CellnucleusandChromosomesHistoryRobertBrown1773-1858-Discoveredin1831byScottishbotanistRobertBrown-Suggestedthenucleusplayedakeyroleinfertilizationanddevelopmentoftheembryoinplants-Name(nucleus)derivedfromtheLatinwordforkernel/nutIntroductionofnucleusThenucleusisthemostobviousorganelleinanyeukaryoticcell.NucleusisthestorageofthegeneticmessageofthecellinwhichtheDNAreplication,transcriptionoccurs.Itissurroundedbyadoublemembrane.Itcommunicateswiththesurroundingcytosolvianumerousnuclearpores.Thenucleusis,therefore,thecontrolcenterofthecell.Theprominentstructureinthenucleusisthenucleolus.Thenucleolusproducesribosomes,whichmoveoutofthenucleustoroughendoplasmicreticulum.Therearethreemaintypesofinstructionsthenucleusdoes.

Thefirstisthatitdirectscellularreproduction.Thesecondisthatthenucleuscontrolsacell'sdifferentiation.Thethirdtypeofinstructionofthenucleusisthatitregulatesthemetabolicactivitiesofthecell.Generally,nucleusissphericalandcentrallylocatedinthecell.Itsvolumeisabout10%ofthatofthecellanditsdiameteris5-10um.Usuallyeachcellhasasinglenucleus,whereassomecellssuchasosteoclastspossessseveralnuclei.Stillothercellsasredbloodcellshaveextrudedtheirnuclei.Constituents

ofthenucleus:

DNA<20%

nucleoproteinshistonesnonhistones

enzymes

RNAmRNA,tRNA,rRNAStructureofthenucleus:nuclearenvelope:twolipidmembraneschromatin:thegeneticmaterialofthecellNuclearmatrixnucleolus:

thecenterforrRNAsynthesisSectionINuclearenvelopeThenuclearenvelope,composedoftwoparallelunitmembranes,separatedfromeachotherbya10-30nmspace,theperinuclearcisterna.Thenuclearenvelopeisperforatedatvariousintervalsbynuclearpores,whichpermitthecommunicationbetweenthecytoplasmandthenucleus.Thenuclearenvelopehelpscontrolthemovementofmacromoleculesbetweenthenucleusandthecytoplasmandassistinorganizingthechromatin.

Two-membranestructure

Thenuclearenvelopehastwomembranes,eachwiththetypicalunitmembranestructure.Theyencloseaflattenedsacandareconnectedatthenuclearporesites.

(I)structureofnuclearenvelope(I)structureofnuclearenvelopeOuternuclearmembraneInnernuclearmembranePerinuclearspaceNuclearporecomlex1.OuternuclearmembraneTheouternuclearmembranefacesthecytoplasm.Itiscontinuouswiththeroughendoplasmicreticulum.Itscytoplasmicsurfaceusuallypossessesribosomesactivelysynthesizingtransmembraneproteins.Itscytoplasmicsurfaceissurroundedbyathinloosemeshworkofintermediatefilaments,vimentin.2.InnernuclearmembraneTheinnernuclearmembranefacesthenuclearcontents.Itisinclosecontactwiththenuclearlamina,aninterwovenmeshworkofintermediatefilaments,80-100nmthick.Thenuclearlaminahelptoorganizeandprovidesupporttothebilayernuclearmembranesandtheperinuclearchromatin.3.PerinuclearspacePerinuclearspacethespacebetweentheouterandinnermembranes20～40nmcontinuouswithrERspace.

