>> WE WOULD LIKE TO WELCOME ADRIAN HAYDAY TO NIH AND I FEEL THAT WE HAVE A COUPLE OF CONNECTIONS BECAUSE ONE OF MY FORMER POST DOCTORAL FELLOWS HAS BEEN IN HIS DEPARTMENT AT KING'S COLLEGE LONDON FOR A PERIOD OF TIME AND BEEN EXTREMELY GOOD TO HER. AND THEN OF COURSE I DID THE SEMIFEAUP A UX OF OFFERING CLAUDIA A JOB, AFTER SHE LEFT HIM BUT HE SAID WHATEVER IS BEST FOR HER, SHE IS WONDERFUL BUT ANYWAY, ADRIAN IS A NEW LEADER AT THE CRICK INSTITUTE AND CONTINUES TO HAVE HIS CHAIR AT KING'S COLLEGE LONDON AS WELL AS CO LEADING A CLINICAL ACADEMIC GROUP'S AT KING'S HEALTH PARTNERS. UNDERGRADUATE CAMBRIDGE AND GOT HIS Ph.D. AT UNIVERSITY OF LONDON, CAME TO THE U.S. AT M. I.T. WHERE HE AND HIS COLLEAGUES DISCOVERED THE TCR GAMMA CHAIN WHICH LAUNCHED HIS CAREER CHAIN RELATED TO DELTA GAMMA CELLS DHS --WHICH IS THE FOCUS OF HIS TALK TODAY. LET ME JUST SAY THAT HE IS THE RECIPIENT OF MANY AWARDS INCLUDING WHILE AT YALE THE WILLIAM DEVAIN MEDAL WHICH IS THEIR HIGH HONOR SCHOLARSHIP FOR A FELLOW AND AS WELL AS ELECTED TO THE ACADEMY OF SCIENCES AT IT IS ROYAL INSTITUTE AND HEADING IMMUNOLOGY. HE'S A DYNAMIC SPEAKER AND HOPEFULLY HE WILL NOT TELL YOU ABOUT HIS DIFFICULTIES OF TRAVEL COME COMING TO THE NIH BUT INSTEAD ONLY ABOUT SCIENCE. SO ... >> [ APPLAUSE ] >> THANK YOU. SO IT'S A REAL PLEASURE TO BE HERE. YOU KNOW WE'VE BEEN GOING THROUGH SOME POLITICAL PROBLEMS IN BRITAIN LATELY AND IT SEEMS TO ME, YOU HAVE, TOO. I JUST WANT TO MAKE SURE THAT IF SOMEONE HANDS ME A NOTE AFTER 20 MINUTES, I WILL COMPLETE THE SEMINAR, I'M NOT GOING TO LEAVE OFFICE IMMEDIATELY. SO I'M GOING TO TALK TO YOU TODAY ABOUT EPITHELIAL MOLECULES SHAPING IMMUNE SURVEILLANCE BY LOCAL T-CELLS AND I THOUGHT THAT YOU KNOW I REALLY IN FRONT OF THIS WONDERFUL AUDIENCE HERE AND THE SCHOLARSHIP AND THE INSTITUTION, YOU KNOW I CAN'T DO THAT IN ISOLATION OF THE CONTEXT IN WHICH IMMUNOLOGY HAS MOVED. SO, IT'S THE TRANSFORMATION IN TUMOR IMMUNOLOGY WHICH OF COURSE, AS YOU CAN SEE HERE AND I COULD HAVE PICKED ANY NUMBER OF OTHER EXAMPLES, HAS CAPTURED THE PUBLIC IMAGINATION, CAPTURED PRESS ATTENTION AND CAPTURED A VAST AMOUNT OF INVESTMENT AND IT'S ABSOLUTELY CLEAR THAT ALPHABETTA T-CELLS PLAY SIGNIFICANT ROLES IN THESE ENDEAVORS. AND AT A CERTAIN LEVEL YOU CAN SAY, HAVE PROVEN TO BE PRETTY GOOD DRUGS. SO, THE ISSUE OF COURSE IS WHAT REALLY IS THE UNDERLYING BASIS FOR THAT. SO ON THE ONE HAND ON A SIMPLE LEVEL, THIS IS VERY ATTRACTIVE APPROACH TO DEALING WITH CANCER BECAUSE IT OFFERS THE OPPORTUNITY OF AN ADAPTIVE ANTIGEN SPECIFIC RESPONSE THAT IS COMMITTED TO MEMORY AND THEREFORE MAY ERADICATE THE POTENTIAL THOUGHT CANCER CELL RESURGENCE. AND IN THAT REGARD, PEOPLE ARE UNDERSTANDABLY CONFLATED WITH THE NOTIONS ORIGINALLY PUT PERIOD BY PAUL EHRLICH AND CHAMPIONED BY Mc FAR LANE BURNETT THAT THERE IS A NATURAL EVER OPERATIVE IMMUNE O SURVEILLANCE SYSTEM THAT IS CONSTANTLY MONITORING THE POTENTIAL FOR THE DEVELOPMENT AND GROWTH OF TRANSFORMED CELLS AND ESSENTIALLY CUTTING IT OFF EARLY ON BUT IN FACT, I WOULD ARGUE THAT THE EVIDENCE THAT ALPHABETTA T-CELLS COMPOSE SUCH A COMPARTMENT IS REALLY RATHER SCANT AND ONE OF THE REASONS THAT I THINK I CAN SAY THAT IS IT'S VERY EVIDENT THAT THE CONSIDERABLE SUCCESS IN ALPHABETTA T-CELL MEDIATED IMMUNOTHERAPY IS ONLY REALIZED WHEN YOU USE PHARMACOLOGICAL MODULATIONS TO PREVENT THE NATURAL STATE WHICH IS ONE OF QUIESCENCE. AND SO, IF YOU THINK ABOUT THIS CROWD OF INDIVIDUALS, I DON'T THINK ANY OF US CAN REALLY ARGUE THAT THERE'S ANY INDIVIDUAL IN THIS PICTURE WHO WE CAN KNOW FOR SURE HAS NATURALLY SUPPRESSED A TUMOR THROUGH AN ALPHABETTA T-CELL MEDIATED IMMUNE T-CELL SYSTEM WITHOUT SOME FORM OF FARM PHARMACOLOGICAL PREVENTION PROMOTING THAT ACTIVITY. WELL YOU MIGHT SAY, OKAY, WELL, ADRIAN, IF YOU FEEL THIS WAY, MAYBE WE SHOULD LOOK AT DATA I BIT MORE SCIENTIFICALLY. AND OF COURSE, THE SORT OF DATA WE WOULD LOOK AT WOULD BE DATA WHETHER THERE'S ALPHABETTA T-CELL IMMUNE O DEFICIENCIES AND IF YOU LOOK AT THOSE INDIVIDUALS UNFORTUNATE ENOUGH TO HAVE ACQUIRED IMMUNE O DEFICIENCY, THEY SHOW INCREASED CANCER BUT AS WE ALL KNOW THOSE CANCERS ARE EXTRAORDINARILY ENRICHED IN THOSE WITH VIRAL ETIOLOGY. AND SO YOU MIGHT SAY, WELL, LIKEWISE, PEOPLE IN THE TRANSPLANT WARD UNDERGO SUPPRESSION SHOW INCREASED CANCER, THAT'S UNDOUBTEDLY THE IN SQUEAMOUS CELL CARC NOAMS AND UNDOUBTLY HAVE HPV. DESPITE MANY, MANY ATTEMPTS THERE HAS BEEN LITTLE CAPACITY TO LINK MHC GENERATEDET INCREASE IN BODYS WITH THE NATURAL DEVELOPMENT OF CANCER AND THERE HAVE BEEN A NUMBER OF SYSTEMS THAT PEOPLE MAY CHOOSE TO IGNORE IF THEY WISH BUT THEY'RE NOT TERRIBLY BAD LYE UNDERTAKEN EXPERIMENTS SUCH AS THIS NATURE PAPER FROM VAL INKSY AND BLANKENSTEIN AND WHICH UNDERLYING 184 DEVELOPING CANCERS IN MICE AND IN EVERY CASE, THE TUMOR ANTIGENS INDUCED TOLERANCE TO THESE TUMORS DURING THEIR DEVELOPMENT. SO THERE ISN'T REALLY A WHOLE LOT OF SUPPORTIVE EVIDENCE FOR THE IMMUNE O SURVEILLANCE THEORY THERE, SO NOW YOU CAN GO TO ALPHABETTA T-CELL DEFICIENT MICE AND IT MAY SURPRISE SOME YOUNG PEOPLE WHO MAY JUST ARKTS SIEWM THAT IF YOU TAKE ALPHABETTA T-CELL DEFICIENT MICE THEY MUST BE MORE SUSCEPTIBLE TO CANCERS BECAUSE THESE ARE THE TOOLS WE ARE USING IN THE CLINIC BUT IN FACT THE EVIDENCE THAT ALPHABETTA T-CELL MICE SHALL INCREASE SEPTORSIBILITYIBILITY TO CANCER IS SCANT, NOT NONEXISTENT BUT SCANT. WE PUBLISHED THIS OURSELVES AND WE PUBLISHED IN THIS PAPER THAT IN SOME MODELS YOU CAN SEE AN EFFECT BUT VERY OFTEN, MOST OFTEN YOU CAN'T. SO THAT OF COURSE LEADS TO A QUESTION WHICH IS, IS THERE A NATURAL IMMUNE O SURVEILLANCE