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LET'S GET STARTED.

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I AM SO DELIGHTED TODAY TO

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INTRODUCE TODAY'S SPEAKER SIR

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STEVEN JACKSON.

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SO STEVE, STEVE IS THE

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UNIVERSITY OF CAMBRIDGE

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PROFESSOR OF QUICK BIOLOGY AND

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SENIOR GROUP LEADER AT THE

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CANCER UK INSTITUTE AT

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CAMBRIDGE.

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HIS RESEARCH IDENTIFIED KEY

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PRINCIPLES BY WHICH CANCER CELLS

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RESPOND TO AND REPAIR DNA DAMAGE

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AND HOW THEY YIELD CANCER AND

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OTHER AGE RELATED DISEASES.

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IN ADDITION TO THE VARIOUS

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COMPANIES THAT HE HAS FOUNDED TO

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TAKE SOME OF HIS RESEARCH

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FINDINGS TO THE CLINIC, STEVE IS

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A FELLOW OF THE ROYAL SOCIETY,

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UK ACADEMY OF MEDICAL SCIENCES,

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THE EUROPEAN ACADEMY OF CANCER

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SCIENCES AND [INDISCERNIBLE].

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HE HAS RECEIVED NUMEROUS

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NATIONAL AND INTERNATIONAL

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PRIZES AND IN 2023 WAS AWARDED A

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KNIGHT HOOD FOR HIS SERVICES TO

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INNOVATION AND RESEARCH.

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I AM VERY EXCITED TO HEAR WHAT

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HE HAS TO SAY TODAY BUT BEFORE

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WE START, I REMIND YOU ALL TO

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PUT YOUR QUESTIONS IN THE CHAT

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BOX AND WE WILL READ THEM OUT

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AFTER HE COMPLETED HIS

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PRESENTATION.

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STEVE, IF YOU CAN START WHEN

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YOU'RE READY?

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>> GREAT.

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CAN YOU HEAR ME?

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>> WE CAN.

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>> GREAT.

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I WOULD LIKE TO START BY

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THANKING KAREN, CHUN ZHANG, AND

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COLLEAGUES FOR INVITING ME TO

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THIS GROUP TODAY.

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I WOULD LIKE TO THANK THE NIH

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PER FUTURE--POTENTIALING THESE

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SEMESTER NEARS ON, IT WAS

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CRUCIAL DURING COVID TIMES BUT

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IT STILL PLAYED AN AMAZING

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IMPORTANT ROLE IN THE DNA REPAIR

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FIELD AND IS IMPACTING WORLD

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WIDE SO THANKS AGAIN FOR THAT.

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SO I KNOW I WILL BE PREACHING TO

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THE CONVERTED HERE BUT TO START

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WITH, I'M JUST GOING TO GIVE A

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BRIEF SUMMARY OF CELLULAR

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RESPONSES TO DNA DAMAGE AND DNA

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REPAIR.

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POINT OUT IT'S MEDICAL

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IMPORTANCE AND CLINICAL

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POTENTIAL AND THEN I WILL ANY ON

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TO TALK ABOUT SOME RECENT WORK

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IN MY LAB WHICH PROVIDED NEW

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INSIGHTS INTO THESE PATHWAYS,

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AND ALSO PROVIDE FURTHER LINKS

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TO HUMAN DISEASE.

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SO WE'RE MORE AWARE OF THE FACT

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THAT DNA IN OURSELVES AND EVERY

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ORGANISMS' CELLS IS CONTINUALLY

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BEING BOMBARDED BY A RANGE OF

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[INDISCERNIBLE] AS WELL AS

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EXOGENOUS DNA DAMAGING AGENTS

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GIVING RISE TO A WIDE RANGE OF

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DNA DAMAGE THAT NEEDS TO BE

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DEALT WITH BY THE DNA REPAIR

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SYSTEMS AND THE SYSTEMS.

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APOLOGIES I HAVE A BIT OF A COLD

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TODAY BUT HOPEFULLY MY VOICE

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WILL HANG TOGETHER.

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WHAT I WILL FOCUS ON TODAY IS A

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BIT UNUSUAL FOR ME, I USUALLY

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TALK ON DOUBLE STRAND BREAK

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MECHANISMS BUT I WILL TALK TODAY

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ABOUT DNA CROSS LINKS.

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THESE ARE VERY, VERY IMPORTANT

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LESIONS IN EMERGING ITS OF

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BIOLOGICAL AND MEDICAL

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IMPLICATIONS, OF COURSE DNA

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CROSS LINK WILL BE A SEVERE

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IMPEDIMENT TO PROCESS SUCH AS

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TRANSCRIPTION AND REPLICATION

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EMPLOY NOW WHEN WE THINK ABOUT

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DNA REPAIR, WE CLEARLY WANT TO

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KNOW HOW THE DNA REPAIR PROTEIN

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COMPLEXES WORK BUT WE NEED TO

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THINK ABOUT DNA REPAIR OR ALSO

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THINK ABOUT DNA REPAIR IN THE

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CONTEXT OF A BROADER DNA DAMAGE

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RESPONSE AS IS OFTEN TERMED

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WHICH LEADS US TO CONSIDER DNA

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REPAIR AND THE CONTEXT BRINGS UP

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CHROMATIN STRUCTURE, DNA

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REPLICATION AND HOW IT'S

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IMPACTED BY VARIOUS OTHER THINGS

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INCLUDING NUCLEAR ARCHITECTURE

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AND DNA METABOLISM AND OTHER

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EVENTS.

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WE ALSO NEED TO THINK ABOUT THE

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SIGNALING PATHWAYS THAT ARE

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TRIGGERED IN RESPONSE TO VARIOUS

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FORMS OF DNA DAMAGE, AND HOW

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THAT CAN LEAD TO INDUCTION OF

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REPAIR FACTORS, TRANSCRIPTIONAL

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PROGRAMS, CELL CYCLE, CONTROLS

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INCLUDING CELL CYCLE CHECK

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POINTS, AS WELL IN OTHER CASES

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TRIGGERS APOPTOSIS OR

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SENESCENCE.

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AND WE ALSO NEED TO THINK ABOUT

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HOW PROTEINS ARE INVOLVED IN

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OTHER PROCESSES SUCH AS TELOMERE

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MAINTENANCE UNDER

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NORMAL/ABNORMAL CIRCUMSTANCES.

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OVER THE YEARS WE LEARNED THAT

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THESE RESPONSES ARE GOVERNED BY

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THE ACTIONS OF A WIDE VARIETY OF

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REVERSIBLE POST TRANSLATION

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MODELS INCLUDE, UBIQUITINNATION,

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POLYSATURATION, AND

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HBB -ELATION, I THINK WHERE WE

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STAND HOW THE DNA FIELD IS QUITE

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A MATURE 1 AND WE KNOW NOW

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THERE'S OVER 1000 HUMAN PROTEINS

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WITH STRONG DNA DAMAGE RESPONSE

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AND DNA REPAIR LINKS BUT MANY OF

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THESE ARE STILL LARGELY

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UNCHARACTERRIZED.

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WE ALSO KNOW THAT DNA,

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UNDOUBTEDLY OTHER DDR PROTEINS

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AND REGULATORS TO BE IDENTIFIED

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AND I THINK THE BIGGEST

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CHALLENGE AND OPPORTUNITY FOR US

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RIGHT NOW MOVING FORWARD IS TO

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THINK ABOUT THIS IN TERMS OF THE

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FUNCTIONAL INTERACTION LANDSCAPE

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WHICH IS LARGELY UNEXPLORED AND

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WHAT I MEAN BY THAT IS NOT

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THINKING ABOUT INDIVIDUAL

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COMOPPOSITE BEHAVIORIAL

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PHENOTYPENTS OR PATHWAYS IN

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ISOLATION BUT THINKING HOW THEY

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CONNECT, REGULATE AND IMPACT ON

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1 ANOTHER IN A BROADER RESPONSE,

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AND WE NOW HAVE TOOLS AND

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TECHNIQUES TO PICK THIS APART

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AND I WILL BE HIGHLIGHTING SOME

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OF THOSE IN MY TALK TODAY.

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NOW WE KNOW THAT DNA DAMAGE AND

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CELLULAR RESPONSE FOR DNA DAMAGE

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UNDERLINE MANY DECS AND YOU WILL

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BE AWARE OF JUST ABOUT ALL OF

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THESE BUT I WANT TO HIGHLIGHT,

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BUT THE OUTFIELD IS AMAZING IN

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TERMS OF HUMAN BIOLOGY AND

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DISEASE.

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THESE FACTORS GOVERN HYPER

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SENSITIVITY TO RADIATION AND

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CHEMO THERAPIES USED IN CANCER.

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THERE ARE DEFECTS IN THESE

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PATHWAYS GIVE RISE TO

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DEVELOPMENT DEFECTS,

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DEFICIENCIES, NEURODEGENERATION,

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PREMATURE AGING, INFERTILITY AND

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DIABETES APPROXIMATE A RANGE OF

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OTHER THINGS BUT MOST OF THESE

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IS IN THE REALM OF CANCER WHERE

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DNA DAMAGE AND THE DNA DAMAGE

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RESPONSE AND REPAIR MECHANISMS

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HAS HUGE RELEVANCE.

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WE KNOW THAT DNA DAMAGE CAN GIVE

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RISE TO MUTATIONS WHICH IS

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CRUCIAL FOR CAR SIN O GENESIS,

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AND WE KNOW THAT MANY DDR

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DEFECTS INHERITED OR ACQUIRED

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PROMOTE CANCER.

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THE MAIN STAYS OF CANCER THERAPY

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REMAIN IONINIZING RADIATION AND

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MOST CHEMO TERRAPY THERAPEUTICS

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WORK BY INFLECTING DNA DAMAGE.

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OTHER AREAS WHERE DNA DAMAGE

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IMPACT ON CANCER BUT OVER THE

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LAST COUPLE DECADES WE LEARNED

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THE DDR COMPARE MECHANISMS

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CONTAIN VARIOUS ATTRACTED DRUG

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TARGETS AND I IMAGINE, IN MY

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VIEW, THE POSTER CHILD FOR THIS,

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AT LEAST AT THIS TIME, ARE PARP

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INHIBITORS INCLUDING PARP

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INHIBITORS 1 AND 2, TAKEN BY THE

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FIRST PATIENTS BY MY COMPANY

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[INDISCERNIBLE] WHICH WAS

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ACQUIRED BY AFT RAZENECCA AND

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IT'S REACHING WELL OVER A

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HUNDRED THOUSAND PATIENTS RIGHT

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NOW TO RECEIVE THIS DRUG AND I

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SHOULD ALSO POINT OUT THERE ARE

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OTHER PARTNERSHIP --PARP

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INHIBITORSS THAT ARE MAKING

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INROADS.

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THIS IS EXITING.

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SOPHISTICATED WHAT IS

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PARTICULARLY EXCITING IS HAS

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PARP INHIBITORS BECOME MORE

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ENTRENCH INDEED CANCER THERAPY

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AND THEY'RE BEING BROADENED,

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THEY'RE USED EARLIER AND EARLIER

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AND IF WE LOOK AT THE SLIDE

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GOING LEFT TO RIGHT, WE CAN SEE

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THAT PARP INHIBITORS SUCH AS

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[INDISCERNIBLE] ARE NOW MOVING

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TOWARDS EVEN IN FIRST LINE

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TREATMENT AND WHAT CLINICIANS

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ARE SEEING IS THAT THE EARLIER

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PARP INHIBITORS ARE USED WITH

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THE RIGHT TYPES OF PATIENTS

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THOSE WITH HRD, MORGS

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DEFICIENCY, RECOMBINATION FOR

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EXAMPLE, THE IMPACT ON THESE

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PATIENT COHORT SYSTEM BECOMING

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MORE EVER PRONOUNCED AND ON THE

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RIGHT HAND SIDE, IT'S REALLY,

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REALLY EXCITING DATA SHOWING

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THAT THE [INDISCERNIBLE]

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PROGRESSION FREE SURVIVAL IS

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MUCH, MUCH BETTEROT TREATMENT

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ARM AND OF COURSE YOU WILL NOTE

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THAT THESE CURVES ARE FLAT

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LINING OR STARTED TO FLAT LINE

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TOWARDS THE END WHICH INDICATES

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THAT MANY OF THESE PATIENTS ARE

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BENEFITING FOR CONSIDERABLE

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LINKS OF TIME AND INDEED THERE

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MAY BE IN DUE COURSE, CERTAIN

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INDIVIDUAL WHO IS COULD BE

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CLAIMED AS CURES THROUGH THESE

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KIND OF APPROACHES.

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GOING ON FOR DNA REPAIR AND

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ENZYMES IS NOW CAUGHT ON AND

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THERE ARE A RAINCHL OF COMPANIES

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AND OF COURSE ACADEMIC LABS IN

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THIS SPACE IDENTIFYING SYNTHETIC

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LETHAL AS SOMEWHAT OTHER

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OPPORTUNITIES WHICH OFFER HOPE

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IN THE NOTE TOO DISTANT FUTURE

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FURTHER FDA AND E, INA APPROVED

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MEDICINES FOR TREATING CANCER.

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SO GOOD LUCK WITH ALL THOSE

259
00:08:42,829 --> 00:08:44,664
PROGRAMS AND HOPEFULLY THERE

260
00:08:44,664 --> 00:08:46,265
WILL BE POSITIVE NEWS IN VARIOUS

261
00:08:46,265 --> 00:08:47,700
FRONTS OVER THE COMING YEARS.

262
00:08:47,700 --> 00:08:48,901
BUT THAT'S IMPORTANT BUT AS I

263
00:08:48,901 --> 00:08:50,770
MENTIONED A FEW MINUTES AGO,

264
00:08:50,770 --> 00:08:53,306
MOST CANCERS ARE TREATED BY

265
00:08:53,306 --> 00:08:56,509
SURGERY OR BY AND/OR RADIATION

266
00:08:56,509 --> 00:08:58,945
AND CONVENTIONAL CHEMO THERAPIES

267
00:08:58,945 --> 00:09:00,046
OR TRADITIONAL ANTICANCER CHEMO

268
00:09:00,046 --> 00:09:01,347
THERAPIES AND WE DON'T ACTUALLY

269
00:09:01,347 --> 00:09:04,050
REALLY UNDERSTAND HOW MANY OF

270
00:09:04,050 --> 00:09:05,585
THESE TRADITIONAL CANCER CHEMO

271
00:09:05,585 --> 00:09:07,420
THERAPY ACTUALLY WORK AND HOW

272
00:09:07,420 --> 00:09:08,821
THEY SOMETIMES SUCCEED BUT OFTEN

273
00:09:08,821 --> 00:09:10,490
FAIL.

274
00:09:10,490 --> 00:09:13,926
NOW 1 OF THESE THERAPIES IS

275
00:09:13,926 --> 00:09:16,596
BASED ON A COMPOUND CALLED

276
00:09:16,596 --> 00:09:17,463
ATOPOCIDE WHICH IS SHOWN ON THE

277
00:09:17,463 --> 00:09:20,299
LEFT HERE AND THIS IS USED IN A

278
00:09:20,299 --> 00:09:21,968
RANGE OF OF DIFFERENT LIQUID AS

279
00:09:21,968 --> 00:09:23,402
WELL AS SOLID CANCERS BUT THERE

280
00:09:23,402 --> 00:09:26,105
ARE ISSUES WITH THESE

281
00:09:26,105 --> 00:09:28,074
TREATMENTS, THERE'S A

282
00:09:28,074 --> 00:09:32,311
2-3 PERCENT RISK OF DEVELOPING

283
00:09:32,311 --> 00:09:35,047
SECONDARY LEUKEMIAS, SHOWING

284
00:09:35,047 --> 00:09:36,516
CHROMOSOMAL TRANSWELLICATIONS BY

285
00:09:36,516 --> 00:09:38,918
DNA REPAIR MECHANISMS BY NOT YET

286
00:09:38,918 --> 00:09:39,719
FULLY UNDERSTOOD MECHANISMS.

287
00:09:39,719 --> 00:09:42,154
AND OF COURSE CHALLENGES TO

288
00:09:42,154 --> 00:09:43,322
MAXIMIZE EFFICACY, REDUCE SIDE

289
00:09:43,322 --> 00:09:44,590
EFFECTS AND OVERCOME RESISTANCE

290
00:09:44,590 --> 00:09:46,158
TO THESE KINDS OF DRUGS AND

291
00:09:46,158 --> 00:09:46,993
MAYBE SIMILAR TO WHAT WE'RE

292
00:09:46,993 --> 00:09:52,532
THINKING ABOUT IN THE CONTEXT OF

293
00:09:52,532 --> 00:09:54,367
PARP INHIBITORS TO TAILOR

294
00:09:54,367 --> 00:09:55,735
THERAPIES TO RIGHT CANCERS TO

295
00:09:55,735 --> 00:09:58,170
THE RIGHT TIME TO THE RIGHT

296
00:09:58,170 --> 00:09:58,638
PATIENTS.

297
00:09:58,638 --> 00:10:00,806
SO WHAT DOES ATOPOCIDE DO?

298
00:10:00,806 --> 00:10:05,845
IT BINDS TO AND TRAPS DNA

299
00:10:05,845 --> 00:10:07,880
POLYISOMMER ACE 2, TOP 2 ALPHA

300
00:10:07,880 --> 00:10:11,250
AND TOP 2 BETA, IT TRAPS THESE

301
00:10:11,250 --> 00:10:13,753
IN A CLEAVAGE COMPLEX

302
00:10:13,753 --> 00:10:15,254
INTERMEDIATE DURING ITS

303
00:10:15,254 --> 00:10:16,088
ENZYMATIC CYCLE.

304
00:10:16,088 --> 00:10:19,492
SO TOP 2 ENZYMES MODULATE DNA

305
00:10:19,492 --> 00:10:22,695
TOPOLOGY DURING DNA REPLICATION,

306
00:10:22,695 --> 00:10:25,264
RECOMBINATION, AND SISTER

307
00:10:25,264 --> 00:10:25,831
CHROMATIN DECANTONNATION.

308
00:10:25,831 --> 00:10:28,100
AND THE WAY TOP 2 ENZYMES WORK

309
00:10:28,100 --> 00:10:29,669
IS TO GENERATE A DOUBLE STRAND

310
00:10:29,669 --> 00:10:31,604
BREAK IN 1 DNA DOUBLE HELIX AND

311
00:10:31,604 --> 00:10:34,140
THEN PASS THAT THROUGH ANOTHER

312
00:10:34,140 --> 00:10:36,075
DNA DOUBLE HELIX, THEN RESEAL

313
00:10:36,075 --> 00:10:38,444
THAT DOUBLE STRAND BREAK AND

314
00:10:38,444 --> 00:10:40,513
THEREFORE OBVIOUSLY IN THIS

315
00:10:40,513 --> 00:10:42,481
SCENARIO HERE CAN DISENTANGLE OR

316
00:10:42,481 --> 00:10:45,918
WE MOVE TOPOLOGY FROM DNA

317
00:10:45,918 --> 00:10:46,752
MOLECULES.

318
00:10:46,752 --> 00:10:48,654
AND WHAT ATOPOCIDE DOES, IT

319
00:10:48,654 --> 00:10:51,591
BINDS TO THIS REACTION ISHT

320
00:10:51,591 --> 00:10:53,893
MEDIATE WHERE THERE'S A CLEAVAGE

321
00:10:53,893 --> 00:10:56,762
COMPLEX, AND IN THIS CLEAVAGE

322
00:10:56,762 --> 00:10:59,532
COMPLEX, TOP 2 IS ATTACHED TO A

323
00:10:59,532 --> 00:11:01,334
LINKAGE TO THE 5 PRIME ENDS OF

324
00:11:01,334 --> 00:11:02,401
THE DOUBLE STRAND BREAK.

325
00:11:02,401 --> 00:11:04,870
SO WE HAVE A DOUBLE STRAND BREAK

326
00:11:04,870 --> 00:11:06,739
IN THE DNA, BUT IT'S NOT NAKED,

327
00:11:06,739 --> 00:11:10,209
IT'S AN ENZYME COUPLED COVALENT

328
00:11:10,209 --> 00:11:11,844
INTERMEDIATE AND OF COURSE

329
00:11:11,844 --> 00:11:13,479
TRAPPING THESE WITH ATOPOCIDE

330
00:11:13,479 --> 00:11:14,780
POSES MAJOR PROBLEMS TO CELLS,

331
00:11:14,780 --> 00:11:16,349
THIS IS CLEARLY GOING TO BE A

332
00:11:16,349 --> 00:11:20,119
MAJOR BARRIER TO THINGS SUCH AS

333
00:11:20,119 --> 00:11:21,420
REPLICATION OR TRANSCRIPTION.

