1 00:00:06,680 --> 00:00:11,120 I'M I AM HEADING THE SECTION OF 2 00:00:11,120 --> 00:00:12,640 THE MOLECULAR TRANSACTION AT 3 00:00:12,640 --> 00:00:14,080 NICHD. 4 00:00:14,080 --> 00:00:17,680 IT IS MY PRIVILEGE TO WELCOME 5 00:00:17,680 --> 00:00:18,200 TODAY'S SPEAKER, DR. TOBIAS 6 00:00:18,200 --> 00:00:19,880 WALTHER. 7 00:00:19,880 --> 00:00:23,480 DR. WALTHER IS AN HHMI 8 00:00:23,480 --> 00:00:26,040 INVESTIGATOR, NEWLY APPOINTED 9 00:00:26,040 --> 00:00:27,320 CHAIR, PROGRAM CHAIR, OF THE 10 00:00:27,320 --> 00:00:37,280 CELL BIOLOGY DEPARTMENT AT 11 00:00:37,280 --> 00:00:39,280 SLOAN-KETTERING, PROFESSOR IN 12 00:00:39,280 --> 00:00:41,680 BIOCHEMISTRY AT WEILL CORNELL 13 00:00:41,680 --> 00:00:42,360 SCHOOL OF MEDICINE. 14 00:00:42,360 --> 00:00:46,160 SOME OF YOU MIGHT REMEMBER AND 15 00:00:46,160 --> 00:00:48,360 BE SURPRISED DR. WALTHER IS NOT 16 00:00:48,360 --> 00:00:49,440 AT HARVARD. 17 00:00:49,440 --> 00:00:54,400 HE RECENTLY MOVED FROM HARVARD 18 00:00:54,400 --> 00:00:55,800 WHERE HE WAS PROFESSOR AT THE 19 00:00:55,800 --> 00:00:59,640 CHAN SCHOOL OF PUBLIC HEALTH, 20 00:00:59,640 --> 00:01:01,360 AND ASSOCIATE MEMBER OF THE 21 00:01:01,360 --> 00:01:02,480 BROAD INSTITUTE. 22 00:01:02,480 --> 00:01:04,440 THESE POSITIONS UNTIL LAST YEAR. 23 00:01:04,440 --> 00:01:07,120 SO, THIS HAS BEEN A BUSY YEAR 24 00:01:07,120 --> 00:01:11,440 FOR HIM BECAUSE HE MOVED TO 25 00:01:11,440 --> 00:01:12,440 SLOAN-KETTERING, WHERE HE'S 26 00:01:12,440 --> 00:01:21,080 RUNNING A LAB TOGETHER WITH DR. 27 00:01:21,080 --> 00:01:23,040 ROBERT FORESER JR., FOR A LONGER 28 00:01:23,040 --> 00:01:29,680 PERIOD OF TIME. 29 00:01:29,680 --> 00:01:35,480 THEY INVESTIGATE CELL LIPID, IN 30 00:01:35,480 --> 00:01:36,440 PARTICULAR MECHANISM OF NEUTRAL 31 00:01:36,440 --> 00:01:37,520 FAT STORAGE. 32 00:01:37,520 --> 00:01:40,720 MORE BROADLY, HIS LAB IS 33 00:01:40,720 --> 00:01:44,560 INTERESTED IN HOW CELLS REGULATE 34 00:01:44,560 --> 00:01:47,320 THE SUPPLY OF FAT, AND ABUNDANCE 35 00:01:47,320 --> 00:01:50,040 OF LIPIDS, HOW THE CELLS STORE 36 00:01:50,040 --> 00:01:57,840 AND MOBILIZE LIPIDS UPON NEED, 37 00:01:57,840 --> 00:01:59,800 AND HOW PROCESSES HELP 38 00:01:59,800 --> 00:02:02,040 PHYSIOLOGY AND MEMBRANE BIOLOGY 39 00:02:02,040 --> 00:02:03,440 AND DYNAMICS. 40 00:02:03,440 --> 00:02:06,400 OF COURSE, IT IS IMPORTANT THAT 41 00:02:06,400 --> 00:02:09,680 ALL OF THESE FUNCTIONS ARE 42 00:02:09,680 --> 00:02:13,000 RELATED TO HUMAN DISEASES, SO 43 00:02:13,000 --> 00:02:15,520 THEY ARE VERY INVOLVED. 44 00:02:15,520 --> 00:02:18,480 THEY USE NOTABLE APPROACHES, 45 00:02:18,480 --> 00:02:24,320 SOME BIOPHYSICAL, SOME IMAGING, 46 00:02:24,320 --> 00:02:25,920 BIOCHEMISTRY, PROTEOMICS, 47 00:02:25,920 --> 00:02:28,040 LIPIDOMICS TO DISCOVER INSIGHTS 48 00:02:28,040 --> 00:02:29,000 INTO LIPID METABOLISM. 49 00:02:29,000 --> 00:02:33,880 HE IS A NATIVE OF GERMANY, 50 00:02:33,880 --> 00:02:39,360 UNDERGRADUATE DEGREE IN 51 00:02:39,360 --> 00:02:40,240 MOLECULAR GENETICS IN MUNICH, 52 00:02:40,240 --> 00:02:43,960 GERMANY, AND DID HIS Ph.D. 53 00:02:43,960 --> 00:02:46,280 STUDIES IN BIOCHEMISTRY AT THE 54 00:02:46,280 --> 00:02:49,120 MOLECULAR BIOLOGY LAB IN 55 00:02:49,120 --> 00:02:54,360 HEIDELBERG, DID A POSTDOCTORAL 56 00:02:54,360 --> 00:02:57,640 STUDY IN THE UCSF IN 2002-2006, 57 00:02:57,640 --> 00:03:00,720 RETURNED TO GERMANY TO MAX 58 00:03:00,720 --> 00:03:02,240 PLANCK INSTITUTE AT GROUP LEADER 59 00:03:02,240 --> 00:03:04,640 FOR A SHORT PERIOD. 60 00:03:04,640 --> 00:03:07,480 HE THEN MOVED BACK TO THE U.S. 61 00:03:07,480 --> 00:03:11,440 IN 2010, AND BECAME AN ASSOCIATE 62 00:03:11,440 --> 00:03:15,480 PROFESSOR IN CELL BIOLOGY AT 63 00:03:15,480 --> 00:03:16,280 YALE. 64 00:03:16,280 --> 00:03:19,080 FROM THERE, IN 2014, HE JOINED 65 00:03:19,080 --> 00:03:21,200 HARVARD AND THE BROAD INSTITUTE. 66 00:03:21,200 --> 00:03:24,920 I THINK THIS IS JUST ENOUGH TO 67 00:03:24,920 --> 00:03:28,160 LIST ALL OF THE MOVEMENTS, I'M 68 00:03:28,160 --> 00:03:31,240 REALLY IMPRESSED BY THE AMOUNT 69 00:03:31,240 --> 00:03:32,160 OF MOVEMENTS HE DID. 70 00:03:32,160 --> 00:03:36,280 BUT IT SEEMS TO BE ALL GOING 71 00:03:36,280 --> 00:03:36,760 UPWARD. 72 00:03:36,760 --> 00:03:42,520 SO, ASIDE FROM RECOGNITION FROM 73 00:03:42,520 --> 00:03:46,360 THE HHMI INSTITUTE, AND SEVERAL 74 00:03:46,360 --> 00:03:50,400 NIH GRANTS AND AWARDS, I ONLY 75 00:03:50,400 --> 00:03:53,040 HAVE TO INTRODUCE HIS TALK, 76 00:03:53,040 --> 00:03:56,000 WHICH THE TITLE IS FAT MECHANISM 77 00:03:56,000 --> 00:04:02,040 AND PHYSIOLOGY OF LIPID STORAGE. 78 00:04:02,040 --> 00:04:04,960 PLEASE WELCOME DR. TOBIAS 79 00:04:04,960 --> 00:04:05,160 WALTHER. 80 00:04:05,160 --> 00:04:12,400 [APPLAUSE] 81 00:04:12,400 --> 00:04:13,280 >> THANK YOU FOR THE 82 00:04:13,280 --> 00:04:14,640 INTRODUCTION AND ALL OF YOU FOR 83 00:04:14,640 --> 00:04:16,360 SHOWING UP FOR THE SEMINAR 84 00:04:16,360 --> 00:04:16,680 TODAY. 85 00:04:16,680 --> 00:04:18,640 IT'S A GREAT PLEASURE TO BE 86 00:04:18,640 --> 00:04:19,040 HERE. 87 00:04:19,040 --> 00:04:23,480 I THINK I HAVE HOPEFULLY THE 88 00:04:23,480 --> 00:04:25,040 NEXT COUPLE MINUTES TO HOPEFULLY 89 00:04:25,040 --> 00:04:27,640 IMPRESS YOU WITH ANYTHING ELSE 90 00:04:27,640 --> 00:04:30,600 BUT MY ABILITY TO MOVE. 91 00:04:30,600 --> 00:04:33,040 SO, BEFORE I GET STARTED, I 92 00:04:33,040 --> 00:04:34,760 WANTED TO BRIEFLY MENTION 93 00:04:34,760 --> 00:04:36,120 SOMETHING THAT TOMAS ALREADY 94 00:04:36,120 --> 00:04:36,840 INTRODUCED, WHICH IS ALL THE 95 00:04:36,840 --> 00:04:38,520 WORK THAT I'M GOING TO PRESENT 96 00:04:38,520 --> 00:04:41,600 TODAY IS THE RESULT OF A 97 00:04:41,600 --> 00:04:44,840 SCIENTIFIC PARTNERSHIP BETWEEN 98 00:04:44,840 --> 00:04:46,320 MYSELF AND BOB FARESE, WORKED 99 00:04:46,320 --> 00:04:48,880 TOGETHER FOR 15 TO 20 YEARS NOW, 100 00:04:48,880 --> 00:04:51,640 WORKED AS A SCIENTIFIC 101 00:04:51,640 --> 00:04:52,160 PARTNERSHIP. 102 00:04:52,160 --> 00:04:53,440 WE RECENTLY TRIED TO REFLECT ON 103 00:04:53,440 --> 00:04:54,640 OUR EXPERIENCES HERE A LITTLE 104 00:04:54,640 --> 00:04:59,480 BIT ON HOW IT IS TO RUN A LAB 105 00:04:59,480 --> 00:05:00,160 TOGETHER. 106 00:05:00,160 --> 00:05:04,480 NOW, WHAT I WANTED TO INTRODUCE 107 00:05:04,480 --> 00:05:09,000 TO YOU IS REALLY WONDERFUL, HOW 108 00:05:09,000 --> 00:05:10,680 MUCH LIFE REQUIRES IN TERMS OF 109 00:05:10,680 --> 00:05:10,920 ENERGY. 110 00:05:10,920 --> 00:05:14,520 IT'S JUST IF YOU TAKE A REGULAR 111 00:05:14,520 --> 00:05:16,960 HUMAN BEING, WE GENERATE ABOUT 112 00:05:16,960 --> 00:05:20,360 OUR BODY WEIGHT IN ATP, 113 00:05:20,360 --> 00:05:21,800 OBVIOUSLY TURNOVER, BUT ROUGHLY 114 00:05:21,800 --> 00:05:23,440 BODY WEIGHT IN ATP EVERY DAY TO 115 00:05:23,440 --> 00:05:25,680 DO ALL KINDS OF WONDERFUL 116 00:05:25,680 --> 00:05:26,880 THINGS, MOVE AROUND, DIGEST, 117 00:05:26,880 --> 00:05:31,080 THINK, WHAT NOT. 118 00:05:31,080 --> 00:05:33,240 SO, A CLOSE IS BALANCE BY ENERGY 119 00:05:33,240 --> 00:05:35,000 INTAKE, YOU HAVE TO FUEL THE 120 00:05:35,000 --> 00:05:42,000 SYSTEM TO ALLOW THAT TO HAPPEN. 121 00:05:42,000 --> 00:05:44,080 NOW, IN MOST SYSTEMS, I CAN'T 122 00:05:44,080 --> 00:05:45,360 FIGURE OUT WHICH MICROPHONE, 123 00:05:45,360 --> 00:05:45,560 SORRY. 124 00:05:45,560 --> 00:05:48,120 IN MOST SYSTEMS, OF COURSE, THIS 125 00:05:48,120 --> 00:05:51,200 BALANCE NEEDS TO HAPPEN, DOESN'T 126 00:05:51,200 --> 00:05:52,280 PERFECTLY OCCUR OVER TIME BUT 127 00:05:52,280 --> 00:05:54,440 THERE ARE FLUCTUATIONS BOTH IN 128 00:05:54,440 --> 00:05:56,760 THE NEED TO EXPEND METABOLIC 129 00:05:56,760 --> 00:05:58,840 ENERGY AND IN THE ABILITY TO 130 00:05:58,840 --> 00:06:01,600 TAKE UP ENERGY FROM THE 131 00:06:01,600 --> 00:06:02,040 ENVIRONMENT. 132 00:06:02,040 --> 00:06:03,680 AND BECAUSE OF THOSE 133 00:06:03,680 --> 00:06:07,960 FLUCTUATIONS THERE'S A NEED TO 134 00:06:07,960 --> 00:06:09,720 BUFFER TEMPORARILY THE ABILITY 135 00:06:09,720 --> 00:06:13,320 TO EXPAND ENERGY AND WE DO THAT 136 00:06:13,320 --> 00:06:14,960 PREDOMINANTLY BY TEMPORARILY 137 00:06:14,960 --> 00:06:15,640 STORING METABOLIC ENERGY. 138 00:06:15,640 --> 00:06:17,360 AND BY AND LARGE THIS OCCURS IN 139 00:06:17,360 --> 00:06:19,200 THE FORM OF LIPIDS. 140 00:06:19,200 --> 00:06:21,400 THIS IS DRAWN ROUGHLY TO SCALE 141 00:06:21,400 --> 00:06:23,800 FOR A HEALTHY HUMAN, AND SHOWS 142 00:06:23,800 --> 00:06:25,240 ROUGHLY HOW MUCH METABOLIC 143 00:06:25,240 --> 00:06:30,600 ENERGY WE TEMPORARILY STORE IN 144 00:06:30,600 --> 00:06:33,560 THE FORM OF LIPIDS VERSUS 145 00:06:33,560 --> 00:06:36,400 CARBOHYDRATES,' VOLVED AS A NEAR 146 00:06:36,400 --> 00:06:37,840 UNIVERSAL FORM OF METABOLIC 147 00:06:37,840 --> 00:06:41,320 ENERGY STORAGE BECAUSE MOLECULES 148 00:06:41,320 --> 00:06:44,960 OF LIPIDS ARE STORED IN A 149 00:06:44,960 --> 00:06:48,160 MOLECULE CHARACTERIZED BY 150 00:06:48,160 --> 00:06:50,240 GLYCEROL BACKBONE WITH THREE 151 00:06:50,240 --> 00:06:51,120 CHAINS. 152 00:06:51,120 --> 00:06:53,600 THAT MAKES THAT MOLECULE 153 00:06:53,600 --> 00:06:55,160 EXTREMELY HYDROPHOBIC, DOESN'T 154 00:06:55,160 --> 00:06:56,440 MIX WITH WATER. 155 00:06:56,440 --> 00:06:59,520 AS A CONSEQUENCE THIS IS THE 156 00:06:59,520 --> 00:07:01,480 DENSEST FORM YOU CAN STORE 157 00:07:01,480 --> 00:07:03,040 CARBOHYDRATES IN BECAUSE YOU 158 00:07:03,040 --> 00:07:03,800 DON'T HAVE THE WATER. 159 00:07:03,800 --> 00:07:05,840 THIS IS THE REASON PRESUMABLY 160 00:07:05,840 --> 00:07:08,520 THIS HAS EVOLVED TO BECOME MORE 161 00:07:08,520 --> 00:07:12,560 OR LESS UNIVERSAL FORM OF ENERGY 162 00:07:12,560 --> 00:07:12,800 STORAGE. 163 00:07:12,800 --> 00:07:15,520 IT CREATES A PROBLEM. 164 00:07:15,520 --> 00:07:19,640 SO BECAUSE OUR CHEMISTRY HAPPENS 165 00:07:19,640 --> 00:07:23,400 PREDOMINANTLY IN AQUEOUS SPACE, 166 00:07:23,400 --> 00:07:30,080 WHEN YOU PUT TRIDELAYS TRIGLYCY 167 00:07:30,080 --> 00:07:33,480 STAY SEPARATE. 168 00:07:33,480 --> 00:07:35,360 YOU CAN RECONSTITUTE, IN A GLASS 169 00:07:35,360 --> 00:07:36,320 ON MY PORCH. 170 00:07:36,320 --> 00:07:37,640 OIL DOESN'T MIX WITH WATER, SO 171 00:07:37,640 --> 00:07:39,080 HERE'S WHERE YOU WILL AT ENERGY 172 00:07:39,080 --> 00:07:41,120 IS STORED IN A CELL, IF YOU 173 00:07:41,120 --> 00:07:41,320 WILL. 174 00:07:41,320 --> 00:07:42,680 THE WATER IS REALLY WHERE ALL 175 00:07:42,680 --> 00:07:45,040 THE LIFE HAPPENS. 176 00:07:45,040 --> 00:07:46,480 AT THE INTERFACE A LOT OF 177 00:07:46,480 --> 00:07:48,280 BIOLOGY NEEDS TO HAPPEN TO ALLOW 178 00:07:48,280 --> 00:07:51,440 THE POSITION OF THE ENERGY INTO 179 00:07:51,440 --> 00:07:53,080 THE OIL PHASE, SOME OF YOU MAY 180 00:07:53,080 --> 00:07:55,280 REMEMBER THAT IN OLD DAYS IN 181 00:07:55,280 --> 00:07:57,240 ORDER TO EXTRACT DNA WE USED 182 00:07:57,240 --> 00:07:58,440 SUCH INTERFACES TO DENATURE ALL 183 00:07:58,440 --> 00:08:00,320 THE PROTEINS AND GET RID OF THEM 184 00:08:00,320 --> 00:08:06,200 TO EXACT THE DNA IN THE WATER 185 00:08:06,200 --> 00:08:07,440 SOLUBLE PHASE, CREATING UNIQUE 186 00:08:07,440 --> 00:08:07,680 CHALLENGE. 187 00:08:07,680 --> 00:08:10,480 IN CELLS THE OIL IS NOT STORED 188 00:08:10,480 --> 00:08:13,320 AT MASSIVE LAYER OF FAT BUT 189 00:08:13,320 --> 00:08:17,680 INSTEAD IN WHAT WE KNOW AS 190 00:08:17,680 --> 00:08:22,960 ORGANELLES, LIPID DROPLETS, VERY 191 00:08:22,960 --> 00:08:26,480 UNUSUAL COMPARED TO MITOCHONDRIA 192 00:08:26,480 --> 00:08:28,000 OR RETICULUM, WITH A CORE OF 193 00:08:28,000 --> 00:08:29,560 NEUTRAL LIPIDS THAT FORMS 194 00:08:29,560 --> 00:08:31,640 ORGANIC PHASE, BOUNDED BY A 195 00:08:31,640 --> 00:08:33,960 MONOLAYER, NOT A BILAYER 196 00:08:33,960 --> 00:08:34,800 MEMBRANE, PHOSPHOLIPIDS INTO 197 00:08:34,800 --> 00:08:37,000 WHICH SPECIFIC PROTEINS ARE 198 00:08:37,000 --> 00:08:39,120 EMBEDDED. 199 00:08:39,120 --> 00:08:40,840 AND REALLY THE DISCOVERY THAT 200 00:08:40,840 --> 00:08:42,600 THERE ARE SPECIFIC PROTEINS TO 201 00:08:42,600 --> 00:08:44,280 THIS SURFACE REALLY STARTED TO 202 00:08:44,280 --> 00:08:46,080 PUT THIS ORGANELLE ON THE MAP AS 203 00:08:46,080 --> 00:08:48,440 SOMETHING THAT'S NOT JUST AN 204 00:08:48,440 --> 00:08:51,040 INERT BLOB OF FAT. 205 00:08:51,040 --> 00:08:52,760 BUT AN ORGANELLE IN CELLS WITH 206 00:08:52,760 --> 00:08:54,880 ALL THE USUAL DYNAMIC BEHAVIORS. 207 00:08:54,880 --> 00:08:59,000 YOU CAN SEE LIPID DROPLETS 208 00:08:59,000 --> 00:09:01,400 INTERACT WITH OR ORGANELLES SUCH 209 00:09:01,400 --> 00:09:03,040 AS MITOCHONDRIA OR ENDOPLASMIC 210 00:09:03,040 --> 00:09:03,320 RETICULUM. 211 00:09:03,320 --> 00:09:07,120 IF YOU TAKE A SINGLE LIPID 212 00:09:07,120 --> 00:09:10,600 DROPLET IF IT'S 300 TO 500 213 00:09:10,600 --> 00:09:12,520 NANOMETERS CAN STORE EQUIVALENT 214 00:09:12,520 --> 00:09:15,040 OF 23 BILLION ATP EQUIVALENTS, 215 00:09:15,040 --> 00:09:16,760 THAT'S REALLY WHERE A LOT OF 216 00:09:16,760 --> 00:09:19,920 METABOLIC ENERGY IS TEMPORARILY 217 00:09:19,920 --> 00:09:20,160 STORED. 