SYSTEM. AND, YOU KNOW, I MAY NOT BE ABLE TO ANSWER THAT BY THE END OF THIS SEMINAR BUT I WILL GIVE YOU SOME THINGS TO THINK ABOUT, I HOPE AND GIVEN WAR REPUBLICAN'S INTRODUCTION, IT WON'T SURPRISE YOU TO KNOW THAT THIS IS THE FOCUS FOR THIS CONCONJECTURE. SO THIS WAS A PARTNERSHIP WE PUBLISHED IN 2001 WHEN THE PURSUIT OF CANCER IMMUNOLOGY WAS SOMETHING EQUIVALENT TO SAN FRANCISCO TANNIC--SAN FRANCISCO FRANCISCO--SATANIC WITCH CRAFT AND WE WERE BEHIND BOB IN PRODUCING GAMMA INTERFERON FROM LYMPHOCYTES. AND THERE IS NO DOUBT WHATSOEVER THAT IN THE SYSTEMS THAT WE USE,& WHICH WERE PRIMARILY CAR SIN O GENERATED--CARCINOGEN SCWAIMOUS CELL CARCINOMA WAS INVARIABLE TIED TO CANCER. WELL YOU MAY BE SAYING ADRIAN THAT A LOT'S MOVED ON AND ACTUALLY CAN YOU FIND PAPERS IN RELATIVELY HIGH IMPACT JOURNALS SUCH AS THIS ONE HERE THAT SEEM TO SUGGEST COMPLOATLY THE OPPOSITE TO WHAT I JUST SUGGESTED TO YOU AND IN FACT GAMMA DELTA T-CELL CAN PROMOTE CANCER AND OF COURSE, WE KNOW THESE PAPERS WELL AND IN THAT REGARD, I JUST LIKE TO MAKE A DISTINCTION THAT IS GOING TO BE IMPORTANT FOR US DURING THIS SEMINAR AND OUR EXPLORATION OF THE IDEA THAT A TISSUE SURVEILLANCE COMPARTMENT MIGHT ACTUALLY EXIST AND THAT IS THIS ONE HERE, SO IF YOU LOOK IN A MOUSE SKIN, YOU WILL FIND GAMMA DELTA T-CELLS SAT WITHIN THE CELL SURFACE AND YOU MAY FIND THAT THEY HAVE INTERFERON GAMMA DELTA AND THEY MAKE LIMP FOE TACTIN AND THEY NEVER REALLY MAKE IL17. AND YET, JUST BENEATH THE BASEMENT MEMBRANE IN THE TERMIS YOU--DERMIS YOU WILL FIND THE CELLS WHOSE ROLE IN LIFE IS TO MAKE IL17. THIS MICROANATOMICAL THING MAKES THIS IN THE GUT, THE EPITHELIAL CELLS DON'T MAKE IL17 AND THE ONES WITH THE LAM MA PROARP RIA, DO. AND THE TUMOR OF THE MOUSE HAS BEEN ALMOST EXCLUSIVELY DESIGNED TO THOSE THAT MAKE IL17 AS YOU JUST SAW. WHAT I WANT TO FOCUS ON TODAY ARE THESE CELLS OUT HERE THAT SIT WITHIN THE EPITHELIAL TISSUES BY AND LARGE THROUGHOUT LIFE IN WHICH DON'T MAKE IL17 AND WHICH I AM GOING TO SUGGEST MAY BE A NATURAL IMMUNE SURVEILLANCE MECHANISM THAT KEEPS TISSUES IN GOOD SHAPE. SO HERE ARE EXAMPLES OF THEM, THESE WILL BE FAMILIAR TO MANY OF YOU, THIS IS A VIEW IN PLAN OF THE MOUSE SKIN. THE LEVEL THAT WE'RE FOCUSING ON HERE IS THE--IS THE SUPER BASAL EPIDERMIS AND YOU SEE THESE EXTREMELY ATTRACTIVE LOOKING DENDRITIC CELLS THESE ARE NOT LONGER HAN CELLS, THESE CAN BE SEEN IN RED, THESE ARE DENDRITIC T-CELLS. EPIDERMAL DENDRITIC T-CELLS. AND THESE ARE IN THE SMALL INTESTINE ARE INTEREPITHELLIAL GAMMA DELTA T-CELLS. AND IT WAS THESE LOCAL CELLS, THESE GAMMA DELTA T-CELLS SHOWN HERE THAT WE SHOWED SEVEN YEARS AFTER THAT SCIENCE PAPER. IT'S THE LOCAL T-CELLS ENTIRELY THAT ARE RESPONSIBLE FOR THE GAMMA DELTA PROTECTION IN THOSE CARCINOGEN MODELS, NOT THE SYSTEMIC CELLS. SO, SO, THAT WE COULD ESTABLISH. NOW THERE ARE TWO PRETTY STUPID ORGANISMS IN THIS PICTURE AND IT'S THIS ONE HERE THAT I WANT TO FOCUS ON BECAUSE THIS AS MANY OF YOU WILL KNOW IS THE LAMP RAY AND THIS IS DEFINITELY THE MOST HANDSOME SIDE. THE LAMP RAY PUTS ITS ADAPTIVE IMMUNE SYSTEM TOGETHER IN A COMPLETELY DIFFERENT WAY. TO US AS YOU KNOW IT DOES NOT USE A RECOMBIN ACE ACTIVATING GENE. IT USING THE ASSEMBLY OF MOLECULES CALLED VARYING LUCE RECEPTORS BUT WHAT HAS BEEN STRIKING BY THESE GROUPS SEVERAL GROUPS WHERE THEY PROBABLY TAKE THE FOREMOST ROLE IS THAT JUST LIKE OUR OWN USE OF RECOMBIN ACE ACTIVATING GENES THERE ARE THREE PRODUCTS OF THIS PROCESS, A, B, & C AND FOR US IT WOULD BE ALPHABETTA T-CELLS, B-CELLS AND GAMMA DELTA CELLS AND INDEPENDENT, BERM AND COOPER WORKING TOGETHER PUBLISHED IN NATURE, A FEW YEARS AGO THIS VERY INTERESTING ANALYSIS, THIS--I REALIZE MANY OF YOU WILL NOT KNOW IMMEDIATELY THAT HAD IS THE LAMP RAY SMALL INTESTINE, BUT THAT'S WHAT IT IS AND THIS IS THE BASEMENT MEMBRANE HERE AND YOU CAN SEE THIS EPITHELIUM BEYOND THE BASEMENT MEMBRANE AND WHAT YOU SEE IS THIS EXTRAORDINARY ENRICHMENT IN THE EPITHELIAL LAYER OF THE VLRC POSITIVE CELLS THAT ARE STAINED IN GREEN WHICH BOEHM AND COOPER REGARD AS GAMMA DELTA LIKE T-CELL EQUIV LEBTS WHICH I WOULD BE HAPPY TO DISCUSS LATER. AND WITH REGARD TO THESE CELLS IN WHICH THEY ARE ENRICHED THROUGHOUT THE SYSTEMIC PARTNER. AND MUCH IS TRUE IN THE SAME SKIN. SO THIS ASSOCIATION, THIS INTIMATE ASSOCIATION OF THESE GAMMA DELTA T-CELLS WITH THE EPITHELIAL CELLS SITTING OUT ABOVE THE BASEMENT MEMBRANE MAY REFLECT A VERY CONSERVED COMPONENT OF TISSUE IMMUNITY AND IT WOULD THEREFORE BE A BIT SURPRISING IF SOMETHING THAT WAS CONSERVED IN SOME WAY SHAPE OR FORM FROM HAG FISH ALL THE WAY THROUGH MICE IS NOT SURPRISING IN HUMANS. THAT WOULD BE SLIGHTLY SURPRISING AND WE DON'T BELIEVE IT IS THE EXAMPLE, THIS IS OUR CAPACITY TO OVERCOME A TECHNICAL BARRIER WHICH IS TO VISUALIZE THESE GAMMA DELTA T-CELLS IN SITUE, IN TISSUE, AND THIS IS TAKEN FROM A COLON AND YOU SEE IN THESE CRYPTS THAT THEY HAVE THESE ASSOCIATED WITH THEM, THESE CELLS WITH THE GAMMA DELTA T-CELLS AND YOU CAN PICK THEM UP QUITE EASILY AND WE'RE ABLE TO ISOLATE THESE CELLS AND LEARN A LOT ABOUT THEM. NOW IT'S BEEN KNOWN FOR A VERY LONG TIME THAT ONE OF THE PROBLEMS WITH PUTTING ADAPTIVE CELLS, LYMPHOCYTES WITH POTENTIALLY ADAPTIVE RECEPTORS WITHIN TISSUES IS THE SAMPLING.