THANKS FOR COMING IN PERSON
AND THE NUMBER IS 7657 AND THERE
WILL BE SLIDE.
MY NAME IS JOHN.
I'M A PROGRAM OFFICER AT NIDA
AND MEMBER OF THE ECONOMIST FOR
THE PROGRAM AND ON BEHALF OF THE
PROGRAM JENNIFER PHILLIPS
CREMINS IS AN ASSOCIATE
PROFESSOR IN GENETICS AT THE
UNIVERSITY OF PENNSYLVANIA AND
SHE OBTAINED AN NIH NEW
INNOVATOR AWARD AND NUCLEOME
AWARD AND THE PRESTIGIOUS NIH
PIONEER AWARD AMONG OTHER
GRANTS.
THAT'S THE EXAMPLE OF HOW NIH
AND THE COMMON FUND CAN SERVE AS
AN INCUBATOR SPATE FOR TALENT.
TODAY SHE'LL TELL US ABOUT THE
SCIENCE OF CONNECTIONS BRIDGING
CHROMATIN FOLDING AND SYNAPTIC
PHYSIOLOGY.
TAKE IT AWAY, JEN.
>> HELLO, EVERYONE.
IT'S A GREAT PRIF LEN --
PRIVILEGE TO BE HERE AND WANT I
WANT TO THANK FOR THE INVITATION
FOR THE LECTURE.
I'M GOING TO TALK TO ABOUT
MEMORY.
IT'S IMPORTANT TO ALL OF US AND
IT UNDER LIES OUR ABILITY TO
READ, WRITE, ACQUIRE NEW SKILLS,
MAINTAIN RELATIONSHIPS.
IT'S AN IMPORTANT QUESTION
BECAUSE IT'S A UNIFIER OF THE
HUMAN CONDITION.
NO MATTER WHAT BACKGROUND WE
HOLD SPACE WITH US TO CARRY
WONDERFUL MEMORIES AND DIFFICULT
EVENTS LIKE ISOLATION, LOSS OF A
LOVED ONE OR TRAUMA.
LOSS OF A MEMORY UNDER LIES
DISEASES LIKE ALZHEIMER'S AND
EXACERBATION OR RESILIENCE IS
CRITICAL FOR PTSD.
SO WHAT ENCODES MEMORY?
ONE LEADING LINE OF THOUGHT IS
IN THE LIES IN NEURONS AND THE
CONNECTIONS BETWEEN THEM AND
SYNAPSES AND THE MAMMALIAN BRAIN
THERE'S BILLIONS OF NEURONS
CONNECTED BY TRILLIONS OF
SYNAPSES AND WHAT'S SO
INTERESTING IS THEY'RE NOT
STATIC BUT RATHER DYNAMICALLY
ALTERED IN A CONTINUOUS MANNER
ACROSS THE LIFE TIME OF AN
INDIVIDUAL IN RESPONSE TO
EXPERIENCES.
SO I'D LIKE TO SHARE THIS VIDEO
OF DATA FROM JIM McGEE'S LAB
BECAUSE IT HIGHLIGHTS THE
DIFFICULTY AND EXCITEMENT OF THE
PROBLEM SPACE FOR SCIENTISTS
STUDYING MEMORY.
THEY'RE USING CALCIUM CENTER TO
TRACK LONGITUDINALLY POTENTIAL
FIRING IN NEURONS IN THE MOUSE
BRAIN.
THE MOUSE IS IN THE DARK AND
EXPOSED TO A SENSORY STIMULUS.
THIS HIGHLIGHTS HOW DIFFICULT
IT'S GOING TO BE TO UNDERSTAND
WHAT UNDER LIES AND MAINTAINS
MEMORY OVER THE LONG TERM
BECAUSE CLEARLY THE NEURONS ARE
CHANGING OVER TIME AND EVEN THE
SAME NEURON MIGHT BE HAVING
DIFFERENT PATTERNS OF
STIMULATION TEMPORALLY.
SO WE LIKE TO TAKE WHAT I CALL A
MULTI-SCALE APPROACH.
THE IDEA IS THE BRAIN IS
REGULATED AT MANY DIFFERENT
BIOLOGICAL SCALES AND PART OF
THE COMPLICATION IN FINDING
PATTERNS THAT LEAD TO BIOLOGICAL
INSIGHT THROUGH ALL THESE
CHALLENGES IS THE SCALES ALSO
MUST BE INTEGRATED TO UNDERSTAND
HOW NEURAL CIRCUITS WORK.
OF COURSE THERE'S LARGE SCALE
ANATOMY AND PHYSICAL CONNECTIONS
BETWEEN NEURONS AND LABELLING OF
N GRAMS AND PREVIOUS FIRING OF
ACTION POTENTIALS AND
PHYSIOLOGICAL PATTERNS STRUCTURE
AND BIO CHEMICAL AND SYNAPTIC
PLASTICITY AND NUCLEUS WHERE WE
THINK POST TRANSLATIONAL
CHROMATIN AND FIBERS AND
VARIATION IN THE GENETIC
SEQUENCE WHICH MAY CONTRIBUTE TO
DIFFERENCES IN CIRCUITS.
WE AIM TO STUDY TWO OR THREE
MODEL SYSTEMS AND BEGIN TO
GATHER THE INFORMATION IN THE
MODEL SYSTEMS AND INTEGRATE TO
LEAD TO INSIGHT.
THIS IS WHERE WE'RE GOING, GOING
FORWARD.
I HAVE TO SAY OVER THE FIRST 10
YEARS OF THE LAB FOCUSES ON THE
NUCLEUS AND HOW HEADING OUT TO
THE SYNAPSE.
WHERE WE FOCUSSED IS ON
UNDERSTANDING SO-CALLED
EPIGENETIC OR CHROMATIN
MECHANISMS TO DNA OR HISTONES
DYNAMICALLY ALTERED THROUGH THE
LIFE TIME OF A CIRCUIT.
THE CLASSICAL APPROACH OF WHAT
CHROMATIN MARKS IS FUNCTIONAL IS
TO DO A TYPE OF GENOME EDITING
AND LOOK AT HOW THE LOSS OF
FUNCTION MUTATION AFFECT
EXPRESSION OF A NEARBY GENE.
WE'VE TAKEN WHAT I CALLED A
STRUCTURED APPROACH IN WHICH WE
HAVE MAPPED THE HIGHER ORDER
FOLDING WITHIN THE NUCLEUS AND
COMPUTATIONALLY IDENTIFIED THE
SPATIAL OR PHYSICAL TOUCH DOWN
POINTS BETWEEN DISTAL DNA
FRAGMENTS AND STUDIED HOW GENES
ARE REGULATED FROM LONG RANGE
MECHANISMS.
SO, WHEN I STARTED IN THE FIELD
AS A POSTDOC BACK IN 2008 OR SO,
THIS WAS THE TECHNOLOGY AND
WHERE THE FIELD WAS AT THAT
TIME.
WE COULD USE A VERSION OF DNA
PHISH TO DETERMINE THERE WAS
INDEPENDENT LOCATIONS IN THE
NUCLEUS.
AT THE OTHER END OF THE SPECTRUM
USE DNA PHISH AND PAIR-WISE
PROBES TO LOOK AT FOCAL DOTS IN
THE NUCLEUS AND BEGIN TO MAKE
INFERENCE OF THE OVERLAP OF THE
DOTS AND IT WOULD REPRESENT A
LONG-RANGE INTERACTION BETWEEN
TWO DISTAL FRAGMENTS.
EVERYTHING ELSE IN BETWEEN WAS
THEORETICAL OR ABSTRACTION OF
POSSIBLE FOLDING PATTERNS.
AND SO WHAT I THINK I'D LIKE TO
SAY HERE IS THAT OVER THE LAST
10 YEARS OR SO IT'S BEEN A WILD
RIDE I THINK FROM ALL COLLEAGUES
IN THE 40 NUCLEOME AND HAVE
FILLED IN THE GAPS CONFIRMING
TERRITORIES AND ALSO FINDING AB
COMPARTMENTS, SUBCOMPARTS AND
NANO COMPARTMENTS TOP LOGICALLY
ASSOCIATED DOMAINS AND TAGS
WITHIN TAGS AND EVEN NANO TAGS
AND LONG-RANGE LOOPING
INTERACTIONS.
