>> GOOD AFTERNOON, EVERYONE. WELCOME BACK TO THOSE WHO HAVE BEEN HERE FOR MUCH OF THE DAY, A WONDERFUL SYMPOSIUM ON MITOCHONDRIAL BIOLOGY AND DISEASE. WELCOME TO THOSE WHO HAVEN'T BEEN HERE ALL DAY & HAVE COME
FOR THE WEDNESDAY AFTERNOON LECTURE BY DR. DOUGLAS WALLACE A MITOCHONDRIAL ETIOLOGY OF METABOLIC AND DEGENERATIVE DISEASES, CANCER AND AGING. I'VE HAD THE PLEASURE OF KNOWING DOUG SINCE WE'VE SHARED INTERESTS IN THE FIELD OF HUMAN GENETICS. HE GOT HIS UNDERGRAD DEGREE AT CORNELL, TWO YEARS IN THE PUBLIC HEALTH SERVICE, HERE? NO, I WAS HOPING YOU WERE IN THE NIH GANG OF THE YELLOW BERETS, BUT WENT ON TO GET HIS Ph.D. AT YALE. AFTER THAT, HE SPENT SEVEN YEARS AT STANFORD, I GUESS HE'S ONE OF THOSE PEOPLE WHO GETS RESTLESS, ENDED UP AT EMORY WHERE HE WAS FOR A SIGNIFICANT PERIOD OF TIME IN A PRODUCTIVE PERIOD OF TIME, ALMOST 19 YEARS, THEN MOVED TO UNIVERSITY OF CALIFORNIA, IRVINE, AND MOST RECENTLY FOUR YEARS AGO HAS MOVED TO THE UNIVERSITY OF PENNSYLVANIA AND CHILDREN'S HOSPITAL OF PHILADELPHIA WHERE HE HAS PERHAPS ONE OF THE LONGEST TITLES I'VE SEEN IN A WHILE, MICHAEL AND CHARLES BARNETT ENDOWED CHAIR IN PEDIATRIC MITOCHONDRIAL DISEASE. HE HAS DEFINED THE FIELD OF HUMAN MITOCHONDRIAL GENETICS, MITOCHONDRIAL DNA, BACK THERE A FEW YEARS AGO DEMONSTRATING THE VERY FIRST EXAMPLE OF A DISEASE THAT IS INHERITED BECAUSE OF MUTATIONS IN MITOCHONDRIAL DNA AND THEREFORE INHERITED THROUGH THE MATERNAL LINE. A LONG LIST OF OTHER ACHIEVEMENTS FOR WHICH HE'S BEEN RECOGNIZED BY ELECTION TO THE NATIONAL ACADEMY OF SCIENCES, INSTITUTE OF MEDICINE, AND MOST RECENTLY RECEIVED THE HIGHEST PRIZE GIVEN IN GENETICS, THE GRUBER FOUNDATION GENETICS PRIZE IN 2012. WE'RE FORTUNATE TO HAVE HIM HERE TO TALK TO US THIS AFTERNOON. PLEASE JOIN ME IN WELCOMING DR. DOUG WALLACE. [APPLAUSE] >> I WANT TO REALLY THANK YOU, FRANCIS, FOR BEING HERE, FOR THE GRACIOUS INTRODUCTION. THANK YOU VERY MUCH, STEVE, FOR ALL THE EFFORT YOU PUT IN HAVING THE FABULOUS SPEAKERS WE HEARD TODAY, FOR ALL OF YOU FOR BEING HERE THIS AFTERNOON. WE'RE AT AS FRANCIS MENTIONED TO ME A UNIQUE TIME. THERE'S NO TIME IN WESTERN MEDICINE, BIOMEDICAL SCIENCE WITH MORE TOOLS AND MORE OPPORTUNITIES TO MAKE REAL IMPACT ON THE HEALTH OF OUR SOCIETY, AT THE SAME TIME WE'RE LOOKING AT CONTINUAL DECLINE IN THE SUPPORT FOR OUR SCIENCES, WITH PARTICULAR ANXIETY AND CONCERN FOR THE FUTURE OF OUR YOUNG SCIENTISTS WHO OF COURSE ARE THE FUTURE FOR THIS FIELD. AND WE HAVE TO ASK OURSELVES AT THIS POINT WHY IS IT THAT OUR SOCIETY IS TURNING AWAY FROM THE CONCEPT THAT BIOMEDICAL RESEARCH COULD ACTUALLY BE A MAJOR ASSET TO THEM, WHY ARE THEY THEN PULLING BACK FROM THEIR SUPPORT? AND I THINK THE REASON FOR THAT IS THAT UNFORTUNATELY, THERE IS AN INVERSE CORRELATION BETWEEN THE FREQUENCY OF THE DISEASES THAT IMPACT OUR SOCIETY, AND OUR ABILITY TO DIAGNOSE THEM EFFECTIVELY AND TREAT THEM. AND THIS MEANS THAT FOR THE VAST MAJORITY OF THE AMERICAN CITIZENS AND ALSO FOR THOSE THROUGHOUT THE WORLD THERE'S A GROWING DISSOLUTIONMENT WITH WHAT MEDICINE CAN DO, SINCE THESE ARE THE VERY PROBLEMS THAT THEY ARE CONFRONTED WITH. SO HAVING THIS PROBLEM IN FRONT OF US WHICH I THINK WE MUST ADMIT IS TRUE WE HAVE TO ASK OURSELVES THEN WHY IS IT WITH ALL THE EFFORT THAT YOU ALL ARE PUTTING IN AND WITH ALL THE SUPPORT THAT GOVERNMENTS AND PRIVATE INDUSTRY HAVE PROVIDED THROUGHOUT THE WORLD, WHY HAVEN'T WE BEEN ABLE TO DO BETTER? WHY HAVEN'T WE CURED A SINGLE CASE OF ALZHEIMER'S DISEASE? WHY IS DIABETES INCREASING? WHY IS OBESITY AN EPIDEMIC, ET CETERA, ET CETERA. AND THERE WAS A PHILOSOPHER OF SCIENCE, TIMUS KUHN WHO PROPOSED A NUMBER YEARS AGO WHEN SCIENTIFIC EFFORT IS INVESTED MORE AND MORE HEAVILY IN TRYING TO SOLVE SOME PROBLEMS, AND YET PROGRESS SEEMS TO NOT BE MOVING FORWARD AT THE RATE YOU WOULD EXPECT, IT MAY BE TIME TO ASK, WHAT ARE THE BASIC PREMISES ON WHICH THE EVERY DID HE EVER DID
HE ENDEAVO
RS ARE ADDRESSED AND ARE THEY THE RIGHT ONES AND WHAT ARE THE PROBLEMS WE'RE CONCERNED ABOUT? THERE ARE ALL THE NEURO PSYCHIATRIC DISEASES, BEING IN PEDIATRICS, I WENT TO LOOK AT PEDIATRIC SEDUCES, ONE OF THE MOST OBVIOUS IS AUTISM, NOW ASSUMED TO BE ONE IN 88 BOYS BEING AFFECTED, OR ALZHEIMER'S, PARKINSON'S, MIGRAINE, DEPRESSION, SCHIZOPHRENIA, OBSESSIVE COMPULSIVE, MYALGIA, CHRONIC DISEASE, VISCERAL DISEASES, GASTROINTESTINAL, OR METABOLIC, TYPE 2 DIABETES, OBESITY, HYPERTENSION, INFLAMMATORY DISEASE, LUPUS, CANCER AND AGING, ALL OF THESE ARE IN FACT THE MAJOR CONCERNS WE HAVE AND YET THEY ARE THE ONES THAT WE SEEM TO BE LEAST ABLE TO ADDRESS. IF KUHN IS RIGHT THAT MAYBE THERE'S SOMETHING WRONG WITH HOW WE APPROACH, WE SHOULD ASK WHAT ARE THE BASIC ASSUMPTION? I'D LIKE TO PROPOSE WESTERN MEDICAL PHILOSOPHY HAS BEEN BASED ON TWO BASIC IDEAS, ONE THAT GOES BACK TO A MAN NAMED VISALIUS 500 YEARS AGO IN IT EL, THE FIRSITALY,THE FIRST TO
DEFINE THE ANAT
OMY OF THE HUMAN BODY, A BREAKTHROUGH, AND ALL FUTURE PHYSICIANS AND FUTURE INVESTIGATORS THEN BEGAN TO SPECIALIZE IN DIFFERENT PARTS OF THE BODY. SO NOW TODAY WE HAVE OPHTHALMOLOGISTS, NEPHROLOGISTS, ALL OR BEGAN-SPECIFIC SPECIALTIES. IT LED TO AN UNSPOKEN COROLLARY, IF YOU HAVE A HEADACHE, YOU GET REFERRED TO THE NEUROLOGIST BECAUSE THE ASSUMPTION IS THERE'S SOMETHING WRONG WITH YOUR HEAD. THAT IS THE IDEA IS IF YOU HAVE A SPECIFIC TISSUE-SPECIFIC SYMPTOM THIS MUST BE DUE TO A TISSUE-SPECIFIC DEFECT. THE OTHER IDEA GOES BACK TO GREGOR MENDEL, OBSERVED A SUBSET FOLLOWED A PATTERN WHERE IT SEEMED LIKE THE ADULT PLANT HAD TWO COPIES OF SOMETHING, EACH SEX CELL GOT ONE OF THOSE COPIES, AND THEN OFFSPRING GOT TWO COPIES, WE CALL THAT THE LAWS OF MENDELIAN INHERITANCE. THE TURN OF THE CENTURY, STUDYING THE ANATOMY OF THE FRUIT FLY, ALMOST EVERY ONE OF THEIR ANATOMICAL PROBLEMS WAS INHERITED ACCORDING TO THE PATTERN AND SOON LEARNED THE CHROMOSOMES FOLLOWED THOSE SAME IDEAS AND THAT LED TO WHAT WE CALL THE MENDELIAN LAWS OF INHERITANCE. THE IDEA THERE'S A LAW OR LAWS OF INHERITANCE HAS AN UNSTATED COROLLARY, IF SOMETHING IS TRANSMITTED IN A FAMILY ACCORDING TO THE LAWS OF MENDEL, GENETIC. IF NOT, IT'S ENVIRONMENTAL. THE TWO IDEAS HELD US IN GOOD STEAD, WE MADE PROGRESS, BUT MAYBE THEY EXHAUSTIONED SOM