218 00:09:20,160 --> 00:09:21,480 AS I MENTIONED IN CELLS YOU HAVE 219 00:09:21,480 --> 00:09:23,320 MULTIPLE OF THESE LIPID 220 00:09:23,320 --> 00:09:30,240 DROPLETS, MOST OFTEN. 221 00:09:30,240 --> 00:09:34,480 AND THEIR PRESENCE CONVERTS TO 222 00:09:34,480 --> 00:09:38,760 EMULSION, DROPLETS FORM, AND 223 00:09:38,760 --> 00:09:40,760 SURFACTANTS ARE THERE TO 224 00:09:40,760 --> 00:09:42,320 BASICALLY DELINEATE TWO SURFACES 225 00:09:42,320 --> 00:09:43,800 AND ALLOW ALL THE TRAFFIC IN AND 226 00:09:43,800 --> 00:09:47,240 OUT, ALL THE BIOLOGY THAT I'M 227 00:09:47,240 --> 00:09:48,560 GOING TO TALK ABOUT. 228 00:09:48,560 --> 00:09:58,200 NOW, IN MULTI-CELLULAR 229 00:09:58,200 --> 00:10:00,040 ORGANISMS, FAT IS STORED? 230 00:10:00,040 --> 00:10:00,680 ADIPOSE TISSUE. 231 00:10:00,680 --> 00:10:05,720 YOU CAN SEE THIS IS A SINGLE 232 00:10:05,720 --> 00:10:12,600 LIPID DROPLET THAT OCCUPIES MOST 233 00:10:12,600 --> 00:10:17,400 OF THE ADIPOCYTE ORGANELLES ARE 234 00:10:17,400 --> 00:10:18,000 SQUEEZED. 235 00:10:18,000 --> 00:10:19,520 LIPID DROPLETS ARE NOT 236 00:10:19,520 --> 00:10:21,240 RESTRICTED TO ADIPOCYTES, THEY 237 00:10:21,240 --> 00:10:22,880 OCCUR IN PRETTY MUCH EVERY OTHER 238 00:10:22,880 --> 00:10:28,360 CELL TYPE PEOPLE HAVE LOOKED AT 239 00:10:28,360 --> 00:10:31,200 INCLUDING WHERE YOU SEE AFTER 240 00:10:31,200 --> 00:10:32,400 FATTY ACIDS COME FROM THE MEAL, 241 00:10:32,400 --> 00:10:37,520 AND THE LIVER WHERE THEY ARE 242 00:10:37,520 --> 00:10:39,080 TRAFFICKED FROM EITHER THE 243 00:10:39,080 --> 00:10:41,480 ADIPOSE TISSUE, STORED IN LIPID 244 00:10:41,480 --> 00:10:41,760 DROPLETS. 245 00:10:41,760 --> 00:10:44,760 THE HEART RUNS ON FATTY ACIDS, 246 00:10:44,760 --> 00:10:46,840 METABOLIC FUEL, TEMPORARILY CAN 247 00:10:46,840 --> 00:10:48,800 STORE LIPID DROPLETS. 248 00:10:48,800 --> 00:10:50,800 FOR INSTANCE MILK HAS A LOT OF 249 00:10:50,800 --> 00:10:52,400 FAT, THOSE ARE LIPID DROPLETS 250 00:10:52,400 --> 00:10:56,880 THAT GET SHED FROM THE 251 00:10:56,880 --> 00:10:58,680 EPITHELIAL CELL, STILL POORLY 252 00:10:58,680 --> 00:11:00,320 UNDERSTOOD MECHANISM. 253 00:11:00,320 --> 00:11:01,440 LIPID DROPLETS ARE PRESENT 254 00:11:01,440 --> 00:11:03,600 THROUGHOUT PHYSIOLOGY, NOT ONLY 255 00:11:03,600 --> 00:11:05,760 IN PHYSIOLOGY BUT 256 00:11:05,760 --> 00:11:06,320 PATHOPHYSIOLOGY. 257 00:11:06,320 --> 00:11:07,320 SO, FOR INSTANCE, MANY OF YOU 258 00:11:07,320 --> 00:11:10,280 KNOW OF COURSE THAT WE'RE FACING 259 00:11:10,280 --> 00:11:11,320 AN UNPARALLELED EPIDEMIC OF 260 00:11:11,320 --> 00:11:13,880 OBESITY IN THIS COUNTRY AND 261 00:11:13,880 --> 00:11:19,000 ELSEWHERE WHERE REALLY MASSIVE 262 00:11:19,000 --> 00:11:20,000 ACCUMULATION OF TRIGLYCERIDES 263 00:11:20,000 --> 00:11:21,000 INTERFERE WITH PHYSIOLOGY OF 264 00:11:21,000 --> 00:11:23,240 HUMANS. 265 00:11:23,240 --> 00:11:27,000 WHAT'S LESS KNOWN THERE'S ALSO A 266 00:11:27,000 --> 00:11:29,960 DIFFERENT CONDITION KNOWN AS 267 00:11:29,960 --> 00:11:31,360 LIPODYSTROPHY, FAILURE TO 268 00:11:31,360 --> 00:11:32,120 MAINTAIN ADIPOSE TISSUE LEADS TO 269 00:11:32,120 --> 00:11:33,440 SEVERE DISEASE. 270 00:11:33,440 --> 00:11:36,200 IN BOTH CASES ONE CAN THINK FOR 271 00:11:36,200 --> 00:11:37,640 DIFFERENT REASON ADIPOSE TISSUE 272 00:11:37,640 --> 00:11:39,840 GETS OVERWHELMED WITH FATTY 273 00:11:39,840 --> 00:11:44,640 ACIDS, ALTHOUGH THIS PERSON HERE 274 00:11:44,640 --> 00:11:46,720 LOOKS PHENOTYPICALLY HEALTHY THE 275 00:11:46,720 --> 00:11:49,880 TWO CONDITIONS SHARE SIMILAR 276 00:11:49,880 --> 00:11:55,600 HALLMARKS IN TERMS OF DISEASE. 277 00:11:55,600 --> 00:11:56,240 ON THE BIOCHEMICAL LEVEL, 278 00:11:56,240 --> 00:11:59,200 TRIGLYCERIDES ARE SYNTHESIZED BY 279 00:11:59,200 --> 00:12:01,480 TWO ENZYMES, KNOWN INITIALLY 280 00:12:01,480 --> 00:12:02,640 DISCOVERED BY EUGENE KENNEDY, 281 00:12:02,640 --> 00:12:06,520 CO-WORKERS IN THE 19 50s, 282 00:12:06,520 --> 00:12:10,680 PURIFIED AND NAMED AFTER THEIR 283 00:12:10,680 --> 00:12:15,280 ACTIVITIES, KNOWN AS D GAT 1 AND 284 00:12:15,280 --> 00:12:15,600 2. 285 00:12:15,600 --> 00:12:21,600 AND THOSE TWO TAKE A POLAR 286 00:12:21,600 --> 00:12:29,920 MOLECULE, AND MAKE IT APOLAR. 287 00:12:29,920 --> 00:12:31,480 AFTER BOB DISCOVERED DGAT 1, 288 00:12:31,480 --> 00:12:37,200 THIS IS A FANTASTIC DRUG TARGET, 289 00:12:37,200 --> 00:12:39,560 IF YOU'RE A MOUSE. 290 00:12:39,560 --> 00:12:42,320 IF YOU DELETE OR INHIBIT MICE 291 00:12:42,320 --> 00:12:44,920 LIVE 30% LONGER, RESISTANT TO 292 00:12:44,920 --> 00:12:47,560 DIET-INDUCED OBESITY, AND THEY 293 00:12:47,560 --> 00:12:49,120 HAVE ESSENTIALLY ALL POSITIVE 294 00:12:49,120 --> 00:12:51,840 CHARACTERISTICS OF METABOLISM. 295 00:12:51,840 --> 00:12:53,800 SO THIS HAS INSPIRED A LOT OF 296 00:12:53,800 --> 00:12:54,800 PHARMACEUTICAL COMPANIES TO 297 00:12:54,800 --> 00:12:55,360 DEVELOP INHIBITORS. 298 00:12:55,360 --> 00:12:57,320 I'LL SAY A LITTLE BIT ABOUT THAT 299 00:12:57,320 --> 00:12:58,400 IN A SECOND. 300 00:12:58,400 --> 00:13:03,320 TURNS OUT THAT WHEN YOU THINK 301 00:13:03,320 --> 00:13:04,760 ABOUT TRIGLYCERIDE SYNTHESIS TWO 302 00:13:04,760 --> 00:13:09,880 PARTS OF THE EQUATION ARE WORTH 303 00:13:09,880 --> 00:13:10,320 CONSIDERING. 304 00:13:10,320 --> 00:13:13,760 ONE IS THEY MAKE AND ARE PUT IN 305 00:13:13,760 --> 00:13:17,880 DROPLETS BUT REMOVE DIGLYCEROL, 306 00:13:17,880 --> 00:13:19,480 TWO SIDES OF THE COIN HERE. 307 00:13:19,480 --> 00:13:21,720 WHEN YOU FOR INSTANCE GENERATE A 308 00:13:21,720 --> 00:13:24,320 MOUSE THAT DOES NOT HAVE BOTH 309 00:13:24,320 --> 00:13:26,160 ENZYMES SPECIFICALLY IN ADIPOSE 310 00:13:26,160 --> 00:13:28,480 TISSUE, THIS MOUSE IS 311 00:13:28,480 --> 00:13:30,040 SURPRISINGLY HEALTHY, DOESN'T 312 00:13:30,040 --> 00:13:31,680 HAVE ANY PARTICULAR PROBLEM, YOU 313 00:13:31,680 --> 00:13:34,200 CAN SEE IT HAS ADIPOSE TISSUE, 314 00:13:34,200 --> 00:13:35,960 BUT THE ADIPOSE TISSUE DOESN'T 315 00:13:35,960 --> 00:13:40,000 HAVE ANY FAT. 316 00:13:40,000 --> 00:13:41,400 IT'S JUST A SIMPLE WAY TO 317 00:13:41,400 --> 00:13:43,160 VISUALIZE THE EFFECT. 318 00:13:43,160 --> 00:13:44,680 THE MICE ARE FINE AS LONG AS 319 00:13:44,680 --> 00:13:46,560 THEY HAVE FOOD, BUT IF YOU DON'T 320 00:13:46,560 --> 00:13:48,640 HAVE FOOD AND PUT THEM IN THE 321 00:13:48,640 --> 00:13:50,600 COLD, A MAJOR METABOLIC STRESSOR 322 00:13:50,600 --> 00:13:53,280 IN MICE, THEY CAN NOT MAINTAIN 323 00:13:53,280 --> 00:13:54,120 BODY TEMPERATURE. 324 00:13:54,120 --> 00:13:55,560 SO THIS WILL ARGUE AS YOU 325 00:13:55,560 --> 00:13:58,120 PROBABLY KNOW, WHAT I ARGUED FOR 326 00:13:58,120 --> 00:14:00,000 IS INTEREST TRIGLYCERIDES 327 00:14:00,000 --> 00:14:01,200 FUNCTION AS ENERGY SOURCE. 328 00:14:01,200 --> 00:14:03,600 WHAT ALSO HAPPENS IS THAT IN 329 00:14:03,600 --> 00:14:10,960 MANY CASES SUCH AS OBESITY THIS 330 00:14:10,960 --> 00:14:11,800 EXCESS LIPIDS OVERWHELM 331 00:14:11,800 --> 00:14:16,200 MACHINERY AND FLOW INTO OTHER 332 00:14:16,200 --> 00:14:18,000 METABOLITES THAT WHEN ACCUMULATE 333 00:14:18,000 --> 00:14:19,360 CAN TRIGGER E.R. STRESS THAT 334 00:14:19,360 --> 00:14:22,000 LEADS TO CELL DEATH OR 335 00:14:22,000 --> 00:14:22,280 ADAPTATION. 336 00:14:22,280 --> 00:14:24,720 TURNS OUT IN HUMANS THAT'S WHAT 337 00:14:24,720 --> 00:14:26,800 HAPPENS IN THE INTESTINE BECAUSE 338 00:14:26,800 --> 00:14:31,320 THOSE MICE I MENTIONED EXPRESS 339 00:14:31,320 --> 00:14:34,240 DGAT1 AND 2, WE ONLY EXPRESS 340 00:14:34,240 --> 00:14:34,600 DGAT1. 341 00:14:34,600 --> 00:14:36,640 WHEN WE DISCOVERED PATIENTS WITH 342 00:14:36,640 --> 00:14:38,840 MUTATIONS IN THIS ENZYME, THEY 343 00:14:38,840 --> 00:14:46,280 HAVE A SEVERE CONGENITAL 344 00:14:46,280 --> 00:14:46,920 DIARRHEA SYNDROME, ESSENTIALLY 345 00:14:46,920 --> 00:14:50,760 WHAT WE THINK IS GOING ON IS 346 00:14:50,760 --> 00:14:53,480 THAT WHEN THE ORGANISM IS 347 00:14:53,480 --> 00:14:57,120 CONFRONTED WITH A LOT OF FATTY 348 00:14:57,120 --> 00:14:59,200 ACIDS FROM THE MEAL ENTEROCYTE 349 00:14:59,200 --> 00:15:02,160 CANNOT TEMPORARILY BUFFER THE 350 00:15:02,160 --> 00:15:06,880 INFLUX IN LIPID DROPLETS, AS A 351 00:15:06,880 --> 00:15:08,640 RESULT TRIGGERS METABOLIC -- 352 00:15:08,640 --> 00:15:09,720 DYSFUNCTION AND STRESS, 353 00:15:09,720 --> 00:15:11,920 CONSISTENT WITH THIS WE NOW CAN 354 00:15:11,920 --> 00:15:15,520 FIGURE OUT HOW TO -- SORRY, HOW 355 00:15:15,520 --> 00:15:16,960 TO DEAL WITH THIS, WITH 356 00:15:16,960 --> 00:15:18,400 BYPASSING THE INTESTINE FOR 357 00:15:18,400 --> 00:15:20,120 THOSE PATIENTS. 358 00:15:20,120 --> 00:15:22,520 MORE BROADLY, THIS HAS LED TO A 359 00:15:22,520 --> 00:15:24,040 CONCEPT PROPOSED BY US AND 360 00:15:24,040 --> 00:15:25,680 OTHERS THAT LIPID DROPLETS HAVE 361 00:15:25,680 --> 00:15:27,560 A PROTECTIVE FUNCTION IN MANY 362 00:15:27,560 --> 00:15:29,760 CONTEXTS SUCH AS LIVER 363 00:15:29,760 --> 00:15:31,280 MACROPHAGES FOR MUSCLE AND 364 00:15:31,280 --> 00:15:32,480 HEART, WHERE THEY OFTEN PROTECT 365 00:15:32,480 --> 00:15:34,800 THE TISSUE FROM THE ACCUMULATION 366 00:15:34,800 --> 00:15:36,080 OF METABOLIC DISEASE. 367 00:15:36,080 --> 00:15:45,560 AND WE THINK THIS IS ALSO WHY 368 00:15:45,560 --> 00:15:46,440 DGAT1 INHIBITORS, KILLS THE 369 00:15:46,440 --> 00:15:47,000 PROTECTIVE FUNCTION. 370 00:15:47,000 --> 00:15:49,160 WE KNOW FROM WORK OF MANY 371 00:15:49,160 --> 00:15:51,800 LABORATORIES INCLUDING OURS THAT 372 00:15:51,800 --> 00:15:54,400 THE PROCESSES INVOLVED IN LIPID 373 00:15:54,400 --> 00:15:56,920 STORAGE ARE INTIMATELY LINKED TO 374 00:15:56,920 --> 00:15:57,720 INHERITED METABOLIC DISEASES 375 00:15:57,720 --> 00:16:00,240 BECAUSE WHEN YOU HAVE MUTATIONS 376 00:16:00,240 --> 00:16:02,600 IN ENZYMES INVOLVED IN THE 377 00:16:02,600 --> 00:16:06,080 SYNTHESIS OF TRIGLYCERIDE YOU 378 00:16:06,080 --> 00:16:08,960 GET A NUMBER OF SEVERE DISEASES. 379 00:16:08,960 --> 00:16:10,720 NOW, WITH THAT, YOU COULD ASK IS 380 00:16:10,720 --> 00:16:11,920 ALL HOPE LOST? 381 00:16:11,920 --> 00:16:13,520 TURNS OUT THERE'S ALSO 382 00:16:13,520 --> 00:16:14,880 INHIBITORS FOR THIS OTHER 383 00:16:14,880 --> 00:16:15,840 ENZYME, THERE'S A LESSON HERE 384 00:16:15,840 --> 00:16:17,480 BECAUSE WHEN WE INITIALLY FOUND 385 00:16:17,480 --> 00:16:19,600 THAT ENZYME AND MADE A DELETION 386 00:16:19,600 --> 00:16:21,640 IT WAS LETHAL IN MICE. 387 00:16:21,640 --> 00:16:23,720 MICE DIE SOON AFTER BIRTH. 388 00:16:23,720 --> 00:16:25,920 TURNS OUT THAT NOW THIS HAS 389 00:16:25,920 --> 00:16:27,120 BECOME A FRONTRUNNER FOR 390 00:16:27,120 --> 00:16:28,080 TREATMENT OF FATTY LIVER 391 00:16:28,080 --> 00:16:30,280 DISEASE, AGAIN BECAUSE OF THE 392 00:16:30,280 --> 00:16:31,720 SPECIES DIFFERENCE. 393 00:16:31,720 --> 00:16:35,240 AND WE FIND THAT IF YOU JUST 394 00:16:35,240 --> 00:16:37,720 RESTRICT THE EFFECT TO MICE, 395 00:16:37,720 --> 00:16:39,600 SORRY, TO LIVER, THAT PROTECTS 396 00:16:39,600 --> 00:16:41,160 THE ANIMALS FROM FATTY LIVER 397 00:16:41,160 --> 00:16:41,400 DISEASE. 398 00:16:41,400 --> 00:16:45,120 ONE OF THE MAJOR SIDE EFFECTS OF 399 00:16:45,120 --> 00:16:46,120 OBESITY BECAUSE NOW FATTY ACIDS 400 00:16:46,120 --> 00:16:48,280 ARE NOT ONLY IN THE ADIPOSE 401 00:16:48,280 --> 00:16:49,280 TISSUE ANYMORE BUT SPILL OVER TO 402 00:16:49,280 --> 00:16:50,160 THE LIVER. 403 00:16:50,160 --> 00:16:52,120 THIS PROTECTS FROM THAT AND HAS 404 00:16:52,120 --> 00:16:55,600 LED TO MOVING THOSE DRUGS INTO 405 00:16:55,600 --> 00:16:56,720 CLINICAL TRIALS WITH MARKEDLY 406 00:16:56,720 --> 00:16:57,600 REDUCED LIVER FAT. 407 00:16:57,600 --> 00:17:00,840 THIS IS INITIALLY DONE BY 408 00:17:00,840 --> 00:17:01,320 PFIZER. 409 00:17:01,320 --> 00:17:02,840 TURNS OUT WHAT I TOLD YOU 410 00:17:02,840 --> 00:17:06,440 SHOULDN'T MAKE SENSE, RIGHT? 411 00:17:06,440 --> 00:17:08,480 YOU STILL HAVE THE PROTECTIVE 412 00:17:08,480 --> 00:17:11,280 EFFECT, WHY IS ANY NOT A PROBLEM 413 00:17:11,280 --> 00:17:13,240 WITH DGAT2? 414 00:17:13,240 --> 00:17:16,280 YOU NOT ONLY BLOCK TRIGLYCERIDE 415 00:17:16,280 --> 00:17:17,160 SYNTHESIS BUT A FEEDBACK 416 00:17:17,160 --> 00:17:19,600 REGULATION SHUTS OFF SYNTHESIS 417 00:17:19,600 --> 00:17:22,400 OF MORE FATTY ACIDS, PREVENTS 418 00:17:22,400 --> 00:17:24,800 ACCUMULATION OF ANY LIPID TOXIC 419 00:17:24,800 --> 00:17:25,160 INTERMEDIATE. 420 00:17:25,160 --> 00:17:27,920 ALL OF WHAT I TOLD YOU IN A 421 00:17:27,920 --> 00:17:28,800 RELATIVELY SHORT PROGRESSION IS 422 00:17:28,800 --> 00:17:31,600 JUST MEANT TO GIVE AN 423 00:17:31,600 --> 00:17:32,280 INTRODUCTION THAT LIPID DROPLETS 424 00:17:32,280 --> 00:17:33,640 CAN HAVE TWO MAJOR FUNCTIONS IN 425 00:17:33,640 --> 00:17:33,840 CELLS. 426 00:17:33,840 --> 00:17:37,760 THEY CAN BE USED TO STORE 427 00:17:37,760 --> 00:17:45,640 METABOLIC EMERGENCY AND DETOXIFY 428 00:17:45,640 --> 00:17:48,120 INTERMEDIATES OF METABOLISM. 