1 00:00:04,507 --> 00:00:08,010 WELCOME, 2 00:00:08,010 --> 00:00:08,277 EVERYBODY. 3 00:00:08,277 --> 00:00:10,780 IT'S A PLEASURE TO BE ABLE TO BE 4 00:00:10,780 --> 00:00:13,449 HERE TODAY INTRODUCING HENRY 5 00:00:13,449 --> 00:00:14,750 COLECRAFT FOR THE NEUROSCIENCE 6 00:00:14,750 --> 00:00:15,551 SEMINAR SERIES. 7 00:00:15,551 --> 00:00:16,619 HENRY'S BEEN A GOOD FRIEND FOR 8 00:00:16,619 --> 00:00:18,454 MANY YEARS AND WE WERE 9 00:00:18,454 --> 00:00:19,822 REMINISCING LAST NIGHT ABOUT HOW 10 00:00:19,822 --> 00:00:21,691 WE MET AND IT WAS A LITTLE WHILE 11 00:00:21,691 --> 00:00:23,326 AGO AND I'M NOT GOING TO SAY HOW 12 00:00:23,326 --> 00:00:25,728 LONG BECAUSE I DON'T WANT TO 13 00:00:25,728 --> 00:00:33,035 DATE MYSELF, BUT, HENRY HAS HAD 14 00:00:33,035 --> 00:00:35,238 AN INTERESTING WANDERING CAREER, 15 00:00:35,238 --> 00:00:37,874 HE WAS BORN IN GANNAA BUT THEN 16 00:00:37,874 --> 00:00:40,510 MOVED TO ENGLAND FOR HIS 17 00:00:40,510 --> 00:00:41,777 UNDERGRADUATE STUDIES AT KING'S 18 00:00:41,777 --> 00:00:43,513 COLLEGE IN LONDON BUT THEN, 19 00:00:43,513 --> 00:00:45,381 ACTUALLY AND I THINK THIS IS A 20 00:00:45,381 --> 00:00:47,250 FEATURE OF MANY OF THE BEST 21 00:00:47,250 --> 00:00:50,153 STORYTELLERS IN SCIENCE, HENRY 22 00:00:50,153 --> 00:00:51,988 ACTUALLY RETURNED TO GHANA AND 23 00:00:51,988 --> 00:00:53,289 TAUGHT HIGH SCHOOL FOR A FEW 24 00:00:53,289 --> 00:00:54,957 YEARS BEFORE HE WENT TO GRADUATE 25 00:00:54,957 --> 00:00:56,125 SCHOOL AND I THINK YOU SEE THAT 26 00:00:56,125 --> 00:00:57,426 IN THE QUALITY OF HIS TALK. 27 00:00:57,426 --> 00:00:59,162 IF YOU CAN TEACH SOMETHING TO 28 00:00:59,162 --> 00:01:01,230 HIGH SCHOOL KIDS, YOU CAN TEACH 29 00:01:01,230 --> 00:01:01,797 ANYBODY. 30 00:01:01,797 --> 00:01:03,866 AND THEN, HENRY CAME TO THE U.S. 31 00:01:03,866 --> 00:01:05,968 AFTER THAT TO GO TO -- AND DID 32 00:01:05,968 --> 00:01:07,370 HIS GRADUATE WORK AT THE 33 00:01:07,370 --> 00:01:11,574 UNIVERSITY OF ROUGH ATOM CHESTER 34 00:01:11,574 --> 00:01:14,443 WHERE HE FOUND COLD AND THEN 35 00:01:14,443 --> 00:01:16,546 WENT TO HOPKINS TO WORK WITH 36 00:01:16,546 --> 00:01:18,047 DAVID [INDISCERNIBLE] AS A POST 37 00:01:18,047 --> 00:01:19,448 DOC AND THAT'S WHERE HE MET UP 38 00:01:19,448 --> 00:01:20,750 WITH THE CALCIUM CHANNELS HE'S 39 00:01:20,750 --> 00:01:23,386 BEEN WORKING ON EVER SINCE. 40 00:01:23,386 --> 00:01:25,054 HE THEN BECAME FACULTY IN 41 00:01:25,054 --> 00:01:26,422 BIOENGINEERING DIRECTLY IN 42 00:01:26,422 --> 00:01:28,758 HOPKINS BEFORE BEING RECRUITED 43 00:01:28,758 --> 00:01:31,027 IN 2007, IS THAT RIGHT? 44 00:01:31,027 --> 00:01:34,430 TO COLUMBIA WHERE HE'S BEEN IN 45 00:01:34,430 --> 00:01:36,165 THE PHARMACOLOGY DEPARTMENT EVER 46 00:01:36,165 --> 00:01:38,534 SINCE WORKING ON JUST REALLY 47 00:01:38,534 --> 00:01:41,938 DOING BEAUTIFUL INSIGHTFUL WORK 48 00:01:41,938 --> 00:01:44,907 ON CALCIUM CHANNELS AND THE 49 00:01:44,907 --> 00:01:45,841 REGULATION, THEIR MECHANISM AND 50 00:01:45,841 --> 00:01:52,048 AS YOU'LL SEE TODAY, REALLY HAS 51 00:01:52,048 --> 00:01:53,282 ROBUSTLY MADE THE TRANSITION TO 52 00:01:53,282 --> 00:01:56,719 INVESTIGATING WHAT CAN WE DO 53 00:01:56,719 --> 00:01:58,354 PHARMACOLOGICALLY TO INTERVENE 54 00:01:58,354 --> 00:02:00,189 IN THE SYSTEM AND TRY TO DREET 55 00:02:00,189 --> 00:02:01,824 DISEASES OF IT AND THAT'S WHAT 56 00:02:01,824 --> 00:02:03,125 WE WILL HEAR ABOUT TODAY BUT I 57 00:02:03,125 --> 00:02:05,261 WILL NOT TRY TO ENUMERATE 58 00:02:05,261 --> 00:02:06,195 ACCOMPLISHMENTS OR ANYTHING LIKE 59 00:02:06,195 --> 00:02:08,297 THAT, IT'S EASY TO SEE HOW MUCH 60 00:02:08,297 --> 00:02:11,400 HENRY HAS DONE BUT I JUST WANT 61 00:02:11,400 --> 00:02:13,536 TO END BY TELLING YOU GUYS ABOUT 62 00:02:13,536 --> 00:02:16,606 A QUOTE THAT I FOUND IN AN 63 00:02:16,606 --> 00:02:18,674 INTERVIEW HENRY DID A FEW YEARS 64 00:02:18,674 --> 00:02:20,977 AGO THAT WAS -- HE WAS TALKING 65 00:02:20,977 --> 00:02:25,548 ABOUT HOW YOU GET INSIGHTS INTO 66 00:02:25,548 --> 00:02:27,617 THESE CHANNEL MECHANISMS, MODEL 67 00:02:27,617 --> 00:02:28,751 CITIZEN LEAK LECULAR MECHANISMS 68 00:02:28,751 --> 00:02:30,219 IN GENERAL AND YOU KNOW WHAT YOU 69 00:02:30,219 --> 00:02:32,655 HAVE DO IS MAKE YOURSELF REALLY 70 00:02:32,655 --> 00:02:33,589 SMALL AND PRETEND YOU'RE IN THE 71 00:02:33,589 --> 00:02:35,825 CELL WITH IT AND I THINK HENRY 72 00:02:35,825 --> 00:02:37,994 HAS BEEN EXTREMELY SUCCESSFUL AT 73 00:02:37,994 --> 00:02:39,095 MAKING HIMSELF SMALL BUT IN A 74 00:02:39,095 --> 00:02:42,231 BIG WAY. 75 00:02:42,231 --> 00:02:42,498 SO HENRY? 76 00:02:42,498 --> 00:02:47,536 [ APPLAUSE ] 77 00:02:47,536 --> 00:02:49,805 >> WELL, THANKS A LOT JOE. 78 00:02:49,805 --> 00:02:50,973 FIRST TIME ANYBODY'S TOLL ME I'M 79 00:02:50,973 --> 00:02:54,043 SMALL BUT I WILL TAKE THAT 80 00:02:54,043 --> 00:02:55,077 ANYTIME OF DAY. 81 00:02:55,077 --> 00:02:57,079 THANKS FOR THE INVITATION, IT'S 82 00:02:57,079 --> 00:02:58,714 REALLY WONDERFUL TO BE HERE, JOE 83 00:02:58,714 --> 00:03:02,218 WON'T SAY IT BUT I'VE KNOWN HIM 84 00:03:02,218 --> 00:03:05,054 FOR OVER 20 YEARS AND ALSO A FEW 85 00:03:05,054 --> 00:03:06,522 OTHER COLLEAGUESY HERE AND IT'S 86 00:03:06,522 --> 00:03:09,091 REALLY GREAT AND I'VE ALREADY 87 00:03:09,091 --> 00:03:10,826 LEARNED SOME REALLY GREAT 88 00:03:10,826 --> 00:03:12,194 SCIENCE THIS MORNING. 89 00:03:12,194 --> 00:03:14,597 SO I'M TYPICALLY TALKING TO 90 00:03:14,597 --> 00:03:15,798 QUITE A BROAD AUDIENCE SO I 91 00:03:15,798 --> 00:03:18,334 START MY TALKS OFF WITH 2 92 00:03:18,334 --> 00:03:21,837 METAPHORS, WHICH REALLY CAPTURE 93 00:03:21,837 --> 00:03:23,339 THE SCOPE AND UNIQUE CHALLENGES 94 00:03:23,339 --> 00:03:25,875 OF A PROBLEM I GOT INTERESTED IN 95 00:03:25,875 --> 00:03:28,678 THIS THE LAST 6 OR 7 YEARS OR 96 00:03:28,678 --> 00:03:28,844 SO. 97 00:03:28,844 --> 00:03:30,313 SO THE FIRST METAPHOR STARTs 98 00:03:30,313 --> 00:03:33,082 IN THE CHESS BOARD, SO IN CHESS 99 00:03:33,082 --> 00:03:34,717 1 WINS BY CAPTURING THE 100 00:03:34,717 --> 00:03:36,686 OPPONENTS OF THE KING, BUT IN 101 00:03:36,686 --> 00:03:38,521 ACTUALITY, THE MOST POWERFUL 102 00:03:38,521 --> 00:03:40,723 PIECE IS THE QUEEN AND THAT'S 103 00:03:40,723 --> 00:03:42,425 BECAUSE UNLIKE ANY OTHER PIECE, 104 00:03:42,425 --> 00:03:44,327 IT CAN MOVE UNCONSTRAINED ACROSS 105 00:03:44,327 --> 00:03:44,794 THE BOARD. 106 00:03:44,794 --> 00:03:46,896 SO ON THE CHESS BOARD OF 107 00:03:46,896 --> 00:03:47,830 BIOLOGICAL MOLECULES I WOULD 108 00:03:47,830 --> 00:03:49,699 LIKE TO POSIT AS 1 PROTEIN THAT 109 00:03:49,699 --> 00:03:51,801 IS PROBABLY THE MOST POWERFUL OF 110 00:03:51,801 --> 00:03:53,436 THEM ALL WITH RESPECT TO YOUR 111 00:03:53,436 --> 00:03:54,837 PARTICULAR PROTEIN THAT YOU'RE 112 00:03:54,837 --> 00:03:56,005 INTERESTED IN, THAT PROTEIN IS 1 113 00:03:56,005 --> 00:03:57,540 OF THE SMALLEST THERE IS, IT'S 114 00:03:57,540 --> 00:03:58,407 CALLED CRUCIATE LIGAMENT CIBOL 115 00:03:58,407 --> 00:03:58,741 WITTEN. 116 00:03:58,741 --> 00:04:01,344 AND THE REASON WHY I SAY 117 00:04:01,344 --> 00:04:03,245 UBIQUITIN IS THE QUEEN OF 118 00:04:03,245 --> 00:04:04,313 BIOLOGICAL MOLECULES IS THAT IT 119 00:04:04,313 --> 00:04:07,083 CONTROLS THE LIFE AND DEATH OF 120 00:04:07,083 --> 00:04:07,650 EVERY OTHER PROTEIN. 121 00:04:07,650 --> 00:04:10,186 SO ONCE A PROTEIN IS MADE, A 122 00:04:10,186 --> 00:04:18,060 STEADY STATE CONCENTRATION IS 123 00:04:18,060 --> 00:04:18,561 [INDISCERNIBLE] BETWEEN 124 00:04:18,561 --> 00:04:20,096 UBIQUITINNIZATION, AND THE MOLL 125 00:04:20,096 --> 00:04:23,599 CUE IS KEY WHICH DIRECTION THAT 126 00:04:23,599 --> 00:04:24,333 GOES. 127 00:04:24,333 --> 00:04:25,401 SO THERE ARE LITERALLY HUNDREDS 128 00:04:25,401 --> 00:04:26,469 OF THOUSANDS OF DISEASES WHERE 129 00:04:26,469 --> 00:04:28,537 WE HAVE TOO MUCH OR TOOLET OF A 130 00:04:28,537 --> 00:04:31,006 PROTEIN SO IN PRINCIPAL THE 131 00:04:31,006 --> 00:04:33,309 UBIQUITIN SYSTEM HAS A POTENTIAL 132 00:04:33,309 --> 00:04:34,477 SOLUTION TO ALL THOSE KINDS OF 133 00:04:34,477 --> 00:04:36,112 DISEASES BECAUSE IT SHOULD BE 134 00:04:36,112 --> 00:04:37,580 POSSIBLE TO REGULATE THE HALF 135 00:04:37,580 --> 00:04:40,349 LIFE OF A PROTEIN AND THAT'S A 136 00:04:40,349 --> 00:04:41,817 STEADY STATE CONCENTRATION 137 00:04:41,817 --> 00:04:43,719 BECAUSE IT'S A REGULATED AMOUNT 138 00:04:43,719 --> 00:04:45,488 THAT A PROTEIN HAS BUT THAT'S 139 00:04:45,488 --> 00:04:47,590 NOT REALLY THE WORLD WE LIVE IN. 140 00:04:47,590 --> 00:04:51,093 AND UBIQUITIN IS NOT BEING 141 00:04:51,093 --> 00:04:53,162 UTILIZED, IS NOT BEING USED TO 142 00:04:53,162 --> 00:04:54,597 TARGET TOO MANY DISEASES AT THE 143 00:04:54,597 --> 00:04:56,599 MOMENT AND THE REASON WHY HAS TO 144 00:04:56,599 --> 00:04:57,566 DID WITH MAY SECOND METAPHOR. 145 00:04:57,566 --> 00:05:00,035 SO CONSIDER A MAN WHO'S DYING OF 146 00:05:00,035 --> 00:05:02,104 THIRST IN THE MIDDLE OF THE 147 00:05:02,104 --> 00:05:03,739 OCEAN, THE IRONY IS THAT THE 148 00:05:03,739 --> 00:05:04,807 APPARENT SOLUTION TO HIS PROBLEM 149 00:05:04,807 --> 00:05:07,076 IS ALL AROUND HIM, IT'S WATER 150 00:05:07,076 --> 00:05:09,612 BUT THAL CASE, THE WATER IS A 151 00:05:09,612 --> 00:05:10,946 POISON CHALICE, IT HAS A LOT OF 152 00:05:10,946 --> 00:05:13,582 SALT IN IT, IF YOU DRANK IT 153 00:05:13,582 --> 00:05:14,950 WOULD SPEED UP DEHYDRATION IS 154 00:05:14,950 --> 00:05:15,151 DEATH. 155 00:05:15,151 --> 00:05:19,088 IN THE SAME WAY, UBIQUITIN WE 156 00:05:19,088 --> 00:05:20,756 THINK IS A POTENTIAL KEY TO MANY 157 00:05:20,756 --> 00:05:22,892 DISEASES BUT THE PROBLEM IS IT'S 158 00:05:22,892 --> 00:05:25,661 EVERYWHERE, EVERY SINGLE PROTEIN 159 00:05:25,661 --> 00:05:28,397 IS UTILIZING UBIQUITIN TO 160 00:05:28,397 --> 00:05:29,732 MAINTAIN HOMEOSTASIS. 161 00:05:29,732 --> 00:05:31,467 THE CHALLENGE BECOMES HOW DO YOU 162 00:05:31,467 --> 00:05:33,903 TARGET A PROTEIN AND SPECIFICITY 163 00:05:33,903 --> 00:05:35,504 WITHOUT DISTURBING THE WHOLE 164 00:05:35,504 --> 00:05:38,674 GLOBAL PROTEOME THAT WOULD THEN 165 00:05:38,674 --> 00:05:39,141 BE POTENTIALLY TOXIC? 166 00:05:39,141 --> 00:05:41,177 SO WHAT I WOULD LIKE TO DO, THIS 167 00:05:41,177 --> 00:05:45,147 OPPORTUNITY TO TALK ABOUT 3 168 00:05:45,147 --> 00:05:46,348 STORIES THAT REALLY POINT TO OUR 169 00:05:46,348 --> 00:05:48,451 WAYS TO DEAL WITH THAT ESSENTIAL 170 00:05:48,451 --> 00:05:52,621 CHALLENGE AND TRY TO MAKE THE 171 00:05:52,621 --> 00:05:55,391 UBIQUITIN SYSTEM IN A WAY MORE 172 00:05:55,391 --> 00:05:56,225 TRACTABLE FOR DESIGNER 173 00:05:56,225 --> 00:05:56,559 THERAPIES. 