334
00:11:21,420 --> 00:11:24,457
SO TO DEAL WITH THESE KINDS OF

335
00:11:24,457 --> 00:11:26,525
LESIONS AND PRESUMABLY NATURALLY

336
00:11:26,525 --> 00:11:29,462
ARISING STORE CLEAVAGE COMPLEXES

337
00:11:29,462 --> 00:11:30,262
WHICH PROBABLY TAKING PLACE ALL

338
00:11:30,262 --> 00:11:32,832
THE TIME IN THE ABSENCE OF THE

339
00:11:32,832 --> 00:11:34,900
ATOPOCIDE IN RESPONSE TO BASE

340
00:11:34,900 --> 00:11:37,937
DAMAGE OR OTHER CHEMICALS, THERE

341
00:11:37,937 --> 00:11:40,039
ARE A RANGE OF DNA REPAIR

342
00:11:40,039 --> 00:11:41,440
SYSTEMS THAT DEAL WITH THESE,

343
00:11:41,440 --> 00:11:43,209
NOW 1 OF THESE SYSTEMS SHOWN ON

344
00:11:43,209 --> 00:11:45,077
THE LEFT HERE, IS MEDIATED, THE

345
00:11:45,077 --> 00:11:50,883
TOP OF THE PATHWAY IS A PROTEIN

346
00:11:50,883 --> 00:11:56,389
CALLED ZNF451, ALSO CALLED ZAT,

347
00:11:56,389 --> 00:12:00,393
AND IT ATTACHING SUMO UNITS TO

348
00:12:00,393 --> 00:12:01,260
TOP 2.

349
00:12:01,260 --> 00:12:05,531
THIS THEN SERVES AS AN ATTRACT

350
00:12:05,531 --> 00:12:09,235
ANT FOR TDP2.

351
00:12:09,235 --> 00:12:12,271
TDP2, HAS SAME MOTIF, SAME

352
00:12:12,271 --> 00:12:13,973
INTERACTION MOTIFS WHICH

353
00:12:13,973 --> 00:12:17,543
RECOGNIZE THE SUMO RELATED TOP

354
00:12:17,543 --> 00:12:19,311
2, TDB2 RECRUITMENT USES

355
00:12:19,311 --> 00:12:20,413
ENZYMATIC ACTIVITY TO CLEAVE THE

356
00:12:20,413 --> 00:12:23,749
FOSTER NURSED FOCUSED ON

357
00:12:23,749 --> 00:12:24,550
TYROSOLLINKAGES, RELEASING THE

358
00:12:24,550 --> 00:12:27,753
DOUBLE STRAND BREAK THAT CAN BE

359
00:12:27,753 --> 00:12:30,022
REPAIRED BY JOINING MECHANISMS

360
00:12:30,022 --> 00:12:31,557
OFTEN IN A GREAT FASHION.

361
00:12:31,557 --> 00:12:33,092
THERE'S AN ALTERNATIVE TYPE OF

362
00:12:33,092 --> 00:12:35,828
SET OF PATHWAYS THAT CAN ALSO

363
00:12:35,828 --> 00:12:37,363
ARISE AT STABILIZED TOP 2

364
00:12:37,363 --> 00:12:38,431
CLEAVAGE COMPLEXES AND AT THE

365
00:12:38,431 --> 00:12:42,301
TOP OF THIS PATHWAY IS A SUMO

366
00:12:42,301 --> 00:12:45,504
TARGETED UBIQUITIN LIGASE OR

367
00:12:45,504 --> 00:12:49,108
STUBBLE CALLED RNF4, WHICH

368
00:12:49,108 --> 00:12:52,111
ASSOCIATES WITH THE ASSIMILATED

369
00:12:52,111 --> 00:12:54,146
TOP 2, MEDIATES UBIQUITINNATION

370
00:12:54,146 --> 00:12:56,315
WHICH THEN LEADS TO PROTEOSOME

371
00:12:56,315 --> 00:12:56,615
DEGRADATION

372
00:12:56,615 --> 00:12:57,583
GRATES--GRATUEIGATION EMPLOY NOW

373
00:12:57,583 --> 00:12:59,985
UNLIKE THE SITUATION WITH TDP2

374
00:12:59,985 --> 00:13:03,456
WHICH CAN CLEANLY REMOVE THE

375
00:13:03,456 --> 00:13:05,091
COVALENTLY ATTACHED TOP 2, RNF 4

376
00:13:05,091 --> 00:13:07,626
DOES AND THE PATHWAY WILL NIBBLE

377
00:13:07,626 --> 00:13:09,395
AWAY AND EAT AWAY AT THAT BUT IS

378
00:13:09,395 --> 00:13:10,930
UNABLE TO REMOVE THE FINAL

379
00:13:10,930 --> 00:13:13,032
PEPTIDE AND THEY MUST BE REMOVED

380
00:13:13,032 --> 00:13:14,633
BEFORE DMA REPAIR TAKES PLACE

381
00:13:14,633 --> 00:13:17,169
AND THOSE PEPTIDE BLOCKED DOUBLE

382
00:13:17,169 --> 00:13:19,405
STRAND BREAKS ARE PROCESSED BY

383
00:13:19,405 --> 00:13:20,439
TDP2, FOLLOWING INTO THE PATHWAY

384
00:13:20,439 --> 00:13:23,275
IN THE MIDDLE OR THROUGH VARIOUS

385
00:13:23,275 --> 00:13:25,344
NUCLEACES WHICH ALLOW MORE ARROW

386
00:13:25,344 --> 00:13:28,814
PRONE PATHWAYS SUCH AS

387
00:13:28,814 --> 00:13:30,649
NORMAL--SORRY ALTERNATIVE AND

388
00:13:30,649 --> 00:13:32,585
JOINING POSSIBLY ENJOINING AS

389
00:13:32,585 --> 00:13:34,787
WELL AS BY TDP2, FOLLOWING ON

390
00:13:34,787 --> 00:13:37,857
INTO THE PATHWAY IN THE MIDDLE

391
00:13:37,857 --> 00:13:40,159
ALL THROUGH VARIOUS NUCLEACES

392
00:13:40,159 --> 00:13:43,028
WHICH ALLOW ERROR PRONE PATHWAYS

393
00:13:43,028 --> 00:13:45,431
SUCH AS ALTERNATIVE END JOINING

394
00:13:45,431 --> 00:13:47,500
[INDISCERNIBLE] END JOINING AS

395
00:13:47,500 --> 00:13:49,101
WELL AS TDP2, FOLLOWING INTO THE

396
00:13:49,101 --> 00:13:51,437
PATHWAY IN THE MIDDLE OR THROUGH

397
00:13:51,437 --> 00:13:54,173
VARIOUS NUCLEACES WHICH ALLOW

398
00:13:54,173 --> 00:13:57,042
MORE EROR PRONE PATHWAYS SUCH AS

399
00:13:57,042 --> 00:13:58,744
[INDISCERNIBLE] ALTERNATIVE AND

400
00:13:58,744 --> 00:14:00,379
JOINING POSSIBLY CANNED WHAT

401
00:14:00,379 --> 00:14:02,715
JOINING AS WELL AS TDP2,

402
00:14:02,715 --> 00:14:03,883
FOLLOWING INTO THE PATHWAY IN

403
00:14:03,883 --> 00:14:06,285
THE MIDDLE OR THROUGH VARIOUS

404
00:14:06,285 --> 00:14:11,490
NUCLEACES WHICH ALLOW MORE ERROR

405
00:14:11,490 --> 00:14:13,292
PRONE PATHWAYS [INDISCERNIBLE]

406
00:14:13,292 --> 00:14:14,794
ALTERNATIVE END JOINING AND

407
00:14:14,794 --> 00:14:17,329
[INDISCERNIBLE] END JOINING AS

408
00:14:17,329 --> 00:14:18,397
WELL AS TDP2 FOLLOWING IN THE

409
00:14:18,397 --> 00:14:20,933
PATHWAY IN THE MITSD ALL THROUGH

410
00:14:20,933 --> 00:14:22,101
VARIOUS NUCLEACES WHICH ALLOW

411
00:14:22,101 --> 00:14:25,171
MORE ERROR PRONE PATHWAYS SUCH

412
00:14:25,171 --> 00:14:26,639
AS NON[INDISCERNIBLE]

413
00:14:26,639 --> 00:14:27,840
ALTERNATIVE END JOINING AND

414
00:14:27,840 --> 00:14:31,343
POSSIBLY [INDISCERNIBLE] AS WELL

415
00:14:31,343 --> 00:14:33,746
AS IN THE PATHWAY IN THE MIDDLE

416
00:14:33,746 --> 00:14:36,916
ALL THROUGH VARIOUS NUCLEACES TO

417
00:14:36,916 --> 00:14:47,426
ALLOW FOR ERROR PRONE PATHWAYS

418
00:14:49,449 --> 00:14:53,753
--YOU CAN SEE THERE'S NF451 BUT

419
00:14:53,753 --> 00:14:55,254
WE ALSO IDENTIFIED 2 FACTORS

420
00:14:55,254 --> 00:14:58,925
THAT WE HIGHLIGHT IN RED, RAD 54

421
00:14:58,925 --> 00:15:01,060
L2 EXPW ERCC6 L2 WHICH AT THAT

422
00:15:01,060 --> 00:15:02,195
TIME WERE UNCHARACTERRIZED IN

423
00:15:02,195 --> 00:15:05,865
TERMS OF THEIR DVR ROLES.

424
00:15:05,865 --> 00:15:08,701
THESE ARE BOTH ATP ACE HELIX

425
00:15:08,701 --> 00:15:12,105
CASE ENZYMES. 

426
00:15:12,105 --> 00:15:16,075
NOW FOLLOWING ENSUING WORK BY US

427
00:15:16,075 --> 00:15:16,809
ESTABLISHED THAT RAV54 KNOCK OUT

428
00:15:16,809 --> 00:15:21,080
CELLS WE CAN SPECIFICALLY MAKE

429
00:15:21,080 --> 00:15:28,287
IN 1 RPA AND 1 UTOS BACKGROUNDS

430
00:15:28,287 --> 00:15:32,191
WITH CRISPR CAS9 TECHNOLOGY, AND

431
00:15:32,191 --> 00:15:33,926
WHAT WE FIND FOR THE KNOCK OUT

432
00:15:33,926 --> 00:15:35,895
CELL SYSTEM THAT THEY'RE

433
00:15:35,895 --> 00:15:37,530
SPECIFICALLY HYPER SENSITIVE TO

434
00:15:37,530 --> 00:15:37,897
ATOPOCIDE.

435
00:15:37,897 --> 00:15:41,901
SO KNOCK OUT CLONES FOR RAV54 L2

436
00:15:41,901 --> 00:15:43,636
ARE MORE SENSITIVE THAN CONTROLS

437
00:15:43,636 --> 00:15:44,837
TO ATOPOCIDE BUT WE DON'T SEE

438
00:15:44,837 --> 00:15:47,273
THIS FOR A RANGE OF DNA OTHER

439
00:15:47,273 --> 00:15:48,808
DAMAGINGLY AGENTS WE USED AND

440
00:15:48,808 --> 00:15:51,577
THIS ACTUALLY CONTRASTED TO THE

441
00:15:51,577 --> 00:15:53,546
SITUATION FOR ERCC 6 L2 WHICH WE

442
00:15:53,546 --> 00:15:54,947
FOUND WERE HYPER SENSITIVE TO A

443
00:15:54,947 --> 00:15:56,382
RAINCHL OF DNA DAMAGING AGENTS

444
00:15:56,382 --> 00:15:59,585
AND HAVE BEEN NOW PUBLISHED BY

445
00:15:59,585 --> 00:16:04,157
LABS OF [INDISCERNIBLE] AND

446
00:16:04,157 --> 00:16:06,425
[INDISCERNIBLE] TO HAVE LINKAGES

447
00:16:06,425 --> 00:16:09,629
AND MORE IN THE ENJOINING

448
00:16:09,629 --> 00:16:09,996
PROCESSES.

449
00:16:09,996 --> 00:16:13,766
WE CHOSE TO FOCUS ON RAD 54 L2

450
00:16:13,766 --> 00:16:16,202
TO SEE HOW DNA INDUCED DAMAGE

451
00:16:16,202 --> 00:16:17,370
COULD BE BROUGHT ABOUT AND A KEY

452
00:16:17,370 --> 00:16:19,438
SET OF EXPERIMENTS THAT REALLY

453
00:16:19,438 --> 00:16:21,941
SET US ON PATH TO UNDERSTAND

454
00:16:21,941 --> 00:16:23,476
THANKSGIVING FACTOR WAS THROUGH

455
00:16:23,476 --> 00:16:28,347
A COLLABORATION WITH

456
00:16:28,347 --> 00:16:29,115
PETRA [INDISCERNIBLE] AND

457
00:16:29,115 --> 00:16:30,516
[INDISCERNIBLE] IN GERMANY.

458
00:16:30,516 --> 00:16:33,786
SO WE PROVIDED PETRA'S LAB WITH

459
00:16:33,786 --> 00:16:37,990
CELL LINES EXPRESSING RFP ATTACK

460
00:16:37,990 --> 00:16:41,160
EXPRESSING CELLS OR EXPRESSING

461
00:16:41,160 --> 00:16:46,199
PROTEIN LINKED TO RAV54 L2. 

462
00:16:46,199 --> 00:16:48,367
WE THEN ISOLATED THE THESE

463
00:16:48,367 --> 00:16:49,569
PROTEINS BY IMMUNO PRECIPITATION

464
00:16:49,569 --> 00:16:54,440
AND THESE WERE THEN SUBJECT TO

465
00:16:54,440 --> 00:16:57,210
MASS SPEC. PROTEOMICS BY PETRA'S

466
00:16:57,210 --> 00:16:57,443
LAB.

467
00:16:57,443 --> 00:16:58,611
NOW SATISFYING IF WE FIRST LOOK

468
00:16:58,611 --> 00:17:03,316
AT THE DATA WE OBTAINED WITH NO

469
00:17:03,316 --> 00:17:05,017
ATOPOCIDE WITH THE CELLS

470
00:17:05,017 --> 00:17:08,454
UNTREATED WE FIND RAVPOOR L2

471
00:17:08,454 --> 00:17:09,422
UNCONTROLLED, WE PULL IT DOWN,

472
00:17:09,422 --> 00:17:13,426
WE SEE THAT, BUT YOU ALSO NOTE

473
00:17:13,426 --> 00:17:15,828
THAT WE SEE SUMO 1, 2, 3, AND

474
00:17:15,828 --> 00:17:18,164
THE TOP 2 IN HUMAN CELLS AND

475
00:17:18,164 --> 00:17:20,633
ALPHA TOP 2 BETA AND ALPHA.

476
00:17:20,633 --> 00:17:22,969
SO THIS DID SAY TO US THAT THIS

477
00:17:22,969 --> 00:17:24,070
FACTOR GIVING THE SENSITIVITY IS

478
00:17:24,070 --> 00:17:25,471
CONNECTED TO THE TYPES OF THINGS

479
00:17:25,471 --> 00:17:28,975
THAT WE KNOW NEED TO OPERATE TO

480
00:17:28,975 --> 00:17:31,477
MEDIATE TOP 2 CLEAVAGE COMPLEX

481
00:17:31,477 --> 00:17:31,844
REPAIR.

482
00:17:31,844 --> 00:17:32,445
THINGS BECOME EVEN MORE

483
00:17:32,445 --> 00:17:35,548
INTERESTING WHEN WE LOOKA THE

484
00:17:35,548 --> 00:17:36,649
ATOPOCIDE TREATED CELLS.

485
00:17:36,649 --> 00:17:38,851
WE ALSO GET TOP 2 ALPHA AND TOP

486
00:17:38,851 --> 00:17:39,218
2 BETA.

487
00:17:39,218 --> 00:17:41,554
YOU WILL SEE NOW A VERY, VERY

488
00:17:41,554 --> 00:17:43,623
STRONG INTERACTOR HERE IS AN

489
00:17:43,623 --> 00:17:45,391
F451 WHICH IS NOT A STRONG HIT

490
00:17:45,391 --> 00:17:47,260
IN THE UNTREATED CONDITION.

491
00:17:47,260 --> 00:17:49,795
SO, THIS SUGGESTED TO US THAT

492
00:17:49,795 --> 00:17:52,665
RAV54 L2 MUST BE CONNECTED TO

493
00:17:52,665 --> 00:17:56,569
THE SUMO E3 LIGASE, AS AN F451,

494
00:17:56,569 --> 00:17:58,337
WHICH HAS BEEN SHOWN A FEW

495
00:17:58,337 --> 00:18:01,307
MOMENTS AGO TO SUMO ELATE TOP 2

496
00:18:01,307 --> 00:18:03,576
ALPHA AND TOP 2 BETA.

497
00:18:03,576 --> 00:18:05,811
SO, WHAT WAS RAV 54 L2 LOOK

498
00:18:05,811 --> 00:18:08,648
LIKE, THIS IS A DEPICTION ON THE

499
00:18:08,648 --> 00:18:11,250
LEFT-HAND SIDE, IT'S AN ATP ACE

500
00:18:11,250 --> 00:18:12,818
HELIX CASE ENWRIEM AND IT ALSO

501
00:18:12,818 --> 00:18:14,687
HAS A RANGE OF MOTIFS THAT ARE

502
00:18:14,687 --> 00:18:16,889
PREDICTED TO IRPT ACT WITH SUMO.

503
00:18:16,889 --> 00:18:19,225
THESE ARE CALLED SUMO

504
00:18:19,225 --> 00:18:20,660
INTERACTING MOTIFS OR SIMS, SO

505
00:18:20,660 --> 00:18:22,795
THIS ALLOWED US TO GO AHEAD AND

506
00:18:22,795 --> 00:18:25,531
MAKE MUTATED VERSIONS OF RAV54

507
00:18:25,531 --> 00:18:27,767
L2, .2 MUTATE INDEED THE

508
00:18:27,767 --> 00:18:29,235
ANALYTIC SITE TO KILL THE

509
00:18:29,235 --> 00:18:31,037
ACTIVITY OR MUTATE INDEED

510
00:18:31,037 --> 00:18:32,204
MULTIPLE SIMS PRESUMABLY,

511
00:18:32,204 --> 00:18:33,139
HOPEFULLY THEN TO PREVENT THE

512
00:18:33,139 --> 00:18:36,876
ABILITY TO IRPT ACT WITH SUMO

513
00:18:36,876 --> 00:18:38,077
RELATED PROTEINS.

514
00:18:38,077 --> 00:18:38,778
SO THROUGH COMPLEMENTATION

515
00:18:38,778 --> 00:18:41,414
TODAYS AS YOU WILL SEE IN THE

516
00:18:41,414 --> 00:18:43,249
GRAPH BELOW, COMPARED TO

517
00:18:43,249 --> 00:18:47,953
WILD-TYPE CELLS, SHOWN IN BLUE,

518
00:18:47,953 --> 00:18:50,690
CELLS COMPLEMENTED--SORRY, CELLS

519
00:18:50,690 --> 00:18:53,592
LACKING RAV54 L2 ARE HYPER

520
00:18:53,592 --> 00:18:55,328
SENSITIVE TO ATOPOCIDE AND

521
00:18:55,328 --> 00:18:56,495
IMPLEMENTATION OF THESE CELLS

522
00:18:56,495 --> 00:18:58,564
WITH THE CATALYTIC DEAD OR SAME

523
00:18:58,564 --> 00:19:02,768
MUTANT PROTEIN DOES NOT RESTORE

524
00:19:02,768 --> 00:19:03,336
RESISTANCE.

525
00:19:03,336 --> 00:19:04,804
BY CONTRAST, COMPLEMENTING THESE

526
00:19:04,804 --> 00:19:09,208
CELLS WITH WILD-TYPE RAV54 L2

527
00:19:09,208 --> 00:19:10,242
DOES.

528
00:19:10,242 --> 00:19:12,211
AND IN FACT, WE CAN SEE THESE

529
00:19:12,211 --> 00:19:14,413
CELLS ARE MORE RESISTANT THAN

530
00:19:14,413 --> 00:19:15,314
THE WILD-TYPE CELLS.

531
00:19:15,314 --> 00:19:18,117
SO THESE CELLS ARE INDICATING

532
00:19:18,117 --> 00:19:20,986
THAT THE ATP-CASE, RAV54 L2 AS

533
00:19:20,986 --> 00:19:22,955
WELL AS SIEWM OR INTERACTION

534
00:19:22,955 --> 00:19:25,591
DOMAINS ARE IMPORTANT FOR ITS

535
00:19:25,591 --> 00:19:26,392
ACTIVITY.