& THAT IS TO SAY IF YOU HAVE IMMENSE DIVERSITY IN THE RECEPTOR, WHAT IS THE LIKELIHOOD YOU ARE GOING TO FIND IN PROXIMITY, AN ANTIGEN THAT FITS THAT BILL. SO ONE SOLUTION FOR THE GAMMA DELTA T-CELLS THAT SIT WITHIN TISSUES IS FOR THEM TO ADOPT AN ALTERNATIVE MODEUS OPERATING GLOBALLY RANDOMIZED TRIAL I, AND THAT WE WERE ABLE TO SHOW IN A SERIES OF PAPERS THAT I WILL SUMMARIZE NOW BECAUSE I WANT WANT TO DEVOTE MOST OF MY ATTENTION TO UNPUBLISHED DATAY. SO THIS WOULD BE A SCHEMATIC OF ONE OF THESE T-CELLS AND IN ADDITION TO THE T-CELL CELL RECEPTOR, THESE ACTIVATE THE RECEPTOR NGKTWO D THAT WE FIRST LEARNED ABOUT IN THE NK CELLS. EXPH WHAT IS STRIKING IS THAT WHEN WE LOOK AT T-CELLS THAT ARE TAKEN FROM THE SKIN OR GUT, OR HUMAN SKIN OR GUT OR MAMMARY EPITHELIUM. THESE GAMMA DELTA CELLS WILL RESPOND FULLY TO THE ENGAGEMENT OF A LIGAND FOR THIS NGGTWO D RECEPTOR. THIS IS AN INNATE FORM OF ACTIVATION WHEREAS CONVENTIONAL CD-EIGHT T-CELLS COME FROM EXACTLY THE SAME TISSUE STILL ARE CONTINGENT UPON TCR SIGNALING. AND THIS IS ACQUIRED DEVELOPMENTALLY AFTER CELLS ARE SELECTED ON THE BASIS OF THEIR T-CELL RECEPTOR TO ENTER THE TISSUE. SO IT'S SORT OF LIKE A TWO-STEP PROCESS OF SPECIFICITY BEING ESTABLISHED DURING DEVELOPMENT WHERE UPON ESSENTIALLY INNATE LIKE OR CO-STIMULATION IS SUFFICIENT TO DRIVE THE RESPONSE. SO THE QUESTION IS, WHAT KIND OF DISREGULATION DRIVES THAT RESPONSE? AND THAT WAS STUDIED BY A VERY TALENTED POST DOC IN MY LAB WHO PUBLISHED THIS PAPER A COUPLE YEARS AGO, SHOWING THAT THE MAJOR DRIVER IN HUMAN EPITHELIA, FOR THE UPREGULATION OF LIGANDINGS, MICA, FOR NKGTWO D IS DISREGULATION OF THE RECEPTOR THAT IS INVOKED BY A NUMBER STRESSORS INCLUDING OXIDATIVE OZ MOTTIC STRESS. NOW YOU WILL BE AWARE OF THE FACT THAT EGF DISREGULATION IS EXTREMELY IMPORTANT IN CARC FOAMA AND THERE WAS AN ENORMOUS ANALYSIS PUBLISHED BY MEDICINE A YEAR AND HALF AGAIN WHERE THEY USE TECHNOLOGY WHICH IS BIER NO MEANS PERFECT BUT THEY ANALYZE CANCER PATIENTS AND THE SEVENTH MOST POSITIVELY CORRELATING GENE WITH OVERALL SURVIVAL WAS NKGTWO D, BUT WHO WAS MOST SURPRISING OF OVERALL SURVIVAL WAS A HUMAN GAMMA DELTA T-CELL SIGNATURE ASSOCIATED WITH THE TUMOR. CAN THAT TO US CREATES NOT ONLY A SCIENTIFIC INTEREST BUT ALMOST A SOCIAL OBLIGATION TO UNDERSTAND BETTER THE WAY IN WHICH THESE CELLS GET INTO TISSUES AND THE WAY THEY'RE ATTAINED THERE AND REGULATED AND I NOW WANT TO USE THE DATA SLIDES TO SHOW YOU HOW WE APPROACH THAT. SO IN SUM, IF YOU ASK TO LOOK AT GAMMA T-CELL DEFICIENT MICE, THEN THOSE WHICH LACK LYMPHOCYTES ARE INVARIABLY SUSCEPTIBLE TO INCREASE CANCERS OF THOSE SITES AND ALTHOUGH HUMANS WITH GAMMA DELTA DEFICIENCY HAVE NOT BEEN DESCRIBED IN ANY LENGTH, THE IMPLICATION OF CYBER SORT IS THE INSUFFICIENCIES OF GAMMA DELTA T-CELL SIGNALS LEADS TO DECREASE CANCER SURVIVAL. SO, THE PROBLEM YOU COULD ARGUE WITH THE MODEL THAT I'VE SHOWN YOU FOR STRESS SURVEILLANCE IS THAT IT'S VERY GENERIC SO THE IDEA WOULD BE THAT THE NKGTWO D COULD BE ACTIVATED ON AN EPITHELIAL CELL AND IT DOESN'T MATTER IF THAT IS IN THE SKIN OR GUT OR PREOF THE OR WHATEVER AND THE NKGTWO D POSITIVE CELL WILL RESPOND. THE INCOMPLETE ASPECT OF THAT IS WE'VE KNOWN FOR LITERALLY ABOUT TWO AND HALF DECADES THAT THESE INTERHEPATITIS E THELLIAL COMPARTMENTS ARE ACTUALLY ORPG AN SPECIFIC.--AND IN THE INTESTINE AS WE'L L SEE IN A MOMENT, V-GAMMA SEVEN. SO THAT OF COURSE LED TO THE HYPOTHESIS THAT THERE MUST BE REGULATION OR THERE IS PERHAPS REGULATION OF THESE COMPARTMENTS BY ORGAN SPECIFIC SELECTING ELEMENTS AND MANY IN THE FIELD WERE--WERE STRUGGLING WITH THIS ISSUE AND TRYING TO MAKE IN-ROADS INTO THIS ISSUE FOR MANY, MANY YEARS WHICH LEADS ME TO INTRODUCE AN OLD FASHIONED IDEA IN ACADEMIA WHICH IS ONE OF SEBATICLE. SO I DON'T KNOW HOW GENEROUS YOU BOSSES ARE AT SUPPORTING YOUR APPLICATION FOR SEBATICLES BUT IT TURNS OUT THAT THEY'RE USEFUL. NOW SOME OF YOU MAY RECOGNIZE THIS IS BOB, HE'S SORT OF SMILING BECAUSE HE LIVESOT BEACH AND HE KNOWS I DON'T BUT I WAS VISITING HIM AND MORE IMPORTANTLY THAT FOLLOWED ON HIM VISITING US ON SEBATICLE IN LONDON WHEN THE TWO OF US MADE THIS DISCOVERY AND THIS IS NOW WELL KNOWN BUT IT'S TURNED OUT TO BE AN IMPORTANT PLATFORM FOR THE SCIENCE THAT FOLLOWS. SO I MENTIONED TO YOU BRIEFLY THAT THE CELLS IN THE EPIDERMIS OF THE V-GAMMA FIVE POSITIVE AND THIS IS A CLONE O TYPIC ANTIBODY THAT BOB'S LAB RAISED MANY YEARS AGO THAT DETECTS THESE CELLS AND THESE ARE THE MICE FROM THE JACKSON LAB AND WHAT BOB AND I DISCOVERED WAS THAT DURING THE REDERIVATION OF THE FEB STRAIN TAKEN--THEY CONIC FARMS HAD A BOTTLENECK HAD ARISEN AND THE V-GAMMA FIVE V-DELTA ONE CELLS WERE LOST FROM THE SKIN REPLEASED BY GAMMA DELTA CELLS USING A DIFFERENT COMPARTMENT. SO WE THOUGHT MAYBE THIS WAS GOING TO BE THE ROOT IN TO UNDERSTANDING HOW THESE SKIN SPECIFIC T-CELL COMPARTMENTS ARE ESTABLISHED. AND TO MAKE A LONG STORY SHORT, THAT DID IMPROVE--INDEED PROVE USEFUL. WE AND OTHERS WERE ABLE TO SHOW THAT THE PROGENITORS OF THE SKIN CELLS INTO THE FETAL LIVER WHERE THERE'S A CRITICAL INTERACTION THAT ENABLINGS THE CELLS TO MATURE AND ENTER THE SKIN PREPARED FOR INNATE RESPONSIVENESS OF THE KIND THAT I DESCRIBED AND IT TURNS OUT IF YOU LOOK AT THE TAKEN--THEY CONIC STRAIN OF THE MICE, IT DOESN'T WORK AND IT DOESN'T WORK NOT BECAUSE OF SOMETHING WRONG WITH THE PROGENITORS BUT BECAUSE THERE'S SOMETHING WRONG WITH THE THYMIC EPITHELIUM. AND IT BECAME CLEAR THIS WAS A MONOGENIC RECESSIVE PROBLEM SO WE DECIDED TO GO ABOUT CLONING THIS MOLECULE AND BECAUSE THIS WAS A--LONG BEFORE DEEP SEQUENCING, THIS WAS DONE IN THE CLASSICAL GENETIC FASHION, WITH CONSIDERABLY INPUT FROM RICK LIFTON AND THE RESULT WAS SOMETHING COMPLETELY NEW. SO THIS IS A SCHEME OF THE MOLECULE WE CALL SKIM ONE. YOU MIGHT THINK THAT'S SKIM T, BUT ACTUALLY THIS WAS 2008, WHICH WAS THE CREDIT CRISIS, SO NO ONE HAD ANY MONEY, EVERYBODY WAS SKIM--SCINT SO WE WE LIKE THAT IDEA BUT THIS LINKS THE DOMAINS TO THE RATHER