EXHAUSTED THEI
R CAPACITY TO SERVE US BECAUSE ALL OF THE MENDELIAN GENES ON CHROMOSOMES ARE WHERE THE ANATOMICAL GENES ARE, THERE WAS AN INTERN CONSIS CONSISTENCY. LIFE IS NOT JUST ABOUT ANATOMY, IT'S ABOUT BEING ANIMATED. IN FACT, YOU'RE THE MOST ANIMATED THING IN OUR KNOWN ENVIRONMENT. AND NEWTON INDICATED, AGAIN HALF A MILLENIA AGO, ANY KIND OF MALTER WAS INANIMATE UNLESS IT WAS ACTED ON BY ENERGY. YOU'RE HIGHLY ANIMATED, ONE OF THE MOST IMPORTANT THINGS ABOUT YOU IS THE FLOW OF ENERGY. SO THEREFORE WE CAN REALIZE LIFE IS REALLY INTERACTION OF ANATOMY, ENERGY AND INFORMATION FOR ANATOMY AND INFORMATION FOR ENERGY. ONCE WE REALIZE ENERGY IS SOMETHING WE SHOULD ALSO BE LOOKING AT, THAT ENWE CAN REALIZE THAT DIFFERENT ORGANS RELY ON ENERGY TO DIFFERENT EXTENTS. SO YOU COULD THEN IMAGINE A SITUATION WITH A PARTIAL ENERGETIC DEFECT AND IT WOULD SPECIFICALLY AFFECT THOSE ORGANS THAT HAD THE HIGHEST ENERGY DEMAND. AND THAT IN FACT TURNS OUT TO BE THE BRAIN, HEART, MUSCLE, RENAL AND ENDOCRINE SYSTEMS, AND SOME ARE NOT INHERIT AND NOT ACCORDING TO THE LAWS OF MENDEL AND THEREFORE THIS WHOLE ASPECT OF INHERITANCE HAD BEEN OVERLOOKED FROM A CLINICAL POINT OF VIEW. SO NOW WE REALLY HAVE BASICALLY FOUR SETS OF PARADIGMS, WHEN WE ADD THESE TOGETHER I THINK, I HOPE TO BE ABLE TO CONVINCE YOU WE'LL HAVE A MORE COHERENT VIEW OF THE DISEASE PROCESS. SO WHERE DID THIS DICHOTOMY OF ENERGY AND ANATOMY COME FROM? IN A SYMBIOSIS TWO BILLION YEARS AGO BETWEEN A BACTERIA AND AN OXIDATIVE BACTERIA, THE ALPHA PROTEAL MATERIAL THAT CAME TOGETHER TO FORM A RELATIONSHIP, THAT ULTIMATELY BECAME TIGHT ENOUGH THAT THEY MERGED TOGETHER. THE GENOME WHICH WAS THE SAME SIZE AS THE ORIGINAL BACTERIA BEGAN TO ACCUMULATE GENES BECAUSE BACTERIA EXCHANGE GENES. IT'S WHAT THEY DO. THERE WAS A GENE FLOW FROM THE OXIDATIVE BACTERIA TO THE METHIANOGEJ NUCLEUS. WHY WOULD THERE BE A GENE CELL? TURNS OUT THAT IT TAKES ABOUT ONE BACTERIA SIZE CELL'S ENERGY TO REPLICATE ITS DNA, TRANSCRIBE INTO RNA AND TRANSLATE INTO PROTEIN. THE REASON BACTERIA DON'T HAVE BIGGER GENOMES IS THEY CAN'T MAKE ENOUGH ENERGY. ENERGY IS LIMITING THROUGH THE BIOENERGETIC PROCESS. THROUGH THE BIOSYNTHETIC PROCESS. IF ALL THE BACTERIA WOULD USE UP ALL THE ENERGY, THERE WOULD BE NO ADVANCES. BUT IF IN FACT A GENE FROM THE
THE BACTERIUM WAS TRANSFERRED TO THE NUCLEUS, INSTEAD OF HAVING TO HAVE A THOUSAND COPIES, ONE IN EVERY BACTERIUM, YOU COULD HAVE ONLY TWO. NOW THOSE TWO IN THE NUCLEUS COULD BE REPLICATED, TRANSCRIBED, TRANSLATED WITH A THOUSAND-FOLD SAVINGS OF ENERGY. BY THIS TRANSFER THEN OF GENES FROM THE MITOCHONDRIA TO THE NUCLEUS, THERE WAS A TREMENDOUS ENERGY SAVINGS. AND THAT ENERGY SAVINGS THEN CREATED ENOUGH RAW ENERGY TO ALLOW THE NUCLEUS TO ACCUMULATE 25,000 GENES AND THUS ALLOW MULTI-CELLURITY TO OCCUR FOR ARMS, LEGS, QUESTIONS. WE HAVE TWO ORGANISMS THAT HAVE UNDERGONE A SPECIALIZATION, SPECIALIZED IN ANATOMY AND IN ENERGY. HOW DOES THIS ENERGY FLOW THROUGH OUR CELLS? FOR US THAT LIVE ON THE SURFACE OF THE EARTH, HIGH ENERGY PHOTONS IMPINGE AND TAKES THE ENERGY FROM THE SUNLIGHT AND SPLITS WATER INTO HYDROGEN AND OXYGEN, RELEASES OXYGEN IN THE ATMOSPHERE AND THEN THE PLANT THEN USES THAT HYDROGEN CONDENSED WITH CARDON T CARBON
TO GIVE YOU SUGAR. WE GET THE ENERGY OF SUNLIGHT IN THE STARCH IN THE PLANTS, WHY WE CLEARED THE MIDWEST. WE EAT THE STARCH, IT GOES INTO OUR CELLS, SPLIT INTO TWO, THE MITOCHONDRIA STRIPS THE HYDROGEN TO GIVE YOU THE WATER BACK AND RELEASE ENERGY IN CHEMICAL FORM AS ATP, KINETIC FORM AS HEAT. THE IMPORTANT POINT, ALL ENERGY FLOW IS NOT THROUGH THE NUCLEAR CYTOCOLIC ORGANISM BUT BACTERIA. UNTIL WE UNDERSTAND THE FLOW, WE CAN'T UNDERSTAND THE PATHOPHYSIOLOGY OF COMPLEX DISEASE. SO THIS IS MITOCHONDRIAL BIOCHEMISTRY LESSON FROM HELL. I NEVER HAD A MEDICAL STUDENT THAT LIKED IT BUT IN FACT I LOVE IT AND THEREFORE YOU'LL BE TORTURED. WE HAVE AN OUTER MEMBRANE, INNER MEMBRANE SPACE, MATRIX, THIS IS THE MITOCHONDRIA, GLUCOSE HAS GONE THROUGH GLYCOLYSIS, THE CYCLE STRIPS THE HYDROGEN OFF THE CARBON AND PUTS THEM INTO THIS CARRIER, NAD, TO GIVE YOU THE REDUCED FORM, NADH. WE'RE GOING TO BURN THOSE WITH OXYGEN YOU'RE BREATHING, ELECTRONS FLOW DOWN THE WIRE, REDUCING TO A MOLECULE OF WATER. THE PRISO PROBLEM WITH ELECTRON
FLOW, IT'S DIFFICULT TO STORE THE ENERGY BECAUSE IT'S KINETIC. HOW DID MOTHER NATURE SOLVE THIS PROBLEM? SHE SOLVED THAT BY CREATING A CAPACITOR, IT PUMPS POSITIVE CHARGES TO THE INSULATOR INTO THE MEMBRANE SPACE TO GIVE YOU POSITIVE ON THE OUTSIDE, ALKALINE AND NEGATIVE ON THE INSIDE, THE GREAT INSIGHT OF PETER MITCHELL. THIS STORED ENERGY CAN THEN BE USED FOR LOTS OF PURPOSES, ONE OF WHICH CAN BE USED TO ADP INTO ATP WHICH CAN BE EXCHANGED, GOES THROUGH THE VOLTAGE CHANNEL AND THE ATP CAN DO WORK. WE'VE COUPLED OXIDATION WITH PHOSPHORYLATION. NOW EVERYBODY IN THIS ROOM HAS A DIFFERENT EFFICIENCY AT OX-PHOS. SOME ARE EFFICIENT AND CONVERTING HYDROGEN INTO ATP, TIGHTLY COUPLED. EVERY CALORIE YOU EAT IS ONE UNIT OF HEAT, THE MOST EFFICIENT PEOPLE EAT THE LESS LEAF CALORIES FOR MAXIMUM AMOUNT OF WORK AND GENERATE LEAST AMOUNT OF HEAT. OTHER PEOPLE ARE LESS EFFICIENT, AND THESE PEOPLE ARE LOOSELY COUPLED, AND THEREFORE TO MAKE THE SAME AMOUNT OF ATP THEY HAVE TO EAT MORE CALORIES. BY BURNING MORE CALORIES THEY ARE MAKING PROPORTIONALLY MORE