429 00:17:48,120 --> 00:17:52,880 THREE TOPICS IN THE BASIC 430 00:17:52,880 --> 00:17:54,360 BIOLOGY OF ORGANELLES, HOW THEY 431 00:17:54,360 --> 00:17:58,280 ARE MADE, WHAT ARE ENZYMATIC 432 00:17:58,280 --> 00:18:00,080 MECHANISMS, HOW DO YOU PACKAGE 433 00:18:00,080 --> 00:18:02,280 THEM INTO LIPID DROPLETS AND HOW 434 00:18:02,280 --> 00:18:03,240 ARE PROTEINS THAT MEDIATE 435 00:18:03,240 --> 00:18:04,880 EXCHANGE WITH THE REST OF THE 436 00:18:04,880 --> 00:18:08,160 CELL ACTUALLY GETTING THERE, AND 437 00:18:08,160 --> 00:18:10,040 FINALLY HOW DO YOU EXTRACT AND 438 00:18:10,040 --> 00:18:13,240 MOBILIZE THINGS FROM A LIPID 439 00:18:13,240 --> 00:18:14,520 DROPLET AGAIN. 440 00:18:14,520 --> 00:18:18,920 FIRST PART, HOW ARE CELLS 441 00:18:18,920 --> 00:18:20,000 SYNTHESIZED TRIGLYCERIDES, 442 00:18:20,000 --> 00:18:21,560 THERE'S TWO ENZYMES BUT THEY 443 00:18:21,560 --> 00:18:23,600 ONLY SHARE A NAME. 444 00:18:23,600 --> 00:18:27,760 OTHERWISE THOSE ARE 445 00:18:27,760 --> 00:18:30,760 EVOLUTIONARIAL UNRELATED, DGAT 1 446 00:18:30,760 --> 00:18:34,360 IS A MEMBRANE PROTEIN, 2 AS A 447 00:18:34,360 --> 00:18:35,880 HAIRPIN DOMAIN, OTHERWISE 448 00:18:35,880 --> 00:18:36,640 CYTOSOLIC. 449 00:18:36,640 --> 00:18:40,240 WE SOLVED THE STRUCTURE BY 450 00:18:40,240 --> 00:18:41,440 MICROSCOPY, REVEAL AS DIMER 451 00:18:41,440 --> 00:18:44,280 HERE, IF YOU FOCUS ON ONE SIDE 452 00:18:44,280 --> 00:18:46,920 WHAT WE FOUND IS THE CATALYTIC 453 00:18:46,920 --> 00:18:47,880 CENTER IS DEEPLY BURIED WITHIN 454 00:18:47,880 --> 00:18:53,160 THE PLANE OF THE E.R. MEMBRANE. 455 00:18:53,160 --> 00:18:53,680 ACCESSIBLE FROM BOTH THE 456 00:18:53,680 --> 00:18:55,120 CYTOSOLIC SIDE THROUGH A TUNNEL 457 00:18:55,120 --> 00:18:57,320 AND ALSO THROUGH A LATERAL GATE 458 00:18:57,320 --> 00:18:58,520 WITHIN THE MEMBRANE. 459 00:18:58,520 --> 00:19:00,800 AND BASED ON MANY EXPERIMENTS WE 460 00:19:00,800 --> 00:19:02,680 NOW BELIEVE THIS CATALYTIC 461 00:19:02,680 --> 00:19:06,160 CENTER IS ACCESSED FROM THE SITE 462 00:19:06,160 --> 00:19:07,560 OF THE CYTOPLASM, LOOKING LIKE 463 00:19:07,560 --> 00:19:13,400 THIS CARROT HERE, ACCESSES 464 00:19:13,400 --> 00:19:19,080 CATALYTIC SITE, A MOIETY 465 00:19:19,080 --> 00:19:20,200 ACCESSES CATALYTIC CENTER, THE 466 00:19:20,200 --> 00:19:21,720 MOLECULE IS FORMED THAT CAN BE 467 00:19:21,720 --> 00:19:25,760 RELEASED INTO THE PLANE OF THE 468 00:19:25,760 --> 00:19:26,280 MEMBRANE. 469 00:19:26,280 --> 00:19:28,600 INTRIGUINGLY, JUST AS A SIDE 470 00:19:28,600 --> 00:19:34,960 NOTE, THIS ENZYME IS A MEN OF A 471 00:19:34,960 --> 00:19:36,280 FAMILY MEMBRANE BOUND 472 00:19:36,280 --> 00:19:42,520 TRANSFERASES, A LARGE GROUP THAT 473 00:19:42,520 --> 00:19:45,120 ARE INVOLVED IN THE FORMATION OF 474 00:19:45,120 --> 00:19:47,600 IMPORTANT SIGNALING MOLECULES 475 00:19:47,600 --> 00:19:50,240 SUCH AS HEDGEHOG OR WINGLESS. 476 00:19:50,240 --> 00:19:51,600 WE NOW HAVE STRUCTURES FOR A 477 00:19:51,600 --> 00:19:54,000 NUMBER BUT I'LL SKIP THIS. 478 00:19:54,000 --> 00:19:57,200 WE HAVE MUCH LESS INFORMATION ON 479 00:19:57,200 --> 00:20:00,080 DGAT2, BUT IT WAS ONE OF THE 480 00:20:00,080 --> 00:20:02,720 MAJOR PROTEINS HIGHLIGHTED IN 481 00:20:02,720 --> 00:20:05,320 THE INITIAL ALPHA FOLD 482 00:20:05,320 --> 00:20:07,000 PREDICTION, TO SHOW THIS IS HOW 483 00:20:07,000 --> 00:20:09,080 WE THINK THIS ENZYME IS LOOKING 484 00:20:09,080 --> 00:20:12,480 LIKE BASED ON THE PREDICTION, 485 00:20:12,480 --> 00:20:14,360 AND INTRIGUINGLY HAS AN ACTIVE 486 00:20:14,360 --> 00:20:17,800 SITE, NOT IN THE MEMBRANE BUT ON 487 00:20:17,800 --> 00:20:20,880 THE CYTOSOLIC SIDE OF THE 488 00:20:20,880 --> 00:20:21,520 MEMBRANE. 489 00:20:21,520 --> 00:20:27,640 SO NOW THAT WE UNDERSTAND THAT 490 00:20:27,640 --> 00:20:28,920 PARTICULARLY THIS ENZYME 491 00:20:28,920 --> 00:20:31,080 RELEASES THE TRIGLYCERIDE, THE 492 00:20:31,080 --> 00:20:32,920 NEXT QUESTION BECOMES HOW THEN 493 00:20:32,920 --> 00:20:36,320 DO YOU FORM A LIPID DROPLET? 494 00:20:36,320 --> 00:20:37,560 AFTER THE TRIGLYCERIDE IS 495 00:20:37,560 --> 00:20:38,880 RELEASED INTO THE MEMBRANE? 496 00:20:38,880 --> 00:20:40,520 FOR MANY YEARS THE GENERAL 497 00:20:40,520 --> 00:20:42,240 THINKING WAS, WELL, YOU JUST 498 00:20:42,240 --> 00:20:45,400 RELEASE IT INTO THE MEMBRANE, 499 00:20:45,400 --> 00:20:46,840 AND WE KNOW FROM WORK OF 500 00:20:46,840 --> 00:20:48,520 HAMILTON AND OTHERS THAT THERE'S 501 00:20:48,520 --> 00:20:54,640 A MAXIMUM SOLUBILITY, AROUND 3. 502 00:20:54,640 --> 00:20:58,080 UP TO 3% OF TRIGLYCERIDE, THEY 503 00:20:58,080 --> 00:20:59,440 JUST SOLUBILIZE IN THE MEMBRANE, 504 00:20:59,440 --> 00:21:02,040 AND IF YOU THINK ABOUT IT, VERY 505 00:21:02,040 --> 00:21:06,720 SIMILAR TO SIDE CHAINS OF A 506 00:21:06,720 --> 00:21:07,080 PHOSPHOLIPID. 507 00:21:07,080 --> 00:21:08,600 ABOUT 3% PHASE SEPARATION OCCURS 508 00:21:08,600 --> 00:21:10,480 AND AN OIL PHASE STARTS 509 00:21:10,480 --> 00:21:10,720 EMERGING. 510 00:21:10,720 --> 00:21:11,760 FOR MANY YEARS IT WAS THOUGHT 511 00:21:11,760 --> 00:21:15,360 THAT THAT'S JUST A PASSIVE 512 00:21:15,360 --> 00:21:16,240 PROCESS THAT OCCURS. 513 00:21:16,240 --> 00:21:18,320 BUT WE AND OTHERS HAVE FOUND 514 00:21:18,320 --> 00:21:19,760 THAT ACTUALLY CELLS DON'T LEAVE 515 00:21:19,760 --> 00:21:22,160 THIS PROCESS UP TO CHANCE BUT 516 00:21:22,160 --> 00:21:23,920 INSTEAD HAVE EVOLVED DEDICATED 517 00:21:23,920 --> 00:21:25,680 PROTEIN MACHINERY THAT ORGANIZED 518 00:21:25,680 --> 00:21:27,120 FORMATION OF LIPID DROPLETS OUT 519 00:21:27,120 --> 00:21:28,960 OF THE E.R. 520 00:21:28,960 --> 00:21:30,520 SPECIFICALLY, A KEY PROTEIN FOR 521 00:21:30,520 --> 00:21:33,120 THIS IS A PROTEIN CALLED SAPIN, 522 00:21:33,120 --> 00:21:35,280 NAMED AFTER ONE OF THE 523 00:21:35,280 --> 00:21:36,840 DISCOVERERS OF A MUTATION IN THE 524 00:21:36,840 --> 00:21:40,880 '50s THAT LEADS TO THIS LIPID 525 00:21:40,880 --> 00:21:41,440 DYSTROPHY. 526 00:21:41,440 --> 00:21:48,320 SO, THIS PROTEIN IS CALLED SAPIN 527 00:21:48,320 --> 00:21:53,440 OR BSCL 2, AND MUTATIONS 528 00:21:53,440 --> 00:21:55,760 ESSENTIALLY LEAD TO A COMPLETE 529 00:21:55,760 --> 00:21:57,720 LACK OF ADIPOSE TISSUE, AND THIS 530 00:21:57,720 --> 00:22:00,360 IS THOUGHT TO OCCUR BECAUSE 531 00:22:00,360 --> 00:22:02,640 DURING THE DIFFERENTIATION OF 532 00:22:02,640 --> 00:22:04,640 ADIPOSE TISSUE MASSIVE SYNTHESIS 533 00:22:04,640 --> 00:22:06,600 OF TRIGLYCERIDES OCCURS, WHEN 534 00:22:06,600 --> 00:22:08,480 THE MACHINERY THAT I WILL SHOW 535 00:22:08,480 --> 00:22:10,680 YOU INVOLVED IN TRIGLYCERIDE 536 00:22:10,680 --> 00:22:12,120 PACKAGING INTO LIPID DROPLETS IS 537 00:22:12,120 --> 00:22:15,720 DEFICIENT YOU GET E.R. STRESS, 538 00:22:15,720 --> 00:22:20,720 TOXICITY, PRESUMABLY THAT'S WHY 539 00:22:20,720 --> 00:22:22,240 THOSE CELLS DON'T SURVIVE. 540 00:22:22,240 --> 00:22:24,360 FIFTY YEARS LATER THE GENE 541 00:22:24,360 --> 00:22:25,600 ENCODING THIS PROTEIN WAS 542 00:22:25,600 --> 00:22:29,080 CLONED, AND IT WAS SHOWN BY A 543 00:22:29,080 --> 00:22:30,800 NUMBER OF LABS THAT THE 544 00:22:30,800 --> 00:22:33,200 DEFICIENCY IN THIS PERTURBED 545 00:22:33,200 --> 00:22:35,320 LIPID DROPLETS IN YEAST, PLANTS, 546 00:22:35,320 --> 00:22:37,040 WORMS, MANY DIFFERENT LABS, 547 00:22:37,040 --> 00:22:42,280 INCLUDING OUR OWN, WHERE WE 548 00:22:42,280 --> 00:22:45,600 FOUND INITIALLY IN IN MAMMALIAN 549 00:22:45,600 --> 00:22:46,880 CELLS GENE ENCODING THIS, THIS 550 00:22:46,880 --> 00:22:49,400 IS A REPRESENTATION LOOKING FOR 551 00:22:49,400 --> 00:22:50,640 PHENOTYPES ASSOCIATED WITH 552 00:22:50,640 --> 00:22:52,480 DEPLETION OF EACH GENE GIVES ONE 553 00:22:52,480 --> 00:22:54,320 OF THE STRONGEST PHENOTYPES ON 554 00:22:54,320 --> 00:22:55,600 LISPED DROPLETS. 555 00:22:55,600 --> 00:22:57,960 SO THAT PHENOTYPE CAN BE 556 00:22:57,960 --> 00:22:59,720 SUMMARIZED IN SHORT, IN A FLY 557 00:22:59,720 --> 00:23:03,880 CELL HERE, THAT IN NORMAL CELLS 558 00:23:03,880 --> 00:23:06,520 THERE'S A FEW EXISTING 559 00:23:06,520 --> 00:23:07,280 PREEXISTING LIPID DROPLETS, 560 00:23:07,280 --> 00:23:11,960 SHOWN IN THE GREEN COLOR, 561 00:23:11,960 --> 00:23:15,160 SURROUNDED BY LIPID DROPLET, 562 00:23:15,160 --> 00:23:15,680 LIVEDROP. 563 00:23:15,680 --> 00:23:16,280 THERE'S PREEXISTING DROPLETS, 564 00:23:16,280 --> 00:23:21,160 WHEN YOU ADD FATTY ACIDS, 565 00:23:21,160 --> 00:23:21,680 EQUIVALENT OF McDONALD'S, 566 00:23:21,680 --> 00:23:24,240 THERE'S A FEW LIPID DROPLETS 567 00:23:24,240 --> 00:23:25,880 FORMING IN AN ORGANIZED WAY AT 568 00:23:25,880 --> 00:23:30,360 SOME SITES IN THE LIPID DROPLET, 569 00:23:30,360 --> 00:23:31,640 IN THE E.R. 570 00:23:31,640 --> 00:23:41,840 WHEN YOU DO NOT HAVE SEIPIN 571 00:23:41,840 --> 00:23:43,160 PREEXISTING DROPLETS, CAN YOU 572 00:23:43,160 --> 00:23:49,880 SEE SMALL BLISTERS OCCURRING, 573 00:23:49,880 --> 00:23:52,120 OBSERVED ACROSS DIFFERENT 574 00:23:52,120 --> 00:23:53,880 SPECIES, THAT PHENOTYPE. 575 00:23:53,880 --> 00:23:57,280 WE TACKED IT AT ENDOGENOUS LOCUS 576 00:23:57,280 --> 00:23:58,560 WITH FLUOROPHORE, TRACKED. 577 00:23:58,560 --> 00:24:02,600 YOU CAN SEE AN E.R. SIGNAL IN 578 00:24:02,600 --> 00:24:02,760 RED. 579 00:24:02,760 --> 00:24:03,960 AND SEIPIN IN GREEN. 580 00:24:03,960 --> 00:24:07,000 WHAT BECOMES OBVIOUS IS THAT THE 581 00:24:07,000 --> 00:24:08,440 PROTEIN FORMS FOCI, COMPLEXES IN 582 00:24:08,440 --> 00:24:09,560 THE E.R. MEMBRANE. 583 00:24:09,560 --> 00:24:10,960 AGAIN, YOU CAN SEE THIS EASY IN 584 00:24:10,960 --> 00:24:14,200 A DROSOPHILA CELL THAT IS THE 585 00:24:14,200 --> 00:24:15,600 SAME AS TRUE IN MAMMALIAN CELL, 586 00:24:15,600 --> 00:24:17,720 IN THIS CASE A HUMAN CELL. 587 00:24:17,720 --> 00:24:18,680 THAT OF COURSE IMMEDIATELY 588 00:24:18,680 --> 00:24:20,440 BROUGHT US TO WHAT ARE THESE 589 00:24:20,440 --> 00:24:21,560 COMPLEXES, HOW DO THEY WORK, 590 00:24:21,560 --> 00:24:22,880 WHAT DO YOU HAVE TO DO AND WHY 591 00:24:22,880 --> 00:24:24,720 DO YOU GET THAT PHENOTYPE? 592 00:24:24,720 --> 00:24:28,000 SO IN ORDER TO UNDERSTAND THAT 593 00:24:28,000 --> 00:24:29,320 WE SOLVED THE STRUCTURE OF SUCH 594 00:24:29,320 --> 00:24:31,320 A COMPLEX. 595 00:24:31,320 --> 00:24:33,800 THIS IS BY CRYO-ELECTRON 596 00:24:33,800 --> 00:24:34,120 MICROSCOPY. 597 00:24:34,120 --> 00:24:36,880 WHAT BECOMES OBVIOUS, THIS FORMS 598 00:24:36,880 --> 00:24:38,160 A LARGE OLIGOMERIC RING 599 00:24:38,160 --> 00:24:42,640 STRUCTURE IN THE ENDOPLASMIC 600 00:24:42,640 --> 00:24:44,960 RETICULUM MEMBRANE WITH A 601 00:24:44,960 --> 00:24:47,280 LUMINAL DOMAIN, AND MANY 602 00:24:47,280 --> 00:24:48,360 TRANSMEMBRANE SEGMENTS. 603 00:24:48,360 --> 00:24:50,760 SO TO GIVE A BETTER VIEW WHAT 604 00:24:50,760 --> 00:24:53,600 YOU CAN SEE IS THERE'S A CASE 605 00:24:53,600 --> 00:24:57,120 STRUCTURE, AGAIN OPEN TO THE 606 00:24:57,120 --> 00:24:59,280 PLANE OF THE MEMBRANE AND 607 00:24:59,280 --> 00:25:00,640 RELATIVELY CLOSE. 608 00:25:00,640 --> 00:25:03,560 WHAT WE THINK HAPPENS HERE IS 609 00:25:03,560 --> 00:25:04,760 THAT THIS PROVIDES A SPACE IN 610 00:25:04,760 --> 00:25:08,600 THE CENTER OF THIS, NOT UNLIKE 611 00:25:08,600 --> 00:25:13,200 FOR INSTANCE GRUEL E L DURING 612 00:25:13,200 --> 00:25:14,080 PROTEIN FOLDING, TRIGLYCERIDE 613 00:25:14,080 --> 00:25:15,600 MOLECULES CAN DIFFUSE IN AND 614 00:25:15,600 --> 00:25:17,160 FIND A SPACE THAT EXCLUDES 615 00:25:17,160 --> 00:25:19,760 PHOSPHOLIPIDS AND THERE BY HELPS 616 00:25:19,760 --> 00:25:20,880 CATALYZING PHASE TRANSITION, IN 617 00:25:20,880 --> 00:25:24,680 OTHER WORDS THE LIKELIHOOD OF 618 00:25:24,680 --> 00:25:25,600 THREE TRIGLYCERIDE MOLECULES TO 619 00:25:25,600 --> 00:25:27,440 MEET RATHER THAN INTERACT WITH 620 00:25:27,440 --> 00:25:28,640 PHOSPHOLIPID. 621 00:25:28,640 --> 00:25:31,600 WE TESTED THIS IN MOLECULAR 622 00:25:31,600 --> 00:25:34,760 DYNAMIC SIMULATIONS, FINDING FOR 623 00:25:34,760 --> 00:25:36,720 INSTANCE TOGETHER WITH 624 00:25:36,720 --> 00:25:39,480 UNIVERSITY OF CHICAGO WE FIND 625 00:25:39,480 --> 00:25:43,520 ONLY WHEN SEIPIN IS PRESENT, 626 00:25:43,520 --> 00:25:44,160 LIPID DROPLET FORMATION OCCURS 627 00:25:44,160 --> 00:25:47,000 IN THE CENTER OF THE VERY LARGE 628 00:25:47,000 --> 00:25:47,600 COMPLEX. 629 00:25:47,600 --> 00:25:51,840 AND THEN YOU CAN SEE IN THE 630 00:25:51,840 --> 00:25:52,920 DYNAMIC SIMULATIONS THAT THE 631 00:25:52,920 --> 00:25:56,240 COMPLEX OPENS AND LIPID DROPLET 632 00:25:56,240 --> 00:25:58,480 FORMS, TO THE CYTOPLASM. 633 00:25:58,480 --> 00:25:59,680 WE CAN SEE, IMPORTANTLY, ONLY 634 00:25:59,680 --> 00:26:02,480 WHEN WE HAVE SEIPIN CAN YOU 635 00:26:02,480 --> 00:26:05,920 START NUCLEATING LIPID DROPLET 636 00:26:05,920 --> 00:26:07,600 OR PHASE TRANSITION OF 637 00:26:07,600 --> 00:26:11,640 CONCENTRATIONS BELOW THAT 638 00:26:11,640 --> 00:26:12,520 SOLUBILITY LEVEL. 639 00:26:12,520 --> 00:26:13,720 THAT APPEARS TO BE A CRUCIAL 640 00:26:13,720 --> 00:26:15,880 PART OF THE MOLECULE AT LEAST IN 641 00:26:15,880 --> 00:26:18,160 SIMULATIONS TO HELP WITH 642 00:26:18,160 --> 00:26:18,480 NUCLEATION. 