08:03,293 GENE HAD BEEN MODULATED. 179 00:08:03,293 --> 00:08:05,094 THIS COULD THEN BE DONE FOR 180 00:08:05,094 --> 00:08:10,900 EXAMPLE IN SCREENS TO ALLOW AND 181 00:08:10,900 --> 00:08:14,304 IDENTIFY GENES THAT CONFER 182 00:08:14,304 --> 00:08:18,141 RESISTANCE TO PRESSURE. 183 00:08:18,141 --> 00:08:21,377 FOR EXAMPLE ANANTI-CANCER DRUG 184 00:08:21,377 --> 00:08:23,780 YOU CAN LOOK FOR GENES WHEN YOU 185 00:08:23,780 --> 00:08:28,218 KNOCKED OUT CONFERRED RESISTANCE 186 00:08:28,218 --> 00:08:33,256 TO THAT THERAPEUTIC, RESISTANCE 187 00:08:33,256 --> 00:08:35,491 MECHANISMS OR ACT 188 00:08:35,491 --> 00:08:36,459 SYNERGISTICALLY AND LOOK AT THE 189 00:08:36,459 --> 00:08:40,129 NEXT DRUG TO ADD ON TO THAT. 190 00:08:40,129 --> 00:08:42,131 AND YOU MAY BE INVOLVED IN THE 191 00:08:42,131 --> 00:08:45,068 BIOLOGICAL PROFESSOR. 192 00:08:45,068 --> 00:08:48,137 WE WERE INTERESTED IN STRESS AND 193 00:08:48,137 --> 00:08:49,606 THEN BUILD A FLUORESCENCE 194 00:08:49,606 --> 00:08:51,741 REPORTER TO REPORT ON THE LEVEL 195 00:08:51,741 --> 00:08:56,079 OF STRESS OF AND THEN DO A FACT 196 00:08:56,079 --> 00:08:59,182 SCREEN FOR AND IDENTIFY ALL THE 197 00:08:59,182 --> 00:09:02,785 GENES WHEN YOU TURN ON OR OFF 198 00:09:02,785 --> 00:09:06,956 CAUSE DISRUPT FOLDING IN THE ER 199 00:09:06,956 --> 00:09:10,226 AND THIS COLLECTIVELY THESE AND 200 00:09:10,226 --> 00:09:12,061 PARALLEL EFFORTS WITH CRISPR 201 00:09:12,061 --> 00:09:14,163 CUTTING OPENED UP MAMMALIAN 202 00:09:14,163 --> 00:09:17,033 BOARD GENETICS IN A WAY THAT HAS 203 00:09:17,033 --> 00:09:19,435 PROVEN TO BE QUITE 204 00:09:19,435 --> 00:09:19,802 TRANSFORMATIVE. 205 00:09:19,802 --> 00:09:23,006 THERE WAS SHRNA APPROACH BEFORE 206 00:09:23,006 --> 00:09:24,607 AND THOSE WERE GROUNDBREAKING 207 00:09:24,607 --> 00:09:26,876 FOR THE DESIGN MUCH THESE. 208 00:09:26,876 --> 00:09:31,547 BUT THE SPECIFICITY AND EFFICACY 209 00:09:31,547 --> 00:09:38,588 OF THE TOOLS HAD NOW ROOM 210 00:09:38,588 --> 00:09:41,491 COMMODITIZED THE CRISPR SCREEN 211 00:09:41,491 --> 00:09:43,693 AND NOW IT'S FIGURE 1A THE 212 00:09:43,693 --> 00:09:46,562 STARTING POINT TO LAUNCH INTO 213 00:09:46,562 --> 00:09:56,706 BIOLOGY. 214 00:09:58,374 --> 00:10:00,610 YOU HAD TO DECIDE WHAT PART OF 215 00:10:00,610 --> 00:10:02,545 BIOLOGY YOU'RE INTERESTED IN AND 216 00:10:02,545 --> 00:10:05,348 IF I'M LOOKING AT THE MUTATED 217 00:10:05,348 --> 00:10:08,618 CANCER CELLS I THEN DO A CRISPR 218 00:10:08,618 --> 00:10:10,953 SCREEN FOR THINGS THAT MODULATE 219 00:10:10,953 --> 00:10:11,788 THE TOXICITY OR IF I'M 220 00:10:11,788 --> 00:10:15,091 INTERESTED AS I TALKED ABOUT IN 221 00:10:15,091 --> 00:10:17,660 THE PROTEIN FOLDING IN THE ER I 222 00:10:17,660 --> 00:10:21,464 BUILD A CELL LINE TO REPORT ON 223 00:10:21,464 --> 00:10:22,498 PROTEIN FOLDING AND HAD TO 224 00:10:22,498 --> 00:10:24,167 DEFINE AHEAD OF TIME WHAT THE 225 00:10:24,167 --> 00:10:27,470 BIOLOGY WAS AND THE BIGGER 226 00:10:27,470 --> 00:10:30,306 PROBLEM WAS AT THE END YOU WOULD 227 00:10:30,306 --> 00:10:33,343 GET A LIST OF SCREENS AND WHEN 228 00:10:33,343 --> 00:10:34,877 DONE WE WILL HAVE A HIGH 229 00:10:34,877 --> 00:10:37,146 CONFIDENCE LIST OF GENES AND 230 00:10:37,146 --> 00:10:41,551 DECONVOLUTING HOW THEY'RE WORK 231 00:10:41,551 --> 00:10:46,289 IS AN ENORMOUS CHALLENGE AND 232 00:10:46,289 --> 00:10:49,158 CELLS LIVE IN A HIGH DIMENSIONAL 233 00:10:49,158 --> 00:10:53,029 MANI FOLD OF TRANSCRIPTIONAL 234 00:10:53,029 --> 00:10:54,797 STATES AND WHEN YOU DO SINGLE 235 00:10:54,797 --> 00:10:56,933 READ OUT ONE DIMENSIONAL SCREEN 236 00:10:56,933 --> 00:10:59,235 IS DOING A DIMENSIONALITY 237 00:10:59,235 --> 00:11:00,036 REDUCTION PREDICTING THE TYPES 238 00:11:00,036 --> 00:11:02,505 ON THE SINGLE DIMENSION AND 239 00:11:02,505 --> 00:11:05,742 DOING THIS -- IT'S NOT A U MAP 240 00:11:05,742 --> 00:11:07,710 OR ANY UNDERSTOOD WAY. 241 00:11:07,710 --> 00:11:11,848 IT'S A HORRIBLY NON LINEAR AND 242 00:11:11,848 --> 00:11:12,548 IRREVERSIBLE APPROACH. 243 00:11:12,548 --> 00:11:14,817 TO GO BACK FROM WHERE YOU STAND 244 00:11:14,817 --> 00:11:16,819 HERE IN YOUR ONE DIMENSIONAL 245 00:11:16,819 --> 00:11:18,187 PHENOTYPE TO WHAT IT'S DOING 246 00:11:18,187 --> 00:11:20,323 UNDERSTANDING WHAT'S IT'S DOING 247 00:11:20,323 --> 00:11:22,759 TO THE CELL ENDS UP BEING OFTEN 248 00:11:22,759 --> 00:11:25,094 A Ph.D. THESIS. 249 00:11:25,094 --> 00:11:27,430 SO IT DIDN'T SCALE WITH THE 250 00:11:27,430 --> 00:11:28,097 NUMBER OF INTERESTING GENES THAT 251 00:11:28,097 --> 00:11:28,998 WERE COMING OUT OF THESE TYPES 252 00:11:28,998 --> 00:11:35,271 OF SCREENS. 253 00:11:35,271 --> 00:11:37,607 THIS MOTIVATE THE US AND OTHERS 254 00:11:37,607 --> 00:11:41,511 TO MAP THE EFFECTED PHENOTYPE IN 255 00:11:41,511 --> 00:11:44,614 A RICHER PHENOTYPIC SPACE AND 256 00:11:44,614 --> 00:11:49,419 APPROACH THAT WE AND WONDERFUL 257 00:11:49,419 --> 00:11:50,319 COLLABORATION AND MY LAB WITH 258 00:11:50,319 --> 00:11:54,157 THOSE WHO ARE NOW FACULTY 259 00:11:54,157 --> 00:11:57,093 MEMBERS AT PRINCETON AND 260 00:11:57,093 --> 00:11:58,294 MEMORIAL KETTERING RESPECTIVELY 261 00:11:58,294 --> 00:12:01,130 AND WONDERFUL COLLABORATION WITH 262 00:12:01,130 --> 00:12:10,306 A LAB TO DEVELOP THE PERTURBED 263 00:12:10,306 --> 00:12:13,142 SYC APPROACH WITH READOUT. 264 00:12:13,142 --> 00:12:15,178 AND THE FRAMEWORK OF THESE 265 00:12:15,178 --> 00:12:20,450 EXPERIMENTS WERE WE'D START WITH 266 00:12:20,450 --> 00:12:27,557 THE POOLED SCREEN AND NOW YOU 267 00:12:27,557 --> 00:12:33,162 WOULD DO A DROPLET BASED SINGLE 268 00:12:33,162 --> 00:12:35,465 CELL RNA SEQ AND THE GUIDE RNA 269 00:12:35,465 --> 00:12:37,133 PRESENT IN THE CELL WOULD TELL 270 00:12:37,133 --> 00:12:40,069 YOU WHAT THE PERTURBATION IN THE 271 00:12:40,069 --> 00:12:42,171 CELL WAS AND READ OUT THE 272 00:12:42,171 --> 00:12:44,474 TRANSCRIPTIONAL RESPONSE. 273 00:12:44,474 --> 00:12:46,008 IN ESSENCE THIS LET YOU MAP OUT 274 00:12:46,008 --> 00:12:47,376 FOR EACH OF THE PERTURBATIONS 275 00:12:47,376 --> 00:12:53,583 HOW YOU WERE PUSHING THE CELL IN 276 00:12:53,583 --> 00:12:56,752 THE TRANSCRIPTIONAL LANDSCAPE OR 277 00:12:56,752 --> 00:12:58,821 PHENOTYPIC LANDSCAPE OF THE 278 00:12:58,821 --> 00:12:59,021 CELL. 279 00:12:59,021 --> 00:13:00,857 AND STEPPING BACK IT'S PART OF A 280 00:13:00,857 --> 00:13:05,895 BROADER EFFORT WE AND OTHERS 281 00:13:05,895 --> 00:13:08,464 HAVE DONE TO TAKE SINGLE CELL 282 00:13:08,464 --> 00:13:09,799 RNA SEQ AND SEE THE STATE OF THE 283 00:13:09,799 --> 00:13:13,269 CELL AND MAKE IT ESSENTIALLY AN 284 00:13:13,269 --> 00:13:15,104 EXPERIMENTAL TOOL IN WHICH EACH 285 00:13:15,104 --> 00:13:16,572 CELL BECOMES A TEST TUBE AND YOU 286 00:13:16,572 --> 00:13:17,974 HAVE THE NATURE OF THE 287 00:13:17,974 --> 00:13:20,943 EXPERIMENT IN THIS CASE THE 288 00:13:20,943 --> 00:13:21,577 PERTURBATION THE GENE YOU'RE 289 00:13:21,577 --> 00:13:26,282 TURNING ON AND OFF AND THE 290 00:13:26,282 --> 00:13:30,052 RESULT READ OUT BY THE SINGLE 291 00:13:30,052 --> 00:13:32,388 CELL RNA SEC FROM HUNDREDS TO 292 00:13:32,388 --> 00:13:34,290 THOUSANDS AND NOW ROUTINELY A 293 00:13:34,290 --> 00:13:36,959 MILLION CELLS WE COULD DO THESE 294 00:13:36,959 --> 00:13:38,294 RICH EXPERIMENTS IN A MASSIVELY 295 00:13:38,294 --> 00:13:43,299 PARALLEL WAY. 296 00:13:43,299 --> 00:13:46,702 SO WHEN WE STARTED THE SCALE WAS 297 00:13:46,702 --> 00:13:50,606 A FEW HUNDRED GENES AND THE 298 00:13:50,606 --> 00:13:54,143 ORIGINAL VISION OF WHAT WE DO IS 299 00:13:54,143 --> 00:13:56,979 DO THE POOLED SCREEN AND GET 300 00:13:56,979 --> 00:13:59,749 FAVORITE HITS AND PUT IT INTO A 301 00:13:59,749 --> 00:14:01,083 PERTURBED SEQ TO UNDERSTAND THE 302 00:14:01,083 --> 00:14:01,817 FUNCTIONAL RELATIONSHIP BETWEEN 303 00:14:01,817 --> 00:14:02,752 THE TWO. 304 00:14:02,752 --> 00:14:03,619 THAT DIDN'T SOLVE THE FIRST 305 00:14:03,619 --> 00:14:06,155 PROBLEM I TALKED ABOUT WHICH IS 306 00:14:06,155 --> 00:14:07,990 YOU HAD TO STILL PREDEFINE WHAT 307 00:14:07,990 --> 00:14:10,660 THE FIRST POOLED SCREEN WAS. 308 00:14:10,660 --> 00:14:14,897 THAT MOTIVATED US AND US BEING 309 00:14:14,897 --> 00:14:19,569 BRIT ADAMSON AND A TERRIFIC 310 00:14:19,569 --> 00:14:24,140 STUDENT NOW HAS GONE ON TO DO 311 00:14:24,140 --> 00:14:26,909 MEDICAL TRAINING M.D. Ph.D. 312 00:14:26,909 --> 00:14:30,680 STUDENT TO ALLOW THEM TO BE DONE 313 00:14:30,680 --> 00:14:34,450 ACROSS GENOME PERTURBATIONS. 314 00:14:34,450 --> 00:14:37,486 SO IN THE FIRST EXPERIMENT HE 315 00:14:37,486 --> 00:14:39,622 DID ALL THE EXPRESSED GENES IN 316 00:14:39,622 --> 00:14:44,760 ONE CANCER CELL LINE K562 AND 317 00:14:44,760 --> 00:14:46,796 ACROSS FOUR CELL LINES INCLUDING 318 00:14:46,796 --> 00:14:50,833 BOTH CANCER AND NON-TRANSFORMED 319 00:14:50,833 --> 00:14:52,935 IMMORTALIZED CELL LINES. 320 00:14:52,935 --> 00:14:55,838 THIS IS ABOUT 6 MILLION CELL 321 00:14:55,838 --> 00:14:56,138 EXPERIMENT. 322 00:14:56,138 --> 00:14:58,307 AT THE TIME NOW A FEW YEARS AGO, 323 00:14:58,307 --> 00:15:01,577 WAS FAIRLY HEROIC BUT AGAIN 324 00:15:01,577 --> 00:15:02,878 BECAUSE TECHNOLOGY IS ADVANCING 325 00:15:02,878 --> 00:15:05,648 SO RAPIDLY THIS WOULD BE STILL A 326 00:15:05,648 --> 00:15:09,785 LARGE SCALE EXPERIMENT BUT 327 00:15:09,785 --> 00:15:12,154 NOWHERE NEAR AS HEROIC AS IT WAS 328 00:15:12,154 --> 00:15:13,389 JUST A COUPLE YEARS AGO. 329 00:15:13,389 --> 00:15:18,160 WHAT YOU GOT OUT OF THIS WAS A 330 00:15:18,160 --> 00:15:23,799 LIST OF THE GENETIC 331 00:15:23,799 --> 00:15:26,135 PERTURBATIONS, KNOCKING DOWN 332 00:15:26,135 --> 00:15:28,304 IRE1 OR PROTEIN RESPONSE SENSOR 333 00:15:28,304 --> 00:15:29,839 AND THE TRANSCRIPTIONAL RESPONSE 334 00:15:29,839 --> 00:15:31,707 FOR EACH OF THESE PERTURBATIONS. 