HEAT PER UNIT WORK. THIS COUPLING EFFICIENCY AS I'M GOING TO ARGUE IS A VERY, VERY IMPORTANT COMPONENT OF THE VARIATION OF PEOPLE IN THE ROOM AND IN THE WORLD. NOW, THIS SYSTEM IS LIKE ANY FURNACE, IT HAS INCOMPLETE OXIDATION, COMBUSTION, THAT'S PERSONIFIED BY THE ELECTRONS AND COMPLEX ONE AND THREE, BEING ABLE TO DONATE DIRECTLY TO O 2 TO GIVE YOU AN UNPAIRED ELECTRON WHICH WANTS TO OXIDIZE LIPIDS, PROTEINS AND DNA, TWO WILL BE DISMUTATED INTO HYDROGEN PEROXIDE, YOU CAN GET ANOTHER ELECTRON TO GIVE YOU A RADICAL WHICH AN UNPAIRED ELECTRON, OXIDIZING YOUR MITOCHONDRIA AND YOUR CELLS, THE OXYGEN RADICALS. BUT YOU'LL ALSO SEE THEY ARE CRITICAL IN SIGNALING. THE MITOCHONDRIA BEING A CAPACITOR PICKS UP POSITIVE CHARGE REGULATING CALCIUM AND PERMEABILITY TRANSITION PORE, SELF DESTRUCT SYSTEM, NORMALLY A CLOSED DOOR, MAINTAINING THE MEMBRANE PR POTENTIAL BUT WHEN
IT DECLINES, CALCIUM OVERLOAD BECOMES HIGH, OXIDATIVE STRESS IS HIGH, IT GOES FROM CLOSED DOOR TO OPEN CHANNEL, SHORTS THE CIRCUIT, MEMBRANE POTENTIAL, THE STRENGTH OF THE MATRIX CAUSES FLUIDS TO FLOW IN, INNER MEMBRANE SWELLS, BAX AND BAT FORM A NEGATIVE A CHANNEL, RESTORING PROTEIN, DEGRADING THE NUCLEUS AND CYTOSOL WHEN MITOCHONDRIAL ENERGETICS IS IMPAIRED. THE MITOCHONDRIA GENERATION
URGES ENERGY, REGULATES REDOX, AT HIGH LEVELS IS DAMAGING, REGULATES CALCIUM, CELL DEATH, REGULATES THINGS LIKE ATP. BASICALLY THEN WHAT WE CAN THINK OF THE CELL AS TWO LIFE FORMS, NUCLEAR CNTOSOLIC LIFE FORMS, GENES IN THE NUCLEUS, AND ABOUT ONE TO TWO THOUSAND OF THE NUCLEAR ENCODED PROTEINS WERE ORIGINALLY FROM THE MITOCHONDRIAL DNA AND ARE NOW MADE ON THE CYTOSOL AND TRANSPORTED TO MAKE THE ANATOMY OF THE MITOCHONDRIA. THESE ARE TRANSLATED ON MITOCHONDRIAL-SPECIFIC RIBOSOMES, SENSITIVE, AND THESE 13 PROTEINS ARE SEVEN OF THE 45 PROTEINS OF COMPLEX 1, ONE OF THE 11 PROTEINS OF COMPLEX 3, 3 OF THE 13 PROTEINS OF 4, TWO OF THE 17 OF COMPLEX FIVE. YOU MIGHT ASK, IF YOU PUT 2000 GENES IN THE NUCLEUS WHY KEEP 13? THE ANSWER, I BELIEVE, COMES FROM THE FACT THAT EVERY ONE OF THE PROTEINS IS COMPLEX 1, 3, 4 AND 5. AND WHAT DO 1, 3, 4 AND 5 HAVE IN COMMON? THE MEMBRANE POTENTIAL. SO THESE PROTEINS THEN ARE THE WIRING DIAGRAM OF THE POWER PLANT. IF ANY ONE OF THESE COMPLEXES BECAME LEAKY FOR PROTON, IT WOULD SHORT THE CAPACITOR, ONCE THAT COLLAPSES YOU LOSE YOUR POTENTIAL FOR ENERGY, STOP BREATHING, BECOME INERT, WE CALL THAT DEAD. THIS THEN IS CRITICAL THIS SYSTEM BE MAINTAINED, AS ALL OF THESE ENZYMES HAVE TO CO-EVOLVE AND BE BALANCED TOGETHER. SO HOW DOES THAT OCCUR? WELL, THE PROBLEM THEN IS YOU CAN'T HAVE RECOMBINATION, BECAUSE IF WE MIXED AND MATCH MITOCHONDRIAL DNA FROM ANY TWO OF YOU, YOU HAVE SLIGHTLY DIFFERENT CIRCUIT DIAGRAMS, WE GET A MISMATCHED CIRCUIT THAT WOULD SHORT. SO IN FACT WE CANNOT ALLOW RECOMBINATION. THAT'S WHAT THE NUCLEUS DOES. SO MOTHER NATURE HAD TO SOLVE THIS PROBLEM. BEING A WOMAN, SHE SOLVED THE IN A LOGICAL SAY WEIGHING THE MITOCHONDRIAL DNA WOULD BE INHERITED ONLY FROM THE MOTHER, SO IT'S INHERITED TO HER CHILDREN, DAUGHTERS TO THEIR CHILDREN, A MALE'S MITOCHONDRIAL ARE SELECTIVELY THROWN OUT AND DESTROYED. MEN HAVE BEEN THROWN OUT FOR TWO BILLION YEARS, DON'T FEEL BAD. [ LAUGHTER ] EACH CELL THEN HAS THOUSANDS OF THESE MITOCHONDRIAL DNA'S AND THEY ARE CONSTANTLY REPLICATING INSIDE YOUR CELL. SO THEY ARE REPRESENT INDICATING, INCREASING IN NUMBER. THEY ARE BEATING EATEN UP, CREATING MUTEAND AND NORMAL BACTERIA. RED IS MUTANT, BLUE IS NORMAL. IF THE CELL DIVIDED DOWN THIS WAY, BOTH CELLS WOULD HAVE MUTE QUANTITY ANMUTANT AND NORMAL. POINT IS MITOCHONDRIAL DNA GENOTYPES SEGREGATE DURING MITOSIS. A HETEROPLASTIC SKY GOA ZYGOTE
WILL GIVE RISE TO TWINS WITH DIFFERENT PHENOTYPES, OR IF HETEROPASS PLASTIC, SOME WOULD BE PRIMARILY MUTANT, AND THE MORE THERE WERE, THE LESS ENERGY, THE EQUIVALENT OF METROPOLITAN BROWNOUT. ORGAN SYSTEMS WOULD MALFUNCTION, THAT TURNS OUT TO BE THE BRAIN. WE HAVE A HIGH MUTATION RATE. WE'LL TALK ABOUT WHY THAT IS BUT THE MUTAGEN IS PROBABLY ROS. DID DID TRANSFER RNA'S PUNCTUATE GENES, AND COMPLEX ONE, COMPLEX FOUR, SIX AND EIGHT FOR COMPLEX FIVE WITH A CONTROL REGION THAT REGULATION TRANSCRIPTION AND REPLICATION. SO IT HAS A VERY HIGH MUTATION RATE. ONE OF THE THINGS THAT WE GET IS MA TERNALLY INHERITED DISEASE. THIS MUTATION, A NUCLEOTIDE POSITION, IF YOU INHERIT THAT FROM YOUR MOTHER YOU'RE FINE UNTIL MID-LIFE AND YOU'LL GO DEAF. THIS GIVES YOU DIABETES, A HIGH PERCENTAGE GIVES YOU STROKE, EVEN HIGHER LEVEL WILL KILL YOU AS AN INFANT. MUTATION IN THE TRNA GENE AT 8344 GIVES YOU A KIND OF EPILEPSY, THERE ARE HUNDREDS OF THESE PROTEIN SYNTHESIS MATERNALLY INHERITED GENES. WITH ONE YOU'LL BE FINE UNTIL MID-LIFE AND LOSE VISION IN ONE EYE. UP HERE, THE SAME MUTATION, 14484 GIVES YOU THE SAME PHENOTYPE, 14459, WHEN THE MUTATION AT 8993, 70% GIVES YOU RETINAIZED PIGMENTATION, SOME OF THE SUBUNIT 1 GENES IN PROSTATE CANCER. WE HEARD SEVERAL NICE LECTURES IN THAT AREA SO I DROPPED THEM FOR TIME'S SAKE. THE MITOCHONDRIAL GENOME INVOLVES THOUSANDS OF ON ONE TO
TWO THOUSAND NUCLEAR GENES, 35 MITOCHONDRIAL GENES, MA TERNALLY INHERITED DISEASES. EVERYBODY IN THE ROOM HAS A DIFFERENT SEQUENCE. SOME ARE COMMON, THIS ONE IS FOUND IN 3/4 OF SUBSAHARAN AFRICANS, THIS AROSE IN ASIA, CROSSING THE BEHRING LAND BRIDGE, AND THESE I'M GOING TO ARGUE CHANGED THE MITOCHONDRIAL PHYSIOLOGY TO ALLOW PEOPLE TO ADAPT TO DIFFERENT ENVIRONMENTS. OKAY. SO ONCE WE BEGIN TO THINK ENERGETICALLY, GETTING CLOSE TO THE INTRODUCTION, BE PATIENT, ONCE WE BEGIN TO THINK ENERGETICALLY, THEN WE CAN REALIZE WE CAN THINK ABOUT ANATOMY FROM AN ENERGETIC POINT OF VIEW. HIGH ENERGY TISSUES LIKE THE BRAIN HAS THE HIGHEST ENERGY DEMAND, LOWEST RESEARCH, 2% OF THE BODY WEIGHT, USESE USES 20%