643 00:26:18,480 --> 00:26:23,120 WHEN YOU LOOK MORE CAREFUL AT A 644 00:26:23,120 --> 00:26:26,680 STRUCTURE IT HAS A RING OF 645 00:26:26,680 --> 00:26:28,040 LUMINAL DOMAINS, PROTRUDING IN 646 00:26:28,040 --> 00:26:30,640 THE MEMBRANE, WE THINK THIS 647 00:26:30,640 --> 00:26:32,840 HELPS EXCLUDE PHOSPHOLIPIDS, AND 648 00:26:32,840 --> 00:26:33,960 THESE ARE VERY IMPORTANT, ALSO 649 00:26:33,960 --> 00:26:36,120 SHOWN BY OTHERS FOR NUCLEATION 650 00:26:36,120 --> 00:26:39,760 AND TRANSMEMBRANE DOMAINS. 651 00:26:39,760 --> 00:26:42,280 WHAT WE DIDN'T REALIZE, THESE 652 00:26:42,280 --> 00:26:43,920 HELIXES THAT PUSH APART 653 00:26:43,920 --> 00:26:45,320 PHOSPHOLIPIDS ARE NOT ONLY 654 00:26:45,320 --> 00:26:47,320 CONSERVED IN THEIR CHARACTER BUT 655 00:26:47,320 --> 00:26:49,480 ALSO IN EVOLUTION ARE CONSERVED 656 00:26:49,480 --> 00:26:54,360 WITH THEIR -- IN THEIR SPECIFIC 657 00:26:54,360 --> 00:26:54,640 SEQUENCE. 658 00:26:54,640 --> 00:26:56,160 THAT'S SOMEWHAT UNUSUAL. 659 00:26:56,160 --> 00:26:59,320 WHY IS THERE ANY SPECIFIC 660 00:26:59,320 --> 00:27:01,000 SEQUENCES CONSERVED? 661 00:27:01,000 --> 00:27:03,040 SO THE LAB POSTULATED WE MAY BE 662 00:27:03,040 --> 00:27:05,760 MISSING A CO-FACTOR AND SHE DID 663 00:27:05,760 --> 00:27:08,720 SOME EXPERIMENTS, I'LL SHOW ONE 664 00:27:08,720 --> 00:27:09,480 HERE. 665 00:27:09,480 --> 00:27:17,600 SHE EITHER HAD THESE HYDROPHOBIC 666 00:27:17,600 --> 00:27:18,280 HELICES, FOUND ONE PROTEIN 667 00:27:18,280 --> 00:27:18,920 SPECIFICALLY INTERACTED WITH 668 00:27:18,920 --> 00:27:22,240 THIS FORM OF THE COMPLEX BUT NOT 669 00:27:22,240 --> 00:27:23,440 THAT ONE, UNCHARACTERIZED, A 670 00:27:23,440 --> 00:27:30,800 MEMBRANE PROTEIN WE HAVE RENAMED 671 00:27:30,800 --> 00:27:34,200 TMEM FROM TMEM 159. 672 00:27:34,200 --> 00:27:38,000 WHEN WE TAKE CELLS, TACK SEIPIN 673 00:27:38,000 --> 00:27:41,240 ENDOGENOUS LOCUS AND NEW PROTEIN 674 00:27:41,240 --> 00:27:43,680 LDAF, AND WE FOLLOW LIPID 675 00:27:43,680 --> 00:27:46,640 DROPLET FORMATION BY A THIRD 676 00:27:46,640 --> 00:27:48,600 COLOR KNOCK-IN, AND USING A 677 00:27:48,600 --> 00:27:51,280 MARKER PROTEIN THAT LIPID 678 00:27:51,280 --> 00:27:54,440 DROPLETS ONLY FORM AT SITES THAT 679 00:27:54,440 --> 00:28:01,000 HAVE BOTH SEIPIN AND THIS LDAF 680 00:28:01,000 --> 00:28:01,280 PROTEIN. 681 00:28:01,280 --> 00:28:03,200 AND YOU CAN -- YES, THAT'S WHY 682 00:28:03,200 --> 00:28:05,480 WE RENAMED THE TWO PROTEINS 683 00:28:05,480 --> 00:28:07,120 TOGETHER, FORMED THE LIPID 684 00:28:07,120 --> 00:28:08,520 DROPLET ASSEMBLY COMPLEX. 685 00:28:08,520 --> 00:28:15,200 ONE OF THE THINGS THAT WAS 686 00:28:15,200 --> 00:28:17,320 GRATIFYING, IS SEIPIN WOULD BE 687 00:28:17,320 --> 00:28:21,360 SUFFICIENT TO MEDIATE, WHEN WE 688 00:28:21,360 --> 00:28:22,240 PURIFIED SEIPIN BIOCHEMICALLY 689 00:28:22,240 --> 00:28:23,240 NEVER FOUND ASSOCIATE TO 690 00:28:23,240 --> 00:28:28,800 TRIGLYCERIDE, ONLY WHEN YOU 691 00:28:28,800 --> 00:28:30,880 CO-PURIFY AND RECONSTITUTE YOU 692 00:28:30,880 --> 00:28:32,080 CAN SEE IT'S TRIGLYCERIDE 693 00:28:32,080 --> 00:28:35,920 BECAUSE IT IS SENSITIVE TO THE 694 00:28:35,920 --> 00:28:41,720 INHIBITION AND CAN BE RAISED AT 695 00:28:41,720 --> 00:28:44,720 EXTRA FATTY ACIDS. 696 00:28:44,720 --> 00:28:49,400 THIS APPEARS TO START SORT OF 697 00:28:49,400 --> 00:28:49,920 TRANSFORMING THE MICELLE. 698 00:28:49,920 --> 00:28:54,960 IF YOU LOOK AT SEIPIN IT FORMS A 699 00:28:54,960 --> 00:28:55,880 FLAT MICELLE. 700 00:28:55,880 --> 00:28:58,360 WE SEE THERE'S TWO THINGS IN 701 00:28:58,360 --> 00:29:01,320 HERE, ADDITIONAL DENSITY OF 702 00:29:01,320 --> 00:29:02,640 PROTEIN, VERY POORLY RESOLVED, 703 00:29:02,640 --> 00:29:03,600 WE CAN TALK BIT, AND 704 00:29:03,600 --> 00:29:04,200 TRIGLYCERIDE. 705 00:29:04,200 --> 00:29:07,360 WE THINK THIS IS THE EARLIEST 706 00:29:07,360 --> 00:29:08,320 INTERMEDIATE OF LIPID DROPLET 707 00:29:08,320 --> 00:29:10,840 FORMATION WE CAN VISUALIZE WHERE 708 00:29:10,840 --> 00:29:12,400 WE START HAVING THIS COMPLEX AND 709 00:29:12,400 --> 00:29:15,080 SOME TRIGLYCERIDE TRAPPED IN THE 710 00:29:15,080 --> 00:29:15,520 MIDDLE. 711 00:29:15,520 --> 00:29:18,840 FASCINATINGLY AS LIPID DROPLETS 712 00:29:18,840 --> 00:29:19,880 FORM THE COMPLEX DISASSOCIATES, 713 00:29:19,880 --> 00:29:23,000 SEIPIN STAYS IN THE E.R. 714 00:29:23,000 --> 00:29:24,520 MEMBRANE, AND LF1 STARTS TO 715 00:29:24,520 --> 00:29:27,040 DECORATE THE SURFACE OF LIPID 716 00:29:27,040 --> 00:29:27,280 DROPLETS. 717 00:29:27,280 --> 00:29:29,120 SO IF I PUT THIS TOGETHER INTO A 718 00:29:29,120 --> 00:29:33,720 MODEL, WHAT I TOLD YOU ABOUT, IT 719 00:29:33,720 --> 00:29:35,840 WILL LOOK LIKE THIS. 720 00:29:35,840 --> 00:29:46,360 YOU HAVE ENZYMES THAT TAKE THE 721 00:29:47,160 --> 00:29:48,640 FATTY ACID, RELEASED INTO THE 722 00:29:48,640 --> 00:29:51,560 PLANE OF THE MEMBRANE, DIFFUSE 723 00:29:51,560 --> 00:29:54,080 INTO THE CAGE SHAPE OF THE 724 00:29:54,080 --> 00:29:56,600 SEIPIN LDAF COMPLEX, THAT ALLOWS 725 00:29:56,600 --> 00:29:57,680 PHASE TRANSITION. 726 00:29:57,680 --> 00:29:59,480 AND THEN ONCE YOU START THAT, 727 00:29:59,480 --> 00:30:01,200 THE COMPLEX OPENS AND GOES 728 00:30:01,200 --> 00:30:02,640 TOWARDS THE CYTOPLASM. 729 00:30:02,640 --> 00:30:04,480 AT THAT POINT ADDITIONAL 730 00:30:04,480 --> 00:30:06,120 MONOMERS OF TRIGLYCERIDES ARE 731 00:30:06,120 --> 00:30:09,640 INTEGRATED INTO THIS BY A 732 00:30:09,640 --> 00:30:10,600 PROCESS EQUIVALENT THAT'S EASIER 733 00:30:10,600 --> 00:30:13,240 TO GET INTO THE PHASE THAN TO 734 00:30:13,240 --> 00:30:14,000 GET OUT. 735 00:30:14,000 --> 00:30:15,360 THAT'S OUR CURRENT UNDERSTANDING 736 00:30:15,360 --> 00:30:16,560 OF HOW LIPID DROPLET WORKS. 737 00:30:16,560 --> 00:30:21,200 THERE'S A LOT OF MYSTERY STILL 738 00:30:21,200 --> 00:30:21,400 HERE. 739 00:30:21,400 --> 00:30:22,480 NOW, ONCE IT FORMS TO INTERACT 740 00:30:22,480 --> 00:30:24,960 WITH THE REST OF THE CELL IT IS 741 00:30:24,960 --> 00:30:26,480 REALLY IMPORTANT TO HAVE THE 742 00:30:26,480 --> 00:30:28,920 SPECIFIC SET OF MEMBRANE 743 00:30:28,920 --> 00:30:30,320 PROTEINS ON THE SURFACE. 744 00:30:30,320 --> 00:30:32,840 AND A KEY QUESTION IS HOW ARE 745 00:30:32,840 --> 00:30:34,040 PROTEINS TARGETED TO THE 746 00:30:34,040 --> 00:30:34,920 SURFACE? 747 00:30:34,920 --> 00:30:36,960 WE STILL DON'T UNDERSTAND THAT 748 00:30:36,960 --> 00:30:37,640 COMPLETELY. 749 00:30:37,640 --> 00:30:39,520 WE BELIEVE THIS IS A FUNDAMENTAL 750 00:30:39,520 --> 00:30:40,840 QUESTION IN BIOLOGY BECAUSE IT'S 751 00:30:40,840 --> 00:30:43,320 NOT SIMILAR TO OTHER THINGS. 752 00:30:43,320 --> 00:30:44,560 THERE'S NO IMPORT MACHINERY 753 00:30:44,560 --> 00:30:47,160 KNOWN FOR LIPID DROPLETS SUCH AS 754 00:30:47,160 --> 00:30:48,240 FOR E.R., MITOCHONDRIA. 755 00:30:48,240 --> 00:30:50,000 THERE ARE NO SPECIFIC LIPIDS 756 00:30:50,000 --> 00:30:53,200 THAT MARK THE SURFACE SUCH AS 757 00:30:53,200 --> 00:30:55,640 ENDOSOME OR PLASMA MEMBRANE. 758 00:30:55,640 --> 00:30:57,840 BUT INSTEAD THE STRUCTURE OF 759 00:30:57,840 --> 00:30:59,120 THIS SURFACE IS QUITE DIFFERENT. 760 00:30:59,120 --> 00:31:01,200 I WILL TELL YOU ABOUT THIS IN A 761 00:31:01,200 --> 00:31:01,560 SECOND. 762 00:31:01,560 --> 00:31:05,040 WHAT THIS LEADS TO IS THAT IN 763 00:31:05,040 --> 00:31:06,520 PROTEOMIC STUDIES WE KNOW 764 00:31:06,520 --> 00:31:08,640 THERE'S UPWARDS OF 100 PROTEINS 765 00:31:08,640 --> 00:31:10,320 SPECIFIC TO LIPID DROPLETS 766 00:31:10,320 --> 00:31:12,320 DEPENDING ON CELL TYPE, ET 767 00:31:12,320 --> 00:31:12,560 CETERA. 768 00:31:12,560 --> 00:31:13,880 THESE PROTEINS COME IN DIFFERENT 769 00:31:13,880 --> 00:31:18,560 FLAVORS, SOME OF THEM YOU MIGHT 770 00:31:18,560 --> 00:31:21,640 EXPECT SYNTHESIS OR DEGRADATION, 771 00:31:21,640 --> 00:31:22,840 THAT MAKES SENSE. 772 00:31:22,840 --> 00:31:25,800 THERE'S OTHERS WHICH YOU CAN 773 00:31:25,800 --> 00:31:28,080 STILL RATIONALIZE SUCH AS 774 00:31:28,080 --> 00:31:31,720 PHOSPHOLIPID REMODELING ENZYME 775 00:31:31,720 --> 00:31:32,600 AND TRANSCRIPTION FACTORS THAT 776 00:31:32,600 --> 00:31:34,840 WE START TO UNDERSTAND INVOLVED 777 00:31:34,840 --> 00:31:35,960 IN REGULATION OF METABOLISM, 778 00:31:35,960 --> 00:31:41,520 THERE'S ENZYMES INVOLVED IN 779 00:31:41,520 --> 00:31:42,960 GLYCOSYLATION REACTIONS MANY 780 00:31:42,960 --> 00:31:45,360 LABS FOUND AND NOBODY KNOWS WHAT 781 00:31:45,360 --> 00:31:47,760 THEY ARE DOING AND WHY THAT 782 00:31:47,760 --> 00:31:48,120 MATTERS. 783 00:31:48,120 --> 00:31:51,520 IF YOU'RE INTERESTED WE BUILT A 784 00:31:51,520 --> 00:31:54,840 KNOWLEDGE PORTAL WITH THE BROAD 785 00:31:54,840 --> 00:31:55,800 INSTITUTE THAT ALLOWS YOU TO 786 00:31:55,800 --> 00:31:56,880 BROWSE THE INFORMATION THAT AT 787 00:31:56,880 --> 00:31:58,720 LEAST WE AND SOME OTHER LABS 788 00:31:58,720 --> 00:32:01,560 HAVE GENERATED ON THE LIPID 789 00:32:01,560 --> 00:32:01,840 DROPLETS. 790 00:32:01,840 --> 00:32:04,920 HOW DO ALL THESE PROTEINS GET 791 00:32:04,920 --> 00:32:05,120 THERE? 792 00:32:05,120 --> 00:32:06,840 AS I MENTIONED WE DON'T REALLY 793 00:32:06,840 --> 00:32:08,880 KNOW ANY OF THE PATHS. 794 00:32:08,880 --> 00:32:12,280 SO WHAT WE HAVE LEARNED SO FAR 795 00:32:12,280 --> 00:32:14,240 IS THAT GENERALLY THERE APPEAR 796 00:32:14,240 --> 00:32:16,800 TO BE TWO PATHWAYS, ONE PATHWAY 797 00:32:16,800 --> 00:32:19,520 YOU MAKE PROTEINS IN THE CYTOSOL 798 00:32:19,520 --> 00:32:21,680 AND DIRECTLY BIND THE SURFACE. 799 00:32:21,680 --> 00:32:23,760 IN A SECOND PATHWAY, PROTEINS 800 00:32:23,760 --> 00:32:24,880 ARE INITIALLY INSERTED IN THE 801 00:32:24,880 --> 00:32:29,440 E.R. AND GO TO THE SURFACE OF 802 00:32:29,440 --> 00:32:29,800 LIPID DROPLETS. 803 00:32:29,800 --> 00:32:40,360 IN THAT -- WE CALL THEM C YTOLS. 804 00:32:42,560 --> 00:32:43,040 REGULATORS, TRANSCRIPTION 805 00:32:43,040 --> 00:32:45,080 FACTORS, I'LL TELL YOU ABOUT ONE 806 00:32:45,080 --> 00:32:45,760 OTHER PROTEIN LATER. 807 00:32:45,760 --> 00:32:47,880 HOW DO THEY GET TO THE SURFACE 808 00:32:47,880 --> 00:32:50,240 IS NON-TRIVIAL BECAUSE THEY HAVE 809 00:32:50,240 --> 00:32:54,320 A SEQUENCE, A HELIX THAT HAS 810 00:32:54,320 --> 00:32:55,560 PROPENSITY TO BIND THE MEMBRANE. 811 00:32:55,560 --> 00:32:56,960 WHY DO THEY GO TO THIS RATHER 812 00:32:56,960 --> 00:32:59,880 THAN THAT, THAT IS VASTLY MORE 813 00:32:59,880 --> 00:33:01,840 ABUNDANT, MORE E.R. SURFACE THAN 814 00:33:01,840 --> 00:33:03,280 LIPID DROPLETS IN CELLS, EVEN 815 00:33:03,280 --> 00:33:06,240 THOUGH THEY HAVE THE SAME LIPID 816 00:33:06,240 --> 00:33:08,440 COMPOSITION? PART OF THE 817 00:33:08,440 --> 00:33:10,440 SOLUTION IS THE UNDERLYING 818 00:33:10,440 --> 00:33:12,400 TRIGLYCERIDE CHANGES THE SURFACE 819 00:33:12,400 --> 00:33:13,480 PROPERTIES OF THE MONO LAYER. 820 00:33:13,480 --> 00:33:15,880 I DON'T HAVE TIME TO SHOW ALL 821 00:33:15,880 --> 00:33:19,400 THE DATA BUT ESSENTIALLY IF YOU 822 00:33:19,400 --> 00:33:20,920 IMAGINE A MEMBRANE HAS TO BE 823 00:33:20,920 --> 00:33:22,680 TIGHTLY PACKED, NEXT TO EACH 824 00:33:22,680 --> 00:33:22,920 OTHER. 825 00:33:22,920 --> 00:33:24,440 IN A MONOLAYER YOU CAN PUSH THEM 826 00:33:24,440 --> 00:33:27,080 APART A LITTLE BIT, THE 827 00:33:27,080 --> 00:33:29,920 TRIGLYCERIDE CAN PUSH APART THE 828 00:33:29,920 --> 00:33:32,880 CHAINS A LITTLE BIT, PACKING 829 00:33:32,880 --> 00:33:33,560 DEFECTS, PROVIDES SURFACES THAT 830 00:33:33,560 --> 00:33:35,520 CAN BE READ OUT BY PROTEINS. 831 00:33:35,520 --> 00:33:38,040 THESE PACKING DEFECTS ARE MORE 832 00:33:38,040 --> 00:33:44,160 ABUNDANT, MORE PERSIST ENT LARGR 833 00:33:44,160 --> 00:33:49,160 ON THE SURFACE, MODELS WITH 834 00:33:49,160 --> 00:33:49,480 TRIGLYCERIDES. 835 00:33:49,480 --> 00:33:50,800 WHAT WE THINK HAPPENS FOR MANY 836 00:33:50,800 --> 00:33:53,040 PROTEINS, I DON'T HAVE TIME TO 837 00:33:53,040 --> 00:33:54,280 SHOW THE DATA, THEY HAVE 838 00:33:54,280 --> 00:34:02,200 SEQUENCES THAT ARE DISORDERED IN 839 00:34:02,200 --> 00:34:07,000 CYTOSOL, PROPENSITY TO BIND 840 00:34:07,000 --> 00:34:08,680 THROUGH HYDROPHOBICS SUCH AS 841 00:34:08,680 --> 00:34:12,160 TRYPTOPHANS, LEADING TO THE 842 00:34:12,160 --> 00:34:19,240 FOLDING OF HELICES , LEAVING THS 843 00:34:19,240 --> 00:34:20,040 PROCESS IRREVERSIBLE, REQUIRING 844 00:34:20,040 --> 00:34:21,640 A DIFFERENT PROCESS TO GET THE 845 00:34:21,640 --> 00:34:23,040 PROTEINS OFF THE SURFACE LATER 846 00:34:23,040 --> 00:34:23,360 ON. 847 00:34:23,360 --> 00:34:26,080 HOW ABOUT THAT OTHER PATHWAY? 848 00:34:26,080 --> 00:34:28,320 SOME PROTEINS WE KNOW ARE 849 00:34:28,320 --> 00:34:30,360 INSERTED INTO THE E.R., AND THEY 850 00:34:30,360 --> 00:34:35,280 END UP ON THE LIPID DROPLET, 851 00:34:35,280 --> 00:34:43,280 INCLUDING METABOLIC ENZYMES, G 852 00:34:43,280 --> 00:34:45,120 PAT AND ASCL, INCLUDES THE RISK 853 00:34:45,120 --> 00:34:51,840 FACTOR FOR DEVELOPMENT OF FATTY 854 00:34:51,840 --> 00:34:54,840 LIVER DISEASE FROM HEPATITIS, 855 00:34:54,840 --> 00:34:55,120 HSD17B13. 