335 00:15:31,707 --> 00:15:34,710 WHAT YOU COULD DO THEN IS EITHER 336 00:15:34,710 --> 00:15:37,246 CLUSTER ON THE GENES THE 337 00:15:37,246 --> 00:15:40,983 TRANSCRIPTIONAL RESPONSES AND IN 338 00:15:40,983 --> 00:15:44,620 THIS WAY IN A SYSTEMATIC MANNER 339 00:15:44,620 --> 00:15:46,288 LOOK AT THE TRANSCRIPTIONAL 340 00:15:46,288 --> 00:15:47,790 RESPONSES IN THE CELLS. 341 00:15:47,790 --> 00:15:50,159 FOR EXAMPLE, INTEGRATE STRESS 342 00:15:50,159 --> 00:15:51,594 RESPONSE AND THE PROTEIN 343 00:15:51,594 --> 00:15:54,163 RESPONSE AND FALLOUT IN AN 344 00:15:54,163 --> 00:15:57,033 AUTOMATIC WAY AND DO OTHER 345 00:15:57,033 --> 00:16:00,169 STRESS RESPONSE AND OTHER 346 00:16:00,169 --> 00:16:01,103 TRANSCRIPTIONAL RESPONSES THAT 347 00:16:01,103 --> 00:16:02,171 HAVEN'T BEEN AS WELL STUDIED OR 348 00:16:02,171 --> 00:16:06,142 HAVE A NAME ON THEM. 349 00:16:06,142 --> 00:16:13,215 THE SECOND APPROACH IS TO ORDER 350 00:16:13,215 --> 00:16:15,484 THE GENETIC PERTURBATIONS AND 351 00:16:15,484 --> 00:16:17,319 ORDER GENES BASED ON THE 352 00:16:17,319 --> 00:16:18,187 SIMILARITY OF HOW THE CELL 353 00:16:18,187 --> 00:16:19,855 RESPONDS TO THE LOSS OF THAT 354 00:16:19,855 --> 00:16:21,090 GENE. 355 00:16:21,090 --> 00:16:24,193 SO GENES THAT ARE HAVING SIMILAR 356 00:16:24,193 --> 00:16:25,961 FUNCTIONS WILL THEN CLUSTER NEXT 357 00:16:25,961 --> 00:16:30,633 TO EACH OTHER. 358 00:16:30,633 --> 00:16:35,871 THIS IS REALLY POWERFUL APPROACH 359 00:16:35,871 --> 00:16:38,340 IN AN OBJECTIVE SYSTEMATIC WAY 360 00:16:38,340 --> 00:16:41,444 OF ORDERING THE GENES AND GROUPS 361 00:16:41,444 --> 00:16:42,712 AND DIDN'T PRESUPPOSE YOU HAVE 362 00:16:42,712 --> 00:16:45,381 TO DEFINE WHAT THE GROUPS ARE. 363 00:16:45,381 --> 00:16:50,286 IT ALLOWS A FORM OF UNBIASSED 364 00:16:50,286 --> 00:16:52,421 HYPOTHESIS GENERATING 365 00:16:52,421 --> 00:16:54,190 EXPERIMENTS ON A DIFFERENT SCALE 366 00:16:54,190 --> 00:16:56,525 THAN PREVIOUSLY POSSIBLE. 367 00:16:56,525 --> 00:17:00,196 LET'S JUST GIVE YOU A BIT OF 368 00:17:00,196 --> 00:17:02,832 EXAMPLES OF THIS. 369 00:17:02,832 --> 00:17:06,102 HERE'S JUST CLUSTERING 370 00:17:06,102 --> 00:17:08,270 PERTURBATION CORRELATION AND 371 00:17:08,270 --> 00:17:10,873 MATRIX WITH EVERY PERTURBATION 372 00:17:10,873 --> 00:17:12,508 ORDERED WITH ONES MOST SIMILAR 373 00:17:12,508 --> 00:17:12,808 TO IT. 374 00:17:12,808 --> 00:17:16,045 THEN BLOWUP AND YOU CAN SEE HOW 375 00:17:16,045 --> 00:17:17,146 MUCH BIND STRUCTURE THERE IS 376 00:17:17,146 --> 00:17:18,481 HERE AND WHEN YOU GET TO THE 377 00:17:18,481 --> 00:17:21,717 POINT YOU SEE GENE NAMES YOU CAN 378 00:17:21,717 --> 00:17:24,987 SEE THIS IS ABLE TO PUT TOGETHER 379 00:17:24,987 --> 00:17:29,125 A WELL STUDIED PROCESSES AND 380 00:17:29,125 --> 00:17:31,494 METALATION AND ELONGATER AND 381 00:17:31,494 --> 00:17:33,462 INTEGRATOR COMPLEX AND A TINY 382 00:17:33,462 --> 00:17:36,232 CORNER OF THIS ENTIRE MAP. 383 00:17:36,232 --> 00:17:39,235 SO BASICALLY WHAT WE'RE ABLE TO 384 00:17:39,235 --> 00:17:40,936 DO IS RECAPITULATE THOUSANDS OF 385 00:17:40,936 --> 00:17:43,572 KNOWN RELATIONSHIPS AND BECAUSE 386 00:17:43,572 --> 00:17:45,908 AS YOU'LL SEE IN A VERY SPECIFIC 387 00:17:45,908 --> 00:17:47,276 EXAMPLE BECAUSE THERE'S NEWER OR 388 00:17:47,276 --> 00:17:49,478 POORLY CHARACTERIZED GENES THAT 389 00:17:49,478 --> 00:17:51,046 OFTEN CLUSTER WITHIN THESE KNOWN 390 00:17:51,046 --> 00:17:53,783 FUNCTIONAL CLUSTERS IT ALLOWS 391 00:17:53,783 --> 00:17:55,684 YOU TO DISCOVER OR GENERATE 392 00:17:55,684 --> 00:17:58,487 HYPOTHESES ABOUT NEW ELEMENTS. 393 00:17:58,487 --> 00:18:01,457 IN ADDITION TO THIS GROUPING 394 00:18:01,457 --> 00:18:03,425 GENES INTO FUNCTION THOUGH IT 395 00:18:03,425 --> 00:18:08,097 CAN BE USED TO DISSECT A LARGE 396 00:18:08,097 --> 00:18:08,731 MULTI-FUNCTIONAL COMPLEX INTO 397 00:18:08,731 --> 00:18:11,333 FUNCTIONAL MODULES. 398 00:18:11,333 --> 00:18:13,369 I'LL SPEND A COUPLE SLIDES 399 00:18:13,369 --> 00:18:15,004 SHOWING A NICE EXAMPLE CENTERED 400 00:18:15,004 --> 00:18:20,743 AROUND THIS INTEGRATOR COMPLEX. 401 00:18:20,743 --> 00:18:24,246 THE INTEGRATOR COMPLEX THIS IS A 402 00:18:24,246 --> 00:18:25,548 PIECE. 403 00:18:25,548 --> 00:18:30,252 IT'S A METAZONE TRANSCRIPTIONAL 404 00:18:30,252 --> 00:18:32,121 COMPLEX STUDIED OVER THE LAST 405 00:18:32,121 --> 00:18:33,956 DECADE OR SO THE SUBJECT OF 406 00:18:33,956 --> 00:18:34,657 INTENSE STRUCTURE AND 407 00:18:34,657 --> 00:18:37,560 MECHANISTIC STUDIES. 408 00:18:37,560 --> 00:18:39,261 WHAT WE FOUND IS THAT EVEN 409 00:18:39,261 --> 00:18:43,499 THOUGH THIS IS A SINGLE THEN 410 00:18:43,499 --> 00:18:45,668 PROTEIN AND REALLY A 15 PROTEIN 411 00:18:45,668 --> 00:18:47,503 COMPLEX, THE FUNCTIONAL MAP 412 00:18:47,503 --> 00:18:50,539 DISSECTED THIS INTO AT LEAST 413 00:18:50,539 --> 00:18:53,609 THREE DISTINCT FUNCTIONAL 414 00:18:53,609 --> 00:18:53,843 MODULES. 415 00:18:53,843 --> 00:18:55,945 AND WE HAD A STRUCTURE OF MOST 416 00:18:55,945 --> 00:18:58,280 THE COMPONENTS OF THE INTEGRATOR 417 00:18:58,280 --> 00:19:03,752 AND THE FUNCTIONAL MODULES AS 418 00:19:03,752 --> 00:19:13,829 DEFINED BY OUR PERTURB SEQ WENT 419 00:19:13,829 --> 00:19:16,131 TO THE COMPLEX AND THIS IS PART 420 00:19:16,131 --> 00:19:19,068 OF THE BACKBONE AND SHOULDER AND 421 00:19:19,068 --> 00:19:22,271 THE ENDONUCLEASE SUB COMPLEX 422 00:19:22,271 --> 00:19:25,307 RESPONSIBLE FOR CLEAVING NASCENT 423 00:19:25,307 --> 00:19:25,774 TRANSCRIPTS. 424 00:19:25,774 --> 00:19:27,910 WE FOUND THREE SUB UNITS AND ONE 425 00:19:27,910 --> 00:19:30,246 THEN COMPLETELY UNCHARACTERIZED 426 00:19:30,246 --> 00:19:33,415 PROTEIN AS A DISTINCT FUNCTIONAL 427 00:19:33,415 --> 00:19:33,649 MODULE. 428 00:19:33,649 --> 00:19:36,218 AND SO WE THEN AS YOU'LL SEE 429 00:19:36,218 --> 00:19:46,929 WHY, WE RENAMED THIS UNNAME D AD 430 00:19:52,268 --> 00:19:57,072 RUBIN SAUNDERS SHOWED THROUGH A 431 00:19:57,072 --> 00:20:02,912 PULL DOWN THIS FORMED THE 432 00:20:02,912 --> 00:20:07,116 SUBCOMPLEX THE SUB UNITS. 433 00:20:07,116 --> 00:20:09,919 IT ALSO SHOWED THE SUB COMPLEX 434 00:20:09,919 --> 00:20:14,256 WAS NOT REQUIRED FOR THE 435 00:20:14,256 --> 00:20:14,723 ENDNUCLEASE ACTIVITY. 436 00:20:14,723 --> 00:20:17,760 AND THIS LED RUBIN TO 437 00:20:17,760 --> 00:20:18,694 HYPOTHESIZE THE REASON THE 438 00:20:18,694 --> 00:20:24,266 SUBCOMPLEX WAS MISSING FROM THE 439 00:20:24,266 --> 00:20:26,302 OVER ALL STRUCTURE WAS BECAUSE 440 00:20:26,302 --> 00:20:30,239 THEY WERE MISSING THE SUB UNIT. 441 00:20:30,239 --> 00:20:32,808 AND GRATIFYINGLY A MONTH OR SO 442 00:20:32,808 --> 00:20:37,179 AGO PATRICK KRAMER PUBLISHED THE 443 00:20:37,179 --> 00:20:39,381 FULL INTEGRATOR COMPLEX 444 00:20:39,381 --> 00:20:44,687 EXPLICITLY PURIFYING AND 445 00:20:44,687 --> 00:20:49,825 GRACEFULLY NOTING RUBIN'S 446 00:20:49,825 --> 00:20:53,595 OBSERVATIONS AND ADDED IT TO THE 447 00:20:53,595 --> 00:20:55,698 REST AND FOUND IT FORMED WHAT IS 448 00:20:55,698 --> 00:21:00,235 CALLED THE TAIL DOMAIN AND IT 449 00:21:00,235 --> 00:21:03,472 RESEMBLED A SCORPION TAIL AND 450 00:21:03,472 --> 00:21:07,910 ALSO SHOWED THE SUB UNIT THAT 451 00:21:07,910 --> 00:21:09,411 WITH US MISSING FROM OTHER 452 00:21:09,411 --> 00:21:10,579 COMPLEXES WAS CRITICAL FOR 453 00:21:10,579 --> 00:21:13,615 STAPLING THE TAIL ON TO THE REST 454 00:21:13,615 --> 00:21:19,054 OF THE COMPLEX AND THEN PATRICK 455 00:21:19,054 --> 00:21:20,756 SHOWED THIS TAIL COMPLEX WAS NOT 456 00:21:20,756 --> 00:21:23,592 REQUIRED FOR THE ENDONUCLEUS 457 00:21:23,592 --> 00:21:26,028 ACTIVITY BUT FOR REMOVAL TO 458 00:21:26,028 --> 00:21:29,231 AFTER THE CLEAVAGE EVENT. 459 00:21:29,231 --> 00:21:30,966 I GO THROUGH THIS EXAMPLE AND A 460 00:21:30,966 --> 00:21:33,402 LITTLE BIT OF DETAIL BECAUSE IT 461 00:21:33,402 --> 00:21:37,272 SHOWS HOW FROM A GENERIC 462 00:21:37,272 --> 00:21:39,975 EXPERIMENT TO BE HONEST WE 463 00:21:39,975 --> 00:21:41,310 BARELY KNEW WHAT THE INTEGRATOR 464 00:21:41,310 --> 00:21:43,645 WAS WHEN WE CONDUCTED THE 465 00:21:43,645 --> 00:21:47,516 STUDIES WE WERE ABLE IN A FAIRLY 466 00:21:47,516 --> 00:21:49,318 SYSTEMATIC AND SCALABLE WAY GET 467 00:21:49,318 --> 00:21:52,488 TO A SPECIFIC ENOUGH HYPOTHESIS 468 00:21:52,488 --> 00:21:55,591 THAT IT COULD HAND OFF DIRECTLY 469 00:21:55,591 --> 00:21:58,293 AND BE OF DIRECT UTILITY TO 470 00:21:58,293 --> 00:22:00,229 PEOPLE WITH MORE FOCUSSED 471 00:22:00,229 --> 00:22:07,403 INTEREST IN THIS APPROACH. 472 00:22:07,403 --> 00:22:11,907 I'VE TOLD YOU FROM THE GENOTYPE, 473 00:22:11,907 --> 00:22:14,376 PHENOTYPE MAPS YOU CAN DISCOVER 474 00:22:14,376 --> 00:22:16,412 TRANSCRIPTIONAL PROGRAMS AND 475 00:22:16,412 --> 00:22:17,046 FUNCTIONALLY CHARACTERIZE AND 476 00:22:17,046 --> 00:22:21,450 DISSECT PROTEIN COMPLEXES. 477 00:22:21,450 --> 00:22:23,719 THE FINAL AND I FEEL IN MANY 478 00:22:23,719 --> 00:22:26,055 WAYS THE MOST EXCITING ASPECT OF 479 00:22:26,055 --> 00:22:27,456 THE POTENTIAL FOR THE ANALYSIS 480 00:22:27,456 --> 00:22:31,627 IS THAT YOU CAN THEN TAKE THESE 481 00:22:31,627 --> 00:22:34,897 GENOTYPE/PHENOTYPE MAPS AND 482 00:22:34,897 --> 00:22:37,066 ESSENTIALLY CREATE A SYNTHETIC 483 00:22:37,066 --> 00:22:39,435 PHENOTYPE IN SILICO AND QUERY 484 00:22:39,435 --> 00:22:41,937 THAT TO DO THE SCREEN AND IN A 485 00:22:41,937 --> 00:22:45,474 NICE EXAMPLE OF THIS JOSEPH AND 486 00:22:45,474 --> 00:22:47,242 RUBIN ESSENTIALLY CREATED FOR 487 00:22:47,242 --> 00:22:50,245 EACH PERTURBATION A SCORE OF HOW 488 00:22:50,245 --> 00:22:51,113 THEY AFFECTED CHROMOSOME 489 00:22:51,113 --> 00:22:56,185 STABILITY AND HOW THEY AFFECT 490 00:22:56,185 --> 00:22:59,755 WILL THE POTENTIAL FOR 491 00:22:59,755 --> 00:23:01,623 ANEUPLOIDITY AND DID A GENOME 492 00:23:01,623 --> 00:23:03,559 WIDE SCREEN FOR FACTORS THAT 493 00:23:03,559 --> 00:23:07,796 MODULATED THE INPUT PROCESS. 