OF OXYGEN. A 10% REDUCTION IN OXYGEN WILL GIVE YOU A BAD HEADACHE. WE HAVE RESULTS ON HEARTS, RENAL, ENDOCRINE, THEY CAN BE MORE TOLERANT. WE HAVE ENERGY STORAGE TISSUES, ADIPOSE STORES FAT. WE HAVE AN ENERGY HOMEOSTASIS TISSUE, THE LIVER, WHY DOES IT REGULATE BLOOD GLUCOSE? THAT'S YOUR CONNECTION WITH SUNLIGHT. AN ENERGY SENSITIVE TISSUE, PANCREATIC BETA CELL TELLS THE SWELLS TO SWITCH TO TOR GLYCOLYSIS, OR TURN ON CYCLIC AMP TO BURN FAT. ALL OF THIS LEADS TO THIS CONCEPT THAT IF WE NOW MOVE ANATOMY OUT OF THE CENTER OF OUR THINKING AND PUT IN ENERGETICS, ALL OF THE COMMON COMPLEX DISEASES COULD BE UNDERSTOOD FROM THE SAME PATHO PHYSIOLOGICAL MESSAGE. THIS IS THE MAJORITY OF THE ENERGY AND THE MOST SENSITIVE. SO WE COULD HAVE VARIATION IN NUCLEAR GENES, MUTATIONS OR POLY MORPHISMS, CHANGES IN THE EXPRESSION THAT WOULD HAVE CHANGED ENERGETICS. THIS IS HOW YOU PROCESS ALL YOUR CALORIES, AND THE OXYGEN THAT YOU'RE BREATHING. IT'S WHERE THE SOURCE OF ENERGY TO GROW, MATURE AND REPRODUCE ARE. IT'S ALSO THE MOST SENSITIVE TO TOXINS, SO IF YOU DON'T LIKE YOUR NEIGHBOR, YOU WANT HER TO STOP BOTHERING YOU, PUT A LITTLE CYANIDE IN YOUR TEA. SHE'LL STOP BOTHERING YOU. ALMOST ALL THE MAJOR TOXINS ARE DIRECT INHIBITORS EVER MITOCHONDRIA. WHY? THAT'S YOUR ACHILLES HEEL. IF YOU INHIBIT FUNCTION, YOU WILL ACCUMULATE SOMATIC MUTATION BECAUSE REPLICATION IS NOT GOING WELL GIVE YOU A DECLINE IN FUNCTION. AND THAT GIVES YOU THE DELAYED ONSET OF PROGRESSIVE COURSE WE SEE IN ALL COMPLEX DISEASES AND AGING AND WE BELIEVE THAT'S THE AGING CLOCK. WHAT WOULD BE THE INTERMEDIATATES, PARTIAL DEFECTS? THEY ARE GOING TO AFFECT THE BRAIN, HEART, MUSCLE AND RENAL, WITH THE COMPLEX DISEASES. IT WILLEIT WILL ALSO MESH
PERTURB THE FLOW
OF ENERGY IN YOUR SYSTEM. EVERY TIME A CELL BREAKS OPEN, IF THE MITOCHONDRIA ARE NOT DIGESTED, YOU'RE GOING TO RELEASE THOSE POLY PEPTIDES AND OTHER THINGS AND YOU'RE GOING TO AMASS A MASSIVE INFLAMMATION RESPONSE. THIS IS A VERY GROWING AREA RIGHT NOW, THE SO-CALLED DAMPS, THE INFLAMMATION SYSTEM SECONDARY TO MOST OF THE DEGENERATIVE DISEASES. FINALLY, HOW YOU MANAGE ENERGY IS CRITICAL TO WHETHER CANCER GROWS, IT'S NOT THAT ONE SYSTEM IS DEFECT ACTIVE, CANCER CELL HAS THE ABILITY TO SWITCH. THAT'S THE BACKGROUND. NOW WE'RE GOING TO HAVE EXAMPLES. THIS IS JUST THE FAMILY WHERE THIS PERSON WENT BLIND, RELATED TO THIS FEMALE, TO THIS PERSON WENT BLIND, THESE PEOPLE, TO THIS FEMALE, THESE MEN WENT BLIND AND THIS WOMAN, AND THEN TO THIS FEMALE, THIS BLIND PERSON AND IT'S A LATE ONSET DISEASE, YOUNG PEOPLE DIDN'T SHOW IT. THIS IS NEUROPATHY, THIS IS A MUTATION. SOME OF THE THINGS YOU CAN IMMEDIATELY SEE ABOUT THIS, NOT EVERYBODY ON THE MATERNAL LINE IS BLIND. PEOPLE THOUGHT THIS WAS AN EXCELLENEXCELLENT GENE
ORIGINALLY, THERE'S A 4 TO 1 PREFERENCE IN MALES. WHY WOULD YOU HAVE THAT WITH MALE BIAS? MALE BIAS IS IN FACT A THEME. ONE OF THE INTERESTING THINGS IS THAT YOU ALSO SEE IT IN AUTISM, IN PARKINSON'S DISEASE, MANY OF THESE COMPLEX DISEASES SHOW THIS MALE BIAS WHICH WE THINK IS A SEX LIMITED FEATURE OF ENERGETICS. STILL, THAT ASKS THE QUESTION, WHY IS IT THERE'S VARIABILITY IN A MITOCHONDRIAL DNA DISEASE GENE? ONE OF THE MAJOR FACTORS IS THE BACKGROUND IN WHICH THAT MUTATION OCCURS. THE MITOCHONDRIAL DISEASE, EVERYBODY HERE HAS A DIFFERENT MITOCHONDRIAL DNA. A MUTATION WOULD HAVE DIFFERENT EFFECTINGS OEFFECTS ON YOU. HERE ARE THREE, THERE ARE NOW DOZENS. THIS ONE CAUSES A SEVERE COMPLEX ONE DEFECT, 1178, A MODERATE DEFECT, AND A MILD, YET THEY ALL GIVE THE SAME PHENOTYPE. WHY WOULD THAT BE? THE ANSWER IS THAT THE BACKGROUND IS IMPORTANT. THIS IS GROUP J FOUND IN 20% OF PEOPLE OF EUROPEAN BACKGROUND. THIS SEVERE MUTATION DOESN'T MATTER YOUR BACKGROUND, WHETHER IT'S J OR NOT. YOU'RE GOING TO GO BLIND. THIS MORE MODERATE ONE, ONE-THIRD OF THE PEOPLE ARE ALSO J, SO J HAD TO INCREASE THE PENETRANCE, AND FOR THIS MILD ONE ALMOST 3/4 ARE J. SO IN FACT J INCREASED THE PENETRANCE OF THE MILDER MUTATIONS. HERE IS ANOTHER MUTATION. THAT ALSO WILL INCREASE THE PENETRANCE OF THE MILDER MUTATIONS. SO WHAT ARE THESE LINEAGES? THESE LINEAGES ARE IN FACT CONTINENT AND REGIONAL SPECIFIC MITOCHONDRIAL DNA LINES. SO IF I SEQUENCE THE MITOCHONDRIAL DNA OF EVERYBODY IN THE ROOM, ANY TWO OF YOU, WITH NUCLEOTIDE SUBSTITUTIONS, WOULD BE PROPORTIONAL IF YOU SHARED A COMMON MOTHER, BY USING THAT LOGIC, AND SEQUENCING THE MITOCHONDRIAL DNA OF INDIGENOUS PEOPLE AROUND THE WORLD WE FOUND IN FACT MITOCHONDRIAL DNA INITIATED IN AFRICA ABOUT 150,000 YEARS AGO, GAVE RISE TO L-1 AND L-2, PI PYGMIES,
SOUTHEAST ASIA, THE TROPICS, DOWN TO AUSTRALIA, BUT DID SOMETHING UNIQUE, MOVING NORTH TO THE TEMPERATE ZONE, AND H, J, TU, V AND M MOVED NORTH, AND 20,000 YEARS AGO THE AMERICAS WERE COLONIZED. ALL ALELES ARE FOUND THROUGHOUT THE WORLD. THIS WOMEN NEED TO BE ABLE TO RUN AWAY FROM LIONS, OTHERWISE THEY GET EATEN. HOW DO THEY RUN FROM LIONS? THEY HAVE TO BE EFFICIENT. UP HERE THE LIONS FROZE TO DEATH. HOW ARE THEY GOING TO SOLVE THAT PROBLEM? ACCUMULATING MITOCHONDRIAL DNA MUTATIONS THAT DECREASE COUPLING EFFICIENCY. NOW THEY HAVE TO EAT A HIGH FAT DIET FOR THE SAME AMOUNT OF ATP GENERATING MORE INTERNAL HEAT. THEY WILL HAVE TO KILL MARINE MAMMALS, EVERYBODY WILL LOOK DOWN ON THEM. THAT'S THE WAY IT IS. THE POINT IS THAT THESE MITOCHONDRIAL DNA LINEAGES BECAME ADAPTIVE AND THEY ALLOWED OUR ANCESTORS TO LIVE IN THESE DIFFERENT GEOGRAPHICAL REGIONS, THIS SHOWS OUT OF AFRICA, LINEAGE M WHICH STAYED IN THE TOPICS HAS SYNONYMOUS MUTATIONS, RANDOM MUTATIONS, BUT N WHICH MOVED TO THE TEMPERATE ZONE, HAS THREE. WHAT DOES THAT DO? DECREASED THE MEMBRANE POTENTIAL AND ALTERED CALCIUM METABOLISM MAKING IT LESS EFFICIENT. AN AMIN AMINO ACID, THIS