856 00:34:55,120 --> 00:34:56,160 WHAT'S THE MECHANISM? 857 00:34:56,160 --> 00:34:57,680 WE KNOW PROTEINS DON'T JUMP IN 858 00:34:57,680 --> 00:34:58,840 AND OUT OF MEMBRANES, THIS CAN'T 859 00:34:58,840 --> 00:34:59,480 BE IT. 860 00:34:59,480 --> 00:35:01,880 INSTEAD WHAT WE FOUND IS THAT 861 00:35:01,880 --> 00:35:07,960 WHEN YOU ANALYZE LIPID DROPLET 862 00:35:07,960 --> 00:35:09,640 FORMATION, VISUALIZE RETICULUM 863 00:35:09,640 --> 00:35:12,000 IN GREEN, ENZYMES IN RED AS CAR 864 00:35:12,000 --> 00:35:16,000 GO, THEY START IN THE SAME CASE, 865 00:35:16,000 --> 00:35:17,840 SAME PLACE, AND OVERLAP IN 866 00:35:17,840 --> 00:35:18,200 COLOR. 867 00:35:18,200 --> 00:35:23,080 WHEN YOU MAKE LIPID DROPLETS, 868 00:35:23,080 --> 00:35:24,520 COLORS SEPARATE AND ENZYME ENDS 869 00:35:24,520 --> 00:35:32,480 UP ON DROP DROPLET, ER STAYS B. 870 00:35:32,480 --> 00:35:37,400 WE CAN DO LABELING WITH ANTIBODY 871 00:35:37,400 --> 00:35:38,200 AGAINST ENDOGENOUS PROTEIN, 872 00:35:38,200 --> 00:35:39,480 DECORATES THE SURFACE OF THE 873 00:35:39,480 --> 00:35:44,200 LIPID DROPLET BUT NOT THE 874 00:35:44,200 --> 00:35:46,360 ENDOPLASM RETICULUM CLOSE BY. 875 00:35:46,360 --> 00:35:47,520 FIRST THOUGHT, IT'S SIMPLE, 876 00:35:47,520 --> 00:35:48,920 WHAT'S THE BIG HOOPLA? 877 00:35:48,920 --> 00:35:52,520 YOU START WITH A PROTEIN, MAKE 878 00:35:52,520 --> 00:35:54,160 THE LIPID DROPLET, PROTEINS GOES 879 00:35:54,160 --> 00:35:56,000 ON WHEN IT STARTS, RIGHT? 880 00:35:56,000 --> 00:35:57,320 SOMETHING LIKE THAT. 881 00:35:57,320 --> 00:36:00,840 DURING FORMATION PROTEIN GOES ON 882 00:36:00,840 --> 00:36:02,920 THE LIPID DROPLET. 883 00:36:02,920 --> 00:36:07,080 WELL, SOME PROTEINS CAN DO THIS 884 00:36:07,080 --> 00:36:09,120 SUCH AS THIS LIVEDROP, YOU CAN 885 00:36:09,120 --> 00:36:12,320 SEE DURING FORMATION THEY GO ON. 886 00:36:12,320 --> 00:36:13,440 FOR THIS PARTICULAR PROBE WE 887 00:36:13,440 --> 00:36:17,000 KNOW THIS IS DUE TO A CHANGE IN 888 00:36:17,000 --> 00:36:19,640 CONFORMATION OF ENZYME THAT 889 00:36:19,640 --> 00:36:22,920 ALTERS ITS ENERGY STATE BETWEEN 890 00:36:22,920 --> 00:36:24,240 BILAYER AND MONOLAYER. 891 00:36:24,240 --> 00:36:26,000 CURRENT MODEL WHEN THESE THINGS 892 00:36:26,000 --> 00:36:30,800 ARE INSERTED INTO THE E.R. 893 00:36:30,800 --> 00:36:31,800 MEMBRANE, THAT INSERTION 894 00:36:31,800 --> 00:36:33,000 PLACES -- ANCHORS PROTEIN TO 895 00:36:33,000 --> 00:36:35,960 BOTH SIDES OF THE MEMBRANE, WITH 896 00:36:35,960 --> 00:36:37,480 THAT PLACES LARGE HYDROPHOBIC 897 00:36:37,480 --> 00:36:41,960 TRYPTOPHANS IN THE MIDDLE OF A 898 00:36:41,960 --> 00:36:42,520 BILAYER, HIGHLY ENUSUAL, 899 00:36:42,520 --> 00:36:45,040 UNFAVORABLE, WHEN THEY COME TO 900 00:36:45,040 --> 00:36:47,600 LIPID DROPLETS SWIM UP, 901 00:36:47,600 --> 00:36:48,240 TRYPTOPHANS BECOME INTERFACIAL, 902 00:36:48,240 --> 00:36:50,160 WE BELIEVE THAT DRIVES HOW THEY 903 00:36:50,160 --> 00:36:50,600 ACCUMULATE. 904 00:36:50,600 --> 00:36:52,280 I SHOULD SAY THIS IS ALL A MODEL 905 00:36:52,280 --> 00:36:54,160 BECAUSE TO DATE THERE'S NOT YET 906 00:36:54,160 --> 00:36:57,000 A STRUCTURE OF A LIPID DROPLET 907 00:36:57,000 --> 00:36:57,200 PROTEIN. 908 00:36:57,200 --> 00:36:58,840 TURNS OUT ONLY SOME PROTEINS CAN 909 00:36:58,840 --> 00:36:59,960 DO THIS. 910 00:36:59,960 --> 00:37:02,440 WE THOUGHT CASE CLOSED, DONE. 911 00:37:02,440 --> 00:37:04,000 AS YOU FOLLOW ENDOGENOUS ENZYME 912 00:37:04,000 --> 00:37:05,520 AND NOT THE FRAGMENT OFF IT 913 00:37:05,520 --> 00:37:07,120 THERE'S A MYSTERY. 914 00:37:07,120 --> 00:37:09,040 YOU CAN FOLLOW LIPID DROPLET 915 00:37:09,040 --> 00:37:11,560 FORMATION FOR HOURS AND NOTHING 916 00:37:11,560 --> 00:37:11,800 HAPPENS. 917 00:37:11,800 --> 00:37:12,880 UNFORTUNATELY THIS PROJECTOR, 918 00:37:12,880 --> 00:37:15,080 YOU CAN'T SEE THE GREEN DIFFUSE 919 00:37:15,080 --> 00:37:16,920 BACKGROUND SIGNAL VERY WELL. 920 00:37:16,920 --> 00:37:19,120 WHAT YOU CAN APPRECIATE IS THAT 921 00:37:19,120 --> 00:37:20,920 AGAIN 350 MINUTES, NOTHING 922 00:37:20,920 --> 00:37:21,200 HAPPENED. 923 00:37:21,200 --> 00:37:23,360 WITHIN 10 MINUTES ONE OF THESE 924 00:37:23,360 --> 00:37:24,960 DROPLETS BECAME ACCESSIBLE AND 925 00:37:24,960 --> 00:37:26,960 TARGETS THAT ENZYME TO IT. 926 00:37:26,960 --> 00:37:30,600 AND THEN VERY RAPIDLY LIKE IN 927 00:37:30,600 --> 00:37:33,200 THE TRIGGERED FASHION THIS LIPID 928 00:37:33,200 --> 00:37:34,520 DROPLET ACQUIRES THIS PROTEIN. 929 00:37:34,520 --> 00:37:36,200 HOW DOES THAT WORK? 930 00:37:36,200 --> 00:37:37,920 THE FIRST QUESTION IF IT WAS IN 931 00:37:37,920 --> 00:37:40,120 THE E.R. WHY DOESN'T IT GET IN 932 00:37:40,120 --> 00:37:40,640 HERE? 933 00:37:40,640 --> 00:37:41,840 WELL, IT'S NOT HARD TO IMAGINE 934 00:37:41,840 --> 00:37:44,920 THAT IF YOU HAVE THIS LARGE MEGA 935 00:37:44,920 --> 00:37:48,680 DALTON COMPLEX AT THE BASE OF 936 00:37:48,680 --> 00:37:50,720 FORMATION IT DOESN'T LET 937 00:37:50,720 --> 00:37:51,400 EVERYTHING THROUGH, LARGER CARGO 938 00:37:51,400 --> 00:37:52,520 CAN THEN GO THERE. 939 00:37:52,520 --> 00:37:54,320 THIS IS THE CASE BECAUSE THE 940 00:37:54,320 --> 00:37:56,640 EXPERIMENT I JUST SHOWED YOU 941 00:37:56,640 --> 00:37:58,080 HERE SIMPLIFIED SAYS THAT, OKAY, 942 00:37:58,080 --> 00:38:00,040 IN A WILDTYPE CELL AFTER HALF AN 943 00:38:00,040 --> 00:38:05,280 HOUR EVEN THOUGH DROPLETS ARE 944 00:38:05,280 --> 00:38:08,800 FORM ENZYME IS NOT THERE, LATER 945 00:38:08,800 --> 00:38:12,520 IT IS, TAKE OUT SEIPIN, IT HOPS 946 00:38:12,520 --> 00:38:13,440 ON. 947 00:38:13,440 --> 00:38:14,400 THERE IS A PATHWAY BECAUSE AT 948 00:38:14,400 --> 00:38:16,480 THE END THEY END UP ON LIPID 949 00:38:16,480 --> 00:38:18,440 DROPLETS, THAT'S A REAL MYSTERY 950 00:38:18,440 --> 00:38:19,520 FOR US. 951 00:38:19,520 --> 00:38:21,720 IT LED US TO HYPOTHESIZE THERE 952 00:38:21,720 --> 00:38:23,640 MUST BE TWO PATHWAYS, ONE EARLY 953 00:38:23,640 --> 00:38:25,760 THAT ALLOWS THE ACCESS OF SOME 954 00:38:25,760 --> 00:38:29,360 PROTEINS TO LIPID DROPLETS, AND 955 00:38:29,360 --> 00:38:31,760 ONE LATER ONE THAT MAYBE USES 956 00:38:31,760 --> 00:38:32,560 INDEPENDENT BRIDGES. 957 00:38:32,560 --> 00:38:34,160 TURNS OUT AT LEAST IN TERMS OF 958 00:38:34,160 --> 00:38:36,200 THE TIMING THERE'S A WHOLE SLEW 959 00:38:36,200 --> 00:38:37,800 OF METABOLIC ENZYMES THAT APPEAR 960 00:38:37,800 --> 00:38:39,000 TO USE THAT. 961 00:38:39,000 --> 00:38:41,000 YOU MIGHT WONDER WHY SO 962 00:38:41,000 --> 00:38:42,440 COMPLICATED, I'M HAPPY TO TALK 963 00:38:42,440 --> 00:38:45,040 ABOUT THAT. 964 00:38:45,040 --> 00:38:46,400 SO TURNS OUT REALLY THERE ARE 965 00:38:46,400 --> 00:38:49,400 MULTIPLE MEMBRANE BRIDGES, THIS 966 00:38:49,400 --> 00:38:51,080 IS AN EXPERIMENT, IMMEDIATE TIME 967 00:38:51,080 --> 00:38:51,280 POINT. 968 00:38:51,280 --> 00:38:53,200 THERE'S SIGNAL ALREADY ON THE 969 00:38:53,200 --> 00:38:53,800 LIPID DROPLET. 970 00:38:53,800 --> 00:38:55,560 AT THE BEGINNING OF THE 971 00:38:55,560 --> 00:38:56,520 EXPERIMENT WE BLEACH THAT. 972 00:38:56,520 --> 00:38:59,160 WHAT YOU CAN SEE THE SIGNAL 973 00:38:59,160 --> 00:39:00,920 RECOVERS AND APPEARS TO RECOVER 974 00:39:00,920 --> 00:39:02,240 THROUGH MULTIPLE CONNECTIONS 975 00:39:02,240 --> 00:39:06,360 BETWEEN THE LIPID DROP LET 976 00:39:06,360 --> 00:39:09,600 SURFACE AND ENDOPLASM RETICULUM 977 00:39:09,600 --> 00:39:10,840 AND VISUALIZED BY 978 00:39:10,840 --> 00:39:17,320 CRYOTOMOGRAPHY, YOU CAN SEE 979 00:39:17,320 --> 00:39:18,400 E.R.s STUDDED WITH RIBOSOME, 980 00:39:18,400 --> 00:39:22,360 THIS IS COLORED FOR BETTER 981 00:39:22,360 --> 00:39:22,800 IMAGINATION. 982 00:39:22,800 --> 00:39:24,560 THAT TRIGGERS HOW WOULD YOU 983 00:39:24,560 --> 00:39:25,880 GENERATE SUCH MEMBRANE BRIDGES 984 00:39:25,880 --> 00:39:28,160 BETWEEN THE E.R. AND LIPID 985 00:39:28,160 --> 00:39:30,720 DROPLET INDEPENDENT OF THAT 986 00:39:30,720 --> 00:39:32,320 INITIAL FORMATION SITE? 987 00:39:32,320 --> 00:39:34,400 WE BANGED OUR HEAD FOR MANY 988 00:39:34,400 --> 00:39:35,960 YEARS FRANKLY AND HAVE A NUMBER 989 00:39:35,960 --> 00:39:39,120 OF IDEAS. 990 00:39:39,120 --> 00:39:40,760 EVENTUALLY DECIDED TO DO A 991 00:39:40,760 --> 00:39:42,600 VISUAL SCREEN KNOCKING OUT ONE 992 00:39:42,600 --> 00:39:44,920 GENE AT A TIME, EVERY GENE IN 993 00:39:44,920 --> 00:39:48,400 THE GENOME, ASKING WHICH ONES 994 00:39:48,400 --> 00:39:49,400 SPECIFICALLY ABOLISH THE 995 00:39:49,400 --> 00:39:50,400 RECRUITMENT OF LIPID DROPLET 996 00:39:50,400 --> 00:39:54,240 PROTEINS TO THIS BUT NOT OTHER 997 00:39:54,240 --> 00:39:58,840 CARGO, LIPID DROPLETS OR OTHER 998 00:39:58,840 --> 00:40:01,440 SECRETORY PATHWAYS, AND FOUND A 999 00:40:01,440 --> 00:40:03,880 CLASS OF TETHERING COMPONENTS, 1000 00:40:03,880 --> 00:40:05,720 MEMBRANE FUSION MACHINERY, AND 1001 00:40:05,720 --> 00:40:06,520 RECYCLING OF THAT MEMBRANE 1002 00:40:06,520 --> 00:40:08,560 FUSION THAT'S SPECIFICALLY 1003 00:40:08,560 --> 00:40:09,840 REQUIRED FOR THIS PATHWAY. 1004 00:40:09,840 --> 00:40:12,360 WE CAN SEE THOSE COMPONENTS NOT 1005 00:40:12,360 --> 00:40:13,360 ONLY ARE REQUIRED BUT THEY ARE 1006 00:40:13,360 --> 00:40:15,880 AT THE RIGHT TIME BECAUSE WE CAN 1007 00:40:15,880 --> 00:40:17,200 DO A SOMEWHAT COMPLICATED 1008 00:40:17,200 --> 00:40:22,840 EXPERIMENT HERE BUT JUST TO SHOW 1009 00:40:22,840 --> 00:40:24,240 WITHOUT EXPLAIN IN DETAIL, HAPPY 1010 00:40:24,240 --> 00:40:25,640 TO ANSWER QUESTIONS BIT, BUT WE 1011 00:40:25,640 --> 00:40:28,840 SET UP A SYSTEM WHERE YOU CAN 1012 00:40:28,840 --> 00:40:30,800 BASICALLY TEMPORARILY SEGREGATE 1013 00:40:30,800 --> 00:40:31,480 TRAFFICKING FROM FORMATION, AND 1014 00:40:31,480 --> 00:40:34,280 WHAT WE FOUND AS SOON AS WE 1015 00:40:34,280 --> 00:40:35,840 BASICALLY TRIGGERED, THESE ARE 1016 00:40:35,840 --> 00:40:37,280 MINUTES IN A CELL FUSION 1017 00:40:37,280 --> 00:40:38,120 EXPERIMENT, THE POTENTIAL FOR 1018 00:40:38,120 --> 00:40:40,520 THE PROTEIN TO GO ON TO LIPID 1019 00:40:40,520 --> 00:40:42,080 DROPLET, RIGHT BEFORE THAT, THE 1020 00:40:42,080 --> 00:40:44,160 PROTEIN WILL END UP ON THE LIPID 1021 00:40:44,160 --> 00:40:46,120 DROPLET, WE SEE THE MACHINERY WE 1022 00:40:46,120 --> 00:40:47,840 DISCOVERED, THIS IS ONE EXAMPLE, 1023 00:40:47,840 --> 00:40:50,600 WILL SHOW UP RIGHT AT WHAT WILL 1024 00:40:50,600 --> 00:40:52,320 BECOME A MEMBRANE FUSION SITE 1025 00:40:52,320 --> 00:40:53,440 BETWEEN THOSE TWO. 1026 00:40:53,440 --> 00:40:55,840 OUR BEST MODEL RIGHT NOW IS THAT 1027 00:40:55,840 --> 00:40:58,640 THERE IS AN INDEPENDENT 1028 00:40:58,640 --> 00:41:00,800 MACHINERY THAT INVOLVES E.R. 1029 00:41:00,800 --> 00:41:02,640 EXIT SITES AND MEMBRANE TETHERS 1030 00:41:02,640 --> 00:41:04,280 THAT ALLOWS FORMATION OF 1031 00:41:04,280 --> 00:41:06,120 SECONDARY CONTACT OR NOT 1032 00:41:06,120 --> 00:41:06,920 ACTUALLY CONTACT, MEMBRANE 1033 00:41:06,920 --> 00:41:08,480 BRIDGES BETWEEN THE E.R. AND 1034 00:41:08,480 --> 00:41:09,000 LIPID DROPLETS. 1035 00:41:09,000 --> 00:41:14,320 IF YOU THINK ABOUT IT, OF 1036 00:41:14,320 --> 00:41:16,520 COURSE, THIS MAY MAKE SOME 1037 00:41:16,520 --> 00:41:19,600 SENSE, ALLOWS YOU TO UNCOUPLE 1038 00:41:19,600 --> 00:41:21,200 PROTEIN COUPLING, THERE ARE MANY 1039 00:41:21,200 --> 00:41:23,200 CELL TYPES THAT DON'T MAKE A LOT 1040 00:41:23,200 --> 00:41:25,040 OF LIPID DROPLETS BUT STILL HAVE 1041 00:41:25,040 --> 00:41:26,040 TO CHANGE PROTEOME, MAYBE THIS 1042 00:41:26,040 --> 00:41:28,560 IS A PATHWAY REQUIRED FOR THAT. 1043 00:41:28,560 --> 00:41:30,000 HOWEVER, THERE'S MORE OPEN 1044 00:41:30,000 --> 00:41:32,280 QUESTIONS THAN ANSWERS HERE. 1045 00:41:32,280 --> 00:41:33,920 SO WE DON'T UNDERSTAND REALLY 1046 00:41:33,920 --> 00:41:34,800 HOW THIS BARRIER WORKS. 1047 00:41:34,800 --> 00:41:37,080 I CAN TELL YOU THE SIZE DOESN'T 1048 00:41:37,080 --> 00:41:37,560 MATTER. 1049 00:41:37,560 --> 00:41:38,840 IT'S NOT THE SINGLE 1050 00:41:38,840 --> 00:41:39,400 DISTINGUISHER HERE. 1051 00:41:39,400 --> 00:41:41,120 WE DON'T KNOW HOW THIS PATHWAY 1052 00:41:41,120 --> 00:41:43,120 HERE IS TRIGGERED AT ALL. 1053 00:41:43,120 --> 00:41:44,520 WE DON'T KNOW HOW PERSISTENT 1054 00:41:44,520 --> 00:41:46,640 BRIDGES ARE, WHETHER THEY ARE 1055 00:41:46,640 --> 00:41:48,600 ONE WAY OR TWO-WAY STREETS. 1056 00:41:48,600 --> 00:41:51,080 AND HOW THEY PARTICIPATE IN 1057 00:41:51,080 --> 00:41:51,840 PHYSIOLOGY. 1058 00:41:51,840 --> 00:41:54,400 SO WE'RE STILL BUSY FOR A WHILE. 1059 00:41:54,400 --> 00:41:55,920 WITH THIS I WANT TO SWITCH GEARS 1060 00:41:55,920 --> 00:41:56,800 FOR A SECOND HERE. 1061 00:41:56,800 --> 00:41:59,760 I TOLD YOU HOW TO MAKE A LIPID 1062 00:41:59,760 --> 00:42:03,000 DROPLET, GET THE PROTEINS ON 1063 00:42:03,000 --> 00:42:03,360 THAT MATTER. 