4Nuclearpore

FoundbyH.G.CallanS.G.Tomlinin1949.NamedbyM.L.Wastonin1959.Nuclearporesareformedatsiteswheretheinnerandoutermembranesofthenuclearenvelopearejoined,leavingaspacefilledwithfilamentousmaterial.PoresizeisvariableaccordingtothecelltypesandtissueIngeneral40-100nmNumbersofporesingivenarealowincellswithslowmetabolismandattimesoflowactivityduringcellcyclehighincellsaftercelldivisionandwithhigheractivityofRNAtransportandproteinsynthesisNuclearporedensityofnumbersofthenuclearporespernucleusNuclearporecomplex(NPC)Thenuclearporecomplexisabout80-100nmindiameterandspansthetwonuclearmembranes.It'sthoughttobecomposedoffourelements:StructureofNuclearporecomplexICytoplasmicring

IInuclearringIIISpoke

columnsubunitluminalsubunitannularsubunitIVCentralplug

i

Cytoplasmicring

BecauseitfacetocytoplasmItwasalsocalledouterring.Thereare8symmetricthickfilamentsdistributeonthering.Itissuggestedthatthesefilamentsmayactasastagingareaforthebindingoftheproteinsthataretobetransportedintothenucleus.iinuclearringItfacetotheinnerofthenucleus,andwasalsocalledinnerring.Nuclearringwasmorecomplexthanouterring.Therearealso8filamentsextendinsideto50-70nm.Itformedalittleringonthebottomofthefilamentswhichcomposedof8particles.Thusthewholenuclearringlookslikeafishtrap.Sosomepeoplecalledit

muclearbasket.iiiSpokeThespokefixedonthenuclearmembraneandfacetothecenterofthenuclearpore.Itisalsosymmetricandcomplex.Indetail,spokemaypartintothreesubunit.①columnsubunit

Distributeattheedgeofnuclearpore.Itconnecttheouterandinnerringandsupportthewholenuclearpore.②luminalsubunit

Thedomainwhichlinkedthenuclearmembranewascalledluminalsubunit.Itperforatethenuclearmembraneandextendintotheperinuclearcisterna.③annularsubunitiscomposedof8particlesandformedachanneltoexchangesubstances.

ivCentralplugTheparticlethatliedinthecenterofthenuclearpore.Itwasalsotermedcentralgranule.Thecurrentunderstandingisthatthetransporterfunctionsintransportofmaterialintoandoutofthenucleus.Butnotallthenuclearporescouldbeseenthisstructure.Sosomescientiststhinkthatcentralplugisnotastructurepartofnuclearporebutaparticlethatjustbeingtransportedthroughthenuclearpore.35nmindiameter,transporterNuclearporetransport

Duetothestructuralconformationofthesubunitsofthenuclearporecomplex,thereareseveral9-11nmwidechannelsavailableforsimplediffusionforionsandsmallmolecules.Substanceslargerthan11nmareselectivelytransportedviaareceptor-mediatedtransportprocess.Moleculesenterandexitthenucleus

throughnuclearporecomplexBidirectionaltrafficNuclearproteintransportmechanismbasicdefinition

◆nuclearprotein◆nuclearlocalizationsignals,NLS◆nuclearexportsignals,NES◆importin◆exportin

MaterialsexchangeTransportinandoutofthenucleuscanoccurinseveralways.Passivetransport-diffusionActiveTransportPassivetransport—passivelydiffuse3000-4000NPC/cell(mammalian);EachNPCcontainsoneormoreopenaqueouschannels:9nmindiameterand15nmlong<10nmindiameter<60kdglobularproteinAbletoenterthenucleusActivetransportTransportoflargeproteinsintonucleusneedsnuclearlocalizationsignal(NLS)NucleoplasminNuclearLocalizationSignals(NLS)basicorclassicNLSImportingproteinsfromthecytoplasmintothenucleusTheproteinbearingaNLSbindstotheimportinα/βtoformatransportingcomplex.Transportingcomplexbindswithacytoplasmicfilamentwithimportinβ.Thereceptor-cargocomplexmovesthroughthenuclearpore.ThecargocomplexinteractswithRan-GTPanddissociates.Theimportinsubunit,inassociationwithRan-GTP,istransportedbacktothecytoplasm,wheretheRan-GTPishydrolyzedRan-GDPissubsequentlytransportedbacktothenucleus,whereitisconvertedtoRan-GTP.Conversely,importinistransportedbacktothecytoplasm.ImportedmanytypesofproteinsDNApolymerase,RNApolymeraseandotherproteinsareimportedthroughnuclearpore.NuclearexportNuclearexportworkslikenuclearimport,butinreverse