174 00:05:56,559 --> 00:05:58,627 AND THE PARTICULAR PLAY GROWBD I 175 00:05:58,627 --> 00:06:00,629 PLAY IN IS ION CHANNELS SO THE 176 00:06:00,629 --> 00:06:03,098 STORY OF THE ION CHANNEL IS 177 00:06:03,098 --> 00:06:04,800 CENTRIC, BUT THE UBIQUITIN 178 00:06:04,800 --> 00:06:05,634 SUGGESTS REGULATES EVERY PROTEIN 179 00:06:05,634 --> 00:06:07,203 AND THIS COULD BE APPLIED TO ANY 180 00:06:07,203 --> 00:06:09,939 PROTEIN AS L. SO THE FIRST STORY 181 00:06:09,939 --> 00:06:11,373 STARTS WITH THE REALIZATION WE 182 00:06:11,373 --> 00:06:12,641 HAVE, THERE ARE SEVERAL DISEASES 183 00:06:12,641 --> 00:06:15,845 THAT CAN BE CHARACTERIZED AS 184 00:06:15,845 --> 00:06:18,280 RARE, THEY'RE DEVASTATING THINGS 185 00:06:18,280 --> 00:06:20,649 LIKE ARRHYTHMIAS THAT HAVE 186 00:06:20,649 --> 00:06:22,184 SUDDEN CARDIAC DEATH, CYSTIC 187 00:06:22,184 --> 00:06:24,687 PHOBEROSIS THAT OCCURS IN THE 188 00:06:24,687 --> 00:06:26,322 LUNGS AND EPILEPSY AND THE LIST 189 00:06:26,322 --> 00:06:27,122 GOES ON. 190 00:06:27,122 --> 00:06:28,724 THESE ARE VERY DIFFERENT 191 00:06:28,724 --> 00:06:29,325 DISEASES AFFECTING DIFFERENT 192 00:06:29,325 --> 00:06:31,026 ORGAN SYSTEMS AND AT FIRST 193 00:06:31,026 --> 00:06:32,795 GLANCE IT DOESN'T SEEM LIKE 194 00:06:32,795 --> 00:06:33,863 THERE'S MUCH THAT CONNECTS THEM 195 00:06:33,863 --> 00:06:36,832 BUT THAT COULD BE POTENTIALLY 196 00:06:36,832 --> 00:06:38,033 COMMON SOLUTIONS FOR FURTHER, 197 00:06:38,033 --> 00:06:40,436 BUT WHAT WE REALIZE UPON DEEPER 198 00:06:40,436 --> 00:06:41,737 REFLECTION IS THAT MANY 199 00:06:41,737 --> 00:06:43,506 INSTANCES OF THESE DISEASES, ARE 200 00:06:43,506 --> 00:06:44,940 DUE TO MUTATIONS IN ION 201 00:06:44,940 --> 00:06:46,475 CHANNELS, THAT CAUSE A DECREASE 202 00:06:46,475 --> 00:06:47,977 IN THE FUNCTIONAL EXPRESSION, 203 00:06:47,977 --> 00:06:48,410 OKAY? 204 00:06:48,410 --> 00:06:50,513 SO FROM THAT PERSPECTIVE, THESE 205 00:06:50,513 --> 00:06:52,081 DISEASES HAVE THE SAME VERY 206 00:06:52,081 --> 00:06:55,584 DIFFERENT DISEASES HAVE THE SAME 207 00:06:55,584 --> 00:06:56,852 COMMON BIOLOGICAL PROBLEM AND 208 00:06:56,852 --> 00:06:58,821 THESE SIMPLE BIG IDEA THAT 209 00:06:58,821 --> 00:06:59,989 EMANATES FROM THAT IF WE COULD 210 00:06:59,989 --> 00:07:02,558 FIGURE OUT A WAY TO STABILIZE 211 00:07:02,558 --> 00:07:03,893 THESE PROTEINS YOU COULD HAVE 212 00:07:03,893 --> 00:07:05,294 SOMETHING THAT COULD BE 213 00:07:05,294 --> 00:07:08,697 GENERALIZABLE TO MANY DEFINITE 214 00:07:08,697 --> 00:07:08,964 DISEASES. 215 00:07:08,964 --> 00:07:10,499 SO THE ION CHANNELS THAT WE'RE 216 00:07:10,499 --> 00:07:12,568 INTERESTED IN, WORK AT THE CELL 217 00:07:12,568 --> 00:07:15,037 SURFACE, AND THE SURFACE DENSITY 218 00:07:15,037 --> 00:07:16,205 IS REALLY REGULATED BY 3 219 00:07:16,205 --> 00:07:17,172 PROCESSES ISSUES RATE OF 220 00:07:17,172 --> 00:07:19,475 DELIVERY, RATE OF REMOVAL AND 221 00:07:19,475 --> 00:07:21,076 RATE OF DEGRADATION OF THE OTHER 222 00:07:21,076 --> 00:07:26,415 PROTEIN AND FOR WILD-TYPE 223 00:07:26,415 --> 00:07:27,349 PROTEINS THE UBIQUITIN TAGGING 224 00:07:27,349 --> 00:07:34,723 ON TO THE PROTEIN DICTATES WHERE 225 00:07:34,723 --> 00:07:35,591 THE EQUILIBRIUM RELIES, SO IF 226 00:07:35,591 --> 00:07:37,660 YOU HAVE THIS, YOU GET THIS TO 227 00:07:37,660 --> 00:07:37,860 FIRST. 228 00:07:37,860 --> 00:07:39,728 SO THE REALIZE WE HAD WAS THAT 229 00:07:39,728 --> 00:07:41,564 THERE WERE MANY MUTANT ION 230 00:07:41,564 --> 00:07:44,133 CHANNELS THAT CAUSE THESE LIFE 231 00:07:44,133 --> 00:07:45,301 THREATENING DISEASES, WHERE THE 232 00:07:45,301 --> 00:07:47,136 MUTATION CAUSED THE CHANNEL TO 233 00:07:47,136 --> 00:07:48,370 BE TRAPPED INSIDE THE CELL, SO 234 00:07:48,370 --> 00:07:50,005 IN THAT WAY, IT'S VERY SIMILAR 235 00:07:50,005 --> 00:07:52,241 TO A WILD-TYPE CHANNEL WITH A 236 00:07:52,241 --> 00:07:54,543 LOT OF UBIQUITIN ON IT, SO THE 237 00:07:54,543 --> 00:07:55,411 SIMPLE HYPOTHESIS WE HAD THEN 238 00:07:55,411 --> 00:07:58,280 WAS THAT WE COULD POTENTIALLY 239 00:07:58,280 --> 00:07:59,348 STRIP UBIQUITIN FROM MUTANT 240 00:07:59,348 --> 00:08:01,450 CHANNELS AND CAUSE THEM TO BE 241 00:08:01,450 --> 00:08:03,519 STABILIZED AND GET TO THE CELL 242 00:08:03,519 --> 00:08:06,722 SURFACE, AND POETIC TEBTIA WILY 243 00:08:06,722 --> 00:08:08,457 RESCUE FUNCTION. 244 00:08:08,457 --> 00:08:09,558 -- POTENTIALLY RESCUE FUNCTION. 245 00:08:09,558 --> 00:08:11,594 I'VE ALREADY TOLD YOU THE 246 00:08:11,594 --> 00:08:12,761 ESSENTIAL CHALLENGE TO DO THAT, 247 00:08:12,761 --> 00:08:15,064 YOU HAVE TO DO THAT AT 248 00:08:15,064 --> 00:08:16,599 SPECIFICITY SO YOU DON'T HAVE 249 00:08:16,599 --> 00:08:17,366 TOXICITY ASSOCIATED WITH THAT. 250 00:08:17,366 --> 00:08:18,434 SO THE WAY WE SOLVE THAT PROBLEM 251 00:08:18,434 --> 00:08:21,003 IS TO COME UP WITH A NEW TYPE OF 252 00:08:21,003 --> 00:08:25,007 MOLECULE, WE CALL THESE 253 00:08:25,007 --> 00:08:26,475 ENGINEERED DECIBOL WITTEN ACES 254 00:08:26,475 --> 00:08:27,576 AND THEY'RE BI VALENTINEDENT 255 00:08:27,576 --> 00:08:31,347 MOLECULES THAT HAVE A TARGET 256 00:08:31,347 --> 00:08:33,782 MOEITY, A NANO BODY THAT ALLOWS 257 00:08:33,782 --> 00:08:38,387 TO YOU HAVE SPECIFIC CELLS OF 258 00:08:38,387 --> 00:08:40,990 CHOICE. 259 00:08:40,990 --> 00:08:47,763 WE FELL THAT CATALYTIC DOMAINS 260 00:08:47,763 --> 00:08:51,667 UBIQUITIN AS OR DUBS, SO WITH 261 00:08:51,667 --> 00:08:53,602 THAL SIGN FEATURE YOU HAVE 262 00:08:53,602 --> 00:08:54,937 SPECIFICITY BY THE NANO BODY, 263 00:08:54,937 --> 00:08:56,105 THE ISS IS MODULAR, YOU SHOULD 264 00:08:56,105 --> 00:08:58,073 BE ABLE TO SWAP OUT NANO BODIES 265 00:08:58,073 --> 00:08:59,041 FOR DIFFERENT TARGETS AND WE 266 00:08:59,041 --> 00:09:00,442 EXPECTED TO BE HIGHLY POTENT 267 00:09:00,442 --> 00:09:03,412 BECAUSE OF THE CATALYTIC 268 00:09:03,412 --> 00:09:04,346 ACTIVITY OF THE ENZYME. 269 00:09:04,346 --> 00:09:06,081 SO THIS IS THE CARTOON CONCEPT, 270 00:09:06,081 --> 00:09:11,754 COULD WE GET THIS TO WORK IN 271 00:09:11,754 --> 00:09:12,521 CELLS? 272 00:09:12,521 --> 00:09:15,024 SO SCOTT CANON IS THE STUDENT 273 00:09:15,024 --> 00:09:17,092 WHO DEVELOPED THE IDEA IN THE 274 00:09:17,092 --> 00:09:20,396 LAB, TO SEE THE PROOF OF CONCEPT 275 00:09:20,396 --> 00:09:22,164 OF THE TECHNOLOGY, HE CAME UP 276 00:09:22,164 --> 00:09:24,233 WITH A BIOCHEMICAL ASSAY TO TEST 277 00:09:24,233 --> 00:09:24,433 THAT. 278 00:09:24,433 --> 00:09:32,474 SO IN HEX CELLS WE CAN EXPRESS A 279 00:09:32,474 --> 00:09:34,677 QKKCM ATTACHED TO YFP AND THIS 280 00:09:34,677 --> 00:09:37,112 IS A POTASSIUM CHANNEL AND SO IF 281 00:09:37,112 --> 00:09:39,782 YOU CO-EXPRESS IT THE CHANNEL 282 00:09:39,782 --> 00:09:41,650 GETS UBIQUITINNATED AND GETS 283 00:09:41,650 --> 00:09:43,052 ENDOSIGNIFYITOSED AND A LOT OF 284 00:09:43,052 --> 00:09:44,353 IT GETS DEGRADED EMPLOY SO NOW 285 00:09:44,353 --> 00:09:47,556 WHAT WE'RE ABLE DO THEN IS WE 286 00:09:47,556 --> 00:09:50,225 COULD THEN CO-EXPRESS AN NDUB, A 287 00:09:50,225 --> 00:09:52,528 NANO BODY THAT TARGETS THE YFP, 288 00:09:52,528 --> 00:09:57,066 BRINGS THE DUB TO THE TARGET 289 00:09:57,066 --> 00:09:58,467 DECIBOL WITTENATEED AND WE COULD 290 00:09:58,467 --> 00:10:02,905 GET A REVERSAL OF THAT PROCESS. 291 00:10:02,905 --> 00:10:04,039 SO HERE'S BIOCHEMICAL PROOF OF 292 00:10:04,039 --> 00:10:04,540 THE CONCEPT. 293 00:10:04,540 --> 00:10:06,842 WE EXPRESS THE CHANNEL Q1, AND 294 00:10:06,842 --> 00:10:09,044 THE CONTROL CONDITIONS WITH THE 295 00:10:09,044 --> 00:10:09,578 NAKED NANO BODY. 296 00:10:09,578 --> 00:10:10,913 YOU PULL DOWN THE CHANNEL AND 297 00:10:10,913 --> 00:10:13,515 PROBE IT, YOU FIND 4 BANDS 298 00:10:13,515 --> 00:10:15,584 REPRESENTING THE MONOMER, 299 00:10:15,584 --> 00:10:16,385 TRIEMER, TETRAMERIC SPECIES OF 300 00:10:16,385 --> 00:10:17,519 THE CHANNEL. 301 00:10:17,519 --> 00:10:20,355 IF YOU CO EXPRESS NET FOIL THE 302 00:10:20,355 --> 00:10:21,957 CHANNEL, THE DENSITY OF THOSE 303 00:10:21,957 --> 00:10:24,693 GOES DOWN AS THE CHANNEL IS 304 00:10:24,693 --> 00:10:25,327 ENDOSIGNIFYITOSED AND DEGRADED. 305 00:10:25,327 --> 00:10:27,996 SO WHAT WE CAN THEN DO IS STRIP 306 00:10:27,996 --> 00:10:30,399 THIS BLOT AND PROBE FOR 307 00:10:30,399 --> 00:10:32,101 UBIQUITIN AS YOU WOULD EXPECT, 308 00:10:32,101 --> 00:10:33,035 UNDER BASE LINE CONDITIONS 309 00:10:33,035 --> 00:10:34,837 THERE'S A BIT MORE, AND THAT 310 00:10:34,837 --> 00:10:37,973 GETS MORE IN THE E-3 LIGASE, SO 311 00:10:37,973 --> 00:10:40,242 THIS ARM OF THE PATHWAY IS 312 00:10:40,242 --> 00:10:42,878 ESTABLISHED AND NOW WE CAN HAVE 313 00:10:42,878 --> 00:10:44,146 AN NDUB AND SEE WHAT HAPPENS. 314 00:10:44,146 --> 00:10:46,014 WHEN WE DO THAT YOU RESTORE THE 315 00:10:46,014 --> 00:10:49,251 PROTEIN EXPRESSIONS AND THAT'S 316 00:10:49,251 --> 00:10:51,587 ACCOMPANIED BY THE UBIQUITIN 317 00:10:51,587 --> 00:10:51,854 PROTEIN. 318 00:10:51,854 --> 00:10:53,021 SO BIOCHEMICALLY THIS APPEARS TO 319 00:10:53,021 --> 00:10:53,722 FUNCTION. 320 00:10:53,722 --> 00:10:58,127 WHAT WE NEEDED TO KNOW THEN WAS 321 00:10:58,127 --> 00:10:59,361 WHETHER THESE DEUBIQUITINNATED 322 00:10:59,361 --> 00:11:02,064 CHANNELS GET TO THE CELL SURFACE 323 00:11:02,064 --> 00:11:03,031 ARE FUNCTIONAL? 324 00:11:03,031 --> 00:11:05,734 TO DO THAT WE DEVELOPED A FLOW 325 00:11:05,734 --> 00:11:07,870 CYTOMETRY ASSAY, THAT ALLOWS TO 326 00:11:07,870 --> 00:11:10,072 US TO SIMULTANEOUSLY LOOK THEA 327 00:11:10,072 --> 00:11:11,807 SURFACE CHANNELS USING A 328 00:11:11,807 --> 00:11:13,108 FLUORESCENT TOXIN THAT BINDS IN 329 00:11:13,108 --> 00:11:14,309 THE EPITOPE THAT'S ENGINEERED 330 00:11:14,309 --> 00:11:16,678 INTO THE SURFACE OF THE ION 331 00:11:16,678 --> 00:11:19,548 CHANNEL, AND THEN WE HAVE THE 332 00:11:19,548 --> 00:11:22,751 YFP TO GIVE US A MEASURE OF THE 333 00:11:22,751 --> 00:11:23,719 TOTAL PROTEIN EXPRESSION, OKAY. 334 00:11:23,719 --> 00:11:25,954 SO WHEN YOU APPLY THAT TO FLOW 335 00:11:25,954 --> 00:11:27,256 CYTOMETRY, WHAT YOU CAN SEE IN 336 00:11:27,256 --> 00:11:29,124 THE CONTROL CONDITIONS IS THAT 337 00:11:29,124 --> 00:11:32,461 YFP POSITIVE CELLS HAVE A LOT OF 338 00:11:32,461 --> 00:11:33,996 REGINALSOT CELL SURFACE, THE 339 00:11:33,996 --> 00:11:37,533 CHANNELS ARE RIGHT AT THE 340 00:11:37,533 --> 00:11:38,233 CHANNEL MEMBRANE, THE DENSITY 341 00:11:38,233 --> 00:11:41,103 GOES DOWN AS WELL AS YFP 342 00:11:41,103 --> 00:11:43,338 INTENSITY GOES DOWN AS PROTEIN 343 00:11:43,338 --> 00:11:44,373 IS DEGRADED AND 344 00:11:44,373 --> 00:11:44,807 ENDOSIGNIFYITOSED. 345 00:11:44,807 --> 00:11:47,142 SO NOW WHEN WE ADD THE NDUB, WE 346 00:11:47,142 --> 00:11:49,278 FIEBD A RESCUE OF BOTH THE 347 00:11:49,278 --> 00:11:50,979 SURFACE DENSITY AND THE TOTAL 348 00:11:50,979 --> 00:11:51,480 PROTEIN EXPRESSION. 349 00:11:51,480 --> 00:11:54,550 SO PROEN TOOS DO MAKE IT TO THE 350 00:11:54,550 --> 00:11:55,717 CELL SURFACE, TO DETERMINE 351 00:11:55,717 --> 00:11:56,451 WHETHER THEY'RE FUNCTIONAL WE 352 00:11:56,451 --> 00:11:59,521 USE THE PATCH CLAMP, WHOLESALE 353 00:11:59,521 --> 00:12:02,558 PATCH CLAMP TECHNIQUE. 354 00:12:02,558 --> 00:12:04,526 CONTROL CHANNELS EXPRESS ROBUST 355 00:12:04,526 --> 00:12:05,727 OUTWARD OCCURRENCE, CO EXPRESSED 356 00:12:05,727 --> 00:12:07,329 BEFORE ALL THE CHANNELS ARE GONE 357 00:12:07,329 --> 00:12:10,933 AND WHEN YOU ADD THE ENDUB, YOU 358 00:12:10,933 --> 00:12:14,269 RESCUE THOSE CURRENTS, SO THE 359 00:12:14,269 --> 00:12:15,237 DECIBOL WITTENATEED CHANNELS ARE 360 00:12:15,237 --> 00:12:15,437 FINE. 361 00:12:15,437 --> 00:12:18,974 DID A NUMBER OF CONTROL 362 00:12:18,974 --> 00:12:20,242 EXPERIMENTS TO FIND OUT THAT 363 00:12:20,242 --> 00:12:23,712 THIS REQUIRED IT TO BE TARGETED 364 00:12:23,712 --> 00:12:26,081 TO THE END OF CHOICE AND THOSE 365 00:12:26,081 --> 00:12:27,316 ARE TARGETABLE EFFECT CHES. 366 00:12:27,316 --> 00:12:28,483 SO WITH THAT ESTABLISHED WE ARE 367 00:12:28,483 --> 00:12:31,186 READY TO APPLY THE SYSTEM TO A 368 00:12:31,186 --> 00:12:33,956 RARE DEC. 369 00:12:33,956 --> 00:12:37,893 THE FIRST 1 WE LOOKED AT IS 370 00:12:37,893 --> 00:12:43,165 [INDISCERNIBLE] SYNDROME, 1 IN 2 371 00:12:43,165 --> 00:12:43,999 BIRTHS ASHING THE 400,000 SUDDEN 372 00:12:43,999 --> 00:12:45,701 DEATH KNOW DEATH CASES IN THE 373 00:12:45,701 --> 00:12:48,637 U.S. AND MUCH MORE BEYOND THE 374 00:12:48,637 --> 00:12:48,871 U.S. 375 00:12:48,871 --> 00:12:50,305 SO LOSS OF FUNCTION MUTATIONS IN 376 00:12:50,305 --> 00:12:52,407 THE CHANNEL HAVE BEEN TALKING 377 00:12:52,407 --> 00:12:54,009 ABOUT CALCIUM K1, COME FROM 378 00:12:54,009 --> 00:12:57,479 THERE 35% OF THESE LQTS CASES, 379 00:12:57,479 --> 00:13:00,782 SO THIS CHANNEL GIVES RISE TO 380 00:13:00,782 --> 00:13:02,050 SLOWLY ACTIVATED POTASSIUM 381 00:13:02,050 --> 00:13:04,353 CURRENT THAT'S REALLY IMPORTANT 382 00:13:04,353 --> 00:13:06,421 FOR THE REPOLARIZATION PHASE OF 383 00:13:06,421 --> 00:13:07,289 THE CARDIAC ACTION POTENTIAL SO 384 00:13:07,289 --> 00:13:09,892 WHEN YOU HAVE LESS OF IT, YOU 385 00:13:09,892 --> 00:13:11,727 HAVE A PROLONGATION OF THE 386 00:13:11,727 --> 00:13:13,896 ACTION PROTENTIAL THAT SHOWS UP 387 00:13:13,896 --> 00:13:16,798 AS A PROLONGATION OF THE LQTS 388 00:13:16,798 --> 00:13:17,733 CARDIOGRAM AND THE BAD THING 389 00:13:17,733 --> 00:13:19,902 ABOUT IT IS THAT PEOPLE WHO HAVE 390 00:13:19,902 --> 00:13:23,205 ARE PREDISPOSED TO GETTING 391 00:13:23,205 --> 00:13:24,806 LETHERAL VENTRICULAR ARRHYTHMIA 392 00:13:24,806 --> 00:13:26,308 WHEN THEY TWERICIZE, SO IT TURNS 393 00:13:26,308 --> 00:13:32,080 OUT MOST OF THOSE MUTATIONS AND 394 00:13:32,080 --> 00:13:33,815 YOU CAN USE ENDUBS TO STRIP THEM 395 00:13:33,815 --> 00:13:34,783 FROM THESE CHANNELS AND CAUSE 396 00:13:34,783 --> 00:13:36,051 THEM TO BE RESCUED. 