536
00:19:26,392 --> 00:19:27,326
CONSISTENT WITH THIS, AND ALSO

537
00:19:27,326 --> 00:19:29,228
CONSIST WENT THE PROTEOMICS

538
00:19:29,228 --> 00:19:32,965
DATA, WE FIND THROUGH IP-IP

539
00:19:32,965 --> 00:19:34,700
WESTERN BLOT STUDIES AS YOU CAN

540
00:19:34,700 --> 00:19:38,637
SEE HERE THAT RAV 54 L2, AND IT

541
00:19:38,637 --> 00:19:40,906
IRPT ACTING IN A CONSTITTATIVE

542
00:19:40,906 --> 00:19:42,808
MANNER WITH ALPHA TOP 2 ALPHA

543
00:19:42,808 --> 00:19:45,511
AND BETA, BUT IN RESPONSE TO

544
00:19:45,511 --> 00:19:47,880
ATOPOCIDE TREATMENT, WHICH

545
00:19:47,880 --> 00:19:49,515
GENERATES A MODIFIED FORM, TOP

546
00:19:49,515 --> 00:19:52,518
2, THIS FACTOR BINDS TO THOSE

547
00:19:52,518 --> 00:19:54,120
FORMS THE PROTEIN, HOWEVER, IF

548
00:19:54,120 --> 00:19:56,856
WE LOOK AT THIS DELTA SAME

549
00:19:56,856 --> 00:19:58,958
MUTANT SHOWN TO THE RIGHT HERE,

550
00:19:58,958 --> 00:20:00,860
THAT'S INTERACT WITH THE TOP 2,

551
00:20:00,860 --> 00:20:04,397
ALPHA AND TOP 2 BETA, BUT NOT

552
00:20:04,397 --> 00:20:05,898
THE MODIFIED FORMS, EVEN THOUGH

553
00:20:05,898 --> 00:20:08,401
THEY'RE EXISTING IN THIS

554
00:20:08,401 --> 00:20:08,667
SCENARIO.

555
00:20:08,667 --> 00:20:10,336
AND YOU WILL ALSO LIKE FROM THE

556
00:20:10,336 --> 00:20:12,471
BOTTOM, ALSO CONSISTENT WITH THE

557
00:20:12,471 --> 00:20:18,477
MASS SPEC DATA WE ALSO SEE AN

558
00:20:18,477 --> 00:20:19,545
INTERACTION WITH ZEBF451 WHICH

559
00:20:19,545 --> 00:20:22,982
IS REDUCED AND THIS IS LARGELY

560
00:20:22,982 --> 00:20:26,252
AGGREGATED IN THE DELTA MUTANT.

561
00:20:26,252 --> 00:20:28,154
SO WHILE WE'RE AT THAT STAGE, WE

562
00:20:28,154 --> 00:20:31,157
CLEARLY CONNECTED IT INTO A

563
00:20:31,157 --> 00:20:32,324
ATOPOCIDE SENSITIVITY AND

564
00:20:32,324 --> 00:20:34,093
PRESUMABLY TOP 2 CLEAVAGE

565
00:20:34,093 --> 00:20:34,894
COMPLEX PROCESSING AND/OR REPAIR

566
00:20:34,894 --> 00:20:36,862
BUT WE WANT TO FIGURE OUT WHERE

567
00:20:36,862 --> 00:20:38,998
IT MAPPED WITHIN THE NETWORK OF

568
00:20:38,998 --> 00:20:39,965
PROTEINS ALREADY KNOWN.

569
00:20:39,965 --> 00:20:41,901
SO THE APPROACH WE USED HERE AND

570
00:20:41,901 --> 00:20:43,869
THIS IS AN APPROACH WE'RE USE

571
00:20:43,869 --> 00:20:45,037
NOTHING MANY PROJECTS IN MY

572
00:20:45,037 --> 00:20:48,808
GROUP IS TO CARRY OUT NOT 1

573
00:20:48,808 --> 00:20:50,643
CRISPR KREENS BUT A RANGE OF

574
00:20:50,643 --> 00:20:51,777
THEM ACROSS GENETIC BACKGROUNDS.

575
00:20:51,777 --> 00:20:53,312
SO IF YOU THINK OF THIS SCENARIO

576
00:20:53,312 --> 00:20:55,581
HERE WE MIGHT HAVE A WILD-TYPE

577
00:20:55,581 --> 00:20:58,484
CELL BACKGROUND, CARRY OUT A

578
00:20:58,484 --> 00:20:59,285
CRISPR SCREEN, FOR THE

579
00:20:59,285 --> 00:21:00,453
PARTICULAR DRUG AND WE IESHES

580
00:21:00,453 --> 00:21:02,888
DEBTIFY A SET OF GENES A-G.

581
00:21:02,888 --> 00:21:06,025
OF COURSE IF WE CARRY OUT

582
00:21:06,025 --> 00:21:09,562
ANOTHER SCREEN IN A CELL THAT'S

583
00:21:09,562 --> 00:21:10,830
KNOCKED OUT FOR GENE A WE WOULD

584
00:21:10,830 --> 00:21:17,269
NOT EXPECT TO SEE ANY FURTHER

585
00:21:17,269 --> 00:21:18,404
SENSITIZATION FOR THE CRISPR

586
00:21:18,404 --> 00:21:20,973
SCREEN, AND IN THIS SCENARIO

587
00:21:20,973 --> 00:21:22,475
HEREY WE SEE THAT GENE C IS

588
00:21:22,475 --> 00:21:23,576
NOTHING LIKE THAT DROPPING OUT

589
00:21:23,576 --> 00:21:26,345
BECAUSE IT'S WORKING ON THE SAME

590
00:21:26,345 --> 00:21:28,981
PATHWAY AS L ORA, WHERE B IS.

591
00:21:28,981 --> 00:21:31,851
SO WE'RE ABLE TO DRAW UP

592
00:21:31,851 --> 00:21:33,018
INTERACTION NETWORKS FOR THE

593
00:21:33,018 --> 00:21:35,221
DRUG AT HAND BUT WE'RE STUDYING

594
00:21:35,221 --> 00:21:37,389
AND THEREBY DRAW LINES OR

595
00:21:37,389 --> 00:21:38,090
CONNECTIONS BETWEEN DIFFERENT

596
00:21:38,090 --> 00:21:39,492
PATHWAYS THAT MIGHT BE BEING

597
00:21:39,492 --> 00:21:42,561
USED IN DIFFERENT CONTEXT.

598
00:21:42,561 --> 00:21:44,296
SO WHEN WE CARRY THIS OUT IN

599
00:21:44,296 --> 00:21:45,130
THIS PARTICULAR EXPERIMENT HERE,

600
00:21:45,130 --> 00:21:47,833
THIS WORK WAS CARRIED OUT BY

601
00:21:47,833 --> 00:21:49,301
[INDISCERNIBLE] A POST DOC IN

602
00:21:49,301 --> 00:21:54,940
THE GROUP WITH HELP FROM

603
00:21:54,940 --> 00:21:55,508
[INDISCERNIBLE], A

604
00:21:55,508 --> 00:21:56,208
BIOMATHEMATICIAN WHO JUST MOVED

605
00:21:56,208 --> 00:21:57,776
DOWN THE ROAD FROM US, WHAT WE

606
00:21:57,776 --> 00:21:59,645
SEE HERE ARE THE RESULTS FROM

607
00:21:59,645 --> 00:22:01,614
PLOTS IN 2 DIFFERENT

608
00:22:01,614 --> 00:22:07,386
BACKGROUNDS, WE HAVE THE

609
00:22:07,386 --> 00:22:09,755
WILD-TYPE CRISPR ALONG THE

610
00:22:09,755 --> 00:22:12,658
BOTTOM AND A IMK OUT IN THE

611
00:22:12,658 --> 00:22:15,628
Y-AXIS AND ANYTHING IN THE LOWER

612
00:22:15,628 --> 00:22:17,363
LEFT QUADRANT ARE DRIVING

613
00:22:17,363 --> 00:22:18,230
SENSITIVITY.

614
00:22:18,230 --> 00:22:19,632
VERY NICE CONTROL IN THIS

615
00:22:19,632 --> 00:22:23,469
EXPERIMENT IS THAT THIS RAV54

616
00:22:23,469 --> 00:22:25,337
L2, ITS, IT'S SERVING AS A GOOD

617
00:22:25,337 --> 00:22:26,972
HIT IN THE WILD-TYPE BACKGROUND

618
00:22:26,972 --> 00:22:29,942
BUT NOT DROPPING OUT IN THE

619
00:22:29,942 --> 00:22:31,343
RAV54 L2 BACKGROUND BECAUSE IT'S

620
00:22:31,343 --> 00:22:35,648
ALREADY KNOCKED OUT IN THAT BACK

621
00:22:35,648 --> 00:22:36,015
IMROWND.

622
00:22:36,015 --> 00:22:37,182
BUT WHILE WE'RE STRIKING IT,

623
00:22:37,182 --> 00:22:38,484
EVERY OTHER FACTOR WE'RE SEEING

624
00:22:38,484 --> 00:22:39,985
IN THIS SCREEN IS STILL LYING

625
00:22:39,985 --> 00:22:41,053
CLOSE TO THE DIAGONAL.

626
00:22:41,053 --> 00:22:44,924
AND THIS IS HIGHLIGHTING TO US

627
00:22:44,924 --> 00:22:47,226
THAT RAV 54 L2 SEEMS TO BE WORKS

628
00:22:47,226 --> 00:22:49,328
IN A DISTINCT MANNER FROM THE

629
00:22:49,328 --> 00:22:53,766
BULK OF KNOWN RESPONDERS TO TOP

630
00:22:53,766 --> 00:22:55,467
2 COMPLEX CANNED WHAT.

631
00:22:55,467 --> 00:22:56,902
BUT 1 THING I NEED TO HIGHLIGHT

632
00:22:56,902 --> 00:22:58,837
TO US IS I'VE LEARNED THAT THE

633
00:22:58,837 --> 00:23:01,473
LIGHT WE USED FOR CRISPR

634
00:23:01,473 --> 00:23:03,309
SCREENING THAT THE GAINS ARE NOT

635
00:23:03,309 --> 00:23:05,711
VERY GOOD AND I WILL COME BACK

636
00:23:05,711 --> 00:23:07,513
TO THAT IN A MOMENT AND I WILL

637
00:23:07,513 --> 00:23:09,782
COME BACK TO THAT IN A MOMENT

638
00:23:09,782 --> 00:23:12,451
BECAUSE THEY DO INDEED CONNECT.

639
00:23:12,451 --> 00:23:15,654
SO VALIDATING THOSE CRISPR DATA

640
00:23:15,654 --> 00:23:17,056
INDICATING THAT RAV54 L2 WORKS

641
00:23:17,056 --> 00:23:18,557
IN A DISTINCT WAY TO OTHER

642
00:23:18,557 --> 00:23:19,858
FACTORS WE'VE KNOWN ABOUT, IF

643
00:23:19,858 --> 00:23:23,295
WHAT WE'RE SEEING ON THE LEFT

644
00:23:23,295 --> 00:23:24,797
PLOT HERE, ARE SURVIVAL CURVES

645
00:23:24,797 --> 00:23:29,201
WITH A RAINCHL OF DIFFERENT

646
00:23:29,201 --> 00:23:31,837
KNOCK OUTS AND IF WE KNOCK OUT

647
00:23:31,837 --> 00:23:35,774
BOGHT RAV 54, L2, MDP2, WHICH

648
00:23:35,774 --> 00:23:43,048
RECORDS PLAYING KEY ROLE, IN

649
00:23:43,048 --> 00:23:44,416
REMOVING THE CLEAVAGE COMPLEX,

650
00:23:44,416 --> 00:23:46,185
WE GET MUCH MORE SENSITIVITY

651
00:23:46,185 --> 00:23:48,053
THAN THE SINGLE MUTANTS ALONE

652
00:23:48,053 --> 00:23:49,588
EMPLOY THE SENSITIVITY HERE IS A

653
00:23:49,588 --> 00:23:51,957
LOW CONCENTRATIONS AND THAT

654
00:23:51,957 --> 00:23:53,425
THESE CONCENTRATIONS ACTUALLY

655
00:23:53,425 --> 00:23:55,394
NOT VERY MUCH SENSITIVE IN THE

656
00:23:55,394 --> 00:24:00,566
KNOCK OUT ON HIS OWN, AND IT IS

657
00:24:00,566 --> 00:24:04,203
WORKING AT A DISTINCT PATHWAY

658
00:24:04,203 --> 00:24:07,106
FROM TDP 2, AND SIMILARLY, FROM

659
00:24:07,106 --> 00:24:08,307
THE COMPETITION ASSAYS, WE LOOK

660
00:24:08,307 --> 00:24:13,746
TO SEE IF RAV 54 L2 WAS WORKING

661
00:24:13,746 --> 00:24:17,750
INSIDE OR OUTSIDE OR HR RELATED

662
00:24:17,750 --> 00:24:20,352
PROCESSES HOMOLOGOUS COMBINATION

663
00:24:20,352 --> 00:24:22,221
ASSOCIATED PROCESSES AND WE SEE

664
00:24:22,221 --> 00:24:30,195
THAT IT'S ADD.

665
00:24:30,195 --> 00:24:33,032
SO RAV54 L2 IS ALSO HAVING AN

666
00:24:33,032 --> 00:24:34,099
IMPACT ON THOSE PROCESSES SO

667
00:24:34,099 --> 00:24:35,501
WHAT IS IT DOING.

668
00:24:35,501 --> 00:24:40,105
THROUGH SIMILAR PRPLTS -

669
00:24:40,105 --> 00:24:42,374
-EXPERIMENTS THAT WHILE

670
00:24:42,374 --> 00:24:48,781
KNOCKING OUT RAV54 L2, 

671
00:24:48,781 --> 00:24:49,648
SENSITIZING CELLS, AND THAT'S

672
00:24:49,648 --> 00:24:51,116
SHOWN ON THE LEFT BUT I THINK

673
00:24:51,116 --> 00:24:54,086
IT'S ACTUALLY EVEN MORE STRIKING

674
00:24:54,086 --> 00:24:55,554
ON THE RIGHT, AND THIS IS

675
00:24:55,554 --> 00:24:58,290
ACTUALLY PART OF THE STUDY THAT

676
00:24:58,290 --> 00:25:06,331
IS AS YET UNPUBLISHED AND

677
00:25:06,331 --> 00:25:08,634
SPEARHEADED BY [INDISCERNIBLE]

678
00:25:08,634 --> 00:25:09,902
BONITEZ, IN MY GROUP AND SHE'S

679
00:25:09,902 --> 00:25:13,439
CARRYING THIS OUT AND A RANGE OF

680
00:25:13,439 --> 00:25:15,074
BACKGROUNDS A HERCULEAN EFFORT,

681
00:25:15,074 --> 00:25:16,375
AND LOOKING AT CELLULAR

682
00:25:16,375 --> 00:25:18,711
RESPONSES TO TOP 2 AND PROVIDING

683
00:25:18,711 --> 00:25:19,745
A GENETIC INTERACTION MAP AND

684
00:25:19,745 --> 00:25:21,480
THIS IS JUST 1 OF THOSE EXAMPLES

685
00:25:21,480 --> 00:25:23,449
OF THESE KREENS OR 2 OF THEZ

686
00:25:23,449 --> 00:25:25,217
KREENS AND WE'RE PLOTTING THEY

687
00:25:25,217 --> 00:25:27,720
RESULTS OF A GENOME WIDE SCREEN

688
00:25:27,720 --> 00:25:30,189
IN THE WILD-TYPE CELLS IN THE

689
00:25:30,189 --> 00:25:32,925
ACCESS COMPARED TO NFR 451

690
00:25:32,925 --> 00:25:34,760
BACKGROUND AND I HOPE YOU AGREE

691
00:25:34,760 --> 00:25:36,528
WITH ME, IT'S A STRIKING FINDING

692
00:25:36,528 --> 00:25:38,897
HERE THAT IT'S DROPPING OUT

693
00:25:38,897 --> 00:25:40,766
QUITE STRONGLY IN THE WILD-TYPE

694
00:25:40,766 --> 00:25:42,401
SETTING BUT BASICALLY IS SITTING

695
00:25:42,401 --> 00:25:45,504
ON THE 0 LINE IN THAT 451.

696
00:25:45,504 --> 00:25:51,510
SO IT'S WORKING DOWN STREAM OF

697
00:25:51,510 --> 00:25:53,746
ZNF451, OR IT'S EPISTATIC TO

698
00:25:53,746 --> 00:25:59,151
RAD54 L2 IN TERMS OF THIS

699
00:25:59,151 --> 00:25:59,418
PHENOTYPE.

700
00:25:59,418 --> 00:26:03,522
SO IN TERMS OF TRYING TO FIGURE

701
00:26:03,522 --> 00:26:06,158
OUT PRESLICELY WHAT RAD54 L2 WAS

702
00:26:06,158 --> 00:26:08,794
DOING WE WERE BENEFIT FRIDAY A

703
00:26:08,794 --> 00:26:10,295
COLLABORATION WITH JOHN AND

704
00:26:10,295 --> 00:26:14,533
KAREN NITRIC  NITRIC OXIDE--KNIT

705
00:26:14,533 --> 00:26:16,835
THE UNIVERSITY OF ILLINOIS ROCK

706
00:26:16,835 --> 00:26:17,102
FORD.

707
00:26:17,102 --> 00:26:18,303
AND JOHN'S LAB TOGETHER WITH US

708
00:26:18,303 --> 00:26:20,739
CARRIED OUT A SET OF ASSAYS ON

709
00:26:20,739 --> 00:26:22,040
OUR CELLS INCLUDING DUST ASSAYS

710
00:26:22,040 --> 00:26:23,509
WHICH ARE WELL ESTABLISHED

711
00:26:23,509 --> 00:26:27,179
ASSAYS LOOKING FOR BASICALLY DNA

712
00:26:27,179 --> 00:26:28,580
PROTEIN CROSS LINKS THROUGH

713
00:26:28,580 --> 00:26:30,315
WESTERN BLOT APPROACH AND THIS

714
00:26:30,315 --> 00:26:31,717
ALLOWED US TO RECOGNIZE THAT

715
00:26:31,717 --> 00:26:35,654
COMPARED TO CONTROL CELLS RAD54

716
00:26:35,654 --> 00:26:39,258
L2 KNOCK OUT CELLS HAVE HIGHER

717
00:26:39,258 --> 00:26:40,592
LEVELS OF SUMO RELATED PROTEINS

718
00:26:40,592 --> 00:26:45,397
AS WELL AS TOP 2 PROTEINS ON

719
00:26:45,397 --> 00:26:46,899
CHROMATIN, ON DNA,

720
00:26:46,899 --> 00:26:49,568
CONVALENTINEDDENTLY ATTACHED TO

721
00:26:49,568 --> 00:26:50,335
D NA CONTROLS.

722
00:26:50,335 --> 00:26:52,704
THIS IS SOMEHOW SAYING THAT THE

723
00:26:52,704 --> 00:26:54,773
PRESENCE OF RAD54 L2 IS

724
00:26:54,773 --> 00:26:55,574
COUNTERING THESE COMPLEXES AND

725
00:26:55,574 --> 00:26:58,243
THAT COULD BE VENTING THE

726
00:26:58,243 --> 00:27:04,516
FORMATION OR PROMOTING THEIR

727
00:27:04,516 --> 00:27:04,750
REPAIR.