hnRNPproteinscontainanuclear-exportsignal(NES)Reference:Cell92:327,1998mRNA,tRNA,subunitofribosomecouldtransportoutofthenucleustocytoplasm.Structureandfunctionof

nuclearlaminaUnderlyingtheinnernuclearmembraneisthenuclearlamina.Thenuclearlaminaisadense(~30to100

nmthick)fibrillarnetworkinsidethenucleusofaeukaryoticcell.Itiscomposedofintermediatefilamentsandmembraneassociatedproteins.CompositionofnuclearlaminaLaminsA:nexttoNuclearskeletonlaminsC:nexttoNuclearskeletonlaminsB:neartheinnernuclearmembrane.Theymaybindtointegralproteinsinsidethatmembrane.laminsB1laminsB2Functionalofnuclearlamina

(1)providingstructuralsupporttothenucleusbybindingtoinnernuclearmembraneproteins(2)Playingaroleinnucleusassemblyanddisassemblybeforeandaftermitosis.NuclearlaminadisassemblyinMphase

(3)ServeasasiteofchromatinattachmentChromatinorganizationDNAreplicationStructureofthenucleus:nuclearenvelopechromatinnucleolusnucleoplasmSectionIIChromatinandChromosome1WaltherFlemmingnamedchromatin(stronglyabsorbedbasophilicdyes)in1879.Flemmingsurmisedforthefirsttimethatallcellnucleicamefromanotherpredecessornucleus2.WilhelmvonWaldeyer-HartzNamedthechromosomes(meaningcolouredbody)in1888HistoryChromatin:Chromatincarrythegeneticinformationoforganismwhichliesininterphase.ThecomplexofDNAandproteinwhichmakeupthecontentsofthenucleusofacell.Cromatiniscomposedof:DNAHistonesNonhistonesRNAWhatischromatinandchromosome?Chromosome:InM-phase,thechromatinfibersareextensivelycondensedtoformchromosomes,structuresthatarevisiblewiththelightmicroscope.Whatischromatinandchromosome?ReplicationoriginCentromereTelomereThreeimportantDNAsequencesofchromatin◆ThehistonesaresmallproteinscontainingahighproportionofbasicaminoacidswithpositivechargethatfacilitatebindingtothenegativelychargedDNAmolecule.HistonesareRichinLysineandArginine◆Therearetwoclassesofhistones:NucleosomalhistonesH2A,H2B,H3,andH4whicharehighlyconserved.H1-likeHistone

VariableHistonesareboundwithDNAinanon-specificway.HistonesStructuralOrganizationoftheCoreHistonesNonhistones:

ThisDNA-associatedproteinsbindDNAinaspecificway,thereforenamedsequencespecificDNAbindingprotein,whichcanformseveralspecificstructuresandmayalsoinvolvedintheregulationofDNAactivities.NonhistonesNonhistonesEuchromatinand

HeterochromatinDependingonitstranscriptionalactivity,chromatinmaybecondensedasheterochromatinorextendedaseuchromatin.

Euchromatin:scatteredthroughoutthenucleusandnotvisiblewithlightmicroscope,whichistheactiveformofchromatinwherethegeneticmaterialoftheDNAmoleculesisbeingtranscribedintoRNA.

LieisthecenterofthenucleusReplicationatearlystageofSphase.Heterochromatin:itisvisibleunderlightmicroscope,whichlocatedmostlyattheperipheryofthenucleusbutalsoformsirregularclumpsthroughoutthenucleus.ItiscondensedinactiveformwhichmeansitisnotactiveinRNAsynthesis.ItcanbedividedintoConstitutiveheterochromatinandFacultativeheterochromatin.NotranscriptionalactivityConstitutiveheterochromatin:Exceptforreplication,thistypeofheterochromatinisalwaysinacondensedinactiveformduringthewholecellcycle.Facultativeheterochromatin:Insometypesofcellsorincertaindevelopmentalphases,someeuchromatinbecomescondensed,losingtheirtranscriptionalfunctionandturnsintoheterochromatin.Itisawaytoclosegeneactivity.Forexample:XChromosomeBarrBody（