SO FOR THOSE THAT ARE IN A
BROADER AUDIENCE I WANTED TO
SHOW THE TYPE OF DATA THAT'S
BEEN ABLE TO DRIVE DISCOVERIES
IN THE FIELD OVER THE LAST 10
YEARS.
THIS IS A SO-CALLED CHROMOSOME
CONFIRMATION CAPTURE MAP THROUGH
PRINCIPLES OF PROXIMAL LOCATION
ORIGINALLY BACK AROUND 2002.
THE IDEA IS IF YOU POST THE
GENOME FRAGMENT OF INTEREST ON
BOTH THE X AND THE Y AXIS, ONE
CAN LOOK AT THE PIXEL
INTERSECTING THE PARTICULAR
GENOMIC LOCI AND LOOK AT THE
READS IN THE PIXEL.
THAT REPRESENT THE NUMBER OF
TIMES THOSE TWO PARTICULAR
FRAGMENTS WERE DETECTS AS
ARTIFICIALLY
>>Y GLUED TOGETHER IN
THE PHYSICAL OR NUCLEAR SPACE.
WHAT ONE CAN SEE IMMEDIATELY IS
THERE'S EXCITING AND ALSO VERY
DIFFICULT TO UNDERSTAND FOLDING
PATTERNS THAT THESE MAPS
REVEALED.
AND ONE I WANT TO HIGHLIGHT HAVE
THE SO-CALLED FOCAL DOTS.
THEY HAVE BEEN VERY DIFFICULT
WORK TO DETECT AND ANALYZE AND
NOW WE HAVE THE ABILITY TO
DETECT THEM GENOME WIDE AND THEY
ARE REPRESENT BIO LOGICALLY IS
AN INTERACTION WHERE THE TWO
DISTAL FRAGMENTS CONTACT EACH
OTHER AND THROUGH LOOP EXTRUSION
WHERE AN ATP DEPENDENT MANNER
MEDIATEDS THE BOUNDARIES.
WHEN YOU SEE FOCAL DOTS OF THIS
NATURE THEY'RE IMPORTANT BECAUSE
THEY REPRESENT A SO-CALLED LONG
RANGE LOOPING INTERACTION.
WHEN MY LAP STARTED WE COULD SEE
THE TECHNOLOGY WAS ADVANCING
RAPIDLY AND KNEW REFERENCE MAPS
WERE BEGINNING TO BE CREATED A
WIDE RANGE OF CELL TYPES.
THE QUESTION WE HAD IS WHAT IF
WE SORT OF MOVED TO THE NEXT
THING THAT WAS GOING TO BE
DIFFICULT AND THAT IS IF WE KNOW
THE GENOME FOLDS INTO MANY
COMPLICATED STRUCTURES HOW DO WE
FIGURE OUT WHAT ASPECTS OF
STRUCTURE ARE FUNCTIONAL AND
GIVE INSIGHT TO WHAT THE GENOME
IS DOING?
I WANTED TO SHARE THIS CAME OUT
IN THE MEETING WITH THE STUDENTS
AND POSTDOCS OVER LUNCH, HOW
DOES ONE PICK A BIOLOGICAL
PROBLEM AND FOR US WHAT WE TRIED
TO DO IS FIND A QUESTION
TIMELESS AND SO I'D LIKE TO USE
THIS EXAMPLE OF THE EARLIEST
POSSIBLE REFERENCE I COULD FIND
FROM THOMAS HUNT MORGAN BACK
WHEN NATURE PAPERS WERE SINGLE
PAGE AND ONE LAND DRAWN FIGURE.
THERE WAS NO SOCIAL MEDIA BUT
CLEARLY NO ABSENCE OF SNARK AND
THERE WAS AN ARRANGEMENT OF
LINKED GENES IN THREE DIMENSIONS
BASED ON THE ASSUMPTION THE
DISTANCE BETWEEN TWO LOCI IS
PROPORTIONAL TO THE OBSERVED
CROSS OVER VALUE AND BELIEVES
THERE'S A BETTER AGREEMENT
BETWEEN OBSERVATION AND
EXPECTATION THAN THE HYPOTHESIS
OF A STRICTLY LINEAR ARRANGEMENT
WE HAVE DEVELOPED.
SO I THINK PROBABLY JOHN CAN
SPEAK TO THIS AS WELL AS MANY IN
THE 3-D GENOME FIELD I SEE PEDRO
HERE, THESE TYPES OF DEBATES ARE
STILL ONGOING AT THIS EXACT TIME
ADVANCING ALL THE WAY TO 2024
AND WILL CERTAINLY BE HAPPENING
AT THE CLOSING MEETING OF THE 40
NUCELEOME PHASE 2 AND THERE'S SO
MUCH FOR THE NEXT WAVE OF
SCIENTISTS AFTER US.
AFTER THE YEARS WE'VE CONDUCTED
A VARIETY OF STUDIES.
FIRST IN MODELS OF NEURONS
THROUGH NEURAL DIFFERENTIATION
AND REPROGRAMMING AND MORE
RECENTLY NEURAL CIRCUIT
STIMULATION IN VITRO AND IN
VIVO.
INDEED LOOPS ARE HIGHLY DYNAMIC
ACROSS ALL THE SYSTEMS.
MOST IMPORTANTLY CHANGING ON
FAST TIME SCALES DURING
ACTIVATION OF NEURAL CIRCUITS
AND ALSO NECESSARY AND ESSENTIAL
FOR THE ACTIVATION OF CRITICAL
SYNAPTIC, IMMEDIATE, EARLY AND
DELAYED RESPONSE GENES IN VIVO
AND IN VITRO AND PROTEINS TO GET
AT THE GENOME STRUCTURE
RELATIONSHIP RAND THIS IS AN
EXAMPLE WHERE WE CREATED
SOMETHING CALLED LIGHT ACTIVATED
DYNAMIC LOOPING TO FORM LOOPS ON
DEMAND AND ASK IF THEY WERE
SUFFICIENT FOR EN LANCER
PROMOTER AND -- ENHANCER AND
GENE EXPRESSION AND WE WORKED
WITH THE GILBERT LAB TO
UNDERSTAND COHESIVE MEDIATED
LOOP EXTRUSION WAS NEEDED FOR
LOCATIONS IN THE GENOME IN EARLY
S PHASE.
THESE ARE ALL ALUMNI WHO WORKED
ON THE EARLY STAGES AND THEY NOW
HAVE HIGH FLYING CAREERS IN
START-UP COMPANIES IN ARTIFICIAL
INTELLIGENCE AS WELL AS IN THE
DEVELOPMENT OF THERAPEUTICS.
GOOD KIM AND REGE BOTH MY FIRST
POSTDOCS IN THE LAB AND NOW EACH
RUN LABORATORIES IN SOUTH KOREA
AS WELL AS IN MUMBAI AND IN THE
REPLICATION STUDY DR. EMMERSON
IS A POSTDOC AND DR. COOK AT THE
FDA AND OTHERS ARE IN THE JOB
MARKET AND FABULOUS
COMPUTATIONAL BIOLOGISTS.
ANOTHER THING WE'VE DONE IN THE
RECENT FUTURE IS THINK OF
DISEASE AS NATURE'S PERTURBATION
AND WONDER WHEN WE BROKE THE 3-D
GENOME WHAT WOULD HAPPEN TO THE
GENE EXPRESSION IN A PATH LOGIC
SITUATION.
WE CHOSE WHAT WE THOUGHT WAS A
DISORDER KNOWN IN THE TEXTBOOKS
TO BE DRIVEN BY A SINGLE
MUTATION RAY SHORT REPEAT TRACK
IN THE FML1 GENE IN THE UTR.
FOR REASONS THAT ARE STILL
POORLY UNDERSTAND, THIS SEQUENCE
GROWS OUT OF CONTROL.
IT REACHES A LENGTH OF 200CGGs
AND THEN BECOMES FULL MUTATION
AND AT THAT TIME THERE IS
SO-CALLED EPIGENETIC SILENCING
OF THE FMR1 GENE AND QUITE
SEVERE DISEASE.
THE TEAM FOUND IN THE SYNDROME
THERE'S ISN'T JUST A LOCAL
SILENCING OF THE FMR1 BUT IN THE
PCs AND NEURONS THAT HAVE THIS
FRAGILE DOMAINS AMONG AUTOSOME.
