>>WELCOME TO
THE WEDNESDAY AFTERNOON LECTURE
SERIES. MY NAME IS MARY DAWSON
SENIOR INVESTIGATOR THE
NATIONAL INSTITUTE OF HUMAN
HEALTH DEVELOPMENT AND I'M
EXCITED TO INTRODUCE TODAY'S
SPEAKER DR. YAMASHITA, A
PIONEER IN THE STUDY OF GENOME
ORGANIZATION PARTICULARLY IN
THE MAIL GERMLINE AND SHE WOULD
PRESENTING THE 2023 PITTMAN
LECTURE AS PART OF THE
WEDNESDAY AFTERNOON SERIES. THE
PITTMAN LECTURE IS GIVEN BY A
LECTURE DEDICATED TO ADVANCING
THE IMPROVEMENT OF CAREERS OF
WOMEN IN SCIENCE. SINCE 1994
WHEN THIS LECTURE SERIES BEGAN,
EVERY SPEAKER HAS EXEMPLIFIED
THE INTELLIGENCE, SCIENTIFIC
EXCELLENCE, AND DRIVE THAT MADE
MARGARET PITTMAN A LEADER AT
THE FIRST FEMALE LABORATORY LAB
CHIEF AT NIH.
I WILL POINT OUT THAT DR.
YAMASHITA WAS NOMINATED FOR
THIS HONOR BY THE NIA WOMEN IN
SCIENCE ADVISORS.
IS A GREAT HONOR
TO HAVE YUKIKO YAMASHITA WITH
US TODAY AND SHE IS CURRENTLY
AN HHMI INVESTIGATOR PROFESSOR
OF BIOLOGY AT MIT AND IS A
MEMBER OF THE WHITEHEAD
INSTITUTE WHERE SHE HELD THE
SUSAN LINDQUIST CHAIR FOR WOMEN
IN SCIENCE AND SHE BEGAN HER
SCIENTIFIC CAREER AT KYOTO
AND STANFORD
RESPECTIVELY BEFORE
HER OWN LAB AT THE
UNIVERSITY OF MICHIGAN AT ANN
ARBOR AND BECAME A FULL
PROFESSOR IN THE DEPARTMENT OF
CELL AND DEVELOPMENT OF BIOLOGY
MICHIGAN WHERE SHE ULTIMATELY
HELD A PROFESSORSHIP IN THE
LIFE SCIENCES INSTITUTE AND WAS
A DIRECTOR OF THE MICHIGAN LAB
SCIENCE FELLOWS PROGRAMS. SHE
MOVED TO MIT IN 2020 WHERE IN
ADDITION TO THE TITLES THAT I
MENTIONED EARLIER SHE SERVES AS
THE COCHAIR OF THE BIOLOGY
DIVERSITY EQUITY AND INCLUSION
COMMITTEE.
DR. YAMASHITA'S WORKERS
BROADEN OUR UNDERSTANDING
REGARDING FUNDAMENTAL ASPECTS
OF MULTICELLULAR ORGANISMS
INCLUDING HOW CELLS ARE
DEMYSTIFIED BY ASYMMETRIC CELL
DIVISION AND GENETIC
INFORMATION IS TRANSMITTED
THROUGH THE GERMLINE.
HER WORK HAS BEEN PARTICULARLY
IMPORTANT FOR THE ELUCIDATION
OF STRUCTURAL GENOMES AND
EXPLORING SUCH IMPORTANT
QUESTIONS AS A MEASURE OF
SATELLITE DNA PREVIOUSLY
CONSIDERED TO BE CONSIDER
GENOMICS JUNK AND HOW IT MIGHT
CONTRIBUTE TOWARDS SPECIATION.
HER WORK IS ALSO BROUGHT UP
LIKABILITY TO UNDERSTANDING
ASYMMETRICAL TELEVISION
QUESTIONS REGARDING THE
IMMORTALITY OF CELLS WITHIN THE
GERMLINE AND I BELIEVE SHE IS
GOING TO ADDRESS SOME OF THESE
QUESTIONS TODAY AND AS YOU
WOULD EXPECT FROM SUCH AN
UPSETTING SCIENTIST TO MENTOR.
IT INCLUDES THE AWARD FROM
MACARTHUR FELLOWSHIP IN
THOUSAND 11 AND THE -- AND
AWARD FROM NAGOYA UNIVERSITY
2016 AND BEING NAMED AS A
FELLOW OF THE AMERICAN SOCIETY
OF CELL BIOLOGY IN 2021 AND SHE
IS CURRENTLY SERVING AS THE
BOARD OF SCIENTIFIC COUNSELORS,
MEMBER FOR THE NICHD INTRAMURAL
PROGRAM. WITHOUT FURTHER ADO
THE TITLE OF YUKIKO'S TALK IS
GERMLINE IMMORTALITY AND
ASYMMETRIC STEM CELL DIVISION.
ONCE AGAIN BECAUSE THEY WANTED
TO EMPHASIZE IT, THE CME CODE
IS 44455.
LOOKING VERY MUCH AND THANKS
SO MUCH FOR BEING HERE.
>> [APPLAUSE]
>>I'M REALLY EXCITED TO BE BACK
THANK YOU SO MUCH FOR THE
INVITATION.
TODAY I'M GOING TO TELL YOU
SOMETHING. IT TOOK 10 YEARS FOR
US TO GET SOME SORT OF THE
FRAME LOG, OR THE WHAT ARE WE
TALKING ABOUT. THAT IS QUITE A
JOURNEY. I AM VERY EXCITED TO
SHARE.
ALMOST AS AN EXCUSE I PUT THIS
QUOTE FROM -- "I DOUBT IT COULD
HAVE BEEN ANTICIPATED..." ABOUT
HOW IT DISCOVERED THE
TRANSPOSALS, AND MANY THINGS
UNEXPECTED IN SCIENCE.
TODAY I AM GOING TO TELL YOU
ABOUT THE GERMLINE IMMORTALITY.
IF YOU REALLY THINK ABOUT THIS,
IT IS QUITE REMARKABLE.
IF YOU ARE A TALKING ABOUT
SOMATIC CELLS, ANY CELLS IN
YOUR BODY-- AND EVEN ACTUALLY
-- TEND TO DIVIDE. IN THE FACE
OF THIS LIMITED LIFESPAN FOR
ANY KIND OF A CELL, IS QUITE
REMARKABLE TO TO THINK ABOUT
THIS LINEAGE. EACH ONE OF YOU
USED TO BE A GERM CELL FROM OUR
PARENTS, SEVERAL DECADES AGO.
AND YOUR PARENTS WERE ALSO
AGAIN, THE TWO GERM CELLS,
SEVERAL DECADES AGO, IT KEEPS
GOING BACK AND DURING THE
JOURNEY OF THE GERM CELLS, THE
GERM CELLS NEVER STOPPED
DECEASING. THAT IS WHAT YOU AND
I ARE HERE. GERM CELLS LASTED
1.5 BILLION YEARS.
IT IS A FASCINATING QUESTION.
HOW DOES GERMLINE ACHIEVE THIS
IMMORTALITY?I DO NOT HAVE A
FULL ANSWER. BUT THERE IS
RESEARCH THAT LEADS TO PART OF
THE QUESTION AND THIS RESEARCH
STARTED WHEN I WAS AN ASSISTANT
PROFESSOR AT THE UNIVERSITY OF
MICHIGAN. THIS IDEA OF THE
NONRANDOM SISTER CHROMATID
SEGREGATION HAS BEEN OUT THERE
QUITE A BIT OF TIME. LET ME
EXPLAIN WHAT IT MEANS BY
CHROMATID SEGREGATION. HERE IS
THE DNA STRAND. DURING DNA
REPLICATION THESE TWO STRANDS
ARE GOING TO SEPARATE FROM EACH
OTHER. SO THE PLUS STRAND WILL
TEMPLATE THE MINUS STRAND, THE
MINUS STRAND TEMPLATES THE
POSITIVE STRAND. IT IS CALLED
SISTER CHROMATIDS.
