Multivariate statistical analysis of epigenetic regulation with application to the analysis of human embryonic stem cells [Elektronische Ressource] / Lukas W. Chavez Wurm

Sujets

• Naturwissenschaften
• Biologie

Informations

• Publié par: freie_universitat_berlin
• Publié le: 01 janvier 2010
• Nombre de lectures: 9
• Langue: English
• Poids de l'ouvrage: 5 Mo

Extrait

GMultivaereicriate(Dr.statisticalUnivanalysisderofFepigeneticInformatikregulationErlangungwitheinesapplicationhaftetonat)thehanalysisMathematikofFhumanBerlinembdesryradesonicDoktorsstemNaturwissensccellsnLukrer.asamW.acChabvhezundWderurmreien2010ersit?tDissertationzurMolecularSupdisputation:ervisor:alterProf.VingronDr.Prof.MartinSaarlandes)VingronDr.(Max-Planck-Institutek-InstitutenforJ?rnMolecularersit?tGeofnProf.etics)MartinA(Max-Plancdvisor:forDr.GeRalfetics)HerwigDr.(Max-PlancWk-Institute(UnivfordesMolecularDateGenetics)theReview15.12.2010er:43Contents.1.Introvdu.c.t.iCpGo.n.53.3.51.162Humanorkem.bry.onic.stem.cells.(hES.Cs)......of...man.and.................er..........5eth1.2.T.ranscri56ptional.regulation.ofagepluri.pdepotency4.1.68...con...deriv..........................6441.3.DNA.meth.ylation3.3.2.......3.3.3.......3.3.4.......score.....of...57.ylated...w.......4.dermal.stem.core.trolling.tal.....4.39.1.4.In.troMeDIP-Seqductionprolesto.metho3.2.2dsanalysisand.limitations.for.d.etec.ti.ng40DNAhmenmeth.ylation...10.1.5.Aims.of.theNormalizationthesis................Reads.(rpm).............factors...............curv...............e12score2.Exp.erimental.techniques.16.2.1methDNA.arra.ys......Ev.data.......Iden.tially.(DMRs).....MEDIPS.pac...............and.during.of.ry.ells.tifying.net.OCT4.otency.Exp.ced.metho..16.2.2.Statistical.testingqualitometricsf.dieren.tial.expression....4.4.W.meth...77...33.Co.erage..............19.2.3.Second.generation.sequen.c.i.ng3.2.3.enric.t.............................3.3................24.2.4.Chromatin.imm.unoprecipitation.(ChIP)........3.3.1.p.million.........................4425Coupling2.5.Meth.ylated.DNA.imm.unoprecipitation.(MeDIP)................45.Calibration.e........27.2.6.Alternativ.e.metho.ds.for.detecting.DNA.meth48ylationRelativ.m.ylation.(rms)..................28523AbsoluteMoylationdelling(ams)of.MeDIP-Seq.data.30.3.1.Genome.v.ector....3.3.6.aluation.MeDIP-Seq.normalization.............3.4.tication.dieren.meth.regions...........3.5.soft.are.k................30.3.2.Qualit.y.con.trol65metricsGenetic.epigenetic.endencies.endo.dierentiation.hu-.emb.onic.c.68.Iden.a.regulatory.w.of.con.plurip...4.2.erimen.pro.ures.analysis.ds..........3.3.3.2.170SaturationMeDIP-Seqanalysisy.trol.......................74.Comparing.and.GSBS.ed.ylation.......3.endixContiten.ts.4.5ciatesPromoter.meth.ylationBibliography...histone.....alterations.on...110...hmen.en...........ylation...8.net...91.Abb.111.endix...4.10.analysis.ylation79to4.6dicationsMeth.ylation.patterns.of.transcrip.tion.factor.binding.si4.11telsgene.......Epigenetic.OCT4.ork......79Conclusion4.7NotationIden109ticationcknoof112dieren114tially116meth.ylated.regions85(DMRs)Enric.t.asso.de-meth.ev.ts.functional.mo....79.4.8.Genome.wide.distri.bution.of.DMRs................85.Dieren.a.meth.and.expression...........8.4.12.eects83the4.9regulatoryDierenwtial.meth.ylation.at.TFBSs......5.94.98.and.reviations.Publications.A.wledgments.Zusammenfassung.App.1.App.2.4.stem1patienIntrohromatinductionthe1.1fromHumanedembtheryeonicadultstemwcellscell-replacemen(hESCs)donor-hostInhES1989,aThstudoformson[96]andindepcolleaguestheirpresenyteddiforfuturetheyrstcausedtimealternativthebderivcolleaguesationedofotenhESCspluripfromtran-themolecularinnerakcelltimemassSubsequenofpresenhBothumanharacteristicblativainstotcystsfor[98].First,Fareorcthis,incleaikvgenomicageline.stageforhcellsumandevem-abryeos,canprowithoutduceddevbotherysomaticincellsvitrofos)fertilizationcellsforeclinical1.1).purpsoses,demonstratedwberebcultured2007,tooratoriesblastoiPSCscysttiatedstageeandthetheorinnertiationcellthreemassesiPSCswgeneeremethisolatedo(seecuFiguree1.1).therapiThomsonputativandapplications,colleaguesallodenedbthreetaminatedessentstialculture.careharacteristicstoofyprimatekgroundsemandbryortunatelyonicapproacstemationcellshES-lik[98].vTheserecenareed.(i)hand,derivaahatiwnohnefromsingletheterferingpreimplbryatalnOntationsevoronptoeriimplanttationhaemthatbryao,factors(ii)sucienprolongeddierenun-todierensimilartibrya(seeted2006,proliferationh(andstemnessv),theandiPSCs(iii)generatedstablebdevinelopmen,talwptotenvtialhtoerivformideriv[95,ativhaeswnofcellsallotencythreehESCs.emtheybrySCsonictogermoflagermyFersbleevmorphologyen,afterandprolon5gwedmainculture(ltiespluripputativotencyfuture).tHumanes.ESCsforoereinsighclinicaltshESCsinnottowdevtoelopmenetalonevbenreagentsusedwhiccellhSecond,cannotrejectionsbleelystudiedarisedibredieringctlbacyofintthecellinFtact,heumanhesemderivbryofoandbutethathahaeveenetlyimpelopoOnrtanonetKilmanskconsequencesyiandn[50]clinicalvareas.shoFthortexample,ESCsunbderstandingderivthefrommecblastomershanismsinthatwithconemtrolonicdierenelopmentiationpenablestial.directedthedif-hand,fereneraltiiesareprogrammingticellsoinducednotenofstemhES(iPSCs)cellsvtodemonstratedsptransductioneciconlycellfewtscriptionyp(TFesis[98].tHoresettingwtiatedeviner,aderivstateationtoofmESCsonicfromcellshFigureumanInblastoTcystsaforaclinicalhiapplicationscolleagueshashatoebforerstconsideredthatcriticalcanbeecausefromofethicalroandlaststecmouse.hntlyicalinreasons.tTheoethicalendenconcernslabarehaobevious,tedbumanecausedisolationedofdtheereninnerbroblastscell107].massgroupsbvyshodestructionthatofiPSthefulllempluripbrycoofinFvthis,erselysholeadsiP-todierentheinabderivortionesofallemprimarybrylaogenesis.ers.Furthermore,romresemthehESCstectheirhnical,pexpressionoinctstateofDNAview,y-therearehESCs1kInthattrotransductionranscriptioFigureis1.1:cellsEmubryhanismsonicexpressedstemnetcellsonandereprahashiogramming.stillEmeects.bryvonictostemdomainceofllscoareOCT4derionicvevedthefromrstheeminneretcellX2,massconsideredofossiblemammalianer,blastoservcysts.analyzingTheytargetedhaypvpluripeOCT4thennerdevItelself-renopmenytalvpwnotengtialSevtosomaticdierenroletiateplinbrytoaderivforativmolecularescells.ofandalliPSthreeTFemapplicationsbrybonicbgermoflsirableawyresearcersiPSCseasvresourceenmolecularafteronsibleprolongedtiationculturecell(1.2pluripregulationotencyThe).scriptionSevhigheralthestudiesmaonblastoreprogrammingsuppsomaticotencycellsaltotrolledinducedtranscriptionpluriporkotenbtthatstemofcellsfor(iPSCs)emhatiationvstuehdemonstratedtothatemphasizetheOCT4transductionregulatorofotencyonlymammalianaHere,feofwtranscriptiontranscriptionsucienfactorsdieren(TFtos)similarisonicsucienTtal.forernresetting[104]dierenbtiateofdOCT4,cellsnicalinhastotoaemolecularcritical,stecaasetpeunde-similarsidetoHoemevbryforonich,standealreadymecells.a(Individualaluableimaforgestheillustratingmectherespblastoforcyst,dierenretroinviruses,functionalandtthees.vitruvianTmannalwofereotencytakPOUentranfromfactorhisttp://wilykiniipcelledia.org/.)sslationmammalianccysts.haracteristicsis[104].osedTpluripakandenewtogether,isthesenstudiesbdemoanregulatorystratewdgothaternedhESCsyorandiPSCsdocan-regulationbOCT4ecrucialginitiatinenearlyerabryteddierenwithout[6].beraleingdiesburdenedreprogrammingbumanycellstheiPSCsethicalenconcernstheofofdestructivaseeyhESCofgeneration.uripTheinpearlyossibilitemyo.tothgeneratetransductionpatienonlytfewspfactoecicisiPSCstwillresettingputativtiatedelyinreduceadonor-hoststaterejectionstoinbryfstemutureWhilecell-replacemenaktettherapies.[95]NevWertheless,igtheal.utiliobtainedzcellsationyofductiontransfectedthesomaticscellsSOfor6clilin1yInresultingtroindingductionerimenKLF4,oandvidesC-MYC,theYwnuOCT4etapprsoacal.homeo[107]wnacandhievetedal.similarerimenresultsactivwithpiecesategratedtranscriptionoffactoridensetteractscompwosedcomplexofsequencesOCT4,enSOabilitX2,wNANOG,onandscriptionLIN28.suppressionInexpressionterestinglyb,edonlytrophoblasttheandTFordshaOCT4withandtrollingSOeacXv2oareTTTGCAcommonfactorsinTFbmotifothX2/OCT4approacMappinghes.motifsMoreoOCT4vder,hipHuangfuhaetputatival.hniques[39]transcriptionaldemonstratedbindingthatIniPSCsOCT4cantargetbedeinderivtheedyatchighercecienciestob,ydierentheChIP-on-ctransductionromofprothesetranscriptionaltidenworkoerformedfactorshin(seecomnetbinationhESCswithgeneshistoneindividualdeacet1.2).ylasebinhibitorT-varealpmotifrothatiPOUceacid.conThe,TFasOCT4ognizedisSknoeterown18,astheseafactorktheeyputativregulatorgenesfevidenceobindingrAlthoughmainerimentainimethongepluriptootencyprotein-DNAinthesemammaliannotemofbryendenciesoseen[73,the81,expression1]toandinuenceistheexclusivofelyBabaieexpressedpininmammaOCT4lH1ianandemglobalbryhangesonicystemTcells.induFOCT4ordexample,expitgeneswpluripasdshoupwnalongthatineageOCT4ipis,expressedsequenceneitherinincomplemenfunctionalinformationsomaendencies.tircacellsnetnhESCs,oreinanadultofstemdatacells,sequenceb4.1).yOCT4aorkmicroarraotencyycompbasedsetgenebexpressionofproleerimenco(seemphaaerieensotied.nwbexamplesetthewctamereenAhESCs,Tadultinlivwitherdomainprogenitorlikcells,theanddomainhepatotainingcytesOCT4[as4ell8].aTherec-HMG-bbothexOconhtainingdimerTF[93,SO87].Xof2knointranscriptionteractsbindingwithtoOCT4promoterandoftheeSOtargetX2/OCT4proheteroadditionaldimerforcomplexirectisevablets.toChIP-on-cpro-expmotetsselectivsequence-basededsgenevactivtheationyordetectrepressioneduringbmammaliansites,emtecbrydoogenesisallo[82,inference12directional,dep74].bFwunctionalDNAdataandoneectOCT4generegulatoryregulation.actionorderistestaregulatoryvofailabletofromtran-heterogeneousratesources:itstogenes,reveteal[6]DNA-ProteinerformbindingRNAevterference-mediatedenoftsfunctionoftheOCT4,hESCSOeX2analyzedandresultingofgenethecpluripbotemicroarranexpcyts.assoranscriptionalci-hangesatedcedTFyNANOG,knockhromatinoimmareuno-precipitationectedfolloincludewlinkedwithbotencyyanmicroarragenesyatedexponerimentiationtsthe(ChIP-on-clhip)[6].hashbexpeentspperformedoterusinganalysishESCsRNA[13].terferenceComplemenvidetarytarytoofOCT4onbindidepngIneveentotstifyincorehESCs,regulatorywweinhaIvvepiden[17]tiedinOCT4analysisbindingsucevhigh-throughputenalongtspromoterinanalysishsectionumanTheemcorebryregulatoryonicwcarcinomaconcellspluripbinyisChIP-on-Chiposedexptheerimenoftsdetected[49].yMoreohvtheer,expOCT4talsphesecicFiguresequence7motifsb1