397 00:13:36,051 --> 00:13:38,487 SO WITH THAT IN MIND, WE MADE A 398 00:13:38,487 --> 00:13:42,624 NUMBER OF PATIENT MUTATIONS AND 399 00:13:42,624 --> 00:13:46,795 USING THE ASSAY TO LOOK FOR 400 00:13:46,795 --> 00:13:48,730 SURFACE TRAFFICKING, WE ASSESS 401 00:13:48,730 --> 00:13:50,599 THE IMPACT OF THOSE MUTATIONSOT 402 00:13:50,599 --> 00:13:53,068 SURFACE SENSITY OF THE CHANNELS, 403 00:13:53,068 --> 00:13:54,803 SO THE DOTTED LINE REPRESENTS 404 00:13:54,803 --> 00:13:58,440 WHAT YOU GET WITH WILD-TYPE MANY 405 00:13:58,440 --> 00:13:59,942 OF THOSE CHANNELS CO EXPRESS 406 00:13:59,942 --> 00:14:01,343 WITH THE NCI--AND NANO BODY, 407 00:14:01,343 --> 00:14:04,246 HAVE A DEFICIENCY IN TRAFFIC TO 408 00:14:04,246 --> 00:14:05,013 THE CELL SURFACE. 409 00:14:05,013 --> 00:14:08,250 HOWEVER, WHEN WE ADD THE ENDUB, 410 00:14:08,250 --> 00:14:11,486 WE ARE ABLE TO RESCUE MANY OF 411 00:14:11,486 --> 00:14:12,821 THEM, NOT ALL OF THEM BUT A 412 00:14:12,821 --> 00:14:14,790 LARGE FRACTION OF THEM TO 413 00:14:14,790 --> 00:14:16,658 ESSENTIALLY WILD-TYPE LEVELS OF 414 00:14:16,658 --> 00:14:17,492 THE SURFACE DENSITY. 415 00:14:17,492 --> 00:14:19,895 IF WE -- AND YOU CAN LOOK TO SEE 416 00:14:19,895 --> 00:14:21,396 THAT PICTORIALLY HERE, SO HERE 417 00:14:21,396 --> 00:14:23,265 WE ARE STAINING THE SURFACE 418 00:14:23,265 --> 00:14:24,700 CHANNELS WITH THE FLUORESCENT 419 00:14:24,700 --> 00:14:26,501 TOXIN, IN THIS MUTANT, THAT 420 00:14:26,501 --> 00:14:28,270 STANDARD IS GONE, BUT YOU CO 421 00:14:28,270 --> 00:14:29,204 EXPRESS WITH THE ENDUB AND YOU 422 00:14:29,204 --> 00:14:30,205 GET IT RIGHT BACK. 423 00:14:30,205 --> 00:14:31,807 ALL RIGHT, SO WE'VE BEEN ABLE TO 424 00:14:31,807 --> 00:14:34,076 RESCUE, A CHANNEL THAT'S TRAPPED 425 00:14:34,076 --> 00:14:36,378 INSIDE THE CELL, THE NEXT 426 00:14:36,378 --> 00:14:37,879 CHALLENGE IS WHETHER THOSE 427 00:14:37,879 --> 00:14:39,848 CHANNELS WERE FUNCTIONAL AS THEY 428 00:14:39,848 --> 00:14:42,117 GET TO THE CELL SURFACE. 429 00:14:42,117 --> 00:14:42,851 SO USING ELECTROPHYSIOLOGY HERE 430 00:14:42,851 --> 00:14:44,786 IS WHAT YOU GET WITH THE 431 00:14:44,786 --> 00:14:45,354 WILD-TYPE CURRENT AGAIN. 432 00:14:45,354 --> 00:14:47,823 HERE'S 1 OF THOSE MUTANTS ARE 433 00:14:47,823 --> 00:14:49,424 591 H, YOU CAN SEE THAT THE 434 00:14:49,424 --> 00:14:54,062 CURRENT IS ALMOST TOTALLY 435 00:14:54,062 --> 00:14:55,931 ABOLISHED, ML277 IS A SMALL 436 00:14:55,931 --> 00:14:57,399 MOLECULE AGONIST THAT IF YOU 437 00:14:57,399 --> 00:14:59,568 HEAD ADD THE IN YOUITATION, YOU 438 00:14:59,568 --> 00:15:00,435 INDICATE THAT SOME OF THE 439 00:15:00,435 --> 00:15:02,537 CHANNELS GET TO THE SURFACE, SO 440 00:15:02,537 --> 00:15:04,206 HERE'S WHAT HAPPENS WHEN WE CO 441 00:15:04,206 --> 00:15:05,841 EXPRESS THE MUTANT, AND YOU GET 442 00:15:05,841 --> 00:15:07,275 A 50% RESCUE OF THE CURRENT. 443 00:15:07,275 --> 00:15:07,976 THIS IS ACTUALLY QUITE 444 00:15:07,976 --> 00:15:09,344 INTERESTING BECAUSE IF YOU WE 445 00:15:09,344 --> 00:15:12,280 LEAK AT THE SURFACE DENSITY, IT 446 00:15:12,280 --> 00:15:14,049 DOES RESCUE A HUNDRED PERCENT IS 447 00:15:14,049 --> 00:15:17,119 SO WHAT THIS VEALS IN THE 448 00:15:17,119 --> 00:15:18,653 TRAPPED DEFICIENCY, THE MUTATION 449 00:15:18,653 --> 00:15:19,821 CAUSES A [INDISCERNIBLE], AND 450 00:15:19,821 --> 00:15:20,989 DEFICIT AS L. HOWEVER, NOW ON 451 00:15:20,989 --> 00:15:23,925 TOP OF THAT IF WE ADD THE SMALL 452 00:15:23,925 --> 00:15:25,761 MOLECULE AGONIST, YOU CAN GET A 453 00:15:25,761 --> 00:15:26,595 SUPER RECUSE OF THE 454 00:15:26,595 --> 00:15:27,295 [INDISCERNIBLE], AND YOU CAN 455 00:15:27,295 --> 00:15:29,531 ASHES TACH THE SMALL MOLECULE TO 456 00:15:29,531 --> 00:15:31,366 GET WILD-TYPE CURRENTS BACK. 457 00:15:31,366 --> 00:15:32,567 SO THIS WAS ENCOURAGING. 458 00:15:32,567 --> 00:15:35,170 IN ORDER TO SEE WHETHER THIS 459 00:15:35,170 --> 00:15:38,507 COULD HAVE A POTENTIAL 460 00:15:38,507 --> 00:15:39,141 THERAPEUTIC POTENTIAL, WAS IT 461 00:15:39,141 --> 00:15:40,609 IMPORTANT TO GO FROM THESE MODEL 462 00:15:40,609 --> 00:15:43,278 SYSTEM OF HEX CELLS INTO THE 463 00:15:43,278 --> 00:15:45,147 HEART ITSELF. 464 00:15:45,147 --> 00:15:47,082 WE COULDN'T TEST THE EFFICACY OF 465 00:15:47,082 --> 00:15:49,084 THESE IN MICE, IN MICE BECAUSE 466 00:15:49,084 --> 00:15:50,519 THEY ARE ACTUALLY POTENTIALS ARE 467 00:15:50,519 --> 00:15:53,121 TOO SHORT, SO THIS POTASSIUM 468 00:15:53,121 --> 00:15:54,056 CURRENT ACTUALLY DOESN'T PLAY A 469 00:15:54,056 --> 00:15:55,957 ROLE IN THE REPOSEARIZATION OF 470 00:15:55,957 --> 00:15:57,826 THAT, SO THE MODEL WE CAME UP 471 00:15:57,826 --> 00:16:01,196 WITH WITH WAS TO TAKE A GINNY 472 00:16:01,196 --> 00:16:03,398 PIG MONOCYTE WHICH DOES EXPRESS 473 00:16:03,398 --> 00:16:07,269 ENDOGENOUS K1 AND WE CAN USE AN 474 00:16:07,269 --> 00:16:08,603 ADENOVIRUS TO EXPRESS MORE ON 475 00:16:08,603 --> 00:16:10,806 THAT, IT COMBINES WITH THE 476 00:16:10,806 --> 00:16:11,139 ENDOGENOUS 1. 477 00:16:11,139 --> 00:16:13,341 IF YOU DO A ROUND PROTOCOL ON 478 00:16:13,341 --> 00:16:14,609 THESE GINNY PIG MICE, YOU GET 479 00:16:14,609 --> 00:16:18,380 THIS WAVE FORM OF CURRENT AND 480 00:16:18,380 --> 00:16:19,681 THE ICS, CURRENT, CONTRIBUTE TO 481 00:16:19,681 --> 00:16:21,349 THE POTASSIUM CHANNEL IS THIS 482 00:16:21,349 --> 00:16:24,786 LATE PHASE OF THE WAVE FROM -- 483 00:16:24,786 --> 00:16:26,688 AND THIS THE NORMAL LOOKING 484 00:16:26,688 --> 00:16:28,390 POTENTIAL YOU CAN RECORD IN 485 00:16:28,390 --> 00:16:28,890 THESE MICE. 486 00:16:28,890 --> 00:16:31,893 SO THEN WHAT WE CAN DO THEN IS 487 00:16:31,893 --> 00:16:35,497 TO USE AN ADENOVIRUS TO EXPRESS 488 00:16:35,497 --> 00:16:38,366 THE MUTATION, AND THIS MIMICS 489 00:16:38,366 --> 00:16:39,534 THE AUTOSOMAL DOMINANT NATURE OF 490 00:16:39,534 --> 00:16:42,237 THIS DEC IN HUMANS SO THE MUTANT 491 00:16:42,237 --> 00:16:44,005 WILL MIX WITH ENDOGENOUS 1 AND 492 00:16:44,005 --> 00:16:45,540 IF YOU LOOK AT CURRENT, CAN YOU 493 00:16:45,540 --> 00:16:47,642 SEE IT'S ABOUT HALF, SO IT HAS A 494 00:16:47,642 --> 00:16:50,479 DOMINANT NEGATIVE EFFECT ON THE 495 00:16:50,479 --> 00:16:50,712 CURRENT. 496 00:16:50,712 --> 00:16:52,013 SO WHAT -- AND THE RESULT OF 497 00:16:52,013 --> 00:16:54,549 THAT IS THIS CLASSICAL SIGNATURE 498 00:16:54,549 --> 00:16:56,384 OF LONG QT SYNDROME, 499 00:16:56,384 --> 00:16:58,120 PROLONGATION OF THE ACTION 500 00:16:58,120 --> 00:16:59,888 POTENTIAL IN THIS SYSTEM, AND SO 501 00:16:59,888 --> 00:17:01,923 WE CAN ADD AN ENDUB TO THIS AND 502 00:17:01,923 --> 00:17:07,395 SEE IF WE CAN RESCUE THE CURRENT 503 00:17:07,395 --> 00:17:11,299 POETIC TEBTIAL AND THE ANSWER IS 504 00:17:11,299 --> 00:17:11,800 YES. 505 00:17:11,800 --> 00:17:14,102 SO THIS RESCUES THE OUTWARD 506 00:17:14,102 --> 00:17:15,937 CURRENT AND NORMALIZES THE 507 00:17:15,937 --> 00:17:16,771 POTENTIAL DURATION, SO PREDICT 508 00:17:16,771 --> 00:17:18,540 THAT IF YOU DID THIS IN THE 509 00:17:18,540 --> 00:17:20,675 HUMAN THAT THAT WOULD BE 510 00:17:20,675 --> 00:17:23,645 BENEFICIAL FOR THE LONG QT 511 00:17:23,645 --> 00:17:23,912 SYNDROME. 512 00:17:23,912 --> 00:17:26,314 SO 1 OF THE THINGS THAT WE WERE 513 00:17:26,314 --> 00:17:27,249 MOST EXCITED ABOUT, THE 514 00:17:27,249 --> 00:17:30,085 TECHNOLOGY IS THE IDEA THAT IT 515 00:17:30,085 --> 00:17:31,887 SHOULD BE GENERALLY APPLICABLE 516 00:17:31,887 --> 00:17:34,322 TO BASICALLY ANY PROTEIN. 517 00:17:34,322 --> 00:17:36,725 AND SO TO SEE WHETHER THAT'S THE 518 00:17:36,725 --> 00:17:39,494 CASE FOR THE ION CHANNELS, WE 519 00:17:39,494 --> 00:17:42,130 LOOKED AT ANOTHER ION CHANNEL 520 00:17:42,130 --> 00:17:43,765 DISEASE, THIS CASE, 521 00:17:43,765 --> 00:17:45,700 CYSTIC FIBROSIS, FOR JUST REALLY 522 00:17:45,700 --> 00:17:47,702 THE POSTER CHILD OF ION CHANNEL 523 00:17:47,702 --> 00:17:49,604 MUTATION CAUSING A DEC. 524 00:17:49,604 --> 00:17:51,373 SO IN THIS CASE, THE MUTATION 525 00:17:51,373 --> 00:17:53,141 OCCURS IN THE CHLORIDE CHANNEL, 526 00:17:53,141 --> 00:17:55,544 THAT LINES THE AIR WAY 527 00:17:55,544 --> 00:17:57,312 EPITHELIAL CELLS, IN THE AIR 528 00:17:57,312 --> 00:17:58,947 WAYS, THIS CHANNEL NORMALLY 529 00:17:58,947 --> 00:18:04,152 ALLOWS CHLORIDE TO COME OUT OF 530 00:18:04,152 --> 00:18:05,887 THE EPITHELIAL CELLS AND THAT 531 00:18:05,887 --> 00:18:09,291 DRAGS WATER AND THAT'S VERY 532 00:18:09,291 --> 00:18:10,358 IMPORTANT FOR HYDRATING THE 533 00:18:10,358 --> 00:18:11,993 AIRWAYS, WHEN YOU HAVE A LOSS OF 534 00:18:11,993 --> 00:18:13,528 FUNCTION MUTATION, YOU GET LESS 535 00:18:13,528 --> 00:18:15,130 WATER COMING UP SO YOU GET A 536 00:18:15,130 --> 00:18:18,466 THICK BUILD UP OF MUCUS IN THE 537 00:18:18,466 --> 00:18:21,303 AIRWAYS AND THAT TRAPS BACTERIA, 538 00:18:21,303 --> 00:18:22,671 CAUSES ROUNDS OF INFECTION AND 539 00:18:22,671 --> 00:18:24,773 SO FORTH THAT ULTIMATELY DESTROY 540 00:18:24,773 --> 00:18:25,073 THE LUNGS. 541 00:18:25,073 --> 00:18:27,108 SO PEOPLE SUFFERING FROM THIS 542 00:18:27,108 --> 00:18:28,710 HAVE A LOW QUALITY OF LIFE AND 543 00:18:28,710 --> 00:18:31,880 ALSO A SHORTER LIFE SPAN. 544 00:18:31,880 --> 00:18:36,084 SO WE ASK WHETHER ENDUBS COULD 545 00:18:36,084 --> 00:18:39,221 BE USED TO HELP OUT 546 00:18:39,221 --> 00:18:39,588 CYSTIC FIBROSIS. 547 00:18:39,588 --> 00:18:41,389 IN ORDER TO DO THAT WE GENERATED 548 00:18:41,389 --> 00:18:46,361 NANO BODIES TO THE CFTR CHANNEL, 549 00:18:46,361 --> 00:18:47,896 WE PURIFIED THE NBD1 DOMAIN AND 550 00:18:47,896 --> 00:18:50,065 USED IT AS BAIT TO FISH OUT 551 00:18:50,065 --> 00:18:52,467 BINDERS FROM A SYNTHETIC YEEOF 552 00:18:52,467 --> 00:18:54,002 THE NANO BODY LIBRARY THAT 553 00:18:54,002 --> 00:18:55,737 CONTAINS A HUNDRED MILLION 554 00:18:55,737 --> 00:18:57,505 UNIQUE NANO BODIES AND AFTER 555 00:18:57,505 --> 00:19:00,041 ROUNDS OF THEM, 2 OR 3 ROUNDS OF 556 00:19:00,041 --> 00:19:02,277 THE SELECTION, WHAT YOU CAN SEE 557 00:19:02,277 --> 00:19:06,381 HERE IN THIS FLOW CYTOMETRY, FAX 558 00:19:06,381 --> 00:19:11,953 EXPERIMENT IS YOU GET AN 559 00:19:11,953 --> 00:19:13,088 EVOLUTION OF BINDERS RIGHT HERE, 560 00:19:13,088 --> 00:19:15,657 SO WE HAVE A NUMBER TO THE 561 00:19:15,657 --> 00:19:17,993 CFTRAD LEAST TO THE BD1. 562 00:19:17,993 --> 00:19:20,195 TO DETERMINE IF IT BINDS, WE USE 563 00:19:20,195 --> 00:19:22,831 A FRET ASSAY, AND THIS SHOWS 564 00:19:22,831 --> 00:19:24,699 THAT THE BINDER THAT BINDS THE 565 00:19:24,699 --> 00:19:26,701 FULL PROTEIN COMPARED TO A 566 00:19:26,701 --> 00:19:29,104 NEGATIVE CONTROL. 567 00:19:29,104 --> 00:19:29,337 OKAY. 568 00:19:29,337 --> 00:19:32,274 SO WITH THE BANDER WE'RE ABLE TO 569 00:19:32,274 --> 00:19:34,476 ARM THAT AND CONVERT IT INTO AN 570 00:19:34,476 --> 00:19:37,912 ENDUB AND SEE IF WE COULD RESCUE 571 00:19:37,912 --> 00:19:48,356 METRICSITANCY OF CRT CFTR 572 00:19:49,124 --> 00:19:49,624 CHANNELS. 