IN DNA DUPLICATION YOU ARE
COPYING FROM THE TEMPLATE EVERY
TIME YOU DIVIDE. YOU ARE GOING
TO BE COMPLETELY DEAD OR BE
SOMETHING TO BELIEVE DIFFERENT
IF IT DID NOT DID THAT. DESPITE
THIS IDEA OF THIS REPLICATION
IS ASSOCIATED WITH SOMETHING
YOU NEED, SUCH AS EPIGENETIC
MARKS OR INDUCED MUTATIONS OR
WHATEVER. THERE IS SOMETHING
ELSE.
IF THAT KIND OF SPECIAL
INFORMATION IS ASSOCIATED WITH
ONLY ONE SISTER CHROMATID, AND
IT ONLY GOES TO ONE CELL
DIVISION THAT MIGHT LEAD TO A
MUTATION AND THERE ARE MANY
PAPERS THAT SUPPORT THIS IDEA.
MY GRADUATE STUDENT WANTED TO
TEST THIS IDEA. THEY TESTED
THE POSSIBILITY AS PART OF THE
DNA THAT ALL THE STRAND OF US
HAS THE NEW STRAND OF THE
ENTIRE GENOME. SHE SAW NO
EVIDENCE OF THIS.
BUT THE SAME STUDENT,
CURRENTLY AN ASSISTANT
PROFESSOR AT THE UNIVERSITY OF
MICHIGAN, SWATHI YADLAPALLI,
SHE WANTED TO GET TO THE BOTTOM
AND ADDRESSES POSSIBILITY. SHE
WONDERED IF SHE COULD ADDRESS
THIS AT THE HIGHEST RESOLUTION
POSSIBLE.
IN THIS METHOD, WHEN THE DNA
REPLICATION IS HAPPENING, YOU
ARE GOING TO PUT THE BRDU AND
THE ANALOG DURING THE DNA
REPLICATION AND WHEN THAT
HAPPENS ONLY ONE SIDE OF THE
STRAND IS GOING TO BE BRDU
POSITIVE. UNLESS THE DNA
CONTAINS BRDU, AFTER YOU
DISSECT THE TISSUE IN THIS
EXAMPLE YOU CAN IRRADIATE THE
AFFECTED TISSUE. IF YOU MAKE
THE NEW STRAND YOU CAN DIGEST
THEM AWAY.
THAT LEADS ONLY OLDER
TEMPLATES AS A SINGLE-STRAND
DNA. AND HOW YOU CAN
DIFFERENTIATE THE TWO STRANDS
IN A DIFFERENT-- SO YOU CAN
TELL WHICH CELLS INHERITED THE
CY3 STROBE ANDYOU SEE THE CY5
IN THE BLUE STRAND. THIS IS
CALLED CHROMOSOMAL ORIENTATION
FISH, DEVELOPED BEFORE SWATHI
DECIDED TO DO THIS BUT SHE
DECIDED TO DESIGN AND PRODUCE
CHROMOSOME SPECIFIC.
SHE WAS ABLE TO DESIGN THE X
CHROMOSOME SPECIFIC IN THE RED
AND BLUE AND THE -- --.
AND THEN WE HAVE THE SYSTEM
CALLED THE -- STEM CELL WHERE
WE CAN IDENTIFY THE STEM CELL
DIFFERENTIATED FROM THE DOTA,
SO WE KNOW-- WE HAVE SEEN THE
TISSUE WHICH IS THE STEM CELL,
THE MEDIATED DORA. IN THIS
CONTEXT YOU CAN APPLY THE FISH,
YOU CAN SEE WHICH STRAND, RED
STRAND OR BLUE STRAND WENT TO
THE SIDE OF THE STEM CELL
DIVISION.
THIS IS WHAT WE GET WHEN WE DO
THIS. THIS IS AN EXAMPLE. AS I
SAID YOU CAN IDENTIFY STEM CELL
DIFFERENTIATING CELL IN A
SINGLE RESOLUTION AND IF IT
GETS RED AND THE -- GETS BLUE
YOU CAN HAVE THE
(INCOMPREHENSIBLE) PART OVER
HERE AS WELL.
WHEN SWHATHI DID FISH, THIS IS
WHAT YOU GOT.
FIRST IN THEORY IS THE DNA
STRAND WERE INDEED A COPY OF
EACH STRAND, IT DOES NOT MATTER
IF IT GETS A RED STRAND OR A
BLUE STRAND. IT DOES NOT MATTER.
AND THESE PATTERNS SHOULD BE
OBSERVED WITH EQUAL FREQUENCY
LIKE 50% AND 50%. THAT IS WHAT
YOU EXPECT.
HOWEVER IN THE REAL
EXPERIMENT, WHERE SHE TOOK A
LOOK AT THE X CHROMOSOME THE
SISTER CHROMATID SEGREGATION
PATTERN, ETC., THEN SHE SAW
STRIKING BIAS IN THE
SEGREGATION PATTERN FOR JUST
THE X AND Y CHROMOSOMES.
OF COURSE AT THE BEGINNING WE
WORRIED THAT THIS HAD TO BE
SOME SORT OF ARTIFACT. THIS
SHOULD NOT BE HAPPENING.
HOWEVER, WHEN WE DID EXACT
SAME EXPERIMENT FOR THE
CHROMOSOMES THERE IS NO BIAS.
AT THAT POINT 10 YEARS AGO WE
HAD NO IDEA WHY OR HOW THIS WAS
HAPPENING. THE MAIN PART OF
THE TALK TODAY IS A JOURNEY
OVER THE LAST 10 YEARS.
THEN BACK THEN, GEORGE WATASE
WE JUST LEFT THIS WEEK, THIS
WEEK WAS AS LAST WEEK AND NOW
HE'S HEADING TO HIS NEXT
SEMI-INDEPENDENT POSITION WHERE
HE CAN DO IS INDEPENDENT WORK
CONTINUING THIS WORK.
WHEN GEORGE STARTED, THE FIRST
THING HE DECIDED WAS, WE SHOULD
REALLY MAP SOME SORT OF
CHROMOSOME SPECIFIC PROCESS;
WHEN WE STARTED THIS PROCESS IS
THE ONLY ASSUMPTION WE MADE IS
THAT THERE HAD TO BE A PROBLEM
SPECIFIC (INCOMPREHENSIBLE),
BECAUSE THERE WAS A CHROMOSOME
SPECIFIC PHENOMENA.
WHEN GEORGE DELETED THIS BIG
CHUNK CALL THE BB CHROMOSOME,
THE X CHROMOSOME WAS SEGREGATED
COMPLETELY.
SO WE THOUGHT THERE IS
SOMETHING IN THERE. WHAT IS IN
THEIR?
THE STUDY OF THE SISTER
CHROMATID, YOU GIVE US AN
ANSWER AND WHEN IS THE
(INCOMPREHENSIBLE) -- IF
ANYBODY ASKED ME TO BET MONEY
ON THE -- AND I WOULD'VE SET
MONEY ON THE DNA. BUT FOR THIS
PARTICULAR CASE, I WAS WRONG!
AND IT TURNED OUT TO BE NOT
RIBOSOME DNA. THAT ANSWER CAME
FROM LOOKING AT THE Y
CHROMOSOME WHICH ALSO HAS YBB
DNA. SUCH Y CHROMOSOME ALSO
SEGREGATED.
WE REALLY DID NOT KNOW WHAT TO
MAKE OF THIS. BECAUSE RIBOSOME
DNA IS NOT SPECIFIC TO ANYTHING
THAT HAS A SEX CHROMOSOME. WE
WERE VERY, VERY PUZZLED. BUT
WE HAD TO SAY THIS KIND OF
MAKES SENSE, LOGICALLY
SPEAKING, BECAUSE TWO AUTOSOMES
DO NOT SEGREGATE AND
(INCOMPREHENSIBLE). MORE
RECENTLY WHEN GEORGE TOOK A
LOOK AT THE TRANSLOCATION
CHROMOSOME, WHERE THE SECOND
CHROMOSOME GOT Y CHROMOSOMES
FRAGMENTED THAT CONTAINED
RIBOSOME DNA, THE SECOND
CHROMOSOME NOW SEGREGATES --.