XChromosome）:microscopicstudyofinterphasenucleiofcellsfromfemaledisplaysaverytightlycoiledclumpofchromatin,thesexchromatin(BarrBody),theinactivecounterpartofthetwoXChromosomes.Itisafacultativeheterochromatin,whichisoneofthetwoXchromosomesbecomingcondensedrandomlyatembryonic16daysoffemale.XchromosomeinactivationinmammalsXXXYDosagecompensationXAvnerandHeard,Nat.Rev.Genetics20012(1):59-67Xist(X-inactivespecifictranscript)ThefirstlncRNA(longnon-codingRNA)ChromatinpackagingEachhumancellcontainsabout2mofDNAwithinnucleusifstretchedend-to-end,yetthenucleusofahumancellitselfisonlyabout6umindiameter.Compactionratio=nearly10000-fold.FromDNAtochromosomeneedsfour-steppackaging.Step1Nucleosome

ThebasicUnitofChromatin

Thefirststepofcondensation,from2nmto11nm.

Evidence:

(1)ElectronmicrographsofchromatinfibersIsolatedfrominterphasenucleus:30nmthickChromatinunpacked,showtheunclesomeEvidence:

(2)Nucleasedigestion(Ratliverchromatin)WhatisNucleosome?Nucleosomeisthebasicunitofchromatin.Eachnucleosomeismadeupofanoctomerofproteins,duplicatesofeachoffourtypesofhistones(H2A,H2B,H3andH4).Thenucleosomeisalsowrappedwith1.75turnsoftheDNAmoleculethatcontinuesaslinkerDNAextendingtothenext“bead”.Thespacingbetweeneachnucleosomeisabout200basepairs.AhistoneoctamerformsthenucleosomecoreHistoneoctamer:(H2A-H2B)-(H3-H4)-(H3-H4)-(H2A-H2B)WhereisthehistoneH1?H1moleculesareassociatedwiththelinkerregion.146+15~50bplinkerDNA200bpDNA:LinkerDNA:15-50bpNucleosomalDNA:146bptowrap1.65timesaroundthehistonecore.CyclicDiagramfornucleosomeformationanddisruptionChromatinRemodelingChromatinremodelingisthedynamicmodificationofchromatinarchitecturetoallowaccessofcondensedgenomicDNAtotheregulatorytranscriptionmachineryproteins,andtherebycontrolgeneexpression.CovalentModificationofcorehistonetailsAcetylationoflysinesMythylationoflysinesPhosphorylationofserinesHistoneacetyltransferase(HAT)Histonedeacetylase(HDAC)StepIISolenoidSolenoid:(Chromatinfiberofpackednucleosomes)Thesecondstepofcondensation,30nm.Packagingofchromatininto30nmisbelievedtooccurbyhelicalcoilingofconsecutivenucleosomesatsixnucleosomesperturnofthecoilandcooperativelyboundtherewithHistoneH1.StepIIISupersolenoidSupersolenoid:Thethirdstepofcondensation,300nm.Tightercondensingofthechromatinmaterialisaccomplishedbyloopingthecoiled30nmfibersinto300nmloopsheldtogetherbyspecificprotein/DNAboundcomplexeslocatedattheirbases.RadicalloopmodelChromosomes:Theforthstepofcondensation,700nm.Furthercoilingofthe300nmloopsintotightlywoven700nmhelicalloopsformsthemaximallycondensedchromosomesobservedinthemetaphasestageofmitosisormeiosis.chromatidchromosomeStepIVmetaphasechromosomesDNAnucleosome

solenoidsupersolenoidchromosomeChromosomeinmetaphaseKaryotypeisthenumberandappearanceofchromosomesinthenucleusofaeukaryoticcell.HumanmitoticchromosomesandkaryotypeChromsomescanbedividedintofourtypesdependonCentromerepositionsThereisnotelocentricchromosomeinhumancellsHumanchromosomeNo.14