1064 00:42:03,360 --> 00:42:05,720 ALL OF THIS OF COURSE PRESUMABLY 1065 00:42:05,720 --> 00:42:07,480 ONLY MATTERS IN CONTEXT WHERE 1066 00:42:07,480 --> 00:42:10,680 YOU ALSO HAVE MECHANISMS TO 1067 00:42:10,680 --> 00:42:11,760 MOBILIZE FATTY ACIDS, YOU WANT 1068 00:42:11,760 --> 00:42:13,280 TO STORE, PUT THINGS IN THE 1069 00:42:13,280 --> 00:42:15,560 BANK, YOU NEED TO WITHDRAW AT 1070 00:42:15,560 --> 00:42:16,800 LATER TIME POINT. 1071 00:42:16,800 --> 00:42:18,440 NOW, GENERALLY THERE ARE TWO 1072 00:42:18,440 --> 00:42:19,960 DISTRICT PATHWAYS HOW THIS CAN 1073 00:42:19,960 --> 00:42:21,160 BE ACHIEVED IN ALL THE CELLS 1074 00:42:21,160 --> 00:42:22,840 THAT PEOPLE HAVE LOOKED AT. 1075 00:42:22,840 --> 00:42:25,560 THE FIRST ONE IS THAT ENZYMES 1076 00:42:25,560 --> 00:42:27,880 SUCH AS APGL THAT SHOWED UP ON 1077 00:42:27,880 --> 00:42:30,080 MY SLIDES ALREADY CAN TARGET THE 1078 00:42:30,080 --> 00:42:35,000 SURFACE OF LIPID DROPLETS AND 1079 00:42:35,000 --> 00:42:36,080 ESSENTIALLY HYDROLASES THAT 1080 00:42:36,080 --> 00:42:39,600 CLEAVE ESTER BONDS, PARTICULARLY 1081 00:42:39,600 --> 00:42:42,000 THE FIRST ONE, HYDROLYZE 1082 00:42:42,000 --> 00:42:42,760 TRIGLYCERIDE AND RELEASE FATTY 1083 00:42:42,760 --> 00:42:46,040 ACID, THREE PACES TAKE THE 1084 00:42:46,040 --> 00:42:48,880 FIRST, SECOND, THIRD ACEEL 1085 00:42:48,880 --> 00:42:50,640 CHANGE, DISCOVERED BY RUDY 1086 00:42:50,640 --> 00:42:52,720 TECHNER, AND WHO HAS DONE 1087 00:42:52,720 --> 00:42:55,560 BEAUTIFUL WORK SHOWING HOW THIS 1088 00:42:55,560 --> 00:42:56,840 WORKS, PARTICULARLY IN 1089 00:42:56,840 --> 00:42:58,080 ADIPOCYTE, ESSENTIALLY NOT 1090 00:42:58,080 --> 00:42:59,720 UNDERSTOOD IN ANY OTHER CELL 1091 00:42:59,720 --> 00:43:00,240 TYPE. 1092 00:43:00,240 --> 00:43:03,000 THERE'S A SECOND PATHWAY THAT 1093 00:43:03,000 --> 00:43:04,520 YOU CAN IMAGINE WHICH WORKS 1094 00:43:04,520 --> 00:43:05,960 DIFFERENT, WHERE A LIPID DROPLET 1095 00:43:05,960 --> 00:43:11,640 OR PART OF A LIPID DROPLET IS 1096 00:43:11,640 --> 00:43:18,080 INTERNALIZED INTO AN 1097 00:43:18,080 --> 00:43:28,600 AUTOPHAGOSOME STRUCTURE, ACID 1098 00:43:30,960 --> 00:43:31,880 LIPase, THIS LIPO PHAGEY WAS 1099 00:43:31,880 --> 00:43:34,080 MORE THAN TEN YEARS AGO, 1100 00:43:34,080 --> 00:43:36,160 REPORTED IF YOU KNOCK OUT 1101 00:43:36,160 --> 00:43:37,160 AUTOPHAGY IN LIVER YOU 1102 00:43:37,160 --> 00:43:41,400 ACCUMULATE A LOT OF LIPID 1103 00:43:41,400 --> 00:43:42,160 DROPLETS. 1104 00:43:42,160 --> 00:43:43,840 THERE'S HUNDREDS OF PAPERS AFTER 1105 00:43:43,840 --> 00:43:44,360 THAT. 1106 00:43:44,360 --> 00:43:46,520 IT'S BEEN REALLY DIFFICULT TO 1107 00:43:46,520 --> 00:43:48,000 ASK, OKAY, HOW MUCH HAPPENS, HOW 1108 00:43:48,000 --> 00:43:49,440 DOES IT WORK AND REGULATE AND TO 1109 00:43:49,440 --> 00:43:51,840 ME AT LEAST MOST IMPORTANTLY HOW 1110 00:43:51,840 --> 00:43:53,320 DOES THIS CONTRIBUTE TO 1111 00:43:53,320 --> 00:43:53,640 PHYSIOLOGY. 1112 00:43:53,640 --> 00:43:55,520 BECAUSE THE ONLY TOOLS THAT WE 1113 00:43:55,520 --> 00:43:58,040 HAVE WERE BLUNT LIKE WE KNOCK 1114 00:43:58,040 --> 00:43:59,360 OUT AUTOPHAGY BUT NOTHING HAS 1115 00:43:59,360 --> 00:44:02,240 BEEN KNOWN SPECIFICALLY HOW DO 1116 00:44:02,240 --> 00:44:06,520 YOU CONNECT THE LIPID DROPLETS 1117 00:44:06,520 --> 00:44:07,280 TO THE AUTOPHAGESOME, AN 1118 00:44:07,280 --> 00:44:10,360 INTERESTING QUESTION. 1119 00:44:10,360 --> 00:44:13,920 LUCKILY ALSO TO BOB AND CHUNG. 1120 00:44:13,920 --> 00:44:18,440 THE DISCOVERY WAS SERENDIPITOUS, 1121 00:44:18,440 --> 00:44:20,280 STARTED WITH ASKING RANDOMLY FOR 1122 00:44:20,280 --> 00:44:25,440 LIPID DROPLET PROTEIN WHERE IS 1123 00:44:25,440 --> 00:44:26,400 IT LOCALIZED? 1124 00:44:26,400 --> 00:44:28,680 THIS PROTEIN IS CALLED SPARTAN. 1125 00:44:28,680 --> 00:44:30,560 WHAT WAS PECULIAR WAS THAT WHEN 1126 00:44:30,560 --> 00:44:33,320 WE LOCALIZE WITH A KNOCK-IN 1127 00:44:33,320 --> 00:44:35,600 ENDOGENOUS LOCUS WE FOUND THAT 1128 00:44:35,600 --> 00:44:38,560 IT LOCALIZED TO LIPID DROPLETS, 1129 00:44:38,560 --> 00:44:40,440 BECAUSE THEY STAIN WITH NEUTRAL 1130 00:44:40,440 --> 00:44:41,600 LIPID DYES, BUT DOESN'T LOCALIZE 1131 00:44:41,600 --> 00:44:49,720 TO THE SAME LIPID DROPLETS AS A 1132 00:44:49,720 --> 00:44:51,160 SURFACE MARKER PLIN3. 1133 00:44:51,160 --> 00:44:52,040 PARTICULARLY WE ASKED WHAT'S 1134 00:44:52,040 --> 00:44:53,040 DIFFERENT ABOUT THESE LIPID 1135 00:44:53,040 --> 00:44:55,960 DROPLETS THAN THE REST OF THE 1136 00:44:55,960 --> 00:44:56,880 LIPID DROPLETS? 1137 00:44:56,880 --> 00:45:00,000 WHAT HE DID WAS CO-LOCALIZED 1138 00:45:00,000 --> 00:45:04,400 WHAT MIGHT BE DIFFERENT AND 1139 00:45:04,400 --> 00:45:08,440 FOUND QUICKLY THAT THE SPARTIN 1140 00:45:08,440 --> 00:45:13,440 DROPLETS LOCALIZE FLEX TO A 1141 00:45:13,440 --> 00:45:17,080 LYSOSOME, AND ARE ALMOST ALWAYS 1142 00:45:17,080 --> 00:45:20,000 MARKED WITH AN LC3 PROTEIN, AND 1143 00:45:20,000 --> 00:45:24,440 CORE PART OF THE AUTOPHAGY 1144 00:45:24,440 --> 00:45:25,160 MACHINERY. 1145 00:45:25,160 --> 00:45:27,040 AND I WILL TELL YOU ABOUT 1146 00:45:27,040 --> 00:45:28,600 SPARTIN NOW. 1147 00:45:28,600 --> 00:45:30,880 WHAT IS THIS PROTEIN? 1148 00:45:30,880 --> 00:45:34,600 IT IS A PROTEIN THAT HAS A 1149 00:45:34,600 --> 00:45:36,920 COUPLE OF SUGGESTED DOMAINS. 1150 00:45:36,920 --> 00:45:38,640 THERE'S SOMETHING IN THE 1151 00:45:38,640 --> 00:45:40,800 N-TERMINUS THAT'S CALLED A 1152 00:45:40,800 --> 00:45:44,960 MICROTUBULE INTERACTING DOMAIN 1153 00:45:44,960 --> 00:45:46,600 WHICH MOSTLY LIKELY DOES NOT 1154 00:45:46,600 --> 00:45:48,240 INTERACT WITH TUBULES. 1155 00:45:48,240 --> 00:45:58,880 THE YOUR -- THERE'S A UBIQUITID 1156 00:46:01,120 --> 00:46:02,040 SENESCENCE DOMAIN, ASSOCIATED 1157 00:46:02,040 --> 00:46:03,360 THROUGH COMPLEX PHENOTYPE. 1158 00:46:03,360 --> 00:46:05,960 WHAT WE POSTULATED IS THAT MAYBE 1159 00:46:05,960 --> 00:46:08,600 THIS PROTEIN SERVES OR 1160 00:46:08,600 --> 00:46:10,320 HYPOTHESIZED AS A PROTEIN THAT 1161 00:46:10,320 --> 00:46:14,720 BRIDGES THE LIPID DROPLET AND 1162 00:46:14,720 --> 00:46:15,320 LYSOSOME OR AUTOPHAGIESOME. 1163 00:46:15,320 --> 00:46:16,800 I DON'T HAVE TIME TO SHOW THE 1164 00:46:16,800 --> 00:46:19,120 DATA BUT WHAT WE FOUND IS THAT 1165 00:46:19,120 --> 00:46:20,960 THE SENESCENCE DOMAIN IS 1166 00:46:20,960 --> 00:46:25,320 COMPOSED OF A SERIES OF REPEATED 1167 00:46:25,320 --> 00:46:26,840 HELICES, YOU KNOW, THERE'S A 1168 00:46:26,840 --> 00:46:28,480 WHOLE SET OF THEM IN HERE. 1169 00:46:28,480 --> 00:46:31,680 WHAT WE FOUND IS THAT THESE 1170 00:46:31,680 --> 00:46:33,640 PROTEINS ARE IN CELLS SUFFICIENT 1171 00:46:33,640 --> 00:46:36,440 TO TARGET A PROTEIN TO LIPID 1172 00:46:36,440 --> 00:46:37,360 DROPLETS, CONSISTENT WITH WORK 1173 00:46:37,360 --> 00:46:40,640 FROM OTHERS THAT WAS PUBLISHED 1174 00:46:40,640 --> 00:46:41,560 BEFORE. 1175 00:46:41,560 --> 00:46:44,480 IN ADDITION, THIS DOMAIN IN THE 1176 00:46:44,480 --> 00:46:46,240 MIDDLE REALLY LOOKS LIKE A PH 1177 00:46:46,240 --> 00:46:51,640 DOMAIN, AND CAN INTERACT IN AN 1178 00:46:51,640 --> 00:46:53,120 UNCANONICAL MOTIF WITH LC3. 1179 00:46:53,120 --> 00:46:54,400 AND WE'VE SHOWN DIRECTLY, I 1180 00:46:54,400 --> 00:46:58,680 DON'T HAVE TIME TO SHOW ALL OF 1181 00:46:58,680 --> 00:47:00,640 THIS, THAT THIS PROTEIN BINDS 1182 00:47:00,640 --> 00:47:01,400 DIRECTLY IN VITRO. 1183 00:47:01,400 --> 00:47:03,840 BASED ON THIS WE THINK THIS 1184 00:47:03,840 --> 00:47:05,360 PROTEIN SPARTIN MIGHT BE 1185 00:47:05,360 --> 00:47:08,440 FUNCTIONING AS RECEPTOR THAT 1186 00:47:08,440 --> 00:47:11,640 BRIDGES THE LIPID DROPLET 1187 00:47:11,640 --> 00:47:14,080 SURFACE TO THE AUTOPHAGYIC 1188 00:47:14,080 --> 00:47:14,360 MACHINERY. 1189 00:47:14,360 --> 00:47:16,320 AND AGAIN, THE PAPER WILL BE 1190 00:47:16,320 --> 00:47:17,160 PUBLISHED SHORTLY SO YOU CAN 1191 00:47:17,160 --> 00:47:19,920 HAVE A LOOK AT THAT. 1192 00:47:19,920 --> 00:47:29,560 BUT THIS BRINGS UP THE QUESTION 1193 00:47:29,560 --> 00:47:30,520 PROTEIN-PROTEIN INTERACTIONS ARE 1194 00:47:30,520 --> 00:47:33,160 NICE BUT WHAT DO THEY DO. 1195 00:47:33,160 --> 00:47:36,240 WE FUSED A FLUOROPHORE TO A 1196 00:47:36,240 --> 00:47:37,000 PROTEIN, PECULIAR FLUOROPHORE IN 1197 00:47:37,000 --> 00:47:40,240 AS MUCH AS IT HAS A DIFFERENT 1198 00:47:40,240 --> 00:47:45,880 EXCITATION MAXIMUM IN NEUTRAL 1199 00:47:45,880 --> 00:47:47,480 AND ACIDIC PHs, YOU CAN 1200 00:47:47,480 --> 00:47:48,960 DISTINGUISH IN THE CYTOSOL. 1201 00:47:48,960 --> 00:47:50,880 IT WILL BE -- HAS A GREEN 1202 00:47:50,880 --> 00:47:53,400 MAXIMUM, IF IT GOES TO ACIDIC 1203 00:47:53,400 --> 00:47:54,920 ENVIRONMENT SHIFTS TO RED 1204 00:47:54,920 --> 00:47:56,000 MAXIMUM, TRANSITION YOU WOULD 1205 00:47:56,000 --> 00:47:58,880 EXPECT FROM SOMETHING BEING 1206 00:47:58,880 --> 00:47:59,760 DELIVERED TO A LYSOSOME. 1207 00:47:59,760 --> 00:48:01,680 , FROM GREEN TO RED HERE. 1208 00:48:01,680 --> 00:48:05,080 WHAT WE FOUND, IF YOU TAKE 1209 00:48:05,080 --> 00:48:10,160 CELLS, LOAD WITH FATTY ACID, 1210 00:48:10,160 --> 00:48:11,800 WITHDRAW THE FATTY AS I.D. TO 1211 00:48:11,800 --> 00:48:16,920 FORCE THEM TO MOBILIZE STORES, 1212 00:48:16,920 --> 00:48:20,320 APPEARS TO DELIVER TO 1213 00:48:20,320 --> 00:48:21,520 COMPARTMENT BLUNTED WITHOUT 1214 00:48:21,520 --> 00:48:21,760 SPARTIN. 1215 00:48:21,760 --> 00:48:24,640 THIS IS A CRISPR KNOCKOUT. 1216 00:48:24,640 --> 00:48:27,000 CELLS THAT DO NOT HAVE SPARTIN 1217 00:48:27,000 --> 00:48:28,640 ACCUMULATE TRIGLYCERIDE BECAUSE 1218 00:48:28,640 --> 00:48:30,600 NOW THEY CAN'T MOBILIZE AS WELL. 1219 00:48:30,600 --> 00:48:36,400 THIS ACCUMULATION IS NOT DUE TO 1220 00:48:36,400 --> 00:48:38,240 DIFFERENCE IN SYNTHESIS BUT 1221 00:48:38,240 --> 00:48:39,800 DEGRADATION, ROUGHLY ON THE SAME 1222 00:48:39,800 --> 00:48:43,200 SCALE THAT YOU OBSERVED WITH 1223 00:48:43,200 --> 00:48:45,640 DEFECT, THAT YOU OBSERVE WITH 1224 00:48:45,640 --> 00:48:46,960 THE MUTATION OF AUTOPHAGY 1225 00:48:46,960 --> 00:48:48,560 MACHINERY THAT'S NOT ON THE 1226 00:48:48,560 --> 00:48:50,280 SLIDE, IF YOU MUTATE BOTH 1227 00:48:50,280 --> 00:48:51,600 TOGETHER THERE'S NO ADDITIONAL 1228 00:48:51,600 --> 00:48:51,880 PHENOTYPE. 1229 00:48:51,880 --> 00:48:53,680 WITH ALL OF THIS THE CURRENT 1230 00:48:53,680 --> 00:48:56,960 WORKING MODEL THAT WE HAVE IS 1231 00:48:56,960 --> 00:49:00,560 THAT SPARTIN DECORATES THE 1232 00:49:00,560 --> 00:49:02,520 SURFACE AND INTERACTS DIRECTLY 1233 00:49:02,520 --> 00:49:07,760 WITH LC3 AND DELIVERS THE 1234 00:49:07,760 --> 00:49:10,120 PROTEIN -- DELIVER LIPID 1235 00:49:10,120 --> 00:49:11,000 DROPLETS TO LYSOSOME FOR 1236 00:49:11,000 --> 00:49:15,600 DEGRADATION A LAYINGS, NOW 1237 00:49:15,600 --> 00:49:17,360 -- FOR 1238 00:49:17,360 --> 00:49:17,840 DEGRADATION. 1239 00:49:17,840 --> 00:49:20,280 WHY IS THIS CALLED SPARTIN? 1240 00:49:20,280 --> 00:49:24,480 IT'S ALSO KNOWN AS SPG20, FOUND 1241 00:49:24,480 --> 00:49:27,960 IN HUMAN MUTATIONS THAT CAUSE A 1242 00:49:27,960 --> 00:49:28,720 COMPLEX SPASTIC PARAPLEGIA, 1243 00:49:28,720 --> 00:49:32,680 TROYER SYNDROME. 1244 00:49:32,680 --> 00:49:37,360 THAT IS CHARACTERIZED BY 1245 00:49:37,360 --> 00:49:41,080 AUTOSOMAL PARAPLEGIA WITH MUSCLE 1246 00:49:41,080 --> 00:49:44,280 WEAKNESS, SMALL STATURE AND 1247 00:49:44,280 --> 00:49:45,280 NEUROLOGICAL DEFECTS, SPREAD ALL 1248 00:49:45,280 --> 00:49:47,440 OVER THE PROTEIN QUITE FRANKLY 1249 00:49:47,440 --> 00:49:49,200 BUT CONCENTRATE ALSO TO SOME 1250 00:49:49,200 --> 00:49:51,280 DEGREE ON THE MIDDLE DOMAIN AND 1251 00:49:51,280 --> 00:49:54,880 SOMEWHAT IN THE SENESCENCE 1252 00:49:54,880 --> 00:49:56,080 DOMAIN. 1253 00:49:56,080 --> 00:49:57,280 CONSISTENT WITH THIS IDEA, 1254 00:49:57,280 --> 00:50:00,160 SOMEHOW HAS SOMETHING TO DO WITH 1255 00:50:00,160 --> 00:50:02,040 NEURONS, HIGHLY EXPRESSED IN THE 1256 00:50:02,040 --> 00:50:05,960 BRAIN AND PARTICULARLY IN 1257 00:50:05,960 --> 00:50:06,280 NEURONS. 1258 00:50:06,280 --> 00:50:08,720 FRANKLY, THIS IS THE LAST PLACE 1259 00:50:08,720 --> 00:50:10,000 I SUSPECTED LIPID DROPLETS TO 1260 00:50:10,000 --> 00:50:11,000 MATTER. 1261 00:50:11,000 --> 00:50:12,960 IF YOU THINK ABOUT THE BRAIN, 1262 00:50:12,960 --> 00:50:15,240 NOT NECESSARILY THE FIRST PLACE 1263 00:50:15,240 --> 00:50:16,240 ADIPOSE TISSUE DELIVERED, THAT'S 1264 00:50:16,240 --> 00:50:18,000 WHERE PEOPLE LOOK, WHAT ABOUT 1265 00:50:18,000 --> 00:50:19,200 THE BRAIN? 1266 00:50:19,200 --> 00:50:21,040 WE MADE INITIALLY AS A FIRST 1267 00:50:21,040 --> 00:50:24,120 EXPERIMENT MADE NEURONS IN VITRO 1268 00:50:24,120 --> 00:50:25,320 FROM iPS CELLS, DIFFERENTIATE 1269 00:50:25,320 --> 00:50:27,160 EITHER WILD TYPE OR SPARTIN 1270 00:50:27,160 --> 00:50:28,160 KNOCKOUTS AND FOUND THAT ONLY 1271 00:50:28,160 --> 00:50:33,880 WHEN YOU DON'T HAVE SPARTIN YOU 1272 00:50:33,880 --> 00:50:34,880 APPEAR TO ACCUMULATE LIPID 1273 00:50:34,880 --> 00:50:37,040 DROPLETS IN THE CELL BODY. 1274 00:50:37,040 --> 00:50:38,760 THIS IS CELL CULTURE, I MOTOR 1275 00:50:38,760 --> 00:50:40,520 NEURONS ARE MAYBE A MODEL OF 1276 00:50:40,520 --> 00:50:41,920 NEURONS BUT ARE NOT IN THE 1277 00:50:41,920 --> 00:50:42,400 BRAIN. 