THEY COME TOGETHER TO PHYSICALLY
CONTACT EACH OTHER AND THE GENES
UNDER THE LOCATIONS ARE LARGELY
SYNAPTIC GENES SILENCED AND
LINKED TO FRAGILE X AND AUTISM.
WHEN WE LOOK AT THE BASE OF
THOSE PARTICULAR GENOMIC
COORDINATES WE FOUND GENOME
INSTABILITY WE DIDN'T EXPECT IN
A MONOGENIC DISORDER AND CALL
THEM BEACONS OF REPEAT EXPANSION
BY CONTACTING HETEROCROW
PLAT -- HOTERO CHROMATIN AND A
GROUP IN A RARE FORM OF
PEDIATRIC BRAIN CANCER FOUND A
SIMILAR PATTERN THEY ARE CALL
TULIPS.
THE SAME IDEA THERE'S LARGE
SCALE HETEROCHROMATINS AND THINK
IT WILL CAN MEAN WE HAVE NEW
IDEAS FOR DRUGGABLE TARGETS IN
THUS FAR INTRACTABLE
NEUROLOGICAL DISORDERS.
THIS WORK WAS LED BY A TEAM OF
WONDERFUL PEOPLE WHO ARE STILL
IN THE LAB.
AND DURING THE PANDEMIC I THINK
WE ALL WENT THROUGH SOME FORM OF
EXISTENTIAL CRISIS AND I STARTED
TO THINK ABOUT THE QUESTIONS THE
LAB WANTS TO THINK IF GOING
FORWARD ONCE WE'RE ALL ABLE TO
GO BACK TOGETHER AND I WAS
IMPACTED BY THE BOOK WRITTEN BY
ERIC KANDEL AND TEACHES PEOPLE
THE SEQUENCE OF EVENTS OR
HISTORICAL EVENTS OF MANY OF THE
CORE DISCOVERIES.
AND HAD US THINKING ABOUT THE
CONCEPT OF MEMORY AND CERTAINLY
THE FOCUS HERE WAS A LOT OF ON
SYNAPTIC PLASTICITY AND THOUGHT
THERE'D BE IMPORTANT WAYS WHERE
CRHROMATIN CAN PLAY A ROLE AND
THEY'RE SCORED ON TIME SCALES OF
DECADES AND BIO MOLECULES, RNA
PROTEINS, ETCETERA, HAVE HALF
LIVES ON THE MANNER OF HOURS UP
TO A FEW DAYS.
SO THERE HAS TO BE A CONNECTING
POINT THAT MAINTAINS THINGS IN
THE LONG TERM.
ONE THING I WANTED TO SHARE IS
OFTEN TIMES THE CLASSIC STUDIES
OF NEURAL PHYSIOLOGY AND
SYNAPTIC PLASTICITY MAY OVERLAP
THE CONTRIBUTION OF CRHROMATIN
AND IF WE CONNECT THE TIME
SCALES WE CAN STUDY HOW THE
NUCLEUS AND SYNAPSE INTERPLAY TO
KEEP THINGS GOING IN THE LONG
TERM.
THERE'S CRITICAL DATA THROUGHOUT
THE FIELD THAT FORM THE
FOUNDATION FOR THIS IDEA.
THERE'S A CLASSIC EM STUDY
LOOKING AT A SINGLE NUCLEUS FROM
A MAMMALIAN NEURON AND WE SEE
MARK RESTRUCTURING OF CRHROMATI
AND WE SEE RECOVERY AS ONE MIGHT
EXPECT BUT THERE ARE POCKETS
THAT REMAIN THAT OF COURSE WE
DON'T KNOW FROM THIS IMAGE WHAT
THAT GENETIC SEQUENCE IS BUT
WE'RE EXCITED TO FIGURE THIS
OUT.
THE OTHER FOUNDATIONAL DATA IS
THERE'S LESSER KNOWN STUDIES
THAT HAVE LOOKED MULTIPLE YEARS
AGO NOW ALMOST 10 YEARS OF CDC
KNOCKOUT MICE AND ANIMALS AND
KNOW THEY ARE HAVE DEFECTS IN
GENOME WIDE FOLDING AND THEY
DEMONSTRATED THERE WAS SEVERE
LEARNING DEFECTS BOTH INABILITY
OF ANIMALS TO LEARN THE WATER
AND FEAR CONDITIONING.
AT THE NEURAL PHYSIOLOGICAL
LEVEL THERE WERE LONG-TERM
DEFECTS IN POTENTIATION AND AT
THE CELLULAR LEVEL THE KNOCKOUT
LEADS TO DECREASED IN DENDRITIC
AND SPINE SIZE AND THE DEFECTS
ARE ACROSS SCALES AND WE THINK
THERE'S A FOUNDATIONAL BASIS FOR
THIS.
THE OTHER IMPORTANT POINT IS
JUST THAT IN THE CLASSIC
LITERATURE AND SUMMARIZED BY
KANDEL NOT ONLY DOES ANY FORM OF
ACTION POTENTIAL LEAD TO CHANGES
AT THE SYNAPSE BUT RAPID CHANGES
IN THE NUCLEUS AT THE PRESYNAPSE
THERE'S RAY RELEASE OF NEURAL
TRANSMITTERS SUCH AS DOPAMINE OR
GLUTAMATE AND LEADING TO
STRENGTHENING OF SPECIFIC
SYNAPSES OR SPINES AND INFLUX OF
CALCIUM GOING RAPIDLY TO THE
NUCLEUS TO FIRE AND SECONDARY
RESPONSE GENES FOLLOW SHORTLY
THEREAFTER.
ONE EXCITING THOUGHT IN THE
FIELD IS OF SYNAPTIC TAGGING AND
CAPTURE.
THE IDEA IS DURING THE LEARNING
THERE'S THE EVER ELUSIVE BIO
CHEMICAL SYNAPSE AND MARKS THE
SPINES AND TRANSCRIPTIONAL
PROGRAMS OR CERTAIN RNAs GO BACK
TO THE SPINE AND LEAD TO THE BIO
CHEMICAL MECHANISMS THAT
PROPAGATE THE TAGS OVER THE LONG
TERM.
WE'RE EXCITED TO WORK ON THIS
PROBLEM AND FIGURE OUT SOME
DETAILS AMONG OTHER COLLEAGUES
ALREADY INVESTED AND WORKING ON
THE PROBLEM.
AND WE BOILED IT DOWN TO THREE
THINGS WE FIRST HAVE TO PROVE TO
OURSELVES.
FIRST IS FIND SOMETHING IN THE
CHROMATIN THAT'S CONSISTENT AND
REMAIN FOR A LONG TIME UPWARDS
OF MONTHS OR MAYBE EVEN YEARS.
THE SECOND IS WHATEVER THE
PERSISTENT THING IT HAS TO GIVE
A FLASHLIGHT OR BEACON TO WHAT
RNAs MAY BE CHANGING AND QUAN WE
FIND THE ONES BACK IT OUR
SYNAPSE AND WE'D HAVE TO
COLLABORATE WHAT IS GOING ON FOR
THE PROPAGATION OF THE TAG BUT
THINK WE HAVE TO FIND BOTH
THINGS AND DISRUPTED TO SOME
DEGREE IN A DISORDER WHERE
THERE'S LOSS OR GAIN OF MEMORY.
THE LAB IS DIVIDED AMONG FOUR
AREAS.
WE'RE STUDYING HOW CRHROMATIN I
CHANGED IN IN VIVO AND LEARNING
IN IN VITRO WE'RE BUILDING
IMAGING TECHNOLOGIES TO CREATE
OVER MANY YEARS SYNAPSE
RESOLUTION ACTIVITY DEPENDENT
NEURAL CONNECTOME MAPS AND OTHER
CONNECTIVITY PROBLEM AND STUDY
NUCLEUS TO SYNAPSE.
USING OUR TECHNOLOGIES TO
UNDERSTAND WHICH RNAs HAD BACK
OUT FROM NUCLEUS TO THE FINE AND
ARE THEY ARE FUNCTIONING FOR
BEHAVIOR AND IS THE GENOME
STRUCTURE MISWIRED IN
NEUROLOGICAL DISORDERS.
ONE WORK IS HIGHLIGHTING THINGS
WE PUBLISHED IN THE AREA OF
NEURAL STIMULATION.
AND SHOW YOU NEW SINGLE CELL
WORK FROM MY GROUP.