573 00:19:49,624 --> 00:19:51,526 THE ATIN03 K IS THE ABOUT THE 574 00:19:51,526 --> 00:19:52,894 THIRD OR FOURTH MOST COMMON 575 00:19:52,894 --> 00:19:53,161 MUTATION. 576 00:19:53,161 --> 00:19:55,830 WHAT ARE YOU LOOKING AT HERE, IN 577 00:19:55,830 --> 00:19:58,366 BLACK ARE THE WILD-TYPE CURRENTS 578 00:19:58,366 --> 00:19:59,968 WITH THE WILD-TYPE RELATIONSHIP, 579 00:19:59,968 --> 00:20:02,170 WITH WILD-TYPE AND CHLORIDE 580 00:20:02,170 --> 00:20:03,471 CHANNEL CFTR, IN RED ARE THE 581 00:20:03,471 --> 00:20:03,705 MUTANT. 582 00:20:03,705 --> 00:20:07,542 SO YOU CAN SEE THAT AT ANY 583 00:20:07,542 --> 00:20:10,011 [INDISCERNIBLE]. 584 00:20:10,011 --> 00:20:11,413 THEY HAVE MINIMUM CURRENT GOING 585 00:20:11,413 --> 00:20:12,247 THROUGH THEM. 586 00:20:12,247 --> 00:20:13,081 ANOTHER IMPORTANT DIFFERENCE IN 587 00:20:13,081 --> 00:20:15,617 THE MUTATIONS IS THAT THERE'S 588 00:20:15,617 --> 00:20:18,787 SMALL MOLECULE MUTATION THERAPY 589 00:20:18,787 --> 00:20:20,221 FROM VERTEX PHARMACEUTICALS THAT 590 00:20:20,221 --> 00:20:23,658 HAVE BEEN APPROVED FOR THE F508 591 00:20:23,658 --> 00:20:26,261 DEL, BUT IS INEFFECTIVE FOR THE 592 00:20:26,261 --> 00:20:27,562 N1303 K. 593 00:20:27,562 --> 00:20:32,734 SO WHEN YOU ADD TRI TRIKAFTA, 594 00:20:32,734 --> 00:20:34,803 YOU GET A SMALL INCREASE IN THE 595 00:20:34,803 --> 00:20:36,438 CURRENT, NOWHERE NEAR THE 596 00:20:36,438 --> 00:20:38,540 WILD-TYPE BUT SUDDENLY 597 00:20:38,540 --> 00:20:39,674 RATIONALIZES WHY THE PATIENTS 598 00:20:39,674 --> 00:20:43,878 FEEL BETTER ON THIS DRUG. 599 00:20:43,878 --> 00:20:45,580 THE N1303 K DOES MARGINALLY 600 00:20:45,580 --> 00:20:47,248 WORSE THAN THIS AND THAT'S ALSO 601 00:20:47,248 --> 00:20:48,583 CONSISTENT WITH WHY THE PATIENTS 602 00:20:48,583 --> 00:20:50,618 DON'T DO AS WELL. 603 00:20:50,618 --> 00:20:52,020 WHAT WE'RE REALLY EXCITED ABOUT 604 00:20:52,020 --> 00:20:55,490 IS WHEN WE COMBINED THE 605 00:20:55,490 --> 00:21:01,996 TRIKAFTA, WITH THE END OF IT, 606 00:21:01,996 --> 00:21:04,132 YOU GET A WILD-TYPE, THE 1 607 00:21:04,132 --> 00:21:07,635 THAT'S BEEN APPROVED FOR FDA FOR 608 00:21:07,635 --> 00:21:09,738 TRIKAFTA, AS WELL AS WHAT HASN'T 609 00:21:09,738 --> 00:21:11,439 AND THAT'S SUFFICIENT TO CAUSE A 610 00:21:11,439 --> 00:21:12,907 RESCUE OF THOSE CURRENTS. 611 00:21:12,907 --> 00:21:15,844 AND SO WE'VE BEEN USING THESE 612 00:21:15,844 --> 00:21:17,245 ENDUBS IN VARIOUS DIFFERENT ION 613 00:21:17,245 --> 00:21:19,214 CHANNELS AS WELL AS OTHER 614 00:21:19,214 --> 00:21:21,816 DEFINITE PROTEINS, AND YOU KNOW, 615 00:21:21,816 --> 00:21:23,918 IT'S VERY HARD TO -- SO FAR WE 616 00:21:23,918 --> 00:21:25,220 HAVEN'T FOUND ANYTHING THAT IT 617 00:21:25,220 --> 00:21:25,787 DOESN'T WORK FOR. 618 00:21:25,787 --> 00:21:29,591 SO I'LL LEAVE IT THERE. 619 00:21:29,591 --> 00:21:29,924 OKAY. 620 00:21:29,924 --> 00:21:32,494 SO I'LL NOW GO ON TO A SECOND 621 00:21:32,494 --> 00:21:34,529 STORY, THAT WE'VE BEEN WORKING 622 00:21:34,529 --> 00:21:37,432 ON AND THAT TO SET UP THAT 623 00:21:37,432 --> 00:21:39,033 STORY, I WILL IMAGINE, ASK YOU 624 00:21:39,033 --> 00:21:40,435 TO IMAGINE BEING PUT INTO THE 625 00:21:40,435 --> 00:21:42,771 MIDDLE OF A VERY COMPLICATED 626 00:21:42,771 --> 00:21:44,939 CITY AND BEING TOLD TO GET FROM 627 00:21:44,939 --> 00:21:45,907 POINT A TO POINT B WITHOUT THE 628 00:21:45,907 --> 00:21:48,777 AID OF A MAP OR A GPS. 629 00:21:48,777 --> 00:21:50,545 FOR ME, EVEN WITH A GPS, IT 630 00:21:50,545 --> 00:21:55,150 MIGHT BE A PROBLEM, OKAY IN BUT 631 00:21:55,150 --> 00:21:57,185 THIS SAME SORT OF PROBLEM IS 632 00:21:57,185 --> 00:22:00,388 WHAT I THINK IS FACED BY ION 633 00:22:00,388 --> 00:22:01,990 CHANNELS THAT I MADE INSIDE THE 634 00:22:01,990 --> 00:22:04,959 CELL AND REALLY HAVE TO NAVIGATE 635 00:22:04,959 --> 00:22:06,327 A VERY COMPLEX INTRACELLULAR 636 00:22:06,327 --> 00:22:09,063 ENVIRONMENT IN ORDER TO GET TO 637 00:22:09,063 --> 00:22:13,034 THE CELL SURFACE WHERE THEY 638 00:22:13,034 --> 00:22:13,601 FUNCTION. 639 00:22:13,601 --> 00:22:14,836 SO WHAT WE'VE BEEN INTERESTED IN 640 00:22:14,836 --> 00:22:16,504 OR 1 OF THE QUESTIONS WE'VE BEEN 641 00:22:16,504 --> 00:22:17,238 INTERESTED IN ADDRESSING IS YOU 642 00:22:17,238 --> 00:22:20,008 KNOW WHAT ARE THE DIRECTIONS OR 643 00:22:20,008 --> 00:22:21,776 SIGNALS THAT ENABLE ION CHANNELS 644 00:22:21,776 --> 00:22:23,278 TO REALLY TRAVERSE THIS VERY 645 00:22:23,278 --> 00:22:25,046 COMPLICATED SYSTEM TO GET TO THE 646 00:22:25,046 --> 00:22:28,316 CELL SURFACE AND WE WILL USE 647 00:22:28,316 --> 00:22:30,051 THIS TOOL THAT I'VE ALREADY 648 00:22:30,051 --> 00:22:32,120 SPOKEN TO YOU ABOUT THE CHANNEL 649 00:22:32,120 --> 00:22:35,290 WHERE WE ARE AN EPITOPE TAG, AND 650 00:22:35,290 --> 00:22:37,625 LOOK AT ALSO THE YFP SO WE CAN 651 00:22:37,625 --> 00:22:40,228 LOOK AT THE TOTAL PROTEIN 652 00:22:40,228 --> 00:22:40,528 EXPRESSION. 653 00:22:40,528 --> 00:22:43,665 SO IF YOU EXPRESS THIS ENGINEER 654 00:22:43,665 --> 00:22:46,034 CHANNEL INSIDE THE HEX CELL AND 655 00:22:46,034 --> 00:22:47,435 LOOK FOR WHERE IT'S DISTRIBUTED, 656 00:22:47,435 --> 00:22:49,504 YOU FIND IT IN IN OTHER PLACE 657 00:22:49,504 --> 00:22:52,807 WHERE YOU FIND IT, IN THE ER, 658 00:22:52,807 --> 00:22:54,742 GOLGI, ENDOSTUDIES OF MULTIPLE 659 00:22:54,742 --> 00:22:56,010 ENDOCRINES AND LIES SOSOMES SO 660 00:22:56,010 --> 00:22:57,545 IT GOES THROUGH THE TRAFFICKING 661 00:22:57,545 --> 00:22:57,879 PATHWAY. 662 00:22:57,879 --> 00:22:59,414 AND THE QUESTION WE'RE 663 00:22:59,414 --> 00:23:00,849 INTERESTED IN IS WHAT ACTUALLY 664 00:23:00,849 --> 00:23:03,751 DICTATES THE ABILITY OF THOSE 665 00:23:03,751 --> 00:23:07,222 CHANNELS TO REALLY GO THROUGH 666 00:23:07,222 --> 00:23:08,456 THOSE PATHWAYS, RIGHT? 667 00:23:08,456 --> 00:23:10,325 AND THE HYPOTHESIS WE HAD WAS 668 00:23:10,325 --> 00:23:11,526 THAT THE UBIQUITIN SUGGESTS WAS 669 00:23:11,526 --> 00:23:13,261 INVOLVED IN THAT, BECAUSE 670 00:23:13,261 --> 00:23:14,629 UBIQUITIN IS MOST WELL KNOWN FOR 671 00:23:14,629 --> 00:23:17,499 ITS ROLE IN PROTEIN DEGRADATION, 672 00:23:17,499 --> 00:23:19,901 BUT THERE ARE INDICATIONS IT 673 00:23:19,901 --> 00:23:21,102 COULD POTENTIALLY ALSO BE 674 00:23:21,102 --> 00:23:23,404 INVOLVED IN TRAFFICKING AND SO 675 00:23:23,404 --> 00:23:23,638 FORTH. 676 00:23:23,638 --> 00:23:27,642 SO HOW CAN THAT HAPPEN, THE SAME 677 00:23:27,642 --> 00:23:31,579 SIGNAL BE UTILIZED FOR THESE 678 00:23:31,579 --> 00:23:32,146 MANY DIFFERENT EFFECTS? 679 00:23:32,146 --> 00:23:35,984 WELL 1 THING THAT WE -- THAT 680 00:23:35,984 --> 00:23:38,887 COULD BE THE CASE IS THAT 681 00:23:38,887 --> 00:23:42,123 UBIQUITIN IS NORMALLY ATTACHED 682 00:23:42,123 --> 00:23:43,958 TO LYSINE SUBSTRATES BUT IT 683 00:23:43,958 --> 00:23:47,095 TURNS OUT THE UBIQUITIN ITSELF 684 00:23:47,095 --> 00:23:51,666 HAS 7 DISTINCT LYSINEs AND 685 00:23:51,666 --> 00:23:53,268 THOSE COULD BE UBIQUITINNATED AS 686 00:23:53,268 --> 00:23:53,635 WELL. 687 00:23:53,635 --> 00:23:58,172 SO DEPENDING ON THE TYPE OF 688 00:23:58,172 --> 00:24:00,942 UBIQUITIN THAT'S ATTACHED TO A 689 00:24:00,942 --> 00:24:03,978 PARTICULAR LYSINE, CAN YOU GET 690 00:24:03,978 --> 00:24:04,712 UBIQUITIN THAT LOOK VERY 691 00:24:04,712 --> 00:24:05,780 DIFFERENT IN TERMS OF SIZE AND 692 00:24:05,780 --> 00:24:08,149 SHAPE AND SO COULD BE RECOGNIZED 693 00:24:08,149 --> 00:24:10,151 DIFFERENTLY WITHIN THE CELL. 694 00:24:10,151 --> 00:24:14,455 SO K48 IS THE CANNONICLE 695 00:24:14,455 --> 00:24:17,125 DEGRADATION SIGNAL WHERE AS 696 00:24:17,125 --> 00:24:19,861 CASES FOR EXAMPLE, STARTED TO BE 697 00:24:19,861 --> 00:24:20,361 NONDEGRADATION 698 00:24:20,361 --> 00:24:20,895 GRATES--GRATUEIDATIVE AND 699 00:24:20,895 --> 00:24:21,162 FUNCTIONS. 700 00:24:21,162 --> 00:24:22,997 AND THE SYSTEM GETS MORE 701 00:24:22,997 --> 00:24:24,299 COMPLICATED BECAUSE MASS SPEC 702 00:24:24,299 --> 00:24:26,901 HAS INDICATED YOU CAN FORM 703 00:24:26,901 --> 00:24:28,036 DIFFERENT POLYUBIQUITIN CHAINS 704 00:24:28,036 --> 00:24:29,203 THAT HAVE VERY DIFFERENT 705 00:24:29,203 --> 00:24:29,771 ARCHITECTURE SYSTEMS AND SO 706 00:24:29,771 --> 00:24:30,772 FORTH SO YOU CAN IMAGINE THAT 707 00:24:30,772 --> 00:24:32,540 MOST OF THE CELLS THAT ARE -- 708 00:24:32,540 --> 00:24:34,909 MOST OF THE PROTEINS INSIDE OF 709 00:24:34,909 --> 00:24:37,445 CELL REALLY HAVE A TON OF 710 00:24:37,445 --> 00:24:39,314 UBIQUITIN ON THEM AND WE HAVE 711 00:24:39,314 --> 00:24:43,318 ABSOLUTELY NO IDEA THE 712 00:24:43,318 --> 00:24:45,119 FUNCTIONAL CONSEQUENCES OF THIS. 713 00:24:45,119 --> 00:24:46,721 SO OUR HIGHWAY PUTTING SIS WAS 714 00:24:46,721 --> 00:24:49,257 THAT THIS SOMEHOW WAS INVOLVED 715 00:24:49,257 --> 00:24:50,925 IN THE REGULATING ION CHANNELS 716 00:24:50,925 --> 00:24:54,228 AND THAT'S THE THING THAT WE 717 00:24:54,228 --> 00:24:54,963 WILL INVESTIGATE. 718 00:24:54,963 --> 00:24:56,497 SO TO ADDRESS THAT, THE FIRST 719 00:24:56,497 --> 00:24:58,132 THING WE ASKED IS WHETHER THE 720 00:24:58,132 --> 00:25:00,568 CHANNEL EXPRESSED IN HEX CELLS 721 00:25:00,568 --> 00:25:02,403 DOES HAVE ANY POLYUBIQUITIN ON 722 00:25:02,403 --> 00:25:04,939 IT, SO WE -- WE PULL DOWN THE 723 00:25:04,939 --> 00:25:06,207 CHANNEL EXPRESSING HEX CELLS, 724 00:25:06,207 --> 00:25:09,344 AND THEN WE COULD CUT OUT THE 725 00:25:09,344 --> 00:25:10,178 MONOMERAND THE DIME MERRIC 726 00:25:10,178 --> 00:25:12,447 SPECIES AND HAVE THEM IN MASS 727 00:25:12,447 --> 00:25:17,385 SPECT AND IN ADDITION TO THE 728 00:25:17,385 --> 00:25:18,753 PEPTIDES REPRESENTING THE 729 00:25:18,753 --> 00:25:20,588 CHANNEL, WHAT WE ALSO SAW IN 730 00:25:20,588 --> 00:25:22,223 THERE WERE PEPTIDES THAT 731 00:25:22,223 --> 00:25:24,225 CORRESPONDED TO UBIQUITIN AND. 732 00:25:24,225 --> 00:25:26,995 UBIQUITIN DEPENDING ON THE 733 00:25:26,995 --> 00:25:30,264 LYSINE THAT IS UBIQUITINNATED IT 734 00:25:30,264 --> 00:25:32,467 LEAVES BETWEEN A DISCIPLINARY 735 00:25:32,467 --> 00:25:33,701 GLAISIN SIGNATURE, SO BASED ON 736 00:25:33,701 --> 00:25:35,870 WHERE THE DISCIPLINARY GLIESIN 737 00:25:35,870 --> 00:25:38,640 IS ON THE LYSINES, YOU COULD 738 00:25:38,640 --> 00:25:41,175 FIGURE OUT WHICH 1S WERE 739 00:25:41,175 --> 00:25:41,542 UBIQUITINNATED. 740 00:25:41,542 --> 00:25:45,213 SO WHAT WE DID NOW IS WAS ON THE 741 00:25:45,213 --> 00:25:47,348 UBIQUITIN CALCIUM CHANNEL WE 742 00:25:47,348 --> 00:25:50,251 FOUND OUT 40% OF REPRESENTED 743 00:25:50,251 --> 00:25:51,786 K48, ABOUT 24%, WAS THE K AND 744 00:25:51,786 --> 00:25:54,288 THERE WAS A MINOR'S SPECIES OF 745 00:25:54,288 --> 00:25:55,123 ABOUT 4%. 746 00:25:55,123 --> 00:25:58,726 SO K48 IS THE CANNONICLE 747 00:25:58,726 --> 00:26:06,167 DEGRADATION SIGNAT SIGNATURES E 748 00:26:06,167 --> 00:26:07,835 LITERATURE. 749 00:26:07,835 --> 00:26:09,804 ALL RIGHT, SO, THE QUESTION NOW 750 00:26:09,804 --> 00:26:12,573 BECOMES, SO, WE DO HAVE 751 00:26:12,573 --> 00:26:16,044 POLYUBIQUITINNATION OF THE 752 00:26:16,044 --> 00:26:16,277 CHANNEL. 753 00:26:16,277 --> 00:26:17,345 WHAT DOES IT DO? 754 00:26:17,345 --> 00:26:18,746 THERE'S NO WAY THAT RIGHT NOW 755 00:26:18,746 --> 00:26:21,516 THAT YOU CAN ADDRESS THAT IN A 756 00:26:21,516 --> 00:26:22,050 LIVE CELL. 757 00:26:22,050 --> 00:26:24,619 BUT WE THOUGHT WE COULD 758 00:26:24,619 --> 00:26:25,553 POTENTIALLY EXTEND THE ENDUB 759 00:26:25,553 --> 00:26:26,821 APPROACH THAT WE THEN DEVELOPED 760 00:26:26,821 --> 00:26:28,890 TO BEGIN TO ADDRESS THOSE KINDS 761 00:26:28,890 --> 00:26:29,691 OF QUESTIONS. 