FROM THERE, I MEAN, LED THE
DEFINED RIBOSOME DNA, WE WERE
DOWN TO THE SPECIFIC -- --
WHICH I'M GOING TO COME INTO A
LITTLE BIT LATER. SO THAT IS
SOMETHING WE FIND VERY HARD.
FROM THERE, IT TOOK LIKE FIVE
YEARS. GEORGE ISOLATED A
PROTEIN WHICH WE NAMED INDRA.
IT WAS ISOLATED BASED ON ITS
ABILITY TO BIND TO RIBOSOME DNA.
LATER WE FOUND THAT THIS
REQUIRED NONRANDOM SISTER
CHROMATID SEGREGATION AND IN
ADDITION TO THIS ELEMENT WE
HAVE A TRANS FACTOR THAT SEEMS
TO DO SOMETHING ABOUT THE
SISTER CHROMATID SEGREGATION.
YOU HAVE A MOLECULAR MECHANISM
FOR SOME SAY THAT YOU DON'T
EVEN KNOW THE MEANING. THAT IS
A WEIRD SITUATION. WHAT IS
DIFFERENT BETWEEN SISTER
CHROMATID, EVEN THOUGH WHEN WE
KNOW IT IS RIBOSOME DNA AFTER
IT (INCOMPREHENSIBLE)-- WHAT IS
THE DIFFERENCE? THAT IS A
REMAINING LINGERING QUESTION.
TO ADDRESS THIS QUESTION, I
HAVE TO STEP A LITTLE BIT TO
EXPLAIN WHAT RIBOSOME DNA LOOKS
LIKE.
IT IS A PIECE OF THE MANY
COPIES OF THE RIBOSOME DNA
GENE. OR RIBOSOMAL RNA GENES.
80% OF THE ENTIRE
TRANSCRIPTION IN THE CELL IS
GENERATED RIBOSOME RNA. FOR
THAT REASON YOU CANNOT JUST
MAKE A SINGLE COPY. THAT IS WHY
EVERY SINGLE ORGANISM HAS
HUNDREDS AND HUNDREDS--
SOMETIMES THOUSANDS OF COPIES
OF RIBOSOME RNA. AND IN THE
CASE OF -- ABOUT 200 COPIES OF
EACH X CHROMOSOME AND Y
CHROMOSOME.
IN HUMANS, I THINK THERE IS A
5 RNA ROSI OUT OF
(INCOMPREHENSIBLE) AND EACH
CONTAINS HUNDREDS OF THE
RIBOSOME DNA COPIES.
THAT IS CRITICALLY IMPORTANT
TO SUPPORT OUR CELLULAR, THE
DEMAND OF THE RIBOSOME
BIOGENESIS. THE PROBLEM IS --
IF YOU HAVE REPEATING COPIES
EVERYWHERE BECAUSE THAT IS THE
SAME THINGS, THEY CAN EASILY
FIND HOMOLOGY BETWEEN THE
DIFFERENT COPIES WITH EACH
OTHER. AND IF THEY END UP
RECOMBINING OVER HERE, YOU LOSE
A COPY NUMBER TWO THE EXTRA
CHROMOSOME OR SACO, REDUCING
THE COPY NUMBER ON YOUR
CHROMOSOME.
THIS COPY NUMBER DEDUCTION IS
KNOWN TO BE ONE OF THE MAJOR
CAUSES OF (INCOMPREHENSIBLE) AT
LEAST IN THE EAST. BUT NOT
MUCH OF IT WAS KNOWN ABOUT WHAT
IS HAPPENING IN THE
MULTICELLULAR ORGANISM.
MY FORMER GRADUATE STUDENT
KEVIN LU DECIDED TO TAKE A LOOK
AT THIS COPY, LOOKING AT THE
(INCOMPREHENSIBLE).
HERE I AM SHYING RDNA COPY
NUMBER ON THE XY AXIS AND THE X
AXIS IS A TIME OR GENERATION.
IF YOU LOOK AT YOUNGER MALES,
THEY START WITH THE GOOD COPY
NUMBER, IS SUFFICIENT COPY
NUMBER HERE. BUT THEN IF YOU
FOLLOW THESE FLIES AS THEY AGE,
THIS GONAD LOSES AN RDNA COPY
NUMBER THAT MEANS ALL THE MALES
END UP HAVING DEDUCED COPY
NUMBERS, BECAUSE THIS HAPPENS
IN THE GERM. THAT MEANS THAT IT
IS PASSING REDUCED RDNA. YOU
CAN SEE THIS YOUNG SON-- THEY
ARE YOUNG!-- BUT THEIR DNA COPY
IS REDUCED AND IF THIS KEEPS
HAPPENING IN THE FUTURE, WE
WILL BE LOSING THE ABILITY TO
KEEP GOING.
SO OBVIOUSLY IT HASN'T
HAPPENED LIKELY TO US, OR TO
FLIES EITHER. HOW IS IT?
ONE SHOCKING OBSERVATION
INDEED THE SON WHO STARTED WITH
THE REDUCED COPY NUMBER, HIS
GRANDSON HAD RECOVERED. TWO
GENERATIONS FROM BEGINNING.
THEIR PROGENY HAS NOW AN OKAY
RDNA COPY NUMBER.
THAT MEANS THERE'S A
FLUCTUATION OF THE COPY NUMBER.
WHEN WE FOUND THIS AND WE SAW
THIS DATA, WE REALIZED WAIT A
MINUTE! THIS POD, IT IS NOT
UNHEARD OF WITH THE RDNA COPY
RECOVERY. THAT GOES BACK 50
YEARS. SO THEN, BACK THEN, THIS
ORIGINAL DISCOVERY OF THE
DEDUCED RDNA COPY NUMBER ON THE
CHROMOSOME WAS DISCOVERED KIND
OF BY (INCOMPREHENSIBLE) A
CENTURY AGO.
LATER, SOMETIMES THE STARTING
CONDITIONS THAT THE DUES THE
MUTANT CHROMOSOME WITH MINIMUM
RDNA COPY NUMBER CAN BE INDUCED
IN THE RECOVERY COPY NUMBER AND
THIS PROCESS IS CALLED RDNA
MAGNIFICATION.
THAT IS A PHENOMENON BACK THEN
THAT PEOPLE WERE STUDYING
BECAUSE IT WAS AN INTERESTING
PHENOMENON. BUT BACK THEN
PEOPLE DID NOT KNOW IF THERE
WERE ANY BIOLOGICAL VARIANCES.
NOW WE THINK THAT THIS
MAGNIFICATION IS A PROCESS TO
RECOVER ANY RDNA THAT LOST A
COPY NUMBER BECAUSE OF THE
AGING. OKAY?
AND THE YOUNGEST PERSON BEFORE
US IF WE STUDY THE PHENOMENA IS
HAWLEY. AND THIS MECHANISM, WE
THINK THAT THIS IS THE
MECHANISM FOR THE TRANS
GENERATION, MAINTENANCE OF THE
RDNA COPY NUMBER IN FLIES.
THE GOOD THING IS PRAYER
STUDIES FIGURED QUITE A BIT OF
THE MOLECULAR MECHANISM AND HOW
THIS MIGHT HAVE HAPPENED. THAT
WAS SOMETHING NEW THAT WE
REALLY BENEFITED FROM, BY GOING
INTO THE LITERATURE. WE
DISCOVERED THIS PHENOMENON IN
THE -- FLIES.
THIS IS HOW NORMATIVE FLIES
RECOVERED THE COPY NUMBER. I
AM SURE IT IS TO EXEMPLIFY,
SHOWING JUST A COPIES BUT IS
TYPICALLY MANY MORE.
LET'S SAY IF YOU HAVE TO EIGHT
COPIES OF RDNA, AFTER
DUPLICATION HAVE A SISTER
CHROMATID HERE.
THIS MISALIGN ON PUPAS. AND
THEY GO ON THE SISTER CHROMATID
EXCHANGE HERE. AFTER THIS,
SISTER CHROMATID STEALS A COPY
FROM IT SISTER. THAT MEANS ONE
SISTER CHROMATID RECOVERS A
COPY NUMBER AT THE EXPENSE OF
ITS SISTER. RIGHT?
