分子生物学10-22-核小体.ppt

染色体与DNA,FigureCO:Xchromosomeinblue,染色体,2.1.1染色体概述,染色体(chromosome)是细胞核中载有遗传信息的物质，在显微镜下呈丝状或棒状，由核酸和蛋白质组成，在细胞发生有丝分裂时期容易被碱性染料着色，因此而得名。同一物种内每条染色体所带DNA的量是一定的，但不同染色体或不同种中间变化很大。,原核细胞染色体外裹着稀疏的蛋白质，其中一部分与DNA的折叠有关，另一些则参与DNA复制、重组及转录过程。真核细胞的染色体中，DNA与蛋白质完全融合在一起，其蛋白质与相应DNA的质量比约为1:1.,chromosomeAdiscreteunitofthegenomecarryingmanygenes.EachconsistsofaverylongmoleculeofduplexDNAandanapproximatelyequalmassofproteins.Itisvisibleasamorphologicalentityonlyduringcelldivision.,Athinsectionshowsthebacterialnucleoidasacompactmassinthecenterofthecell,原核细胞染色体(类核Nucleoid),ThenucleoidspillsoutofalysedE.colicellintheformofloopsofafiber,ThebacterialgenomeconsistsofalargenumberofloopsofduplexDNA,FIGURE08:AnunrestrainedsupercoilintheDNApathcreatestension,butnotensionistransmittedalongDNAwhenasupercoilisrestrainedbyproteinbinding,Thesisterchromatidsofamitoticpaireachconsistofafiber(30nmindiameter)compactlyfoldedintothechromosome,真核细胞染色体,类核；染色质；包装比例,nucleoidThestructureinaprokaryoticcellthatcontainsthegenome.TheDNAisboundtoproteinsandisnotenclosedbyamembrane.chromatinThestateofnuclearDNAanditsassociatedproteinsduringtheinterphase(betweenmitoses)oftheeukaryoticcellcycle.packingratioTheratioofthelengthofDNAtotheunitlengthofthefibercontainingit.,SpecificSequencesAttachDNAtoanInterphaseMatrix（分裂间期基质）,DNAisattachedtothenuclearmatrixatspecificsequencescalledMARsorSARs.TheMARsareA-T-richbutdonothaveanyspecificconsensussequence.chromosomescaffold（支架）Aproteinaceousstructureintheshapeofasisterchromatidpair,generatedwhenchromosomesaredepletedofhistones.,ChromatinIsDividedintoEuchromatinandHeterochromatin,Regionsofheterochromatinremaindenselypackedthroughoutinterphase.chromocenterAnaggregateofheterochromatinfromdifferentchromosomes.,Regionsofcompactheterochromatinareclusterednearthenucleolusandnuclearmembrane,常染色质和异染色质EuchromatinandHeterochromatin,DNAIsOrganizedinArraysofNucleosomes（核小体）,ThecoreDNAisthelengthof146bpthatisfoundonthecoreparticlesproducedbyprolongeddigestionwithMNase.LinkerDNAistheregionof8to114bpthatissusceptibletoearlycleavagebynucleases.,FIGURE05:Micrococcalnucleaseinitiallycleavesbetweennucleosomes,TheNucleosomeIstheSubunitofAllChromatin,Anucleosomecontains200bpofDNAandtwocopiesofeachcorehistone(H2A,H2B,H3,andH4).DNAiswrappedaroundtheoutsidesurfaceoftheproteinoctamer（八聚体）.ThehistoneoctamerhasastructureofanH32-H42tetramerassociatedwithtwoH2A-H2Bdimers.,FIGURE06:ThenucleosomeconsistsofapproximatelyequalmassesofDNAandhistones(includingH1),TheNucleosomeIstheSubunitofAllChromatin,TheNucleosomeIstheSubunitofAllChromatin,ThenucleosomeisacylinderwithDNAorganizedinto12/3turnsaroundthesurface,TheNucleosomeIstheSubunitofAllChromatin,Eachhistoneisextensivelyinterdigitatedwithitspartner.Allcorehistoneshavethestructuralmotifofthehistonefold.N-andC-terminalhistonetailsextendoutofthenucleosome.H1isassociatedwithlinkerDNAandmaylieatthepointwhereDNAentersorexitsthenucleosome.,PhotoscourtesyofE.N.Moudrianakis,JohnsHopkinsUniversity.,FIGURE10ab:Thecrystalstructureofthehistonecoreoctamerisrepresentedinaspace-fillingmodel,TheNucleosomeIstheSubunitofAllChromatin,Thehistonefoldconsistsoftwoshorta-helicesflankingalongera-helix,Histonepairs(H3+H4andH2A+H2B)interacttoformhistonedimers,StructuresfromProteinDataBank1HIO.G.Arents,etal.,Proc.Natl.Acad.Sci.USA88(1991):10145-10152.,TheNucleosomeIstheSubunitofAllChromatin,Thehistonefolddomainsofthehistonesarelocatedinthecoreofthenucleosome,NucleosomesAreCovalentlyModified,Histonesaremodifiedbymethylation,acetylation,phosphorylation,andothermodifications.Combinationsofspecifichistonemodificationsdefinethefunctionoflocalregionsofchromatin;thisisknownasthehistonecode.,Histonetailshavemanysitesofmodification,NucleosomesAreCovalentlyModified,PhotocourtesyofSeanD.Taverna,JohnsHopkinsUniversitySchoolofMedicine,andHaitaoLi,MemorialSloan-KetteringCancerCenter.AdditionalinformationatS.D.Taverna,etal.,Nat.Struct.Mol.Biol.14(2007):1025-1040.,Numerousproteinmotifsrecognizemethylatedlysines,NucleosomesAreCovalentlyModified,Acetylationduringreplicationoccursonspecificsitesonhistonesbeforetheyareincorporatedintonucleosomes,NucleosomesAreCovalentlyModified,Acetylationassociatedwithgeneactivationoccursbydirectlymodifyingspecificsitesonhistonesthatarealreadyincorporatedintonucleosomes,DNAStructureVariesontheNucleosomalSurface,DNAiswrapped1.67timesaroundthehistoneoctamer.DNAonthenucleosomeshowsregionsofsmoothcurvatureandregionsofabruptkinks.ThestructureoftheDNAisalteredsothatithasanincreasednumberofbasepairs/turninthemiddle,butadecreasednumberattheends.,StructuresfromProteinDataBank:1P34.U.M.Muthurajan,etal.,EMBOJ.23(2004):260-271.,ThePathofNucleosomesintheChromatinFiber,10nmchromatinfibersconsistofastringofnucleosomes.30nmfibershavesixnucleosomes/turn,whichareorganizedintoatwo-starthelix.HistoneH1,histonetails,andincreasedionicstrengthallpromotetheformationofthe30nmfiber.,ThePathofNucleosomesintheChromatinFiber,The10nmfiberisacontinuousstringofnucleosomes,The30nmfiberisatwostarthelixconsistingoftworowsofnucleosomescoiledintoasolenoid,Levelsofchromatinpackaging,ReplicationofChromatinRequiresAssemblyofNucleosomes,Histoneoctamersarenotconservedduringreplication,butH2A-H2BdimersandH32-H42tetramersareconserved.Therearedifferentpathwaysfortheassemblyofnucleosomesduringreplicationandindependentlyofreplication.,ReplicationofChromatinRequiresAssemblyofNucleosomes,Nucleosomeassemblyinvivo,NucleosomesAreDisplacedandReassembledDuringTranscription,Mosttranscribedgenesretainanucleosomalstructure,thoughtheorganizationofthechromatinchangesduringtranscription.Someheavilytranscribedgenesappeartobeexceptionalcasesthataredevoidofnucleosomes.RNApolymerasedisplaceshistoneoctamersduringtranscriptioninvitro,butoctamersreassociatewithDNAassoonasthepolymerasehaspassed.,NucleosomesAreDisplacedandReassembledDuringTranscription,AnexperimenttotesttheeffectoftranscriptiononnucleosomesshowsthatthehistoneoctamerisdisplacedfromDNAandrebindsatanewposition,NucleosomesAreDisplacedandReassembledDuringTranscription,Nucleosomesarereorganizedwhentranscriptionpassesthroughagene.AdditionalfactorsarerequiredbothforRNApolymerasetodisplaceoctamersduringtranscriptionandforthehistonestoreassembleintonucleosomesaftertranscription.,染色体带型G（Giemsa)-bandinggeneratesacharacteristiclateralseriesofbandsineachmemberofthechromosomeset,ThehumanXchromosomecanbedividedintodistinctregionsbyitsbandingpattern,每条带约10000Kb,Amagnifiedviewofbands87Aand87CshowstheirhybridizationinsituwithlabeledRNAextractedfromheat-shockedcells,Chromosomesarepulledtothepolesviamicrotubulesthatattachatthecentromeres,有丝分裂中期,有丝分裂后期,有丝分裂末期,着丝点,粘结蛋白,微管,Amodeloftheoverallstructureofaregionalcentromere,FIGURE26:AlargeproteincomplexassemblesattheCDEsequencesandconnectsthechromosometomicrotubules,AtypicaltelomerehasasimplerepeatingstructurewithaG-T-richstrandthatextendsbeyondtheC-A-richstrand,端粒有简单的重复序列AtypicaltelomerehasasimplerepeatingstructurewithaG-T-richstrandthatextendsbeyondtheC-A-richstrand,ThecrystalstructureofashortrepeatingsequencefromthehumantelomereformsthreestackedGquartets,端粒封堵线性染色体的末端,AloopformsattheendofchromosomalDNA,染色体末端形成环状结构,AloopformsattheendofchromosomalDNA,端粒的功能保护染色体末端：端粒延长协助减数分裂中同源染色体配对,ThemeiotictelomereclusterisvisualizedbytelomereFISH,端粒对生存是必需的,1.在分裂细胞中存在端粒酶活性。2.端粒酶突变后，每次分裂，染色体变短。3.端粒消失后，细胞不能分裂。,Mutationintelomerasecausestelomerestoshortenineachcelldivision,Telomerelengthismaintainedat350bpinwild-typeyeast,