AND ALSO SUMMARIZE AN ANY STORY
OF LOSS OF MEMORY IN THE DISEASE
OF ALZHEIMER'S.
SO THE OTHER UNTIL WORK IN MY
LAB IN NEURAL STIMULATION WAS
LED BY JOHN BEACON AND UPON
CALCIUM INFLUX WHEN ACTION
POTENTIAL FIRED THERE'S THE
EARLY GENES AND SECONDARY
RESPONSE GENES.
FOR CHROMATIN STRUCTURE PEOPLE
IT'S EXCITING BECAUSE THINGS
HAPPEN SO FAST.
SYNAPTIC PLASTICITY IN TIME
SCALES OF MILLISECONDS AND
SECONDARY RESPONSE AGAINST ON
HOURS TO A DAY.
THIS HELPS GET FASTER Ph.D.s
BECAUSE THE STUDENTS AREN'T
WAITING TO ARE A YEAR TO PROCESS
A SAMPLE AND LAD NO PROBLEM TO
RECRUITING TO THIS PARTICULAR
PROJECT.
THIS IS ONE EXAMPLE OF DATA
JOHN CREATED AND INDUCERS AS
FUNCTIONALLY REPRODUCED BY MANY
NEUROSCIENT SCIENSCIENTISTS IN D
IT STAYS UP TO 60 MONTHS AND
THEN BEGINS TO DISSIPATE AS THE
GENE IS TURNED OFF.
I WANTED TO SHOW THIS TIME
COURSE BECAUSE IN THE EARLY DAYS
WE REALLY HAD NO IDEA IF LOOPS
MATTERED FOR LOOP EXPRESSION AND
WAITED UP AGAINST THE GENE
EXPRESSION AND LOOPS FORM
QUICKER THAN mRNA LEVELS AND
NEVER WENT BACK TO LOOK EVEN
EARLIER.
SO WE KNOW THEY FORM FAST AND
THEY'RE ACTIVITY DEPENDENT.
I'M REDUCING PERHAPS FOUR YEARS
TO A SINGLE SLIDE BUT ALSO SAW
PATTERNS OF GENE EXPRESSION AND
ACTIVATED SUPER FAST AND TURNED
OFF JUST AS FAST.
BY CONTRAST THE SECONDARY
RESPONSE TEAM FORMS A COMPLEX
NETWORK OF MULTIPLE LOOPING
INTERACTIONS AND THINK IT MIGHT
CONTRIBUTE TO THE FURTHER
DELAYED GENE EXPRESSION.
AND SO MORE LOOPS, LONGER LOOPS
AND MORE ENHANCERS AT LEAST IN A
NEURAL ACTIVATION PARADIGM OFTEN
LEADS TO DELAYED DYNAMIC LOOP
EXPRESSION.
ANOTHER QUESTION IS DO LOOPS
MATTER FOR GENE EXPRESSION AND
THIS IS CRITICAL WORK KICKED OFF
BY AIDAN AND LIDS COLLEAGUES AND
OTHERS GOING UP TO EIGHT YEARS
AGO.
IN THIS STUDY THERE WAS A DEGRON
USED IN CANCER CELL LINES.
COHESIN WAS TAKEN AND ALL THE
LOOPS WERE GONE.
WHAT SHOCKED THE FIELD AND LED
TO DISMAY IS LOOKING AT THE GENE
EXPRESSION THERE WAS ESSENTIALLY
NO CHANGE WHETHER THE CANCER
CELL LINE LAD LOOPS OR NOT.
SO LOOPS DON'T METER FOR GREEN
EXPRESSION MAINTENANCE AT LEAST
IN THIS SYSTEM.
ONE IDEA WE MHAD IS WE HAD TO
THINK OF THE ESTABLISHMENT OF
NEW GENE EXPRESSION PATTERNS.
WE TEAMED UP WITH MATHIAS AND WE
ACTIVATED THE NEURONS AND
ASSESSED DO THEY ARE HAVE THE
ABILITY TO ACTIVATE THE
IMMEDIATE EARLY GENES AND SECOND
RESPONSE GENES AND I'M SUMMARIZE
AND THE IMMEDIATE EARLY GENES
TURNED ON JUST FINE EVEN WHEN
LOOPS WERE GONE, HOWEVER A LARGE
RANGE OF PLASTICITY AND RESPONSE
GENES HAD NO ABILITY TO TURN ON
ONCE WE ASKED THE NEURONS TO DO
SOMETHING NEW.
MAYBE THEY'RE JUST NOT NEEDED TO
MAIN TRAIN WHAT YOU HAVE BUT
NEED TO GO SOMEWHERE NEW AS A
CELL YOU NEED NEW LOOPING
INTERACTIONS TO GET TO WHERE YOU
WANT TO GO NEXT.
SO WHERE ARE WE GOING NOW?
WE'RE WORKING AS LAB WITH TWO
AMAZING NEURO BIOLOGISTS AND
SCIENTISTS.
WE WANTED TO GO IN VIVO AND USE
CONTEXTUAL FEAR CONDITIONING AND
THE IDEA IS EXPOSE THE MOUSE TO
A NEUTRAL AND AVERSIVE STIMULUS
IN THIS CASE SHOCK AND LOOK FOR
CHROMATIN PERSISTENCE AND HAVE A
SPAN OF DAYS TO A MONTH RAND
HAVE RECALL TO EXPOSE THE MOUSE
TO A MONTH IN AND RAY ONE-SHOT
BEHAVIORAL DEAL AND HAVE
ISOLATION IN VARIOUS PARTS OF
THE BRAIN RELEVANT FOR ENCODING,
CONSOLIDATION OR RECALL OF FIELD
MEMORY.
HOW DO WE GET ALL THE
COMPLICATED PRAT EARNS AND GET
THE NEURONS AT THE EXACT RIGHT
PERIOD OF TIME?
WE USED AN EXCITING AND ELEGANT
MOUSE MODEL CALLED THE TRAP 2
MOUSE.
WHAT THIS MOUSE DOES IN BRIEF IS
USES AN ACTIVITY-DRIVEN CRAY TO
GENETICALLY MARK IN A PERMANENT
MANNER THE NEURONS FIRED ONLY
DURING THE EXACT WINDOW OF THE
BEHAVIOR DURING THE INJECTION.
THEN SUBSEQUENTLY WE CAN WAIT
LONG PERIOD OF TIME AND SORT OUT
THE NUCLEAR ALMOST A MONTH LATER
BECAUSE THEY ARE WERE ACTIVE
ONLY TO OUR KNOWLEDGE BACK AT
THE ORIGINAL ENCODING.
WE USED A NEW MULTIMODAL
TECHNOLOGY CALLED SINGLE NUCLEUS
3C SEQ AND WHAT WE CAN DO IS NOW
CLU
CLUSTER THE CELLS AND GRAB ALL
THE NEURONS THAT LOOK EXACTLY
LIKE THEY HAVE THE SAME
METHYLATION PROFILE AND MATCHED
THE CELL TYPE AND CREATE ONE
LESS SPARSE AND NOW THERE'S A
PROXY PAUSE IT'S A PSEUDO BULK
DATA AND THIS IS HOT OFF THE
PRESS OF TWO YEARS OF EFFORT.
THIS IS AROUND THE MMD RAF
RECEPTOR GENE AND THERE'S NO
LOOP FORM IN THE NEURONS IN THE
CONTROL AND SEE IT FORMED IN THE
TRAPPED NEURONS.
IT REMAINS AT THREE DAYS AND
THROUGH DAY 28.
THIS IS JUST ONE EXAMPLE JUST IN
THE LAST FEW DAYS WHERE WE THINK
IS A LAB WE ARE GOING TO BE ABLE
TO HAVE THE POTENTIAL TO SEARCH
FOR PERSISTENT LOOPING
INTERACTIONS AND THE HYPOTHESIS
WE'RE CHASING OR TRYING TO DID
PROF IS DURING MEMORY ENCODING
THERE'S DEPENDENT LOOPS THAT
STAY STUCK AND PERSISTENT AND
REMAIN OVER LONG PERIOD OF TIME
EVEN IF THE GENE EXPRESSION
COMES BACK DOWN.
AND WHEN THAT NEURON IS CALLED
PON UPON THERE'S A FASTER
RESPONSE AND HAVE TO FIGURE OUT
THE FUNCTIONALITY OF HOW THAT
ENABLES THIS EXPRESSION DURING
RECALL.