AWAY WE DREW THIS DIAGRAM WE
WERE THUNDERSTRUCK. WOW! THIS
IS THE SYMMETRY WE WERE LOOKING
FOR, TO EXPAND SISTER CHROMATID
SEGREGATION. WE WERE VERY
EXCITED AND GEORGE STARTED TO
TAKE A LOOK AT THE COPY NUMBER
DIFFERENCE POTENTIALLY IN THE
STEM CELL DIVISION.
DURING THE STEM CELL DIVISION
THE STEM CELLS GOING TO CREATE
A DIFFERENTIATION, CAN WE SEE
THE COPY DIFFERENT?
DIFFERENTIATED CELLS ARE STEM.
WERE KIND OF AGNOSTIC BACK THEN
AND THEN THE DNA FISH CAPTURED
THE CELL DIVIDING ITSELF. IF
YOU WERE TRYING TO LOOK AT A
DIFFERENT-- (INCOMPREHENSIBLE)
-- WE WERE VERY DISAPPOINTED,
THERE IS NO DIFFERENCE!
MAYBE THIS IS BEYOND THE
RESOLUTION OF THE DNA FISH.
BECAUSE A 5% IN THE DIFFERENCE
IN THE COPY NUMBER IS STILL
MEANINGFUL. WE WERE VERY
DISAPPOINTED ABOUT THE -- MAYBE
WE SAID, I DON'T KNOW WHAT IS
THE NEXT STEP FOR THIS PROJECT.
ONE DAY, GEORGE CAME TO ME. I
THINK WE HAVE BEEN LOOKING AT--
HE TOLD ME-- THE WRONG
CONDITION. THIS IS THE
CONDITION WHERE IT IS NOT
TEMPTED TO INCREASE THE COPY
NUMBER. THEY ARE FINE. WITH
THAT IDEA GEORGE WENT TO TAKE A
LOOK AT THIS MAGNIFYING
CONDITION THAT HAS MINIMAL RDNA
COPY NUMBER AND NOW WE WERE
STARTING TO SEE THE DIFFERENCE
BETWEEN THE STEM CELL SITE AND
THE DIFFERENTIATING SIDE. THE
STEM CELL WAS ALWAYS INHERITING
A LITTLE BIT MORE RDNA. YOU
CAN QUANTIFY OVER HERE. THE
SITUATION ON THE COPY NUMBER
SITUATION WE SEE A NUMBER. AND
ONCE THERE IS A MAGNIFYING
CONDITION HERE.
I AM NOT SURE IF I WILL
BELIEVE THAT THIS IS THREE
TIMES DIFFERENT WITH THE FISH,
IF THIS WERE THE FASTER DATA WE
GOT THERE WE SPENT MANY MONTHS
WANTED TO SEE SOMETHING OVER
HERE AND NO MATTER WHAT WE
WANTED TO SEE WE DID NOT SEE
IT.
AFTER THIS VERY SQUEAKY
NEGATIVE DATA WE WERE ABLE TO
BELIEVE THIS.
ON TOP OF THAT THIS INDRA THAT
IS REQUIRED FOR SISTER
CHROMATID SEGREGATION NOW
REDUCES ASYMMETRY COMPLETELY.
SO I'M GOING TO COME BACK.
THIS IS ACTUALLY HINTING ALSO
AT THE -- FUNCTION. AT THIS
POINT THE INDRA, THE MUTANT
CANNOT DO THIS ASYMMETRIC
SEGREGATION. THAT IS WHAT WE
THINK NOW. SO JUST TO
SUMMARIZE, UP UNTIL THIS POINT,
THIS IS KIND OF LIKE A VERY
EXPECTED AND MEANINGFUL SISTER
CHROMATID SEGREGATION. WE
FOUND FIRST OF ALL RDNA AND IS
BINDING PROTEIN INDRA, PART OF
THE SISTER CHROMATID
SEGREGATION. AND INSTEAD OF
THE GENETIC INFORMATION
DISTRIBUTED ACROSS THE ENTIRE
CHROMOSOMES, WE THINK THIS
ELEMENT IS LOCALIZED, THE RDNA,
AND I EXPECT THAT IS A COPY
NUMBER.
WHAT ARE THEY DOING THIS? WE
THINK IT IS FOR THE SAKE OF
GERM IMMORTALITY OR TO MAKE AN
RDNA COPY NUMBER FOR MANY
GENERATIONS SO THEY DO NOT LOSE
THIS INTEGRITY.
THE GENOME INDRA COMES FROM
THE HINDU GOD WHO LOSSES
IMMORTALITY BECAUSE OF CASTE.
SO THERE IS GERMLINE
IMMORTALITY AND THAT IS WHY WE
NAMED THE GENE INDRA.
BEFORE GOING TO THE NEXT
SECTION OF HOW THE INDRA MIGHT
BE FUNCTIONING IN THE
MECHANISM-- IT IS FAIRLY NEW,
LAST MONTH OR SO-- ALL OF A
SUDDEN WE STARTED GETTING
REALLY INTERESTING DATA.
I WANT TO TELL YOU A LITTLE
BIT ABOUT THE SLIGHTLY
DIFFERENT ASPECT OF THE
MODIFICATION. ANOTHER THING
THAT WE FOUND IS THIS IS WORK
BY POSTDOC JONATHAN NELSON WHO
IS HEADING TO HIS OWN LAB AT
THE STONY BROOK SOMETIME SOON
IN THE NEXT FEW MONTHS.
TO INDUCE THIS SISTER
CHROMATID EXCHANGE INCREASE
COPY NUMBER THE FASTER STEP
THAT YOU HAVE TO DO IS TO BREAK
THE DNA. OTHERWISE NOTHING
WOULD HAPPEN.
JONATHAN FOUND THAT THIS IS
ACTUALLY TRANSPOSAL. THAT HAS
ONLY BEEN KNOWN IN THE RDNA,
AND THAT IS THE KEY TO INDUCE A
MODIFICATION. THAT IS
INTERESTING. THE PHENOMENA
WHERE THE GENETIC PARASITE, THE
TRANSPOSAL HAS ADOPTED AND NOW
HAS A SYMBIOTIC RELATIONSHIP AS
A HOST.
WHEN WE STARTED THIS WORK WE
WERE PUZZLED. HOW WOULD THE
HOST WANT ANY TRANSPOSAL TO GO
SPECIFICALLY INTO THE MOST PART
OF THE GENOME? IT TURNED OUT
INDEED THAT IT IS CO-OPTED AND
INDUCED THE MODIFICATION ONLY
WHEN IT IS REQUIRED.
SO THEN, NOW THERE IS LITTLE
BIT OF A QUESTION, AT THE
BEGINNING WERE AGNOSTIC ABOUT
WHICH CELLS, OR WHICH
DIFFERENTIATED CELLS INHERITED
THE COPY NUMBER.
MANY PEOPLE CAME TO ME AND
SAID HEY IT DOES NOT MAKE ANY
SENSE. YOU HAVE TO BE WRONG!
THE STEM CELL INHERITING MORE
COPIES? THAT DOES NOT MAKE ANY
SENSE BECAUSE THIS
DIFFERENTIATING CELL -- IS MUCH
MORE IMPORTANT THAN THE SPERM
GENERATION CELLS.
IF YOU THINK ABOUT IT IT MAKES
SENSE. IF STEM CELL MAGNIFIES
THE COPY NUMBER, THE STEM CELL
IS GOING TO MAKE THE
DIFFERENTIATED CELL IN THE NEXT
CELL DIVISION.
THIS KEEPS GOING AND THE
DIFFERENTIATED CELL DOWNSTREAM
OF ALL OF THOSE POPULATION
START INHERITING THE RECOVERY
COPY NUMBER.
ON THE OTHER HAND, IF ONE
TIME IN THE INCREASE OF THE
COPY NUMBER GOES TO
DIFFERENTIATING CELL, THIS
DIFFERENTIATING CELL DOES NOT
SUSTAIN. THAT MEANS THAT WITH
THE CELL DIFFERENTIATION KEEPS
GOING DOWN, AND YOU WILL BE
DOWN ONE MAGNIFICATION.