494 00:23:07,796 --> 00:23:09,965 AND AGAIN IT WAS THE FACT TH%T 495 00:23:09,965 --> 00:23:11,934 YOU COULD DO THESE COMPLEX IN 496 00:23:11,934 --> 00:23:18,240 WHAT WOULD OTHERWISE BE A 497 00:23:18,240 --> 00:23:24,913 DIFFICULT AND HAVING THE FULL 498 00:23:24,913 --> 00:23:29,485 DATA SET AND DO AND ILLUSTRATES 499 00:23:29,485 --> 00:23:34,289 THE POTENTIAL FOR THE MAPS AS A 500 00:23:34,289 --> 00:23:36,225 DISCOVERY TOOL. 501 00:23:36,225 --> 00:23:39,728 I SAY THIS AND IT SOUNDS LIKE 502 00:23:39,728 --> 00:23:40,696 WE'VE SOLVED ALL THE QUESTION 503 00:23:40,696 --> 00:23:42,731 WHICH IS BY FAR NOT THE CASE. 504 00:23:42,731 --> 00:23:45,167 THERE'S TWO BIG LIMITATIONS. 505 00:23:45,167 --> 00:23:47,936 ONE IS MANY GENES -- IN FACT 506 00:23:47,936 --> 00:23:50,272 MOST GENES WHEN WORKING AT LEAST 507 00:23:50,272 --> 00:23:52,441 IN CANCER CELLS JUST DON'T GIVE 508 00:23:52,441 --> 00:23:54,710 YOU A GOOD TRANSCRIPTIONAL 509 00:23:54,710 --> 00:23:54,977 PHENOTYPE. 510 00:23:54,977 --> 00:23:56,044 IT'S NOT THE RIGHT WAY TO LOOK 511 00:23:56,044 --> 00:23:57,412 AT THE FUNCTION. 512 00:23:57,412 --> 00:23:59,081 WE'RE NOT ASSESSING THE RIGHT 513 00:23:59,081 --> 00:24:01,216 PHYSIOLOGY AND THERE'S FEATURES 514 00:24:01,216 --> 00:24:02,351 NOT WELL READ OUT BY 515 00:24:02,351 --> 00:24:05,754 TRANSCRIPTION AND WE HAVE 516 00:24:05,754 --> 00:24:09,024 MULTIPLE EXAMPLES BUT ONE 517 00:24:09,024 --> 00:24:12,261 CONCRETE ONE THE TOR PATHWAY IS 518 00:24:12,261 --> 00:24:16,732 CRITICAL FOR THE SENSOR OF 519 00:24:16,732 --> 00:24:18,433 CELLULAR STRESS AND AS YOU CAN 520 00:24:18,433 --> 00:24:22,304 TURN IT UP AND DOWN SECONDARY 521 00:24:22,304 --> 00:24:24,239 SIGNATURES FROM TRANSCRIPTION 522 00:24:24,239 --> 00:24:27,442 REALLY ISN'T READ OUT WELL AND 523 00:24:27,442 --> 00:24:29,912 READING THE LEVELS YOU HAVE ROOM 524 00:24:29,912 --> 00:24:31,413 TO MONITOR THE TOR PATHWAY. 525 00:24:31,413 --> 00:24:33,615 IT'S NOT JUST ENOUGH TO HAVE A 526 00:24:33,615 --> 00:24:34,750 LOT OF DIMENSIONS WE HAVE TO 527 00:24:34,750 --> 00:24:36,018 WATCH ALL THE RIGHT FUNCTIONS OF 528 00:24:36,018 --> 00:24:38,787 THE CELLS. 529 00:24:38,787 --> 00:24:41,390 WE HAVE TO GO TO MORAL 530 00:24:41,390 --> 00:24:42,724 MULTI-MODAL READOUTS THE SECOND 531 00:24:42,724 --> 00:24:44,793 IS EVERYTHING DEPENDS ON YOU 532 00:24:44,793 --> 00:24:49,865 HAVING THE RIGHT MODEL SYSTEM. 533 00:24:49,865 --> 00:24:53,435 THIS IS NOT MY QUOTE AND WE 534 00:24:53,435 --> 00:24:55,771 DON'T WANT FUNCTIONAL GENOMICS 535 00:24:55,771 --> 00:24:58,273 TO GET US TO THE WRONG ANSWER 536 00:24:58,273 --> 00:24:58,540 FASTER. 537 00:24:58,540 --> 00:25:00,008 CELLS LIVE IN COMMUNITIES. 538 00:25:00,008 --> 00:25:02,277 WE NEED TO MOVE AWAY FROM 539 00:25:02,277 --> 00:25:05,847 STUDYING MONOCULTURES OF CANCER 540 00:25:05,847 --> 00:25:06,048 CELLS. 541 00:25:06,048 --> 00:25:08,717 THERE'S GREAT OPTIONS OF 542 00:25:08,717 --> 00:25:10,118 STUDYING ORGANOIDS AND OTHER 543 00:25:10,118 --> 00:25:14,289 ASSEMBLIES OR IPS DERIVED CELLS 544 00:25:14,289 --> 00:25:19,161 AND THEY'RE POWERFUL APPROACHES. 545 00:25:19,161 --> 00:25:22,698 WHAT WE'VE BEEN FOCUSSING ON 546 00:25:22,698 --> 00:25:26,301 THOUGH, RUBIN WHO WORKED WITH 547 00:25:26,301 --> 00:25:32,941 JOSEPH ON THE PERTURB SEQ AND A 548 00:25:32,941 --> 00:25:36,311 JUNIOR FELLOW WHO WILL SOON BE 549 00:25:36,311 --> 00:25:37,446 AN ASSISTANT PROFESSOR AT 550 00:25:37,446 --> 00:25:40,415 STANFORD RAN A WONDERFUL 551 00:25:40,415 --> 00:25:42,284 COLLABORATION WITH A POSTDOC AND 552 00:25:42,284 --> 00:25:43,485 HAVE BEEN DEVELOPING APPROACHES 553 00:25:43,485 --> 00:25:47,222 THAT ABUT US TO DELIVER VERY 554 00:25:47,222 --> 00:25:50,292 COMPLEX GENOME WIDE 555 00:25:50,292 --> 00:25:51,693 PERTURBATIONS IN VIVO TO AN 556 00:25:51,693 --> 00:25:53,061 INTACT LIVE ANIMAL. 557 00:25:53,061 --> 00:25:54,329 FOR EXAMPLE TO LIVER OR TO THE 558 00:25:54,329 --> 00:25:55,964 BRAIN OR TO THE HEART. 559 00:25:55,964 --> 00:26:01,937 YOU ESSENTIALLY CREATE A GENETIC 560 00:26:01,937 --> 00:26:02,170 MOSAIC. 561 00:26:02,170 --> 00:26:03,205 FOR EXAMPLE, WE'VE BEEN 562 00:26:03,205 --> 00:26:05,874 FOCUSSING MOST OF OUR PRIMARY 563 00:26:05,874 --> 00:26:07,876 WORK BUILDING ON EARLY 564 00:26:07,876 --> 00:26:10,279 APPROACHES FOR DELIVERY 565 00:26:10,279 --> 00:26:17,686 INCLUDING SEMINOLE WORK AND IN 566 00:26:17,686 --> 00:26:20,622 EACH HEPATOCYTE YOU GET 567 00:26:20,622 --> 00:26:22,291 DIFFERENT PERTURBATIONS OR SET 568 00:26:22,291 --> 00:26:23,759 OF PERTURBATIONS AND IN THE 569 00:26:23,759 --> 00:26:25,460 ANIMAL YOU CAN PROFUSE THE 570 00:26:25,460 --> 00:26:30,365 ANIMAL TO LOCK IN THE PHENOTYPIC 571 00:26:30,365 --> 00:26:32,968 STATE AND DISASSOCIATE THE CELLS 572 00:26:32,968 --> 00:26:35,470 AND DO PERTURB SEQ FROM THE IN 573 00:26:35,470 --> 00:26:38,907 VIVO SAMPLES OR TAKE SECTIONS OF 574 00:26:38,907 --> 00:26:45,447 TISSUES AND THEN USING THE 575 00:26:45,447 --> 00:26:45,847 APPROACH. 576 00:26:45,847 --> 00:26:47,582 WE'RE ABLE TO READ OUT WHAT THE 577 00:26:47,582 --> 00:26:49,451 PERTURBATION IS AND WHAT THE 578 00:26:49,451 --> 00:26:52,321 EFFECTS ON TRANSCRIPTS ARE AND 579 00:26:52,321 --> 00:26:54,189 ALSO BY COMING WITH ANTIBODIES 580 00:26:54,189 --> 00:26:55,691 AND LOOK AT A HIGH RESOLUTION 581 00:26:55,691 --> 00:26:57,225 VIEW OF HOW THE FUNCTION OF THE 582 00:26:57,225 --> 00:26:58,794 CELL IS MODULATED. 583 00:26:58,794 --> 00:27:04,299 SO THESE ARE JUST A FEW OF THE 584 00:27:04,299 --> 00:27:06,301 DIFFERENT EXAMPLES WHERE YOU GET 585 00:27:06,301 --> 00:27:12,207 RICH PHENOTYPES TO MONITOR NOT 586 00:27:12,207 --> 00:27:12,808 JUST TRANSCRIPTIONALLY BUT IN 587 00:27:12,808 --> 00:27:18,947 OVER ABUNDANCE OR MORPHOLOGY OF 588 00:27:18,947 --> 00:27:21,850 ORGANELLES AND COMBINE THESE SO 589 00:27:21,850 --> 00:27:24,086 FOR EACH CELL WE BUILD IN A 590 00:27:24,086 --> 00:27:26,288 MATRIX OF FEATURES WHAT THE 591 00:27:26,288 --> 00:27:28,123 PERTURBATION IS AND WHAT AFFECTS 592 00:27:28,123 --> 00:27:30,859 ON THE RNA AND PROTEINS AND WHAT 593 00:27:30,859 --> 00:27:31,893 NEIGHBORS ARE DOING. 594 00:27:31,893 --> 00:27:33,962 IN THIS WAY START TO UNDERSTAND 595 00:27:33,962 --> 00:27:36,231 THINGS LIKE CELLULAR STRESS 596 00:27:36,231 --> 00:27:40,669 RESPONSES NOT JUST AS A CELL 597 00:27:40,669 --> 00:27:46,308 AUTONOMOUS EFFECT BUT AS AFFECT 598 00:27:46,308 --> 00:27:47,642 ON CELL COMMUNITY. 599 00:27:47,642 --> 00:27:49,111 MORE GENERALLY WE THINK THIS 600 00:27:49,111 --> 00:27:51,012 WILL BE A GREAT TOOL FOR 601 00:27:51,012 --> 00:27:55,450 DISCOVERY AND REALLY COMES AT A 602 00:27:55,450 --> 00:27:56,385 VERY PROPITIOUS TIME IN BIOLOGY 603 00:27:56,385 --> 00:27:58,286 WHERE THERE'S BEEN A GREAT DEAL 604 00:27:58,286 --> 00:28:04,826 OF EXCITEMENT ABOUT MARRYING 605 00:28:04,826 --> 00:28:05,594 MACHINE LEARNING APPROACHES WITH 606 00:28:05,594 --> 00:28:09,297 DATA RICH BIOLOGICAL DATA SETS. 607 00:28:09,297 --> 00:28:12,534 AND THIS IS NICELY ENCAPSULATED 608 00:28:12,534 --> 00:28:17,506 BY A WONDERFUL BUT BOLD QUOTE IF 609 00:28:17,506 --> 00:28:20,375 HE'D NOT JUST SOLVED THE PROTEIN 610 00:28:20,375 --> 00:28:23,578 FOLDING PROBLEM MAY SEEM LIKE A 611 00:28:23,578 --> 00:28:26,281 GRANDIOSE STATEMENT BUT WITH A 612 00:28:26,281 --> 00:28:28,583 NOD TO GALILEO'S COMMENT THE 613 00:28:28,583 --> 00:28:31,953 LANGUAGE OF THE UNIVERSITY IS 614 00:28:31,953 --> 00:28:34,689 MATHEMATICS HE PROPOSED THAT IT 615 00:28:34,689 --> 00:28:38,293 MAY TURN OUT TO PLAY A SIMILAR 616 00:28:38,293 --> 00:28:39,194 ROLE FOR BIOLOGY AND THE 617 00:28:39,194 --> 00:28:43,765 COMPLEMENT TO THIS IS A QUOTE 618 00:28:43,765 --> 00:28:47,969 FROM DAPHNE KOLLOR FOUNDER AND 619 00:28:47,969 --> 00:28:52,274 CEO OF INSITRO A COMPANY 620 00:28:52,274 --> 00:28:53,975 DEDICATED TO APPLYING MACHINE 621 00:28:53,975 --> 00:28:55,811 LEARNING FOR DRUG DISCOVERY AND 622 00:28:55,811 --> 00:28:57,446 A LEADER IN MACHINE LEARNING 623 00:28:57,446 --> 00:29:00,315 THAT REALLY WHAT THEY'RE 624 00:29:00,315 --> 00:29:03,084 LIMITATION IS DOES NOT COMPUTE 625 00:29:03,084 --> 00:29:05,987 BUT THOUGH THE COMPUTE IS 626 00:29:05,987 --> 00:29:07,556 IMPORTANT BUT THEY NEED A 627 00:29:07,556 --> 00:29:08,757 THOUSAND TIMES THE AMOUNT OF 628 00:29:08,757 --> 00:29:12,761 DATA AND I THINK IN JUST THE 629 00:29:12,761 --> 00:29:16,665 QUANTITY BUT A QUALITATIVE 630 00:29:16,665 --> 00:29:23,738 DIFFERENT TYPE OF DETE -- DATA 631 00:29:23,738 --> 00:29:25,674 AND HOPE THIS WILL HELP GENERATE 632 00:29:25,674 --> 00:29:29,978 THIS TYPE OF DATA AND MORE 633 00:29:29,978 --> 00:29:31,613 BROADLY THAT THEY'LL HELP TO 634 00:29:31,613 --> 00:29:33,615 CREATE FOUNDATION MODELS OF 635 00:29:33,615 --> 00:29:34,883 CELLS AND WITH SOME OF THE 636 00:29:34,883 --> 00:29:38,687 FOLLOWING FEATURES THAT WE HAVE 637 00:29:38,687 --> 00:29:43,191 MULTI-MODAL TO CREATE EMBEDDINGS 638 00:29:43,191 --> 00:29:44,359 AND ENCOMPASS DNA AND 639 00:29:44,359 --> 00:29:46,294 TRANSCRIPTS AND CELL-CELL 640 00:29:46,294 --> 00:29:49,397 INTERACTIONS AND THEY'LL BE 641 00:29:49,397 --> 00:29:50,131 GENERATIVE CAPABILITIES. 