VARIANT, THIS AMINO ACID IS CONSERVED DOWN. THESE MUTATIONS IN THE B GENE, THIS ONE IS CONSERVED IN ALL MESOZOANS ALL THE WAY TO E. COLI. WAIT A MINUTE. THE AMINO ACID CHANGES CONSERVED THROUGHOUT SPECIES EVOLUTION, POLY MORPHIC IN THIS ROOM, THAT'S UPSIDE DOWN FROM EVERYBODY WE'VE BEEN TALK ABOUT MOLECULAR EVOLUTION. HOW COULD THIS BE? REMEMBER, THE MITOCHONDRIAL DNA HAS INCREDIBLY HIGH MUTATION RATE. HOW WOULD IT WORK THEN? THE IDEA THAT WE HAVE IS THAT WHEN -- SO THIS IS THE MEAN ENVIRONMENT, THE MOST COMMON ENVIRONMENT. WE HAVE THEN A MITOCHONDRIAL LINEAGE THAT HAS BECOME ADAPTED TO AN EXTREME ENVIRONMENT, TWO STANDARD DEVIATIONS, IT THEN IS GOING TO ACQUIRE A MUTATION, SOME ALREADY FUNCTIONAL, CREATING LINEAGES THAT MOVE BACK TOWARD THE MODAL ENVIRONMENT, ULTIMATELY FIXING THESE FUNCTIONAL MUTATIONS WITH RANDOM VARIATION. THAT THEN WILL RADIATE AND THEN ACQUIRE THE FUNCTIONAL MUTATIONINGMUTATIONSAND CREATE A
MODE ENOU
GH TO CREATE A NEW SPECIES, YOU GO BACK THROUGH THE SYSTEM AGAIN. WHEN YOU TAKE THE AVERAGE OF THE SPECIES, THE ENVIRONMENT HAS SELECTED FOR CONVERGENT EVOLUTION ON THE SAME AMINO ACID CHANGES. AT EXTREME ENDS, WE HAVE THEN THE FUNCTIONAL VARIATIONS THAT WE SEE IN THE ARCTIC OR EXTREME AFRICA. I THINK THEN THIS IS THE DRIVING FORCE FOR ADAPTATION WITHIN SPECIES WHEREAS NUCLEAR VARIATION IS THE DRIVING FORCE FOR ADAPTATION BETWEEN SPECIES. THIS IS STUDIES IN TIBET. WE SEQUENCED MITOCHONDRIAL DNA FROMTY YETIAN TIBETANS AT MANY
ALTITUDES. THERE'S LINEAGE M AND N. OUT OF AFRICA IS M AND N. WHAT WE FOUND IN TIBET IS THAT THIS MUTATION, T-339-4C AROSE THREE INDEPENDENT TIMES. IF WE ASK THE FREQUENCY OF THAT MUTATION VERSES ALTITUDE, WE SEE AS THE ALTITUDE INCREASES, SO THIS THIS. THERE IS AN ODDS RATIO, THIS OCCURS BY CHANCE OVER 25. SO WHAT THIS MEANS IS THIS MUTATION IS BEING SELECTED BY ALTITUDE. BUT WAIT A MINUTE. THAT'S THE SAME VARIANT WE SAID INCREASES PENETRANCE OF LABORS. HOW COULD IT BE BAD FOR LABORS, GOOD FOR ALTITUDE? OKAY. THIS IS ANOTHER VARIANT. WE'LL COME BACK TO THAT. THIS IS ANOTHER LINEAGE, LINEAGE FOR F-4, THIS IS JUST LOOKING AT OBESITY, ONE OF MANY, MANY STUDDIZE WE'VSTUDIES WE'VE BEEN
DOING. IT HAS A SIGNIFICANT EFFECT, INCREASED BMI LEVELS, WE CAN DO THAT FOR DIABETES, CARDIOVASCULAR DISEASE, ON AND ON. THAT I VARIANT OCCURRED SEVEN TO SEVEN THOUSAND YEARS AGO IN EUROPE, IT'S ONLY ABOUT .4% OF EUROPEANS, BUT IF YOU NOW LOOK AT PEOPLE WITH ALZHEIMER'S AND PARKINSON'S DISEASE, 3% HAVE ALZHEIMER'S, 5% -- 3% OF ALZHEIMER'S HAVE THE MUTATION. IT PREDISPOSES TO LATE ONSET NEURODEGENERATIVE DISEASE. A MUTATION SPONTANEOUS, 3397, THIS IS A CODE, BUT THAT'S THE ADJACENT CODON TO THE MUTATION, THIS ONE GIVES YOU ADPD. HOW DO WE MAKE SENSE OF THIS? WHAT WE REALIZE SENTENCE PHENOTYPE DEPENDS ON CONTEXT. OUT OF AFRICA, WE HAVE L, AND THAT GIVES RISE TO N WITH THESE TWO FOUNDING MUTATIONS, AND THE TRNA GLUTAMIN IS PREDISPOSED, WE CAN GET THIS, THAT'S TOXIC AND GIVES YOU ADPD. HOWEVER, YOU CAN GET THE 3394 MUTATION, THAT IS AT A HIGHER FREQUENCY INCREASING YOUR RISK OF LABORS BUT ALSO OBESITY AND DIABETES. BUT OVER HERE, WHEN WE SKIPPED THESE MUTATIONS, IN THAT CONTEXT, HIGH ALTITUDE, THIS PARTICULAR MUTATION IS NOW COMMON AND IT'S ADAPTIVE TO ALTITUDE BECAUSE THIS IS ONLY BAD WHEN IT'S IN THE CONTEXT OF THESE. SO HOW CAN WE SHOW YOU THAT'S FUNCTIONALLY IMPORTANT? WE TAKE BLOOD PLATELETS, WE HAVE A CELL LINE THAT LACKS ITS OWN MITOCHONDRIA, WE SELECT FOR THESE SOMATIC SELF HYBRIDS, AND WE ASSAYS. F IS THE DIABETES, YOU CAN SEE WHEN WE HAVE THIS C MUTATION, WE GET A 15 TO 30% REDUCTION IN COMPLEX 1, SO IT'S FUNCTIONALLY IMPORTANT. LOOK AT THE RELATIONSHIP BETWEEN THE T ALELE FOR F AND B. B AND F NOT ONLY ARE RELAVENT TO EACH OTHER, RELAVENT TO THE NEW MUTATIONS. FINALLY IF WE LOOK AT THE LINEAGE, THE C IS AS GOOD AS THE T MUTATION ON B. BACKGROUND IS EVERYTHING. CON TEXT IS WHAT MATTERS WITH MITOCHONDRIAL ENERGETICS. WE CAN THEN USE THESE LINEAGES TO DO A LOT OF ASSOCIATION, AND THEY HAVE CORRELATED WITH RISK FOR ALZHEIMER'S, AIDS, OSTEOPOROSIS, AGING, CANCERS, ATHLETIC PERFORMANCE. OKAY. ALL RIGHT. SO HOW CAN I PROVE TO YOU IN FACT THESE ARE RELEVANT TO DISEASE? WHAT WE DID IS WE CREATED A MOUSE WHICH HAS A SPECIFIC MITOCHONDRIAL DNA POINT MUTATION, WE MADE THIS ORIGINALLY TO LOOK AT WHETHER WE COULD SHOW THAT THESE MUTATIONS IN FACT CAUSED THE DISEASE. IN ONE LINE, WITH A FRAME SHIFT, IN THIS THIS, WE TAKE THE FOR EXAMPLEMENTFRAGMENTAND TAKE
THIS, PUT IT I
NTO A FOSTER MOTHER, GOT CHIMERIC FEMALE AND BREAD THROUGH THE FEMALE LINE, THUS PICKING UP THE MITOCHONDRIAL DNA. OKAY. SO THIS SHOWS IN FACT THE MUTATION WHICH WAS INSERTION OF EXTRA C, THROWING IT OUT OF FRAME. THIS EMBRYONIC STEM CELL DELETED THE ADJACENT T AN AND REVERTED
BACK TO WILDTYPE AND CARRY THE MUTATION IN THE CO-1. AFTER MANY TRIES WE GOT THIS PARTICULAR EMBRYONIC STEM CELL LINE AND THE FEMALE WITH THIS LINE HAD IN DIFFERENT ORGANS DIFFERENT PERCENTAGES OF THE MUTANT, REPLICATED SEGREGATION. WE CROSSED THIS FEMALE WITH A NUCLEAR MATCH CONTROL, SHE STARTED WITH 47%, HER FIRST AND SECOND LITTERS WERE 16%, THE THIRD LITTER WAS 6%, FOURTH, FIFTH AND SIXTH WERE 0%, THERE WAS A SELECTIVE LOSS OF THE FRAME SHIFT. WE TOOK THE 16% AND MATED THEM, SIX OR ZERO. WE SUPE SUPER OVULATED THEM. THIS IS A TRUNCATED DISTRIBUTION, IN THE OVARY THERE'S A SYSTEM THAT SELECTS AGAINST DILATARIOUS MUTATIONS. WHY WOULD THAT BE IMPORTANT? IF YOU HAVE AN EXQUISITELY HIGH MUTATION RATE, FOR GENES ABSOLUTELY CRITICAL FOR LIFE, YOU LET ALL THOSE MUTATIONS INTO THE POPULATION, THE POPULATION WOULD GO EXTINCT. SINCE MITOCHONDRIA CAN FUNCTION IN A SINGLE CELL, YOU CAN NOW HAVE SELECTIVE PRESSURE ON THE CELL BEFORE YOU EVER GET A ZYGOTE, ALLOWING US TO HAVE A VERY HIGH MUTATION RATE WITHOUT GENETIC LOAD. LET'S SAY