728
00:27:04,750 --> 00:27:05,350
DISTINGUISHING THOSE 2

729
00:27:05,350 --> 00:27:06,618
POSSIBILITIES WE CARRIED OUT A

730
00:27:06,618 --> 00:27:08,153
RANGE OF STUDIES SHOWING THEM ON

731
00:27:08,153 --> 00:27:09,988
THE RIGHT, AND WHAT WE'RE

732
00:27:09,988 --> 00:27:11,957
PLOTTING HERE ACROSS A RANGE OF

733
00:27:11,957 --> 00:27:13,258
GENETIC BACKGROUNDS WITH OR

734
00:27:13,258 --> 00:27:14,793
WITHOUT THE TOPOCIDE TREATMENT

735
00:27:14,793 --> 00:27:16,261
OR TOPOCIDE TREATMENT AND

736
00:27:16,261 --> 00:27:25,604
RECOVERY, ARE GAMMA H 2 X

737
00:27:25,604 --> 00:27:26,071
 2 FOCI,

738
00:27:26,071 --> 00:27:27,706
AND THIS IS A DOUBLE STRAND

739
00:27:27,706 --> 00:27:31,510
BREAK MARKER BUT THIS WILL ONLY

740
00:27:31,510 --> 00:27:34,146
TAKE PLACE WHEN YOU GENERATE AN

741
00:27:34,146 --> 00:27:36,281
ACCESSIBLE DOUBLE STRAND BREAK,

742
00:27:36,281 --> 00:27:39,251
SO GAMMA H2 X IS NOT FORMED WHEN

743
00:27:39,251 --> 00:27:42,387
YOU HAVE A COVALENTLY ATTACHED

744
00:27:42,387 --> 00:27:43,455
TOP 2 CLEAVAGE COMPLEX, THIS

745
00:27:43,455 --> 00:27:45,557
WILL BE FORMED WHEN THE TOP 2

746
00:27:45,557 --> 00:27:48,093
GETS CLEAVED OFF, THE BREAK CAN

747
00:27:48,093 --> 00:27:48,927
BE DIRECTLY RECOGNIZED BY

748
00:27:48,927 --> 00:27:51,396
MEMBERS OF THE ATM, APR, AND THE

749
00:27:51,396 --> 00:27:53,699
PK FAMILY AND LEAD TO

750
00:27:53,699 --> 00:27:55,000
PHOSPHORYLATION OF H2 AX UNDER

751
00:27:55,000 --> 00:27:55,834
THE MARKERS.

752
00:27:55,834 --> 00:27:57,536
SO JUST TO BRIEFLY TAKE YOU

753
00:27:57,536 --> 00:27:59,938
THROUGH THIS, IN A CONTROL

754
00:27:59,938 --> 00:28:02,908
SETTING I'VE SHOWN HERE IN

755
00:28:02,908 --> 00:28:05,510
PURPLE, WE CLEARLY GET GAMMA H2

756
00:28:05,510 --> 00:28:07,045
X INDUCTION AFTER TREATMENT, THE

757
00:28:07,045 --> 00:28:08,814
TOPOCIDE AND AS WE MOVE THROUGH

758
00:28:08,814 --> 00:28:11,216
THE RECOVERY PERIOD, THESE ARE

759
00:28:11,216 --> 00:28:12,017
LARGELY REPAIRED.

760
00:28:12,017 --> 00:28:18,056
IN THE ABSENCE OF RAD54, AND

761
00:28:18,056 --> 00:28:20,726
RAD54 L2, WE GET HIGH LEVELS OF

762
00:28:20,726 --> 00:28:22,928
GAMMA RAY H2 X BUT THEY'RE

763
00:28:22,928 --> 00:28:24,997
REPAIRED OVER SEBS QUEBT QUENT

764
00:28:24,997 --> 00:28:26,865
PERIODS OF TIME ONCE WE REMOVE

765
00:28:26,865 --> 00:28:27,432
THIS TOPOCIDE.

766
00:28:27,432 --> 00:28:31,536
SO THIS IS SAYING TO US THAT

767
00:28:31,536 --> 00:28:33,071
WITHOUT RAD54 L2, THESE BREAKS

768
00:28:33,071 --> 00:28:34,439
CAN BE REPAIRED BUT SOMEHOW

769
00:28:34,439 --> 00:28:37,542
THEY'RE A GREATER NUMBER OF

770
00:28:37,542 --> 00:28:40,279
GAMMA H2 X FOCI BEING PRODUCED

771
00:28:40,279 --> 00:28:42,147
IN THIS BACKGROUND COMPARED TO

772
00:28:42,147 --> 00:28:43,815
WILD-TYPE, SO THAT MUST MEAN

773
00:28:43,815 --> 00:28:46,451
THERE'S A GREATER AMOUNT OF TOP

774
00:28:46,451 --> 00:28:48,720
2 DAMAGE AND PROCESSING TAKING

775
00:28:48,720 --> 00:28:49,154
PLACE.

776
00:28:49,154 --> 00:28:51,256
WE GO TO A DIFFERENT SITUATION

777
00:28:51,256 --> 00:28:52,758
IN THE TDP 2 KNOCK OUT HERE

778
00:28:52,758 --> 00:28:56,295
SHOWN IN TURQUOISE WHERE WE GET

779
00:28:56,295 --> 00:28:58,664
THE HIGHER, SOMEWHAT HIGHER

780
00:28:58,664 --> 00:29:03,435
LEVEL OF GAMMA H2 X FOCI BUT

781
00:29:03,435 --> 00:29:04,603
THESE EXIST BECAUSE THEY'RE NOT

782
00:29:04,603 --> 00:29:06,371
ABLE TO BE REPAIRED AND FINALLY

783
00:29:06,371 --> 00:29:07,773
IN ORANGE WE GET THE GREATER

784
00:29:07,773 --> 00:29:12,277
NUMBER OF DOUBLE STRAND BREAKS

785
00:29:12,277 --> 00:29:13,779
AS TRIGGERING GRAMA RAY HTWORKS

786
00:29:13,779 --> 00:29:15,514
X AND THEY'RE NOT COMPARED.

787
00:29:15,514 --> 00:29:18,817
SO HOW CAN WE PUT THESE TOGETHER

788
00:29:18,817 --> 00:29:22,154
IN A MODEL.

789
00:29:22,154 --> 00:29:24,556
SO OUR DATA INDICATE THE RAD54

790
00:29:24,556 --> 00:29:28,093
L2 ISHT ACT WITH THE TOP 2

791
00:29:28,093 --> 00:29:32,297
ALFARIN AND TOP 2 BETA, AND

792
00:29:32,297 --> 00:29:33,665
THEREBY THE SIMULATED VERSIONS

793
00:29:33,665 --> 00:29:37,502
OF THIS PROTEIN AS IN ZNF451 AND

794
00:29:37,502 --> 00:29:39,071
THAT SIMULATION, FUNNELS INTO

795
00:29:39,071 --> 00:29:45,610
THE TDP 2 PATHWAY, FOR THE TOP 2

796
00:29:45,610 --> 00:29:48,547
CLEAVAGE COMPACT REPAIR BUT ALSO

797
00:29:48,547 --> 00:29:50,148
IMPACTS ON RAD54 L2 ACTIVITIES

798
00:29:50,148 --> 00:29:53,585
AND OUR CURRENT MODEL IS THAT

799
00:29:53,585 --> 00:29:56,254
RAD 54 L2 ACTIVITY HELPS TO

800
00:29:56,254 --> 00:29:58,290
RESOLVE TOP 2 CLEAVAGE COMPLEXES

801
00:29:58,290 --> 00:29:59,891
AND PREVENT DOUBLE STRAND BREAK

802
00:29:59,891 --> 00:30:00,292
EXPOSURE.

803
00:30:00,292 --> 00:30:04,930
WHAT WE MEAN BY THAT S&P
THAT

804
00:30:04,930 --> 00:30:08,133
TOGETHER WITH ZNF451, WE BELIEVE

805
00:30:08,133 --> 00:30:11,236
THAT THE RAD54 L2, AND ITS

806
00:30:11,236 --> 00:30:13,872
SOCKED D NA HELIX CASE

807
00:30:13,872 --> 00:30:15,173
ACTIVITIES EITHER COUNTERACTS

808
00:30:15,173 --> 00:30:15,941
THE INITIAL ASSOCIATIONS WITH

809
00:30:15,941 --> 00:30:19,211
TOP 2 AND THE DNA OR WE RATHER

810
00:30:19,211 --> 00:30:21,613
THINK, IT ACTUALLY CAN RECOGNIZE

811
00:30:21,613 --> 00:30:25,050
THESE CLEAVAGE COMPLEXES AND

812
00:30:25,050 --> 00:30:26,118
SOMEHOW ALLOW RNA, ALLOW TOP 2

813
00:30:26,118 --> 00:30:27,819
TO BACK UP THAT INITIAL REACTION

814
00:30:27,819 --> 00:30:29,921
AND BE RELEASED FROM THE DNA,

815
00:30:29,921 --> 00:30:31,089
THEREFORE NOT GENERATING THE

816
00:30:31,089 --> 00:30:32,491
DOUBLE STRAND BREAKS THAT NEED

817
00:30:32,491 --> 00:30:36,461
TO BE REPAIRED THROUGH THE

818
00:30:36,461 --> 00:30:37,195
CANNONICLE REPAIR PATHWAYS AND

819
00:30:37,195 --> 00:30:39,664
THIS IS SOMETHING WE'RE NOW

820
00:30:39,664 --> 00:30:43,235
FURTHER EXPLORING IN A RANGE OF

821
00:30:43,235 --> 00:30:44,369
WAYS INCLUDING THROUGH BY

822
00:30:44,369 --> 00:30:44,903
CHEMICAL STUDIES.

823
00:30:44,903 --> 00:30:46,171
AND I WILL POINT OUT THAT OUR

824
00:30:46,171 --> 00:30:48,607
WORK AND THE RELATED WORK FROM

825
00:30:48,607 --> 00:30:50,308
CHEN AND COLLEAGUES HAS BEEN

826
00:30:50,308 --> 00:30:52,444
PUBLISHED IN SCIENCE ADVANCES.

827
00:30:52,444 --> 00:30:55,814
JUST TO ROUND OFF THIS FIRST OF

828
00:30:55,814 --> 00:30:57,449
MY 2 PARTS OF MY TALK TODAY, I

829
00:30:57,449 --> 00:30:58,717
WANT TO GO BACK TO THIS

830
00:30:58,717 --> 00:31:01,219
OBSERVATION HERE THAT WHEN WE

831
00:31:01,219 --> 00:31:03,588
PUT OR WHEN WE OVEREXPRESS RAD

832
00:31:03,588 --> 00:31:05,257
54 IN WILD-TYPE CELLS WE GET

833
00:31:05,257 --> 00:31:07,092
MORE RESISTANCE AND IN THAT

834
00:31:07,092 --> 00:31:09,027
REGARD, IT'S INTERESTING THAT

835
00:31:09,027 --> 00:31:11,863
THERE ARE DATA IN THE LITERATURE

836
00:31:11,863 --> 00:31:14,099
INDICATING THAT THE EXPRESSION

837
00:31:14,099 --> 00:31:16,368
OF RAD54 L2 IMPACTING ON

838
00:31:16,368 --> 00:31:18,537
PROGRESSION FREE SURVIVAL IN

839
00:31:18,537 --> 00:31:20,272
THIS PARTICULAR AML COHORT.

840
00:31:20,272 --> 00:31:24,176
AND SO, THIS SUGGESTS THAT RAD54

841
00:31:24,176 --> 00:31:25,844
L2 ACTIVITY MAY BE A MECHANISM

842
00:31:25,844 --> 00:31:28,280
OF CANCER CELL RESISTANCE IN

843
00:31:28,280 --> 00:31:30,348
CLINICAL SETTINGS, AND

844
00:31:30,348 --> 00:31:32,551
HIGHLIGHTS RAD54 L2 AS A POETIC

845
00:31:32,551 --> 00:31:36,688
TEBTIAL ANTICANCER DRUG TARGET.

846
00:31:36,688 --> 00:31:38,557
ANOTHER POTENTIAL MEDICAL ANGLE,

847
00:31:38,557 --> 00:31:40,725
WE WERE LED TO WAS THROUGH THE

848
00:31:40,725 --> 00:31:42,227
REALIZATION THAT IT'S BEEN

849
00:31:42,227 --> 00:31:44,162
ESTABLISHED BY KEITH AND

850
00:31:44,162 --> 00:31:45,363
COLLEAGUES THAT TDP2 MUTATIONS

851
00:31:45,363 --> 00:31:46,631
WHICH YOU WILL RECALL IS PLAYING

852
00:31:46,631 --> 00:31:50,335
A KEY ROLE IN TOP 2 CLEAVAGE

853
00:31:50,335 --> 00:31:55,273
COMPLEX REPAIR, IT INHERITED

854
00:31:55,273 --> 00:31:56,908
MUTATIONS IN TDP2, CAUSE

855
00:31:56,908 --> 00:31:57,976
NEUROLOGICAL DISEASE AND WE

856
00:31:57,976 --> 00:32:00,512
THEREFORE WONDER WHETHER OR NOT

857
00:32:00,512 --> 00:32:02,614
RAD54 L2 MUTATIONS MIGHT ALSO

858
00:32:02,614 --> 00:32:04,816
CAUSE SIMILAR DISEASE.

859
00:32:04,816 --> 00:32:06,184
NOW IT'S FAIRLY EARLY DAYSA THE

860
00:32:06,184 --> 00:32:07,586
THIS STAGE BUT WE HAVE

861
00:32:07,586 --> 00:32:08,954
COLLABORATED WITH PETER

862
00:32:08,954 --> 00:32:11,089
[INDISCERNIBLE] AND IRENEA

863
00:32:11,089 --> 00:32:12,390
[INDISCERNIBLE] IN THE

864
00:32:12,390 --> 00:32:14,292
NETHERLANDS WHO HAVE BEEN

865
00:32:14,292 --> 00:32:17,262
WORKING WITH A PATIENT

866
00:32:17,262 --> 00:32:19,664
PRESENTING FAIRLY EARLY ON IN

867
00:32:19,664 --> 00:32:20,932
DEVELOPMENT WITH SEIZURES,

868
00:32:20,932 --> 00:32:22,534
DEVELOPMENTAL DELAY AND

869
00:32:22,534 --> 00:32:25,537
INTELLECTUAL DISABILITY, WITH A

870
00:32:25,537 --> 00:32:30,709
DE NOVO HETEROZYGOUS  VARIANT IN

871
00:32:30,709 --> 00:32:31,877
RAD54 L2.

872
00:32:31,877 --> 00:32:35,747
AND A GRADUATE STUDENT IN MY LAB

873
00:32:35,747 --> 00:32:36,581
CHAD [INDISCERNIBLE] HAS BEEN

874
00:32:36,581 --> 00:32:39,484
ABLE TO MODEL THOSE IN HUMAN RP1

875
00:32:39,484 --> 00:32:40,886
CELLS THROUGH CRISPR GENETIC

876
00:32:40,886 --> 00:32:42,821
ENGINEERING AND BASICALLY SEEING

877
00:32:42,821 --> 00:32:46,858
THAT THAT MUTATION DOES IMPACT

878
00:32:46,858 --> 00:32:48,026
ON ATOPOCIDE SENSITIVITY.

879
00:32:48,026 --> 00:32:50,595
SUGGESTING THAT THE INTELLECTUAL

880
00:32:50,595 --> 00:32:51,863
AND OTHER PATIENT'S ISSUES MAY

881
00:32:51,863 --> 00:33:00,038
BE DRIVEN BY THAT VARIANT.

882
00:33:00,038 --> 00:33:01,139
OKAY, SO WHAT I'VE BEEN TALKING

883
00:33:01,139 --> 00:33:03,375
ABOUT SO FAR IS ABOUT DNA

884
00:33:03,375 --> 00:33:05,043
PROTEIN CROSS LINKS INVOLVING

885
00:33:05,043 --> 00:33:07,479
TOP O ICE SOM RACE 2, 1 CAN FORM

886
00:33:07,479 --> 00:33:08,580
THESE AND THAT'S A WHOLE OTHER

887
00:33:08,580 --> 00:33:10,115
STORY ABOUT YOU WHAT I WILL TALK

888
00:33:10,115 --> 00:33:11,416
ABOUT FOR THE REMAINDER OF MY

889
00:33:11,416 --> 00:33:13,418
TALK NOW IS TO FOCUS ON OTHER

890
00:33:13,418 --> 00:33:15,720
TYPES OF DNA PROTEIN CROSS LINKS

891
00:33:15,720 --> 00:33:17,589
THAT ARE GENERATED BY THINGS

892
00:33:17,589 --> 00:33:21,393
SUCH AS REACTIVE METABOLITES

893
00:33:21,393 --> 00:33:23,461
SUCH AS FORMALDEHYDE AND AGENTS

894
00:33:23,461 --> 00:33:28,433
SUCH AS 5 AZADECOXY SWREEN, ALSO

895
00:33:28,433 --> 00:33:30,302
KNOWN AS DECIDE O BEAN A EABT

896
00:33:30,302 --> 00:33:30,869
CANCER DRUG.

897
00:33:30,869 --> 00:33:32,537
NOW WORK OVER THE YEARS HAVE

898
00:33:32,537 --> 00:33:35,473
SHOWN THAT THESE OTHER CROSS

899
00:33:35,473 --> 00:33:38,310
LINKS WHICH CAN ARISE WITH A

900
00:33:38,310 --> 00:33:40,378
RANGE OF PROTEINS, HISTONES RNA

901
00:33:40,378 --> 00:33:42,447
PROTEINS AND WHATEVER, THE KEY

902
00:33:42,447 --> 00:33:43,848
PRINCIPLES OF REPAIR IN THIS

903
00:33:43,848 --> 00:33:47,085
SCENARIO IS THAT THE DPCs NEED

904
00:33:47,085 --> 00:33:48,620
TO BE DEBULKED, EATEN AWAY IF

905
00:33:48,620 --> 00:33:52,023
YOU LIKE BY SPECIALIZED DTP

906
00:33:52,023 --> 00:33:53,992
PROTEASE SUCH AS THE SPARTAN

907
00:33:53,992 --> 00:33:55,293
ENZYME AND THE PROTEOZOME.

908
00:33:55,293 --> 00:33:59,931
LIKE THE STORY I'VE JUST BEEN

909
00:33:59,931 --> 00:34:03,935
TALKING WITH YOU, THIS DC REPAIR

910
00:34:03,935 --> 00:34:11,343
IS ALSO WITH UBIQUITINNATION AND

911
00:34:11,343 --> 00:34:16,114
SUMO-ELATION, AS WELL AS GLOBAL

912
00:34:16,114 --> 00:34:16,781
GENOME REPAIR MECHANISMS.

913
00:34:16,781 --> 00:34:18,550
ONE THING WE THOUGHT ABOUT A FEW

914
00:34:18,550 --> 00:34:20,819
YEARS AGO NOW WAS WHETHER THERE

915
00:34:20,819 --> 00:34:25,423
MIGHT BE A TRANSCRIPTION COUPLED

916
00:34:25,423 --> 00:34:26,858
VERSION OF DMA CROSS LISTEN

917
00:34:26,858 --> 00:34:29,427
REPAIR, THAT WILL MAKE INTUITIVE

918
00:34:29,427 --> 00:34:31,129
SENSE, DNA CROSS LINKS DON'T

919
00:34:31,129 --> 00:34:33,498
JUST INTERFERE WITH REPLICATION.

920
00:34:33,498 --> 00:34:35,767
THEY ALSO INTERFERE WITH

921
00:34:35,767 --> 00:34:36,101
TRANSCRIPTION.

922
00:34:36,101 --> 00:34:38,870
SO, 1 COULD IMAGINE THAT SUCH A

923
00:34:38,870 --> 00:34:40,372
MECHANISM WOULD HAVE ARISEN

924
00:34:40,372 --> 00:34:42,641
DURING EVOLUTION.

925
00:34:42,641 --> 00:34:45,443
SO NOW I'M GOING TO GO THROUGH A

926
00:34:45,443 --> 00:34:49,114
UNPUBLISHED BUT SOON TO BE

927
00:34:49,114 --> 00:34:50,115
PUBLISHED STORY PIONEERS BY

928
00:34:50,115 --> 00:34:52,817
THESE INDIVIDUALS, 2 STORIES

929
00:34:52,817 --> 00:34:56,988
ACTUALLY, BY THESE POST DOCS AND

930
00:34:56,988 --> 00:34:58,023
AND A GRADUATE STUDENT.

931
00:34:58,023 --> 00:35:00,859
THIS WORK IS ALSO BENEFITED FROM

932
00:35:00,859 --> 00:35:02,027
A FANTASTIC COLLABORATION, 1 OF

933
00:35:02,027 --> 00:35:03,828
SEVERAL COLLABORATIONS THAT

934
00:35:03,828 --> 00:35:05,964
WE'VE GOT WITH JULIAN STINGLER'S

935
00:35:05,964 --> 00:35:10,502
GROUP IN  MUNICH AND THAT HAS

936
00:35:10,502 --> 00:35:13,538
BENEFITED WITH OUR INTERACTIONS

937
00:35:13,538 --> 00:35:14,673
WITH [INDISCERNIBLE] IN

938
00:35:14,673 --> 00:35:15,106
[INDISCERNIBLE].