642 00:29:50,131 --> 00:29:53,235 PREDICTING MUTATIONS OR DRUG 643 00:29:53,235 --> 00:29:54,970 TREATMENTS, IDENTIFYING TARGETS, 644 00:29:54,970 --> 00:29:57,606 MODELLING DISEASES AND EVEN 645 00:29:57,606 --> 00:29:58,440 ALLOWING ACROSS SPECIES 646 00:29:58,440 --> 00:30:00,075 PREDICTION BY COLLECTING THESE 647 00:30:00,075 --> 00:30:01,843 TYPES OF DATA SETS IN DIFFERENT 648 00:30:01,843 --> 00:30:05,413 ANIMAL MODELS AND CELL MODELS. 649 00:30:05,413 --> 00:30:10,285 AND THEN PERHAPS MOST 650 00:30:10,285 --> 00:30:14,956 AMBITIOUSLY TRY TO HAVE 651 00:30:14,956 --> 00:30:20,962 INTERPRETTABLE INTERPRETABLE 652 00:30:20,962 --> 00:30:24,299 APPROACHES TO PREDICT FUNCTION. 653 00:30:24,299 --> 00:30:26,968 SO THAT'S THIS PART OF THE TALK 654 00:30:26,968 --> 00:30:28,370 AND I'LL SWITCH GEARS. 655 00:30:28,370 --> 00:30:35,810 THIS IS REALLY USING HOW WE USE 656 00:30:35,810 --> 00:30:36,945 EPIGENETIC MODULATIONS AND 657 00:30:36,945 --> 00:30:39,447 PERTURBATIONS AS A DISCOVERY 658 00:30:39,447 --> 00:30:40,615 TOOL AND IDENTIFYING POTENTIAL 659 00:30:40,615 --> 00:30:42,284 DRUG TARGETS. 660 00:30:42,284 --> 00:30:44,753 WHAT ABOUT USING THESE AS 661 00:30:44,753 --> 00:30:46,288 EFFECTERS FOR THERAPEUTIC? 662 00:30:46,288 --> 00:30:50,825 SO THE ANSWER IS CRISPR A AS WE 663 00:30:50,825 --> 00:30:52,260 ORIGINALLY DEVELOPED THEM WERE 664 00:30:52,260 --> 00:30:53,128 NOT PARTICULARLY GOOD. 665 00:30:53,128 --> 00:30:55,931 THAT'S BECAUSE THEY WERE 666 00:30:55,931 --> 00:30:56,431 INHERENTLY REVERSIBLE. 667 00:30:56,431 --> 00:30:59,601 YOU COULD TURN DOWN OR UP THE 668 00:30:59,601 --> 00:31:02,270 EXPRESSION BUT ONLY FOR AS LONG 669 00:31:02,270 --> 00:31:06,908 AS YOU EXPRESSED THE DEAD CAS 9 670 00:31:06,908 --> 00:31:08,843 TARGETED TO THE RIGHT GENE. 671 00:31:08,843 --> 00:31:13,081 FOR THIS REASON IT MOTIVATED 672 00:31:13,081 --> 00:31:18,053 JAMES NUNEZ BUILDING OFF OTHER 673 00:31:18,053 --> 00:31:20,655 WORK TO DEVELOP THE PROGRAMMABLE 674 00:31:20,655 --> 00:31:22,223 APPROACH NOT FOR DOING TRANSIENT 675 00:31:22,223 --> 00:31:24,159 REPRESSION OR ACTIVATION OF A 676 00:31:24,159 --> 00:31:28,296 GENE BUT FOR METHYLATING CPG 677 00:31:28,296 --> 00:31:31,499 ISLANDS IN THE DNA AND THIS WAY 678 00:31:31,499 --> 00:31:33,735 PERMANENTLY IN A HIT-AND-RUN 679 00:31:33,735 --> 00:31:34,402 MANNER TURNING OFF EXPRESSION OF 680 00:31:34,402 --> 00:31:35,604 THE GENE. 681 00:31:35,604 --> 00:31:37,906 AND THE REASON THIS WORKS IS 682 00:31:37,906 --> 00:31:42,143 BECAUSE IT BUILDS OFF OF THE 683 00:31:42,143 --> 00:31:44,379 CELL'S NATURAL SYSTEM FOR 684 00:31:44,379 --> 00:31:45,947 SILENCING GENES IN PARTICULAR. 685 00:31:45,947 --> 00:31:50,518 THIS DNA METHYLATION PROCESS AND 686 00:31:50,518 --> 00:31:54,289 THE SPECIFIC BUILDING OFF 687 00:31:54,289 --> 00:32:00,362 MACHINERY THE CELL USES FOR 688 00:32:00,362 --> 00:32:03,732 SILENCING ENDOGENOUS 689 00:32:03,732 --> 00:32:05,734 RETROVIRUSES AND FOUND FOREIGN 690 00:32:05,734 --> 00:32:07,435 ELEMENTS AND MAKE SURE FOR THE 691 00:32:07,435 --> 00:32:09,304 LIFE OF THE ANIMAL THEY DON'T 692 00:32:09,304 --> 00:32:11,606 GET EXPRESSED. 693 00:32:11,606 --> 00:32:13,375 THEY PROPAGATE RELYING ON 694 00:32:13,375 --> 00:32:16,311 ENDOGENOUS MACHINERY AND 695 00:32:16,311 --> 00:32:22,283 BRINGING IN A PROGRAMMABLE 696 00:32:22,283 --> 00:32:24,352 TRANSFERASE IN THE ELEMENT AND 697 00:32:24,352 --> 00:32:29,324 IT BUILDS OFF THE MAINTENANCE 698 00:32:29,324 --> 00:32:30,892 METHYL TRANSFERASE. 699 00:32:30,892 --> 00:32:35,530 WHENEVER IT SEES CPG METHYLATED 700 00:32:35,530 --> 00:32:38,266 IT REMETHYLATED THE SECOND 701 00:32:38,266 --> 00:32:38,733 STRAND. 702 00:32:38,733 --> 00:32:42,303 NOW AS THE CELL DIVIDES YOU GET 703 00:32:42,303 --> 00:32:45,206 A NEW STRAND THAT IS NOT 704 00:32:45,206 --> 00:32:46,841 METHYLATED AND IT COMES IN AND 705 00:32:46,841 --> 00:32:50,278 FILLS THIS IN ALLOWING THIS TO 706 00:32:50,278 --> 00:32:51,279 PROPAGATE INDEFINITELY. 707 00:32:51,279 --> 00:32:56,584 AND INDEED JAMES WAS ABLE TO 708 00:32:56,584 --> 00:32:58,286 SHOW THAT YOU'D GET TRANSIENT 709 00:32:58,286 --> 00:33:00,722 EXPRESSION OF THE CRISPR PROGRAM 710 00:33:00,722 --> 00:33:02,424 TO DEFINE LOCUS THROUGH 711 00:33:02,424 --> 00:33:04,993 ITERATIONS HE WAS ABLE TO FIND 712 00:33:04,993 --> 00:33:07,662 ONES THAT WE'RE NOW ABLE TO ONCE 713 00:33:07,662 --> 00:33:09,664 METHYLATED TO PROPAGATE 714 00:33:09,664 --> 00:33:10,265 INDEFINITELY AND ONE OF THE 715 00:33:10,265 --> 00:33:12,367 EARLY EXPERIMENTS HE CARRIED OUT 716 00:33:12,367 --> 00:33:15,336 A CELL DIVIDING FOR 15 MONTHS 717 00:33:15,336 --> 00:33:18,273 AND SHOWED EVEN 15 MONTHS THE 718 00:33:18,273 --> 00:33:21,009 VAST MAJORITY OF THE PROGENY OF 719 00:33:21,009 --> 00:33:24,345 THE CELL WERE METHYLATED AND 720 00:33:24,345 --> 00:33:24,879 SILENCED. 721 00:33:24,879 --> 00:33:27,515 EVEN MORE RECENTLY ANGELO USING 722 00:33:27,515 --> 00:33:30,885 A CRISPR OFF TARGETED TO SHOW IN 723 00:33:30,885 --> 00:33:35,724 THE MOUSE OVER A YEAR YOU COULD 724 00:33:35,724 --> 00:33:37,625 MAINTAIN THE MENLATION AND 725 00:33:37,625 --> 00:33:39,027 TARGETING OF THE GENE AND REMOVE 726 00:33:39,027 --> 00:33:40,595 THE FRACTION OF THE LIVER AND 727 00:33:40,595 --> 00:33:43,198 THE LIVER BEING ONE FEW ORGANS 728 00:33:43,198 --> 00:33:47,035 THAT CAN REGENERATE AND 729 00:33:47,035 --> 00:33:52,040 REGENERATED LIVER WOULD MAINTAIN 730 00:33:52,040 --> 00:33:54,275 THE SILENCING FROM WHAT WAS 731 00:33:54,275 --> 00:33:58,279 IMPARTED PRIOR TO THE REMOVAL OF 732 00:33:58,279 --> 00:33:59,447 THE PART. 733 00:33:59,447 --> 00:34:01,683 SO FINALLY, THIS WAS REALLY 734 00:34:01,683 --> 00:34:02,517 HIGHLY SPECIFIC. 735 00:34:02,517 --> 00:34:05,120 HERE WE'RE TARGETING THREE 736 00:34:05,120 --> 00:34:07,522 DIFFERENT GENES AND SHOWING YOU 737 00:34:07,522 --> 00:34:12,927 THE RNA SEQ DATA ANALYSIS AND 738 00:34:12,927 --> 00:34:16,498 THIS IS NOT THE FIGURE LEGEND 739 00:34:16,498 --> 00:34:18,433 BUT THE TARGETED GENE IN THE 740 00:34:18,433 --> 00:34:22,270 GENOME. 741 00:34:22,270 --> 00:34:26,174 THE REASON THIS WORKS IS BECAUSE 742 00:34:26,174 --> 00:34:28,343 THIS SHOWS WHEN WE TARGET WE GET 743 00:34:28,343 --> 00:34:30,278 NICE METHYLATION AT THIS POINT 744 00:34:30,278 --> 00:34:34,015 AND NO METHYLATION ON THE STRAND 745 00:34:34,015 --> 00:34:38,186 FOR THE REMAINDER OF THE DNA. 746 00:34:38,186 --> 00:34:39,187 THE REASON IT WORK AND THE 747 00:34:39,187 --> 00:34:42,223 PICTURE TO HAVE IN MIND IS MOST 748 00:34:42,223 --> 00:34:45,026 CPGs IN THE GENOME ARE 749 00:34:45,026 --> 00:34:46,294 METHYLATED. 750 00:34:46,294 --> 00:34:50,265 IT'S ONLY THESE CPG ISLANDS AND 751 00:34:50,265 --> 00:34:52,567 RICH REGIONS IN GENES EXPRESSED 752 00:34:52,567 --> 00:34:56,037 THAT HAVE THIS SMALL WINDOW OF 753 00:34:56,037 --> 00:34:58,273 UNMETHYLATED CPGs. 754 00:34:58,273 --> 00:34:59,374 AND ESSENTIALLY WHAT THE CRISPR 755 00:34:59,374 --> 00:35:03,178 IS DOING IS COMING IN AND FILL 756 00:35:03,178 --> 00:35:05,680 IN THE METHYLATION IN THE CPG 757 00:35:05,680 --> 00:35:08,349 ISLAND BUT IT'S NOT TARGETING 758 00:35:08,349 --> 00:35:12,253 THE CPGs IN THE PROXIMAL GENE. 759 00:35:12,253 --> 00:35:17,158 IN THIS WAY WE'RE ABLE TO 760 00:35:17,158 --> 00:35:20,728 SILENCE IN A HERITABLE AND 761 00:35:20,728 --> 00:35:22,096 STABLE MANNER WITHOUT AFFECTING 762 00:35:22,096 --> 00:35:23,998 THE OTHER GENES. 763 00:35:23,998 --> 00:35:26,034 THAT'S WHERE WE STOOD IN THE 764 00:35:26,034 --> 00:35:26,534 FAIRLY RECENTLY. 765 00:35:26,534 --> 00:35:30,839 WE THOUGHT THIS WOULD BE A GREAT 766 00:35:30,839 --> 00:35:34,175 TOOL FOR UNDERSTANDING 767 00:35:34,175 --> 00:35:38,313 METHYLATION AND SILENCING AND 768 00:35:38,313 --> 00:35:40,882 HAD THERAPEUTIC POTENTIAL BUT 769 00:35:40,882 --> 00:35:43,117 THAT WAS IN MY DISCLOSURES LUKE 770 00:35:43,117 --> 00:35:48,423 GILBERT AND I TOGETHER WITH 771 00:35:48,423 --> 00:35:50,358 ANGELO AND STARTED A COMPANY 772 00:35:50,358 --> 00:35:54,295 THAT WOULD TRY TO USE THIS TO 773 00:35:54,295 --> 00:35:55,430 THERAPEUTIC SILENCING AND 774 00:35:55,430 --> 00:35:56,798 THOUGHT THERE'D BE A SEPARATION 775 00:35:56,798 --> 00:35:58,366 OF CHURCH AND STATE AND IT WOULD 776 00:35:58,366 --> 00:36:00,335 BE GREAT TO DEVELOP THE 777 00:36:00,335 --> 00:36:01,703 THERAPEUTICS AND WE'D TWO OTHER 778 00:36:01,703 --> 00:36:04,706 SCIENCE WE'RE BETTER SUITED TO 779 00:36:04,706 --> 00:36:05,907 DO. 780 00:36:05,907 --> 00:36:12,380 THAT WAS THE CASE UNTIL ALMOST A 781 00:36:12,380 --> 00:36:16,351 LITTLE UNDER TWO YEARS AGO WHEN 782 00:36:16,351 --> 00:36:17,452 ALMOST 10 YEARS TO THE DAY 783 00:36:17,452 --> 00:36:21,789 JENNIFER WALKED IN MY OFFICE, 784 00:36:21,789 --> 00:36:23,558 THEY WALKED IN MY OFFICE AND 785 00:36:23,558 --> 00:36:26,294 SAID DO YOU THINK YOU CAN DO 786 00:36:26,294 --> 00:36:30,164 EPIGENETIC SILENCING TO SILENCE 787 00:36:30,164 --> 00:36:34,269 THE PRION PROTEIN? 788 00:36:34,269 --> 00:36:37,939 SO SONIA AND ERIC WON A LAB IN 789 00:36:37,939 --> 00:36:40,975 THE BROAD INSTITUTE NEXT TO US 790 00:36:40,975 --> 00:36:41,743 DEDICATED DEVELOPING 791 00:36:41,743 --> 00:36:44,545 THERAPEUTICS FOR THE PRION 792 00:36:44,545 --> 00:36:46,047 DISEASE AND AS SONIA SAYS AND I 793 00:36:46,047 --> 00:36:49,751 URGE YOU TO WATCH HER RECENT TED 794 00:36:49,751 --> 00:36:51,920 TALK SHE SAYS THIS PROBLEM WAS 795 00:36:51,920 --> 00:36:52,954 PERSONAL TO THEM. 796 00:36:52,954 --> 00:36:56,190 SONIA WAS A HARVARD TRAINED 797 00:36:56,190 --> 00:36:59,661 LAWYER AND ERIC WAS A CITY 798 00:36:59,661 --> 00:37:06,334 PLANNER AND SONIA'S MOTHER DIED 799 00:37:06,334 --> 00:37:09,737 OF FATAL FAMILIAL PION DISEASE 800 00:37:09,737 --> 00:37:11,439 AND THEY STOPPED WHAT THEY WERE 801 00:37:11,439 --> 00:37:12,907 DOING AND WENT TO GRADUATE 802 00:37:12,907 --> 00:37:15,944 SCHOOL AND 12 YEARS OR SO LATER 803 00:37:15,944 --> 00:37:18,279 HAD THIS LAB AT THE BROAD 804 00:37:18,279 --> 00:37:22,083 DEVELOPING THERAPEUTICS. 