IF STEM CELL ARE INHERITING
THE GOOD COPY, RED IS A COPY
NUMBER AND THE BLUES COPY
NUMBER, YOU START WITH KIND OF
A REDDISH COLOR. AND AFTER
DIFFERENTIATING CELLS THEY GET
A HIT. YOU LOSE THE COPY
NUMBER.
IN THE NEXT DIVISION THE STEM
CELL RECOVERS EVEN MORE AND IN
THE NEXT ROUND OF THE
DIFFERENTIATING CELL, EVEN IF
IT IS SEVERAL DEGREES, IT
STARTS THE TEMPLATE BETTER THAN
PREVIOUS GRADUALLY INCREASES
THE COPY NUMBER AND THE
DIFFERENTIATED CELLS START TO
RECOVER A COPY NUMBER, SEVERAL
CYCLES BACK BUT HE CAN STILL
RECOVER IT.
ON THE CONTRARY IF YOU DO THE
OPPOSITE THEY KEEP LOSING THE
COPY NUMBER, TO GIVE THE BETA
COPY TO THE DIFFERENTIATING
CELL. BUT THE DIFFERENTIATING
CELL CAN KEEP RECEIVING THE
COPIES FROM THE STEM CELL.
THAT HAS A SEVERELY REDUCED
COPY NUMBER; THAT MEANS BOTH OF
THEM KEEP DECREASING THE COPY
NUMBER, EATING THE --.
NO MATTER THE CONDITION THERE
IS NO COPY NUMBER DIFFERENCE
BETWEEN SISTERS, NO MATTER WHAT
HAPPENS. STARTING FROM 200
COPIES, AFTER MANY DIVISIONS
WERE PRETTY MUCH NOTHING
HAPPENS.
BUT IN THE MAGNIFYING
CONDITION, IN THE INITIAL COPY
NUMBER SAY STARTING FROM A
REDUCED 100, BUT THEN THE
SISTER CHROMATID THAT HAPPENS
AFTER 50% IN THE GENERATED STEM
CELL, AFTER THE EXCHANGE STEM
CELLS DO NOT HAVE ANY
PREFERENCE. YOU DON'T GET ANY
COPY NUMBER INCREASE. SOME AND
UP WITH AN INCREASING COPY
NUMBER. SOME OF THEM LOOSE. NET
ZERO INCREASE.
IF STEM CELLS ARE GETTING THE
BETTER COPY, THE -- START TO
RECEIVE A HIGHER COPY NUMBER,
NOT LOSING THE RECOVERED COPIES
AND THE SPERM AFTER TWO WEEKS
-- AVERAGE COPY NUMBER IS SO
MUCH MORE REDUCED. THAT REALLY
TELLS. THIS IS PROBABLY
UNNECESSARY ADDITIONAL
MODELING.
THE KEY MESSAGES THAT IT IS
THE STEM CELL RECEIVES THE
MODIFICATION TO INCREASE THE
COPY NUMBER.
SO IS THIS LIMITED TO -- STEM
CELL? BREAKING THE CELL IS
TRICKY BUSINESS. THAT WAS
ANOTHER THING WE WANTED TO
ADDRESS. WE COULD ACTUALLY
TAKE A LOOK AT THE OCCURRENCE
OF THE SISTER CHROMATID
EXCHANGE BY LOOKING AT THE DNA
ABLUTION BREAK, PROVIDING
PHOSPHORYLATED
(INCOMPREHENSIBLE).
IN A NORMAL CONDITION IN THE
STEM CELL AREA YOU BARELY SEE
THE DNA DAMAGE. IF YOU GIVE
THIS REDUCED COPY NUMBER THE
CONDITION, THE MAGNIFYING
CONDITION YOU START TO SEE THE
DNA DAMAGE AROUND HERE. THIS
IS ONE DATA TO SHOW THAT
TRANSPOSAL IS IMPORTANT. IF YOU
LOOK AT THE TRANSPOSAL TO STOP
SEEING THE DNA DAMAGE. OKAY?
NOT ONLY THAT. THIS COMPARISON
IS BETWEEN STEM CELL AND
DIFFERENTIATED CELL, AND IN
WHAT CONDITION. THE CONTROL
SITUATION, MAGNIFYING
CONDITION, MAGNIFYING PLUS
MINUS 2 IN THIS INHIBITED.
DNA STRAND BREAK HAPPENS.
THERE IS ONE DAY OF
DEREGULATION. ON TOP OF THAT
BY COMPARING THIS OVER HERE,
EVEN UNDER MAGNIFYING
CONDITIONS, DNA BREAK IS PRETTY
MUCH LIMITED TO THE STEM CELL
AND THE DIFFERENTIATING CELLS
DON'T DO IT. AND THAT KIND OF
MAKES SENSE BECAUSE WE
PREVIOUSLY PUBLISHED THAT
DIFFERENTIATING CELLS ARE
ACTUALLY SENSITIVE TO DNA
DAMAGE TO MAKE SURE THAT THE
GENOME INTEGRITY IS PRETTY GOOD
SO THE TEENIEST BIT OF DNA
DAMAGE DOES NOT DO ANYTHING TO
THE SISTER CHROMATID CELLS, BUT
THAT IS ENOUGH TO KILL THE
CELLS.
SO NOT ONLY IS THERE RDNA
BREAKS, AS YOU CAN SEE OVER
HERE ONLY WHEN MAGNIFYING
CONDITIONS HAPPEN, THE RDNA
COPY NUMBERS LOWER, R2 COMES
OUT OVER HEREBUT ONLY IN THE
STEM CELL AND THE EXPRESSION IS
MUCH MORE SUPPRESSED.
SO ONE THING. THE ONE
CHALLENGE WE FACED WHEN
STUDYING THIS, WAS THE OFTEN
MANY PEOPLE ARE SKEPTICAL.
THIS IS TRANSPOSED. THE DATA
GOT PARASITED. THEY CANNOT BE
BENEFICIAL TO THE HOST. BUT
OUR DATA WAS PRETTY CONVINCING,
WE THOUGHT, IN TERMS OF THE
TRANSPOSAL HELPING THE
MODIFICATION AND HELPING RDNA
COPY NUMBER MAINTENANCE.
IF INDEED R2 IS AN IMPORTANT
PART OF THE HOST BIOLOGY, THIS
SHOULD BE SIGNALING PATHWAY
THAT HAS TO BE INTEGRATED,
TOGETHER WITH R2 EXPRESSION SO
WHAT HE DID WITH A SINGLE CELL
RDNA SEQUENCING, WHAT HE DID
WAS COMPARE TWO CONDITIONS.
FIRST OF ALL HE USED A STEM
CELL TUMOR. BECAUSE THIS IS A
SINGLE CELL YOU CAN ISOLATE OR
CONCENTRATE STEM CELL AND ALSO
HE COMPARED LOW RDNA COPY
NUMBER AND THE HIGH RDNA COPY
NUMBER SITUATIONS AND THE
GENIUS UPREGULATED UNDER THE
LOWER DATA COPY NUMBER.
I'M GOING TO GIVE A COUPLE OF
EXAMPLES, DOWN REGULATED RDNA
COPY NUMBER, THE CANDIDATE FOR
THE INHIBITED MODIFICATION, OR
YOU DO NOT HAVE TO MAGNIFY,
THAT IS THE REGULAR DATA.
THAT GIVES A COUPLE OF
INTERESTING GENES. ONE WAS
INSULIN RECEPTOR. WE ARE GOING
TO SKIP OVER THIS, BUT I WILL
TELL YOU THAT IT IS PRETTY
CONVINCING. IF YOU MODULATE
THIS NUTRIENT SENSING PATHWAY,
AND THAT COMPLETELY REGULATES
THIS MAGNIFICATION, THAT MEANS
WE THINK THAT THIS IS PROBABLY
SENSING THE ABILITY TO MAKE
LOTS OF RIBOSOME. IF THAT IS
COMPROMISED THAT IS THE
INCIDENT PATHWAY, IT TELLS R2
TO EXPRESS OR NOT.