JUXTAPOSING POSITION OF TRANSMEMBRANES. AND WHEN YOU SOLVED THE STRUCTURE WHICH WAS--WHICH WE DID WITH BEN WILCOX GROUP, AT LEAST AT NMR WILLFUL, YOU CAN SEE THAT THE V-DOMAIN HERE, WHICH IS SHOWN IN ORANGE OR YELLOW SUPER IMPOSES RATHER STRIKINGLY WITH ANOTHER KNOWN STRUCTURE IN THE DATABASE WHICH IS SHOWN IN RED. INTRIGUINGLY ENOUGH, THAT'S PDL ONE. SO THIS MOLECULE ALTHOUGH IT WAS NOVEL IS NOT COMPLETELY DISSIMILAR TO THOSE THAT ARE RATHER INTENSE EVALUATION PROCESSLY STUDIED. BY LOOKING ACROSS THE WHOLE SERIES OF MOUSE STRAINS THIS BECAME VERY CLEAR THIS IS POLYMORPHIC AND POLYMORPHIC IN AN INTERESTING WAY IN THAT THE MUTATIONS OF ARE DISPROPORTIONALLY ENRICHED IN THOSE THAT GENERATE NONCONSERVATIVE AMINO ACID CHANGES WHICH OF OF COURSE REMINISCENT OF MHC. AND THE LEVEL OF THAT POSITIVE SELECTION IN THIS GENE SO FAR AS WE CAN TELL IS SECOND ONLY TO THAT OF MHC. ALTHOUGH I WOULD STAND TO BE CORRECTED BY SOMEBODY WHO KNOWS THAT MATERIAL BETTER. SO WHAT DOES SCINT ONE DO. THIS IS WORKED OUT BY A VERY TALENTED POST DOC IN MY LAB, TURCHINOVICH AND CONTINUED ON BY MELANIE WENCKER, IT ENGAGES THESE INTEREPITHELLIAL T-CELLS AND AS A CONSEQUENCE THE GENETIC PROGRAM OF THESE CELLS IS CHANGED 180-DEGREES AS IT WERE. GAMMA IS SWITCHED OFF. TBED IS SWITCHED ON, THE COMPETENCES I DESCRIBED TO YOU, IMMERGE BECOME INNATE TO RESPONSE SIGNALS WHERE THEY ARE NOT AND THEY BECOME NOT SURPRISINGLY VERY HIGH IN THE IL15 RECEPTOR CD122. AND WITH THAT LEVEL OF MATURATION, GO OFF TO THE SKIN AND FUNCTION AS WE'VE CONSIDERED. HOWEVER, SCINT ONE IS NOT EXPRESSED SOLELY IN THE THYMIC EPITHELIUM. IT'S EXPRESSED IN SUPER BASAL KERATINOCYTES WHICH ARE EXACTLY THE CELLS THAT THE GAMMA DELTA T-CELLS ARE NEIGHBORING. SO OF COURSE THE QUESTION BECAME PERHAPS AS A FUNCTION FOR THE SCINT ONE EXPRESSED IN THE KERATIN O CITES, THESE ARE THE MATURITY CELL COMPARTMENT. SO IN 2012, TALK THOMAS JYLES GROUP IN TEXAS HAD SHOWN THREE PRETTY FINE IMAGING PROTOCOLS THAT THESE GAMMA DELTA T-CELLS AT ESSENTIALLY STEADY STATE ARE CONSTANTLY ENGAGEING THE EPITHELIAL CELLS AT FOCAL POINTS WHERE THE T-CELL RECEPTOR IS CONCENTRATED AND THESE ARE ACTIVE POINTS BECAUSE THEY'RE CDTHREE ZETA PHOSPHORALATION BUT THE SIGNALING DOES NOT PROGRESS FURTHER SO YOU GET NOR ERK AND YOU GET NO CALCIUM FLUX AND WE WERE ABLE TO VISUALIZE THOSE WHEN WE ADOPTED THESE TECHNIQUES AND THIS IS JUST A FAIRLY STANDARD CONFOCAL. OBVIOUSLY THE EPITHELIAL CELLS ARE MASKED BUT WHAT YOU CAN SEE ARE THESE POINTS WHERE THE T-CELL RECEPTOR ACTIN AND CDTHREE ZETA PHOSPHORALATION CO LOCALIZE AND YOU CAN SEE THAT EACH CELL HAS GOT SOMEWHERE BETWEEN ONE, TWO, OR THREE ADHESION POINTS SITTING IN THE SKIN AS THEY ARE DOING SO THERE. AND DISCIPLINARY MEET RIIN RIIN--DIMITRI, ADAPTED THIS TO A MORE BETTER RESOLUTION USING SKIN AND THIS IS IN THE SKIN INSITUE AND THIS CO LOCALIZES WITH THE ACTING FOCI AND YOU SEE THESE WHITE SPOTS THAT ENABLE US NOW A BETTER LEVEL OF RESOLUTION TO TRY TO UNDERSTAND THEM. BUT THE CRITICAL QUESTION FOR TODAY IS WHAT ARE THEY BINDING TOO THAT'S ON THE EPITHELIAL CELL? SO THIS IS JUST ANOTHER DEPICTION OF THIS. THIS IS EFACTIN, CDTHREE STAINING AND YOU SEE THAT COLOCALIZATION OF THE TWO VERY CLEARLY IN THIS WILD TYPE MOUSE SKIN. IF INTO THAT WILD TYPE MOUSE SKIN WE HAVE INTRODUCED SOME ANTISCINT ONE ANTIBODY, THEN WITHIN HOURS, ALTHOUGH 3D C STAINING IS PRETTY MUCH UNAFFECT SAID, IT THE REASON IT SEEMS DIFFERENT IS ITS CAPACITY TO CO LOCALIZE AND FOCAL POINTS WITH THE FACTIN IN THE CONTROLLED EXPERIMENT. DESTABILIZES THESE POINTS OF INTERACTION, AND GIVEN THAT SCINT ONE IS EXPRESSED ON THE EPITHELIAL CELL, IT SUGGESTS THAT MAY BE THE DOCKING POINT FOR THESE T-CELL RECEPTOR COMPLEXES. NOW THESE T-CELL RECEPTOR COMPLEX DOCKING POINTS ARE ALSO DESTABILIZED BY STRESS SO IF YOU EPICUE TAINIOUSLY STREET ANIMALS WITH A SMALL AMOUNT OF TPA, YOU CAN SEE THE DISCIPLINARY MINEITION OF THE FREQUENCY OF THESE FOCI. WELL, INTERESTINGLY ENOUGH, SCINT ONE UNLIKE MANY GENES THAT WE STUDY IN IMMUNOLOGY, IS ALSO DIMINISHED BY STRESS AS OPPOSE TO BEING INDUCED BY STRESS AND IN FACT IT'S PROVEN IMPOSSIBLE TO MAIP TAIN SCINT ONE EXPRESSION FOR EXAMPLE, IN KEROTIN O CITES THAT WE EXPLANT TO CULTURE. SO SINCE SCINT ONE IS APPARENTLY IMPORTANT FOR THESE INTERACTIONS BETWEEN THE T-CELL AND THE EPITHELIUM, SINCE THOSE INTERACTIONS ARE STRESS LABEL AND SINCE SCINT ONE IS STRESSED LABEL, WE CAN ASK THE QUESTION, IF WE STOPPED SCINT ONE WOULD BE --WE BE ABLE TO STOP THESE FROM BEING DIMINISHED. SO OF COURSE TO DO THIS IS TO SET UP TRANSGENIC MICE WITH PROMOTERS THAT PREVENT THEIR EXPRESSIBILITY AND OF COURSE THIS IS A REPRESENTATIVE EXAMPLE, WHAT YOU NOW FIND IS YOU CAN PUT TPA ON THESE ANIMAL AND NOW THESE CONTACT POINTS ARE MUCH LESS STRAIBL LABELED WHICH ENABLES US TO IMPROVE ON OUR MODEL OF STRESS SURVEILLANCE THAT I INTRODUCE TO YOU 15 MINUTES AGO, BY THE INTRODUCTION OF AT LEAST ONE MORE PLAYER AND THAT PLAYER IS SKINT ONE. SO THE IDEA THAT IN STEADY STATE SHAPE OR FORM, THE T-CELL COMPLEX IS REGULATED ON BY THE SKINT ONE IN ASSOCIATION OF SOME KIND. NK GTWO D OF COURSE IS NOT A LIAISON GANNA THAT EXPRESSED UNDER THE HYPOTHESIS WE WOULD EMPHASIZE THAT IT'S NORMALITY AND MAINTAINING COMPETENCE TO RESPOND RAPIDLY WHEN RESPONSES OCCUR, THEY OCCUR BECAUSE LIGANDS FOR NKGTWO D HAVE BEEN UPREGULATED AND WHETHER OR NOT IT'S FREE TO BE DISTRIBUTED THROUGHOUT THE CELL OR WELL IT ENGAGES ANOTHER LIGAND WHONS, THAT REMAINS TO BE SEEN. BUT WE COULD TEST THIS HYPOTHESIS BY SELECTIVELY REMOVING SKINT ONE FROM THE SKIN AS OPPOSE FROM THE THYMUS. IF WE REMOVE IT