805 00:37:22,083 --> 00:37:24,719 SO IT WAS ALSO A COMPELLING 806 00:37:24,719 --> 00:37:26,354 REASON TO WORK WITH THEM AND 807 00:37:26,354 --> 00:37:27,789 IT'S BEEN WONDERFUL. 808 00:37:27,789 --> 00:37:31,092 THEY'RE TRUE SCIENTIFIC PARTNERS 809 00:37:31,092 --> 00:37:33,027 ON THIS AND IT'S BEEN TERRIFIC 810 00:37:33,027 --> 00:37:36,431 COLLABORATION THEY'VE PROPELLED 811 00:37:36,431 --> 00:37:39,567 FORWARD. 812 00:37:39,567 --> 00:37:46,140 ALSO THERE'S A VERY FROM A WELL 813 00:37:46,140 --> 00:37:49,077 VALIDATED TARGET AND PRIONS ARE 814 00:37:49,077 --> 00:37:54,449 AN INFECTIOUS PROTEIN. 815 00:37:54,449 --> 00:38:00,421 THE CAUSE OF THE SLOW VIRUS THAT 816 00:38:00,421 --> 00:38:01,689 WAS DISCOVERED AND THEN IT WAS 817 00:38:01,689 --> 00:38:05,093 SHOWN IT WAS NOT A VARIETY BUT A 818 00:38:05,093 --> 00:38:07,261 MISFOLDED FORM OF AN ENDOGENOUS 819 00:38:07,261 --> 00:38:09,731 NATURAL PRION PROTEIN. 820 00:38:09,731 --> 00:38:12,467 THE BASIS OF THE PATHOLOGY IS IT 821 00:38:12,467 --> 00:38:16,437 EITHER THROUGH SPORADICALLY, 822 00:38:16,437 --> 00:38:19,674 THROUGH INFECTION FOR EXAMPLE 823 00:38:19,674 --> 00:38:22,076 CONSUMPTION OF INFECTED MEAT IN 824 00:38:22,076 --> 00:38:24,445 THAT CASE MAD COW OR THROUGH 825 00:38:24,445 --> 00:38:27,148 MUTATIONS IN THE CASE OF THINGS 826 00:38:27,148 --> 00:38:34,288 LIKE TSS OR FATAL FAMILIAL 827 00:38:34,288 --> 00:38:36,457 STRAINS AND THE AGGREGATED FORM 828 00:38:36,457 --> 00:38:39,160 FORMED A SEED AND COULD USE 829 00:38:39,160 --> 00:38:42,463 ENDOGENOUS PROTEIN AS A 830 00:38:42,463 --> 00:38:45,733 SUBSTRATE TO CONVERT TO MAKE 831 00:38:45,733 --> 00:38:52,940 MORE OF AND IN THIS WAY YOU GET 832 00:38:52,940 --> 00:38:54,275 AN EXPERIENTIAL PROCESS. 833 00:38:54,275 --> 00:38:58,279 MORE OVER, WE KNEW FROM WORK 834 00:38:58,279 --> 00:39:01,749 STARTED ABOUT 20 YEARS AGO FIRST 835 00:39:01,749 --> 00:39:03,317 MAMMALS CAN SURVIVE WELL WITHOUT 836 00:39:03,317 --> 00:39:04,952 THE PRION PROTEIN. 837 00:39:04,952 --> 00:39:06,954 IT'S NOT EXACTLY CLEAR WHAT IT'S 838 00:39:06,954 --> 00:39:07,155 DOING. 839 00:39:07,155 --> 00:39:09,323 THE LOSS OF FUNCTION PHENOTYPES 840 00:39:09,323 --> 00:39:11,959 ARE NONE OR MILD NEUROPATHY 841 00:39:11,959 --> 00:39:14,829 AFTER EXTENDED PERIOD OF TIME 842 00:39:14,829 --> 00:39:18,332 AND SECOND IS IF YOU EVEN IN THE 843 00:39:18,332 --> 00:39:21,135 ACTIVELY INFECTED MOUSE 844 00:39:21,135 --> 00:39:23,871 SYSTEMATIC FOR THE PRION 845 00:39:23,871 --> 00:39:26,774 DISEASE, IF YOU SHUT OFF IN 846 00:39:26,774 --> 00:39:27,775 NEURONS THE CONTINUED EXPRESSION 847 00:39:27,775 --> 00:39:30,278 OF THE NATURAL PRION PROTEIN YOU 848 00:39:30,278 --> 00:39:33,214 NOW DEPRIVE THE PROCESS OF A NEW 849 00:39:33,214 --> 00:39:34,515 SUBSTRATE TO MAKE MORE 850 00:39:34,515 --> 00:39:37,819 INFECTIOUS PROTEIN AND THE 851 00:39:37,819 --> 00:39:40,988 EXISTING INFECTED PROTEIN 852 00:39:40,988 --> 00:39:44,258 INSTEAD OF GROWING EXPONENTIALLY 853 00:39:44,258 --> 00:39:46,027 DECAYS AND THIS IS ENOUGH TO 854 00:39:46,027 --> 00:39:48,563 INTERFERE AND PREVENT THE 855 00:39:48,563 --> 00:39:52,100 FURTHER DISEASE PROCESS. 856 00:39:52,100 --> 00:39:58,272 SO WE HAD CLEAR THERAPEUTIC 857 00:39:58,272 --> 00:40:00,408 HYPOTHESIS IF WE SILENCE THE 858 00:40:00,408 --> 00:40:04,178 PROTEIN IT WILL HAVE A 859 00:40:04,178 --> 00:40:09,383 THERAPEUTIC AFFECT IN A PREV 860 00:40:09,383 --> 00:40:10,284 PREVENTATIVE FORM FOR PEOPLE 861 00:40:10,284 --> 00:40:12,653 LIKE SONIA WHO INHERIT THE 862 00:40:12,653 --> 00:40:19,327 BETWEEN -- GENETIC MUTATION AND 863 00:40:19,327 --> 00:40:21,662 SOMEONE WHO THE ACTIVE INFECTION 864 00:40:21,662 --> 00:40:23,231 GOING ON. 865 00:40:23,231 --> 00:40:26,701 FIRST THING WE DID AND I SAY WE, 866 00:40:26,701 --> 00:40:29,804 EDWIN NEWMAN A TERRIFIC STUDENT 867 00:40:29,804 --> 00:40:31,172 AND A WONDERFUL POSTDOC IN THE 868 00:40:31,172 --> 00:40:33,407 LAB WORKED CLOSELY ON THE 869 00:40:33,407 --> 00:40:33,641 PROJECT. 870 00:40:33,641 --> 00:40:36,511 FIRST THING THEY DID WAS SEE IN 871 00:40:36,511 --> 00:40:38,713 CELLS CAN WE SILENCE THE PRION 872 00:40:38,713 --> 00:40:38,946 PROTEIN? 873 00:40:38,946 --> 00:40:40,181 THE ANSWER IS YES. 874 00:40:40,181 --> 00:40:42,817 YOU COULD TRANSIENTLY EXPRESS 875 00:40:42,817 --> 00:40:47,588 THE CRISPR OFF TARGETING THE 876 00:40:47,588 --> 00:40:50,291 FWEEN AND THIS WOULD CAUSE 877 00:40:50,291 --> 00:40:53,060 HERITABLE SILENCING OF THE PRION 878 00:40:53,060 --> 00:40:59,634 PROTEIN AND ACCOMPANIED BY 879 00:40:59,634 --> 00:41:01,002 METHYLATION OF THE PRION GENE. 880 00:41:01,002 --> 00:41:06,274 WE LAD A POSSIBILITY OF 881 00:41:06,274 --> 00:41:07,575 GENETICALLY SILENCING THE GENE. 882 00:41:07,575 --> 00:41:09,677 THE CHALLENGE IS NOW HOW DO YOU 883 00:41:09,677 --> 00:41:09,911 DELIVER? 884 00:41:09,911 --> 00:41:12,480 THIS IS THE CHALLENGE OF ALL 885 00:41:12,480 --> 00:41:19,520 GENETS IC -- GENETIC MEDICINE 886 00:41:19,520 --> 00:41:21,022 THE RIGHT CELLS AT THE RIGHT 887 00:41:21,022 --> 00:41:24,225 TIME AND ESSENTIALLY AT THIS 888 00:41:24,225 --> 00:41:27,495 POINT THE ONLY THING VIABLE AND 889 00:41:27,495 --> 00:41:29,230 APPROVED METHOD THAT HAD THE 890 00:41:29,230 --> 00:41:35,403 HELP OF GETTING ACROSS THE CNS 891 00:41:35,403 --> 00:41:41,475 WAS A VIRAL VECTOR. 892 00:41:41,475 --> 00:41:43,277 THE PROBLEM WAS CAS 9 IS A BIG 893 00:41:43,277 --> 00:41:47,448 PROTEIN AND THE GENOME SIZE IS 894 00:41:47,448 --> 00:41:49,417 LIMITED. 895 00:41:49,417 --> 00:41:53,521 THE CRISPR AS WE INITIALLY 896 00:41:53,521 --> 00:41:55,456 DESIGN IT HAD NO CHANCE OF 897 00:41:55,456 --> 00:41:58,292 THINNING IN THE AAD AND YOU MAY 898 00:41:58,292 --> 00:42:04,665 BE ABLE TO BY WORKING WITH 899 00:42:04,665 --> 00:42:06,767 DIFFERENT CASTS YOU MAY GET IT 900 00:42:06,767 --> 00:42:07,168 IN. 901 00:42:07,168 --> 00:42:12,206 IT WAS CLEARLY NOT THE BEST DNA 902 00:42:12,206 --> 00:42:14,842 BINDING DOMAIN TO DELIVER THE 903 00:42:14,842 --> 00:42:16,110 SILENCING. 904 00:42:16,110 --> 00:42:21,215 IN FACT A MORE ATTRACTIVE ONE 905 00:42:21,215 --> 00:42:24,418 WAS AN OLDER TECHNOLOGY WHICH 906 00:42:24,418 --> 00:42:27,455 WAS MUCH SMALLER AND BASED ON 907 00:42:27,455 --> 00:42:28,889 HUMAN PROTAIN. 908 00:42:28,889 --> 00:42:31,425 YOU DON'T HAVE CONCERN ABOUT 909 00:42:31,425 --> 00:42:37,732 ANTIGENISTIC AND WHEN YOU NEED A 910 00:42:37,732 --> 00:42:40,001 BUBBLE TO GET THE AMYLASE IN OR 911 00:42:40,001 --> 00:42:45,906 A NICK FOR THE REVERSE 912 00:42:45,906 --> 00:42:47,942 TRANSCRIPTASE WE'RE LOOKING AT 913 00:42:47,942 --> 00:42:51,145 THE PROTEIN AND IT WAS 914 00:42:51,145 --> 00:42:53,014 INCUMBRANCE TO THE PROCESS WHERE 915 00:42:53,014 --> 00:42:56,250 THE FINGERS SEEM LIKE A MORE 916 00:42:56,250 --> 00:43:00,221 NATURAL DELIVERY FOR OUR 917 00:43:00,221 --> 00:43:06,160 EPIGENETIC SILENCERS. 918 00:43:06,160 --> 00:43:08,296 THIS IS THE SIZE OF THE AAG 919 00:43:08,296 --> 00:43:12,633 GENOME AND WHAT THE FINGER 920 00:43:12,633 --> 00:43:13,968 VERSION WOULD LOOK LIKE. 921 00:43:13,968 --> 00:43:17,471 WHEN YOU WORK CLOSELY WITH THE 922 00:43:17,471 --> 00:43:19,707 SPECIALIST AND WHEN WE MADE OUR 923 00:43:19,707 --> 00:43:21,942 FIRST FING VERSION AND HOE WOULD 924 00:43:21,942 --> 00:43:24,445 MAKE THE AA V FOR US WE GOT AN 925 00:43:24,445 --> 00:43:26,013 E-MAIL BACK SAYING IT LOOKS 926 00:43:26,013 --> 00:43:28,149 GREAT BUT CAN YOU ADD STUFF 927 00:43:28,149 --> 00:43:30,484 BECAUSE IT'S SMALL AND ARAID 928 00:43:30,484 --> 00:43:37,591 OTHER DNA WILL GET IN SO YOU CAN 929 00:43:37,591 --> 00:43:42,663 APPRECIATE THE IRONY OF THAT. 930 00:43:42,663 --> 00:43:49,570 THAT SEEMED SIMPLE WE'D GET OUR 931 00:43:49,570 --> 00:43:51,505 ZINC FINGER AND TARGET THE 932 00:43:51,505 --> 00:43:58,279 PROMOTER AND DELIVER AND THEN WE 933 00:43:58,279 --> 00:44:02,416 GOT A RUDE UNFORTUNATE RESULT 934 00:44:02,416 --> 00:44:05,753 AND THAT YOU WERE HIGHLY TOXIC 935 00:44:05,753 --> 00:44:08,556 EVEN TO CANCER CELL LINES WHEN 936 00:44:08,556 --> 00:44:10,124 WE TRANSIENTLY DELIVERED THEM 937 00:44:10,124 --> 00:44:11,759 DUE TO THE METHYL TRANSFERASE 938 00:44:11,759 --> 00:44:14,261 DOMAIN AND WE MADE ANALOGOUS 939 00:44:14,261 --> 00:44:18,265 VERSIONS WHERE WE REMOVED THE 940 00:44:18,265 --> 00:44:21,602 DOMAIN AND GOT RID OF THIS 941 00:44:21,602 --> 00:44:21,869 TOXICITY. 942 00:44:21,869 --> 00:44:24,138 WHY WAS IT TOXICITY? 943 00:44:24,138 --> 00:44:26,273 I'LL HAVE TO TELL YOU A LITTLE 944 00:44:26,273 --> 00:44:31,779 BIT OF A DETAIL ABOUT AND HOW WE 945 00:44:31,779 --> 00:44:38,285 WERE ABLE TO ENGINEER AROUND THE 946 00:44:38,285 --> 00:44:41,389 TOXICITY AND THERE'S A METHYL 947 00:44:41,389 --> 00:44:46,293 TRANSFERASE AND IT EXISTS IN THE 948 00:44:46,293 --> 00:44:51,832 CELL EN AN AUTO INHIBITED FORM 949 00:44:51,832 --> 00:44:55,369 AND WE'VE REMOVED ALL THE 950 00:44:55,369 --> 00:44:56,937 INHIBITORY DOMAINS AND TAKE THE 951 00:44:56,937 --> 00:45:02,276 TRANS TERRASE AND FUSE IT WITH 952 00:45:02,276 --> 00:45:09,150 THE 3L AND THIS THEN GAVE US A 953 00:45:09,150 --> 00:45:12,086 COMPLETELY UNREGULATED METHYL 954 00:45:12,086 --> 00:45:13,521 TRANSFERASE ONCE YOU METHYLATE 955 00:45:13,521 --> 00:45:16,757 YOU HAVE THE METHYL TRANSFERASE 956 00:45:16,757 --> 00:45:23,264 TO PROPAGATE AND THERE WAS A 957 00:45:23,264 --> 00:45:24,465 MUTIGEN WHERE ANY GENE IT HIT IT 958 00:45:24,465 --> 00:45:28,436 COULD POTENTIALLY TURN OF. 959 00:45:28,436 --> 00:45:32,306 WE ALSO KNEW THANKS TO DECADES 960 00:45:32,306 --> 00:45:36,710 OF FUNCTIONAL WORK THAT HOW THIS 961 00:45:36,710 --> 00:45:38,312 WAS RELEASED. 