939
00:35:15,106 --> 00:35:19,244
SO THE BRAIN WAVE THAT CHRIS

940
00:35:19,244 --> 00:35:22,347
CARNIE A FEW WE'RES AGO WAS TO

941
00:35:22,347 --> 00:35:23,948
REDUCE THE DPCs EXPERIMENTALLY

942
00:35:23,948 --> 00:35:26,484
WITH 2 TYPES OF AGENTS AND THEN

943
00:35:26,484 --> 00:35:28,586
COMPARE AND CONTRAST THE CRISPR

944
00:35:28,586 --> 00:35:29,988
SCREEN OUTPUTS IMENERATED BY

945
00:35:29,988 --> 00:35:30,789
THOSE 2 AGENTS.

946
00:35:30,789 --> 00:35:32,991
SO THE AGENTS THAT THEY USE WERE

947
00:35:32,991 --> 00:35:34,292
FORMALDEHYDE WHICH IS A POTENT

948
00:35:34,292 --> 00:35:37,262
STATE AND THEN THE RAPID DPC

949
00:35:37,262 --> 00:35:37,696
INDUCER.

950
00:35:37,696 --> 00:35:40,332
IT'S CELL PSYCHE WILL BLIND, IT

951
00:35:40,332 --> 00:35:41,733
OPERATES ACROSS THE CELL CYCLE.

952
00:35:41,733 --> 00:35:44,336
IMPORTANTLY IT'S A NATURALLY

953
00:35:44,336 --> 00:35:46,304
OCCURRING METABOLITE.

954
00:35:46,304 --> 00:35:48,973
SO FORMALDEHYDE DNA DAMAGE AND A

955
00:35:48,973 --> 00:35:49,974
REAL PHYSIOLOGICAL RELEVANCE.

956
00:35:49,974 --> 00:35:52,210
INITIALLY IF YOU LIKE,

957
00:35:52,210 --> 00:35:53,445
EXPERIMENTAL ISSUE WITH

958
00:35:53,445 --> 00:35:55,280
FORMALDEHYDE IT CAN GIVE RISE TO

959
00:35:55,280 --> 00:35:57,382
A RAGER OF OTHER ADDUCTS SO IF

960
00:35:57,382 --> 00:36:00,085
YOU SEE A RESPONSE TO

961
00:36:00,085 --> 00:36:01,853
FORMALDEHYDE IT'S DIFFICULT TO

962
00:36:01,853 --> 00:36:03,188
DISENTANGLE WHICH LESIONS ARE

963
00:36:03,188 --> 00:36:04,556
GENERATING THE PHENOTYPE EMPLOY

964
00:36:04,556 --> 00:36:06,391
THE OTHER 18th THAT CHRIS

965
00:36:06,391 --> 00:36:09,794
FOCUSED ON IS 5 AINTRRKS-2 PRIME

966
00:36:09,794 --> 00:36:15,367
DEOXYICIDE O DEAN, OR 5

967
00:36:15,367 --> 00:36:17,202
DECYTOBIEN, THIS COMPOUND SHOWN

968
00:36:17,202 --> 00:36:19,738
HERE IS INTO DNA, AND

969
00:36:19,738 --> 00:36:20,705
IMPORTANTLY, WHEN IT'S

970
00:36:20,705 --> 00:36:26,044
INCORPORATED AT SITES OF DNA

971
00:36:26,044 --> 00:36:29,013
METHYLATION, AT CPGs, WHEN THE

972
00:36:29,013 --> 00:36:29,614
MAINTENANCE METHYLTRANSFERASE

973
00:36:29,614 --> 00:36:32,317
COMES IN TO TRY AND BASICALLY

974
00:36:32,317 --> 00:36:35,987
METHALATE THIS SITE TO MAKE IT

975
00:36:35,987 --> 00:36:38,323
SYMMETRICAL, IT GETS STUCK, THE

976
00:36:38,323 --> 00:36:39,591
CATALYTIC MECHANISM IS DISRUPTED

977
00:36:39,591 --> 00:36:43,628
AND THAT LEADS TO DNMT1,

978
00:36:43,628 --> 00:36:45,997
BECOMING A COVALENTLY ATTACHED

979
00:36:45,997 --> 00:36:48,199
DPC IN THIS SCENARIO WHICH IS

980
00:36:48,199 --> 00:36:51,369
OBVIOUSLY GOING TO GIVE PROBLEMS

981
00:36:51,369 --> 00:36:52,203
TO THE CELL.

982
00:36:52,203 --> 00:36:53,037
EXPERIMENTALLY YOU HAVE TO

983
00:36:53,037 --> 00:36:54,873
REALIZE THIS IS ONLY GOING TO

984
00:36:54,873 --> 00:36:57,609
TAKE PLACE IN S-PHASE AND IT

985
00:36:57,609 --> 00:36:58,710
WILL ONLY REALLY GENERATE THOSE

986
00:36:58,710 --> 00:37:00,912
TYPES OF LESIONS WHERE THERE'S A

987
00:37:00,912 --> 00:37:01,780
CPG METHYLATION TAKING PLACE.

988
00:37:01,780 --> 00:37:04,416
THIS IS USED IN THE CLINIC.

989
00:37:04,416 --> 00:37:06,384
AND SO ANYTHING WE LEARN WITH

990
00:37:06,384 --> 00:37:10,188
THIS AGENT POTENTIAL HAS

991
00:37:10,188 --> 00:37:10,655
CLINICAL RELEVANCE.

992
00:37:10,655 --> 00:37:15,493
SO WHAT I'M SHOWING HERE, IS THE

993
00:37:15,493 --> 00:37:16,861
CRISPR SCREEN OUTPUTS, IN THIS

994
00:37:16,861 --> 00:37:20,165
CASE IT'S NOT A CRISPR CAS 9

995
00:37:20,165 --> 00:37:22,767
GENE, IT'S A CRISPR LIBRARY

996
00:37:22,767 --> 00:37:24,436
SCREEN CARRIED OUT BY CHRIS

997
00:37:24,436 --> 00:37:27,972
USING THE LIBRARY AND CELLS

998
00:37:27,972 --> 00:37:29,007
INITIALLY GENERATED BY

999
00:37:29,007 --> 00:37:29,641
APPROXIMATE [INDISCERNIBLE] IN

1000
00:37:29,641 --> 00:37:32,043
THE GROUP AND THIS IS IN K526

1001
00:37:32,043 --> 00:37:33,478
CELLS AND WE ARE LOOKING FOR

1002
00:37:33,478 --> 00:37:35,413
YEENS THAT WERE KNOCKED OUT GIVE

1003
00:37:35,413 --> 00:37:36,548
RISE TO SENSITIVITY TO

1004
00:37:36,548 --> 00:37:38,149
FORMALDEHYDE AND YOU CAN SEE

1005
00:37:38,149 --> 00:37:39,918
THERE'S A WHOLE RANGE OF THEM

1006
00:37:39,918 --> 00:37:41,519
THERE, WE KNOW THE KREEN WORKS

1007
00:37:41,519 --> 00:37:45,924
IMMEDIATELY BECAUSE SOME OF THE

1008
00:37:45,924 --> 00:37:48,059
STRONGEST SCREENING HITS ARE

1009
00:37:48,059 --> 00:37:52,964
THESE YEEPS AND THEIR PROTEINS

1010
00:37:52,964 --> 00:37:55,800
HERE ESD AND ADH5 AND

1011
00:37:55,800 --> 00:37:58,670
IMPORTANTLY THESE ARE INVOLVED

1012
00:37:58,670 --> 00:37:59,737
IN FORMALDEHYDE DETOXITION, SO

1013
00:37:59,737 --> 00:38:01,539
YOU KNOCK OUT WITH THESE, AND

1014
00:38:01,539 --> 00:38:03,374
YOU KNOCK OUT WITH MORE  

1015
00:38:03,374 --> 00:38:04,142
PERRIST SENT FORM ALD HIGHS

1016
00:38:04,142 --> 00:38:06,010
WHICH WILL GIVE RISE TO MORE

1017
00:38:06,010 --> 00:38:07,479
DAMAGE AND MORE CELL KILLING.

1018
00:38:07,479 --> 00:38:09,013
SO THERE ARE MANY FACTORS HERE

1019
00:38:09,013 --> 00:38:11,783
AND THIS WASN'T ALL THAT

1020
00:38:11,783 --> 00:38:12,917
SURPRISING.

1021
00:38:12,917 --> 00:38:14,419
OTHER GROUPS INCLUDING

1022
00:38:14,419 --> 00:38:14,919
[INDISCERNIBLE]'S PAPER

1023
00:38:14,919 --> 00:38:16,488
HIGHLIGHTED HERE IN THE CELL

1024
00:38:16,488 --> 00:38:18,923
PAPER, HAVE MAPPED OUT CELLULAR

1025
00:38:18,923 --> 00:38:20,925
RESPONSES TO FORMALDEHYDE AND

1026
00:38:20,925 --> 00:38:23,561
SHOWN THE ARRANGE OF PROTEINS

1027
00:38:23,561 --> 00:38:25,730
INVOLVED IN NUCLEOTO THETIC

1028
00:38:25,730 --> 00:38:27,699
EXCISION REPAIR, ANEMIA PATHWAY

1029
00:38:27,699 --> 00:38:30,168
AND OTHER PATH WAGHTS IMPACT ON

1030
00:38:30,168 --> 00:38:31,169
THE--PATHWAYS IMPACT ON THE

1031
00:38:31,169 --> 00:38:32,770
REPAIR OF THOSE LESIONS BUT ALSO

1032
00:38:32,770 --> 00:38:37,308
IT'S KNOWN BY WORKING THROUGH

1033
00:38:37,308 --> 00:38:40,512
KPATEL AND COLLEAGUES THAT

1034
00:38:40,512 --> 00:38:41,479
TRANSCRIPTIONAL STRESS TRIGGERS

1035
00:38:41,479 --> 00:38:43,481
DNA RESPONSE IN MOUSE MODELS AND

1036
00:38:43,481 --> 00:38:45,984
THAT'S PROVIDED INSIGHTS INTO

1037
00:38:45,984 --> 00:38:49,020
POTENTIAL MECHANISMS OF HUMAN

1038
00:38:49,020 --> 00:38:50,889
DISORDERS, THAT I'LL TOUCH ON A

1039
00:38:50,889 --> 00:38:51,456
BIT LATER ON.

1040
00:38:51,456 --> 00:38:53,191
BUT I THINK THINGS GET MORE

1041
00:38:53,191 --> 00:38:56,361
INTERESTING WHEN WE FOCUS

1042
00:38:56,361 --> 00:38:57,962
SPECIFICALLY ON HITS IN OUR

1043
00:38:57,962 --> 00:39:00,031
SCREEN THAT RELATE TO A BATH WAY

1044
00:39:00,031 --> 00:39:02,033
THAT'S BEEN KNOWN FOR MANY

1045
00:39:02,033 --> 00:39:05,570
YEARS, IN FACT DECADES NOW,

1046
00:39:05,570 --> 00:39:06,604
TRANSCRIPTION COUPLED NUCLEOTIDE

1047
00:39:06,604 --> 00:39:07,105
ECCISION REPAIR.

1048
00:39:07,105 --> 00:39:10,108
AS YOU BE AWARE, NUCLEOTIDE

1049
00:39:10,108 --> 00:39:11,509
EXCISION REPAIR REMOVES UV

1050
00:39:11,509 --> 00:39:14,112
INDUCED LESIONS AND OTHER BULKY

1051
00:39:14,112 --> 00:39:22,587
DNAA ADDUCTS FROM DNA AND

1052
00:39:22,587 --> 00:39:27,492
BASICALLY ENDS UP THE SINGLE

1053
00:39:27,492 --> 00:39:28,560
STRANDED REGION AND WORKING ON

1054
00:39:28,560 --> 00:39:30,061
ITS REPAIR AND LIGATION, NOW

1055
00:39:30,061 --> 00:39:35,833
MUCH WORK OVER MANY YEARS AND IT

1056
00:39:35,833 --> 00:39:39,137
WORK MISS 2 DEFICIENT ARENAS AND

1057
00:39:39,137 --> 00:39:47,912
THERE'S TRANSCRIPTION COUPLED

1058
00:39:47,912 --> 00:39:49,914
REPAIR AND AND THE TOP OF THE

1059
00:39:49,914 --> 00:39:55,053
PATHWAY IS A CSB ENCODER BY THE

1060
00:39:55,053 --> 00:39:58,056
ARCC6 GENE AND RECOGNIZING A

1061
00:39:58,056 --> 00:40:01,859
STOLED RNETWORKA POLYMERASE 2,

1062
00:40:01,859 --> 00:40:05,530
ENCOULD YOU WANTERRING A

1063
00:40:05,530 --> 00:40:06,831
NUCLEOTIDE EXCISION OF TAR

1064
00:40:06,831 --> 00:40:07,632
TARGETS.

1065
00:40:07,632 --> 00:40:16,608
THIS RELEADS TO REACCUMULATE OF

1066
00:40:16,608 --> 00:40:18,676
ERCC8 GENE, AND WHICH AMONGST

1067
00:40:18,676 --> 00:40:21,512
OTHER THINGS LEAD TO

1068
00:40:21,512 --> 00:40:25,817
UBIQUITINNATION OF THE PUB UNIT

1069
00:40:25,817 --> 00:40:29,821
OF POLYMERASE AND OTHER FACTORS

1070
00:40:29,821 --> 00:40:31,022
INCLUDING UVSSA, THE DOWN STREAM

1071
00:40:31,022 --> 00:40:33,091
OF THE ECCISION REPAIR

1072
00:40:33,091 --> 00:40:34,158
TRANSCRIPTION COUPLED EXCISION

1073
00:40:34,158 --> 00:40:36,160
REPAIR ARE MEDIATED BY FACTORS

1074
00:40:36,160 --> 00:40:41,232
SUCH AS XBA, AND THE NUCLEACES,

1075
00:40:41,232 --> 00:40:42,200
AND XBF AND XPG.

1076
00:40:42,200 --> 00:40:45,503
AND AS YOU WILL NOTE FROM THE

1077
00:40:45,503 --> 00:40:47,639
LEFT-HAND SIDE, THIS RESPONSE TO

1078
00:40:47,639 --> 00:40:49,841
FORMALDEHYDE, ESSENTIALLY ALL OF

1079
00:40:49,841 --> 00:40:52,143
THOSE TCNER HITS SO THE LESION

1080
00:40:52,143 --> 00:40:54,312
IS GENERATED FOR FORMALDEHYDE

1081
00:40:54,312 --> 00:41:00,918
AND DRIVING TOCKSITY, LARGELY

1082
00:41:00,918 --> 00:41:02,020
BEING THE EFFECTS OF COUNTER

1083
00:41:02,020 --> 00:41:04,322
ACTED BY THAT SET OF FACTORS BUT

1084
00:41:04,322 --> 00:41:09,093
WE'RE NOT SEEING XPC, DROPPING

1085
00:41:09,093 --> 00:41:12,430
OUT IN THIS SCENARIO HERE, IT'S

1086
00:41:12,430 --> 00:41:14,532
INVOLVING GENOME REPAIR BUT NOT

1087
00:41:14,532 --> 00:41:15,667
TRANSCRIPTION COUPLED NER AND

1088
00:41:15,667 --> 00:41:17,468
THIS IS WHERE IT'S INDICATING

1089
00:41:17,468 --> 00:41:21,105
THAT THE TOXICITY GENERATED BY

1090
00:41:21,105 --> 00:41:22,073
FORMALDEHYDE IN THIS SCENARIO

1091
00:41:22,073 --> 00:41:23,675
HERE WILL BE LARGELY THROUGH THE

1092
00:41:23,675 --> 00:41:27,278
ACTIONS ON THE TRANSCRIPTIONALLY

1093
00:41:27,278 --> 00:41:27,712
ACTIVE GENES.

1094
00:41:27,712 --> 00:41:28,646
SO WHAT I'M PLOTTING ON THE

1095
00:41:28,646 --> 00:41:33,985
RIGHT HERE ARE THE RESULTS OF

1096
00:41:33,985 --> 00:41:37,055
CHRIS' CRISPR I-SCREEN WITH

1097
00:41:37,055 --> 00:41:38,256
CELLS TREATED WITH DECYTOBIN,

1098
00:41:38,256 --> 00:41:40,725
AND YOU WILL SEE A RAINCHL OF

1099
00:41:40,725 --> 00:41:41,526
POSITIVE CONTROLS.

1100
00:41:41,526 --> 00:41:44,529
THESE FACTORS SHOWN IN PURPLE.

1101
00:41:44,529 --> 00:41:46,864
SLC 29 A 1 ENCODES A TRANSPORTER

1102
00:41:46,864 --> 00:41:50,168
WHICH IS NECESSARY FOR

1103
00:41:50,168 --> 00:41:52,370
INCORPORATION OR OF THIS DRUG,

1104
00:41:52,370 --> 00:41:53,771
INTO CELLS, OBVIOUSLY IT'S NOT

1105
00:41:53,771 --> 00:41:55,540
GOING TO HAPPENOT IMPACT OF THE

1106
00:41:55,540 --> 00:41:57,408
CELL, IF IT CAN'T GET INTO THE

1107
00:41:57,408 --> 00:42:00,645
CELL AND DCK AND CMP K ARE

1108
00:42:00,645 --> 00:42:02,480
KINASES THAT MEDIATE

1109
00:42:02,480 --> 00:42:05,183
PHOSPHORYLATION OF THE CYTOBIN

1110
00:42:05,183 --> 00:42:05,917
INTO A TRANSPHOSPHATE VERSION

1111
00:42:05,917 --> 00:42:08,820
THAT, LOWS IT TO BE INCORPORATED

1112
00:42:08,820 --> 00:42:11,089
IN DNA AND MEDIATE ITS TOXICITY.

1113
00:42:11,089 --> 00:42:13,791
YOU CAN ALSO SEE THAT IN THIS

1114
00:42:13,791 --> 00:42:16,527
PLOT, I HIGHLIGHTED THE VARIOUS

1115
00:42:16,527 --> 00:42:18,162
NUCLEOTIDE EXCISION REPAIR YEENS

1116
00:42:18,162 --> 00:42:19,564
AND WITH STRIKING OBSERVATION IS

1117
00:42:19,564 --> 00:42:27,705
THAT WE CAN SEE CTSA AND CSB ARE

1118
00:42:27,705 --> 00:42:31,642
DROP OUTS IN THE--IN BOTH

1119
00:42:31,642 --> 00:42:32,443
SCENARIOS, WHEREAS--SORRY, I'VE

1120
00:42:32,443 --> 00:42:34,979
JUST BEEN INVITED TO THIS

1121
00:42:34,979 --> 00:42:35,346
SEMINAR.

1122
00:42:35,346 --> 00:42:36,514
--WHEREAS THE OTHER FABLGHTORS

1123
00:42:36,514 --> 00:42:38,616
THAT ARE GETTING YOU--FACTORS

1124
00:42:38,616 --> 00:42:40,651
THAT GETTING UP AS HITS IN THE

1125
00:42:40,651 --> 00:42:43,020
FORMALDEHYDE SCREENS ARE NOT.

1126
00:42:43,020 --> 00:42:44,322
SO KNOWLEDGING ABOUT THIS MORE,

1127
00:42:44,322 --> 00:42:46,491
WE DECIDED TO GO AHEAD AND

1128
00:42:46,491 --> 00:42:48,893
VALIDATE THESE SCREEN HITS AND

1129
00:42:48,893 --> 00:42:50,261
THESE DO VALIDATE SO YOU CAN SEE

1130
00:42:50,261 --> 00:42:51,496
ON THE LEFT-HAND SIDE HERE THAT

1131
00:42:51,496 --> 00:42:54,031
IF WE COMPARE TO WILD'RE

1132
00:42:54,031 --> 00:42:55,767
WILD-TYPE CELLS, CELLS KNOCKED

1133
00:42:55,767 --> 00:42:59,704
OUT FOR XPA OR CSB, ARE HYPER

1134
00:42:59,704 --> 00:43:01,973
SENSITIVE TO FORMALDEHYDE,

1135
00:43:01,973 --> 00:43:03,141
HOWEVER, WHILE CSB KNOCK OUT

1136
00:43:03,141 --> 00:43:07,812
CELLS ARE HYPER SENSITIVE TO 5

1137
00:43:07,812 --> 00:43:09,981
ACDC, THE EXPA KNOCK OUTS ARE

1138
00:43:09,981 --> 00:43:10,348
NOT.

1139
00:43:10,348 --> 00:43:13,818
WE GET SIMILAR RESULTS IN HAP1

1140
00:43:13,818 --> 00:43:16,354
AND ALSO IN HUMAN RBD1 CELLS.