CONSTITTATIVELY, THE T-CELLS WILL NOT DEVELOP BUT IF WE CAN LIMIT ITS REMOVAL TO THE SKIN, WE CAN ASK WHAT HAPPENS TO THESE CELLS AND WHEN YOU DO THAT. YOU CAN SEE THAT WHAT HAPPENS AFTER ARE NOT VERY LONG PERIOD OF TIME AS THEY BEGIN IT LOSE THEIR EXPRESSION OF CD122. THEY BECOME LESS COMPETENT TO COMPETE FOR THE CRITICAL CYTOKINE MILL MILIEU ON THESE CELLS ABSOLUTELY DEPEND. SO WE THINK THIS IS A MODEL FOR HOW IT SURVEILLANCE COMPARTMENT CAN BE SUSTAINED. THE QUESTION IS: CAN WE FIND DATA FROM OTHER SYSTEMS THAT SUPPORT THIS? OR NOT. AND AS MANY PEOPLE KNOW, THERE HAS FOR MANY YEARS AND CONTINUES TO BE THIS ENIGMATIC GUT-SKIN CONNECTION AND IN OUR CASE, THE INTEREST IN THESE INTRA EPITHELIAL LYMPHOCYTES FOR THE SMALL INTESTINE. NOW AS I MENTIONED EARLIER, THOSE LARGE SCALES PRIMARILY EXPRESS THIS V-GAMMA SEVEN RECEPTOR. SO WHAT RAFACASLE DeMARCO BARS ON AND NATALIE AND SARA DID IN THE LAB FOR THE FIRST TIME WAS VISUALIZE THAT COMPARTMENT. SO AGAIN, THIS IS NOT CELLS JUST SPUN DOWN ON A CYTOSPIN. THIS IS CONFOCAL THROUGH THE EPITHELIUM AND WHAT WERE YOU AWARE SEEING HERE IS A MOUSE THAT'S ABOUT TWO AND HALF WEEKS OF AGE. AND IT HAS A FEW ALPHABETTA CELLS AND THE WHOLE THING IS PRETTY UNIMPRESSIVE. IF YOU LOOK ABOUT FOURDAYS LATER, THERE'S BEEN A COMPLETE TRANSFORMATION IN THIS V-GAMMA SEVEN COMPARTMENT HAS ARRIVED AND TAKEN OVER THE TERRITORY. WELL, WE'RE ABLE OF COURSE NOW TO LOOK AT THE TRANSITION OF GAMMA DELTA CELLS ACROSS THIS PERIOD. AND REMARKABLY ENOUGH WHEN WE DO, WE FIND OUT AND HERE'S SOME OF THE RAW DATA THAT THE GENES THAT ARE UPREGULATED ARE ASSOCIATE WIDE THIS TRANSITION ARE VERY SIMILAR TO THOSE WHICH ARE SKINT DEPENDENT DURING THE MATURATION OF SKIN T-CELL PROGENITORS. SO THIS SEEMS LIKE AN ANALOGOUS PROCESS, THE UPREGULATION OF TBET AND INNATE LIKE RESPONSIVENESS. MORE OVER, DESPITE THE FACT THAT THIS IS ALL APPEARING AT A TIME WHEN YOU WOULD THINK THE ANIMALS ARE BEGINNING TO WEAN, BLAH, BLAH, BLAH, IT OCCURS PERFECTLY FINE EXTRA THYMEICALLY, SO THESE ARE NUDE MICE THAT SHOW THESE POPULATIONS PERFECTLY EQUIV LEAPT TO THE U-THYMIC COUNTERPART SO THAT'S AN ISSUE WE WERE ABLE TO DEAL WITH. IT'S BEEN A LONG STANDING ISSUE AND IT DIDN'T REQUIRE GERMS AND IT DIDN'T REQUIRE SOLID FOOD ANTIGEN EITHER. THESE COMPARTMENTS AREENTIOUS SENTIALLY INTACT AND PROPERLY DIFFERENTIATED WITH FOR EXAMPLE, CD122 HIGH CELLS IN ALL OF THESE CIRCUMSTANCES, SO IF THIS IS NOT BEING DRIVEN BY EXOGENOUS ENVIRONMENTAL ANTIGENS, THEN MAYBE IT IS INDEED BEING DRIVEN JUST AS WE SAW IN THE SKIN BY A SELF-ENCODED MOLECULE EXPRESSED BY THE EPITHELIUM. SO, TO TEST THAT IDEA WE HAD TO CONSIDER THE FACT THAT SCINT ONE WHOSE SIMILARITY I SHOWED TO YOU PDL ONE ALREADY IS ACTUALLY PART OF A GREATER BUT VERY ENICMATIC GENE FAMILY KNOWN AS THE BEAUT ROW FILLINS OR THE BEAUT ROW FILLIN MOLECULE. NOW FOR ALL OF YOU THAT ARE EXCITED ABOUT THE PONLT OF SOMETHING BEING BEAUTERATED, DON'T BE, THESE WERE NAMED BY A CLASSICAL SCHOLAR AT CAMBRIDGE BECAUSE ONE OF THEM IS FOUND IN MILK FAT GLOBULES, WHICH MEANS THEIR BUTS ARE LOVING THAT'S WHERE THEY'RE CALLED BEAUT O FILLS, AND POSSIBLY THEY MIGHT BE RENAMED AT SOME POINT. BUT ANYWAY, SKINT ONE IS HERE, IT SITS HERE WITH MOLECULES AND IT'S GOT RELATIONSHIP TO B-SEVEN OR PDL-ONE AS I SAID BEFORE. IMPORTANTLY, SOME OF THEM AND THIS ONE IN PARTICULAR, IS EXPRESSED VERY HIGHLY IN GUT EPITHELIUM AND WHEN TO CUT A VERY LONG STORY SHORT, WE OBTAINED THREE DIFFERENT LINES OF MICE LACKING BUTROPHYLI N, THESE MICE ARE ALMOST OBLAITED FROM THE GAMMA POSITIVE CELLS BUTENTIOUS SENTIALLY EVERY OTHER T-CELL TYPE AND THERE PERFECTLY NORMALLY. SO THIS IS VERY MUCH LIKE WE SAW FOR SKINT, AND IT SUGGESTS THAT THE EPITHELIA BEYOND TERRA TIN O --KERATINOCYTES USE BUTYROPHILIN TO MOVE THESE PARTS. SOPHISTICATEDY WE DESCRIBED THAT THAT YOU COULD DO THAT TO COMPENSATE FOR THE LOSS OF FUNCTION AND IF WE INTRODUCE SKINT ONE AND DRIVE IT FROM A GUT PROMOTER WE CAN RESCUE THE DWIF RENTIATION SO THIS IS ONE OF THESE PLOLTS WHERE MOLDY PARAMETER PHENOTYPES ARE COMPARED HERE OF WILD TYPE MICE AND MICE RESCUED VERSUS THE MUTANT MICE AND VARIOUS CONTROL STRAINS. SO SWITCHING BTNL-ONE BACK ON, RESCUES THE PHENOTYPE AND AS LONG AS YOU DO IT DURING THE DEVELOPMENTAL WINTER WEATHER WINDOW I DESCRIBED IT WILL ALSO RESCUE A CELL NUMBER. SO THAT TO US WAS A PRETTY REASONABLE DEMONSTRATION THAT THESE CELLS DEPENDED ON BEAUT O FILLIN LIKE ONE AND BTNL ONE COULD RESCUE THE DEFICIENCY. AND IT WAS THEREFORE OF SOME CONCERN TO US THAT WE COULD NEVER MIMIC THIS INVITRO. SO WE COULD NEVER TAKE AN EPITHELIAL CELL AND INTRODUCE INTO IT BTNL ONE AND GET IT TO THE EFFECT A MEANINGFUL INTERACTION IN INTEREPITHELLIAL LYMPHOCYTES. SO WE WENT BACK TO THIS PLOT AND WE REALIZE THAT THERE'S ANOTHER MOLECULE THAT'S ALSO EXPRESSED STRONGLY IN THE GUT. IT'S BTNL SIX, YOU TAKE MODE K AND YOU INTRODUCE INTO IT AN EMPTY VECTOR, BTNL, BTNL SIX OR THE COMBINATION OF THE TWO, AND YOU CO CULTURE EITHER WITH ALPHABETTA CELLS OR V-GAMMA SEVEN DELTA NEGATIVE CELLINGS OR GAMMA POSITIVE CELLS AND YOU SEE A HALLMARK RESPONSE WHICH WE WILL SEE AGAIN SHORTLY WHERE CD25 IS UPREGULATED AND THE TCR SHOWS DOWN REGULATION, NOT SHOWN HERE, CDONE - 22 IS ALSO REGULATED. YOU COULD MIMIC THIS USING A NEURO77 GFP REPORTER WHICH IS INDICATIVE OF AN NFACT RESPONSE. SO THIS REQUIRED BTNL ONE AND SIX, GIVEN THAT WE CAN NOW GENERATE BTNL SIX DEFICIENT MICE, WE'VE BEEN ABLE TO SHOW THAT BTNL SIX IS ALSO REQUIRED FOR THE DEVELOPMENT OF THESE CELLS. SO NOW HAVE YOU A DIMER ACTING AND HERE WE HAD A SINGLE MOLECULE ACTING BUT SKINT ONE IS A