962 00:45:38,312 --> 00:45:42,683 AN IT'S SNIFFING AROUND AT THE 963 00:45:42,683 --> 00:45:51,125 CHROMOSOME STATES ON THE 964 00:45:51,125 --> 00:45:54,261 HISTONES AND THE TAIL IN THE H3 965 00:45:54,261 --> 00:45:58,065 IS THE UNMETHYLATED FORM AND 966 00:45:58,065 --> 00:46:00,067 IT'S AN INDICATION OF AN 967 00:46:00,067 --> 00:46:02,269 INACTIVE PROMOTER AND THE KEY 968 00:46:02,269 --> 00:46:06,006 THAT RELEASES THIS AUTO 969 00:46:06,006 --> 00:46:06,307 INHIBITION. 970 00:46:06,307 --> 00:46:11,111 WE KNEW FROM A RECENT PAPER 971 00:46:11,111 --> 00:46:13,781 ABOUT THE SAME TIME THAT SHOWED 972 00:46:13,781 --> 00:46:24,358 IN VITRO PEPTIDE FROM AGE 33 973 00:46:31,365 --> 00:46:37,838 TAIL AND WE KNEW 3L WAS 974 00:46:37,838 --> 00:46:42,276 SUFFICIENT TO RECRUIT AND 975 00:46:42,276 --> 00:46:47,214 RELYING ON THE ENDOGENOUS DNMT3L 976 00:46:47,214 --> 00:46:52,520 AROUND HYPOTHESIZED IF WE COULD 977 00:46:52,520 --> 00:46:57,825 PUT IN THE HISTONE TAIL WE CAN 978 00:46:57,825 --> 00:47:00,761 RECRUIT THE TRANSFERASE ONLY 979 00:47:00,761 --> 00:47:04,164 WHERE WE HAD PROGRAMMED TO PLACE 980 00:47:04,164 --> 00:47:06,300 THE 3L. 981 00:47:06,300 --> 00:47:10,271 SO HE MADE VARIOUS VERSIONS. 982 00:47:10,271 --> 00:47:14,942 THE ORIGINAL CHRIS PER AN 983 00:47:14,942 --> 00:47:17,745 RECRUIT THE 3L AND WINNER WITH 984 00:47:17,745 --> 00:47:24,552 THE HOSE HIS TONE TAIL. 985 00:47:24,552 --> 00:47:25,886 REMARKABLY IT WORK AND TOOK 986 00:47:25,886 --> 00:47:28,389 ITERATIONS AND WAS ABLE TO GET 987 00:47:28,389 --> 00:47:30,257 TO WORK ROBUSTLY. 988 00:47:30,257 --> 00:47:35,229 CRISPR I IS TRANSIENT AND 989 00:47:35,229 --> 00:47:36,764 PERMANENT AND INHERITABLE AND 990 00:47:36,764 --> 00:47:40,000 THE ONE WITH THE HISTONE TAIL 991 00:47:40,000 --> 00:47:41,368 WORKS IN THIS CASE ALMOST AS 992 00:47:41,368 --> 00:47:42,770 WELL AND I'LL SHOW YOU AFTER 993 00:47:42,770 --> 00:47:44,872 SOME ITERATIONS IF NOT BETTER 994 00:47:44,872 --> 00:47:47,474 THAN THE ORIGINAL CRISPR. 995 00:47:47,474 --> 00:47:50,277 THIS WAS TERRIFIC BUT YOU GET A 996 00:47:50,277 --> 00:47:53,113 NICE RESULT AND HAVE THIS 997 00:47:53,113 --> 00:47:54,214 ELEGANT MODEL AND THINK WHAT 998 00:47:54,214 --> 00:47:55,082 COULD BE WRONG. 999 00:47:55,082 --> 00:47:57,918 YOU HAVE MOMENTS OF ANXIETY. 1000 00:47:57,918 --> 00:47:59,920 IS THIS WORKING THE WAY WE THINK 1001 00:47:59,920 --> 00:48:10,264 AND MAYBE ITION AND IT ENTAILS 1002 00:48:10,264 --> 00:48:14,001 SOMETHING AND KNEW THE LYSIN 4 1003 00:48:14,001 --> 00:48:15,436 WAS REQUIRED AND EDWARD MADE A 1004 00:48:15,436 --> 00:48:19,006 VARIANT AND CHANGED ONLY THE 1005 00:48:19,006 --> 00:48:26,847 SINGLE LYSIN AND THAT'S AB ROW 1006 00:48:26,847 --> 00:48:28,449 FA 1007 00:48:28,449 --> 00:48:33,988 GAITED AND GAVE HIM CONFIDENCE 1008 00:48:33,988 --> 00:48:36,156 AND THOUGHT HE'D OFFER THE 1009 00:48:36,156 --> 00:48:38,258 TRANSFERASE AND THAT'S A 1010 00:48:38,258 --> 00:48:39,893 MOUNLFUL BUT CHARM IS MEMORABLE 1011 00:48:39,893 --> 00:48:46,100 AND MADE MULTIPLE ITERATIONS OF 1012 00:48:46,100 --> 00:48:48,802 CHARM WITH TETHERS AND A KEY 1013 00:48:48,802 --> 00:48:51,372 EVENT WAS TO GO THROUGH THIS AND 1014 00:48:51,372 --> 00:48:53,374 THE EUROPEAN WOOD MOUSE WAS THE 1015 00:48:53,374 --> 00:48:54,274 WINNER. 1016 00:48:54,274 --> 00:48:55,809 THROUGH THE VARIOUS ITERATIONS 1017 00:48:55,809 --> 00:49:02,282 HE WAS ABLE TO GET CRISPR BASED 1018 00:49:02,282 --> 00:49:04,752 CHARM THAT WORKED AS WELL AS THE 1019 00:49:04,752 --> 00:49:11,025 CRISPR ALL IN THE MAGNITUDE OF 1020 00:49:11,025 --> 00:49:14,261 THE EFFECT. 1021 00:49:14,261 --> 00:49:17,998 HE PUT THIS ON A ZINC FINGER AND 1022 00:49:17,998 --> 00:49:22,102 ELIMINATE THE TOXICITY IN CELL 1023 00:49:22,102 --> 00:49:27,474 LINES AND WAS ABLE TO PROMOTE 1024 00:49:27,474 --> 00:49:30,277 AND NICELY INHERITABLY SILENCE 1025 00:49:30,277 --> 00:49:37,785 THE PRION PROTEIN AND 1026 00:49:37,785 --> 00:49:40,487 METHYLATING THE CPG ISLANDS AND 1027 00:49:40,487 --> 00:49:40,988 THE PRION. 1028 00:49:40,988 --> 00:49:42,256 NOW HE WAS FINALLY ABLE TO TO 1029 00:49:42,256 --> 00:49:43,524 THE EXPERIMENT. 1030 00:49:43,524 --> 00:49:48,462 HE HAD A NON-TOXIC CFP CHARM 1031 00:49:48,462 --> 00:49:53,934 THAT READILY FITS INTO AN AAV. 1032 00:49:53,934 --> 00:49:56,036 WE GAVE THE DESIGN TO BEN TO PUT 1033 00:49:56,036 --> 00:50:00,774 IN HIS CNS TARGETING PHP AND 1034 00:50:00,774 --> 00:50:05,112 THIS IS GREAT AT TARGETING THE 1035 00:50:05,112 --> 00:50:07,848 MOUSE FRAME BUT DOESN'T TARGET 1036 00:50:07,848 --> 00:50:08,949 OTHER STRAINS OF MICE AND 1037 00:50:08,949 --> 00:50:10,684 CERTAINLY NOT HUMANS BUT 1038 00:50:10,684 --> 00:50:13,520 RECENTLY BEN HAS A BEAUTIFUL 1039 00:50:13,520 --> 00:50:17,391 PAPER ON THE HUMAN TARGETING AAV 1040 00:50:17,391 --> 00:50:19,893 BASED ON THE TRANSPARENT 1041 00:50:19,893 --> 00:50:23,697 RECEPTOR AND OTHER COMPANIES 1042 00:50:23,697 --> 00:50:28,535 INCLUDING REGENERON AMONG OTHERS 1043 00:50:28,535 --> 00:50:29,903 HAVE TARGETING TO HUMAN BRAIN. 1044 00:50:29,903 --> 00:50:33,507 IT'S STILL A WORK IN PROGRESS 1045 00:50:33,507 --> 00:50:38,078 BUT THINK THERE'S POTENTIAL TO 1046 00:50:38,078 --> 00:50:43,550 BE TRANSLATED TO HUMANS. 1047 00:50:43,550 --> 00:50:52,893 WE USED THE PHB AND THE EMPTY 1048 00:50:52,893 --> 00:50:58,298 AAV OR CF CHARM TARGETING THE 1049 00:50:58,298 --> 00:51:01,902 PRION PROTEIN AND GET 80% TO 90% 1050 00:51:01,902 --> 00:51:06,340 OF THE PRION PROTEIN EXPRESSION 1051 00:51:06,340 --> 00:51:12,146 AND FROM THE MOUSE FRAMEWWORK W 1052 00:51:12,146 --> 00:51:17,518 THROUGH THE FRAMEWORK AND SEE 1053 00:51:17,518 --> 00:51:22,322 THIS ACROSS THE CNS AND SHOWED 1054 00:51:22,322 --> 00:51:25,225 WHEN THE VAST MAJORITY OF 1055 00:51:25,225 --> 00:51:26,026 NEURONS WE ARE SWITCHING ON AND 1056 00:51:26,026 --> 00:51:27,928 OFF THE PROTEIN. 1057 00:51:27,928 --> 00:51:32,332 IT'S NOT A FOREGONE CONCLUSION. 1058 00:51:32,332 --> 00:51:34,168 THERE'S A CRITICAL ROLE IN 1059 00:51:34,168 --> 00:51:37,037 PROPAGATION BUT THAT'S ONLY TRUE 1060 00:51:37,037 --> 00:51:40,841 IF THE CELLS ARE DIVIDING AND 1061 00:51:40,841 --> 00:51:44,778 DNA IS BEING REPLICATE AND 1062 00:51:44,778 --> 00:51:53,954 TARGETING THE POSTMITOTIC GENE. 1063 00:51:53,954 --> 00:51:56,123 AND I TOLD YOU THIS IN THE 1064 00:51:56,123 --> 00:51:56,657 HIT-AND-RUN MANNER. 1065 00:51:56,657 --> 00:52:03,997 WE DON'T WANT THE CHARM TO BE 1066 00:52:03,997 --> 00:52:04,798 EXPRESSED AS LONG AS AD IN THE 1067 00:52:04,798 --> 00:52:06,600 NEURONS AND THEY'RE NOT DIVIDING 1068 00:52:06,600 --> 00:52:08,735 AND WOULDN'T GET DILUTED OUT AND 1069 00:52:08,735 --> 00:52:14,007 THERE WAS A SELF-SILENCING 1070 00:52:14,007 --> 00:52:17,878 STRATEGY IN WHICHD IN THE 1071 00:52:17,878 --> 00:52:18,512 NEURONS AND THEY'RE NOT DIVIDING 1072 00:52:18,512 --> 00:52:19,146 AND WOULDN'T GET DILUTED OUT AND 1073 00:52:19,146 --> 00:52:19,713 THERE WAS A SELF-SILENCING 1074 00:52:19,713 --> 00:52:20,314 STRATEGY IN WHICH INTHE 1075 00:52:20,314 --> 00:52:20,948 NEURONS AND THEY'RE NOT DIVIDING 1076 00:52:20,948 --> 00:52:21,582 AND WOULDN'T GET DILUTED OUT AND 1077 00:52:21,582 --> 00:52:22,149 THERE WAS A SELF-SILENCING 1078 00:52:22,149 --> 00:52:23,283 STRATEGY IN IN THE 1079 00:52:23,283 --> 00:52:23,917 NEURONS AND THEY'RE NOT DIVIDING 1080 00:52:23,917 --> 00:52:24,551 AND WOULDN'T GET DILUTED OUT AND 1081 00:52:24,551 --> 00:52:25,118 THERE WAS A SELF-SILENCING 1082 00:52:25,118 --> 00:52:26,286 STRATEGY IN A IN THE 1083 00:52:26,286 --> 00:52:26,920 NEURONS AND THEY'RE NOT DIVIDING 1084 00:52:26,920 --> 00:52:27,554 AND WOULDN'T GET DILUTED OUT AND 1085 00:52:27,554 --> 00:52:28,121 THERE WAS A SELF-SILENCING 1086 00:52:28,121 --> 00:52:28,689 STRATEGY IN WHICHV IN THE 1087 00:52:28,689 --> 00:52:29,323 NEURONS AND THEY'RE NOT DIVIDING 1088 00:52:29,323 --> 00:52:29,957 AND WOULDN'T GET DILUTED OUT AND 1089 00:52:29,957 --> 00:52:30,524 THERE WAS A SELF-SILENCING 1090 00:52:30,524 --> 00:52:31,158 STRATEGY IN WHICH AND PUTTING 1091 00:52:31,158 --> 00:52:31,825 TWO COPIES, A PERFECT MATCH OR 1092 00:52:31,825 --> 00:52:34,595 SCRAMBLED KINETICS TO MAKE SURE 1093 00:52:34,595 --> 00:52:39,733 IT OCCURRED RAPIDLY PUT NOT 1094 00:52:39,733 --> 00:52:41,001 ENOUGH EXPRESSION TO SILENCE THE 1095 00:52:41,001 --> 00:52:42,069 PRION GENE ITSELF. 1096 00:52:42,069 --> 00:52:44,171 SHE WAS ABLE TO TUNE THIS AND 1097 00:52:44,171 --> 00:52:48,842 THIS IS THE GOLDILOCKS 1098 00:52:48,842 --> 00:52:52,346 EXPERIMENT WITH THE PAPA BEAR 1099 00:52:52,346 --> 00:52:57,651 AND MAMA BEAR AND THE BABY BEAR 1100 00:52:57,651 --> 00:52:58,986 THAT WAS RIGHT AND SILENCED THE 1101 00:52:58,986 --> 00:53:01,889 PROTEIN AND THE PROMOTER ITSELF 1102 00:53:01,889 --> 00:53:04,157 AND SHOWED THIS IN CELL LINES AT 1103 00:53:04,157 --> 00:53:07,461 GOT THIS ALLELIC SERIES OF 1104 00:53:07,461 --> 00:53:08,662 SELF-SILENCING EITHER TOO SLOW 1105 00:53:08,662 --> 00:53:12,299 OR TOO FAST OR JUST RIGHT IN THE 1106 00:53:12,299 --> 00:53:14,301 MOUSE AND THIS WAS ALSO WORKING 1107 00:53:14,301 --> 00:53:18,005 BIG METHYLATING THE AAV ITSELF 1108 00:53:18,005 --> 00:53:21,241 SO IT WOULD BE WE THINK IN A 1109 00:53:21,241 --> 00:53:22,209 PERMANENTLY SHUT OFF STATE. 