I'M NOT GOING TO GET INTO THIS
TO VERY MUCH. ACTUALLY I DON'T
HAVE IT, THAT IS GOOD. THE ONE
THING THE INSULIN RECEPTOR,
THEN WE USE THE INSULIN
RECEPTOR NUTRIENT. IT TOLD IS
THAT THE INSULIN RECEPTOR IS
NEGATIVELY REGULATING THE
MAGNIFICATION PROCESS. THAT
MEANS EVEN WHEN THEY WANT TO
MAGNIFY, IF YOU ACTIVATE
INSULIN PATHWAY -- DON'T CARE.
AND ANOTHER THING IS THAT THE
PATHWAY WAS PUT OUT. THAT MAKES
SENSE. WHY DO YOU HAVE A
PATHWAY TO DEPRESS TRANSPOSAL?
IT IS NOT JUST TO DEPRESS. I
WILL REPHRASE THIS. TO FINE
TUNE THE TRANSPOSAL INSTEAD OF
JUST DEPRESS. IF YOU DON'T NEED
THE TRANSPOSAL TO COME OUT
BECAUSE YOU ARE DEPRESSING IT
THAT IS BECAUSE THE CELL HAS A
SITUATION WHERE YOU WANT TO
HAVE A LITTLE BIT OF THE
TRANSPOSAL TO COME OUT THAT IS
WHY THE PATHWAY BECOMES HANDY.
NOW I'M GOING TO GIVE YOU A
LITTLE BIT IN THE LAST TWO
MINUTES ABOUT THE FUNCTION OF
INDRA. WE KNOW THAT INDRA
BINDS TO RIBOSOME DNA. EXACTLY
HOW DOES THAT WORK? THAT IS
GEORGE'S WORK IN THE LAST TWO
MONTHS OR SO.
THIS ONE I TOLD YOU, THIS IS
THE SAME THING, THE ASYMMETRIC
COPY NUMBER, NORMAL CONDITION,
MAGNIFIED, AND MAGNIFIED MUTANT
CONDITION.
IN THIS WHEN YOU CAN SEE NO
SYMMETRY AT ALL. INDRA IS NOT
QUITE SEGREGATION PER SE, BUT
IT IS ABOUT CREATING ASYMMETRY.
RIGHT? BECAUSE IF IT IS SIMPLY
REQUIRED FOR SEGREGATION, YOU
MIGHT CREATE LOTS OF COPY
NUMBER ASYMMETRY. BUT IT MIGHT
GO THE RANDOM WAY. SO I
FABRICATED THIS DATA BY COPYING
THIS FROM THIS DATA LAST NIGHT
TO SHOW WHAT YOU WOULD EXPECT
IF YOU ARE MAKING THAT COPY
NUMBER ASYMMETRY. BUT THEY ARE
NOT SEGREGATING CORRECTLY.
THIS DOES NOT HAPPEN. THAT
MEANS INDRA WAS CREATING A COPY
NUMBER ASYMMETRY.
SO AS I TOLD YOU, AND I
BRIEFLY MENTIONED, INDRA BINDS
TO THIS ONE SPECIFIC SEQUENCE
IGS. THAT IS WHERE THE
PROMOTER OF RIBOSOME DNA GENE.
CURIOUSLY PEOPLE KNOW THAT
THERE ARE TWO RIBOSOME
PROMOTERS. WHY TWO?
BECAUSE IT IS A PROMOTER WE
DECIDED TO TAKE A LOOK TO SEE
IF INDRA HAS ANY FUNCTION IN
THE DISCRETION OF IGS. IT
TURNED OUT THAT INDEED YES. IN
LABORATORY CONDITIONS YOU GET A
LITTLE BIT OF IGS EXPRESSION.
BUT IN THE INDRA MUTANT GETS
REALLY UPREGULATED OVER HERE.
IF YOU DO THIS, FIRST OF ALL
OVER HERE YOU CAN SEE THE INDRA
MUTANT UPGRADES CONSIDERABLY
IGS. INTERESTINGLY THIS UP
REGULATION IS VERY SPECIFIC TO
STEM CELL. THIS IS INDRA
MUTANT, STILL THIS POPULATION
OBSERVED ONLY IN THE STEM CELL.
THAT MEANS WHATEVER THE
TENDENCY IS TO EXPRESS IGS THAT
ITSELF IS LIMITED TO THE
GERMINIZED STEM CELL.
INDRA'S DEPRESSING THE
EXPRESSION WITH THE ABILITY TO
EXPRESS AT ALL IS LIMITED TO
THE STEM CELL, BUT NOT THE
DIFFERENTIATING CELL. THIS WE
DON'T OBSERVE ANY UP REGULATION
AS PART OF THE RDNA. THIS
PROMOTER IS THE SAME, ETS.
THERE IS NO UP REGULATION AT
ALL IN THESE GUYS. SO IGS UP
RELATION BY INDRA IS STEM CELL
SPECIFIC. WHAT DO WE DO?
WE TOOK THE HINT FROM YEAST.
HERE WE HAVE COPIES OF THE RDNA
COPIES AND HERE IT IS ALSO
DEPRESSED. WHEN THE CELL LOSES
A COPY NUMBER IGS INDEED GETS
UPREGULATED.
IF YOU EXPRESS FROM HERE RIGHT
HERE THERE IS A DNA REPLICATION
ORIGIN, THERE IS A CONFLICT OF
TRANSCRIPTION AND DNA
REPLICATION THAT ENDS UP
BREAKING THE DNA. THAT DNA
BREAK IS ACTUALLY USED IN
SISTER CHROMATID EXCHANGE TO
INCREASE THE COPY NUMBER.
SO TRANSPLANTING THIS IDEA TO
WHAT DO WE KNOW IN DROSOPHILA?
INDRA IS OVER HERE. WE STARTED
THINKING THAT MAYBE INDRA IS
DEPRESSING NORMALLY TO DEPRESS
THE MAGNIFICATION AND THAT
BECAME THE IDEA.
WE TESTED THE IDEA. WE FIRST
TOOK A LOOK AT THE IGS
EXPRESSION IN A NORMAL
SITUATION VERSUS THE MAGNIFIED
CONDITION AND THIS IS NOT THE
INDRA MUTATION IF YOU LOOK AT
OVER HERE, THE STEM CELL, THE
UPREGULATED IGS OVER HERE IN
THE MODIFY CONDITION AND
COMPARED TO THE NORMAL
CONDITION YOU GET UP REGULATION.
IF YOU COMPARE AND YOU
NORMALIZE THE IGS EXPRESSION BY
DIFFERENTIATING CELLS, THE STEM
CELL DIVIDED BY DIFFERENTIATING
CELL, IT GETS EVEN CLEARER. THE
STEM CELL EXPRESSES IGS WE NEED
TO MODIFY THE RDNA COPY NUMBER.
IF YOU OVEREXPRESSED INDRA,
YOU CAN SUPPRESS IT. THAT TELLS
US INDRA IS A NEGATIVE UP
REGULATOR FOR IGS EXPRESSION
AND THAT LEADS TO THE NEXT
QUESTION. DOES INDRA REPRESS
MAGNIFICATION?
I DON'T HAVE A SLOT HERE FOR
YOU BUT WE ALSO DISCOVERED
INDRA PROTEIN ON LONG ITSELF
ACTUALLY GOES DOWN DURING
MAGNIFICATION. THAT MEANS INDRA
PROTEIN AMOUNT IS SENSITIVE TO
THE RDNA COPY NUMBER.
THIS IS AS IT WAS EXPECTED THE
MODIFIED CONDITION YOU DESERVE
THE DNA BREAKING CELL AND IF
YOU EXPRESS INDRA YOU GET THE
SAME THING. THIS IS THE CASE
OF THE MODIFICATION AND THE
MODIFYING CONDITION, YOU ARE
OVEREXPRESSING INDRA, YOU WILL
DEPRESS IT.
ACTUALLY THIS IS THE DATA. THE
INDRA AMOUNT ACTUALLY GOES DOWN
WHEN YOU HAVE A REDUCED RDNA
COPY NUMBER.
IS THAT REPLICATION
TRANSCRIPTION CONFLICT THAT
REGULATES RDNA MAGNIFICATION_
THIS IS A GENE ON TOP OF THAT.