FOUNDER MEMBER OF A FAMILY THAT INCLUDES SKINT TWO, AND RECENTLY, YVETTE YANKA HAS DEVELOPED TWO MICE, AND HERE YOU CAN SEE THEM WITH THE WILD TYPE CELLS AND THE SKINT TWO MOUSE, NOW PHENOCOPIES THE ONE MICE IN BEING VERY SPECIFICALLY DEFICIENT IN THE V-GAMMA FIVE SUBSET, EVEN THOUGH IT HAS PLENTY OF GAMMA DELTA CELLS IN THE SKIN THAT JUST THE WRONG TYPE AND YOU CAN ENUMERATE THIS VERY DRAMATIC MUTANT PHENOTYPE BY DOING CELL COUNTING. SO HOW YEE HAVE INTEREPITHELLIAL COMPARTMENTS BY TIMERS OF THESE BTNL MOLECULES. AND I INDICATED BEFORE THAT WE CAN IDENTIFY CELLS LIKE THIS FROM THE HUMAN COLON AS INDICATED HERE. AND HERE ARE TWO MOLECULES, BTNL THREE AND EIGHT THAT ARE HIGHLY EXPRESSED HUMAN SMALL INTESTINAL EPITHELIUM SO WE DO A SIMILAR CO-CULTURE EXPERIMENT AND NOW WHAT YOU CAN SEE IS IF YOU--IF YOU LOOK AT ALPHALPHABETTA T-CELLS IF YOU LOOK AT BLOOD GAMMA DEALT AT-CELLS OR IF YOU LOOK AT GAMMA DELTA T-CELLS FROM THE HUMAN GUT, THESE CELLS WHICH ARE PREDOMINANTLY EXPRESSING V-GAMMA FOUR SELECTIVELY SHOW TCR DOWN REGULATION WHEN CO-CULTURED WITH CELLS THAT EXPRESS HUMAN BTNL THREE AND BTNL EIGHT. AND SO NOW WE'RE ABLE TO POPULATE THIS GENE FAMILY WITH YET ANOTHER SEEMINGLY DISCIPLINARY MERRIC FUNCTIONAL CAPE ANLT AND THAT'S PROVOCATIVE BECAUSE MANY LABS NOT SO MUCH OUR OWN HAVE BEEN WORKING ON HUMAN PERIPHERAL BLOOD FOR MANY YEARS WHERE THE CELLS MAKE RESPONSES TO WHAT ARE KNOWN AS LOW MOLECULAR WEIGHT FOSTER NURSED FOCUSED ON ANTIGENS AND SHOWN THAT THOSE RESPONSES DEPEND ON A MOLECULE CALLED BTNL THREE A ONE. SO THE PEER IN OUR GROUP FOUND IT HARD TO BELIEVE THAT THIS MOLECULE ACTS IN ISOLATION AND UNDERTOOK SOME BIY CHEMISTRY EXPERIMENTS WHERE WE CAN INTRODUCE FLAG TAG THREEA ONE, AND A MOLECULE THREEA TWO, AND TAGGED WITH HA AND HE CAN SHOW IN THESE HAD IMMUNE O PRECIPITATES FOR A-ONE, HOO CAN DETECT A-TWO IN THOSE PRECIPITTORS SUGGESTING THERE IS A TIMER AND WHEN HE MAKES CELLS THAT ARE GOING TO RESENT FOZ FOCUSED ON ANTIGEN TO THE GAMMA DELTA CELLS AND REMOVES DTNL-THREE AND ONE, HE GETSA I MAJOR APLAIGZ OF FUNCTION WHEN HE REMOVES BTN-THREE-A-TWO. SO THIS IS THE COLOCALIZATION WORKED OUT WITH AN ER MARKER, PDI AND IF YOU EXPRESS THE MOLECULES ON THEIR OWN, THEY STRONGLY CO LOCALIZE WITH PDI, BUT IF YOU ALLOW THEM TO DIMERIZE, THEY DON'T BECAUSE THEN THESE MOLECULES BEGIN TO GET TO THE SURFACE. SO BELIEVE THAT MAY BE A GENERAL RULE FOR HOW THESE DIMERS ARE REGULATING EACH OTHER'S CELLULAR TRAFFICKING. JUST IN THE WAY THAT WE THINK OF THE COMPLICATED CELLULAR TRAFFICKING AND MOLECULES LIKE CD1 OR MHC. AND YOU COULD BEGIN TO SEE THAT THIS IS NOW PROVING TO BE QUITE A HIGHLY POPULATED PROPERTY OF THIS FAMILY, WHICH NOW GIVES US A NEW PERSPECTIVE FOR CONSIDERING FOR EXAMPLE, THE STRONG IMPLICATIONS OF MANY OF THESE GENES BOTH BY GWAS ANALYSIS AND TRANSCRIPT OHMICS IN HUMAN DISEASE, A POINT WHICH I'LL CLOSE WITH SHORTLY. SO I REALLY NOW JUST WANT TO CONSIDER THE ISSUES THEY THINK THIS RAISES. SO HOPEFULLY I'VE BEEN ABLE TO CONVINCE YOU TODAY THAT THERE ARE SETS OF GAMMA DELTA T-CELLS BOTH IN MICE AND IN HUMANS THAT ARE CONSTITTIVELY AND INTIMATELY ASSOCIATED WITH EPITHELIAL CELLS IN VARIOUS TISSUES. SO THE QUESTION IS OF COURSE ARE THESE SELECTING LIGANDS FOR THE GAMMA DELTA T-CELL RECEPTOR. IF THEY ARE, IS THAT THE ONLY LIGAND A GAMMA DELTA T-CELL HAS AND IT HAS A LIGAND AND IF SO HOW ARE THOSE DIFFERENT IN THIS SUPPORT OF THE IDEA THAT THE T-CELL RECEPTOR IS ESSENTIALLY INVOLVED, WE CAN PHENOCOPY THE LACK OF SKINT 1 AND ORGAN CULTURE BY TCR AGANIST SO THEY WILL NOW ACQUIRE THE DEVELOPMENTAL MATURATION THAT NORMALLY DEPENDS ON THIS PATHWAY. I'VE INDICATED TO YOU THAT THE SIGNATURE RESPONSES ARE LIMITED TO THOSE WITH THE SIGNATURE TISSUE ASSOCIATED TCR ISOTYPES. THE RESPONSES THAT YOU GET DOWN REGULATION OF THE TCR UPREGULATION OF CD25, ARE SIGNATURE RESPONSES. AND IT'S POSSIBLE THAT THE INTRA CELLULAR TRAFFICKING AND THE SOPHISTICATION OF THAT TAKEN--THEY WE'RE BEGINNING TO UNCOVER REFLECTS A KEY DWELL TIME THAT THESE MOLECULES REQUIRE IN THE ER. SO FOR EXAMPLE, IF WE PUT THE EXTRA DOMAINS OF BTNL1 AND 6, AND EXPOSE THEM ON THE CELL SURFACE LIKE THAT, THEY DON'T FUNCTION AT ALL. SO THIS IS MORE THAN SIMPLY SURFACE DISPLAY. INDEED HERE IS AN IMPROVEMENTOT FUNCTIONAL ASSAY, DEVELOPED BY GRADUATE STUDENT, YOU CAN NOW BEGIN TO SEE HOW WELL SHE HAS THIS WORKING FOR CD25 UPREGULATION AND TCR DOWN REGULATION. AND THERE ENABLING HER TO PULL OUT THE SEQUENCES OF THE SINGLE CELL LEVEL OF THE CELLS OF THE RESPONDING AND THEY'RE ALL VGAM 7 POS 55 AND THEY ALL HAVE A VERY LIMITED CDR3 ALTHOUGH THAT'S THE NATURE OF MOUSE GAMMA CHAINS ANYWAY. BUT THE DELTA CAN BE QUITE VARIABLE IN THE GENE SEGMENT THAT'S USED AND IN THE CDR3 LINK. SO WE DO INDEED BELIEVE THAT THIS ACTIVITY CORRELATES EXACTLY WITH THIS HIGHLY ORGAN SPECIFIC V-GAMMA CHAIN USAGE. BUT IT DOESN'T PROVE BINDING EXPH IT IS POSSIBLE THAT FOR EXAMPLE, THE BEAUT ROUGH ATOMPHILIANS ARE ENGAGING IN CELL TYPE SPECIFIC KORESEPTORSORS, THAT SAME DAY SIGNAL THROUGH PI-3 KINASE WHICH IS SUFFICIENT TOASM RIIN--DIMITRIIFY A TCR SIGNAL, THAT FOR ALL WE KNOW COULD BE COMING FOR LIGAND MONOPOLIED MULTIPLE RISATION WHICH SOME RECEPTORS AND PRET-CELL RECEPTORS ARE CAPABLE OF AND THAT WOULD OF COURSE FIT WITH THE CLOSE SIMILARITY TO PDL 1 BUT THESE ARE THE ISSUINGS THAT WE HAVE TO SELL. BUT THE FINAL ISSUE I WANT TO LEAVE YOU YOU WITH IS GETTING BACK TO THIS, BECAUSE THERE'S A LOT OF WORDS PEOPLE TALK ABOUT BUT THEY'RE NEVER CLEAR WHAT THEY MEAN. PEOPLE TALK ABOUT STRESS, WHAT DO YOU MEAN BY STRESS? I'M TALKING ABOUT NORMALITY. WHAT