1110 00:53:22,209 --> 00:53:23,977 SO IN THE END WHAT WOULD BE LEFT 1111 00:53:23,977 --> 00:53:26,713 IN THE MOUSE WOULD TRANSIENTLY 1112 00:53:26,713 --> 00:53:28,548 EXPRESS THE CFP CHARM AND THEN 1113 00:53:28,548 --> 00:53:34,288 GO IN AND METHYLATE THE 1114 00:53:34,288 --> 00:53:36,723 ENDOGENOUS GENE AND LEAVE THE 1115 00:53:36,723 --> 00:53:39,459 ANIMAL WITH THE TURNED OFF 1116 00:53:39,459 --> 00:53:42,296 PROTEIN AND UNEXPRESSIBLE PIECE 1117 00:53:42,296 --> 00:53:43,830 OF DNA AS CLOSE TO THE PERFECT 1118 00:53:43,830 --> 00:53:47,334 REPAIR AS WE COULD GET. 1119 00:53:47,334 --> 00:53:49,503 NOW WE'RE ABLE TO DO THE 1120 00:53:49,503 --> 00:53:49,803 EXPERIMENT. 1121 00:53:49,803 --> 00:53:50,938 THESE ARE LONG EXPERIMENTS SO 1122 00:53:50,938 --> 00:53:56,476 IT'S A WORK IN PROGRESS. 1123 00:53:56,476 --> 00:54:02,015 THEY INFECTED A MOUSE AND AFTER 1124 00:54:02,015 --> 00:54:06,787 FOUR MONTHS WHEN IT'S BECOMING 1125 00:54:06,787 --> 00:54:09,523 SYMPTOMATIC THEY EXPRESSED MOCK 1126 00:54:09,523 --> 00:54:11,692 VIRUS OR THE CHARMS BOTH CELL 1127 00:54:11,692 --> 00:54:15,495 SILENCING OR NON-FORM AND BY 190 1128 00:54:15,495 --> 00:54:18,298 DAYS AS WE EXPECT ALL THE 1129 00:54:18,298 --> 00:54:22,402 UNTREATED ANIMALS ARE DEAD AND 1130 00:54:22,402 --> 00:54:24,404 NOW A FEW MONTHS OUT THE LARGE 1131 00:54:24,404 --> 00:54:29,209 MAJORITY BUT NOT ALL THE TREATED 1132 00:54:29,209 --> 00:54:31,111 ANIMALS ARE SURVIVING AND IN 1133 00:54:31,111 --> 00:54:31,645 SOME MEASURE STARTING TO 1134 00:54:31,645 --> 00:54:39,920 IMPROVE. 1135 00:54:39,920 --> 00:54:41,588 THIS IS OUR FIRST HINT IT COULD 1136 00:54:41,588 --> 00:54:42,956 BE A VIABLE THERAPY AND THE 1137 00:54:42,956 --> 00:54:44,691 CHALLENGE OF DELIVERY TO THE 1138 00:54:44,691 --> 00:54:46,193 HUMAN BRAIN SHOULD NOT IN ANY 1139 00:54:46,193 --> 00:54:52,366 WAY BE UNDERESTIMATED BUT WE NOW 1140 00:54:52,366 --> 00:54:54,301 HAVE TAKEN THE HUGE BIOLOGICAL 1141 00:54:54,301 --> 00:54:57,738 PROBLEM OF THIS HERITABLE 1142 00:54:57,738 --> 00:55:06,346 MUTATION, MYSTERIOUS DISEASE AND 1143 00:55:06,346 --> 00:55:10,083 COME TO THIS DISCREET 1144 00:55:10,083 --> 00:55:12,119 ENGINEERING PROBLEM WE HOPE 1145 00:55:12,119 --> 00:55:14,021 WE'LL BE ABLE TO ADDRESS IN THE 1146 00:55:14,021 --> 00:55:14,621 NEAR FUTURE. 1147 00:55:14,621 --> 00:55:18,291 AND MORE GENERALLY WE THINK THE 1148 00:55:18,291 --> 00:55:23,463 CHARMS WILL BE A POWERFUL NEXT 1149 00:55:23,463 --> 00:55:27,534 GENERATION OF EPIDEMIOLOGIC 1150 00:55:27,534 --> 00:55:29,770 SILENCERS AND DON'T REQUIRE 1151 00:55:29,770 --> 00:55:32,973 EXPRESSION OF ACTIVITY AND 1152 00:55:32,973 --> 00:55:37,377 RECRUIT ENDOGENOUS AND NOT 1153 00:55:37,377 --> 00:55:39,746 EXPECTED TO BE PARTICULARLY 1154 00:55:39,746 --> 00:55:44,451 MUTIGENIC AND EFFICACIOUS AND 1155 00:55:44,451 --> 00:55:48,855 FOR ANY MODALITY WHETHER AAV OR 1156 00:55:48,855 --> 00:55:52,426 LNP BECAUSE THEY'RE A SIMPLE 1157 00:55:52,426 --> 00:55:54,061 SINGLE PROTEIN AND DON'T REQUIRE 1158 00:55:54,061 --> 00:55:56,530 GUIDE RNA THEY'LL BE MORE 1159 00:55:56,530 --> 00:55:57,864 READILY DELIVERABLE. 1160 00:55:57,864 --> 00:56:00,867 AND STEPPING BACK THIS IS PART 1161 00:56:00,867 --> 00:56:03,437 OF MUCH BROADER EFFORT OF WHERE 1162 00:56:03,437 --> 00:56:06,306 WE CAN USE CRISPR-BASED 1163 00:56:06,306 --> 00:56:09,209 FUNCTIONAL GENOMICS AS A WAY OF 1164 00:56:09,209 --> 00:56:11,011 IDENTIFYING TARGETS THAT ARE 1165 00:56:11,011 --> 00:56:11,645 DISEASE MODULATING AND 1166 00:56:11,645 --> 00:56:12,779 UNDERSTANDING THEIR MECHANISM ON 1167 00:56:12,779 --> 00:56:14,815 ONE HAND AND ON THE OTHER HAND 1168 00:56:14,815 --> 00:56:18,351 WE CAN THEN COME IN AND USE 1169 00:56:18,351 --> 00:56:24,091 CRISPR OR FINGER-BASED TOOLS TO 1170 00:56:24,091 --> 00:56:26,293 DIRECT A PROGRAM TO ADDRESS AND 1171 00:56:26,293 --> 00:56:31,765 TO AFFECT THE CHANGE THAT WE NOW 1172 00:56:31,765 --> 00:56:38,305 UNDERSTAND IS GENE MODULATORY 1173 00:56:38,305 --> 00:56:40,207 AND GAVE US THE TOOLS TO 1174 00:56:40,207 --> 00:56:41,241 MODULATE. 1175 00:56:41,241 --> 00:56:42,742 WITH THAT I'LL FINISH AND 1176 00:56:42,742 --> 00:56:43,643 IMPORTANTLY I THANK THE PEOPLE 1177 00:56:43,643 --> 00:56:45,112 WHO DID THE WORK. 1178 00:56:45,112 --> 00:56:47,214 I'VE TRIED TO MENTION THEM ALONG 1179 00:56:47,214 --> 00:56:52,419 THE WAY BUT IF IT BEARS 1180 00:56:52,419 --> 00:56:57,023 REPEATING, EDWIN AND TESSA AND 1181 00:56:57,023 --> 00:57:03,263 MANY ELAINE WU WORKING WITH 1182 00:57:03,263 --> 00:57:06,333 SONIA AND BEN LED THE WORK AND 1183 00:57:06,333 --> 00:57:11,738 GOING IN TRE FIRST CONCEPTION 1184 00:57:11,738 --> 00:57:12,939 AND FULL SPEED AHEAD FOCUSSING 1185 00:57:12,939 --> 00:57:22,282 ON THE HUMAN PRION PROTEIN. 1186 00:57:22,282 --> 00:57:30,824 AND RUBEN AND JOSEPH WORKED IN 1187 00:57:30,824 --> 00:57:34,294 COLLABORATION AND REUBEN WITH 1188 00:57:34,294 --> 00:57:36,963 WILL AND OUR COLLABORATION LEAD 1189 00:57:36,963 --> 00:57:40,100 THE NEXT GENERATION OF IN VIVO 1190 00:57:40,100 --> 00:57:40,967 FUNCTIONAL GENOMICS. 1191 00:57:40,967 --> 00:57:42,402 WIN THAT I'LL END AND I'LL TAKE 1192 00:57:42,402 --> 00:57:52,646 ANY QUESTIONS. 1193 00:57:54,181 --> 00:57:59,519 >> FIRST THE CME CODE IS 50128 1194 00:57:59,519 --> 00:58:07,394 AND SONIA AND ERIC GAVE A WALs 1195 00:58:07,394 --> 00:58:09,095 TALK AND THE ARCHIVE IS 1196 00:58:09,095 --> 00:58:12,065 AVAILABLE IF ANYONE WANTS TO 1197 00:58:12,065 --> 00:58:12,265 WATCH. 1198 00:58:12,265 --> 00:58:14,000 >> BEAUTIFUL AS ALWAYS. 1199 00:58:14,000 --> 00:58:17,737 ONE GENERAL QUESTION IS NOW THAT 1200 00:58:17,737 --> 00:58:19,472 YOU HAVE SPECIFIC EDITORS LIKE 1201 00:58:19,472 --> 00:58:21,942 BASE EDITING AND YOU CAN 1202 00:58:21,942 --> 00:58:24,978 TARGET -- WHAT ARE THE 1203 00:58:24,978 --> 00:58:26,313 ADVANTAGES OF THE PERMANENT 1204 00:58:26,313 --> 00:58:28,315 SILENCING OPPOSED TO INTRODUCING 1205 00:58:28,315 --> 00:58:30,283 POINT MUTATION THAT WILL 1206 00:58:30,283 --> 00:58:36,923 INTRODUCE A STOP. 1207 00:58:36,923 --> 00:58:39,593 >> WHY WOULD YOU PUT IN A STOP 1208 00:58:39,593 --> 00:58:41,661 CODE THAT CREATES A DAMAGED 1209 00:58:41,661 --> 00:58:43,663 PROTEIN AND MESSAGE THAT WOULD 1210 00:58:43,663 --> 00:58:44,464 EXPRESSED FOR THE REST OF THE 1211 00:58:44,464 --> 00:58:47,167 LIFE OF THE ORGANISM THAN 1212 00:58:47,167 --> 00:58:47,801 PREVENTING EXPRESSION OF THE 1213 00:58:47,801 --> 00:58:48,301 GENE? 1214 00:58:48,301 --> 00:58:51,504 SO IF YOU HAVE A GENETIC 1215 00:58:51,504 --> 00:58:54,107 MUTATION LIKE SICKLE CELL 1216 00:58:54,107 --> 00:58:56,243 TRAITOR MUTATION IN CFTR AND 1217 00:58:56,243 --> 00:58:58,278 NEED TO CORRECT THAT, FOR SURE 1218 00:58:58,278 --> 00:59:02,282 THE BASE EDITING AND PRIME 1219 00:59:02,282 --> 00:59:06,186 EDITING APPROACHES ARE POWERFUL 1220 00:59:06,186 --> 00:59:08,688 APPROACHES BUT IF YOU WANT TO 1221 00:59:08,688 --> 00:59:14,294 TURN OFF OR TURN UP THE FUNCTION 1222 00:59:14,294 --> 00:59:17,797 OF GENE, THE EPIGENETIC 1223 00:59:17,797 --> 00:59:19,099 MODULATORS ARE A MORE NAH 1224 00:59:19,099 --> 00:59:20,433 NATURAL THING. 1225 00:59:20,433 --> 00:59:24,938 WE HAVE A TOOL KIT OF PRECISION 1226 00:59:24,938 --> 00:59:25,505 MROJ LATERS AND WE'RE NOW 1227 00:59:25,505 --> 00:59:27,240 GETTING TOOLS THAT ALLOW US TO 1228 00:59:27,240 --> 00:59:30,310 PUT IN LARGER PIECES OF DNA AS 1229 00:59:30,310 --> 00:59:33,546 WELL. 1230 00:59:33,546 --> 00:59:35,682 I THINK IT'S BETTER TO SEE THE 1231 00:59:35,682 --> 00:59:38,852 CHANGES WANT TO MAKE AND WHAT 1232 00:59:38,852 --> 00:59:40,487 ARROWS DO WE PULL OUT OF THE 1233 00:59:40,487 --> 00:59:42,789 QUIVER TO DO THIS? 1234 00:59:42,789 --> 00:59:44,291 THAT SAID GENETIC DISEASES ARE 1235 00:59:44,291 --> 00:59:45,558 VERY IMPORTANT BUT THE MECHANISM 1236 00:59:45,558 --> 00:59:54,301 OF DRUG IS TO MODULATE THE 1237 00:59:54,301 --> 00:59:54,934 FUNCTION. 1238 00:59:54,934 --> 00:59:57,237 THE GENETIC MODULATORS ARE AN 1239 00:59:57,237 --> 00:59:59,105 IMPORTANT PART BUT NOT THE WHOLE 1240 00:59:59,105 --> 00:59:59,306 PART. 1241 00:59:59,306 --> 01:00:01,041 >> IS THIS NOW MORE EFFICIENCY? 1242 01:00:01,041 --> 01:00:04,177 SHOULD WE USE THESE TOOLS IN THE 1243 01:00:04,177 --> 01:00:09,249 RAB WHEN WE DO KNOCKOUTS INSTEAD 1244 01:00:09,249 --> 01:00:11,217 OF THE USUAL CAS 9? 1245 01:00:11,217 --> 01:00:14,287 >> SO WE DID DO GENOME WIDE 1246 01:00:14,287 --> 01:00:16,656 CRISPR OFFERS AND I AND IT WAS 1247 01:00:16,656 --> 01:00:17,457 GENERALLY MORE EFFECTIVE. 1248 01:00:17,457 --> 01:00:19,993 WE DON'T HAVE THE SAME -- IT'S 1249 01:00:19,993 --> 01:00:21,594 SOMETHING WE WANT TO DO 1250 01:00:21,594 --> 01:00:22,696 ESPECIALLY WITH THE CHARM FORMAT 1251 01:00:22,696 --> 01:00:24,931 WHERE YOU DON'T HAVE TO EXPRESS 1252 01:00:24,931 --> 01:00:28,068 THE METHYL TRANSFERASE. 