secondaryconstrictionnucleolarorganizingregion,NORsatellitetelomerecentromereAtypicalmitoticchromosomeatmetaphaseChromatidKinetochoremicrotubulesCentromereregionofchromosomeMetaphasechromosomeCohesinproteinsKinetochoreCopyright?TheMcGraw-HillCompanies,Inc.Permissionrequiredforreproductionordisplay.MainstructuresofchromosomeTheCentromereandKinetochore:serveasasitefortheattachmentofspindlemicrotubulesduringmitosisandmeiosisMainstructuresofchromosomeThecentromereisthepartofachromosomethatlinkssisterchromatids.Duringmitosis,spindlefibersattachtothecentromereviatheKinetochore.Thekinetochoreistheproteinstructureonchromatidswherethespindlefibersattachduringcelldivisiontopullsisterchromatidsapart.CentromereKinetochoredomainCentraldomainPairingdomainMainstructuresofchromosomeKinetochoresPowerChromosomeMovementsinMitosisMainstructuresofchromosome

ENDREPLICATIONPROBLEMTelomereMainstructuresofchromosomeToovercomeend-replicationproblem.PreventrandomBindingbetweenchromosomes.TTAGGGinHuman.Between3and20KbinlengthRepetitivesequencesattheendsofchromosomes.CentromereTelomereTelomereMainstructuresofchromosomeTelomereFISH(HumanProbe):TTAGGGMainstructuresofchromosomeTelomereTelomeraseisanenzymewhichaddsDNAsequencerepeats("TTAGGG"inallvertebrates)tothe3'endofDNAstrandsinthetelomereregions,whicharefoundattheendsofeukaryoticchromosomes.MainstructuresofchromosomeTelomeraseTelomere&AgeingInhumans,telomeresinmostsomaticcellsshortenwithage.Thesecelltypesdonothaveenoughtelomeraseactivitytomaintaintheiroriginaltelomerelength.MainstructuresofchromosomeRoleofTelomeresinCancercanceroustumorsacquireindefinitereplicativecapacitybyover-expressingtelomerase.telomeresinhumantumorswereshorterthantelomeresinthenormalsurroundingtissueHayflicklimitisbelievedtohelppreventcancer.MainstructuresofchromosomeHumanchromosomeNo.14

secondaryconstrictionnucleolarorganizingregion,NORsatellitetelomerecentromereAtypicalmitoticchromosomeatmetaphaseSecondaryconstrictionSecondaryconstrictionisseenatthechromosomeinadditiontoprimaryconstriction/centromere.Somepartsoftheseconstrictionsindicatessitesofnucleolusformation.SatellitechromosomeSatellites

AsmallterminalsegmentofthechromosomeConnectedtoitsendbyasecondaryconstrictionisknownassatelliteandchromosomeswithsatellitesareknownassatellitechromosomesorSATchromosomes.

SectionIIINucleolus

Themostprominentsubstructurewithinthenucleusisthenucleolus,whichisthesiteofrRNAtranscriptionandprocessing,andofribosomeassembly.Thenucleolusisobservedonlyduringinterphasebecauseitdissipatesduringcelldivision.Fibrillercenter

Densefibrillarcomponent

Granularcomponent

StructureofthenucleolusFibrillarcenter

TherRNAgenesarelocatedinthefibrillarcenters.(NOR)Nucleolusorganizerregion(NOR)ornucleolarorganizerisachromosomalregionaroundwhichthenucleolusforms.TheregioncontainsseveraltandemcopiesofribosomalDNAgenes.Inhumans,theNORcontainsgenesfor5.8S,18S,and28SrRNAclusteredontheshortarmsofchromosomes13,14,15,21and22NORsinhumanchromosomes:13\14\15\21\22LocalizationofNORsinMetaphaseChromosomesInmetaphasechromosomestheNORsappearinmanyspeciesasparticularlythinregions,the"secondaryconstrictions"(Heitz1931;McClintock1934).MatsuiandSasaki,usingastainingprocedureknownasNbanding,concludedthatthesatellites,notthecentromeres,orstalks(secondaryconstrictions),aretheNORs.Densefibrillarcomponent

SurroundingFC

ContainingnucleolarRNAsbeingtranscribed.CompostionrRNA

RNP

Granularcomponent

MaturingribosomalsubunitsareassembledinGC;Particlediameter:15-20nm

RNP:protein+rRNA

Fun