762 00:26:29,691 --> 00:26:35,930 AND WE'RE GOING TO MAKE USE OF 7 763 00:26:35,930 --> 00:26:39,967 TYPES OF DECIBOL WITTEN ACES 764 00:26:39,967 --> 00:26:42,070 THAT HAVE EXQUISITE LINKAGE 765 00:26:42,070 --> 00:26:45,239 SPECIFICITY AND THE ABILITY TO 766 00:26:45,239 --> 00:26:46,841 HIDE ROLLIZE THE POLYUBIQUITIN 767 00:26:46,841 --> 00:26:47,308 CHAINS. 768 00:26:47,308 --> 00:26:56,284 SO FROM [INDISCERNIBLE]'S LAB, 769 00:26:56,284 --> 00:27:00,288 YOU HAVE DEUBIQUITIN AISS AND SO 770 00:27:00,288 --> 00:27:03,424 THE IDEA WAS THAT WE COULD NOW 771 00:27:03,424 --> 00:27:11,599 HIJACK SOME OF THOSE LINKAGE 772 00:27:11,599 --> 00:27:13,000 SPECIFIC DEUBIQUITIN ASES AND 773 00:27:13,000 --> 00:27:15,970 THAT'S HOW YOU GET THE SELECTIVE 774 00:27:15,970 --> 00:27:16,204 ENDUBS. 775 00:27:16,204 --> 00:27:17,739 SO WE GENERATE THESE MOLECULES, 776 00:27:17,739 --> 00:27:19,173 FIRST WE ASKED WHETHER THEY HAD 777 00:27:19,173 --> 00:27:21,776 ANY EFFECT ON THE BIOCHEMICAL 778 00:27:21,776 --> 00:27:23,077 UBIQUITIN SIGNATURE OF THE PULL 779 00:27:23,077 --> 00:27:26,647 DOWN CHANNEL. 780 00:27:26,647 --> 00:27:29,484 SO COMPARED TO THE BASE LINE 781 00:27:29,484 --> 00:27:33,121 CONTROL, WE SAW THAT ALL THESE 782 00:27:33,121 --> 00:27:34,388 PUTATIVE LINKAGE SELECTIVE 783 00:27:34,388 --> 00:27:35,456 ENDUBS DECREASE THE UBIQUITIN 784 00:27:35,456 --> 00:27:37,158 SIGNAL ON THAT ON THE CHANNEL 785 00:27:37,158 --> 00:27:39,360 AND SO THIS INDICATE THAD THEY 786 00:27:39,360 --> 00:27:40,862 HAD SOME ACTIVITY, THEN WE WERE 787 00:27:40,862 --> 00:27:43,264 SURPRISED THAT THEY ALL SEEM TO 788 00:27:43,264 --> 00:27:44,966 HAVE PRETTY PROBUST ACTIVITY 789 00:27:44,966 --> 00:27:46,601 GIVEN THAT K48 WAS THE MOST 790 00:27:46,601 --> 00:27:47,668 DOMINANT 1. 791 00:27:47,668 --> 00:27:54,008 THAT'S MEDIATED BY THIS 1. 792 00:27:54,008 --> 00:27:55,309 WHEREAS [INDISCERNIBLE] THE 793 00:27:55,309 --> 00:27:56,310 MINOR SPECIES BUT HOWEVER THEY 794 00:27:56,310 --> 00:27:59,180 ALL SEEM TO BE VERY EFFECTIVE IN 795 00:27:59,180 --> 00:28:00,214 HIDE ROLLIZING THE UBIQUITIN 796 00:28:00,214 --> 00:28:02,250 FROM THE PULL DOWN CHANNEL. 797 00:28:02,250 --> 00:28:05,153 ONE POTENTIAL WAY THAT COULD 798 00:28:05,153 --> 00:28:06,854 ADDRESS THAT -- THAT COULD 799 00:28:06,854 --> 00:28:08,623 EXPLAIN THAT THAT THERE'S A LOT 800 00:28:08,623 --> 00:28:09,791 OF BRANCHING IN THE UBIQUITIN 801 00:28:09,791 --> 00:28:10,758 CHAIN ANDS THAT'S SOMETHING WE 802 00:28:10,758 --> 00:28:11,592 CAN GET TO. 803 00:28:11,592 --> 00:28:13,594 HOWEVER, SO, BASIC LE USING THE 804 00:28:13,594 --> 00:28:15,096 BIOCHEMICAL SIGNATURE, WE 805 00:28:15,096 --> 00:28:15,797 COULDN'T REALLY TELL WHETHER 806 00:28:15,797 --> 00:28:19,500 THERE WAS A DIFFERENCE AND HOW 807 00:28:19,500 --> 00:28:28,242 THESE THINGS WERE HYDROLLIZING 808 00:28:28,242 --> 00:28:28,709 FROM THE CHANNEL. 809 00:28:28,709 --> 00:28:30,444 BUT NOW WE CAN LOOK AT 810 00:28:30,444 --> 00:28:33,080 FUNCTIONAL OUTPUTS TO SEE THE 811 00:28:33,080 --> 00:28:33,648 DIFFERENT LINKAGES HAVE 812 00:28:33,648 --> 00:28:34,382 DIFFERENT EFFECTS ON THE 813 00:28:34,382 --> 00:28:35,349 EXPRESSION AND THE TRAFFIC OF 814 00:28:35,349 --> 00:28:37,118 THE CHANNEL, SO FESTER WE LOOK 815 00:28:37,118 --> 00:28:39,086 AT YFP SIGNAL AND THAT GIVES YOU 816 00:28:39,086 --> 00:28:40,388 AN INDICATION OF THE TOTAL 817 00:28:40,388 --> 00:28:44,258 AMOUNT OF PROTEIN THAT'S 818 00:28:44,258 --> 00:28:44,525 EXPRESSED. 819 00:28:44,525 --> 00:28:50,965 SO IF WE START FROM THE RIGHT 820 00:28:50,965 --> 00:28:54,068 HAND SIDE, THIS IS ENDUB THAT 821 00:28:54,068 --> 00:28:55,036 HIDE ROLLIZES ALL UBIQUITIN 822 00:28:55,036 --> 00:28:58,906 CHAINS AND WHEN YOU DO THAT YOU 823 00:28:58,906 --> 00:29:00,975 GET AN INCREASE IN THE AMOUNT OF 824 00:29:00,975 --> 00:29:02,543 PROTEIN EXPRESS TD. 825 00:29:02,543 --> 00:29:05,513 NOT SURPRISING WHEN YOU FIND OUT 826 00:29:05,513 --> 00:29:06,113 THAT UBIQUITIN STABILIZES THE 827 00:29:06,113 --> 00:29:07,615 ABILITY OF THE PROTEIN. 828 00:29:07,615 --> 00:29:09,851 BUT WHAT SURPRISING WAS WHICH 829 00:29:09,851 --> 00:29:10,985 LINKAGES WERE IMPORTANT. 830 00:29:10,985 --> 00:29:13,020 SO THE DOMINANT 1 K48 WHICH IS 831 00:29:13,020 --> 00:29:15,556 CANONICALLY RELATED TO PROTEIN 832 00:29:15,556 --> 00:29:17,225 DEGRADATION, THAT HAD ABSOLUTELY 833 00:29:17,225 --> 00:29:18,993 NO EFFECT ON THE SEPARATE OF THE 834 00:29:18,993 --> 00:29:22,163 PROTEIN, NOR DID THE SECOND MOST 835 00:29:22,163 --> 00:29:24,031 DOMINANT 1 K63 REMOVAL. 836 00:29:24,031 --> 00:29:26,534 THE 1S THAT HAVE THE MOST IMPACT 837 00:29:26,534 --> 00:29:28,436 ON THESE, GREATER PROTEIN WITH 838 00:29:28,436 --> 00:29:31,038 THE MINOR SPECIES, THE K11 AND 839 00:29:31,038 --> 00:29:33,174 K29, K33. 840 00:29:33,174 --> 00:29:34,575 SO THAT WAS A FIRST SURPRISE 841 00:29:34,575 --> 00:29:37,845 LOOKING AT THE TOLL AT PROTEIN 842 00:29:37,845 --> 00:29:38,179 CONCENTRATION. 843 00:29:38,179 --> 00:29:40,448 NEXT WE COULD LOOK AT WAS THE 844 00:29:40,448 --> 00:29:41,549 SURFACE DENSITY OF THE CHANNELS 845 00:29:41,549 --> 00:29:42,750 ISSUES THE ABILITY OF THE 846 00:29:42,750 --> 00:29:44,919 CHANNELS TO GET TO THE CELL 847 00:29:44,919 --> 00:29:46,187 SURFACE AND SO STARTING ONCE 848 00:29:46,187 --> 00:29:47,488 AGAIN FROM THE RIGHT HAND SIDE, 849 00:29:47,488 --> 00:29:50,725 EVEN THOUGH YOU GET MORE 850 00:29:50,725 --> 00:29:52,360 PROTEINS BY REMOVING UBIQUITIN, 851 00:29:52,360 --> 00:29:53,895 THAT DOESN'T NECESSARILY 852 00:29:53,895 --> 00:29:55,096 TRANSLATE INTO MORE OF IT 853 00:29:55,096 --> 00:29:58,299 GETTING TO THE CELL SURFACE. 854 00:29:58,299 --> 00:29:59,667 SO UBIQUITIN IS NOT ONLY 855 00:29:59,667 --> 00:30:02,003 IMPORTANT FOR THE STABILITY, BUT 856 00:30:02,003 --> 00:30:03,271 POTENTIALLY COULD BE IMPORTANT 857 00:30:03,271 --> 00:30:05,206 FOR THE TRAFFICKING OF THE 858 00:30:05,206 --> 00:30:05,673 CHANNEL. 859 00:30:05,673 --> 00:30:08,809 SECOND SURPRISING WAS TO -- WHEN 860 00:30:08,809 --> 00:30:11,312 WE REMOVED THE K48, WE GOT LESS 861 00:30:11,312 --> 00:30:14,181 CHANNELS ON THE CELL SURFACE. 862 00:30:14,181 --> 00:30:16,450 K63 WAS PARTICULARLY NEUTRAL BUT 863 00:30:16,450 --> 00:30:19,987 ONCE AGAIN WE'RE MOVING K11, AND 864 00:30:19,987 --> 00:30:21,589 K29, PROMOTED THE SURFACE 865 00:30:21,589 --> 00:30:23,224 TRAFFICKING OF THE CHANNEL. 866 00:30:23,224 --> 00:30:25,426 SO ALL THESE INDICATED THAT 867 00:30:25,426 --> 00:30:26,928 DIFFERENT LINKAGES WERE 868 00:30:26,928 --> 00:30:28,129 POTENTIALLY INVOLVED IN ASPECTS 869 00:30:28,129 --> 00:30:29,230 OF PROTEINS AND STABILITY AS 870 00:30:29,230 --> 00:30:30,831 WELL AS TRAFFICKING ACCIDENT SO 871 00:30:30,831 --> 00:30:31,632 THESE ARE STEADY STATE 872 00:30:31,632 --> 00:30:33,267 MEASUREMENTS OF THE SURFACE 873 00:30:33,267 --> 00:30:34,902 DENSITY SO WE COULD ALSO ADOPT 874 00:30:34,902 --> 00:30:36,771 OUR FIRST TO LOOK AT THE RATE OF 875 00:30:36,771 --> 00:30:38,406 FORWARD TRAFFIC AS WELL AS RATE 876 00:30:38,406 --> 00:30:41,008 OF REMOVAL AND WHEY DID THAT WE 877 00:30:41,008 --> 00:30:42,410 SAW A VERY DIFFERENT INTEREST IN 878 00:30:42,410 --> 00:30:43,811 THEM AS WELL, SO LOOK AT THE 879 00:30:43,811 --> 00:30:46,247 RATE OF FORWARD TRAFFICKING, THE 880 00:30:46,247 --> 00:30:46,847 BLACK REPRESENTS THE CONTROL 881 00:30:46,847 --> 00:30:49,583 CASE AND WHAT WE SAW WAS THAT 882 00:30:49,583 --> 00:30:51,385 MOVEMENT IN THE MINOR SPECIES, 883 00:30:51,385 --> 00:30:52,887 PROMOTED THE RATE OF FORWARD 884 00:30:52,887 --> 00:30:54,755 TRAFFICKING, SO YOU GET MORE OF 885 00:30:54,755 --> 00:30:58,292 THE CHANNELS GETTING TO THE CELL 886 00:30:58,292 --> 00:31:00,027 SURFACE, REMOVING ALL UBIQUITIN 887 00:31:00,027 --> 00:31:01,529 ACTUALLY HAD A -- YOU GOT A 888 00:31:01,529 --> 00:31:02,596 PROFOUND LOSS OF THE ABILITY OF 889 00:31:02,596 --> 00:31:05,132 THE CHANNEL TO TRAFFIC TO THE 890 00:31:05,132 --> 00:31:05,466 CELL SURFACE. 891 00:31:05,466 --> 00:31:06,767 THE OTHER THING WE COULD LOOK AT 892 00:31:06,767 --> 00:31:08,536 WHAT WAS THE RATE OF REMOVAL 893 00:31:08,536 --> 00:31:10,037 CHANNELS THAT ARE ALREADY AT THE 894 00:31:10,037 --> 00:31:11,906 CELL SURFACE, AND ONCE AGAIN, 895 00:31:11,906 --> 00:31:13,274 THEY WERE REMOVING DIFFERENT 896 00:31:13,274 --> 00:31:14,141 LINKAGES AND HAD DIFFERENT 897 00:31:14,141 --> 00:31:16,310 EFFECTS ON THEM, ON THE ABILITY 898 00:31:16,310 --> 00:31:22,049 OF TURN OUT TO BE 899 00:31:22,049 --> 00:31:22,450 UNDERSIGNIFYITOSE. 900 00:31:22,450 --> 00:31:23,117 PUTTING ALL THAT TOGETHER WITH 901 00:31:23,117 --> 00:31:24,652 THE DATA I DON'T HAVE TIME TO GO 902 00:31:24,652 --> 00:31:26,053 THROUGH, CAME UP WITH THIS 903 00:31:26,053 --> 00:31:30,858 EXCITING MODEL FOR HOW UBIQUITIN 904 00:31:30,858 --> 00:31:32,727 ARE AFFECTING THE TRAFFICKING 905 00:31:32,727 --> 00:31:32,960 PATHWAY. 906 00:31:32,960 --> 00:31:36,030 SO MOST OF THE CHANNELS PRESENT 907 00:31:36,030 --> 00:31:38,933 IN IF THEY WERE LABELED WITH 908 00:31:38,933 --> 00:31:40,835 K11, K29, K33, WHICH IS THE 909 00:31:40,835 --> 00:31:42,536 MINOR SPECIES, THEY ACTUALLY GET 910 00:31:42,536 --> 00:31:44,405 SIDE TRACKS TO THE PROTEIN 911 00:31:44,405 --> 00:31:45,840 COMPLEXIOSOME TO BE DEGRADED. 912 00:31:45,840 --> 00:31:48,442 IN ORDER FOR THE CHANNEL TO 913 00:31:48,442 --> 00:31:50,177 ACTUALLY ADD ATTRACT IT TO THE 914 00:31:50,177 --> 00:31:52,246 CELL SURFACE, IT NEEDS K48 WHICH 915 00:31:52,246 --> 00:31:53,981 ANOTHER PROTEIN IS A DEGRADATION 916 00:31:53,981 --> 00:31:56,217 SIGNAL AND IN THIS CASE, IS A 917 00:31:56,217 --> 00:31:57,685 TRAFFICKING SIGNAL TO GET TO THE 918 00:31:57,685 --> 00:32:01,956 CELL SURFACE, FOR REMOVAL OF THE 919 00:32:01,956 --> 00:32:04,925 CHANNEL, YOU REQUIRE K63, AND IF 920 00:32:04,925 --> 00:32:07,061 YOU REMOVE K63, YOU SCROLL DOWN 921 00:32:07,061 --> 00:32:08,629 THE HYDROLYSIS OF THE CHANNEL, 922 00:32:08,629 --> 00:32:10,564 AND SO FORTH, AND SO WHAT THIS 923 00:32:10,564 --> 00:32:15,302 INDICATE THAD INDEED THE 924 00:32:15,302 --> 00:32:17,138 POLYUBIQUITIN CHAINS OF 925 00:32:17,138 --> 00:32:18,239 DIFFERENT LINKAGES HAVE PROFOUND 926 00:32:18,239 --> 00:32:19,373 DIFFERENT ASPECTS OF THE CALCIUM 927 00:32:19,373 --> 00:32:19,607 CHANNEL. 928 00:32:19,607 --> 00:32:21,776 SO THIS IS WHAT PEOPLE IN THE 929 00:32:21,776 --> 00:32:23,010 UBIQUITIN FIELD CALL THE 930 00:32:23,010 --> 00:32:23,744 UBIQUITIN CODE. 931 00:32:23,744 --> 00:32:24,945 AND 1 THING I WOULD LIKE TO 932 00:32:24,945 --> 00:32:26,447 STRESS IS THAT THE UBIQUITIN IS 933 00:32:26,447 --> 00:32:29,583 MADE UP OF A SYSTEM OF EDITORS 934 00:32:29,583 --> 00:32:30,217 AND SO FORTH. 935 00:32:30,217 --> 00:32:32,019 AND THATIA GOING TO BE DIFFERENT 936 00:32:32,019 --> 00:32:33,754 IN DIFFERENT CELLS, AND SO WE 937 00:32:33,754 --> 00:32:35,890 COULD -- FOR EXAMPLE, WE SHOWED 938 00:32:35,890 --> 00:32:37,825 THAT IF WE OVEREXPRESS, A 939 00:32:37,825 --> 00:32:39,560 PARTICULAR TYPE OF LIGASE, WE 940 00:32:39,560 --> 00:32:40,494 CAN AFFAIRS TEAM LEADERRER THE 941 00:32:40,494 --> 00:32:43,464 UBIQUITIN CODE IN OUR LINKAGE 942 00:32:43,464 --> 00:32:45,166 SELECTIVE ENDUBS ARE ABLE TO 943 00:32:45,166 --> 00:32:45,666 DISTINGUISH THAT. 944 00:32:45,666 --> 00:32:49,403 IF WE DO THE SAME EXPERIMENTS IN 945 00:32:49,403 --> 00:32:50,171 HARP, THE OUTCOME IS DEFINITE 946 00:32:50,171 --> 00:32:51,972 THAN WHAT YOU SEE IN HEX CELLS 947 00:32:51,972 --> 00:32:52,907 BECAUSE THE UBIQUITIN CODE IS 948 00:32:52,907 --> 00:32:53,174 DIFFERENT. 