1278 00:50:42,400 --> 00:50:43,800 WE DEVELOPED A SYSTEM TO TEST 1279 00:50:43,800 --> 00:50:47,120 THIS IN VIVO AND SPECIFICALLY 1280 00:50:47,120 --> 00:50:50,200 WHAT WE DID WAS PARK AT WAYNE 1281 00:50:50,200 --> 00:50:53,560 STATE DEVELOPED A SYSTEM WHERE 1282 00:50:53,560 --> 00:50:55,520 WE INJECT EITHER CONTROL OR 1283 00:50:55,520 --> 00:50:58,680 DOMINANT NEGATIVE FRAGMENT OF 1284 00:50:58,680 --> 00:51:00,320 SPARTIN WE DEVELOPED IN VITRO 1285 00:51:00,320 --> 00:51:01,040 AND ASKED EFFECT, EXPRESSED ONE 1286 00:51:01,040 --> 00:51:02,640 ON ONE SIDE OF THE MOUSE BRAIN, 1287 00:51:02,640 --> 00:51:04,080 THE OTHER ON THE OTHER SIDE OF 1288 00:51:04,080 --> 00:51:05,040 THE MOUSE BRAIN. 1289 00:51:05,040 --> 00:51:09,280 YOU CAN SEE CONTROL EXPRESSES 1290 00:51:09,280 --> 00:51:10,520 MUCH HIGHER, WE ASKED WHAT ARE 1291 00:51:10,520 --> 00:51:11,720 THE CONSEQUENCES. 1292 00:51:11,720 --> 00:51:13,560 WE FOUND THAT SPECIFICALLY CELLS 1293 00:51:13,560 --> 00:51:15,120 THAT EXPRESS THIS DOMINANT 1294 00:51:15,120 --> 00:51:16,640 NEGATIVE SPARTIN START TO 1295 00:51:16,640 --> 00:51:18,160 ACCUMULATE LIPID DROPLETS IN 1296 00:51:18,160 --> 00:51:20,040 THEIR CELL BODY. 1297 00:51:20,040 --> 00:51:23,560 AND THEN WHEN WE GO AND CUT OUT 1298 00:51:23,560 --> 00:51:25,080 THOSE REGIONS SPECIFICALLY WE 1299 00:51:25,080 --> 00:51:32,200 CAN SEE THAT THE BRAIN OF THOSE 1300 00:51:32,200 --> 00:51:33,000 ACCUMULATES TRIGLYCERIDE AND DAG 1301 00:51:33,000 --> 00:51:36,000 BUT THE REST OF THE LIPIDS ARE 1302 00:51:36,000 --> 00:51:37,560 NOT CHANGED. 1303 00:51:37,560 --> 00:51:39,200 WE THINK FOR SOME REASON 1304 00:51:39,200 --> 00:51:42,600 AUTOPHAGY APPEARS TO MATTER IN 1305 00:51:42,600 --> 00:51:44,000 THE NEURONS, REQUIRED FOR NORMAL 1306 00:51:44,000 --> 00:51:46,520 NEURONAL FUNCTION. 1307 00:51:46,520 --> 00:51:48,920 THAT'S SURPRISING, AND I DON'T 1308 00:51:48,920 --> 00:51:50,240 KNOW WHY YET. 1309 00:51:50,240 --> 00:51:51,600 WHAT IS TANTALIZING IS THAT WHEN 1310 00:51:51,600 --> 00:51:53,320 WE GO BACK WHAT I HAVEN'T TOLD 1311 00:51:53,320 --> 00:52:00,000 YOU YET THERE ARE MUTATIONS IN 1312 00:52:00,000 --> 00:52:01,720 SEIPIN, SPG17, THEY ARE 1313 00:52:01,720 --> 00:52:02,480 DIFFERENT MUTATIONS OF THE 1314 00:52:02,480 --> 00:52:06,760 SEIPIN THAT DON'T LEAD TO 1315 00:52:06,760 --> 00:52:07,960 LIPODYSTROPHY, MISSENSE 1316 00:52:07,960 --> 00:52:09,240 MUTATIONS MOSTLY. 1317 00:52:09,240 --> 00:52:13,880 THEY LEAD TO SPASTIC PARAPLEGIA 1318 00:52:13,880 --> 00:52:20,720 SPARTIN I TOLD YOU ABOUT. 1319 00:52:20,720 --> 00:52:23,600 CONCOMITANTLY DDHD2 IS A 1320 00:52:23,600 --> 00:52:25,000 TRIGLYCERIDE HYDROLYZED THAT'S 1321 00:52:25,000 --> 00:52:27,400 BRAIN SPECIFIC, AND THAT 1322 00:52:27,400 --> 00:52:31,040 MUTATION -- SO MUTATIONS IS 1323 00:52:31,040 --> 00:52:32,120 LEADING TO SPASTIC PARAPLEGIA. 1324 00:52:32,120 --> 00:52:34,640 THERE'S A SLEW OF MUTATIONS THAT 1325 00:52:34,640 --> 00:52:39,640 ARE KNOWN TO GIVE YOU SPASTIC 1326 00:52:39,640 --> 00:52:41,640 PARAPLEGIA, OFTEN AFFECT E.R. 1327 00:52:41,640 --> 00:52:43,880 FUNCTION AND LIPID DROPLETS. 1328 00:52:43,880 --> 00:52:47,240 WE'RE NOW QUITE INTERESTED TO 1329 00:52:47,240 --> 00:52:49,680 ASK WHAT MIGHT BE THE FUNCTION, 1330 00:52:49,680 --> 00:52:50,920 PARTICULARLY IN MOTOR NEURONS. 1331 00:52:50,920 --> 00:52:52,200 I'M HAPPY TO TALK BIT, CAN'T 1332 00:52:52,200 --> 00:52:53,440 ANSWER THE QUESTION TODAY. 1333 00:52:53,440 --> 00:52:56,400 WHAT I TOLD YOU TODAY, SOME 1334 00:52:56,400 --> 00:52:57,720 HIGHLIGHTS, I HOPE I HELPED YOU 1335 00:52:57,720 --> 00:53:00,360 UNDERSTAND A LITTLE BIT HOW WE 1336 00:53:00,360 --> 00:53:02,120 CURRENTLY THINK TRIGLYCERIDE 1337 00:53:02,120 --> 00:53:08,800 SYNTHESIS OCCURS IN THE DGAT 1338 00:53:08,800 --> 00:53:09,640 ENZYMES, THERE'S DEDICATED 1339 00:53:09,640 --> 00:53:11,640 MACHINERY THAT FORMS A VERY 1340 00:53:11,640 --> 00:53:13,600 LARGE MEGA DALTON COMPLEX THAT 1341 00:53:13,600 --> 00:53:15,240 ALLOWS FORMATION OF LIPID 1342 00:53:15,240 --> 00:53:16,880 DROPLETS OUT OF THE E.R., THERE 1343 00:53:16,880 --> 00:53:19,840 BY PROTECTING THE CELL FROM 1344 00:53:19,840 --> 00:53:21,080 LIPOTOXICITY OF DISTRIBUTED 1345 00:53:21,080 --> 00:53:24,720 LIPID INTERMEDIATES. 1346 00:53:24,720 --> 00:53:26,600 THERE ARE DISTINCT PATHWAYS TO 1347 00:53:26,600 --> 00:53:28,560 GET ONTO LIPID DROPLETS, SOME WE 1348 00:53:28,560 --> 00:53:30,440 START TO UNDERSTAND, TO SOME 1349 00:53:30,440 --> 00:53:30,880 DEGREE. 1350 00:53:30,880 --> 00:53:32,840 AND THAT THERE'S A FASCINATING 1351 00:53:32,840 --> 00:53:35,360 NEW AREA OF LIPID DROPLET 1352 00:53:35,360 --> 00:53:36,360 BIOLOGY WITH AUTOPHAGY IN THE 1353 00:53:36,360 --> 00:53:41,360 BRAIN WHERE WE START TO HAVE 1354 00:53:41,360 --> 00:53:42,600 MOLECULAR PLAYERS THAT ALLOW US 1355 00:53:42,600 --> 00:53:43,560 HOPEFULLY TO ANALYZE THE 1356 00:53:43,560 --> 00:53:46,280 PHYSIOLOGY OF ALL THESE 1357 00:53:46,280 --> 00:53:46,840 PROCESSES. 1358 00:53:46,840 --> 00:53:48,160 AND WITH THIS, I WANT TO 1359 00:53:48,160 --> 00:53:50,040 PARTICULARLY THANK THE PEOPLE 1360 00:53:50,040 --> 00:53:51,560 WHO HAVE DONE THE WORK. 1361 00:53:51,560 --> 00:53:57,240 I HIGHLIGHTED WORK BY CHUNG, A 1362 00:53:57,240 --> 00:53:58,520 FANTASTIC POSTDOC. 1363 00:53:58,520 --> 00:54:00,840 WHEN YOU'RE SUCCESSFUL AS A 1364 00:54:00,840 --> 00:54:03,040 POSTDOC YOU GO SOMEWHERE ELSE. 1365 00:54:03,040 --> 00:54:03,800 YOUR MENTOR STAYS. 1366 00:54:03,800 --> 00:54:08,720 IN OUR CASE SHE GOT A JOB AT 1367 00:54:08,720 --> 00:54:15,600 HARVARD, SHE STATE, WE LEFT. 1368 00:54:15,600 --> 00:54:17,280 SHE'S GREAT. 1369 00:54:17,280 --> 00:54:20,800 WORK ON MEMBRANE TRAFFICKING, A 1370 00:54:20,800 --> 00:54:24,160 FANTASTIC MD/PHD STUDENT, 1371 00:54:24,160 --> 00:54:25,680 STRUCTURAL BIOLOGY A DGATs AND 1372 00:54:25,680 --> 00:54:29,520 WORK ON SEIPIN, NOW FACULTY IN 1373 00:54:29,520 --> 00:54:34,560 TEXAS AND ARLT IS AN OUTSTANDING 1374 00:54:34,560 --> 00:54:35,400 BIOCHEMIST AT THE DANA-FARBER 1375 00:54:35,400 --> 00:54:36,760 WHO WORKED ON THE SEIPIN 1376 00:54:36,760 --> 00:54:37,280 STRUCTURE. 1377 00:54:37,280 --> 00:54:39,400 I WANT TO THANK OUR FUNDING, 1378 00:54:39,400 --> 00:54:41,040 PARTICULARLY NIH. 1379 00:54:41,040 --> 00:54:44,440 THANK YOU FOR SUPPORT OF OVER 1380 00:54:44,440 --> 00:54:46,600 MANY YEARS, PARTICULARLY NIGMS, 1381 00:54:46,600 --> 00:54:51,680 ALSO HHMI WITH THAT ALL TAKE ANY 1382 00:54:51,680 --> 00:54:53,720 QUESTIONS YOU MIGHT HAVE. 1383 00:54:53,720 --> 00:54:54,880 AND THE BLUEFIELD. 1384 00:54:54,880 --> 00:54:58,760 >> PLEASE USE THE MICROPHONES. 1385 00:54:58,760 --> 00:54:59,880 AND I'LL ANNOUNCE FIRST CME 1386 00:54:59,880 --> 00:55:05,680 CODE, IF I REMEMBER CORRECTLY 1387 00:55:05,680 --> 00:55:06,040 44463. 1388 00:55:06,040 --> 00:55:10,160 AND I GUESS THAT'S IT FOR NOW. 1389 00:55:10,160 --> 00:55:11,000 PLEASE. 1390 00:55:11,000 --> 00:55:12,960 >> VERY NICE PRESENTATION. 1391 00:55:12,960 --> 00:55:14,600 SO TWO-PART QUESTION. 1392 00:55:14,600 --> 00:55:15,720 THE SPARTIN PROTEIN IS THAT 1393 00:55:15,720 --> 00:55:18,400 SPECIFIC ONLY FOR LIPID DROPLETS 1394 00:55:18,400 --> 00:55:20,200 TO DELIVER THOSE TO THE 1395 00:55:20,200 --> 00:55:23,720 AUTOPHAGOSOME OR DO YOU FIND IT 1396 00:55:23,720 --> 00:55:25,880 ON MAYBE OTHER COMPARTMENTS OR 1397 00:55:25,880 --> 00:55:26,200 ORGANELLES? 1398 00:55:26,200 --> 00:55:27,520 >> YEAH, THANKS FOR THAT 1399 00:55:27,520 --> 00:55:27,800 QUESTION. 1400 00:55:27,800 --> 00:55:29,720 I SHOULD HAVE SAID THIS PROTEIN 1401 00:55:29,720 --> 00:55:31,320 WAS ALSO REPORTED IN A NUMBER OF 1402 00:55:31,320 --> 00:55:32,040 OTHER CONTEXTS. 1403 00:55:32,040 --> 00:55:37,720 FOR INSTANCE, IN THE CLEAVAGE OF 1404 00:55:37,720 --> 00:55:42,080 MITOTIC CELLS, WE TAG THE LOCUS 1405 00:55:42,080 --> 00:55:43,520 AND NEVER SEE IT OTHER THAN 1406 00:55:43,520 --> 00:55:48,640 DISTRIBUTED IN THE CELLS OR AT 1407 00:55:48,640 --> 00:55:52,800 LIPID DROPLETS CONSISTED WITH 1408 00:55:52,800 --> 00:55:54,680 WORK AT LOCALIZATION, ALSO 1409 00:55:54,680 --> 00:55:55,000 PROTEOMICS. 1410 00:55:55,000 --> 00:55:56,640 >> WHAT IS THE SIGNAL FOR 1411 00:55:56,640 --> 00:55:58,200 LOCALIZING TO THE LIPID 1412 00:55:58,200 --> 00:55:59,440 DROPLETS? 1413 00:55:59,440 --> 00:56:01,280 IS IT LIKE BY DEFAULT SITTING 1414 00:56:01,280 --> 00:56:03,120 THERE AND, YOU KNOW, TURN 1415 00:56:03,120 --> 00:56:03,360 ON/OFF? 1416 00:56:03,360 --> 00:56:05,480 >> YOU CAUGHT ME, RIGHT? 1417 00:56:05,480 --> 00:56:06,480 I SET THIS UP NICELY, WOULDN'T 1418 00:56:06,480 --> 00:56:08,680 IT BE NICE IF WE COULD 1419 00:56:08,680 --> 00:56:09,560 UNDERSTAND THE REGULATION, 1420 00:56:09,560 --> 00:56:11,520 THAT'S HOW THE REGULATION WORKS. 1421 00:56:11,520 --> 00:56:12,520 I DON'T KNOW. 1422 00:56:12,520 --> 00:56:16,760 IT IS -- YOU CAN SPECULATE 1423 00:56:16,760 --> 00:56:20,840 THERE'S A MONO YOUR BIC 1424 00:56:20,840 --> 00:56:22,040 WILLATION SITE EXTENDED TO POLY 1425 00:56:22,040 --> 00:56:23,240 UBIQUITIN CHAIN, THAT APPEARS TO 1426 00:56:23,240 --> 00:56:26,000 BE REGULATED BY THE METABOLIC 1427 00:56:26,000 --> 00:56:26,320 STATE. 1428 00:56:26,320 --> 00:56:31,040 AND CORRELATES WITH A CHANGE IN 1429 00:56:31,040 --> 00:56:31,360 LOCALIZATION. 1430 00:56:31,360 --> 00:56:34,440 WE DON'T KNOW HOW THAT WORKS, 1431 00:56:34,440 --> 00:56:36,400 WHAT TRIGGERS. 1432 00:56:36,400 --> 00:56:37,400 THERE'S CERTAIN STIMULI BUT ONES 1433 00:56:37,400 --> 00:56:39,000 IN VITRO ARE NOT PHYSIOLOGIC. 1434 00:56:39,000 --> 00:56:41,880 YOU DON'T GO AND WITHDRAW FATTY 1435 00:56:41,880 --> 00:56:45,120 ACIDS, SO WE DON'T KNOW IS THE 1436 00:56:45,120 --> 00:56:47,760 SHORT ANSWER ABOUT IT APPEARS 1437 00:56:47,760 --> 00:56:48,400 REGULATED. 1438 00:56:48,400 --> 00:56:49,840 I DON'T KNOW, THIS PROTEIN 1439 00:56:49,840 --> 00:56:52,040 APPEARS TO BIND SUBSET OF LIPID 1440 00:56:52,040 --> 00:56:53,360 DROPLETS, BUT THE SIGNAL YOU GET 1441 00:56:53,360 --> 00:56:55,560 IS ALL AROUND. 1442 00:56:55,560 --> 00:56:56,560 LIKE ONE PROTEIN BINDS, THERE 1443 00:56:56,560 --> 00:56:58,720 MUST BE SOMETHING DIFFERENT 1444 00:56:58,720 --> 00:56:59,520 ABOUT THESE DROPLETS, RIGHT? 1445 00:56:59,520 --> 00:57:01,240 WHY DO THIS ALL GO TO THAT 1446 00:57:01,240 --> 00:57:02,400 DROPLET AND NOT THE ONE NEXT TO 1447 00:57:02,400 --> 00:57:04,120 IT, AND I DON'T KNOW THAT 1448 00:57:04,120 --> 00:57:13,960 EITHER. 1449 00:57:13,960 --> 00:57:18,640 >> YOU MENTIONED TRANSFERASES, 1450 00:57:18,640 --> 00:57:29,000 AND PLASMA MEMBRANE, MUCH MORE 1451 00:57:29,000 --> 00:57:32,400 THAN ENDOPLASMATIC, IF YOU 1452 00:57:32,400 --> 00:57:34,000 EXTEND THAT TRANSPORTER CELLS, 1453 00:57:34,000 --> 00:57:36,160 THE PLASMA IN THE BRAIN, THE 1454 00:57:36,160 --> 00:57:44,560 THICKNESS OF THESE PLASMA 1455 00:57:44,560 --> 00:57:46,240 MEMBRANES, COMPOSITION VERSUS 1456 00:57:46,240 --> 00:57:48,440 ENDOPLASMIC RETICULUM, ARE 1457 00:57:48,440 --> 00:57:49,040 PROTEINS EMBEDDED HERE? 1458 00:57:49,040 --> 00:57:50,360 >> IF I UNDERSTAND YOUR 1459 00:57:50,360 --> 00:57:51,240 QUESTION CORRECTLY, YOU'RE 1460 00:57:51,240 --> 00:57:53,560 TALKING ABOUT A CONCEPT THAT'S 1461 00:57:53,560 --> 00:57:55,280 ALSO KNOWN AS HYDROPHOBIC 1462 00:57:55,280 --> 00:57:57,280 MISMATCH, THE LENGTH OF A 1463 00:57:57,280 --> 00:57:58,800 CHANNEL MEMBRANE DOMAIN IS 1464 00:57:58,800 --> 00:58:02,960 MATCHED TO NICKNESS OF -- 1465 00:58:02,960 --> 00:58:04,920 THICKNESS OF MEMBRANE, 1466 00:58:04,920 --> 00:58:05,920 INCREASING TOWARDS THE PLASMA 1467 00:58:05,920 --> 00:58:06,640 MEMBRANE, RIGHT? 1468 00:58:06,640 --> 00:58:07,440 IF THAT'S MATCHED. 1469 00:58:07,440 --> 00:58:09,200 IF YOU THINK BIT, LIPID DROPLET 1470 00:58:09,200 --> 00:58:12,840 IS THE EXTREME CASE OF A THICK 1471 00:58:12,840 --> 00:58:13,200 MEMBRANE. 1472 00:58:13,200 --> 00:58:15,560 YOU HAVE ONE SET OF 1473 00:58:15,560 --> 00:58:16,520 PHOSPHOLIPIDS, AND THE OCEAN 1474 00:58:16,520 --> 00:58:17,440 UNDER IT ESSENTIALLY. 1475 00:58:17,440 --> 00:58:19,240 WE DO THINK THAT MATTERS FOR 1476 00:58:19,240 --> 00:58:21,120 THAT MECHANISM BUT IT IS 1477 00:58:21,120 --> 00:58:24,320 IMPORTANT TO REMEMBER YOU CAN'T 1478 00:58:24,320 --> 00:58:26,160 PRESUMABLY HAVE TRANSMEMBRANE 1479 00:58:26,160 --> 00:58:29,120 DOMAIN, PER SE. 1480 00:58:29,120 --> 00:58:35,240 THE TRANS PART, NO OTHER LUMEN, 1481 00:58:35,240 --> 00:58:37,800 THEY ALL DIP. 1482 00:58:37,800 --> 00:58:42,160 THAT MAKES IT FUNDAMENTALLY 1483 00:58:42,160 --> 00:58:43,560 SOMEWHAT DIFFERENT. 1484 00:58:43,560 --> 00:58:45,560 >> PROTEIN TRANSFERASE 1485 00:58:45,560 --> 00:58:47,640 N-TERMINAL END IS TRANSMEMBRANE 1486 00:58:47,640 --> 00:58:48,400 DOMAIN. 