JUST TO SUMMARIZE THIS PART
BEFORE GOING TO THE LAST SESSION
WE FIND IN NEURONS MANY LOOPS
CONNECT IN ACTIVITY OF GENES
DURING STIMULATION AND EN
HANCERS AND SEE THEIR COHESIN
INDEPENDENT AND FIND A COMPLEX
NETWORK OF LONG DISTANCE LOOPS
THAT CONNECT ON LATER TIME
SCALES AND COHESIN DEPENDENT
LOOPS NEEDED FOR MATURATION AND
ACTIVITY DEPENDENT SECONDARY
RESPONSE IN VITRO.
SO FOR THE QUESTION OF DO LOOPS
MATTER IN POST MITOTIC
EXPRESSION THEY ARE DO AND WE
ALSO FIND THROUGH NEW
PRELIMINARY DATA I'VE SHOWN
PERSISTENT LOOPS AND N GRAMS
PLAY EXIST AND WE'RE CONTINUING
TO PURSUE THAT THROUGH THE FINAL
YEAR OF THE 4D NUCLEOME AND NOW
THINKING ABOUT WHAT ABOUT IN A
DISEASE WHERE THERE'S LOSS OF
MEMORY?
FOR THAT WE TURN TO ALZHEIMER'S
DISEASE THERE'S MANY EXPERTS AT
THE NIH I HAD THE GREAT FORTUNE
OF DISCUSSING THIS EARLIER THIS
MORNING AND THE MOST COMMON FORM
OF DEMENTIA AND PLAYS OUT
THROUGH COMPLEX
NEURODEGENERATION.
THE LARGE MAJORITY AND DRIVEN BY
RARE MUTATIONS.
THE COMMONALITY IS THEY ARE
OFTEN HAVE SIMILARITY IN BIO
CHEMICAL HALLMARKS OF AGGREGATES
OF AMYLOID OR
HYPERPHOSPHORYLATION OF TAU.
SO SOMETHING LESS APPRECIATED IS
THAT MULTIPLE CELLS TYPES ALSO
HAVE SEVERE TRANSCRIPTIONAL
DEFECTS AND IT'S LATELY COME
INTO FASHION BECAUSE OF STUDIES
IN POSTMORTEM BRAIN TISSUE AND
WHAT FORMS THE DYSREGULATION?
WHERE IS THERE'S LESS INSIGHT?
MANY STUDIES AND RIGHTFULLY SO
ARE FOCUSED ON POST MORTEM BRAIN
TISSUE SO IT'S DIFFICULT TO GET
BACK TO THE EARLY EVENTS.
MOLECULAR AND CELLULAR DEFECTS
THAT OCCUR DECADES BEFORE THE
CLINICAL PRESENTATIONS WILL BE
MISSED BY FOCUSSING ON THE LATE
STAGE.
LATE STAGE AGAIN MISSES EARLY
DRIVERS OF THE DISEASE AND FOR
SPORADIC ALZHEIMER'S, IPS
STUDIES ARE DIFFICULT BECAUSE
THE DEGENERATION IS LATE ONSET
AND MORE SPORADIC AND VARIABLE
ACROSS INDIVIDUALS.
VERY TECHNICALLY CHALLENGING TO
COVER THE DETAILS IN A
REPRODUCIBLE WAY THAT ARE
REPRESENTING A FETAL STAGE.
OUR GOAL IS TO SEE IF THERE'S
CHROMATIN DYSREGULATION DURING
EARLY ONSET AND BEFORE OBVIOUS
PRESENCE OF AMYLOID PLAQUE OR
TAU TANGLES.
THIS WAS WORK WAS BY AN AMAZING
COMPUTATIONAL BIOLOGIST AND THE
TEAM AND TO DO THIS WORK WE
CREATED IPS LINES OF ALZHEIMER'S
AND WANTED TO FOCUS ON FAMILIAL
ONSET TO GET THE EARLY
MECHANISM.
AND SHE CREATED THE STABLE LINES
IN WHICH THERE'S ISOGEN
BASICALLY MATCHED FOR THE
BACKGROUND IN BOTH ALLELES SO
HOMOZYGOUS AND USED A 2-D
PROTOCOL DOMINICK AND HIS TEAM
OPTIMIZED TO GET A PURE
POPULATION OF EXCITATORY
NEURONS.
IN PARALLEL WE CREATED BRAIN
ORGANOIDS THROUGH THE PROTOCOL
TO LOOK AT 2-D AND 3-D MODELS.
WHAT WAS IMPORTANT WITH THE
LINES AND WHAT WE WERE EXCITED
TO SEE IS WHEN WE TAKE THE
ORGANOIDS CREATE FROM THE IPS
CELLS TO DAY 200 IN VITRO, WE'RE
ABLE TO SEE THE BIO CHEMICAL
HALLMARKS OF EARLY FORMULATION
AND WITH THE SAME BACKGROUND IN
2-D MONOLAYER AND CULTURE THEM
UP TO DAY 65 WE DON'T SEE THE
PRESENCE OF THE HALL MARKS AND
THIS MEANT WE COULD SEE A PURE
POPULATION BEFORE THE OBVIOUS
HALLMARKS.
SO FIRST AND FOREMOST WE DID
GENE EXPRESSION AND LOOKED AT
RNA SEQ ACROSS THE TIME COURSE
OF DIFFERENTIATION OF EARLY AND
LATE NPCs AND CORTICONEURONS AND
WITH THE MUTATION WE SAW
WIDESPREAD GENE EXPRESSION
DYSREGULATION WITH A FOUR FOLD
MORE DECREASE IN GENE
EXPRESSION.
THE RESULT IS REPRODUCEABLE
ACROSS MUTATIONS.
THE SAME GENES DOWN AND UP IN
LARGE PART WERE DISRUPTED AND WE
SAW A SIMILAR PATTERNS IN THE
ORGANOID AND CHANGED AT DAY 200.
WE WANTED TO LOOK AT CRHROMATIN
ACCESSIBILITY.
THE GENES THAT SHOWED
DYSREGULATION AND LOOKING AT THE
PROMOTERS, 99% OF THE TIME IT
DID NOT CHANGE AT ALL.
THOUGH THE GENES WERE
DRAMATICALLY CHANGING IN.
LOOKING AT DISTAL NON-CODING IS
WHERE THE ACCESSIBILITY CHANGED
BUT IN WAYS NOT THAT PREDICTABLE
WITH THE GENE EXPRESSION.
AND SO FOR A LAB FOCUSSED
INITIALLY ON THE 3-D GENOME IT
GAVE INSIGHT WE NEEDED THE LOOPS
TO FIGURE OUT THE PROBLEMS.
HERE'S THE POST-MITOTIC NEURONS
IN WILD TYPE AND GENOTYPE AND
JUST TO GIVE AN EXAMPLE TO SEE
THE DOTS AROUND THE GENE AND HOW
THEY'RE DRAMATICALLY DISRUPTED
IN POST-MITOTIC NEURONS.
AND WE CALLED THESE WITH METHODS
DEVELOPED WITHIN OUR LARGER
COMMUNITY OVER THE YEARS WE
FOUND THE RARE MUTATION 30,000
UNCHANGED AND SOME WITH THE APP
MUTATION.
6:WE SAW THEM DUE TO THE RARE
MUTATION AND GRABBED THE LOOPS
GAINED DUE TO THE RARE MUTATION
AND FOUND THE LOOPS UNCHANGING
AND ALSO GROUPED UP GENES NOT
FORMING ANY STRUCTURE AT ALL AND
WANTED TO ASK SO NOW WHAT
HAPPENS IN GENE EXPRESSION IN
THE UNDIVIDING POST-MITOTIC GENE
EXPRESSION.
THIS ONE YOU WILL NOT BE
SURPRISED ABOUT.
WHEN YOU SEE A GENE THAT LOSES
ONE LOOP IN POST-MITOTIC NEURON
THERE'S NO POWER AND YOU CAN
HAVE EQUAL PROBABILITY OF
UNCHANGED, INCREASING OR
DECREASING EXPRESSION
REPRESENTATIVE OF THE
EXPERIMENTAL RESULTS
DEMONSTRATED IN OUR FIELD.
WHEN WE BEGIN TO STRATIFY BY
NUMBER OF ENHANCER LOOPS THE
MORE LOOPS YOU LOSE, THE MORE
IT'S PREDICTIVE OF THE
DIRECTIONALITY AND EFFECT SIZE
OF THE GENE EXPRESSION.