949 00:32:53,174 --> 00:32:55,242 SO I THINK IT WILL BE A VERY 950 00:32:55,242 --> 00:32:57,745 INTERESTING APPROACH TO BEGIN TO 951 00:32:57,745 --> 00:32:58,679 DECODE THE UBIQUITIN FOR 952 00:32:58,679 --> 00:33:02,183 DIFFERENT PROTEINS INSIDE THE 953 00:33:02,183 --> 00:33:02,383 CELL. 954 00:33:02,383 --> 00:33:03,717 ALL RIGHT, SO THE LAST STORY I 955 00:33:03,717 --> 00:33:06,153 WANT TO TALK ABOUT REALLY, SO 956 00:33:06,153 --> 00:33:07,421 FAR THE 2 STORIES I'VE BEEN 957 00:33:07,421 --> 00:33:11,158 TALKING ABOUT ARE TALKING ABOUT 958 00:33:11,158 --> 00:33:12,560 TARGETED DEUBIQUITIN TO 959 00:33:12,560 --> 00:33:14,495 STABILIZE THE PROTEINS OR TO 960 00:33:14,495 --> 00:33:18,032 LOOK AT ROLES IN ION CHANNEL 961 00:33:18,032 --> 00:33:18,332 TRAFFICKING. 962 00:33:18,332 --> 00:33:19,266 THE NICE THING ABOUT THE 963 00:33:19,266 --> 00:33:19,967 UBIQUITIN SYSTEMS IS THAT YOU 964 00:33:19,967 --> 00:33:22,169 CAN DO THE OPPOSITE, YOU CAN 965 00:33:22,169 --> 00:33:24,038 REMOVE CHANNELS BY ADDING 966 00:33:24,038 --> 00:33:24,505 UBIQUITIN TO THEM. 967 00:33:24,505 --> 00:33:27,141 SO WE'VE BEEN LOOKING AT THAT 968 00:33:27,141 --> 00:33:29,643 ASPECT OF THIS STORY AS WELL, 969 00:33:29,643 --> 00:33:33,514 AND FOR THIS, THE -- THE -- 970 00:33:33,514 --> 00:33:34,915 WHERE WE MADE THE MOST HEAD WAY 971 00:33:34,915 --> 00:33:37,651 IS LOOKING AT MY FIRST LOVE WHEN 972 00:33:37,651 --> 00:33:39,587 IT COMES TO ION CHANNELS THAT 973 00:33:39,587 --> 00:33:42,289 ALLOW CALCIUM TO COME IN AND 974 00:33:42,289 --> 00:33:44,291 EXCITABLE CELLS TO LINK ACTION 975 00:33:44,291 --> 00:33:45,559 POTENTIALS TO BIOLOGICAL 976 00:33:45,559 --> 00:33:47,161 RESPONSES, OKAY? 977 00:33:47,161 --> 00:33:49,363 SO 1 OF THE MAJOR THINGS WE'VE 978 00:33:49,363 --> 00:33:50,631 BEEN INTERESTED IN AND THESE ARE 979 00:33:50,631 --> 00:33:53,834 VERY IMPORTANT AND THERAPEUTIC 980 00:33:53,834 --> 00:33:55,536 TARGETS, L-TYPE CALCIUM CHANNELS 981 00:33:55,536 --> 00:33:57,605 ARE WELL KNOWN HYPE EAR 982 00:33:57,605 --> 00:33:58,772 TENSIVES, THE NEURONAL GET 983 00:33:58,772 --> 00:33:59,840 CALCIUM CHANNELS THAT ARE 984 00:33:59,840 --> 00:34:02,009 TARGETED FOR THINGS LIKE 985 00:34:02,009 --> 00:34:05,613 NEUROPATH WICK PAIN, PARKINSON'S 986 00:34:05,613 --> 00:34:06,747 DEC AND FORTH. 987 00:34:06,747 --> 00:34:09,016 SO THE LIMITATIONS OF SMALL 988 00:34:09,016 --> 00:34:10,384 MOLECULE DRUGS BECAUSE THEY GO 989 00:34:10,384 --> 00:34:12,653 EVERYWHERE AND THIS CHANNEL IS 990 00:34:12,653 --> 00:34:13,687 PREVALENT IS MANY DIFFERENT 991 00:34:13,687 --> 00:34:15,990 TISSUES CAN YOU GET OFF TARGET 992 00:34:15,990 --> 00:34:17,525 OFF TISSUE EFFECTS THAT LIMIT 993 00:34:17,525 --> 00:34:19,226 THE THERAPEUTIC WINDOW AND ALSO 994 00:34:19,226 --> 00:34:21,629 SOME OF THESE CHANNELS LOOK SO 995 00:34:21,629 --> 00:34:24,331 CLOSE TO EACH OTHER IT'S BEEN 996 00:34:24,331 --> 00:34:25,633 DIFFICULT TO FIND SMALL 997 00:34:25,633 --> 00:34:26,667 MOLECULES TO DISTINGUISH THEM. 998 00:34:26,667 --> 00:34:29,136 SO MY LAB'S BEEN VERY INTERESTED 999 00:34:29,136 --> 00:34:31,639 IN MAKING GENETICALLY ENCODED 1000 00:34:31,639 --> 00:34:32,573 CALCIUM BLOCKING CHANNELS THAT 1001 00:34:32,573 --> 00:34:33,874 CAN ADDRESS THESE DEFICITS. 1002 00:34:33,874 --> 00:34:36,310 SO THE CHANNEL IS MADE UP OF A 1003 00:34:36,310 --> 00:34:37,811 [INDISCERNIBLE], BUT MANY OF 1004 00:34:37,811 --> 00:34:41,715 THESE ALSO HAVE AN AUXILLIARY 1005 00:34:41,715 --> 00:34:42,883 BETA SUBSUANT THAT IS REQUIRED 1006 00:34:42,883 --> 00:34:44,485 FOR FORMATION OF THE CHANNEL SO 1007 00:34:44,485 --> 00:34:45,886 TO MAKE THESE CHANNELS 1 OF THE 1008 00:34:45,886 --> 00:34:47,855 FIRST THING WE DID WAS TO MAKE 1009 00:34:47,855 --> 00:34:51,091 AN NANO BODY TO THISAUK 1010 00:34:51,091 --> 00:34:52,426 ILLEGALSIARY BETA SUBUNIT. 1011 00:34:52,426 --> 00:34:55,196 AND WE DID THIS THE OLD 1012 00:34:55,196 --> 00:34:57,398 FASHIONED WEIGH, WE PURIFIED A 1013 00:34:57,398 --> 00:34:59,466 SUBBETTA UNIT, SENT IT TO A FARM 1014 00:34:59,466 --> 00:35:01,302 IN MASSACHUSETTS, THEY INJECTED 1015 00:35:01,302 --> 00:35:03,504 IT INTO A LLAMA AND THEY SENT US 1016 00:35:03,504 --> 00:35:07,041 THE BLOOD AND WE USED A PHASED 1017 00:35:07,041 --> 00:35:08,576 APPROACH TO FISH OUT THE 1018 00:35:08,576 --> 00:35:08,809 BINDERS. 1019 00:35:08,809 --> 00:35:11,912 SO HERE'S A BINDER WE FISHED OUT 1020 00:35:11,912 --> 00:35:13,914 F3, AND THIS INDICATES IT BINDS 1021 00:35:13,914 --> 00:35:15,482 ALL 4 TYPES OF BETA ISOFORMS 1022 00:35:15,482 --> 00:35:17,284 THAT ARE OUT THERE. SO WE HAVE 1023 00:35:17,284 --> 00:35:18,919 THESE NANO BODY BINDERS, WHAT DO 1024 00:35:18,919 --> 00:35:20,387 WE DO WITH THEM, WELL WE WILL DO 1025 00:35:20,387 --> 00:35:22,489 WHAT WE DID WITH THE ENDUBS IN 1026 00:35:22,489 --> 00:35:22,957 REVERSE. 1027 00:35:22,957 --> 00:35:25,926 THIS IN CASE, WE ARE LOOKING AT 1028 00:35:25,926 --> 00:35:28,028 AN E3 LIGASE, AND IT'S A MODULAR 1029 00:35:28,028 --> 00:35:31,732 PROTEIN AND HAS THESE W 1030 00:35:31,732 --> 00:35:33,033 W-DOMAINS THAT ALLOW IT TO BIND 1031 00:35:33,033 --> 00:35:33,901 TO THE CELL. 1032 00:35:33,901 --> 00:35:36,136 HAS A C2 DOMAIN THAT ATTACHES 1033 00:35:36,136 --> 00:35:40,207 TOLET MEMBRANE, BUT THE BUSINESS 1034 00:35:40,207 --> 00:35:42,843 END IS THE HECTODOMAIN WHICH IS 1035 00:35:42,843 --> 00:35:45,179 THE ENZYMATIC E3 LIGASE. 1036 00:35:45,179 --> 00:35:47,081 SO WE HIJACKED THE ENZYMATIC 1037 00:35:47,081 --> 00:35:49,149 PARTS, GET RID OF ALL THE OTHER 1038 00:35:49,149 --> 00:35:50,784 PARTS AND FUSE IT TO OUR NANO 1039 00:35:50,784 --> 00:35:51,018 BODY. 1040 00:35:51,018 --> 00:35:54,521 SO IN THIS WAY, IF WE EXPRESS 1041 00:35:54,521 --> 00:35:56,523 THE TAG NANO BODY WITH A CHANNEL 1042 00:35:56,523 --> 00:36:00,094 COMPLEX, MADE UP OF THE COMPLEX, 1043 00:36:00,094 --> 00:36:02,830 THE IDEA IS THAT THE NANO BODY 1044 00:36:02,830 --> 00:36:05,866 WOULD FIND A SUBUNIT AND CAUSE A 1045 00:36:05,866 --> 00:36:06,734 LOCALLAL UBIQUITIN OF THE 1046 00:36:06,734 --> 00:36:08,002 CHANNEL AND WE PREDICT THAD 1047 00:36:08,002 --> 00:36:10,304 WOULD CAUSE THE CHANNEL TO BE 1048 00:36:10,304 --> 00:36:11,372 DETAINED AND DEGRADED AS WELL. 1049 00:36:11,372 --> 00:36:14,541 SO WE WILL USE THE SAME FLO 1050 00:36:14,541 --> 00:36:15,843 CYTOMETRY ASSAY TO GET THESE TO 1051 00:36:15,843 --> 00:36:18,979 WORK IN THE CALCIUM CHANNELS. 1052 00:36:18,979 --> 00:36:20,247 SO WHEN WE EXPRESS THE CHANNEL 1053 00:36:20,247 --> 00:36:21,915 WITH JUST THE NAKED NANO BODY 1054 00:36:21,915 --> 00:36:24,218 FOR OUR CONTROL, YOU SEE A LOT 1055 00:36:24,218 --> 00:36:26,387 OF REG SIGNALS OF THE CELL 1056 00:36:26,387 --> 00:36:27,621 SURFACE IN YFP POSITIVE CELLS, 1057 00:36:27,621 --> 00:36:30,891 IF YOU ARM THE NANO BODY WITH A 1058 00:36:30,891 --> 00:36:33,927 HEX DOMAIN FROM THAT, THAT 1059 00:36:33,927 --> 00:36:35,262 SIGNAL IS ERASED. 1060 00:36:35,262 --> 00:36:36,797 INTERESTINGLY YOU CAN SEE THAT 1061 00:36:36,797 --> 00:36:38,866 THE PROTEIN IS DEGREATED, IT 1062 00:36:38,866 --> 00:36:40,601 DOESN'T GET TO THE CELL SURFACE, 1063 00:36:40,601 --> 00:36:42,569 AND THIS IS REALLY DEPENDENT ON 1064 00:36:42,569 --> 00:36:46,106 INFORMATION FWE MAKE IT 1065 00:36:46,106 --> 00:36:47,041 CATALYTICALLY DEAD VERSION OF 1066 00:36:47,041 --> 00:36:49,143 THE HEX DOMAIN, THE CHANNELS ARE 1067 00:36:49,143 --> 00:36:53,414 AT THE CELL SURFACE. 1068 00:36:53,414 --> 00:36:55,382 AS YOU EXPECT FROM THE 1069 00:36:55,382 --> 00:36:56,450 CHANNELSOT CELL SURFACE, MEASURE 1070 00:36:56,450 --> 00:36:56,950 THE CURRENT. 1071 00:36:56,950 --> 00:37:00,354 THIS IS 1 OF THESE IMPORTANT 1072 00:37:00,354 --> 00:37:01,055 CALCIUM CHANNEL PROHIBITOR, 1073 00:37:01,055 --> 00:37:02,256 PROFOUNDLY MOVES ALL THE OTHER 1074 00:37:02,256 --> 00:37:03,524 CURRENT. 1075 00:37:03,524 --> 00:37:05,025 THERE ARE MULTIPLE ISOFORMS OF 1076 00:37:05,025 --> 00:37:06,527 THESE FOR THE CALCIUM CHANNELS 1077 00:37:06,527 --> 00:37:10,397 THAT ARE IN THE BETA SUBUNIT, 1078 00:37:10,397 --> 00:37:11,765 AND THIS MOLECULE WAS VERY 1079 00:37:11,765 --> 00:37:13,934 EFFECTIVE FOR ALL OF THEM. 1080 00:37:13,934 --> 00:37:16,670 WE CALLED THEM CAPPA BLAITA AND 1081 00:37:16,670 --> 00:37:20,774 THIS IS WHAT TRAVIS IMAGINED IN 1082 00:37:20,774 --> 00:37:23,711 THE LLAMA WITH THE TERMINATOR. 1083 00:37:23,711 --> 00:37:25,512 SO WHAT IS IT GOOD FOR, 1 AREA 1084 00:37:25,512 --> 00:37:30,184 WE THOUGHT MIGHT BE USEFUL FOR 1085 00:37:30,184 --> 00:37:31,952 IS NEUROPATHIC PAIN CAUSED BY 1086 00:37:31,952 --> 00:37:33,921 LESIONS FROM THE NERVOUS SYSTEM, 1087 00:37:33,921 --> 00:37:34,888 DIABETES, NERVE INJURY, CHEMO 1088 00:37:34,888 --> 00:37:37,224 THERAPY AND SO FORTH, AFFECTS 1089 00:37:37,224 --> 00:37:38,359 MILLIONS OF PEOPLE. 1090 00:37:38,359 --> 00:37:40,194 IT'S A REAL MEDICAL NEED BECAUSE 1091 00:37:40,194 --> 00:37:42,029 IN FIRST AND SECOND LINE 1092 00:37:42,029 --> 00:37:45,299 MEDICATIONS ARE OFTEN INEECTIVE 1093 00:37:45,299 --> 00:37:46,600 OR HAVE VERY SERIES SIDE 1094 00:37:46,600 --> 00:37:48,535 EFFECTS, THE REASON WHY WE THINK 1095 00:37:48,535 --> 00:37:50,037 IT COULD BE USEFUL HERE IS 1096 00:37:50,037 --> 00:37:55,275 CONCENTRATION FROM THE CIRCUITRY 1097 00:37:55,275 --> 00:37:58,545 FOR SOPHISTICATED MAT O SENSORY 1098 00:37:58,545 --> 00:37:58,746 PAIN. 1099 00:37:58,746 --> 00:38:03,684 AND THAT SIGNAL HAS TO BE 1100 00:38:03,684 --> 00:38:04,418 TRANSFERRED, TO THE CENTRAL 1101 00:38:04,418 --> 00:38:05,686 NERVOUS SYSTEM WHERE YOU CAN 1102 00:38:05,686 --> 00:38:07,521 DECODE IT AS PAIN. 1103 00:38:07,521 --> 00:38:09,490 THE CENTRAL THING HERE IS THAT 1104 00:38:09,490 --> 00:38:11,325 THAT TRANSFER OF SIGNAL IS 1105 00:38:11,325 --> 00:38:12,526 DEPENDENT ON THESE CALCIUM 1106 00:38:12,526 --> 00:38:18,665 CHANNELS SO THE IDEA IS IF WE 1107 00:38:18,665 --> 00:38:19,633 COULD EXPRESS THAT FROM 1108 00:38:19,633 --> 00:38:21,335 HAPPENING HERE WE COULD PREVENT 1109 00:38:21,335 --> 00:38:22,503 IT FROM TAKING ROOT. 1110 00:38:22,503 --> 00:38:24,605 SO IF YOU EXPRESS THE NEURONS IT 1111 00:38:24,605 --> 00:38:26,106 COULD WEAPON OUT THE CURRENT. 1112 00:38:26,106 --> 00:38:29,777 SO WITH THAT IN HAND, THEN WE 1113 00:38:29,777 --> 00:38:31,879 LINKED UP WITH MY COLLEAGUE WHO 1114 00:38:31,879 --> 00:38:33,113 STUDIES ANIMAL MODELS OF PAIN 1115 00:38:33,113 --> 00:38:35,983 AND HERE'S THE EXPERIMENTAL 1116 00:38:35,983 --> 00:38:36,417 PARADIGM. 1117 00:38:36,417 --> 00:38:37,151 TWENTY-EIGHT DAYS BEFORE NERVE 1118 00:38:37,151 --> 00:38:38,752 INJURY WE CAN INJECT INTO THE 1119 00:38:38,752 --> 00:38:42,990 HIND PAW OF A MOUSE EITHER A 1120 00:38:42,990 --> 00:38:46,126 CONTROL AAV, 9 EXPRESSING 1121 00:38:46,126 --> 00:38:51,532 TDTOMATO, OR 1 THAT EXPRESSES 1122 00:38:51,532 --> 00:38:57,671 KAVABLATOR, AND THEN LOOK AT 1123 00:38:57,671 --> 00:38:57,871 PAIN. 1124 00:38:57,871 --> 00:39:01,642 SO 3 DAYS AFTER THE INJURY MODEL 1125 00:39:01,642 --> 00:39:03,277 THESE ANIMALS DEVELOP MECHANICAL 1126 00:39:03,277 --> 00:39:05,012 ALOE DEANIA WHICH YOU SEE AS A 1127 00:39:05,012 --> 00:39:07,014 DECREASE IN THE PAW WITH THE 1128 00:39:07,014 --> 00:39:08,715 THRESHOLD AND RESPONSE TO A 1129 00:39:08,715 --> 00:39:12,219 STIMULUS THAT WAS INITIALLY 1130 00:39:12,219 --> 00:39:12,686 [INDISCERNIBLE]. 