1487 00:58:48,400 --> 00:58:50,880 IF YOU EXTEND, THE PROTEIN WOULD 1488 00:58:50,880 --> 00:58:52,000 GO TO PLASMA MEMBRANE. 1489 00:58:52,000 --> 00:58:52,880 >> I'M AWARE OF THAT. 1490 00:58:52,880 --> 00:58:55,280 I DO NOT KNOW WHETHER THAT PLAYS 1491 00:58:55,280 --> 00:58:57,360 A ROLE HERE. 1492 00:58:57,360 --> 00:58:59,520 I DID MENTION BECAUSE -- I 1493 00:58:59,520 --> 00:59:01,120 MENTIONED THIS EVERY ONCE IN A 1494 00:59:01,120 --> 00:59:01,760 WHILE. 1495 00:59:01,760 --> 00:59:03,720 SOMEONE WILL HAVE A CLUE AT SOME 1496 00:59:03,720 --> 00:59:04,160 POINT. 1497 00:59:04,160 --> 00:59:07,160 IF YOU JUST LOOK THROUGH 1498 00:59:07,160 --> 00:59:12,080 LITERALLY NOW HUNDREDS OF 1499 00:59:12,080 --> 00:59:13,160 PROTEOMIC STUDIESU GRIND YOUR 1500 00:59:13,160 --> 00:59:18,280 CELLS, PUT THE DROPLETS ON TOP, 1501 00:59:18,280 --> 00:59:20,760 MASS SPECTROMETRY, IT TELLS YOU 1502 00:59:20,760 --> 00:59:22,560 MORE ABOUT MASS SPECTROMETRY, 1503 00:59:22,560 --> 00:59:23,800 BUT THERE'S LOTS OF OTHER 1504 00:59:23,800 --> 00:59:24,480 THINGS. 1505 00:59:24,480 --> 00:59:26,280 IN HUNDREDS OF STUDIES ONE OF 1506 00:59:26,280 --> 00:59:30,480 THE THINGS THAT REPEATEDLY COMES 1507 00:59:30,480 --> 00:59:32,960 UP IS GLYCOSYLATION MACHINERY 1508 00:59:32,960 --> 00:59:34,160 LOCALIZED BY MICROSCOPY. 1509 00:59:34,160 --> 00:59:36,120 TO THIS DAY I HAVE ZERO IDEA WHY 1510 00:59:36,120 --> 00:59:37,800 THEY ARE THERE. 1511 00:59:37,800 --> 00:59:42,560 YOU CAN SPECULATE, THINK ABOUT 1512 00:59:42,560 --> 00:59:44,000 THE STRUCTURAL, RECEPTOR IN 1513 00:59:44,000 --> 00:59:46,320 LIPID DROPLETS BUT NOBODY KNOWS. 1514 00:59:46,320 --> 00:59:47,840 THAT'S A LONG WINDING ANSWER FOR 1515 00:59:47,840 --> 00:59:48,800 I HAVE NO CLUE. 1516 00:59:48,800 --> 00:59:51,880 >> A QUESTION ONLINE BEFORE YOU 1517 00:59:51,880 --> 00:59:52,040 GO. 1518 00:59:52,040 --> 00:59:52,320 >> SURE. 1519 00:59:52,320 --> 00:59:55,720 >> SO IN YOUR PICTURE OF THE 1520 00:59:55,720 --> 01:00:00,880 ENCAPSULATED LD IN LYSOSOME, TWO 1521 01:00:00,880 --> 01:00:02,240 SURFACES SEEM TO CONTACT. 1522 01:00:02,240 --> 01:00:03,720 DOES THE SNARE ET CETERA 1523 01:00:03,720 --> 01:00:11,720 MACHINERY PLAY A ROLE? 1524 01:00:11,720 --> 01:00:13,320 WHY IS ENCAPSULATION LITHOPHAGEY 1525 01:00:13,320 --> 01:00:13,560 NEEDED? 1526 01:00:13,560 --> 01:00:15,760 >> SO WE DO NOT KNOW ANY OTHER 1527 01:00:15,760 --> 01:00:26,240 MACHINERY AT THIS POINT THAT'S 1528 01:00:27,640 --> 01:00:33,400 REQUIRED FOR SPECIFIC 1529 01:00:33,400 --> 01:00:33,680 LITHOPHAGY. 1530 01:00:33,680 --> 01:00:35,480 NORMALLY PHAGOSOME FORMS AND 1531 01:00:35,480 --> 01:00:35,680 RECUSES. 1532 01:00:35,680 --> 01:00:37,520 WE DON'T UNDERSTAND WHY. 1533 01:00:37,520 --> 01:00:38,840 THE ENCAPSULATION PRESUMABLY IS 1534 01:00:38,840 --> 01:00:44,200 NEEDED TO ALLOW FOR THE ACTION 1535 01:00:44,200 --> 01:00:47,080 OF ACID LIPASE. 1536 01:00:47,080 --> 01:00:50,680 THERE'S A PROTEIN CALLED LAO 1537 01:00:50,680 --> 01:00:53,080 ONLY WORKS IN ACIDIC PH, 1538 01:00:53,080 --> 01:00:56,360 KNOCKOUT MICE ARE IMPORTANT FOR 1539 01:00:56,360 --> 01:00:56,840 MOBILIZING TRIGLYCERIDE. 1540 01:00:56,840 --> 01:00:58,240 THEY DO NOT KNOW THE OTHER 1541 01:00:58,240 --> 01:01:05,480 MACHINERY THAT'S INVOLVED IN 1542 01:01:05,480 --> 01:01:07,680 THIS PROCESS YET. 1543 01:01:07,680 --> 01:01:10,080 >>CURVATURE OF LIPID DROPLETS 1544 01:01:10,080 --> 01:01:11,360 CHANGE A LOT FROM WHEN BORN TO 1545 01:01:11,360 --> 01:01:12,800 WHEN THEY ARE LARGE. 1546 01:01:12,800 --> 01:01:14,520 IS THERE EVIDENCE THAT PLAYS A 1547 01:01:14,520 --> 01:01:15,600 ROLE IN ACTIVATING ENZYMES THAT 1548 01:01:15,600 --> 01:01:18,360 SIT ON THE SURFACE OF THE LIPID 1549 01:01:18,360 --> 01:01:18,600 DROPLET? 1550 01:01:18,600 --> 01:01:24,000 >> SHORT ANSWER IS NO. 1551 01:01:24,000 --> 01:01:26,600 ACTUALLY THERE'S -- IF YOU GO TO 1552 01:01:26,600 --> 01:01:27,680 THE BIOCHEMISTRY OF THE PROCESS 1553 01:01:27,680 --> 01:01:29,200 THERE'S A LOT OF MYSTERY. 1554 01:01:29,200 --> 01:01:32,480 WE HAVE NOT WORKED ON THIS SO I 1555 01:01:32,480 --> 01:01:34,360 DON'T KNOW ABOUT BESIDES WHAT'S 1556 01:01:34,360 --> 01:01:34,640 PUBLISHED. 1557 01:01:34,640 --> 01:01:40,800 IF YOU THINK ABOUT FOR INSTANCE 1558 01:01:40,800 --> 01:01:42,240 APGL, THE LIPASE, SITS ON THE 1559 01:01:42,240 --> 01:01:45,320 SURFACE OF LIPID DROPLETS, 1560 01:01:45,320 --> 01:01:46,920 PERFECTLY CAPABLE OF DOING WHAT 1561 01:01:46,920 --> 01:01:49,120 IT DOES, BUT IT'S INACTIVE. 1562 01:01:49,120 --> 01:01:49,760 THERE'S ANOTHER PROTEIN THAT'S 1563 01:01:49,760 --> 01:01:52,520 ALSO ON THE SURFACE OF LIPID 1564 01:01:52,520 --> 01:01:54,920 DROPLETS, AND THAT NEEDS TO BIND 1565 01:01:54,920 --> 01:02:00,600 THAT PROTEIN TO ACTIVATE THE 1566 01:02:00,600 --> 01:02:01,160 LIPASE. 1567 01:02:01,160 --> 01:02:04,440 THEY ARE BOTH THERE, MASSIVE 1568 01:02:04,440 --> 01:02:07,520 CONCENTRATIONS, RIGHT? 1569 01:02:07,520 --> 01:02:09,240 BECAUSE -- BUT CAN'T INTERACT 1570 01:02:09,240 --> 01:02:10,240 BECAUSE THAT'S BINDING SOMETHING 1571 01:02:10,240 --> 01:02:10,480 ELSE. 1572 01:02:10,480 --> 01:02:12,680 ALL THESE THINGS ARE STILL IN 1573 01:02:12,680 --> 01:02:14,520 THE PHASE WHERE WE HAVE 1574 01:02:14,520 --> 01:02:17,400 ESSENTIALLY SOME VERY HIGH LEVEL 1575 01:02:17,400 --> 01:02:18,400 CONCEPTS BUT REALLY NOBODY 1576 01:02:18,400 --> 01:02:20,560 UNDERSTANDS, NO STRUCTURES FOR 1577 01:02:20,560 --> 01:02:21,880 THESE THINGS, BIOCHEMISTRY IS 1578 01:02:21,880 --> 01:02:25,240 REALLY NOT VERY WELL UNDERSTOOD. 1579 01:02:25,240 --> 01:02:26,560 AND DESPITE THAT THERE ARE DRUGS 1580 01:02:26,560 --> 01:02:31,400 FOR INSTANCE LIKE THE APLG 1581 01:02:31,400 --> 01:02:36,880 INHIBITORS PIONEERED FOR KAKEXIA 1582 01:02:36,880 --> 01:02:39,480 CANCER AND OTHER THINGS. 1583 01:02:39,480 --> 01:02:48,000 I HAVE NO ANSWER, HAVEN'T 1584 01:02:48,000 --> 01:02:48,680 EXPERIMENTED WITH THAT. 1585 01:02:48,680 --> 01:02:49,120 >> THANKS. 1586 01:02:49,120 --> 01:02:54,000 >> GREAT TALK, LOOKING AT DGAT1 1587 01:02:54,000 --> 01:02:56,440 AND 2, 1 HAVING LATERAL GATE DO 1588 01:02:56,440 --> 01:02:59,400 YOU THINK VERSUS CYTOSOLIC 1589 01:02:59,400 --> 01:03:02,560 ORIENTED SITE OF DGAT2 IS IT 1590 01:03:02,560 --> 01:03:04,560 ACCESSING FROM DIFFERENT 1591 01:03:04,560 --> 01:03:07,280 LEAFLETS, WHETHER IN THE 1592 01:03:07,280 --> 01:03:10,560 CYTOPLASMIC OR INNER LEAFLET? 1593 01:03:10,560 --> 01:03:12,840 >> I THINK THE GENERAL IDEA IN 1594 01:03:12,840 --> 01:03:15,040 THE FIELD IS THAT THEY CAN 1595 01:03:15,040 --> 01:03:16,800 FLIP-FLOP FAST BECAUSE THEY HAVE 1596 01:03:16,800 --> 01:03:19,880 ESSENTIALLY NO POLAR HEAD GROUP. 1597 01:03:19,880 --> 01:03:26,520 WHAT IS SPECIFIC ABOUT DGAT1, IT 1598 01:03:26,520 --> 01:03:28,120 TAKES FROM THE E.R. 1599 01:03:28,120 --> 01:03:34,200 WE'VE SHOWN, AN MANY OTHERS, 1600 01:03:34,200 --> 01:03:35,760 SOME IN CANADA, THAT DGAT2 1601 01:03:35,760 --> 01:03:38,160 LOCALIZES IS THE SURFACE OF 1602 01:03:38,160 --> 01:03:47,120 LIPID DROPLETS, WE DON'T DON'TN 1603 01:03:47,120 --> 01:03:48,440 DROPLET OR CONTACT SIDES, 1604 01:03:48,440 --> 01:03:53,880 TEMPTING TO THINK IT'S 1605 01:03:53,880 --> 01:03:55,960 DETOXIFYING BUT DGAT 2 LOCALLY 1606 01:03:55,960 --> 01:03:56,280 EXPANDS. 1607 01:03:56,280 --> 01:03:59,040 HORTON AT SOUTHWESTERN HAS 1608 01:03:59,040 --> 01:04:01,440 BEAUTIFUL WORK SUGGESTING 1609 01:04:01,440 --> 01:04:02,520 CHANNELED BETWEEN LIPOGENESIS TO 1610 01:04:02,520 --> 01:04:04,280 TRIGLYCERIDE SYNTHESIS BUT CELL 1611 01:04:04,280 --> 01:04:06,160 BIOLOGICAL BASIS IS NOT YET 1612 01:04:06,160 --> 01:04:06,520 KNOWN. 1613 01:04:06,520 --> 01:04:07,880 >> THAT LEADS INTO THE 1614 01:04:07,880 --> 01:04:11,280 FOLLOW-UP, TALKED ABOUT 1615 01:04:11,280 --> 01:04:12,840 TARGETING ENZYMES TO DROPLETS, 1616 01:04:12,840 --> 01:04:21,160 GPAT4 IS THE BASIS OF LIVEDROP 1617 01:04:21,160 --> 01:04:26,600 SENSOR, MAKING LISO LIPID. 1618 01:04:26,600 --> 01:04:28,360 DO YOU THINK THAT THEY ARE 1619 01:04:28,360 --> 01:04:30,440 PLAYING A ROLE? 1620 01:04:30,440 --> 01:04:36,560 EXIT SITES ARE ALSO ALLEGEDLY 1621 01:04:36,560 --> 01:04:38,240 ENROICHED IN LISO LIPIDS. 1622 01:04:38,240 --> 01:04:38,960 >> GOOD QUESTIONS AND 1623 01:04:38,960 --> 01:04:40,080 EMBARRASSING I DON'T HAVE 1624 01:04:40,080 --> 01:04:40,400 ANSWERS. 1625 01:04:40,400 --> 01:04:43,360 I CAN TELL YOU IF YOU JUST DO 1626 01:04:43,360 --> 01:04:45,840 CRUDE EXPERIMENTS AND PULL ON 1627 01:04:45,840 --> 01:04:53,200 GPATS AN CELLS YOU'LL FIND THE 1628 01:04:53,200 --> 01:04:53,640 EKPAT. 1629 01:04:53,640 --> 01:04:57,480 PULLING ON DGAT YOU CAN FIND 1630 01:04:57,480 --> 01:04:58,920 THINGS THAT DESATURATE. 1631 01:04:58,920 --> 01:05:01,720 THERE'S A FAIRLY HIGH CHANCE, 1632 01:05:01,720 --> 01:05:03,720 SOME PEOPLE WHO JAY HORTON 1633 01:05:03,720 --> 01:05:06,680 PARTICULARLY, BEAUTIFUL WORK ON 1634 01:05:06,680 --> 01:05:08,160 THE FATTY ACID SYNTHASE COMING 1635 01:05:08,160 --> 01:05:10,720 TOGETHER ON A COMPLEX AROUND 1636 01:05:10,720 --> 01:05:12,240 LIPID DROPLETS, THERE'S A FAIRLY 1637 01:05:12,240 --> 01:05:14,320 HIGH CHANCE THERE'S SOME SORT OF 1638 01:05:14,320 --> 01:05:18,920 CELLULAR ORGANIZATION OF THIS 1639 01:05:18,920 --> 01:05:19,240 PATHWAY. 1640 01:05:19,240 --> 01:05:25,240 BUT IT'S JUST A CHANCE. 1641 01:05:25,240 --> 01:05:26,680 HONESTLY THIS FIELD HASN'T BEEN 1642 01:05:26,680 --> 01:05:27,560 NECESSARILY A CENTER OF 1643 01:05:27,560 --> 01:05:29,080 ATTENTION FOR CELL BIOLOGY FOR 1644 01:05:29,080 --> 01:05:31,200 MANY YEARS, AND THEN THE TOOLS 1645 01:05:31,200 --> 01:05:34,360 ARE ONLY NOW CATCHING UP. 1646 01:05:34,360 --> 01:05:35,880 IF YOU OVEREXPRESS THESE 1647 01:05:35,880 --> 01:05:37,840 ENZYMES, HELL BREAKS LOOSE AND 1648 01:05:37,840 --> 01:05:39,360 EVERYTHING IS EVERYWHERE. 1649 01:05:39,360 --> 01:05:39,800 VERY HARD TO STUDY. 1650 01:05:39,800 --> 01:05:42,960 >> THANK YOU SO MUCH. 1651 01:05:42,960 --> 01:05:44,400 >> TRY TO MAKE IT QUICK. 1652 01:05:44,400 --> 01:05:47,920 SURFACE OF THE LIPID DROPLETS, 1653 01:05:47,920 --> 01:05:50,400 BECAUSE OF NON-CANONICAL NATURE, 1654 01:05:50,400 --> 01:05:51,640 I WOULD IMAGINE THE PROTEINS 1655 01:05:51,640 --> 01:05:54,040 THAT BIND AND PROTEIN STRUCTURES 1656 01:05:54,040 --> 01:05:56,360 ARE ALSO NON-CANONICAL. 1657 01:05:56,360 --> 01:05:57,880 CURIOUS FOR EXAMPLE TWO LIKE 1658 01:05:57,880 --> 01:06:03,440 ALPHA FOLD BASES ON TRAINING 1659 01:06:03,440 --> 01:06:04,040 SETS PROBABLY RATHER 1660 01:06:04,040 --> 01:06:06,000 UNREPRESENTED IN WHAT YOU NEED 1661 01:06:06,000 --> 01:06:07,560 TO PREDICT STRUCTURES AT 1662 01:06:07,560 --> 01:06:07,840 PROTEINS. 1663 01:06:07,840 --> 01:06:11,440 CAN YOU SHARE YOUR THOUGHTS 1664 01:06:11,440 --> 01:06:12,040 ABOUT THAT PROBLEM? 1665 01:06:12,040 --> 01:06:15,440 >> YEAH, THERE ARE NO PUBLICLY 1666 01:06:15,440 --> 01:06:17,920 AVAILABLE STRUCTURES FOR A FULL 1667 01:06:17,920 --> 01:06:19,640 LENGTH LIPID DROPLET PROTEIN. 1668 01:06:19,640 --> 01:06:24,680 THERE'S BEEN SOME BEAUTIFUL WORK 1669 01:06:24,680 --> 01:06:28,040 ON LIPOPROTEINS, THE OTHER LIPID 1670 01:06:28,040 --> 01:06:30,000 DROPLETS, MUCH OLDER AND BETTER 1671 01:06:30,000 --> 01:06:31,640 STUDIED BUT NOT THREE 1672 01:06:31,640 --> 01:06:34,280 DIMENSIONAL STRUCTURES, MORE 1673 01:06:34,280 --> 01:06:37,680 LIKE HOW DOES THIS WORK? 1674 01:06:37,680 --> 01:06:42,280 YEAH, BECAUSE OF THAT WE DON'T 1675 01:06:42,280 --> 01:06:44,240 FIND ALPHAFOLD AS VALUABLE FOR 1676 01:06:44,240 --> 01:06:45,720 THIS PROTEIN AS WE FIND FOR 1677 01:06:45,720 --> 01:06:48,080 OTHER THINGS, E.R. PROTEINS, 1678 01:06:48,080 --> 01:06:50,040 WOW, THIS IS AMAZING, AND THEN 1679 01:06:50,040 --> 01:06:54,120 WE GET CryoEM MAP AND JUST 1680 01:06:54,120 --> 01:06:56,160 LOOKS REALLY BLOWN AWAY BY 1681 01:06:56,160 --> 01:06:56,920 POTENTIAL. 1682 01:06:56,920 --> 01:06:58,360 WE'VE NOT HAD THAT EXPERIENCE 1683 01:06:58,360 --> 01:06:59,800 WITH LIPID DROPLET PROTEINS 1684 01:06:59,800 --> 01:07:02,200 WHERE OFTEN YOU GET SOMETHING 1685 01:07:02,200 --> 01:07:03,640 THAT LOOKS LIKE A RANDOM 1686 01:07:03,640 --> 01:07:06,480 COIL-ISH THING WITH A BUNCH OF 1687 01:07:06,480 --> 01:07:08,520 HELICES LEFT OR RIGHT, OR WHERE 1688 01:07:08,520 --> 01:07:09,880 IT'S HARD TO MAKE SENSE. 1689 01:07:09,880 --> 01:07:10,280 >> THANK YOU. 1690 01:07:10,280 --> 01:07:14,320 >> I DON'T THINK IT'S THAT 1691 01:07:14,320 --> 01:07:15,960 USEFUL YET. 1692 01:07:15,960 --> 01:07:16,960 >> THANK YOU. 1693 01:07:16,960 --> 01:07:19,280 WE DO HAVE SOME OTHER GREAT 1694 01:07:19,280 --> 01:07:22,800 QUESTIONS BUT WE'LL GET THEM TO 1695 01:07:22,800 --> 01:07:24,640 YOU ELSEWHERE FROM OUR 1696 01:07:24,640 --> 01:07:24,960 COLLEAGUES. 1697 01:07:24,960 --> 01:07:25,120 YES? 1698 01:07:25,120 --> 01:07:35,480 >> [OFF MICROPHONE]. 1699 01:07:46,680 --> 01:07:46,920 >> NOT YET. 1700 01:07:46,920 --> 01:07:47,520 >> I WANT TO THANK YOU AGAIN. 1701 01:07:47,520 --> 00:00:00,000 [APPLAUSE]