INDRA IS A TRANSCRIPTION
REGULATED, THAT BECAME A
QUESTION. PERHAPS INDRA IS
REPRESSING RDNA EXPRESSION
ITSELF AND IN THE MUTANT THE
UPREGULATED ATTITUDE AND ALSO
IN MAGNIFYING CONDITIONS R2
SHOULD BE UP BUT THAT CAN BE
REPRESSED BUT OVEREXPRESSING
INDRA.
SO THIS ACTUALLY LEAD TO THE
SORT OF UNIFY MODEL.
PREVIOUSLY THIS IS WHAT PEOPLE
KNEW. WHEN PEOPLE THOUGHT R2 IS
PURELY A PARASITE, PEOPLE
FIGURED THAT THE RDNA INSERTED
IN R2 IS COMPLETELY REPRESSED
BY AN UNKNOWN MECHANISM BUT IT
WAS CLEAR THAT RDNA INSERTED
INTO R2 IS NOT EXPOSING AT ALL
AND THE ONLY GOOD COPY IS
EXPRESSED. NOW UPSTREAM
ADDITIONAL PROMOTER CAME INTO
THE PICTURE. THAT IS IGS
BOUNDED BY INDRA.
AND THIS IS WHAT WE THINK IS
HAPPENING. UNDER MAGNIFYING
CONDITION, THIS PROMOTER IS
ALWAYS REPRESSED UNDER NORMAL
CONDITIONS. THIS NORMALLY DOES
NOT COUNT. ONLY TRANSCRIPTION
THAT ARE HAPPENING BY USING
THIS PROMOTER ETS OVER HERE IS
A NORMAL CONDITION BUT NOW
UNDER MAGNIFYING CONDITION,
WHEN THIS STEM CELL LOSES AN
RDNA COPY NUMBER-- NOW THIS NEW
PROMOTER COMES INTO THE PICTURE
AND WE THINK THIS EXPRESSION IS
NONDISCRIMINATORY OF IGS.
INSERTED IN R2. THIS PROBABLY
RESULTS IN R2 EXPRESSION WITH
THE MAGNIFICATION THROUGH THE
SISTER CHROMATID EXCHANGE AND
THAT IS ESSENTIALLY THE ENTIRE
MESSAGE, LEADING TO
MAGNIFICATION.
LET ME GIVE YOU THE SURPRISE.
SO THIS MODEL SAYS BECAUSE RDNA
TYPICALLY IS IMPORTANT AS S1,
WHAT ABOUT THE SECOND PROMOTER?
WHO IS DOING IT?
IF THE SAME POLYMER IS COMING
INTO THE TWO DIFFERENT
PROMOTERS THERE IS NOT MUCH
DIFFERENTIATION YOU CAN DO AND
THIS IGS UP REGULATION UNDER
MAGNIFIED CONDITIONS ENTIRELY
UP TO POLYMERS TO MODIFY THE
STEM CELL.
THIS IS BUSY DATA. IT IS
SHOCKING. IT IS MUCH MORE
VISUALLY APPEALING. IT IS A
SIMPLE IMMUNOFLUORESCENCE
LOOKING AT THE RDNA POLYMER IN
LABORATORY. THIS IS BORING.
WHEN YOU GIVE THIS MAGNIFYING
CONDITION, THE RDNA COPY NUMBER
GOES DOWN, ALL OF A SUDDEN RDNA
POLYMERS COME AROUND, THIS IS
RDNA SYNTHESIS, THE FACTORY.
ALL OF A SUDDEN THE RDNA COPY
NUMBER GOES DOWN AND RDNA DOES
TOO, GET RECRUITED TO IGS. THAT
IS POSSIBLE BECAUSE INDRA GOES
AWAY AND ALLOWS R2 TO COME IN.
TO SUMMARIZE TODAY'S TALK,
RDNA LOCUS IS VULNERABLE AND
SUPER IMPORTANT TO MAINTAIN THE
CELLS' HELP BECAUSE YOU LOSE A
COPY NUMBER ALL THE TIME. YOU
HAVE TO RECOVER TO MAINTAIN THE
GERMLINE IMMORTALITY TO LAST
THE 1.5 BILLION YEARS OTHER
MULTICELLULAR LIFE AND TO DO
SO, THIS RECOVERY IS MEDIATING
BY AN SISTER CHROMATID
EXCHANGE. THAT IS A REAL KEY TO
REJUVENATE RIBOSOME DNA. I
ALSO TOLD YOU THAT IS DONE IN
THE STEM CELL; THIS IS
HAPPENING SPECIFICALLY IN THE
GERMANIZED CELL.
ALSO THE ACTIVATION OF THE
SPECIFIC PROMOTER THAT BRINGS
IN RDNA POLYMERS TOO.
SO THAT LEADS TO
DIFFERENTIATED COPY NUMBER,
WHICH IS FURTHER REGULATED BY
SISTER CHROMATID SEGREGATION
THAT ALLOWS THE STEM CELL TO
INHERIT A COPY NUMBER AND SO
NOW YOU HAVE HOW IT IS
GERMINATING.
THIS IS MY LAB. IT TOOK 10
YEARS. BOTH OF US ARE VERY
HAPPY. FINALLY WE FIGURED IT
OUT. THE R2 POD WAS DONE BY
JONATHAN NELSON. THANK YOU
VERY MUCH FOR YOUR ATTENTION
AND NOW AM HAPPY TO TAKE ANY
QUESTIONS YOU MIGHT HAVE.
THANK YOU.
>> [APPLAUSE]
>> YOU DON'T WANT-- LIKE
INDRA--
>> DR. YAMASHITA: YOU ARE RIGHT.
FOR LONGEST TIME WE THOUGHT
INDRA WAS REQUIRED FOR
SEGREGATION PER SE. BUT WHAT
IS HAPPENING WITH INDRA IS THAT
IN ABSENCE OF INDRA I JUST
EXPRESS SO MUCH. ACTUALLY THEY
ENDED UP RECOMBINING MANY, MANY
TIMES. AND THAT EQUALIZED TO
CHROMATID AND THAT IS
WHY THE CHROMOSOMES GET
CONFUSED ABOUT WHICH STRAND TO
BRING TO THE CELL SO INDRA DOES
NOT SEEM TO BE THE MACHINERY TO
DIFFERENTIATE THE DNA. WE HAVE
TO REALLY STUDY HOW-- THE
DIVISION APPARATUSES
DIFFERENTIATING ONE COPY FROM
THE OTHER. AND THAT IS STILL A
BIG QUESTION.
>> THANK YOU.
>> HI THAT WAS FASCINATING.
THANK YOU VERY MUCH. I HAVE AN
GENERAL EVOLUTIONARY QUESTION.
IF YOU HAVE 200 COPIES, PER
GENE THERE IS SLOW SELECTION
PRESSURE TO KEEP THAT
CONSISTENT. HOW DOES THAT WORK?
IS THERE ANY DIFFERENCE IN THE
MUTATION RATES AT THE END
VERSUS THE MIDDLE?
>> DR. YAMASHITA: EXACTLY SO
TYPICALLY THE MIDDLE COPIES ARE
WELL MAINTAINED. THAT IS
BECAUSE-- ACTUALLY THAT IS
BECAUSE OF THE SISTER CHROMATID
EXCHANGE AND IF YOU DO THAT
EXCHANGE OVER HERE, THE MIDDLE
COPY GETS DUPLICATED. RIGHT? AS
A RESULT THEY HOMOGENIZE THE
MIDDLE COPY. IF YOU GO TO THE
END, SAME THING FOR EXAMPLE
WITH (INCOMPREHENSIBLE) --
TOWARDS THE END THEY USE STUDY
SEQUENCE THAT ARE DEGRADING BUT
IN THE MIDDLE IT IS WELL
CONTROLLED. THAT MEANS IF
ANYTHING GETS THE MUTATION THAT
IS GOING TO SWEEP MOST OF THE
MIDDLE PART OF THE SEGMENT. IN
CASE OF RDNA, IF YOU'RE MUTATED
IN A WAY THAT IS GOING TO
COMPROMISE OUR RDNA FUNCTION
THAT WILL BE SELECTED OUT.