THE MORE LOOPS A GENE GAINS THE
MORE IT'S PREDICTIVE OF AN UP
REGULATED GENE EXPRESSION
PATTERN.
THERE'S NO PREDICTIVE POWER OVER
GENE EXPRESSION AND GENES THAT
DON'T LOOP AT ALL THERE'S ALSO
NO PREDICTIVE POWER.
SO AS WE LIKE TO SAY IN THE LAB,
MORE LOOPS, MORE PROBLEMS.
WHAT'S AT THE BASE OF ALL THE
LOOPS CHANGING?
I'LL SPARE YOU ALL THE DEFEATS
AND SAY AMONG THE ARCHITECTURE
PROTEINS IN THE OBVIOUS
CANDIDATES, WE LOOKED AT THE
BASE OF THE LOOPS AND CLASSIFIED
THEM IN ALL SORTS OF WAYS AND
THE BOTTOM LINE IS THAT ALL
THESE LOOPS THAT ARE LOST AND
GAINED IN POST-MITOTIC IS
UNCHANGING.
THERE'S SOMETHING ELSE LEADING
TO THESE LOOP DYSREGULATIONS.
AND THE ONE MAIN SIGNAL WE'VE
BEEN ABLE TO FIND OVER THE YEARS
IS THAT AT THE BASE OF THE LOOPS
LOST AND GAINED IN THE
ALZHEIMER'S MUTATION GENETIC
BACKGROUND THEY HAVE STRONG
PRESENCE OF RNA POLL II WE CAN
LOOK UNDER THE BAY AND LOOK AT
THE MOTIFS AND LOOKED AT THE
LOST AND GAINED LOOPS IN THE
NEURONS.
USUALLY WITH MOTIF ENRICHMENT
FLALS YOU CAN FIND ANY NUMBER OF
HITS WITH THIS TYPE OF WORK BUT
IN THIS CASE WE KEPT GETTING ONE
SINGULAR HIT.
RAY IF THE LOOPS ARE LOST UNDER
THOSE DYNAMIC SITES IT'S
PRESENTS IN AN ARRAY IF THEY'RE
GAINED IT'S PRESENT TYPICALLY
ONLY OWN ONE KWOPY.
-- COPY.
THE RESULT YOU GET IS FAIRLY
DISTURBING.
THESE ARE TRANSCRIPTION FACTORS
THAT IS HIGHLY REDUNDANT AND
WOULD BE DIFFICULT TO DO A
SINGULAR LOSS OF GENE EXPERIMENT
BUT SOME STOOD OUT AND WE
SEARCHED OUR GENE EXPRESSION
RESULTS.
AND HERE'S WHERE I THINK WE GOT
LUCKY.
WE SAW THAT FOR EXAMPLE THE
DIFFERENTIATION TIME POINTS THE
mRNA PLUMMET AND TURNED OFF BUT
HIGH IN WILD TYPE.
SIMILARLY BUT OPT SIT, DLX5 AND
6 ARE NOT PRESENT IN WILD TYPE
BUT EXPRESS UP REGULATED DUE TO
THE MUTATION AND SO FOR THAT
REASON IT GIVES CANDIDATES WE
CAN BEGIN TO PURSUE WITH THE
GAIN OF FUNCTION STUDIES.
SO WHAT WE'RE DOING RIGHT NOW IS
OVER EXPRESSING DLX1 WITH AN HH
TAG TO CHIP IT AND LOOKING AT
THE TIME COURSE RAND GENE
EXPRESSION AND THE NEURONS.
I ALSO WANTED TO SHARE WITH
GREAT EXCITEMENT WE SAW RECENTLY
A NEW PRE PRINT DROPPED IN
COLLABORATION AND IN A MODEL
SYSTEM OF THE SONIC HEDGEHOG
LOCUS IN VIVO AND WHAT MIGHT
ENCODE OR ASSIST IN THE
INTERACTIONS.
AND SIMILAR TO OUR WORK BUT
WHERE THEY CAN DO THESE IN VIVO
THEY FOUND THE SAME MOTIF
PARTICIPATING IN THE FORMATION
OR DISRUPTION OF LOOPING
INTERACTION AND OVEREXPRESSING
IN AN IN VITRO MODEL AND DOING
CRISPR MUTATIONS IN AN IN VIVO
MODEL AND WE'RE TRYING TO ATTACK
THIS QUESTION FROM TWO ANGLES IN
COLLABORATION.
OKAY, SO I WANT TO END HERE BY
BRINGING THIS FORWARD AS
SOMETHING THAT I CAN SAY JOHN
SADERLY AND OTHER PROGRAM
OFFICERS HAVE CHALLENGED US IN
THE COMMUNITY WE SPEND TIME IN
EFFORT AND BLOOD, SWEAT AND
TEARS BUILDING THE COMPUTATIONAL
TECHNOLOGIES TO MAP AND
UNDERSTAND HOW THE GENOME FOLDS
AND EVEN IF WE THINK IT MATTERS
FOR GENE EXPRESSION, WHAT IS THE
PHYSIOLOGIC SIGNIFICANCE?
WE WONDERED IF THERE WOULD BE
SOMETHING WITHIN OUR SYSTEM TO
HELP US UNDERSTAND.
WE LOOKED FIRST AT OUR NEURONS
WE KNOW HAVE MASSIVELY DISRUPTED
LOOPS AS WELL GENE EXPRESSION
AND THOSE WHO ARE MICROSCOPISTS
WE SAW NO DIFFERENCE.
WHERE THE INSIGHT CAME WAS WHERE
WE BEGAN TO DO MORE ANALYSIS AT
THE SINGLE CELL LEVEL.
WE DID SINGLE NUCLEUS RNA SEQ
FOR THE NEURONS AND FOUND IN
LARGE PART THE WILD TYPE NEURONS
WERE EXCITATORY IN CHARACTER BUT
IN THE MUTATION THOUGH NOT
OBVIOUS BY MORPHOLOGY THE
NEURONS SHIFTED THEIR
COMPOSITION AND SUBSTATE TO NOW
BE ALMOST ENTIRELY INHIBITORY IN
CHARACTER BUT WOULDN'T HAVE
DETECTED IT.
SO THIS IS VERY SURPRISING TO US
AND WE'RE DOING SUBSEQUENT
FOLLOW-UP STUDIES NOW BUT THE
EMERGING MODEL FROM OUR WORK IS
AS FOLLOWS.
IN WILD TYPE HUMAN NEURONS
DERIVED FROM IPS CELLS WE USED
CRISPR TO ADD HOMOZYGOUS RARE
AUTO SOMAL CASES LINKED TO EARLY
ONSET ALZHEIMER'S AND WE SEE
THERE'S MANY LONG RANGE LOOPING
INTERACTIONS AND CAN BE
DISRUPTED DUE TO THE MUTATION.
AT THE BASE OF THE DISRUPTED
LOOPS ARE THE MOTIF AT THE
ENHANCERS TTA AND WE ALSO SEE
GAINED LOOPS IN THE SAME MODEL
SYSTEM.
THEY ALSO HAVE TAA, TTA AND IF
YOU LOOK RAT THE -- AT THE BASE
OF THE LOOPS IT'S OFTEN
INHIBITORY, EXCITATORY GENES
DISRUPTED AND SHIFTS WITH THE
GENE EXPRESSION TO BE MORE
INHIBITORY IN CHARACTER AND THIS
PLAY BE ONE IMPORTANT PLACE
WHERE THEY MATTER FOR THE
CELLULAR STATE OF NEURONS IN
HUMAN DISEASE AND HAVE A
RELATIONSHIP BETWEEN
DYSREGULATED AND EXCITATORY AND
INHIBITORY GENE EXPRESSION AND
IT APPEARS IN OUR MODEL TO OCCUR
PRIOR TO THE DEPOSITION OF BIO
CHEMI
CHEMICAL HALLMARKS OF AMYLOID
PLAQUE AND TAU.
THIS IS A BEAUTIFUL EDITORIAL
THAT SUMMARIZED THE WORK OF SOME
OF MY COLLEAGUES WHERE THE USED
SIMULATION TO CAPTURE THE
IMAGINATION OF HOW DIFFICULT THE
3-D GENOME FOLDING HAS BEEN OVER
THE LAST 10 YEARS.