WE SEGREGATE THE MUTATION AND THIS CREATES A REDUCTION, GIVES YOU DEGENERATING MUSCLE FIBERS, CARDIOMYOPATHY AND MUSCLE SYMPTOMS. LET'S TALK ABOUT ANOTHER DISEASE, A FAMILY REPORTED, THIS WOMAN HAD 50% OF A MUTATION THAT CHANGED THIS GENE, AND SHE HAD CEREBELLAR. ALL THEIR CHILDREN DIED. WE HAVE THE INHERITANCE OF THIS HETEROPLASMIC MUTATION. AN ARTICLE JUST CAME OUT THAT ARGUES DARWIN'S FAMILY, THE REASON HE WAS SICK, HE HAD A MUTATION, IF YOU TRACE HIS PEDIGREE, EVERYBODY ON THE MATERNAL PEDIGREE IS AFFECTED. THAT'S AN ASIDE. WHAT WE HAVE IS THIS RANDOM SEGREGATION OF HETEROPLASMY. HOW CAN WE SEE IF THIS IS THE CAUSAL MUTATION? WE SCREEN FOR MANY YEARS TO FIND A CELL LINE WITH EXACTLY THAT MUTATION. HERE IT IS. WE TAKE OUR FEMALE EMBRYONIC STEM CELL, PUT IN THE MITOCHONDRIAL DNA, THE FOSTER MOTHER, TRANSMIT THE MUTATION. WHAT DO WE GET? THIS IS LOOKING AT THE OPTIC NERVE, THIS IS THE CONTROL OPTIC NERVE WITH THE NORMAL FIBERS AND YOU CAN SEE IN THE MUTANT SWOLLEN FIBERS. IN FACT, THE SMALL CALIBER HIGH ENERGY FIBERS ARE PREFERENTIALLY AFFECTED. YOU CAN SEE THAT THE RED ARE THE NORMAL MICE, THE BLUE ARE THE MUTANT, AND YOU CAN SEE THERE'S A BIAS TOWARDS SMALL CALIBER FIBERS IN THE WILDTYPE VERSUS BLUE AND THAT GETS WORSE AS THEY AGE. IF WE THEN LOOK AT THESE MORE GENERALLY WE SEE DEMYELENNATION, WHAT YOU WOULD SEE IN MULTIPLE SCLEROSIS. IN FAMILIES, MALES GO BLIND, FEMALES GET MULTIPLE SCLEROSIS. THIS IS JUST LOOKING AT THIS CELL LINE. WE DECLINE IN REACT ACTIVITY IN THE BASAL GANGLIA, INCREASE IN ANXIETY, DEPRESSIVE EFFECT. WE SEE SEVERE EFFECTS ON MOTOR AND WE SEE A LEARNING DISABILITY. AND IF WE NOW DO AN MRI WE CAN SEE WE'RE GETTING SPECIFIC ALTERATIONS IN THE FIBER DENSITY. THIS MITOCHONDRIAL DNA POINT MUTATION NUCLEUS PERFECTLY NORMAL IS GIVING YOU CLINICAL NEUROLOGICAL PHENOTYPES OF WHAT WE WOULD CALL PARKINSON'S DISEASE. SO THAT MEANS THAT THIS MUST BE AN ENERGETIC DISEASE BECAUSE THAT'S THE ONLY CHANGE THAT HAS OCCURRED. OKAY. SO HOW CAN WE TELL MORE ABOUT THIS DISEASE? WELL, NOW THAT WE HAVE A MOUSE WE CAN ISOLATE AND STUDY THE MITOCHONDRIA IN SITU. WE FIND THIS COMPLEX 1 DEFECT REJUICEREDUCES ENERGETICS 30%
HAS NO AFFECT ON ATP LEVEL BUT AFFECTS RESPIRATION AT REST BUT IT CAN GO TO NORMAL BUT THE REACT
REACTIVE OXYGEN GOES TO HIGH LEVELS. NOW, LET'S AGE THESE CO 1 MICE TO TWO MONTHS. NOW WE SEE WE GET INSULIN RESISTANCE. HERE IS THE INSULIN LEVEL OF THE MUTANT AND WILDTYPE AND FOR THE CO-1 WE NOW HAVE A GLUCOSE INTOLERANCE. SO THIS POINT MUTATION THEN IS GENERATING ALL THE PHENOTYPES THAT WE SEE IN COMMON DISEASES. SO, OKAY, THOSE ARE PATHOGENIC MUTATIONS, BUT WHAT ABOUT NATURALLY OCCURRING VARIANTS LIKE YOU HAVE IN WE DECIDED TO TRY AND MIX TWO NATURALLY OCCURRING MITOCHONDRIAL DNA'S, WE MIXED NZB WITH 129, PUT ANYTIME THE ANIMAL, NOW WE HAVE THE FOUNDER FEMALE AND THAT'S THE NZB, 129, THIS IS HER DAUGHTER. WE MATED HER AND ALL OF THEY ARE OFFSPRING, MALES OR FEMALES, HAVE THE HETEROPLASMA AND DAUGHTERS TRANSI TRANSMITTED,
MALES DO NOT. THESE ANIMALS HAVE BEEN BACK-CROSSED 20 GENERATIONINGS ON A COMMON INBRED BACKGROUND SO THERE'S NO NUCLEAR VARIATION WITHIN WHAT WE CAN CONTROL FOR. SO NOW WHAT WE DO IS TAKE THE HETEROPLASMIC ANIMAL AND ASK THE PHENOTYPE. IF WE LOOK AT ACTIVITY, WE FIND THE 129 AND NZB ARE ACTIVE AT NIGHT AS THEY SHOULD BE, BUT THE HETEROPLASMIC ARE HYPO ACTIVE, ASSOCIATED WITH FOOD -- DECREASED FOOD INTAKE. THE RESPIRATORY CONTROL RATIO IS SLIGHTLY LOWER BUT WHEN WE PUT THEM IN AN ENVIRONMENT, THEY BECOME HYPER-EXCITABLE. CAN THEY LEARN? HERE IS BLACK IS NZB, BLUE HAS DIFFERENT BOUTS OF LEARNING, TO TAKE THIS OPEN FIELD WITH ALL THESE HOLES AROUND THE MIDDLE, AND AROUND THE OUTSIDE, AND THERE'S A BLACK BOX THERE, THEY ARE COLORED SYMBOLS TO FIND THEIR WAY, AND THEY LEARN OVER TIME HOW TO FIND THE BOX AND HIDE. SO BOTH -- ALL THREE LINES LEARN, HETEROPLASMIC LEARNS MORE SLOWLY. MEGAN LET THE ANIMALS REST FOR A DAY AND ASKED THEM IF THEY COULD REMEMBER THE TASK, 129 AND NZB IMMEDIATELY RAN IN THE HOLE. THE HETEROPLASMIC ARE COMPLETELY FORGOTTEN. HAVING TWO PERFECTLY NORMAL MITOCHONDRIAL DNA'S WE CAN GET MANY FEATURES OF LEARNING DISABILITIES AND OBSESSIVE COMPULSIVE BEHAVIOR AND DEPRESSION. OKAY. NOW LET'S LOOK AT A NUCLEAR MUTATION, THIS TRANSLOCATOR, AND ISOFORMS MAKE THIS NONLETHAL, THIS OCCURRED IN SWITZERLAND 500 YEARS AGO, WE TRACED A HUGE PEDIGREE IN NORTH AMERICA AND ULTIMATELY WE GET THESE PEOPLE HAVING HEART DISEASE. THAT MAPS THE CHROMOSOME FOUR, A FRAME SHIFT MUTATION BOODLE. THEY HAVE HEART DISEASE AND MYOPATHY. EVERY HEART BEAT IN THE WILDTYPE HUMANS IS NORMAL, BUT OF THE ANT WE SEE HIGHLY DISRHYTHMIC EFFECT. GREEN IS NORMAL, RED IS ABNORMAL. BUT THERE'S A MAJOR DIFFERENCE. SOME MUCH THE INDIVIDUALS CAN LIVE TILL THEIR 40, WITH HYPERTROPHIC CARDIOMYOPATHY. SOME OF THE INDIVIDUALS AT VERY EARLY CHILDHOOD DEVELOP A MASSIVE DILATED CARDIOMYOPATHY REQUIRING A HEART TRANSPLANNED AND HAVTRANSPLANTSAND WOULD DIE. WHY THE DIFFERENCE? IT'S ALL ABOUT THE MITOCHONDRIAL GROUP, H HAS NORMAL, AND THOSE WITH U PROGRESS TO TRANSPLANTATION. TO PROVE TO YOU THAT THAT'S TRUE, WE CREATED A MOUSE WITH THE SAME -- WITH AN ANT DEFECT, NORMAL MOUSE WILL RUN. OXYGEN USE IN RED, CO2 GIVEN OFF, USING AEROBIC EXERCISE, CAN RUN FOR LONG PERIODS OF TIME. THE KNOCKOUT MOUSE RUNS AND FALLS DOWN. IT HAS THEN AN ACCUMULATION OF MITOCHONDRIA, SUCCINATE DEHYDROGENASE, CREATING MYOPATHY. NOW, OOPS, THESE ARE THEN THE HEART ECHOES OF WILDTYPE CO 1, ND-6, ANT, ANT-C OCO 1. THIS IS HEART CAN HARDLY BEAT. THIS IS THE REPERTOIRE WITH H VERSUS U MITOCHONDRIAL DNA. SO WE CAN THEN QUANTIFY THAT BY VELOCITY VECTOR IMAGING, WILDTYPE WE GET UNIFORM HEART BEATS, AND THIS SHOWS THE SAME PHENOTYPE. OKAY. SO WE CAN THEN NOW QUANTIFY ALL THIS SO NOW WE'RE LOOKING AT DIFFERENT CARDIAC EFFECTS. IN THIS CASE, DIASTOLIC DIE AMOUNTER OF THE LEAP VENTRICLE