1131 00:39:12,686 --> 00:39:13,987 WHAT WE WERE GRATIFIED TO SEE 1132 00:39:13,987 --> 00:39:18,692 WAS THE ANIMAL THAT HAD 1133 00:39:18,692 --> 00:39:21,895 KAVATBLATOR EXPRESSED LESS PAIN 1134 00:39:21,895 --> 00:39:23,030 AND THAT PERSISTENT, WE'VE GONE 1135 00:39:23,030 --> 00:39:26,800 UP TO 2 YEARS, AFTER INYEKS AND 1136 00:39:26,800 --> 00:39:30,737 SEEN THAT THE CAVABLATOR IS 1137 00:39:30,737 --> 00:39:32,272 EXPRESSED, SO WE COULD GET THIS 1138 00:39:32,272 --> 00:39:36,910 RIGHT, DO A SINGLE INYEKS AND -- 1139 00:39:36,910 --> 00:39:39,613 INJECTION AND GET YEARS OF PAIN 1140 00:39:39,613 --> 00:39:42,316 RELIEF WITHOUT ANY SED EFFECTS 1141 00:39:42,316 --> 00:39:46,620 AND THE EXTENT OF CAVABLATOR, 1142 00:39:46,620 --> 00:39:49,156 INCLUDES NASAA.COMMIA, HEAT AND 1143 00:39:49,156 --> 00:39:49,389 COLD. 1144 00:39:49,389 --> 00:39:53,627 SO AFTER WE PUBLISHED THIS, 1145 00:39:53,627 --> 00:39:55,195 [INDISCERNIBLE] AND HIS 1146 00:39:55,195 --> 00:39:56,930 COLLEAGUE PUBLISHED THIS IN THE 1147 00:39:56,930 --> 00:40:00,968 SPOTLIGHT IN TIPS, THE CALCIUM 1148 00:40:00,968 --> 00:40:06,373 CHANNEL TERMINATOR H 1149 00:40:06,373 --> 00:40:06,807 ASTA LA VISTA PAIN. 1150 00:40:06,807 --> 00:40:09,409 IF YOU LOOK AT IT, EVEN THOUGH 1151 00:40:09,409 --> 00:40:11,678 WE HAD LESS PAIN THAN THE 1152 00:40:11,678 --> 00:40:15,516 CONTROL, WE WERE STILL PROVING 1153 00:40:15,516 --> 00:40:18,151 SOME PAIN RELATIVE TO THE 1154 00:40:18,151 --> 00:40:19,520 WILD-TYPE CONTROL HERE SO THE 1155 00:40:19,520 --> 00:40:21,321 QUESTION WE HAD WAS WHETHER WE 1156 00:40:21,321 --> 00:40:24,825 COULD PUSH THIS UP TO CAUSE, YOU 1157 00:40:24,825 --> 00:40:26,393 KNOW LESS PAIN WITHOUT ANYTHING 1158 00:40:26,393 --> 00:40:28,195 HAPPENING AND 1 THING THAT A 1159 00:40:28,195 --> 00:40:30,230 PHARMACOLOGIST WILL TELL YOU IS 1160 00:40:30,230 --> 00:40:34,034 YEAH, JUST INJECT MORE DRUG BUT 1161 00:40:34,034 --> 00:40:35,536 GIVEN WHAT CAVABLATOR DOES WE 1162 00:40:35,536 --> 00:40:37,037 WERE WORRIED ABOUT IT SPILLING 1163 00:40:37,037 --> 00:40:38,639 OVER INTO OTHER ORGANS 1164 00:40:38,639 --> 00:40:39,573 ESPECIALLY THE HEART. 1165 00:40:39,573 --> 00:40:42,109 SO THE HEART IF YOU RECORD 1166 00:40:42,109 --> 00:40:43,944 CURRENTS, THE INWARD CURRENT 1167 00:40:43,944 --> 00:40:45,445 UNDER CONTROL CONDITIONS IS VERY 1168 00:40:45,445 --> 00:40:46,947 ESSENTIAL FOR YOU TO HAVE A 1169 00:40:46,947 --> 00:40:50,284 HEART BEAT THIS, IS WHAT 1170 00:40:50,284 --> 00:40:52,019 CAVABLATOR DOES WHEN IT GETS 1171 00:40:52,019 --> 00:40:55,822 INTO THE HEART, WE ARE VERY 1172 00:40:55,822 --> 00:40:58,025 AWARE THAT IF WE KEEP INJECTED 1173 00:40:58,025 --> 00:41:01,995 THIS WE WOULD BE SAYING 1174 00:41:01,995 --> 00:41:03,263 HASTA LA VISTA, POETIC THE 1175 00:41:03,263 --> 00:41:04,364 PATIENT. 1176 00:41:04,364 --> 00:41:06,366 SO HOW DO WE DO THIS WITHOUT 1177 00:41:06,366 --> 00:41:06,900 KILLING THE PATIENT? 1178 00:41:06,900 --> 00:41:08,669 SO 1 OF THE THINGS IS THAT THE 1179 00:41:08,669 --> 00:41:09,903 DIFFERENT BETA ISOFORMS OF THE 1180 00:41:09,903 --> 00:41:10,971 CHANNEL HAVE DIFFERENT TISSUE 1181 00:41:10,971 --> 00:41:11,738 EXPRESSION. 1182 00:41:11,738 --> 00:41:14,408 SO FOR EXAMPLE, THE HEART 1183 00:41:14,408 --> 00:41:15,375 EXPRESSES MOSTLY BETA 2, RIGHT? 1184 00:41:15,375 --> 00:41:22,749 AND O IF YOU COULD MAKE OTHER 1185 00:41:22,749 --> 00:41:24,251 KAVABLATOR ISOFORMS THAT COULD 1186 00:41:24,251 --> 00:41:26,353 BE AN INHIBITOR POTENTIALLY TO 1187 00:41:26,353 --> 00:41:30,157 GET TAKEN--THEY RELIEVE. 1188 00:41:30,157 --> 00:41:31,792 SO TRAVIS SCREENED THE SAMPLE 1189 00:41:31,792 --> 00:41:33,994 FROM THE LLAMA AND WE FOUND THAT 1190 00:41:33,994 --> 00:41:35,729 THE ISOFORM BAND AND NONE THE 1191 00:41:35,729 --> 00:41:38,565 OTHER BETAS AND WE COULD CAN IF 1192 00:41:38,565 --> 00:41:40,233 WE COULD MAKE A INHIBITOR BASED 1193 00:41:40,233 --> 00:41:40,801 ON THAT. 1194 00:41:40,801 --> 00:41:43,470 INDEED IF YOU FUSE THE DOMAIN TO 1195 00:41:43,470 --> 00:41:45,439 THE NANO BODY, YOU GET CHANNELS 1196 00:41:45,439 --> 00:41:47,474 THAT CONSITUTE THE BETA 1 AND 1197 00:41:47,474 --> 00:41:48,342 COMPLETE ELIMINATION OF THE 1198 00:41:48,342 --> 00:41:50,611 CURRENT AND THERE WERE TOTALLY 1199 00:41:50,611 --> 00:41:54,348 INERT AGAINST OTHER ISOFORMS OF 1200 00:41:54,348 --> 00:41:55,182 THE BETA SUBUNIT. 1201 00:41:55,182 --> 00:41:57,417 SO THERE'S A REAL NEW PARADIGM 1202 00:41:57,417 --> 00:42:00,020 FOR INHIBITING THE ION CHANNEL 1203 00:42:00,020 --> 00:42:07,461 BASED ON THE ISOFORM SUBTYPE OF 1204 00:42:07,461 --> 00:42:07,995 THE AUXILLARY SUBUNIT. 1205 00:42:07,995 --> 00:42:09,796 THE OTHER THING WE WERE WORRIED 1206 00:42:09,796 --> 00:42:14,468 ABOUT WITH THE CAVABLATOR, IF 1207 00:42:14,468 --> 00:42:16,737 YOU INJECT TOO, EVEN IF THE 1208 00:42:16,737 --> 00:42:18,672 EXPRESS ENOUGH OF IT WILL HAVE 1209 00:42:18,672 --> 00:42:19,339 OFF TARGET CENTER FOR EXCELLENCE 1210 00:42:19,339 --> 00:42:23,110 ON AGINGS SO WE WERE THINKING OF 1211 00:42:23,110 --> 00:42:23,744 WAYS TO AVOID. 1212 00:42:23,744 --> 00:42:27,314 A AND WE TOOK INSPIRATION FROM 1213 00:42:27,314 --> 00:42:30,617 THE PROTAX FIELD, AND PROTAX ARE 1214 00:42:30,617 --> 00:42:31,585 BINDERS, SMALL MOLECULES THAT 1215 00:42:31,585 --> 00:42:33,520 HAVE 1 PART THAT BINDS A LIGAND 1216 00:42:33,520 --> 00:42:35,422 AND ANOTHER PART THAT BINDS AN 1217 00:42:35,422 --> 00:42:38,058 E3 LIGASE, AND THE IDEA THAT IF 1218 00:42:38,058 --> 00:42:40,127 YOU GET INSIDE THE CELL IT 1219 00:42:40,127 --> 00:42:42,596 RECRUITS THE LIGASE TARGET AND 1220 00:42:42,596 --> 00:42:43,864 GETS DEGRADED AND WONDERED IF WE 1221 00:42:43,864 --> 00:42:46,433 COULD DO THE SAME THING WITH THE 1222 00:42:46,433 --> 00:42:48,902 NONVALENTINEDDENT NANO BODIES WE 1223 00:42:48,902 --> 00:42:50,470 CALL DIVAS, AND NOW GET THROUGH 1224 00:42:50,470 --> 00:42:51,405 TO BE RECRUITED. 1225 00:42:51,405 --> 00:42:52,572 THE ADVANTAGE OF A SMALL 1226 00:42:52,572 --> 00:42:53,974 MOLECULE OF COURSE IS THAT NOW 1227 00:42:53,974 --> 00:42:55,242 YOU CAN EXPRESS THIS IN 1228 00:42:55,242 --> 00:42:57,144 PARTICULAR CELLS THAT YOU ARE 1229 00:42:57,144 --> 00:42:59,012 INTERESTED IN INHIBITING IN THE, 1230 00:42:59,012 --> 00:43:00,447 SO IN ORDER TO DO THAT, THE 1231 00:43:00,447 --> 00:43:01,615 FIRST QUESTION WE HAD TO ADDRESS 1232 00:43:01,615 --> 00:43:03,684 WAS, YOU KNOW WHAT IS THE BEST 1233 00:43:03,684 --> 00:43:05,786 E3 LIGASE TO TARGET FOR ION 1234 00:43:05,786 --> 00:43:08,088 CHANNELS AND LOOKING AT THE 1235 00:43:08,088 --> 00:43:10,290 LITERATURE IT SEEMS LIKE NET 1236 00:43:10,290 --> 00:43:12,259 FORAND TO REALLY WELL KNOWN TO 1237 00:43:12,259 --> 00:43:13,460 REGULATE MULTIPLE ION CHANNELS. 1238 00:43:13,460 --> 00:43:15,929 SO THE QUESTION WAS WHETHER WE 1239 00:43:15,929 --> 00:43:21,134 COULD MAKE A RECRUIT NET 4 AND 1240 00:43:21,134 --> 00:43:22,769 NET 42, WHICH HAVEN'T BEEN USED 1241 00:43:22,769 --> 00:43:25,138 IN THE PROTAX FIELD TO INHIBIT 1242 00:43:25,138 --> 00:43:25,772 ION CHANNELS. 1243 00:43:25,772 --> 00:43:29,976 SO WITH THAT WE RAISED NANO 1244 00:43:29,976 --> 00:43:32,479 BODIES USING THE LIBRARY, 1245 00:43:32,479 --> 00:43:33,780 AGAINST THE HEK 42 DOMAIN. 1246 00:43:33,780 --> 00:43:39,319 WE FOUND 1 MAN NANO BODY C11 1247 00:43:39,319 --> 00:43:41,288 HERE THAT BINDS THEAT42 HECHT 1248 00:43:41,288 --> 00:43:44,925 4-2 DOMAIN AND IT BINDS A FULL 1249 00:43:44,925 --> 00:43:46,426 LENGTH IN HECHT-4-2 AS WELL. 1250 00:43:46,426 --> 00:43:52,065 AND IT OBTAIN A YIE O EM 1251 00:43:52,065 --> 00:43:53,533 STRUCTURE WITH OUR COLLEAGUE 1252 00:43:53,533 --> 00:43:55,435 OLIVER CLARK AND THE STUDENT 1253 00:43:55,435 --> 00:43:55,802 [INDISCERNIBLE]. 1254 00:43:55,802 --> 00:43:57,270 AND FOUND WHERE IT BINDS SO WE 1255 00:43:57,270 --> 00:43:58,672 COULD LOOK AT THE INTERFACE AND 1256 00:43:58,672 --> 00:43:59,773 WHAT WAS IMPORTANT ABOUT THIS IS 1257 00:43:59,773 --> 00:44:02,609 THAT WE WANTED A NANO BODY THAT 1258 00:44:02,609 --> 00:44:04,878 WOULD BIND THE HECHT DOMAIN 1259 00:44:04,878 --> 00:44:06,146 WITHOUT DISRUPTING ITS FUNCTION. 1260 00:44:06,146 --> 00:44:08,582 AND BASED ON WHERE IT'S BIND -- 1261 00:44:08,582 --> 00:44:11,418 WHERE THE NANO BODY BIND, WE 1262 00:44:11,418 --> 00:44:13,720 COULD INFER THAT IT DID IRPT 1263 00:44:13,720 --> 00:44:14,988 FEAR WITH ANY OF THE OTHER PARTS 1264 00:44:14,988 --> 00:44:17,624 THAT WERE IMPORTANT FOR THE 1265 00:44:17,624 --> 00:44:20,026 CATALYTIC ACTIVITY THE ENZYME, 1266 00:44:20,026 --> 00:44:21,995 THE CATALYTIC CYSTINE SITE, THE 1267 00:44:21,995 --> 00:44:23,663 E2 BINDING SITE AS WELL AS 1268 00:44:23,663 --> 00:44:25,432 SOPHISTICATED CALLED CRUCIATE 1269 00:44:25,432 --> 00:44:28,268 LIGAMENT CIBOL WITTEN SITE THAT 1270 00:44:28,268 --> 00:44:29,870 WAS IMPORTANT FOR POLYUBIQUITIN 1271 00:44:29,870 --> 00:44:30,270 CHAIN INFORMATION. 1272 00:44:30,270 --> 00:44:32,105 ALL RIGHT, SO NOW THAT WE HAD A 1273 00:44:32,105 --> 00:44:35,275 NANO BODY HERE WE COULD THEN SET 1274 00:44:35,275 --> 00:44:36,943 UP THIS SAME EXPERIMENT. 1275 00:44:36,943 --> 00:44:40,013 WE HAD A CHANNEL AND YOU HAVE A 1276 00:44:40,013 --> 00:44:41,848 3 CONDITIONS EITHER NAKED NANO 1277 00:44:41,848 --> 00:44:45,285 BODY TO THE BETA SUBUNIT OR YOU 1278 00:44:45,285 --> 00:44:46,353 CAN OVEREXPRESS NET 4 TO SEE IF 1279 00:44:46,353 --> 00:44:47,554 THE CHANNEL IS REGULATE BIDE 1280 00:44:47,554 --> 00:44:49,923 THAT OR YOU COULD RECRUIT 1281 00:44:49,923 --> 00:44:51,658 ENDOGENOUS M4 TO THIS 1282 00:44:51,658 --> 00:44:52,292 DISCIPLINARY VALENTINEDENT NANO 1283 00:44:52,292 --> 00:44:55,662 BODY OR THIS DIVA, OKAY. 1284 00:44:55,662 --> 00:44:58,565 SO THE EXPERIMENT, SO, THE 1285 00:44:58,565 --> 00:45:00,167 WILD-TYPE, IF YOU OVEREXPRESS 1286 00:45:00,167 --> 00:45:02,702 IT, THE RED SHOWS IF YOU GET IT, 1287 00:45:02,702 --> 00:45:04,437 YOU GET ABOUT 60% INHIBITION OF 1288 00:45:04,437 --> 00:45:05,839 THE CURRENT AND JUST PUTTING IN 1289 00:45:05,839 --> 00:45:08,842 THE DIVA TO RECRUIT ENDOGENOUS 1290 00:45:08,842 --> 00:45:10,310 NET 4-2 HAS THE SAME FUNCTIONAL 1291 00:45:10,310 --> 00:45:11,745 EFFECT AS THAT SO THAT WAS 1292 00:45:11,745 --> 00:45:14,581 HIGHLY EFFECTIVE TO DO THAT. 1293 00:45:14,581 --> 00:45:17,517 WE MADE AN AAV AND PUT THAT INTO 1294 00:45:17,517 --> 00:45:20,220 THE NEURONS AND INDEED, 1295 00:45:20,220 --> 00:45:21,054 RECRUITING DISCIPLINARY 1296 00:45:21,054 --> 00:45:23,256 VALENTINEDENT NANO BODIES, 1297 00:45:23,256 --> 00:45:24,457 RECRUITING ENDOGENOUS E32, AND 1298 00:45:24,457 --> 00:45:25,959 ENDOGENOUS CHANNELS IN THE 1299 00:45:25,959 --> 00:45:27,994 NEURONS ALSO CAUSE ABOUT A 70% 1300 00:45:27,994 --> 00:45:29,529 INHIBITION OF THE CURRENT. 1301 00:45:29,529 --> 00:45:30,730 ALL RIGHT, SO I WILL SUMMARIZE 1302 00:45:30,730 --> 00:45:32,699 WHAT I TOLD YOU THIS AFTERNOON, 1303 00:45:32,699 --> 00:45:37,170 I SPOKE ABOUT THE FIRST PART, AN 1304 00:45:37,170 --> 00:45:38,572 INCUB APPROACH, WAY TO RESCUE 1305 00:45:38,572 --> 00:45:40,774 AND MAKE MORE ION CHANNELS OR 1306 00:45:40,774 --> 00:45:44,144 ANY OTHER PROTEIN USING THE 1307 00:45:44,144 --> 00:45:44,911 TARGETED DECIBOL WITTENATION 1308 00:45:44,911 --> 00:45:47,147 STRATEGY, I SPOKE TO YOU ABOUT 1309 00:45:47,147 --> 00:45:49,382 THE CHALSIUM CHANNEL INHIBITOR, 1310 00:45:49,382 --> 00:45:52,118 THAT WE INITIALLY USED TO 1311 00:45:52,118 --> 00:45:53,753 INDICATE COULD BE POTENTIALLY 1312 00:45:53,753 --> 00:45:56,423 USED FOR LONG LASTING RELIEF OF 1313 00:45:56,423 --> 00:45:56,656 PAIN. 1314 00:45:56,656 --> 00:45:58,625 THE CHISELS ALLOW YOU MORE 1315 00:45:58,625 --> 00:46:01,795 ISOFORM SELECTIVITY AND SO THEY 1316 00:46:01,795 --> 00:46:03,763 DON'T HOPEFULLY KILL ANYBODY, 1317 00:46:03,763 --> 00:46:05,498 AND THEN THE DIVAS WE THINK CAN 1318 00:46:05,498 --> 00:46:08,635 BE A VERY GENERALIZED APPROACH 1319 00:46:08,635 --> 00:46:11,104 TO MAKING INHIBITORS FOR ANY ION 1320 00:46:11,104 --> 00:46:13,273 CHANNEL OR FOR OTHER PROTEINS AS 1321 00:46:13,273 --> 00:46:13,473 WELL. 1322 00:46:13,473 --> 00:46:15,542 SO THE FINAL THING IS I HAVE A 1323 00:46:15,542 --> 00:46:16,743 CONFESSION TO MAKE, I DIDN'T DO 1324 00:46:16,743 --> 00:46:19,379 ANY OF THIS WORK MYSELF, DONE BY 1325 00:46:19,379 --> 00:46:21,514 TALENTED GROUP OF STUDENTS AND 1326 00:46:21,514 --> 00:46:22,382 POST DOCS. 1327 00:46:22,382 --> 00:46:26,620 I MENTIONED THEM ALONG THE WAY 1328 00:46:26,620 --> 00:46:27,821 AND CERTAINLY 1 OF THE BEST 1329 00:46:27,821 --> 00:46:29,623 PARTS OF BEING A PROFESSOR AND 1330 00:46:29,623 --> 00:46:30,757 WORKING WITH REALLY SMART AND 1331 00:46:30,757 --> 00:46:31,725 TALENTED PEOPLE. 