I'M SHOWING SOME THINGS THAT IN
THE EARLY DAYS WE DID NOT KNOW
IF THIS WOULD BE A TRACKABLE
PROBLEM AND IT WAS HARD BUT FUN
AND NOW I WANT TO DO IT AGAIN.
AND THE IDEA THAT HAS CALLED OUT
SO US I THINK IS ANOTHER
CONNECTIVITY PROBLEM OF THE
NEURAL CONNECTOME AND WE STARTED
THE PROCESS OF THINKING HOW TO
YOU THE PRINCIPLES COMPUTATIONAL
AND EXPERIMENTAL AND BEGIN TO
THINK OF WAYS TO DO MOLECULAR
MAPS OF THE SYNAPTIC CONNECTOME
AND THAT'S WHAT WE'RE WORKING ON
NOW THINKING OF CREATING SYNAPSE
RESOLUTION ACTIVITY.
IT WILL BE HARD BUT SKIESEXCITE
WORK WITH NEW YOUNG PEOPLE TO
TACKLE THE PROBLEM AND WHEN
THERE'S PERSISTENT CRHROMATIN HS
TO GET BACK AND HOW THEY ARE
CHANGE IN A DYNAMIC MANNER AND
PROVING THE RNAs MATTER IN SOME
WAYS FOR BEHAVIOR AND THOUGH
I'VE SHOWN AN EXAMPLE OF
ALZHEIMER'S WHERE I FOCUS ALMOST
ENTIRELY ON CHROMATIN WE HOPE TO
UNDERSTAND HOW THE LOOPS MIGHT
METER FOR THE THE PLASTICITY MAY
BE DYSREGULATED.
OUR INTEREST IS IN FOCUSSING ON
A FEW MODEL SYSTEMS.
AND THINKING FROM THE NUCLEUS
OUT TO THE SYNAPSE AND OUR HOPE
IS OVER TIME WE CAN PERHAPS IN
THE NEXT 10, 20 YEARS FIND
INSIGHTS TO GIVE GENUINE HELP
FOR THOSE WHO CONTINUE TO SUFFER
AND THE MYRIAD OF PROBLEMS THIS
WORLD FACES IN THE POST-PANDEMIC
WORLD.
I'D LIKE TO CLOSE AND HIGHLIGHT
THE WORK OF MY GROUP PAST AND
PRESENT AND IT'S BEEN A
PRIVILEGE TO WORK WITH THEM AND
KEEP ME YOUNG AND THANK YOU TO
ALL WHO LISTENED AND INVESTED IN
OUR LAB AND WORKING WITH ME AND
ALSO VERY IMPORTANT JOHN ALLUDED
TO JUST THANK YOU VERY MUCH TO
THE NIH AND COLLEAGUES AT TO THE
NIH AND I THINK THE SMALL
CONSISTENT SUPPORT MAYBE IN ONE
SINGLE INSTANCE PLAY NOT AFFECT
A PERSON THAT MUCH BUT SUSTAINS
CAN SUPPORT PEOPLE AND HOW IT'S
IMPACTED ALL IN MY GROUP.
THANK YOU AGAIN FOR HAVING ME
AND I'LL BE HAPPY TO ANSWER
QUESTIONS.
>> THANK YOU.
OF COURSE ANYONE HERE SHOULD BE
USING THE MIKES.
WE HAVE A ROBUSTION -- ROBUST
ONLINE QUESTION.
CAN ANY SAMPLES BE DONE WITH THE
HUMAN ABLATED SAMPLES?
>> WE THINK THINK THEY COULD BE
DONE WITH SURGICALLY ABLATED
SAMPLES.
PERHAPS NOT SOME OF THE GENOME
ENGINEERING BUT MANY OF THE
SINGLE CELL TECHNOLOGY IT'S
POSSIBLE AND TRACING OF THE
CIRCUITS AND GETTING OUT NOT
ONLY THE TRAPPED NEURONS ACTIVE
DURING BEHAVIOR BUT IT'S GETTING
TO THE PHYSIOLOGICALLY RELEVANT
NEURONS THAT FOR US HAS BEEN THE
MOST DIFFICULT PIECE AND WILL BE
CRITICAL AND CHANGE WITH EVERY
BEHAVIOR AND EXPERIMENT AND
EXPERIENCE.
YOU HAVE TO GET IT JUST RIGHT.
LUCKILY THERE'S BEAUTIFUL WORK
TO TRACE AND GET.
6:
>> I'M VERY INTRIGUED BY THE AKC
DERIVED NEURONS FROM SINGLE
MUTATIONS.
AND WONDERING WHETHER YOU HAVE
DONE HETER ZYGOUS CELL LINE WITH
A MORE INHIBITORY NEURON, FIRST
QUESTION.
SECOND QUESTION I THINK THERE'S
MICE FOR THE MUTATION BUT DON'T
REMEMBER DRAMATIC CHANGE OF THE
NEURON PRESENTATION THE
INHIBITORY IS HUGE SO HOW DO YOU
VERIFY THE FINDING OF THE IN
VIVO MODEL.
>> THANK YOU FOR YOUR QUESTION
AROUND TWO PARTS SUMMARIZED FOR
THE LIVE ONLINE AUDIENCE.
WE USE HOMOZYGOUS MUTATION OF A
RARE AUTOSOMAL DOMINANT MUTATION
ON PURPOSE.
WE WANTED TO USE FIRST PASS
BECAUSE WE WANTED TO GET THE
HIGH
HIGHEST SIGNAL TO NOISE AND THEN
MOVE TO RAY DIFFERENT STUDY.
WE SAW SEVERE CHANGES.
THE EXCITATORY TO INHIBITORY
SHIFT WAS DRAMATIC AND SEVERE.
WHAT THE QUESTION THOUGH THEY
DIDN'T RAISE IT WAS PROBABLY
ALLUDED TO IS EARLY ONSET
ALZHEIMER'S YOU DON'T SEE THIS
DRAMATIC SHIFT TO INHIBITORY
NEURONS AND WE PUT IN TWO COPIES
OF THIS RARE AUTOSOMAL DOMINANT
MUTATION AND WE ARE WORKING WITH
DOMINICK TO VALIDATE THE LINES
AND ENSURE THEIR EUKARYOT TYPE
MUTATION AND WE HAVE TO USE
SINGLE CELL TECHNOLOGIES TO
LOOK.
THE SECOND PRART OF THE QUESTION
WAS A DIFFERENT ASPECT WHICH IS
THINK BEING ABOUT MOUSE STUDIES
THAT HAVE -- IF I UNDERSTOOD
CORRECTLY WAS A HUMANIZED MOUSE
THAT KNOCKED IN PERHAPS A
PARTICULAR GENE WITH A MUTATION
AND SAYING WE LOOK BACK AT THAT
LITERATURE AND LOOK AT THE
EXCITATORY TO INHIBITORY
POPULATION AND EXCELLENT
SUGGESTION.
>> I HAVE A QUESTION ON YOUR
LOOPING WORK.
YOU SHOWED THE MORE LOOPS YOU
LOSE YOU SEE A DECREASE IN GENE
EXPRESSION.
ARE MOST GENES REGULATED BY ONE
OR TWO LOOPS?
IS THAT HOW THAT WORKS OR AM I
MISUNDERSTANDING HOW TO REC
RECONCILE THESE TWO?
>> I THINK YOU'RE REFERRING TO
THE WORK IN COLLABORATION WITH
WE KNOCKED OUT COHESSIN AND
LOOKED AT THE DISRUPTION OF
LOOPS.
>> THE WORK IN VIVO VERSUS THE
MULTI-LOOP MODEL WITH
ALZHEIMER'S, IN VIVO WORK EN
ELLY 51, WHEN YOU KNOCK OUT
COHESSIN THERE'S SEVERE GENE
EXPRESSION DYSREGULATION DURING
MATURATION IN VIVO.
WE TOOK THE NEURONS OUT AND IN
VITRO USED PHARMACOLOGIC AGENTS
AND TO FIRE AND THEY ARE DIDN'T
MATTER IN LATER THAT'S WHEN
LOOPS MATTERED QUITE A BIT.
IN THAT CASE WE DIDN'T LOOK AT
THE MULTI-LOOPING IDEA.
IT WAS PUBLISHED BEFORE WE FOUND
IT IN THE LATER STUDY SO CANNOT
COMMENT IF IT'S A PHENOMENON FOR
ACTIVITY EXPRESSION?