THEY ARE UNIFORM. THIS IS BAD. THIS IS EJECTION FRACTION, AND IF WE NOW LOOK AT THE VELOCITY VECTOR IMAGES YOU CAN SEE WE IN FACT HAVE A DIRECT RELATIONSHIP BETWEEN THE MITOCHONDRIAL DNA AND NUCLEUS. AND IF WE LOOK AT LIFESPAN, THESE GUY QUICKLY RELATIVE TO WILDTYPE, THE OTHERS INTERMEDIATATE, OTHERS ARE NORMAL. C, YOU CAN SEE A PROGRESS OF ABNORMAL MITOCHONDRIA. SO WHAT THAT SHOWS THEN IS THAT MITOCHONDRIAL VARIATION NATURALLY OCCURRING HAS ALSO HAD A BIG EFFECT ON THE PENETRANCE OF EVEN CLASSIC MENDELIAN NUCLEAR GENES. LAST POINT TO BE MADE, THIS IS THE FAMILY WITH THE 3243 MUTATION, I SAW THIS FAMILY IN THE EARLY '80s. THIS WOMAN HAD LACK PARTICULAR ACIDOSIS, THESE ARE THE CHILDREN, WE COULD EXAMINE THEM. THESE ARE NOT. THEY ALL DIED IN LATE TEENS OR EARLY 20s, MUSCLE FIBERS WERE DEGENERATING, WOLF PARKINSON WHITE CONDUCTION DEFECTS. WE TOOK THE HETEROPLASMIC CELL LINES AND THIS WITHOUT MITOCHONDRIAL DNA AND FUSED THAT TO THIS CELL CREATING DIFFERENT PERCENTAGES. THIS FAMILY WAS 70% MUTANT HAD CARDIOMYOPATHY, 10 TO 30% TYPE ONE, TYPE TWO DIABETES OR AUTISM, THIS IS BIZARRE, THIS IS SUPPOSED TO BE AN AUTO-IMMUNE DISEASE, THIS IS METABOLIC BUT THE SAME MUTATION CAUSES BOTH DISEASES, YET AT HIGH LEVELS WILL KILL YOU AS AN INFANT. WE COULD ASK WHAT'S DIFFERENT ABOUT THESE DIFFERENT GENOTYPES? WE CAN SEE AS THE PERCENTAGE OF MUTANT MITOCHONDRIAL DNA AS THE PERCENTAGE OF MUTANT INCREASES THE PROTEIN SYNTHESIS INCORPORATION IN THIS MITOCHONDRIAL DNA CODED SUBUNIT DECLINES, THERE'S A THRESHOLD EFFECT, OXYGEN CONSUMPTION STAYS CONSTANT UNTIL 60% AND FALLS OFF. WE SEE ALTERATIONS IN MITOCHONDRIAL MORPHOLOGY AND FOR THE -- FROM ZERO, 20, 30% A REMARKABLE DECLINE IN CELL SIZE. WE'RE SEEING SIMPLE CHANGES IN MITOCHONDRIAL GENOTYPE, BY ONLY 10%, HAS A BIG EFFECT ON CELL PHYSIOLOGY. TO CUT A LONG STORY SHORT WE'VE HAVE DONE RNA SEQ, IN FACT AUTISM, THAT'S ONE TRANSCRIPTIONAL PROFILE FROM THE NUCLEUS. 50 TO 90%, THAT'S ANOTHER TRANSCRIPTIONAL PROFILE, THE THIRD KILLS THE INFANTS. THIS IS THE NORMAL GENE EXPRESSION PROFILE. THIS IS THE 20 TO 30% HERE. COMPLETELY OPPOSITE IS THE 50 TO 90%, AND THEN 100%. SO WHAT THIS IS SAYING IS SUBTLE CHANGES IN MITOCHONDRIAL BIOENERGETICS IS CAUSING PHASE SHIFTS IN THE EPI GENOME EXPRESSION, AND THIS EXPRESSION WE BELIEVE CREATES THESE PHENOTYPIC CHANGES RELATED TO CANCER, METABOLIC SYNDROME, DIABETES AND SO ON, IT'S THE NUCLEAR CYTO PLASMIC CROSS TALK THAT'S IMPORTANT. THE NUCLEAR ORGANISM, THE ANATOMY OF ENERGYICS, THIS CAN TOLERATE A HIGH MUTATION RATE, THIS IS LOW BECAUSE ALL THE MUTATIONS HAVE TO GO THROUGH DEVELOPMENT BEFORE THEY CAN BE ACTED ON BY SELECTION, AND THEREFORE A HIGH MUTATION RATE WOULD BE DILATARIOUS. THE ENERGETICS. MITOCHONDRIA MAKE THE ATP AND ACETYL, WHY WOULD THAT BE? THE NUCLEUS CAN'T DO ANYTHING WITHOUT ENOUGH ENERGY AND HAS TO KNOW THE ENERGY FLUX THROUGH THE NTSB. MITOCHONDRIA. IT USED HIGH ENERGY INTERMEDIATATES TO CHANGE THE EXPRESSION TO BE REPLICATED, TRANSCRIBED AND TRANSLATED WHEN THERE WAS ENOUGH ENERGY. NOW THAT SAYS IF THIS CONCEPT IS CORRECT, THEN THESE ARE THE MAJOR INTEREST SPECIFIC VARIABLES RELATED TO PHENOTYPE, PRIMARILY INTRASPECIFIC. I'D LIKE TO END BY THANKING PLANE YEARS OF WONDERFUL COLLEAGUES AND I DON'T EVEN KNOW THAT'S THAT CLEAR, BUT PEOPLE HERE, ALICIA, DANIELLE, KIRSTEN, PIETRO, BRIAN MORROW, MARK AND KATRINA, THE STUDIES IN TIBET DONE WITH THE PEOPLE AT THIRD MILITARY MEDICAL UNIVERSITY, AND THE OPHTHALMOLOGY STUDIES, OUR EPI GENOMIC STUDIES BY PAULO CORSI, THE CARDIOLOGY STUDIES, THE FAMILY WE STUDIED IN PENNSYLVANIA, WITH KEVIN STRAUSS OF THE CLINIC FOR SPECIAL CHILDREN AND OUR LONG-TERM ANTHROPOLOGIC STUDIES. THANK YOU VERY MUCH. [APPLAUSE] >> WE HAVE TIME FOR A FEW QUESTIONS. THE MICROPHONES ARE IN THE AISLES. USE THEM SO PEOPLE WATCHING ON THE VIDEO CAN HEAR THE QUESTIONS. WE'RE HOPE FOR QUESTIONS. YOU SHOWED THE DATA WHERE THE HETEROPLASMY WAS THE PROBLEM. THE HOMOPLASMY WAS FINE. >> SO, THANK YOU, FRANCIS. AGAIN, LET ME REITERATE. THE MITOCHONDRIAL DNAS ARE PERFECTLY NORMAL MICE, JUST COLLECTED FROM DIFFERENT ENVIRONMENTS. BUT THEY STILL DIFFER BY ABOUT AS MANY NUCLEOTIDES AS YOU DO. BUT THOSE, SOME OF THOSE ARE AMINO ACID SUBSTITUTIONS. REMEMBER, ANYTHING IN A DIFFERENT ENVIRONMENT WILL HAVE A SLIGHTLY DIFFERENT AMINO ACID SUBSTITUTION. WE'VE TAKEN ONE MITOCHONDRIAL DNA WITH TWO OR THREE CHANGES AND ANOTHER MITOCHONDRIAL DNA WITH THREE OR FOUR DIFFERENT ONES, PUT THEM TOGETHER AND ASKED THEM TO MAKE THE SAME MULTI-SUBUNIT COMPLEXES. AND SO NOW YOU HAVE A PROBLEM OF A UNIFORM NUCLEUS BUT TRYING TO INSERT DIFFERENT POLY PEPTIDES INTO THE SITE. >> OVER HERE. >> SO WHEN YOU HAVE CELLS THAT HAVE SOME MITOCHONDRIA WITH MITOCHONDRIAL DNA WITH AND WITHOUT MUTATIONS HAVE YOU TRIED TO MANIPULATE IN HANDS, MITT OH MITOPHAGY, MANIPULATIONS DONE IN RECEIVVIVO, AUTOPHAGY WOULD HAVE