DO WE MEAN BY NORMALITY? SO HOW DOES A CELL KNOW THAT IT'S NORMAL AND IT DOESN'T NEED TO ACTIVATE THE LOCAL IMMUNE RESPONSE THAT EVERYTHING THIS BE LEFT ALONE TO AVOID CHRONIC INFLAMMATION. WELL WE BELIEVE WE HAVE THE MOLECULES THAT ARE REQUIRED TO ANSWER THAT QUESTION BECAUSE THESE ARE SO KEY ON MAINTAINING THESE CELLS AT NORMALITY. THAT IF WE COULD UNDERSTAND HOW THESE ARE REGULATED, THEY COULD INDICATE TO US THE MOLECULAR LEXICON FOR NORMALITY. SO PIERRE VONTIVOU, HAS BEEN TRYING TO WORK OUT THE REGULATION OF THESE AND HAS BEEN ABLE TO FIND A MOLECULE THAT MAY RELATE TO THE METABOLIC STATE OF THE GUT THAT REGULATES THE EXPRESSION OF THESE GENES AT STEADY STATE. SO HERE'S AN EXAMPLE, SO THESE ARE K-CODE 2 CELLS THAT MANY OF YOU MAY, WITH THESE NEW HUMAN GASTROINTESTINAL CELLS WHEN THEY'RE GROWING FAST, BTNL 3, IS EXPRESSED ALMOST NOT AT ALL. WHEN THE CELLS ARE CONFLUENT IT BECOMES TO COME UP WHEN THEY'RE IN A POLARIZED DIFFERENTIATED MONOLAYER THEY'RE EXPRESSED VERY HIGHLY IN DICKAATIVE OF THE SYNONYMOUS WITH THE DIFFERENTIATED STATE AND THE TRANSCRIPTION FACTOR THAT'S RESPONSIBLE FOR THAT IS HNF 4 ALPHA WHICH INTERESTINGLY ENOUGH WHEN IT WAS CRYSTALLIZED WAS FOUND DEPENDENT ON THE ACTIVITY ON LIPID. AND THE GASTROENTEROLOGY FELLOW IN THE LAB HAS SHOWN IS NOW YOU CAN MASSIVELY SYNERGIZE THE HNF 4A ON THE BEAUT O FILLIN LIKE 3, BY THE SUPPLEMENTATION OF OLAYIC ACID OR LYNN O LAIC ACID SUGGESTING A HYPOTHESIS AT LEAST THAT THE EPITHELIAL CELL IN THE GUT KNOWS EVERYTHING IS OKAY BY MEASURING THE CONCENTRATION OF DIETARY LIPIDS OR INTERMEDIATES IN THAT PATHWAY THAT IT'S ACCESSIBLE TO AND DISPLAYING AS A RESULT OF THAT MEASURE 1 OF THESE MOLECULES WHICH HOLDS THESE CELLS IN A COMPETENT BUT RESTING STATE. SO 9ALLY TO GET BACK TO THE PROSPECT OF SURVEILLANCE THAT WE OBSERVED BEFORE, IS POSSIBLE THAT THESE MOLECULES MIGHT BE MISSING WHEN CANCER IS GETTING OUT OF CONTROL. SO THIS IS ONKA MINE WHICH IS--MANY OF YOU MAY HAVE USED IT, IT COAALATES ISSUES IT'S LIKE A METADATABASE, WHICH COAALATES I A NUMBER OF STUDIES. AND IT SHOWS THE NUMBER OF STUDIES THAT IMPLY THE CHANGE IN TRANSCRIPTIONAL LEVEL IN PARTICULAR GENE. SO HERE'S BTNL 3 AND 8 AND IT'S INTERESTING THAT THERE'S 1 PLACE THAT THEY FIGURE WHICH IS COLO RECTAL CANCER WHERE THERE SEEMS TO BE REAL CONSISTENCY IN THE DOWN REGULATION OF THOSE. AND WHAT IS INTERESTING IS IF YOU NOW LOOK UP HERE AT BREAST THERE'S 1 STRIKING MOLECULE AGAIN WHICH IS BTNLIKE 9 AND NOW WE'RE NOW LOOKING AT THE POSSIBILITY THAT BTNL 9 IS A REGULATOR IN HUMANS AND IN MICE OF BREAST ASSOCIATED GAMMA DELTA CELLS WHICH I SHOWED VERY BRIEFLY ON A PICTURE MUCH EARLIER AND THIS IS WORK OF FERN NANDA KYLE. SO TO FINISH I JUST WANT TO CONCLUDE THAT WE BELIEVE IN THE MOUSE AND SUSPECT IT'S A MODEL FOR HUMANS IN MANY RESPECTS THAT TISSUES DO IN FACT HAVE AN IMMUNE SURVEILLANCE COMPARTMENT. THE DATA RELATING THAT TO CANCER WILL COME FROM OTHER LABS ON THEIR OWN IN TERMS OF HUMAN DISEASE EPIDEMIOLOGY, BUT MECHANISTICALLY, THIS IS AN ORGAN SPECIFIC SURVEILLANCE COMPARTMENT. THE LOCAL GAMMA DEMENTIA T-CELLS SEEM STRIKINGLY EVOCATIVE OF WHAT WE SEE IN THE LAMP RAY. THEIR COMPOSITION FUNCTIONAL RESPONSIVENESS IS DRIVEN ENTIRELY BY THEIR INTERACTIONS WITH THE LOCAL EPITHELIUM, AND THOSE EPITHELIUM WE NOW KNOW ARE USING ORGANIZATIONS SPECIFIC MOLECULES FROM THE BTNL-LIKE FAMILY. SUCH GENES MAY BE MORE GENERALLY USE INDEED REGULATING GAMMA DELTA CELLS AND IN THAT REGARD IT PROVIDES A PERSPECTIVE OF THE IMPLICATIONS BY GWAS ET CETERA IN DISEASE. OF COURSE I DO NOT MEAN FOR 1 MINUTE THESE TO SUGGEST THESE ARE THE MOLECULES AND IF YOU LOOK AT DOWN REGULATION TCR PLOTS HERE'S A NICE REGULATION OF THE DOWN REGULATING THE TCR AND LOOKING AT THE CELLS HERE IN THE GUT, VDELTA 1 POSITIVE THAT HAVEN'T SHIFTED WHETHER THERE'S ANOTHER MOLECULE THAT WOULD DRIVE THOSE, ET CETERA WE DON'T KNOW BUT IT GIVES US FURTHER WAYS TO EXPLORE THE EPITHELIAL DECISION MAKERS THAT ARE DRIVING THE COMPOSITION AND FUNCTIONAL RESPONSIVENESS OF THESE COMPARTMENTS. SO WITH THAT I WILL THANK THE MANY PEOPLE INVOLVED INCLUDING A NUMBER OF COLLABORATORS. THESE ARE PEOPLE WHO WERE IN THE LAB, MADE CONTRIBUTIONS, THESE ARE THE PEOPLE IN THE LAB, I TRIED TO MAIM THEM AS WE WENT THROUGH AND I WOULD LIKE TO THANK YOU FOR YOUR ATTENTION. [ APPLAUSE ] >> THANK YOU ADRIAN. >> THANK YOU. >> I WOULD LIKE TO OFFER A SUGGESTION TO INTERPRET SOME OF THE DATA IF YOU THINK ABOUT THE SKINT 1 PART IF YOU GO OUT THROUGH THE PERIPH RADIOY AND YOU DON'T GET SIGNALING, THAT'S WHAT OTHERS AND MYSELF HAVE REPORLTED FOR THE BETA RECEPTOR WHICH IS PARTIAL ZETA PHOSPHORALATION AND CENTRALIZATION OF THE CELLS BUT NOT SIGNALING. WHEN YOU STARTED TO TALK ABOUT THE GUT BECAUSE HAVE YOU REAL SIGNALING THERE, YOU POINTED OUT YOU DON'T NEED THE THYMUS IN WHICH THEY ARE TUNING THEMSELVES TO THAT LEVEL AND THAT FITS PRECISELY WELL SO IT APPEARS TO ME YOU'RE DESCRIBING A SYSTEM OF SELECTING LIGANDS IN WHICH THE MATURATION OF THE CELLS EXTINGUISH THEIR AGANIST PROPERTY JUOF THE AS HAPPENS DURING THYMIC SELECTION. >> IT'S DIFFERENT. >> IT'S NOT DIFFERENT THEY'RE LIKE THE DOUBLE POSITIVE WHICH GIVE PRODUCTIVE SIGNALING AND THEN THEY GO INTO THE QUIESCENT STATE IN WHICH THEY'RE GETTING THE SUBTHRESHOLD STIMULATION SO IS THERE ANYTHING I AM MISSING FROM THAT KIND OF SCENARIO. >> NO, NO. I THINK IT'S PERFECTLY POSSIBLE, YOU KNOW, I MOON THE DIFFICULTY IS WE KNOW IS WE'RE ALL CONSTRAINTED BY LYNNIAN CLASSIFICATION, THAT HELPS SORTS OUR LIVES OUT AND SO GAMMA DELTA T-CELLS HAVE SPEND THIS EXISTENCE SITTING BETWEEN THE INNATE IMMUNITY