>> DO YOU SEE PLAQUE BUILD UP IN
THE 2-D NEURON CULTURES?
>> THE NEXT QUESTION IS ABOUT
THE FAD AND IF HE QUESTION IS IF
WE WAIT LONG ENOUGH IN 2-D DO WE
EVENTUALLY SEE PLAQUE OR
PHOSPHORYLATED TAU.
I NEVER EVEN TRIED TO CONVINCE
THE STUDENTS TO KEEP THE NEURONS
GOING FOR ANOTHER 100 DAYS.
THERE'S BEAUTIFUL WORK ALREADY
OUT DIFFICULT WITH IPS MODELS OF
ALZHEIMER'S IN 2-D FLAT CULTURE
TO HAVE AN AGGREGATION.
WE THOUGHT THAT WOULD BE
INTRACTABLE AND FORTUNATE IN
TRYING THE ORGANOIDS AND WHEN WE
HARVESTED THERE WAS NOTHING AND
WE WERE GETTING NERVOUS AT THAT
POINT.
>> THANK YOU.
>> VERY INTERESTING TALK.
I WAS TRYING TO CONNECT THE DOTS
BETWEEN THE DIFFERENT PARTS OF
YOUR TALK AND THERE WAS ONE PART
OF THE TALK WHERE YOU WERE
EMPHASIZING THE IMPORTANCE OF
THE FORMING OF THESE DIFFERENT
LOOPS AND CONNECTIONS BETWEEN
DIFFERENT PARTS AND WASN'T SURE
WHETHER THAT WAS IN CONNECTION
WITH RANDOM ACTIVITY AND
LEARNING BUT WAS IT LEARNING OR
RANDOM ACTIVITY?
>> IN THE CASE OF THE ACTIVITY
DEPENDENT LOOPING IS IT RELATED
IF IN VITRO SPONTANEOUS FIRING
OR INVOKED POTENTIAL AND WITH
THE IN VIVO DATA FROM BEHAVIORAL
OR EXPERIENCE-INDUCED OR
OPTOGENETIC STIMULATION.
FOR THE IN VITRO DATA THAT'S
PHARMACOLOGICALLY INDUCED ACTION
POTENTIAL FIRING NOT SPON
TRAINIOUS AND OUR CONTROL IS TT
X.
WE ENSURE USING THAT
PHARMACOLOGIC AGENT FOR FIRING
AS MUCH AS POSSIBLE.
FOR THE IN VIVO SITUATION WE
USED CONTEXTUAL FEAR
CONDITIONING AND USED THAT
BEHAVIOR FOR INJECTION IN THE
TRAP 2 ANIMAL TO FIRE ACTION
POTENTIAL DURING THE WINDOW OF
BEHAVIOR.
>> SO THE QUESTION FOR ME WAS SO
THAT WAS COOL AND EXCITING AND
TALKED ABOUT ALZHEIMER'S AND
MAKING ORGANOIDS AND THE GENES I
SAW AS DEVELOPMENTAL GENES
THOUGH THEY PLAY HAVE OTHER
ROLES.
SO I WAS TRYING TO FIGURE OUT
THE CONNECTION BETWEEN LOOPS
THAT YOU HAD ESTABLISHED IN THE
CONTEXT OF LEARNING OR TASK AND
WHAT'S HAPPENING IN AN ORGANOID.
>> FOR THOSE ONLINE THE QUESTION
IS RELATED TO HOW DO WE CONNECT
THE WORK HAPPENING IN CIRCUITS
WITH THE IN VITRO HUMAN MODEL OF
ALZHEIMER'S DISEASE WHERE WE'RE
CLEARLY AND SURPRISINGLY SEEING
CHANGES IN DEVELOPMENT GENES AND
LOOPING INTERACTIONS.
WHAT I HAVE TO SAY IS OUR PLAN
IN THE FUTURE IS TO NOW IN THE
ALZHEIMER'S MODEL BEGIN TO LOOK
AT ACTION POTENTIAL FIRING AND
THAT'S THE DIRECTION WE'RE GOING
AND YOU COULD PROBABLY GUESS
WHERE WE'RE PUSHING TOWARDS O
ORGANOIDS AND LOOK TO DO
SOMETHING IN VIVO IN BREEDING
WITH THE TAP 2 ANIMAL AND WHAT
I'LL SAY ON THE RECORD IS NO ONE
MORE THAN ME WAS MORE SHOCKED
THAN THE WIDESPREAD NATURE OF
THE LOOP DYSREGULATION AND THE
FACT IT WAS A LOT OF DEVELOPMENT
AND EARLY SYNAPTIC PLASTICITY
GENES AND IT'S LIKELY PARTIALLY
PART OF THE MODEL SYSTEM BECAUSE
WE HAVE A HOMOZYGOUS MUTATION
BUT THERE'S REAL INSIGHT THERE
BECAUSE YOU'RE STARTING TO SEE
IN CONFERENCES ACROSS DISORDERS
THEY'RE STARTING TO BE MISS
MOVEMENT OF HOW SOON DO WE HAVE
TO LOOK IN DEVELOPMENT IN EARLY
ADULTHOOD TO SEE THE INITIAL
UNRAVELLING AND ONSET OF
DISEASE.
IT'S POSSIBLE OVER THE YEARS
WE'LL START TO SEE THE DEFECTS
AT THE MOLECULAR LEVEL HAPPEN
SOONER THAN SOME OF THE SEVERE
CLINICAL PRESENTATIONS.
SO WE'LL KEEP PUSHING IT AND AS
SOMEBODY ELSE MENTIONED MAYBE
BRINGING IN THE HET WILL
UNDERSTAND THE EFFECT SIZE.
>> IT PREDICTS THOSE ASSOCIATED
WITH ALZHEIMER'S MAY
CONNECTED --
>> YES.
THERE'S AN IMPORTANT IDEA RAISED
BY THE QUESTIONER IF INDIVIDUAL
WITH EARLY ONSET ALZHEIMER'S
SOME WHO MAY HAVE THE MOLECULAR
DEFECT MAY MANIFEST IN THE
CLINIC AS HAVING PRESENTATIONS
MORE TYPICALLY EQUATED WITH
NEURODEVELOPMENTAL DISORDERS.
I'M NOT A CLINICIAN SO I DON'T
KNOW IF AS THE FEATURES
MOLECULAR AS WELL AS CLINICAL
PRESENTATIONS AND I THINK IT
WOULD BE VERY EXCITING IF THAT
ENDED UP BEING THE CASE.
>> ONE LAST QUESTION AND WE ARE
PAST THE HOUR.
>> MY NAME IS MIKE.
I'M 90 YEARS OLD AND FOR MANY
YEARS MY LAB CHIEF WAS MAXINE
SINGER WHO UNFORTUNATELY DIED
THIS SUMMER AND I WOULD LIKE TO
CHALLENGE YOUR TIME FRAME WHICH
YOU MENTIONED OF A DECADE WHEN I
LOOK ON TV AT NEWS AND LOOK AT
THE PHARMA COMPANIES WHICH ARE
USING MONOCLONAL ANTIBODIES TO
IDENTIFY SEQUENCES WHICH ARE
ROLE VANT TO ALZHEIMER'S AND
WHICH ARE USING ARTIFICIAL
INTELLIGENCE ENHANCED SEQUENCING
TO FIND LOW MOLECULAR DRUGS
WHICH INFLUENCE ALZHEIMER'S AND
THAT I WOULD SUGGEST IF YOU LOOK
AT THOSE CAREFULLY AND SEE WHAT
THEY'RE DOING YOU'LL SEE YOUR
TIME FRAME SHOULD REALLY BE ON
THE ORDER OF MONTHS RATHER THAN
ON DECADES.
>> THANK YOU.
>> THANK YOU SO MUCH.
THANK YOU, JEN.
I'M HERE TO TELL YOU ALL THAT
I'M TOLD THERE'S A RECEPTION
AFTER THIS AND THERE WILL BE
COOKIES AND POSSIBLY JEN'S
FAVORITE CHEMICAL, CAFFEINE,
POSSIBLY.
WE'D LIKE YOU TO JOIN US TO TALK
TO JEN DURING THE RECEPTION AND
JOIN ME IN A ROUND OF APPLAUSE
FOR AN AWESOME TALK.