A BENEFIT IN ANY PATIENT? >> VERY GOOD QUESTION, MARK. I DIDN'T HAVE TIME, OBVIOUSLY, FOR OBVIOUS REASONS, TO GO INTO OUR EFFORTS TO DEVELOP DIAGNOSTICS AND THERAPEUTICS. THIS IS IN THE THERAPEUTIC AREA. I WOULD LIKE TO MENTION CARLOS' WORK, CREATING ENZYMES THAT WILL SEE SPECIFIC MITOCHONDRIAL DNA MUTATIONS, LET'S SAY PATHOGENIC, PUT THAT ENZYME IN THE NUCLEUS AND THAT PROTEIN GOES INTO THE MITOCHONDRIAL DNA, SEES IT AS FOREIGN AND DIGESTS IT. HE CAN CHANGE THE HETEROPLASMY BY THAT PROBLEM. WE HAVE A PROBLEM, HOW TO DELIVER SUCH A VECTOR TO ALL THE CELLS IN THE BODY BUT WE'RE ACTIVELY WORKING ON WAYS OF DOING THOSE THINGS. ERIC SHONE HAS PUT THEM INTO A KETO GENIC DIET BURNING FAT, HE CAN SHIFT THE HETEROPLASMA THAT WAY, A MORE CREDIBLE WAY OF TREATING PATIENTS BUT WE DON'T HAVE THE SAMPLE SIZE AT THIS POINT. THE NATIONAL INSTITUTE OF CHILD HEALTH HAS BEEN SUPPORTING A MAJOR EFFORT TO BEGIN TO GET A PATIENT REGISTRY SO THAT WE CAN DO CLINICAL TRIALS AND WE'RE GRATEFUL FOR THEIR SUPPORT ON THAT. >> OVER HERE. >> THANK YOU VERY MUCH, PROFESSOR, FOR YOUR PRESENTATION AS WELL AS YOUR CONTRIBUTIONS IN THIS FIELD. I HAVE THE SAME QUESTION AS I ASKED LAST YEAR. HOW TO GET THE DEFINITION OF MITOCHONDRIAL DISEASE, FOR EXAMPLE SOME OF THE DISEASES ARE BASED ON THE MUTATION OF MITOCHONDRIA, ENCODING PROTEINS, WE CAN SEE IT'S A DEFINITE DISEASE, BUT FOR SOME OTHER DISEASES SUCH AS MUTATION OF THE OTHER PROTEINS IN THE MITOCHONDRIA, IT CAN AFFECT THE MITOCHONDRIAL FUNCTION AS WELL AS THE MITOCHONDRIA ACTIVITY. SO HOW DO YOU THINK THOSE DISORDERS BELONG TO MITOCHONDRIA DISEASE OR ARE THEY DISEASES OF THE MITOCHONDRIAL PHENOTYPES? THANK YOU VERY MUCH. >> SAY YOUR NAME? >> [ INAUDIBLE ] >> AND THIS GENTLEMAN IS WORKING WITH WILL BORE AT NIA, DOING A VERY IMPORTANT STUDY, TRYING TO USE INFORMATION SCIENCES TO GO THROUGH THE LITERATURE AND DEFINE THE PHENOTYPES OF KNOWN MITOCHONDRIAL DISEASE TO TRY AND DEVELOP ALGORITHM FOR BETTER DIAGNOSIS. THE -- SO THIS IS AN AREA OF DISCUSSION IN THE FIELD, THERE ARE SOME PEOPLE THAT WOULD LIKE TO SPLIT THE FIELD OF MITOCHONDRIAL MEDICINE INTO WHAT THEY CALL PRIMARY DISEASE AND SECONDARY DISEASE. I ACTUALLY FEEL THIS IS REALLY MORE OF A CONTINUUM, AND THAT THERE ARE DISEASES THAT ARE DUE TO MITOCHONDRIAL DNA MUTATIONS, DISEASES DUE TO NUCLEAR MUTATIONS AND MITOCHONDRIAL GENES, ALSO THE INTERACTIONS OF THE GENES AND OF ANATOMICAL NUCLEAR GENES AS WELL. SO AT THIS PARTICULAR POINT, WE HAVE A BIG PROBLEM, BECAUSE WHAT WE SEE IS DIFFERENT MITOCHONDRIAL DNA MUTATIONS, OR DIFFERENT MUTATIONS IN NUCLEAR GENES, THEY CAN GIVE QUITE RADICALLY DIFFERENT PHENOTYPES. THAT IS ONE OF THE MOST QUESTIONS, WHY WE SPEND SO MANY YEARS TRYING TO MAKE MOUSE MODELS TO TRY AND LOOK AT ONE MUTATION TO FIND THE PHENOTYPE. BUT WHY WOULD THERE BE SUCH PHENOTYPIC VARIATION, WHY IS THE BRAIN AFFECTED IN SOME INDIVIDUALS IN THE HEART, SOME EYES, SOME KIDNEY. I THINK THE MITOCHONDRIA IS PIVOTAL TO SO MANY PARTS OF THE BODY. YOU CAN HAVE ENERGETICS, YOU COULD HAVE CHANGES IN THE MA TAB LIGHTSMETABOLITES, WE WANT
EVERYTHING TO BE LINEAR BUT I THINK THIS IS SUCH A PIVOTAL SYSTEM IT AFFECTS EVERY ORGAN SYSTEM, EVERY INSTITUTE HERE AT NIH, AND WILL AFFECT THEM IN DIFFERENT WAYS, WE CAN ONLY UNDERSTAND THAT BY TRULY UNDERSTANDING THE PHYSIOLOGY OF THIS. THAT'S WHERE WE'RE AT RIGHT NOW. >> GREAT. WE'RE OVER TIME BUT WE'LL TAKE TWO MORE QUESTIONS. HERE? >> ALL RIGHT, THANK YOU VERY MUCH. VERY INTERESTING TALK. I WAS WONDERING ABOUT THE CONNECTION BETWEEN MITOCHONDRIA AND AGING, AND SO IN MITOCHONDRIAL DISEASES YOU NEED SOMETIMES 70% EXPANSION OF THE MUTATION TO GET A PHENOTYPE, WHEREAS IN AGING, IT'S UPON POSTULATED OR PROPOSED WITH AGING YOU GET ACCUMULATION OF DNA DAMAGE THAT LEADS TO MITOCHONDRIAL DYSFUNCTION, BUT THE ACCUMULATIONS THAT YOU SEE IN AGING IS MUCH, MUCH -- ON A MUCH, MUCH LOWER FREQUENCY. IS THAT SOMETHING YOU COULD ELABORATE ON? >> RIGHT. SO THAT'S A REALLY GOOD QUESTION. BUT REMEMBER, THE CELL HAS FIVE THOUSAND FAR GETS. AND SO THE CONCEPT THAT WE HAVE FROM BEING A MENDELIAN GENETICIST, TWO ALLELES, YOU GET TWO, YOU'RE DONE. WE COULD HAVE ONE OUT OF FIVE THOUSAND THAT ARE MUTANT HERE. SO YOU COULD HAVE FIVE THOUSAND MUTANTS, A DIFFERENT ONE IN EVERY MITOCHONDRIAL DNA. AND THAT CONCEPT IS ALSO VERY RELEVANT IN CANCER. WE'RE WORKING WITH CANCER, HOW IT AFFECTS CANCER CELL GROWTH, IT'S A REALLY BIG AREA. HOW DO YOU QUANTIFY THAT WHEN YOU HAVE MANY DIFFERENT MUTATIONS? SO ONE OF THE THINGS WE SPENT A LOT OF TIME AND MONEY ON IS TRYING TO DEVELOP WAYS TO LITERALLY ANALYZE EVERY SINGLE MITOCHONDRIAL DNA, DEFINE ITS MUTATIONS AND ADD THAT UP AS A POPULATION BIOLOGY. AND THEN YOU'LL SEE THAT IN FACT EVEN THOUGH ANY ONE MITOCHONDRIAL MUTATION IS VERY LOW, THE SUM TOTAL IS ACTUALLY QUITE PRODIGIOUS. WE HEARD EARLIER TODAY THE STUDIES BY LARSON AND POCOVV WHERE THEY PUT IN A POLYMERASE CAUSING THE ANIMALS TO AGE PREMATURELILY. THEY DON'T GET ONE MUTATION, THEY GET MANY MUTATIONS, AGAIN, YOU HAVE TO QUANTIFY EVERY MITOCHONDRIAL SEPARATELY. >> THANK YOU. >> THE LAST QUESTION, YOU CAN ANSWER OTHER QUESTIONS IN THE LIBRARY IN A MINUTE. FINAL QUESTION OVER HERE, YES? >> YOU SHOWED IN YOUR MOUSE STUDIES THAT THE DILATARIOUS ALLELEE WERE LESS IN EVERY JAYS BUGENERATION BUT IN HUMANS IT DIDN'T HAPPEN. >> I DIDN'T MAKE THAT AS CLEAR AS I SHOULD HAVE. OUR WORK AT THE CURRENT STATE SHOWS -- IN HUMANS, THE FEMALE GENERATES ABOUT 3 TO 5 MILLION PROTO-OCYTES, OVULATING 400 VIABLE EGGS. WHERE ARE THE REST OF THE CELLS GOING? IT SEEMS EVERY TIME A FEMALE GOES TO THE CYCLE, MANY HUNDREDS, IF NOT A THOUSAND, OF THESE BEGIN TO MATURE. AND THEN IN FACT THEY ARE IN COMPETITION WITH EACH OTHER AND ALMOST ALL OF THEM WILL DIE BY APOPTOSIS, ONLY A FEW WILL FORM A MATURE FOLLICLE AND APOPTOSE. IT TURNS OUT THE ONES WITH THE HIGHEST UNDERGO APOPTOSIS. THE CELLS WITH THE MOST MITOCHONDRIA HAVE THE MOST OXIDATIVE STRESS, SO YOU HAVE AN INTRAOVARIAN COMPETITION, THAT COMPETITION IS NOT ON OR OFF. SO EVOLUTION WANTS VARIABILITY TO GO INTO THE SPECIES. I'M ARGUING IT'S THE EVOLUTIONARY RADIATION TOOL. SO THE THRESHOLD IS NOT AT ZERO. IT'S UP AT A LEVEL. WE GET A LOT OF 3243 MUTATION PATIENTS, THAT'S DILATARIOUS WHEN IT'S HOMOPLASMIC. WHEN IT'S LOW, IT GIVES YOU MIGRAINE HEADACHES. IT'S NOT ENOUGH TO CLEAR THE OVARY BUT NOT TO SEE THE PATIENTS IN THE CLINIC. THAT'S WHY WE HAVE SO MANY SPECIFIC MITOCHONDRIAL MUTATIONS. >> PLEASE JOIN THE CONVERSATION IN THE LIBRARY OVER COFFEE AND COOKIES AND PLEASE THANK OUR SPEAKER AGAIN. [APPLAUSE]