1332 00:46:31,725 --> 00:46:34,127 I HAVE A NUMBER OF WONDERFUL 1333 00:46:34,127 --> 00:46:35,095 COLLABORATORS AND ALSO, FINANCE 1334 00:46:35,095 --> 00:46:37,931 ALSO SO THANK YOU FOR LISTENING. 1335 00:46:37,931 --> 00:46:48,341 READY TO TAKE QUESTIONS. 1336 00:46:54,481 --> 00:46:54,748 [ APPLAUSE ] 1337 00:46:54,748 --> 00:46:56,116 >> THANK YOU FOR THE TALK. 1338 00:46:56,116 --> 00:46:56,383 QUESTIONS? 1339 00:46:56,383 --> 00:46:59,552 I WILL GET STARTED SO IN YOUR 1340 00:46:59,552 --> 00:47:01,755 INTRODUCTION, YOU TALKED ABOUT 1341 00:47:01,755 --> 00:47:05,325 LONG QT AND CFTR AND EPILEPSY, 1342 00:47:05,325 --> 00:47:07,594 DIDN'T LEE TALK ABOUT EPILEPSY 1343 00:47:07,594 --> 00:47:13,633 IN THE TALK, I'M INTERESTED, I 1344 00:47:13,633 --> 00:47:15,802 MEAN JUST THERE'S SO POTENTIALLY 1345 00:47:15,802 --> 00:47:16,937 CELL TYPE SPECIFICITY IN,A 1346 00:47:16,937 --> 00:47:18,204 DITION TO SUBTYPE, HAVE YOU 1347 00:47:18,204 --> 00:47:20,507 THOUGHT ABOUT ADDING CELL TYPE 1348 00:47:20,507 --> 00:47:21,708 INTO THIS? 1349 00:47:21,708 --> 00:47:24,344 AND HOW YOU COULD ADDRESS THAT? 1350 00:47:24,344 --> 00:47:26,346 >> SO, YEAH, 1 OF OUR -- A 1351 00:47:26,346 --> 00:47:30,750 COUPLE OF OUR PROJECTS ARE 1352 00:47:30,750 --> 00:47:32,819 DEALING WITH EPILEPSY AND 1353 00:47:32,819 --> 00:47:39,559 [INDISCERNIBLE] FOR EXAMPLE, IT 1354 00:47:39,559 --> 00:47:41,428 HAPLOSUFFICIENCY THAT OCCURS IN 1355 00:47:41,428 --> 00:47:43,163 NEURONS SO WE'RE APPLYING THIS 1356 00:47:43,163 --> 00:47:44,698 APPROACH TO ENHANCE THE EXPRESS 1357 00:47:44,698 --> 00:47:46,266 OF THE SODIUM CHANNELS IN THE 1358 00:47:46,266 --> 00:47:48,668 NEURONS FOR EXAMPLE, AND I THINK 1359 00:47:48,668 --> 00:47:51,304 THE -- HAVING A GENETICALLY 1360 00:47:51,304 --> 00:47:52,605 ENCODED APPROACH ALLOWS TO YOU 1361 00:47:52,605 --> 00:47:53,940 DO THAT SO CAN YOU DO IT IN A 1362 00:47:53,940 --> 00:48:02,215 SPECIFIC WAY YOU WANT TO. 1363 00:48:02,215 --> 00:48:03,683 >> [INAUDIBLE QUESTION FROM 1364 00:48:03,683 --> 00:48:08,521 AUDIENCE ] 1365 00:48:08,521 --> 00:48:09,422 >> REALLY GREAT QUESTION, SO 1366 00:48:09,422 --> 00:48:10,690 WHEN WE FIRST STARTED THIS 1367 00:48:10,690 --> 00:48:12,158 APPROACH, ALL I KNEW ABOUT 1368 00:48:12,158 --> 00:48:14,461 UBIQUITIN IS THAT YOU, YOU KNOW 1369 00:48:14,461 --> 00:48:16,496 IT DEGRADES THINGS SO WE WERE 1370 00:48:16,496 --> 00:48:18,298 LOOKING FOR DEGRADATION, AND WE 1371 00:48:18,298 --> 00:48:20,700 DID IT AT FIRST FOR A POTASSIUM 1372 00:48:20,700 --> 00:48:22,802 CHANNEL AND ALSO FOR THE CALCIUM 1373 00:48:22,802 --> 00:48:24,304 CHANNEL SO OUR FIRST WE WERE 1374 00:48:24,304 --> 00:48:26,172 JUST LOOKING FOR THE TOTAL 1375 00:48:26,172 --> 00:48:27,107 AMOUNT OF PROTEIN, DIDN'T 1376 00:48:27,107 --> 00:48:27,340 CHANGE. 1377 00:48:27,340 --> 00:48:30,410 HOWEVER, WHEN YOU LOOK AT THE 1378 00:48:30,410 --> 00:48:32,345 AMOUNT OF BURDEN ON THE CELL 1379 00:48:32,345 --> 00:48:35,281 SURFACE THAT WAS TOTALLY GONE, 1380 00:48:35,281 --> 00:48:36,916 SO IT TURNS OUT THAT WHAT THIS 1381 00:48:36,916 --> 00:48:38,084 WAS DOING IS CAUSING THE 1382 00:48:38,084 --> 00:48:39,619 CHANNELS TO BE TRAPPED INSIDE 1383 00:48:39,619 --> 00:48:42,255 THE CELL, RATHER THAN DEGRADING 1384 00:48:42,255 --> 00:48:44,657 THEM, SO IT WAS JUST NOT 1385 00:48:44,657 --> 00:48:45,692 UBIQUITIN PER SE, THAT'S 1386 00:48:45,692 --> 00:48:46,760 IMPORTANT AND I THINK THE SECOND 1387 00:48:46,760 --> 00:48:48,294 PART OF THE STORY WAS TELLING 1388 00:48:48,294 --> 00:48:50,830 THAT, RIGHT IN BECAUSE IT ALSO 1389 00:48:50,830 --> 00:48:51,798 REGULATES DIFFERENT ASPECTS OF 1390 00:48:51,798 --> 00:48:53,600 TRAFFICKING AS WELL, SO WE'RE 1391 00:48:53,600 --> 00:48:55,235 INTERESTED IN LOOKING AT THESE 1392 00:48:55,235 --> 00:48:56,436 SIGNALS THAT REGULATE EVERYTHING 1393 00:48:56,436 --> 00:48:59,072 FROM THE TRAFFICKING PATHWAY TO 1394 00:48:59,072 --> 00:49:00,173 WHAT ACTIVATES RESPONSIBLE FOR 1395 00:49:00,173 --> 00:49:03,410 DEGRADATION, WE THINK IT WILL BE 1396 00:49:03,410 --> 00:49:04,177 DIFFERENT TYPES OF UBIQUITIN 1397 00:49:04,177 --> 00:49:05,078 CHAINS AND SO FORTH. 1398 00:49:05,078 --> 00:49:12,285 >> SO A QUICK FOLLOW UP 1399 00:49:12,285 --> 00:49:13,620 [INDISCERNIBLE] SURFACE AND 1400 00:49:13,620 --> 00:49:17,123 APPROXIMATELY CHANNELIZE 1401 00:49:17,123 --> 00:49:17,757 TARGETING [INDISCERNIBLE]? 1402 00:49:17,757 --> 00:49:18,792 >> I THINK IT'S A COMBINATION OF 1403 00:49:18,792 --> 00:49:19,526 THE 2. 1404 00:49:19,526 --> 00:49:21,261 SO FOR THE SECOND STORY, FOR 1405 00:49:21,261 --> 00:49:23,229 EXAMPLE, WHERE WE HAD A NANO 1406 00:49:23,229 --> 00:49:25,298 BODY TIED TO THE YFP, SO IT'S 1407 00:49:25,298 --> 00:49:28,668 ALWAYS GOING TO BE THE CYTOSOLIC 1408 00:49:28,668 --> 00:49:32,972 PART AND YOU LOOK AT THE RATE OF 1409 00:49:32,972 --> 00:49:35,008 FORWARD TRAFFICKING VERSUS RATE 1410 00:49:35,008 --> 00:49:36,543 OF -- YOU EFFECT BOTH THOSE 1411 00:49:36,543 --> 00:49:38,078 PROCESSES SO WE THINK YOU ARE 1412 00:49:38,078 --> 00:49:42,449 AFFECTING EVERYTHING IN THAT 1413 00:49:42,449 --> 00:49:42,682 SENSE. 1414 00:49:42,682 --> 00:49:43,183 >> QUESTIONS? 1415 00:49:43,183 --> 00:49:44,284 >> [INAUDIBLE QUESTION FROM 1416 00:49:44,284 --> 00:49:54,461 AUDIENCE ] 1417 00:50:01,301 --> 00:50:02,469 >> K63 PREDOMINANTLY. 1418 00:50:02,469 --> 00:50:04,404 MAKES, SENSE. 1419 00:50:04,404 --> 00:50:05,071 RIGHT. 1420 00:50:05,071 --> 00:50:09,943 WE DID SOME -- WE ALSO HAVE A 1421 00:50:09,943 --> 00:50:11,344 NUMBER OF LIGASES AND SOME ARE 1422 00:50:11,344 --> 00:50:14,481 ABLE TO DEGRADE MORE THAN 1423 00:50:14,481 --> 00:50:17,317 OTHERS. 1424 00:50:17,317 --> 00:50:17,984 THAT'S CONSISTENT. 1425 00:50:17,984 --> 00:50:19,752 >> [INAUDIBLE QUESTION FROM 1426 00:50:19,752 --> 00:50:20,253 AUDIENCE ] 1427 00:50:20,253 --> 00:50:21,855 >> THAT'S RIGHT, THAT'S RIGHT. 1428 00:50:21,855 --> 00:50:22,055 YEAH. 1429 00:50:22,055 --> 00:50:25,191 >> YEAH, SO, FOR SOME OF YOUR 1430 00:50:25,191 --> 00:50:27,227 RESIDENT EXPERIMENTS ESPECIALLY 1431 00:50:27,227 --> 00:50:31,531 FOR LQTS, THERE WAS AN F508 1432 00:50:31,531 --> 00:50:33,733 DELETION THAT YOU HAD, DID YOU 1433 00:50:33,733 --> 00:50:35,668 DO ANY PHYSIOLOGICAL ASSAYS TO 1434 00:50:35,668 --> 00:50:36,836 DETERMINE THE EFFECT OF THE PACE 1435 00:50:36,836 --> 00:50:40,039 ON THE HEART AFTER THOSE RESCUE 1436 00:50:40,039 --> 00:50:42,709 EXPERIMENTS, BUT I KNOW YOU HAD 1437 00:50:42,709 --> 00:50:44,310 A RESCUE EFFECT BE, WAS THERE 1438 00:50:44,310 --> 00:50:45,411 ANY EFFECT OF TACHYCARDIA OR 1439 00:50:45,411 --> 00:50:47,046 SOMETHING OF THAT NATURE? 1440 00:50:47,046 --> 00:50:49,182 >> YEAH, SO THE F5, I THINK THAT 1441 00:50:49,182 --> 00:50:50,917 WAS CFTR, THAT WAS THE CHLORIDE 1442 00:50:50,917 --> 00:51:00,693 CHANNEL SO THAT WAS THE CASE OF 1443 00:51:00,693 --> 00:51:01,327 THE CYSTIC FIBROSIS. 1444 00:51:01,327 --> 00:51:02,862 SO FOR THAT PARTICULAR CASE, I 1445 00:51:02,862 --> 00:51:04,464 THINK THE SUPER RESCUE WOULD 1446 00:51:04,464 --> 00:51:05,532 PROBABLY BE FINE BECAUSE, YOU 1447 00:51:05,532 --> 00:51:07,167 KNOW YOU JUST GET MORE HYDRATION 1448 00:51:07,167 --> 00:51:10,537 OF THE AIR WAYS BUT YOU KNOW 1449 00:51:10,537 --> 00:51:12,071 THERE ARE CERTAIN 1450 00:51:12,071 --> 00:51:13,339 CONSIDERATIONS, IF YOU -- THE 1451 00:51:13,339 --> 00:51:16,409 GENERAL QUESTION IS, CAN YOU 1452 00:51:16,409 --> 00:51:17,443 OVERRESCUE SOMETHING, RIGHT IN 1453 00:51:17,443 --> 00:51:18,845 WHAT WE FOUND IN GENERAL IS THAT 1454 00:51:18,845 --> 00:51:20,914 IT'S VERY HARD TO GET MORE THAN 1455 00:51:20,914 --> 00:51:22,682 2-3 FOLD INCREASE IN ANYTHING, 1456 00:51:22,682 --> 00:51:24,918 AND JUST LIKE ANY OTHER DRUG, 1457 00:51:24,918 --> 00:51:28,588 YOU WOULD HAVE TO TITRATE IT TO 1458 00:51:28,588 --> 00:51:32,225 A THERAPEUTIC SORT OF LEVEL. 1459 00:51:32,225 --> 00:51:35,061 THANK YOU. 1460 00:51:35,061 --> 00:51:35,662 >> BEAUTIFUL TALK. 1461 00:51:35,662 --> 00:51:39,132 ACTUALLY A QUESTION ABOUT THE 1462 00:51:39,132 --> 00:51:39,966 SAME PHYSIOLOGICAL THINGS YOU 1463 00:51:39,966 --> 00:51:40,700 HAD SHOWN BEFORE, IT WAS 1464 00:51:40,700 --> 00:51:41,868 INTERESTING THAT YOU SHOWED IT 1465 00:51:41,868 --> 00:51:44,170 WAS NOT ONLY A TRAFFICKING 1466 00:51:44,170 --> 00:51:47,340 DEFICIT BUT ALSO A DEFICIT IN 1467 00:51:47,340 --> 00:51:48,975 THE YOU KNOW TOTAL 1468 00:51:48,975 --> 00:51:50,009 [INDISCERNIBLE] REACHED THE 1469 00:51:50,009 --> 00:51:51,978 SURFACE AND SO, I GUESS WHEN I 1470 00:51:51,978 --> 00:51:53,713 THINK OF UBIQUITINNATION, I 1471 00:51:53,713 --> 00:51:57,817 THINK OF PROTECTING AGAINST 1472 00:51:57,817 --> 00:51:58,885 DEFECTIVE OR NONFUNCTIONING, 1473 00:51:58,885 --> 00:51:59,752 CHANNELS THAT DON'T FUNCTION 1474 00:51:59,752 --> 00:52:00,553 PROPERLY, DO YOU THINK YOU WOULD 1475 00:52:00,553 --> 00:52:02,755 BE ABLE TO SORT OF MAINTAIN THE 1476 00:52:02,755 --> 00:52:03,723 INTEGRITY OF THE CHANNEL THAT 1477 00:52:03,723 --> 00:52:06,626 MAKE ITS TO THE SURFACE IN 1478 00:52:06,626 --> 00:52:07,994 ADDITION TO INCREASING, YOU KNOW 1479 00:52:07,994 --> 00:52:09,529 BEYOND JUST INCREASING SURFACE 1480 00:52:09,529 --> 00:52:11,130 EXPRESSION, MAKING SURE THAT THE 1481 00:52:11,130 --> 00:52:13,433 1S THAT ARE FUNCTIONAL, IS THAT 1482 00:52:13,433 --> 00:52:15,101 SOMETHING WITH NANO BODY 1483 00:52:15,101 --> 00:52:16,869 TECHNOLOGY COULD DO? 1484 00:52:16,869 --> 00:52:19,272 >> YEAH, SO, MAYBE I'LL TAKE AN 1485 00:52:19,272 --> 00:52:21,140 EXAMPLE FROM THE 1486 00:52:21,140 --> 00:52:22,609 CYSTIC FIBROSIS, THE VERTEX 1487 00:52:22,609 --> 00:52:24,911 DRUG, IT'S A TRIPLE COMBINATION 1488 00:52:24,911 --> 00:52:25,278 THERAPY, RIGHT? 1489 00:52:25,278 --> 00:52:27,547 TWO OF THE DRUGS HELP THE 1490 00:52:27,547 --> 00:52:30,650 CHANNEL FOLD BETTER IN TRAFFIC 1491 00:52:30,650 --> 00:52:32,619 AND THIS IS SOMETHING WE FOUND 1492 00:52:32,619 --> 00:52:33,953 AS WELL. 1493 00:52:33,953 --> 00:52:35,521 FOR THE CYSTIC FIBROSIS CHANNEL, 1494 00:52:35,521 --> 00:52:37,657 WE JUST ADD THE ENDUB, WE CAN 1495 00:52:37,657 --> 00:52:39,692 RESCUE THE CURRENT, THE SURFACE 1496 00:52:39,692 --> 00:52:41,327 DENSITY BY ABOUT A HUNDRED 1497 00:52:41,327 --> 00:52:43,396 PERCENT BUT WHEN YOU MEASURE THE 1498 00:52:43,396 --> 00:52:44,931 CURRENT, IT'S TERRIBLE, STILL SO 1499 00:52:44,931 --> 00:52:49,702 BAD, THAT'S WHAT THE MUTATION 1500 00:52:49,702 --> 00:52:50,737 DOES IT AFFECTS THE ABILITY OF 1501 00:52:50,737 --> 00:52:51,771 THE CHANNEL TO FOLD PROPER LE. 1502 00:52:51,771 --> 00:52:54,140 SO 1 OF THE DRUGS THIS HAS BEEN 1503 00:52:54,140 --> 00:52:55,441 SHOWN IN USING STRUCTURAL 1504 00:52:55,441 --> 00:52:57,610 BIOLOGY, IS THAT THE DRUG 1505 00:52:57,610 --> 00:52:59,145 ACTUALLY HELPS THE CHANNEL TO 1506 00:52:59,145 --> 00:53:02,915 FOLD BETTER AND SO, FOR THE 1507 00:53:02,915 --> 00:53:04,083 CYSTIC FIBROSIS, WE NEEDED THE 1508 00:53:04,083 --> 00:53:05,485 MAUL MOLECULE DRUG IN,A DITION 1509 00:53:05,485 --> 00:53:08,054 TO THE ENDUB TO GET THE MAXIMUM 1510 00:53:08,054 --> 00:53:09,389 RESCUE, RIGHT? 1511 00:53:09,389 --> 00:53:11,457 AND SO, IT'LL BE DEFINITE FOR 1512 00:53:11,457 --> 00:53:18,298 DIFFERENT MUTATIONS AND WHAT 1513 00:53:18,298 --> 00:53:19,332 THEY DO. 1514 00:53:19,332 --> 00:53:21,134 >> IF THAT'S EVERYTHING, THEN 1515 00:53:21,134 --> 00:53:23,970 LET'S THANK HENRY AGAIN. 1516 00:53:23,970 --> 00:53:33,970 [ APPLAUSE ]