1253 01:00:28,068 --> 01:00:31,771 WE SAW THE EXPRESSION AND 1254 01:00:31,771 --> 01:00:34,274 TOXICITY AND THAT WAS HESITATION 1255 01:00:34,274 --> 01:00:37,677 TO GO CRISPR OFF SCREEN AND NOW 1256 01:00:37,677 --> 01:00:40,547 WITH THE CHARM IT COULD BE A 1257 01:00:40,547 --> 01:00:43,283 MORE EFFICACIOUS FORM AND MORE 1258 01:00:43,283 --> 01:00:46,353 SWITCH LIKE AND IF YOU WANT TO 1259 01:00:46,353 --> 01:00:47,554 GO AFTER CENTRAL GENES AND LOSS 1260 01:00:47,554 --> 01:00:50,290 OF FUNCTION BUT IF YOU WANT A 1261 01:00:50,290 --> 01:00:52,392 TRUE NUL THE CRISPR WILL BE 1262 01:00:52,392 --> 01:00:54,928 BETTER AND IT'S PRETTY MUCH THE 1263 01:00:54,928 --> 01:01:04,270 SAME LIBRARY WHICH IS A NICE 1264 01:01:04,270 --> 01:01:04,971 VITRO OF THIS. 1265 01:01:04,971 --> 01:01:08,308 >> I'M EXCITED ABOUT YOUR 1266 01:01:08,308 --> 01:01:12,912 VIRTUAL CELL PROJECT OR YOU 1267 01:01:12,912 --> 01:01:14,447 MENTIONED PART OF THAT. 1268 01:01:14,447 --> 01:01:18,718 WHEN IT FINISHES IT WILL MAKE 1269 01:01:18,718 --> 01:01:21,321 HALF OF BIOLOGY OBSOLETE SO 1270 01:01:21,321 --> 01:01:28,194 BASED ON MY CAREER AND ANXIETY 1271 01:01:28,194 --> 01:01:31,197 SO ARE IN THE MULTI-DIMENSIONAL 1272 01:01:31,197 --> 01:01:34,567 SPACE ONE AT A TIME. 1273 01:01:34,567 --> 01:01:36,636 ARE THERE ANY PUBLICALLY 1274 01:01:36,636 --> 01:01:41,641 AVAILABLE INFORMATION ABOUT THAT 1275 01:01:41,641 --> 01:01:48,381 MAPPING AND DID YOU ACHIEVE FIND 1276 01:01:48,381 --> 01:01:50,283 PROPERTIES TO SEE HOW CLOSE THEY 1277 01:01:50,283 --> 01:01:50,517 ARE? 1278 01:01:50,517 --> 01:01:53,052 >> SO FIRST, THE VIRTUAL CELL 1279 01:01:53,052 --> 01:01:57,891 LINE THERE'S A LOT OF PEOPLE OF 1280 01:01:57,891 --> 01:02:00,260 INTEREST IN THIS AND I THINK 1281 01:02:00,260 --> 01:02:01,628 IT'S GOING TO BE A LONGER TERM 1282 01:02:01,628 --> 01:02:04,164 PROJECT SO I THINK WE ALL HAVE A 1283 01:02:04,164 --> 01:02:06,299 JOB FOR AT LEAST A LITTLE WHILE 1284 01:02:06,299 --> 01:02:09,302 AND BY THEN THE A.I. OVERLORDS 1285 01:02:09,302 --> 01:02:11,204 WILL BE FEEDING US ANYWAY. 1286 01:02:11,204 --> 01:02:13,740 IT WILL BE IRRELEVANT. 1287 01:02:13,740 --> 01:02:18,278 I THINK MORE SERIOUSLY THAT THE 1288 01:02:18,278 --> 01:02:20,079 AN ASPIRATIONAL THING BUT THE 1289 01:02:20,079 --> 01:02:23,716 MUCH MORE SPECIFIC THINGS LIKE 1290 01:02:23,716 --> 01:02:26,186 PREDICTING THE SPOKE AND THE 1291 01:02:26,186 --> 01:02:29,689 IMAGES OF CELL FROM TRANSCRIPT 1292 01:02:29,689 --> 01:02:31,658 OR FROM PERTURBATION AND THE 1293 01:02:31,658 --> 01:02:33,593 IMAGES ARE THE MORE DIRECT 1294 01:02:33,593 --> 01:02:35,161 THINGS AND WHETHER WE CAN BUILD 1295 01:02:35,161 --> 01:02:37,464 A FOUNDATION MODEL TO CAPTURE 1296 01:02:37,464 --> 01:02:42,101 ALL THESE I THINK IS AN OPEN 1297 01:02:42,101 --> 01:02:44,771 QUESTION AND OUR STRATEGY THE 1298 01:02:44,771 --> 01:02:47,674 DATA WE HAVE WILL BE USEFUL AND 1299 01:02:47,674 --> 01:02:48,942 MOTIVATIONAL FOR THESE EFFORTS 1300 01:02:48,942 --> 01:02:51,177 BUT HOW FAR WILL THEY GO? 1301 01:02:51,177 --> 01:02:52,912 I THINK WE HAVE A LONG WAY 1302 01:02:52,912 --> 01:02:55,915 BEFORE WE'RE ALL OUT OF A JOB. 1303 01:02:55,915 --> 01:02:57,750 WE PUBLISHED AND IT'S ALL 1304 01:02:57,750 --> 01:03:00,353 AVAILABLE THE MAP WE HAVE. 1305 01:03:00,353 --> 01:03:03,823 I THINK WHAT WE HOPE IS WE'LL BE 1306 01:03:03,823 --> 01:03:05,925 AND WE'RE SURE WE'LL BE 1307 01:03:05,925 --> 01:03:07,560 GENERATING RICHER AND LARGER 1308 01:03:07,560 --> 01:03:08,595 SCALE MAPS FROM THE IN VIVO 1309 01:03:08,595 --> 01:03:09,696 PHENOTYPES. 1310 01:03:09,696 --> 01:03:14,300 I THINK IN MANY WAYS AS WE GO TO 1311 01:03:14,300 --> 01:03:17,637 MORE PHYSIOLOGIC CELLS IPS 1312 01:03:17,637 --> 01:03:19,138 DERIVED CELLS OR IN VIVO, THE 1313 01:03:19,138 --> 01:03:20,607 LOGIC OF THE SYSTEM WILL BE 1314 01:03:20,607 --> 01:03:23,042 CLEARER AND MORE INTERPRETABLE. 1315 01:03:23,042 --> 01:03:25,144 I GUESS I WOULD SAY WE'RE AT A 1316 01:03:25,144 --> 01:03:27,046 PLACE WHERE THE ABILITY TO 1317 01:03:27,046 --> 01:03:30,183 COLLECT DATA HAS INCREASED SO 1318 01:03:30,183 --> 01:03:31,117 RAPIDLY UP SOME WAYS WE SHOULD 1319 01:03:31,117 --> 01:03:35,688 WORK ON THE METHODS BUT ALSO 1320 01:03:35,688 --> 01:03:37,924 MOTIVATING WHAT THE ARE NEXT 1321 01:03:37,924 --> 01:03:39,125 DATA SETS WE'D WANT. 1322 01:03:39,125 --> 01:03:40,426 >> I'D LIKE TO SLIP IN ONE 1323 01:03:40,426 --> 01:03:41,294 QUESTION ONLINE. 1324 01:03:41,294 --> 01:03:41,661 >> SURE. 1325 01:03:41,661 --> 01:03:47,867 >> IT'S A DOOZY. 1326 01:03:47,867 --> 01:03:49,769 SO POOLED IN VIVO. 1327 01:03:49,769 --> 01:03:51,237 PERTURBATION SCREENS ARE 1328 01:03:51,237 --> 01:03:52,705 ATTRACTIVE BUT ALSO SEEMS TO 1329 01:03:52,705 --> 01:03:57,744 PRESENT DIFFICULTY IN DATA 1330 01:03:57,744 --> 01:04:02,982 INTERPRETATION IT'S CONFOUNDING 1331 01:04:02,982 --> 01:04:04,817 THE EFFECT OF THE RNA DEPENDING 1332 01:04:04,817 --> 01:04:06,319 ON THE STATE. 1333 01:04:06,319 --> 01:04:09,756 IN VIVO CULTURES AT LEAST 1334 01:04:09,756 --> 01:04:10,523 PROVIDE REDUNDANT REPLICATES 1335 01:04:10,523 --> 01:04:11,591 WITHIN THE SAMPLE. 1336 01:04:11,591 --> 01:04:12,458 HOW DO YOU ADDRESS THE 1337 01:04:12,458 --> 01:04:12,825 CHALLENGE? 1338 01:04:12,825 --> 01:04:16,729 >> I GUESS I WOULD SAY THAT 1339 01:04:16,729 --> 01:04:18,598 CELLS MOSTLY FUNCTION AND THIS 1340 01:04:18,598 --> 01:04:20,199 IS HOW THEY'VE EVOLVED IN THE 1341 01:04:20,199 --> 01:04:20,867 NORMAL FUNCTION. 1342 01:04:20,867 --> 01:04:26,639 WE HAVE TO DEAL WITH THE 1343 01:04:26,639 --> 01:04:26,940 COMPLEXITY. 1344 01:04:26,940 --> 01:04:30,176 THAT SAID IT'S COMPLEX BUT NOT 1345 01:04:30,176 --> 01:04:30,743 SO COMPLEX. 1346 01:04:30,743 --> 01:04:32,912 YOU HAVE THREE ZONES FROM THE 1347 01:04:32,912 --> 01:04:34,247 PERIPORTAL TO THE PERICENTRAL 1348 01:04:34,247 --> 01:04:35,548 AND WELL DEFINED. 1349 01:04:35,548 --> 01:04:38,284 WE'RE ABLE TO GET GOOD COVERAGE 1350 01:04:38,284 --> 01:04:43,356 MUCH EACH OF THE ZONES AND YOU 1351 01:04:43,356 --> 01:04:45,592 CAN TELL WHICH ZONE WE'RE IN. 1352 01:04:45,592 --> 01:04:48,628 IT HAS A LEVEL OF COMPLEXITY 1353 01:04:48,628 --> 01:04:51,864 WHEREAS THE BRAIN THOUSANDS OF 1354 01:04:51,864 --> 01:04:54,300 CELL TYPES IT'S GOING TO BE A 1355 01:04:54,300 --> 01:04:59,839 REAL CHALLENGE AND I THINK MORE 1356 01:04:59,839 --> 01:05:02,108 CELLS, FEWER PERTURBATIONS IS 1357 01:05:02,108 --> 01:05:04,010 PROBABLY GOING TO BE THE ROUTE. 1358 01:05:04,010 --> 01:05:06,112 YOU NEED A NUMBER OF -- FOR 1359 01:05:06,112 --> 01:05:08,881 EVERY CELL STATE AND 1360 01:05:08,881 --> 01:05:11,651 NEIGHBORHOOD YOU WANT TO HAVE 1361 01:05:11,651 --> 01:05:13,386 EXAMPLES TO REALLY GET A ROBUST 1362 01:05:13,386 --> 01:05:14,854 SIGNAL OUT OF IT. 1363 01:05:14,854 --> 01:05:15,154 >> OKAY. 1364 01:05:15,154 --> 01:05:15,989 MAYBE ONE MORE QUESTION. 1365 01:05:15,989 --> 01:05:18,291 I DON'T WANT TO CUT THIS OFF 1366 01:05:18,291 --> 01:05:19,592 WE'LL HAVE A RECEPTION. 1367 01:05:19,592 --> 01:05:20,326 THIS WILL BE THE LAST QUESTION. 1368 01:05:20,326 --> 01:05:24,964 >> THAT WAS BASICALLY MY 1369 01:05:24,964 --> 01:05:25,398 QUESTION. 1370 01:05:25,398 --> 01:05:26,899 THERE'S A VARIETY OF DIFFERENT 1371 01:05:26,899 --> 01:05:29,669 TRANSCRIPTION STATES THAT MAY OR 1372 01:05:29,669 --> 01:05:30,903 MAY NOT REPRESENT DIFFERENT 1373 01:05:30,903 --> 01:05:32,405 FUNCTIONAL STATES WITHIN VARIOUS 1374 01:05:32,405 --> 01:05:34,507 TISSUES AND THE AFFECT OF A 1375 01:05:34,507 --> 01:05:37,810 SINGLE GENE PERTURBATION MAY 1376 01:05:37,810 --> 01:05:40,747 HAVE DRASTICALLY DIFFERENT 1377 01:05:40,747 --> 01:05:41,681 EFFECTS DEPENDING ON THE STATE 1378 01:05:41,681 --> 01:05:43,683 OF THE CELL IS THE ANSWER TO 1379 01:05:43,683 --> 01:05:46,285 JUST INCREASE THE NUMBER OF 1380 01:05:46,285 --> 01:05:50,289 CELLS YOU DO? 1381 01:05:50,289 --> 01:05:54,594 >> IT'S ABOUT EXPERIMENTAL 1382 01:05:54,594 --> 01:05:57,697 DESIGN AND TO MAKE COMPLEXITY 1383 01:05:57,697 --> 01:05:59,732 WORSE, THE LIVER CHANGES ITS 1384 01:05:59,732 --> 01:06:01,768 PHYSIOLOGY AND FUNCTION 1385 01:06:01,768 --> 01:06:05,138 DRAMATICALLY BASED ON RAPID 1386 01:06:05,138 --> 01:06:11,811 CHANGES LIKE FASTING AND HIGH 1387 01:06:11,811 --> 01:06:18,284 FAT DIETS AND SYNDROMES AND 1388 01:06:18,284 --> 01:06:20,753 FOLLOWING THE EFFECTS OF 1389 01:06:20,753 --> 01:06:22,422 PERTURBATION AND PHYSIOLOGY WILL 1390 01:06:22,422 --> 01:06:24,090 BE COMPLEX. 1391 01:06:24,090 --> 01:06:24,957 YOU HAVE THE COMPLEXITY. 1392 01:06:24,957 --> 01:06:30,296 ON THE OTHER HAND YOU GET 300 1393 01:06:30,296 --> 01:06:31,431 MILLION HEPATOCYTES FROM THE 1394 01:06:31,431 --> 01:06:31,698 LIVER. 1395 01:06:31,698 --> 01:06:34,267 YOU DON'T HAVE TO SPLIT YOUR 1396 01:06:34,267 --> 01:06:34,467 CELLS. 1397 01:06:34,467 --> 01:06:37,470 YOU FEED THE MOUSE CHOW AND KEEP 1398 01:06:37,470 --> 01:06:39,305 THEM IN A PHYSIOLOGIC STATE. 1399 01:06:39,305 --> 01:06:43,509 IN MANY WAYS THE SCALE YOU'RE 1400 01:06:43,509 --> 01:06:44,711 ABLE TO DO AT LEAST FROM 1401 01:06:44,711 --> 01:06:46,312 PRODUCING THE CELLS IS SIMPLER 1402 01:06:46,312 --> 01:06:56,756 FROM THE IN VIVO SETTING. 1403 01:07:15,541 --> 01:07:17,710 AND IT CAN GENERATE THE IMAGE 1404 01:07:17,710 --> 01:07:22,281 BUT IN THE END WE'RE NOT GOING 1405 01:07:22,281 --> 01:07:24,984 TO BE ABLE TO COLLECT ALL THE 1406 01:07:24,984 --> 01:07:32,859 DATA. 1407 01:07:32,859 --> 01:07:35,628 AND THERE'S MORE SOPHISTICATED 1408 01:07:35,628 --> 01:07:36,963 FOUNDATION MODELS AND HOW WE 1409 01:07:36,963 --> 01:07:41,134 FILL OUT THE SPACE THE REST CAN 1410 01:07:41,134 --> 01:07:42,568 BE IMPUTED ARE BIGGER, BROADER 1411 01:07:42,568 --> 01:07:48,741 CHALLENGE. 1412 01:07:48,741 --> 01:07:50,276 >> THERE'S A RECEPTION AND WE 1413 01:07:50,276 --> 01:07:51,577 MIGHT RUN OUT OF FOOD THIS TIME 1414 01:07:51,577 --> 01:07:52,979 BECAUSE OF THE SIZE OF THE 1415 01:07:52,979 --> 01:07:54,981 AUDIENCE SO THAT'S GREAT AND 1416 01:07:54,981 --> 01:07:58,417 THANK YOU ONCE AGAIN. 1417 01:07:58,417 --> 01:08:08,661 >> THANK YOU.