1141
00:43:16,354 --> 00:43:19,123
SO THE DEFERENT YALE IMPACT THAT

1142
00:43:19,123 --> 00:43:20,992
WE'RE SEEING WITH DECYTOBIN AND

1143
00:43:20,992 --> 00:43:25,396
BASICALLY AT THE LEVEL OF THE

1144
00:43:25,396 --> 00:43:29,634
EARLY STAGES OF WHAT WE CALL

1145
00:43:29,634 --> 00:43:30,968
TCNER, INVOLVING TCAND CSA BUT

1146
00:43:30,968 --> 00:43:33,004
NOT THE DOWN STREAM COMPONENTS.

1147
00:43:33,004 --> 00:43:34,172
SO THROUGH STUDY IN

1148
00:43:34,172 --> 00:43:35,339
COMPLEMENTATION STUDIES IN THE

1149
00:43:35,339 --> 00:43:36,507
KNOCK OUT BACKGROUNDS WE'RE ABLE

1150
00:43:36,507 --> 00:43:40,678
TO SHOW THAT THIS FUNCTION

1151
00:43:40,678 --> 00:43:42,213
REQUIRES CSB ATP ACE ACTIVITY

1152
00:43:42,213 --> 00:43:44,782
AND WE SHOW THAT IF YOU LOSE THE

1153
00:43:44,782 --> 00:43:46,617
GLOBAL GENOME PATHWAYS THROUGH

1154
00:43:46,617 --> 00:43:49,453
KNOCKING OUT XPC, YOU SENSITIZE

1155
00:43:49,453 --> 00:43:52,490
CELLS TO UV LIGHT BUT NOT TO

1156
00:43:52,490 --> 00:43:54,292
FORMALDEHYDE OR RAISER OR THE

1157
00:43:54,292 --> 00:43:56,494
CYTOBIEN, AND EVEN IF YOU KNOCK

1158
00:43:56,494 --> 00:44:00,598
OUT XPA TOGETHER WITH XPC ARE,

1159
00:44:00,598 --> 00:44:02,633
YOU GET FORMALDEHYDE SENSITIVITY

1160
00:44:02,633 --> 00:44:09,040
BUT NOT DESIT BIEN SENSITIVITY.

1161
00:44:09,040 --> 00:44:11,342
FOR SPARTAN ON RF4, AND OR

1162
00:44:11,342 --> 00:44:16,747
EXPRESSION OF A PART AN DELTA

1163
00:44:16,747 --> 00:44:17,815
CTERMINAL TRUNK TERMINATED

1164
00:44:17,815 --> 00:44:18,749
CONSTRUCT WHICH IMPAIRS ITS

1165
00:44:18,749 --> 00:44:22,186
ACTIVITY WERE TO SHOW THAT CSB

1166
00:44:22,186 --> 00:44:23,554
IS COMPARING DPC COLERANCE

1167
00:44:23,554 --> 00:44:25,923
THROUGH A PATHWAY THAT ACTS IN

1168
00:44:25,923 --> 00:44:27,525
PARALLEL NOT IN THE SAME PATHWAY

1169
00:44:27,525 --> 00:44:30,862
BUT IN PARALLEL WITH ESTABLISHED

1170
00:44:30,862 --> 00:44:36,801
SPARTAN RF4 DEPENDENT

1171
00:44:36,801 --> 00:44:37,101
MECHANISMS.

1172
00:44:37,101 --> 00:44:38,636
SO WHAT WE THINK IS GOING ON

1173
00:44:38,636 --> 00:44:42,607
HERE IS THAT THE CROSS LINKS

1174
00:44:42,607 --> 00:44:46,244
AREN'T REQUIRING SEN ILLEGALSEN

1175
00:44:46,244 --> 00:44:48,880
CESS ENSEL OF CSA, AND CSB, AND

1176
00:44:48,880 --> 00:44:53,517
BUT NOT CONNECT WIDE NER.

1177
00:44:53,517 --> 00:44:55,620
NO IT'S SHOWN THROUGH CERTAIN

1178
00:44:55,620 --> 00:44:56,921
STUDIES THAT RNA POLYMERASES ARE

1179
00:44:56,921 --> 00:44:59,357
STOLE BY THE ACTIONS OF THE

1180
00:44:59,357 --> 00:45:02,226
DPCs AND IT'S NOT BEING

1181
00:45:02,226 --> 00:45:03,494
EXPORTS VIVO, BUT WHAT I'M

1182
00:45:03,494 --> 00:45:05,529
SHOWING YOU THROUGH THESE

1183
00:45:05,529 --> 00:45:07,331
STUDIES HERE WHICH WERE CARRIED

1184
00:45:07,331 --> 00:45:10,768
OUT LATER, I COMPARE IN THE LAB

1185
00:45:10,768 --> 00:45:13,304
ISSUE IS TO LOOK AT THE EFFECT

1186
00:45:13,304 --> 00:45:16,140
OF TRANSCRIPTIONAL CELLS AFTER

1187
00:45:16,140 --> 00:45:18,509
FORMALDEHYDE TREATMENT AND WE'RE

1188
00:45:18,509 --> 00:45:20,177
LOOKING AT WILD-TYPE CELLS AND

1189
00:45:20,177 --> 00:45:22,013
CSB KNOCK OUT CELLS.

1190
00:45:22,013 --> 00:45:24,415
SO BOTH SCENARIOS, IT WAS TRIEWT

1191
00:45:24,415 --> 00:45:26,717
WIDE FORMALDEHYDE AND LEADS TO

1192
00:45:26,717 --> 00:45:28,085
TRANSCRIPTIONAL SHUT OFF AS

1193
00:45:28,085 --> 00:45:30,187
MEASURED BY INCORPORATION OF EU,

1194
00:45:30,187 --> 00:45:31,355
SO THAT'S MASSIVE TRANSCRIPTION

1195
00:45:31,355 --> 00:45:36,761
BEING MEASURED HERE BUT IF WE

1196
00:45:36,761 --> 00:45:39,030
LOOK AT OVER TIME AFTER THE

1197
00:45:39,030 --> 00:45:40,364
FORMALDEHYDE IS KRAWN AND CELLS

1198
00:45:40,364 --> 00:45:42,266
RECOVER, WE SEE A GRADUAL

1199
00:45:42,266 --> 00:45:43,301
TRANSCRIPTION AND RECOVERY OF

1200
00:45:43,301 --> 00:45:45,136
TRANSCRIPTION IS MUCH MORE PRO

1201
00:45:45,136 --> 00:45:47,004
NOUNSED IN THE WILD-TYPE CELLS

1202
00:45:47,004 --> 00:45:49,006
THAN IN THE CSB KNOCK OUT CELLS,

1203
00:45:49,006 --> 00:45:51,676
SO THIS IS INDICATING THAT CSB

1204
00:45:51,676 --> 00:45:55,746
KNOCK OUT CELLS HAVE A PROBLEM

1205
00:45:55,746 --> 00:45:59,216
IN REACTIVATING TRANSCRIPTION

1206
00:45:59,216 --> 00:45:59,917
AFTER FORMALDEHYDE EXPOSURE AND

1207
00:45:59,917 --> 00:46:01,886
YOU MAY HAVE NOTED THIS THIS,A

1208
00:46:01,886 --> 00:46:04,188
PLIES BOTH TO PAN NUCLEAR

1209
00:46:04,188 --> 00:46:06,924
STAINING AS WELL AS NUCLEAR

1210
00:46:06,924 --> 00:46:08,192
STAINING SO THIS PRESUMABLY

1211
00:46:08,192 --> 00:46:10,127
MEANS THE IMPACT OF CSB IN THIS

1212
00:46:10,127 --> 00:46:12,997
REGARD IS NOT JUST RELATED TO

1213
00:46:12,997 --> 00:46:14,999
POL2 TRANSCRIPTION BUT ALSO POL1

1214
00:46:14,999 --> 00:46:15,333
TRANSCRIPTION.

1215
00:46:15,333 --> 00:46:17,234
AND WE GET SIMILAR RESULTS FOR

1216
00:46:17,234 --> 00:46:19,670
CSA KNOCK OUT CELLS BUT NOT FOR

1217
00:46:19,670 --> 00:46:25,876
XPA OR THE NER INCISION AND

1218
00:46:25,876 --> 00:46:26,344
NUCLEACES.

1219
00:46:26,344 --> 00:46:27,979
AGAIN SEPARATING THE DPC

1220
00:46:27,979 --> 00:46:31,582
ACTIVITIES OF THOSE COMPONENTS

1221
00:46:31,582 --> 00:46:41,125
FROM THOSE OTHER PROTEINS.

1222
00:46:41,125 --> 00:46:42,994
AS WAS SHOWN PREVIOUSLY FOR UV

1223
00:46:42,994 --> 00:46:46,230
DAMAGE FOR EXAMPLE, CSB AND CSA,

1224
00:46:46,230 --> 00:46:48,666
INTERACT WITH RNA POLYMERASE 2

1225
00:46:48,666 --> 00:46:49,367
AFTER FORMALDEHYDE TREATMENT AND

1226
00:46:49,367 --> 00:46:54,572
IN THE CASE OF CSA, ITS

1227
00:46:54,572 --> 00:46:58,175
ASSOCIATION BY CO IMMUNO

1228
00:46:58,175 --> 00:47:05,349
PRECIPITATION IS CSB DEPENDENT.

1229
00:47:05,349 --> 00:47:07,284
SO FINALLY, WHAT WAS SHARED WITH

1230
00:47:07,284 --> 00:47:09,620
YOU AS HOW WE USED AN

1231
00:47:09,620 --> 00:47:10,688
ESTABLISHED TECHNIQUE AND THIS

1232
00:47:10,688 --> 00:47:15,326
BROUGHT THIS INTO THE REALM OF

1233
00:47:15,326 --> 00:47:17,795
NEXT GEN SEQUENCING AND

1234
00:47:17,795 --> 00:47:19,063
APPROACHES AND BUT IT'S BEEN

1235
00:47:19,063 --> 00:47:25,169
KNOWN FOR TIME IF YOU SDS LY

1236
00:47:25,169 --> 00:47:27,405
CELLS AND CARRY OUT THIS, IT

1237
00:47:27,405 --> 00:47:28,606
WILL PRECIPITATE PROTEINS BUT

1238
00:47:28,606 --> 00:47:31,909
NOT DNA BUT IT WILL PRECIPITATE

1239
00:47:31,909 --> 00:47:35,379
DNA IF THAT DNA IS ATTACHED TO

1240
00:47:35,379 --> 00:47:35,646
PROTEINS.

1241
00:47:35,646 --> 00:47:36,914
SO WE THEREFORE REASONED IF WE

1242
00:47:36,914 --> 00:47:42,353
CARRY OUT THAT PROTOCOL ACTUALLY

1243
00:47:42,353 --> 00:47:44,955
SEVERAL ROUNDS OF THE PROTOCOL,

1244
00:47:44,955 --> 00:47:46,123
WE SHOULD SELECTIVELY RETRIEVE

1245
00:47:46,123 --> 00:47:49,060
REGIONS OF THE GENOME WHICH

1246
00:47:49,060 --> 00:47:51,328
CONTAIN DPCs, OF COURSE THOSE

1247
00:47:51,328 --> 00:47:53,931
REGION OF THE GENOME COULD BE

1248
00:47:53,931 --> 00:47:55,399
ANALYZED BY MASSIVELY

1249
00:47:55,399 --> 00:47:56,534
PARALLELLED NEXT GENERATION

1250
00:47:56,534 --> 00:47:57,768
SEQUENCING, SO WE'LL BE ABLE TO

1251
00:47:57,768 --> 00:47:59,970
IN A TBLOABAL WAY IF YOU LIKE,

1252
00:47:59,970 --> 00:48:01,739
LOOK AT REGIONS OF THE GENOME

1253
00:48:01,739 --> 00:48:03,140
THAT HAVE DPCs AND ACTUALLY

1254
00:48:03,140 --> 00:48:05,242
LOOK AT THEIR REPAIR OVER TIME.

1255
00:48:05,242 --> 00:48:08,179
BECAUSE WE'RE CARRYING OUT THESE

1256
00:48:08,179 --> 00:48:09,847
EXPERIMENTS WITH SYNCHRONIZED

1257
00:48:09,847 --> 00:48:11,415
RPE1 CELLS, THESE CELLS ARE IN

1258
00:48:11,415 --> 00:48:12,917
G1, SO WE'RE NOT GOING TO HAVE

1259
00:48:12,917 --> 00:48:16,787
TO THINK ABOUT THINGS SUCH AS

1260
00:48:16,787 --> 00:48:17,988
DNA REPLICATION, WHICH IS A WAY

1261
00:48:17,988 --> 00:48:20,224
TO IMPACT ON OTHER PATH WAYINGS

1262
00:48:20,224 --> 00:48:24,095
OF DPC REPAIR, AND WE CAN TREAT

1263
00:48:24,095 --> 00:48:25,463
THESE CELLS WITH FORMALDEHYDE

1264
00:48:25,463 --> 00:48:27,531
AND WASH IT OFF AND LOOK AT OVER

1265
00:48:27,531 --> 00:48:29,333
TIME THE REPAIR OF DNA DAMAGE.

1266
00:48:29,333 --> 00:48:32,203
SO WE'RE PLOTTING ON THIS--JUST

1267
00:48:32,203 --> 00:48:33,737
HERE, ARE BASICALLY THE RESULTS

1268
00:48:33,737 --> 00:48:38,676
OF 2 SUCH EXPERIMENTS.

1269
00:48:38,676 --> 00:48:45,316
ALONG THE BOTTOM, IS THE KNA

1270
00:48:45,316 --> 00:48:47,084
CROSS LINK COVERAGE IF YOU LIKE

1271
00:48:47,084 --> 00:48:48,385
EMPLOY THE GENOMIC LOCI WHICH

1272
00:48:48,385 --> 00:48:53,591
ARE PULLED DOWN THROUGH THE KCL

1273
00:48:53,591 --> 00:48:55,759
PRECIPITATION PROCEDURE AFTER

1274
00:48:55,759 --> 00:48:58,362
TREATING FORFORMALDEHYDE. 

1275
00:48:58,362 --> 00:49:03,367
AND ON THE Y-AXIS, ANOTHER SET

1276
00:49:03,367 --> 00:49:11,208
OF DATA GENERATED AFTER THE

1277
00:49:11,208 --> 00:49:11,976
CELLS ARE INDUCED.

1278
00:49:11,976 --> 00:49:14,078
SO IF YOU THINK ABOUT AND CELL

1279
00:49:14,078 --> 00:49:16,814
THAT'S NOT REPAIRED SHOULD LIE

1280
00:49:16,814 --> 00:49:18,349
THERE, THE REGION OF THE GENOME

1281
00:49:18,349 --> 00:49:19,550
WHICH IS BEING REPAIRED SHOULD

1282
00:49:19,550 --> 00:49:21,418
BE LYING OFF THE DIAGONAL

1283
00:49:21,418 --> 00:49:25,356
TOWARDS THE BOTTOM, AND THE

1284
00:49:25,356 --> 00:49:26,757
STRIKING FINDING HERE IDENTIFIED

1285
00:49:26,757 --> 00:49:29,193
THE RANGE OF LOCI ACROSS THE

1286
00:49:29,193 --> 00:49:31,962
GENOME THAT WERE PREFERENTIALLY

1287
00:49:31,962 --> 00:49:34,165
REPAIRED AND IF WE MAP ON TO

1288
00:49:34,165 --> 00:49:36,834
THOSE DATA ESTABLISH RNA POL 2

1289
00:49:36,834 --> 00:49:38,969
TO OCCUPANCY DATA AND WE CAN SEE

1290
00:49:38,969 --> 00:49:40,971
THAT THOSE LOCI, TEBD TEND TO BE

1291
00:49:40,971 --> 00:49:44,208
LOCI THAT HAVE A HIGHER RNA POL

1292
00:49:44,208 --> 00:49:44,909
2 OCCUPANCY.

1293
00:49:44,909 --> 00:49:46,377
SO THIS IS A KIND OF APPROACH,

1294
00:49:46,377 --> 00:49:53,450
WE CAN ALSO LOOK AT WILD-TYPE OR

1295
00:49:53,450 --> 00:49:54,051
OTHER CSB BACKGROUNDS.

1296
00:49:54,051 --> 00:49:55,953
BUT I'M SHOWING THE WORK, THE

1297
00:49:55,953 --> 00:49:57,922
ANALYSIS WAS CARRIED OUT BY

1298
00:49:57,922 --> 00:50:01,025
[INDISCERNIBLE] IN THE GROUP, A

1299
00:50:01,025 --> 00:50:03,027
VERY EXPERIENCED AND VERY

1300
00:50:03,027 --> 00:50:09,266
VERSATILE EXPERIMENTAL BIOLOGIST

1301
00:50:09,266 --> 00:50:12,636
AS WELL AS A FANTASTIC

1302
00:50:12,636 --> 00:50:13,037
BIOINFORMATICIAN.

1303
00:50:13,037 --> 00:50:13,771
WE ARE COMPARING ALONG THE

1304
00:50:13,771 --> 00:50:15,172
BOTTOM, THE DAILY BASIS THEA

1305
00:50:15,172 --> 00:50:17,975
AFTER A 6 HOUR RECOVERY OF CSB

1306
00:50:17,975 --> 00:50:20,211
KNOCK OUT CELLS AND ON THE

1307
00:50:20,211 --> 00:50:22,146
Y-AXIS AFTER A 6 HOUR RECOVER

1308
00:50:22,146 --> 00:50:27,518
EVALUATION PROCESS WILD-TYPE

1309
00:50:27,518 --> 00:50:27,985
CELLS.

1310
00:50:27,985 --> 00:50:30,955
SO IN THIS SCENARIO HERE, ANY

1311
00:50:30,955 --> 00:50:33,524
GEANUS IN THE LOCUST PROMOTED BY

1312
00:50:33,524 --> 00:50:36,160
THE CSB WILL FALL BELOW THE

1313
00:50:36,160 --> 00:50:37,228
DIAGONAL HIGHLIGHTED HERE IN

1314
00:50:37,228 --> 00:50:38,429
BLUE AND THAT IDENTIFIED JUST

1315
00:50:38,429 --> 00:50:40,965
OVER A THOUSAND GENES THAT

1316
00:50:40,965 --> 00:50:43,267
WERE--WE BEING DEFINE AS THEIR

1317
00:50:43,267 --> 00:50:46,170
REPAIR BEING CSB DEPENDENT.

1318
00:50:46,170 --> 00:50:48,172
AND INTERESTING WHEN WE LOOK AT

1319
00:50:48,172 --> 00:50:49,373
THOSE GENES AND IN THIS CASE,

1320
00:50:49,373 --> 00:50:52,109
WHAT WE'VE DONE HERE IS TAKE

1321
00:50:52,109 --> 00:50:54,745
THOSE GENES AND BASICALLY PLOT

1322
00:50:54,745 --> 00:50:56,614
THEM IN A METAGENE PLOT, SO ALL

1323
00:50:56,614 --> 00:50:58,716
THE GENES ARE TOGETHER, WE GET

1324
00:50:58,716 --> 00:51:00,551
THESE--WE GET THESE CURVES HERE,

1325
00:51:00,551 --> 00:51:03,220
AND WHAT WE FIND IS THAT THE

1326
00:51:03,220 --> 00:51:08,092
IMPACT OF CSB, ON DPC REPAIR IS

1327
00:51:08,092 --> 00:51:09,260
MOSTLY WITHIN THE TRANSCRIBED

1328
00:51:09,260 --> 00:51:11,228
PORTION OF THE GENE, THE GENE

1329
00:51:11,228 --> 00:51:11,428
BODY.

1330
00:51:11,428 --> 00:51:15,866
SO THAT IS THE DIFFERENTIAL

1331
00:51:15,866 --> 00:51:18,235
BETWEEN THE WILD-TYPE SCENARIO,

1332
00:51:18,235 --> 00:51:20,638
HERE TO HERE, ORANGE TO RED, IS

1333
00:51:20,638 --> 00:51:22,673
LARGER THAN THE DIFFERENTIAL

1334
00:51:22,673 --> 00:51:25,209
HERE LIGHT BLUE TO DARK BLUE,

1335
00:51:25,209 --> 00:51:27,544
THOSE BEING IN THE PRESENCE OF

1336
00:51:27,544 --> 00:51:30,047
WILD-TYPE OR CSB KNOCK OUT

1337
00:51:30,047 --> 00:51:31,348
RESPECTIVELY, AND ANOTHER WAY OF

1338
00:51:31,348 --> 00:51:34,084
SHOWING THESE DAT IS LOOKING AT

1339
00:51:34,084 --> 00:51:35,719
THE DIFFERENTIAL BETWEEN

1340
00:51:35,719 --> 00:51:36,787
WILD-TYPE AND CSB KNOCK OUT ON

1341
00:51:36,787 --> 00:51:38,622
THE RIGHT HERE WHICH CLEARLY

1342
00:51:38,622 --> 00:51:40,424
INDICATING THAT THE IMPACT OF

1343
00:51:40,424 --> 00:51:41,925
CSB ON PROMOTING REPAIR IS

1344
00:51:41,925 --> 00:51:43,794
LARGELY CORRELATED WITH THE GENE

1345
00:51:43,794 --> 00:51:44,561
BODY.