>> THANK YOU.
>> THANKS THAT WAS A GREAT
TALK. HOW MUCH OF THIS IS
CONSERVED IN FEMALES?
>> DR. YAMASHITA: SUPER
QUESTION. WE HAVE NOT BEEN
ABLE TO REALLY DO THE CO-FISH
IN A FEMALE. BUT THERE IS AN
INTERESTING OBSERVATION FROM 50
YEARS AGO. FEMALE DOES NOT
UNDERGO MODIFICATION. THAT IS
REALLY INTERESTING. WHY IS IT?
WE DO NOT KNOW, BUT THE FEMALE
DOES NOT DO IT.
>> THAT WAS FANTASTIC. I AM
CURIOUS SINCE YOU RAISED THE
SPECTER OF -- SATELLITE IS
THERE ANY ROLE FOR INDRA IN
MITOSIS? OR IS IT SPECIFIC JUST
FOR RDNA?
>> DR. YAMASHITA: IT REALLY
LOOKS LIKE IT IS JUST RDNA.
>> SO I WAS PUZZLED BY THAT.
WHY IS IT SO SPECIFIC FOR RDNA?
>> DR. YAMASHITA: WE DON'T
KNOW. WE DO NOT KNOW ABOUT
INDRA. BUT IT IS ONE NEW GENE
THAT WE STARTED WORKING ON. WE
STARTED WORKING ON IT. THIS
GENE (INCOMPREHENSIBLE) -- THIS
PROTEIN BINDS TO ALL THE
OVERSATURATED DNA PLUS THE RDNA
DNA. IT WAS A GOOD CONNECTION.
>> I HAVE A COUPLE OF QUESTIONS
ONLINE. THE PROBLEM IS I DON'T
KNOW WHAT THE PRONOUN "IT"
MEANS. THE QUESTION IS HOW
DOES IT HAPPEN IN HUMAN GERM
CELLS AS WELL?
>> DR. YAMASHITA: I WOULD LOVE
TO KNOW THAT. JUST BY
LOGICALLY DEDUCING IT HAS TO BE
HAPPENING. WE HAVE NOT LOST OUR
DNA COPIES YET OBVIOUSLY. BUT
THE HUMAN AS WELL AS MOUSE
SPERM CELLS, THERE BELIEVED TO
DIVIDE SYMMETRICALLY. WHEN YOU
IDENTIFY SPERM -- STEM CELL,
THEY DON'T EXHIBIT ANY SIGN
THAT THEY ARE DIVINING
ASYMMETRICALLY IN TERMS OF THE
WEIGHT. PEOPLE HAVE DONE
LINEAGE TRACING AND MODELING
USING RODENT SPERMODOGONIA
(SOUNDS LIKE) STEM CELL. SOME
OF THE CELLS TAKE OVER AND THEY
BECOME STEM CELLS, SO THEIR
FATE VERY PLASTIC. ARE YOU A
STEM CELL? ARE YOU
SPERMODOGONIA? DOES NOT MATTER.
BUT IF YOU THINK ABOUT THE RDNA
COPY NUMBER THAT IS NOT PART OF
THE FATE. IF YOU HAVE A LITTLE
BIT MORE OR LESS OF COPY NUMBER
YOU HAVE THE SAME
"TRANSCRIPTION," AND YOU
IDENTIFY THEM AS EQUAL STEM
CELLS.
BUT I BET THERE SHOULD BE SOME
POPULATION THAT IS DOING THIS,
INCREASING THE COPY NUMBER.
THERE IS A BEAUTY IN THE
REPEATEDLY DOING THIS. IF YOU
ARE SUPPOSED TO HAVE 200
COPIES, AND IF YOU GO DOWN TO
50, IF YOU DO ONLY ONE DIVISION
YOU MIGHT RECOVER ONLY 60. AND
THAT'S IT. BUT WHEN THE CELLS
CAN DO THIS REPEATEDLY 60
BECOMES 70 AND BECOMES 80. IN
THAT KIND OF REASONING MAKES ME
WONDER THAT IF YOU ARE LOOKING
AT THE RDNA COPY NUMBER I THINK
THE HUMAN STEM CELL SHOULD BE
DIVIDING ASYMMETRICALLY. THAT
WOULD BE MY GUESS.
>> ONE MORE QUESTION FROM OUR
LINE. IS POLE 2 USED IN MAKING
RIBOSOMES OR DOES IT SERVE
OTHER PURPOSES?
>> DR. YAMASHITA: TYPICALLY FOR
2, IT DOES NOT BELIEVE TO DO
ANY TRANSCRIPTION FOR RDNA. I
MEAN THAT IS WHAT WE KNOW.
BUT NOT MANY PEOPLE HAVE REALLY
LOOKED AT DIFFERENT CONDITIONS.
THERE IS ONE PAPER THAT SAID
DURING STRESS CONDITIONS-- I
DON'T REMEMBER -- IT IS NOT LOW
COPY NUMBER STRESS CONDITION.
-- WAS OBSERVED COMING INTO THE
NUCLEUS. AT THAT POINT R2
TRANSCRIBED INTO RDNA BECOMES
RIBOZYME? THAT I DON'T KNOW.
>> I HAVE A QUESTION IN THE
HOPE THIS DOESN'T MEAN THAT I
DID NOT SYNTHESIZE ALL THE
PARTS BUT I THOUGHT FROM YOUR
CO-FISH IN THE BEGINNING THAT
WAS NOT FROM THE RDNA LOCUS. IT
IS THE METHOD OF MAGNIFICATION
TRANSPACIFIC SO THE LONGER IS
ALWAYS GOING TO HIT ON ONE
STREAM?
>> DR. YAMASHITA: I THINK I DO
HAVE THE SLIDE UP. I TYPICALLY
KEEP THAT SLIDE SOMEWHERE. YES,
FIRST OF ALL WE DISCOVERED --
SEGREGATION.
HOW IT GOT THE SAME COPY
NUMBER, STILL SEGREGATING --?
INDEED I ACTUALLY ALMOST
SKIPPED ONE PIECE OF CRITICAL
INFORMATION WHEN I SAID, OH, WE
REALIZED THERE WAS ASYMMETRY IN
THE COVER NUMBER. TO BE MUCH
MORE PRECISE THAT IS ASYMMETRY
IN THE PROPENSITY OF THE
INCREASE IN THE COPY NUMBER.
THAT MEANS-- WHAT WE THINK IS
THAT BECAUSE RDNA REPLICATE IN
THE UNIQUE DIRECTIONAL WAY,
BECAUSE OF THAT SPECIFICALLY
FOR RDNA LOCUS, THE STRAND IS
(INCOMPREHENSIBLE). IN THE CASE
OF YEAST, NOBODY DETECTED WHICH
COPY GOES TO WHICH SIDE, BUT IN
CASE OF YEAST TYPICALLY THE
BRAKES HAPPENS AT THE LEADING
STRAND AND REVIEWED DREW THE
CARTOON, YOU WOULD SEE THAT IF
YOU'RE LEADING STRAND BREAKS,
YOU CANNOT GO BEYOND THE
REPLICATION WHERE REPLICATION
IS NOT COMPLETED. IF YOU BREAK
AND TRIED TO GO BEYOND THE --
YOU END UP BREAKING THE
CHROMOSOME AND THE ONLY WAY YOU
CAN REPAIR THE BREAKING
CHROMOSOME IS AT THE POINT OF
THE BREAKING THE LEADING STRAND
HAVE TO GO BACK AND INCREASE
THE COPY NUMBER. IF SOMEBODY
WOULD ASK ME TO BED I WOULD BET
THAT THE LEADING STRAND IS
GOING TO THE STEM CELL. THAT
IS PURE SPECULATION BASED ON
WHAT WOULD MAKE SENSE. WE DON'T
KNOW.
>> THANK YOU.
>> I WANT TO END THE SESSION BY
THANKING DR. YAMASHITA. THANK
YOU EVERYONE FOR YOUR
PARTICIPATION.
>> DR. YAMASHITA: THANK YOU SO
MUCH.