BUT IF WE THINK ABOUT ORCHESTRATING IMMUNE O GENERATEDISSITY, WHOO YOU DESCRIBE FOR EXAMPLE, COULD CERTAINLY HELP US UNDERSTAND WHY IMMUNE O GENERATEDISSITY IS SO DIFFERENT IN THESE TISSUES WHEN YOU CHALLENGE WITH THE SAME ANTIJERN AND THE READ OUT IS ADAPTIVE ALPHABETTA T-CELL RESPONSES BUT THE ONLY THING I WANT TO CAUTION IS, I'LL NOT TREADING ON THE TOINGS OF ANY MOMENT WHO OF THE CROWD WHO BELIEVES THAT'S THE TOTAL ORCHESTERATION THAT'S REQUIRED BUT THE KIND OF DISTINCTIONS YOU CAN MAKE CAN ACTUALLY CORRELATE WITH THAT. >> THANK YOU. >> HELLO ADRIAN. SO WHAT DO YOU THINK THAT THE V-GAMMAS HAVE IN CELLS ARE SEEING IN THE THYMUS, I MEAN THEY'RE BEING SELECTED, RIGHT? >> NOT IN THE THYMUS, THOSE DON'T REQUIRE THE THYMUS, THE 1S IN THE SKIN DO. >> THE OTHERS IN THE SKIN, BUT SOME CELLS ARE GENERATED IN THE THYMUS RIGHT? >> YEAH. >> THEY SHOULD BE A LIGAND? >> SO THIS AGAIN WITHOUT MEANING TO HEDGE GETS VERY DIFFICULT. SO 2011 WAS A BIG YEAR NEAR A LOLT OF PEOPLE, I'M SURE BUT 1 OF THE REASONS IT WAS A BIG YEAR, IS YOU KNOW MONOCITE MACROPHAGE GOT TURNED ON ITS HAD WITH THE NOTION THAT SOME OF THESE TISSUE RESIDENT COMPARTMENTS ARE LAID DOWN IN THE YOLK SACK AND ARE INFALLIBLE THEREAFTER AND THERE IS A LOT OF THAT HERE, TOO, SO IT'S KIND OF--I MEAN, THAT'S ANOTHER COMPONENT TO RON'S QUESTION, WHICH IS--WHICH IS ONTOGNY IN THE SENSZ THAT THE SKIN CELLS ONLY NEED THE FETAL THYMUS AND THE GUT CELLS ARE COMPLETED IN MATURATION AFTER THAT PHASE. SO THE DIFFERENCE BETWEEN THE 2 IS NOT SIMPLY THE THYMUS, IT'S ALSO THE TIME AT WHICH THE MATURATION OCCURS. >> SO WE KNOW, WE ARE CONSCIENCE OF THE FACT THAT 1 DAY WE NEED TO GET UP HERE AND SHOW SOME BIOCHEMISTRY AND SAY THIS DOES OR DOES NOT INTERACT. BUT WHAT I WOULD SAY IN NIGH DEFENSE IS THAT A LOT OF PEOPLE WOULD DO THAT BIOCHEMISTRY WITHOUT EVEN REALIZING THESE THINGS EXIST AS DIMERS. AND THAT'S PROBABLY WHY THEY WEREN'T SUCCESSFUL AND NOW THAT WE'RE LEARNING MORE ABOUT THEM, WE WILL TRY TO FIND OUT AND ANSWER YOUR QUESTION ABOUT WHAT THE LIGAND SURFACE INTERFACE LOOKS LIKE BECAUSE THAT CAN THEN HELP US UNDERSTAND WHAT OTHER LIGANDS, SELECTING LIGANDS OR ACTIVATING LIGANDS MIGHT LOOK LIKE. >> JUST ANOTHER QUICK QUESTION, IF YOU WERE TO EXPRESS BTNL1 OR 6 IN THE SKIN WOULD YOU SEE THOSE V-GAMMA 7-- >> YEAH. SO WE HAVE A PLANT TO DO--PLAN TO DO THAT. NOW THAT WE KNOW WHAT'S REQUIRED WE CAN DO THAT IF YOU JUST EXPRESS SKINT 1 IN THE GUT, YOU DON'T GET THEM IN THE GUT BUT YOU NEED SKINT 2 THE. BUT NOW WE DO THAT. IT'S CLASSIC DEVELOPMENTAL BIOLOGY, IF THESE ARE THE REQUIREMENTS YOU SHOULD GET AN ECTOPIC COMPARTMENT AND WE HAVE EVERY INTENTION OF DOING THAT. >> SO TO GO BACK TO THE TIEN TIMING DEVELOPMENT QUESTION, THE GAMMA DELTA CELLS IN THE GUT. YOU SEE THERE IS A THYMUS, DO YOU SEE EVIDENCE IN THE GUT FOR ANTIGEN ARRANGEMENT WITH THE SAME KINETICS? >> SO IT'S A GOOD QUESTION. OF COURSE, SO WE'VE LOOKED QUITE HARD AT THAT. THE ONLY DIFFERENCE WE SEE IS THERE ARE MORE CELLS IN THE ANIMAL THAT'S UTHYMIC BUT IT'S DIFFICULT OF ALL THE OTHER EVIDENCE. >> DO YOU SEE WITH THE PARABOLA, DO YOU SEE THE [INDISCERNIBLE] IF THE GUT? THYMUS? >> SO THIS IS REVISITING THING SOME EXPERIMENTS THAT WERE DONE MANY YEARS AGO, WHICH STIMULATED IMMENSE DEBATE AND CONTROVERSY AND YEAH, SO WE HAVE NOT GONE BACK TO TO DOING THAT AND THANKFULLY OUR REFREES DIDN'T DEMAND WE DID THAT BUT FOR THOSE WHO DID KNOW, FAMILY, FRIENDSHIPS WERE SPLIT UP OVER THE DISCORDANT RESULTS THAT WERE FOR EXAMPLE ACHIEVED BY THE GROUP IN SWEDEN WHICH CLAIMED THERE WAS AN ALTERNATIVE SPLICE FORM OF REG THAT WAS PRESENT IN THE GUT. SO, YEAH WE'RE STICKING WITH THE SELECTION. >> OKAY. SO 2 VERY QUICK QUESTIONS, PERHAPS I MISSED IT BUT THE BTNL-DEFICIENT MICE DO THEY HAVE INCREASED FREQUENCY? >> IT'S METHANE LOW LEVEL DSS PROMOTION SIMPLY BECAUSE IT'S THE BEST MIRROR OF THE BNBATPA RESPONSE. >> AND TO GO BACK TO YOUR VERY UNUSUAL SLIDES THE LAMP RAYS DO THEY THINK IT'S CURRENT OF [INDISCERNIBLE]? >> YEAH, SO BTNL ARE LIMITED TO MAMMALS, BLUE IF YOU LOOK IN CHICKENS FOR EXAMPLE, THERE ARE THENGS CALL THE B-GENES WHICH HAVE BEEN STUDY SAID PRIMARILY OF LATE BY JIM KAUFMAN'S GROUP AT CAMBRIDGE AND THOSE--THEY'RE VERY, VERY SIMILAR AND THOSE LYMPHOCYTE REGULATION AND YES, ABSOLUTELY. SO WE'RE WORKING WITH THIS TO LOOK THROUGH THE LAMP RAY GENOME TO SEE WHAT WE CAN FIND. >> YOUR CARTOON OF SPRINT 1 SUGGESTED IT HAS A 3 TRANSMEMBRANE DOMAINS THAT'S AN UNUSUAL SIGNATURE. DO ANY OF THE OTHER BTNLs HAVE THAT? AND DOES 2 HAVE THAT AS WELL? INVOLVED IN SELECTIVE DIMERIZATION, ET CETERA? >> SO, YEAH, CD47 AS 3 TRANSPLANT DOMAINS WITH A SINGLE IMMUNE O DPLOBUE LYNN DOMAIN ON THE OUTSIDE. WITHIN THE SKINT FAMILY, SEVERAL OF THEM, ALL OF THE SKINT FAMILY HAD ALL TRANSMEMORY DOMAINS, IT'S EITHER 3 OR 5. AND OF COURSE, THE POTENTIAL FOR THOSE WHEN YOU HAVE DIMERS OF THESE MOLECULES, YOU KNOW YOU WILL BRING ENORMOUS PERTERBANCE TO THE--TO THE LIPID BI-LAYER BY FITTING ALL OF THESE TRANSMEMBRANE, SO WE'RE VERY INTRIGUED ABOUT THE ROLE OF THOSE. BUT THEY'RE THE ONLY MEMBERS WITHIN THE BEAUT ROUGH ATOM FILLIN FAMILY THAT HAVE THE SPECIAL ADAPTATION AND WHETHER IT RELATES TO SOMETHING THAT THEY'RE SENSING, SO, YOU KNOW JUST TO BE SORT OF STUPIDLY SPECULATIVE BUT IMAGINE THAT NORMALITY IN THE SKIN WAS MEASURED BY MEMBRANE PRESSURE. YOU KNOW THE SORT OF DIMES THAT THE MEMBRANE IS SENSING. MAY BE THESE MULTIPLE TRANSMEMBRANE DOMAINS THAT ARE SENSOR FOR THAT. WHO KNOWS. SO YEAH, IT'S INTRIGUING? >> SINCE IT'S AFTER 5:15. WHY DON'T PEOPLE JUST COME DOWN WITH MORE QUESTIONS. THANK YOU. >> THANK YOU.