1346
00:51:44,561 --> 00:51:47,197
THIS HAS BEEN A HIGHLIGHT OF

1347
00:51:47,197 --> 00:51:49,033
THIS BIG PEEK HERE AND IT'S

1348
00:51:49,033 --> 00:51:50,668
REFLECTING THE FACT THAT THE

1349
00:51:50,668 --> 00:51:52,002
TRANSCRIPTION START SITES TEND

1350
00:51:52,002 --> 00:51:54,471
TO BE LARGELY OPEN CHROMATIN

1351
00:51:54,471 --> 00:51:56,106
STRUCTURE, AND WE KNOW THROUGH

1352
00:51:56,106 --> 00:51:57,141
PROTEOMIC STUDIES THAT WE

1353
00:51:57,141 --> 00:51:58,475
CARRIED OUT IN CONJUNCTION WITH

1354
00:51:58,475 --> 00:52:01,679
THIS, THAT A LOT OF THAT, THOSE

1355
00:52:01,679 --> 00:52:03,747
DPCs ARE ACTUALLY

1356
00:52:03,747 --> 00:52:05,015
RNA-POLYMERASE 2, BEING CROSS

1357
00:52:05,015 --> 00:52:05,249
LINKED.

1358
00:52:05,249 --> 00:52:07,084
SO THE TAKE HOME MESSAGE HERE IS

1359
00:52:07,084 --> 00:52:09,520
THAT THERE ARE--THERE IS A SET

1360
00:52:09,520 --> 00:52:11,622
OF LOCI ACROSS THE GENOME THAT

1361
00:52:11,622 --> 00:52:12,823
ARE PREVENTIVE RENTIA WILY

1362
00:52:12,823 --> 00:52:16,593
REPAIRED IN TERMS OF DPCs,

1363
00:52:16,593 --> 00:52:17,928
THEY ARE CSB DEPENDENT AND THE

1364
00:52:17,928 --> 00:52:20,030
EFFECT IS LARGELY COMBOYNED TO

1365
00:52:20,030 --> 00:52:22,533
THE TRANSCRIBED PORTION OF THE

1366
00:52:22,533 --> 00:52:23,500
GENE.

1367
00:52:23,500 --> 00:52:25,769
SO TO SUMMARIZE, DPCs CAUSE

1368
00:52:25,769 --> 00:52:26,603
TRANSCRIPTION STRESS, WE THINK

1369
00:52:26,603 --> 00:52:28,872
THIS IS THE FIRST EVIDENT OF

1370
00:52:28,872 --> 00:52:29,840
TRANSCRIPTION COUPLED DECKER PC

1371
00:52:29,840 --> 00:52:33,177
REPAIR AND WE'VE SHOWN THAT CSB

1372
00:52:33,177 --> 00:52:35,813
PROMOTES DC REPAIR INDEPENDENTLY

1373
00:52:35,813 --> 00:52:37,414
OF CLASSIC TCNAR FACTORS.

1374
00:52:37,414 --> 00:52:39,850
THIS WORK HAS BEEN RECENTLY SENT

1375
00:52:39,850 --> 00:52:41,485
TO FOR PUBLICATION IN NATURE

1376
00:52:41,485 --> 00:52:43,020
CELL BIOLOGY AND HOPEFULLY WE

1377
00:52:43,020 --> 00:52:45,222
WILL BE ABLE TO READ IN THE NOT

1378
00:52:45,222 --> 00:52:47,725
TOO DISTANT FUTURE 2 WONDER AND

1379
00:52:47,725 --> 00:52:50,194
AND VERY COMPLIMENTARY STUDIES

1380
00:52:50,194 --> 00:52:51,762
BY THE LAG OF [INDISCERNIBLE]

1381
00:52:51,762 --> 00:52:53,797
AND [INDISCERNIBLE] WHICH WILL

1382
00:52:53,797 --> 00:52:54,732
BE APPEARING HOPEFULLY

1383
00:52:54,732 --> 00:52:56,033
BACK-TO-BACK TO BACK IN THE SAME

1384
00:52:56,033 --> 00:52:56,233
ISSUE.

1385
00:52:56,233 --> 00:52:57,835
FINALLY IN THE LAST 2 MINUTES, I

1386
00:52:57,835 --> 00:53:03,073
JUST WANT TO TRY TO RELATE THIS

1387
00:53:03,073 --> 00:53:04,942
BACK TO THE CLINICAL ARENA

1388
00:53:04,942 --> 00:53:07,344
AGAIN, AS YOU MAY BE AWARE,

1389
00:53:07,344 --> 00:53:09,446
THERE IS EXCISION REPAIR IN THE

1390
00:53:09,446 --> 00:53:10,614
SYNDROME BEING DESCRIBED OVER

1391
00:53:10,614 --> 00:53:12,015
THE YEARS, THERE'S BEEN THERE

1392
00:53:12,015 --> 00:53:14,551
KNOWN FOR MANY YEARS THAT

1393
00:53:14,551 --> 00:53:22,526
PATIENTS WITH MUTATIONS IN THE

1394
00:53:22,526 --> 00:53:24,595
LCC6--SORRY, ERCC6--ERCC8 WHICH

1395
00:53:24,595 --> 00:53:26,663
ENCODE CSBOR CSA RESPECTIVELY

1396
00:53:26,663 --> 00:53:29,199
AND WITH COCK CAIN'S SIN CHROME,

1397
00:53:29,199 --> 00:53:31,268
WHICH IS SEVERE DISORDER

1398
00:53:31,268 --> 00:53:33,470
PROGRESSIVE WEIGHT LOSS, MUSCLE

1399
00:53:33,470 --> 00:53:34,204
LOSS, NEURODEGENERATION AND

1400
00:53:34,204 --> 00:53:35,939
KIDNEY FAILURE AND THAT'S VERY

1401
00:53:35,939 --> 00:53:39,843
DIFFERENT PRACTICES PATIENTS

1402
00:53:39,843 --> 00:53:41,912
WITH GENES MUTATIONS AND IN THE

1403
00:53:41,912 --> 00:53:45,816
DOWN STREAM ASPECTS OF TCNER AND

1404
00:53:45,816 --> 00:53:47,451
THESE INCLUDE THE XPMUTATIONS

1405
00:53:47,451 --> 00:53:49,887
AND WHICH IS CHARACTERIZED BY

1406
00:53:49,887 --> 00:53:52,289
EXTREME UV SENSITIVITY, AND

1407
00:53:52,289 --> 00:53:56,126
INCREASE CANCER RISK.

1408
00:53:56,126 --> 00:54:01,031
SO CLEARLY THE IMPACTS OF CSA,

1409
00:54:01,031 --> 00:54:03,300
AND CSB IN NEUROLOGICAL ISSUE

1410
00:54:03,300 --> 00:54:04,601
TISSUES CANNOT BE THROUGH LIGHT

1411
00:54:04,601 --> 00:54:06,870
EXPOSURE SO THIS HAS TO BE

1412
00:54:06,870 --> 00:54:08,172
DRIVEN BY THINGS THAT DON'T

1413
00:54:08,172 --> 00:54:17,448
INCLUDE WHATY  WE CALL CLASSICAL

1414
00:54:17,448 --> 00:54:18,115
SUBSTRATES.

1415
00:54:18,115 --> 00:54:20,284
[INDISCERNIBLE] GROUP INDUCES

1416
00:54:20,284 --> 00:54:21,485
TRANSCRIPTIONAL STRESS FROM

1417
00:54:21,485 --> 00:54:23,020
FORMALDEHYDE AND DRIVES PAIN

1418
00:54:23,020 --> 00:54:25,489
LIKE SYNDROMES PHENOTYPES IN

1419
00:54:25,489 --> 00:54:26,056
MICE.

1420
00:54:26,056 --> 00:54:27,458
HOWEVER THE RESPONSIBLE

1421
00:54:27,458 --> 00:54:28,425
FORMALDEHYDE INDUCED LESIONS

1422
00:54:28,425 --> 00:54:31,829
HAVE BEEN UNKNOWN, WHAT WE WOULD

1423
00:54:31,829 --> 00:54:34,131
LIKE TO PROPOSE IS THAT OUR

1424
00:54:34,131 --> 00:54:37,334
FINDING SUGGEST THAT EFFECTIVE

1425
00:54:37,334 --> 00:54:38,302
TRANSCRIPTION COUPLED DCP REPAIR

1426
00:54:38,302 --> 00:54:40,871
IS LARGELY CONTRIBUTING TO THE

1427
00:54:40,871 --> 00:54:46,276
UNIQUE NEUROLOGICAL  FEATURES OF

1428
00:54:46,276 --> 00:54:46,443
CS.

1429
00:54:46,443 --> 00:54:47,711
FINALLY, WHAT'S MIXTURE AND I

1430
00:54:47,711 --> 00:54:50,013
WON'T DWELL ON THIS BUT NOW

1431
00:54:50,013 --> 00:54:51,782
WE'RE DELVING FURTHER INTO THIS

1432
00:54:51,782 --> 00:54:54,685
AND OTHER ARENAS BY CARRYING OUT

1433
00:54:54,685 --> 00:54:57,521
MORE CRISPR KREENS IN THE CSP

1434
00:54:57,521 --> 00:54:59,089
KNOCK OUT BACK GROUPEDS WHICH

1435
00:54:59,089 --> 00:55:01,291
ALLOWED US RECENTLY TO IERK

1436
00:55:01,291 --> 00:55:06,897
DENTIFY A NEW DPC FACTOR CALLED

1437
00:55:06,897 --> 00:55:09,900
TOPORS, WHICH WE THINK WILL WELL

1438
00:55:09,900 --> 00:55:11,902
IN PARALLEL WITH RNF4 AND THE

1439
00:55:11,902 --> 00:55:12,603
OTHER FACTORS I TALKED ABOUT

1440
00:55:12,603 --> 00:55:15,572
TODAY AND THIS WORK HAS BEEN

1441
00:55:15,572 --> 00:55:16,306
SPEARHEADED BY CHLOE

1442
00:55:16,306 --> 00:55:17,674
[INDISCERNIBLE] IN THE LAB WITH

1443
00:55:17,674 --> 00:55:18,775
ADVICE FROM CHRIS

1444
00:55:18,775 --> 00:55:19,543
[INDISCERNIBLE] AND OTHERS.

1445
00:55:19,543 --> 00:55:21,778
AND AGAIN WE'RE BENEFITING FROM

1446
00:55:21,778 --> 00:55:24,314
OUR FANTASTIC COLLABORATORS

1447
00:55:24,314 --> 00:55:25,816
INCLUDING JULIAN STINGLE,

1448
00:55:25,816 --> 00:55:26,717
[INDISCERNIBLE] AND OTHERS.

1449
00:55:26,717 --> 00:55:28,051
SO TO ROUND OFF THERE.

1450
00:55:28,051 --> 00:55:31,989
THIS IS A PICTURE OF MY LAB IN

1451
00:55:31,989 --> 00:55:34,191
SUNNY NORTH UP JUST AN HOUR AND

1452
00:55:34,191 --> 00:55:36,660
A HALF NORTH OF SUNNY CAMBRIDGE,

1453
00:55:36,660 --> 00:55:38,161
ALTHOUGH IT'S NOT SUNNY TODAY.

1454
00:55:38,161 --> 00:55:41,732
AFTER 30 YEARS PLUS IN THE

1455
00:55:41,732 --> 00:55:44,501
INSTITUTE MY LAB MOVED POT

1456
00:55:44,501 --> 00:55:46,904
CANCER RESEARCH UK CAMBRIDGE

1457
00:55:46,904 --> 00:55:48,038
INSTITUTE ON THE BIOMEDICAL

1458
00:55:48,038 --> 00:55:51,141
CAMPUS WHICH IS THE LARGEST AND

1459
00:55:51,141 --> 00:55:52,676
ARGUABLY THE BEST BIOMEDICAL

1460
00:55:52,676 --> 00:55:53,744
CAMPUS IN EUROPE, CERTAINLY THE

1461
00:55:53,744 --> 00:55:55,612
LARGEST AND IT'S A FANTASTIC

1462
00:55:55,612 --> 00:55:56,213
PLACE TO BE.

1463
00:55:56,213 --> 00:55:57,948
I WILL LEAVE IT THERE AND I WILL

1464
00:55:57,948 --> 00:56:04,588
TAKE QUESTIONS. 

1465
00:56:04,588 --> 00:56:06,390
THANK YOU.

1466
00:56:06,390 --> 00:56:10,527
>> THANK YOU DR. JACKSON
FOR THE

1467
00:56:10,527 --> 00:56:13,030
AMAZING TALK. 

1468
00:56:13,030 --> 00:56:15,465
LET'S MOVE TO THE Q&A
SESSION,

1469
00:56:15,465 --> 00:56:17,467
WE HAVE A FEW MINUTES, SO

1470
00:56:17,467 --> 00:56:19,503
AUDIENCE PLEASE TYPE IN YOUR

1471
00:56:19,503 --> 00:56:20,904
QUESTIONS IN THE CHAT BOX SO WE

1472
00:56:20,904 --> 00:56:22,172
CAN LOOK AT THAT.

1473
00:56:22,172 --> 00:56:25,809
WE HAVE A QUESTION RIGHT NOW

1474
00:56:25,809 --> 00:56:27,511
FROM [INDISCERNIBLE] ASKING WHAT

1475
00:56:27,511 --> 00:56:33,317
ARE THE STATUS OF RD54 L2 AND

1476
00:56:33,317 --> 00:56:36,853
ZNF451 IN TERMS OF THE MUTATION

1477
00:56:36,853 --> 00:56:40,591
COPY NUMBER?

1478
00:56:40,591 --> 00:56:41,792
ATOPOCIDE MEDIATED

1479
00:56:41,792 --> 00:56:42,359
[INDISCERNIBLE] CANCERS?

1480
00:56:42,359 --> 00:56:43,827
>> THAT'S A GREAT
QUESTION.

1481
00:56:43,827 --> 00:56:46,129
THIS IS A NEW ARENA FOR US AND I

1482
00:56:46,129 --> 00:56:48,565
THINK THAT IS ANOTHER VERY

1483
00:56:48,565 --> 00:56:49,733
INTERESTING THING TO EXPLORE, WE

1484
00:56:49,733 --> 00:56:51,468
MANY IN THE FIELD I THINK ARE

1485
00:56:51,468 --> 00:56:53,470
NOW VERY WELL AWARE OF

1486
00:56:53,470 --> 00:56:56,273
MUTATIONAL SIGNATURES AND

1487
00:56:56,273 --> 00:56:59,810
MOWITATIONS BUT OF COURSE COPY

1488
00:56:59,810 --> 00:57:01,812
NUMBER CHANGES CAN DRIVE

1489
00:57:01,812 --> 00:57:02,646
PRONOUNCED PHENOTYPES AND I KNOW

1490
00:57:02,646 --> 00:57:04,081
THAT'S THE LAB AND OTHER LABS

1491
00:57:04,081 --> 00:57:05,382
ARE GETTING INTO BUT

1492
00:57:05,382 --> 00:57:06,550
UNFORTUNATELY I DON'T HAVE AN

1493
00:57:06,550 --> 00:57:13,724
ANSWER TO THAT QUESTION RIGHT

1494
00:57:13,724 --> 00:57:13,890
YOU.

1495
00:57:13,890 --> 00:57:15,826
>> KEN HAS A QUESTION, DO
YOU

1496
00:57:15,826 --> 00:57:17,694
WANT TO ASK IT DIRECTLY?

1497
00:57:17,694 --> 00:57:21,732
>> THERE WE GO, WONDERFUL
TALK.

1498
00:57:21,732 --> 00:57:23,634
REALLY SPECTACULAR.

1499
00:57:23,634 --> 00:57:25,969
I REALLY ENJOYED THE CHEMO

1500
00:57:25,969 --> 00:57:30,007
THERAPY ASPECTS OF IT AND THE

1501
00:57:30,007 --> 00:57:30,941
NEUROLOGIC XP GENETICS.

1502
00:57:30,941 --> 00:57:36,780
ONE OF THE ISSUES IN XP IS WHY

1503
00:57:36,780 --> 00:57:40,550
THE XBC PATIENTS DON'T HAVE

1504
00:57:40,550 --> 00:57:41,318
NEUROLOGIC DEGENERATION WHEREAS

1505
00:57:41,318 --> 00:57:44,988
THE XPA AND ALSO THOSE WITH XP,

1506
00:57:44,988 --> 00:57:47,391
OR XPV WHICH ARE HELIX CASES AND

1507
00:57:47,391 --> 00:57:50,027
THEY DIDN'T SHOW UP IN YOUR

1508
00:57:50,027 --> 00:57:50,761
SCREEN AT ALL.

1509
00:57:50,761 --> 00:57:56,099
SO EVERYONE'S BEEN LOOKING FOR

1510
00:57:56,099 --> 00:57:59,970
LESIONS THAT ARE NOT REPAIRED BY

1511
00:57:59,970 --> 00:58:02,139
NUCLEOTIDE EXCISION REPAIR AND

1512
00:58:02,139 --> 00:58:04,474
CYCLOPUREINS HAVE BEEN THE

1513
00:58:04,474 --> 00:58:07,177
PROMINENT 1S, YOU'RE SUGGESTING

1514
00:58:07,177 --> 00:58:09,046
THAT MAYBE OTHER 1S THAT HAS--DO

1515
00:58:09,046 --> 00:58:10,681
YOU KNOW OF ANYONE THAT'S BEEN

1516
00:58:10,681 --> 00:58:13,016
COOING THESE STUDIES THAT YOU'VE

1517
00:58:13,016 --> 00:58:15,152
BEEN SUGGESTING, LOOKING FOR

1518
00:58:15,152 --> 00:58:18,021
THESE LESIONS, AND THEN PATIENTS

1519
00:58:18,021 --> 00:58:19,656
WITH THESE NUCLEOTIDE EXCISION

1520
00:58:19,656 --> 00:58:20,924
REPAIR RELATED DECS THAT

1521
00:58:20,924 --> 00:58:27,531
PRESENCE ARE ABSENCE OR

1522
00:58:27,531 --> 00:58:28,331
NEUROLOGIC DEGENERATION.

1523
00:58:28,331 --> 00:58:29,633
>> THAT'S A GREAT
QUESTION, WE

1524
00:58:29,633 --> 00:58:30,934
ALL KNOW WHAT TOOLS, WHAT TOOLS

1525
00:58:30,934 --> 00:58:33,036
WE CAN USE AT THE CELLULAR LEVEL

1526
00:58:33,036 --> 00:58:34,438
TO GENERATE SPECIFIC TYPES OF

1527
00:58:34,438 --> 00:58:35,372
LESIONS BUT I THINK WE'RE

1528
00:58:35,372 --> 00:58:38,341
ALREADY IN A POSITION TO COMPARE

1529
00:58:38,341 --> 00:58:41,044
OUR CRISPR STREAM DATA SETS FOR

1530
00:58:41,044 --> 00:58:43,146
EXAMPLE WITH THOSE OBVIOUSLY

1531
00:58:43,146 --> 00:58:44,414
ARISEN THROUGH OTHER TREATMENTS

1532
00:58:44,414 --> 00:58:47,050
SUCH AS UV AND WHATEVER.

1533
00:58:47,050 --> 00:58:48,685
OF COURSE, SOME MUTATIONS ARE

1534
00:58:48,685 --> 00:58:51,188
NOT ENTIRE LOSS OF FUNCTION

1535
00:58:51,188 --> 00:58:52,989
MUTATIONS THAT THEY COULD BE

1536
00:58:52,989 --> 00:58:55,659
SEPARATION OF FUNCTION MUTATIONS

1537
00:58:55,659 --> 00:58:57,227
IN MANY CASES, MULTIFUNCTIONAL

1538
00:58:57,227 --> 00:58:59,396
PROTEINS, AND TAKEN--THEY'S

1539
00:58:59,396 --> 00:59:00,497
WHOLE--A WHOLE--A WHOLE OTHER

1540
00:59:00,497 --> 00:59:02,599
ARENA THAT NEEDS TO BE EXPLORED

1541
00:59:02,599 --> 00:59:03,500
BUT I AGREE.

1542
00:59:03,500 --> 00:59:05,569
I THINK WE'RE TOUCHING ON

1543
00:59:05,569 --> 00:59:10,373
CERTAIN ASSPEBTS OF THE NUR O

1544
00:59:10,373 --> 00:59:11,374
DEGENERATION AND COCKAYNE'S

1545
00:59:11,374 --> 00:59:12,809
SYNDROME AND OTHER PATIENTS BUT

1546
00:59:12,809 --> 00:59:13,910
WE'RE NOT EXPLAINING OTHER

1547
00:59:13,910 --> 00:59:18,148
ASPECTS OF THESE DISORDERS WHICH

1548
00:59:18,148 --> 00:59:20,117
ARE DEALING WITH A WHOLE RANGE

1549
00:59:20,117 --> 00:59:22,719
OF OTHER LESIONS FROM ENDOGENOUS

1550
00:59:22,719 --> 00:59:26,156
TO EXOGENOUS CHEMICALS.

1551
00:59:26,156 --> 00:59:28,258
SO I'M RAMBLING HERE BECAUSE I

1552
00:59:28,258 --> 00:59:29,760
DON'T REALLY HAVE AN ANSWER BUT

1553
00:59:29,760 --> 00:59:30,827
IT'S A GREAT QUESTION.

1554
00:59:30,827 --> 00:59:33,363
>> WE ARE ACTUALLY
PREPARING A

1555
00:59:33,363 --> 00:59:34,765
MANUSCRIPT, WE HAVE 18 XPA

1556
00:59:34,765 --> 00:59:37,300
PATIENT ANDS THEY RANGE FROM

1557
00:59:37,300 --> 00:59:41,204
SEVERE NEUROLOGIC DEGENERATION

1558
00:59:41,204 --> 00:59:42,305
TO PATIENTS [INDISCERNIBLE]

1559
00:59:42,305 --> 00:59:43,907
COLLEGE, DIFFERENT MUTATIONS IN

1560
00:59:43,907 --> 00:59:44,674
THE SAME GENE.

1561
00:59:44,674 --> 00:59:47,244
>> WOW, I GUESS SOME OF

1562
00:59:47,244 --> 00:59:49,646
THESE--ARE SOME OF THESE

1563
00:59:49,646 --> 00:59:51,748
POSSIBLY DOMINANT NEGATIVES OR

1564
00:59:51,748 --> 00:59:52,883
PARTIALLY FUNCTIONAL PROTEINS

1565
00:59:52,883 --> 00:59:54,484
THAT BASICALLY MIGHT GUM UP THE

1566
00:59:54,484 --> 00:59:56,586
WORK ANDS EVEN HAVE BROADER

1567
00:59:56,586 --> 01:00:00,891
EFFECTS RATHER THAN A KNOCK OUT

1568
01:00:00,891 --> 01:00:01,224
PERHAPS?

1569
01:00:01,224 --> 01:00:03,193
I LOOK FORWARD TO READING YOUR

1570
01:00:03,193 --> 01:00:07,931
PAPER. 

1571
01:00:07,931 --> 01:00:08,565
>> THANK YOU.

1572
01:00:08,565 --> 01:00:12,536
>> DR., DO YOU WANT TO ASK
A

1573
01:00:12,536 --> 01:00:13,436
QUESTION.

1574
01:00:13,436 --> 01:00:15,772
>> THANK YOU, STEVE, WE
REALLY

1575
01:00:15,772 --> 01:00:20,377
ENJOYED IT, SOME OF US WOULD

1576
01:00:20,377 --> 01:00:22,612
ADVOCATE THAT CSV HAS A MAJOR

1577
01:00:22,612 --> 01:00:23,513
MITOCHONDRIA DISFUNCTION THAT

1578
01:00:23,513 --> 01:00:24,748
COULD BE A FACTOR IN THE SYSTEM,

1579
01:00:24,748 --> 01:00:27,350
I WOULD LIKE TO ASK YOU IF YOU

1580
01:00:27,350 --> 01:00:31,354
TRIED THE RESUSMGZ OF RNA

1581
01:00:31,354 --> 01:00:32,222
SYNTHESIS EXPERIMENTS THAT WERE

1582
01:00:32,222 --> 01:00:36,726
CLASSIC IN THE LITERATURE TO SEE

1583
01:00:36,726 --> 01:00:41,131
IF WHEN THERE IS A DELAYING

1584
01:00:41,131 --> 01:00:42,632
RESUSMGZ OF TRANGZ SCRIPGZ WITH

1585
01:00:42,632 --> 01:00:45,268
DAMAGE AFTER THESE AGENTS, THESE

1586
01:00:45,268 --> 01:00:47,204
CROSS LINKERS THAT WOULD

1587
01:00:47,204 --> 01:00:52,409
INDICATE THAT THE ACTIVE GENES

1588
01:00:52,409 --> 01:00:55,312
ARE LATER IN CSA AND B?

1589
01:00:55,312 --> 01:00:56,947
>> ABSOLUTELY, I THINK THE
DATA

1590
01:00:56,947 --> 01:00:58,782
I SHARED WAS INDICATING THAT AT

1591
01:00:58,782 --> 01:01:02,219
THE CELLULAR LEVEL AT LEAST FOR

1592
01:01:02,219 --> 01:01:03,520
RNAPOL2, TRANSCRIPTION.

1593
01:01:03,520 --> 01:01:05,655
THE OOH RENA OF MITOCHONDRIA IS

1594
01:01:05,655 --> 01:01:08,291
NOT SOMETHING WE'VE NOT DELVED

1595
01:01:08,291 --> 01:01:10,460
INTO BUT IT IS VERY IMPORTANT

1596
01:01:10,460 --> 01:01:15,098
AND I WILL, ON THE BACK OF THIS

1597
01:01:15,098 --> 01:01:17,033
QUESTION, I WILL GO BACK IN MY

1598
01:01:17,033 --> 01:01:19,236
LAB AND SEE IF THERE AND

1599
01:01:19,236 --> 01:01:20,170
EXPERIMENTS WE COULD RUN TO

1600
01:01:20,170 --> 01:01:21,705
PROBE INTO THAT BUT IF YOU HAVE

1601
01:01:21,705 --> 01:01:24,207
ANY SPECIFIC QUESTIONS, WILL,

1602
01:01:24,207 --> 01:01:25,442
PLEASE DO E-MAIL US AND I WOULD

1603
01:01:25,442 --> 01:01:27,744
LIKE TO DISCUSS THAT.

1604
01:01:27,744 --> 01:01:31,681
>> GLADLY. 

1605
01:01:31,681 --> 01:01:32,015
YEAH.

1606
01:01:32,015 --> 01:01:32,315
COMPREHEND.

1607
01:01:32,315 --> 01:01:33,783
>> WE HAVE A QUESTION FROM

1608
01:01:33,783 --> 01:01:34,885
[INDISCERNIBLE].

1609
01:01:34,885 --> 01:01:36,753
VERY INTERESTING TALK, THANK

1610
01:01:36,753 --> 01:01:38,121
YOU, I'M WONDERING WHAT'S

1611
01:01:38,121 --> 01:01:40,690
HAPPENING TO THE CELL SET AFTER

1612
01:01:40,690 --> 01:01:41,157
ATOPOCIDE TREATMENT.

1613
01:01:41,157 --> 01:01:44,461
THIS IS EITHER A WILD-TYPE OR

1614
01:01:44,461 --> 01:01:47,731
THE RD54 L2 KA, ARE THEY

1615
01:01:47,731 --> 01:01:55,005
PROLIFERATING OR SEN  SENESCENC?

1616
01:01:55,005 --> 01:01:56,539
>> YES, IF YOU CARRY OUT
THE

1617
01:01:56,539 --> 01:01:57,173
SCREEN, YOU MEASURE

1618
01:01:57,173 --> 01:01:57,974
PROLIFERATION EMPLOY SO THE

1619
01:01:57,974 --> 01:01:59,709
CELLS COULD BE DYING OR THEY

1620
01:01:59,709 --> 01:02:01,011
COULD JUST BE STOPPING

1621
01:02:01,011 --> 01:02:02,078
PROLIFERATING AND THEY WILL

1622
01:02:02,078 --> 01:02:03,680
STILL BEHAVE AS A KNOCK OUT AND

1623
01:02:03,680 --> 01:02:05,248
THE ONLY WAY YOU CAN REALLY

1624
01:02:05,248 --> 01:02:06,516
FIGURE OUT WHAT'S GOING ON IS

1625
01:02:06,516 --> 01:02:09,152
THEN TO GO IN AND HAVE A LOOK AT

1626
01:02:09,152 --> 01:02:11,988
THOSE THINGS IN A SPECIFIC WAY.

1627
01:02:11,988 --> 01:02:18,728
I THINK THE IDEA ISN'T CELL

1628
01:02:18,728 --> 01:02:20,664
DYING AND IF IT ISN'T WHERE'S IT

1629
01:02:20,664 --> 01:02:21,765
ANYTHING OFF.

1630
01:02:21,765 --> 01:02:22,699
THAT'S AN INTERESTING 1 AND

1631
01:02:22,699 --> 01:02:24,367
SOMETIMES IF YOU GET INTO THAT

1632
01:02:24,367 --> 01:02:25,769
IF YOU TAKE A LOOK AT THE

1633
01:02:25,769 --> 01:02:27,070
BACKGROUND AND LOOK AT FACTORS

1634
01:02:27,070 --> 01:02:28,405
THAT MAY GIVE YOU RESISTANCE

1635
01:02:28,405 --> 01:02:30,140
BECAUSE IN SOME CASES, THE THING

1636
01:02:30,140 --> 01:02:32,075
THAT'S ENDING UP KILLING THE

1637
01:02:32,075 --> 01:02:35,378
CELL MAYBE ISN'T THE DAMAGE BUT

1638
01:02:35,378 --> 01:02:36,379
MAYBE CHROMOSOMAL TRANSLOCATIONS

1639
01:02:36,379 --> 01:02:38,782
OR ABERRATIONS THAT ARE CAUSED

1640
01:02:38,782 --> 01:02:40,717
BY DNA REPAIR PROCESSES, SO

1641
01:02:40,717 --> 01:02:42,018
AGAIN IT'S A QUESTION ABOUT HOW

1642
01:02:42,018 --> 01:02:43,787
TO CLEAR OUT FROM ME BUT I THINK

1643
01:02:43,787 --> 01:02:46,556
IT'S A GOOD 1 AND I'M LOOKING

1644
01:02:46,556 --> 01:02:47,724
FORWARD TO DETAIL ABOUT WHAT'S

1645
01:02:47,724 --> 01:02:48,992
HAPPENING TO THESE CELLS THAT

1646
01:02:48,992 --> 01:02:53,396
DIE AND ALSO WHAT HAPPENS TO THE

1647
01:02:53,396 --> 01:02:54,497
CELL SURVIVE, WHAT SURVIVE

1648
01:02:54,497 --> 01:02:55,265
SYSTEM VERY IMPORTANT.

1649
01:02:55,265 --> 01:02:58,868
YOU KNOW WE'VE DONE THIS IN THE

1650
01:02:58,868 --> 01:03:00,303
PAST, 1 POISONS FOR EXAMPLE IN

1651
01:03:00,303 --> 01:03:01,805
THE PRESENCE OF ATM DEFICIENCY

1652
01:03:01,805 --> 01:03:04,140
AND WE FOUND THERE, THAT'S

1653
01:03:04,140 --> 01:03:05,241
ENDURING IS ACTUALLY A PATHWAY

1654
01:03:05,241 --> 01:03:07,877
THAT ENDS UP KILLING THE CELLS,

1655
01:03:07,877 --> 01:03:09,746
THAT CELLS WITHOUT ALL THIS END

1656
01:03:09,746 --> 01:03:11,815
JOIPING IN THE ATM BACKGROUND

1657
01:03:11,815 --> 01:03:14,117
ARE MORE RESISTANT TO TOP O ICE

1658
01:03:14,117 --> 01:03:16,252
ON SOM RACE, AS WELL AS PARP

1659
01:03:16,252 --> 01:03:17,787
INHIBITORS AND MAYBE A SIMILAR

1660
01:03:17,787 --> 01:03:19,489
THING MIGHT BE HAPPENING IN THIS

1661
01:03:19,489 --> 01:03:26,963
ARENA AS WELL. 

1662
01:03:26,963 --> 01:03:27,764
I DON'T KNOW.

1663
01:03:27,764 --> 01:03:29,632
>> OKAY, WE HAVE A
QUESTION FROM

1664
01:03:29,632 --> 01:03:30,800
WILLIE JAMES, COULD YOU EXPAND

1665
01:03:30,800 --> 01:03:32,502
YOUR COMMENTS ON THE

1666
01:03:32,502 --> 01:03:33,770
PESFISSILITY OF THE CRISPR

1667
01:03:33,770 --> 01:03:38,641
TARGETING SYSTEMYING AS MEASURED

1668
01:03:38,641 --> 01:03:39,776
BY NG S?

1669
01:03:39,776 --> 01:03:40,377
>> RIGHT.

1670
01:03:40,377 --> 01:03:42,879
SO, NEXT GENERATION SEQUENCING,

1671
01:03:42,879 --> 01:03:45,682
YOU KNOW, NOW PIONEERED BY

1672
01:03:45,682 --> 01:03:47,317
LUMENNA, DEVELOPED BY MY

1673
01:03:47,317 --> 01:03:50,687
COLLEAGUE NEXT DORDOOR TO ME AND

1674
01:03:50,687 --> 01:03:53,223
THE NEXT DOOR LAB IS THE LAB OF

1675
01:03:53,223 --> 01:03:54,190
[INDISCERNIBLE] WHO TOGETHER

1676
01:03:54,190 --> 01:03:55,658
WITH DAVID GENERATED THIS

1677
01:03:55,658 --> 01:03:56,493
TECHNIQUE.

1678
01:03:56,493 --> 01:03:57,660
IT'S MIND BOGGLING WHAT 1 CAN DO

1679
01:03:57,660 --> 01:04:00,463
NOW WITH THIS TECHNIQUE IF YOU

1680
01:04:00,463 --> 01:04:01,664
HAVE A BIO-INFORMATICS GROUP AND

1681
01:04:01,664 --> 01:04:02,599
TESTIFIES LOOK SIDE.

1682
01:04:02,599 --> 01:04:03,867
SO IT'S DEEP SEQUENCING.

1683
01:04:03,867 --> 01:04:05,535
OBVIOUSLY WHEN YOU CARRY OUT A

1684
01:04:05,535 --> 01:04:06,503
CRISPR SCREEN, I THINK THIS IS

1685
01:04:06,503 --> 01:04:13,009
THE QUESTION, YOU HAVE A WHOLE

1686
01:04:13,009 --> 01:04:14,978
POPULATION, WE PROBE THE LIBRARY

1687
01:04:14,978 --> 01:04:16,513
AT 1000 OR 5000 X AND WE HAVE

1688
01:04:16,513 --> 01:04:20,016
BIG COVERAGE AND THEN YOU JUST

1689
01:04:20,016 --> 01:04:22,619
BASICALLY PC R THAT POPULATION,

1690
01:04:22,619 --> 01:04:24,587
AND THEN BY DEEP SEQUENCING YOU

1691
01:04:24,587 --> 01:04:26,890
JUST SEE HOW MANY READS YOU GET

1692
01:04:26,890 --> 01:04:28,625
PER GUIDE AND COMPARE THEM TO 1

1693
01:04:28,625 --> 01:04:28,992
ANOTHER.

1694
01:04:28,992 --> 01:04:30,493
AND THAT IS BASICALLY TELLING

1695
01:04:30,493 --> 01:04:31,561
YOU THE RELATIVE DISTRIBUTIONS

1696
01:04:31,561 --> 01:04:33,530
OF THOSE GUIDES IN THE

1697
01:04:33,530 --> 01:04:34,097
POPULATION.

1698
01:04:34,097 --> 01:04:35,665
IT'S NOT ENTIRELY ACCURATE, AND

1699
01:04:35,665 --> 01:04:37,600
OF COURSE, THERE'S GOING TO BE

1700
01:04:37,600 --> 01:04:43,006
SOME DRIFT XTHERE'S GOING TO BE

1701
01:04:43,006 --> 01:04:44,441
IN STOI CASTIC SCATTER BUT THE

1702
01:04:44,441 --> 01:04:46,509
LEVEL OF DATA SETS ARE

1703
01:04:46,509 --> 01:04:48,144
SUFFICIENT, THAT THESE DATA ARE

1704
01:04:48,144 --> 01:04:49,646
INCREASINGLY TIGHT.

1705
01:04:49,646 --> 01:04:50,580
AND IT'S VERY--IT'S ALMOST NEVER

1706
01:04:50,580 --> 01:04:53,983
NOW THAT WE GET A DESCENT HIT IN

1707
01:04:53,983 --> 01:04:55,952
A CRISPR SCREEN THAT SCNT

1708
01:04:55,952 --> 01:04:56,219
VALIDATE.

1709
01:04:56,219 --> 01:05:02,258
THEY ALMOST ALL DO.

1710
01:05:02,258 --> 01:05:03,593
>> WE HAVE OUR LAST
QUESTION

1711
01:05:03,593 --> 01:05:07,797
FROM JOHN [INDISCERNIBLE]. 

1712
01:05:07,797 --> 01:05:12,102
THERE'S ALREADY 54 L2 INFECTION

1713
01:05:12,102 --> 01:05:14,003
AFFECT THE TOPO2 ACTIVITY?

1714
01:05:14,003 --> 01:05:14,604
>> THAT'S GREAT QUESTION.

1715
01:05:14,604 --> 01:05:16,306
GREAT YOU ARE ON THE CALLMENT I

1716
01:05:16,306 --> 01:05:17,507
HOPE YOU WERE ATTENDING WHEN I

1717
01:05:17,507 --> 01:05:19,342
TALKED ABOUT OUR COLLABORATION

1718
01:05:19,342 --> 01:05:19,943
WITH YOU.

1719
01:05:19,943 --> 01:05:21,144
I THINK THATIA ANOTHER THING WE

1720
01:05:21,144 --> 01:05:27,217
SHOULD GO AHEAD AND LOOK AT IN

1721
01:05:27,217 --> 01:05:29,085
CELLS, WE HAVEN'T YET GONE ON TO

1722
01:05:29,085 --> 01:05:30,286
THAT BUT THAT'S ANOTHER THING.

1723
01:05:30,286 --> 01:05:31,254
AND MAYBE I WILL LOOK UP WITH

1724
01:05:31,254 --> 01:05:32,889
YOU TO SEE IF WE CAN ENGAGE WITH

1725
01:05:32,889 --> 01:05:37,894
YOU IN THOSE STUDIES AS WELL.

1726
01:05:37,894 --> 01:05:38,595
GREAT.

1727
01:05:38,595 --> 01:05:39,996
SO WITH TIME RESTRAIN, WE HAVE

1728
01:05:39,996 --> 01:05:41,698
TO WRAP UP OVER HERE, PLEASE

1729
01:05:41,698 --> 01:05:43,366
YOIN ME TO THANK DR. JACKSON

1730
01:05:43,366 --> 01:05:44,901
AGAIN FOR THE WONDERFUL TALK AND

1731
01:05:44,901 --> 01:05:47,871
WE ARE LOOKING FORWARD FOR YOUR

1732
01:05:47,871 --> 01:05:51,107
FUTURE WORKS IN THOSE GREAT

1733
01:05:51,107 --> 01:05:52,108
PUBLICATIONS.

1734
01:05:52,108 --> 01:05:53,176
>> THANKS AGAIN FOR THE

1735
01:05:53,176 --> 01:05:53,743
OPPORTUNITY TO TALK.

1736
01:05:53,743 --> 01:05:55,478
IT'S BEEN A PLEASURE AND IT'S

1737
01:05:55,478 --> 01:05:58,848
GREAT MEETING EVERYBODY AGAIN.

1738
01:05:58,848 --> 01:05:59,082
THANKS.

1739
01:05:59,082 --> 01:06:00,183
>> THANK YOU. 

1740
01:06:00,183 --> 01:06:02,152
>> THANK YOU.

1741
01:06:02,152 --> 01:06:12,395
>> BYE-BYE.