生理化學教學課件：Regulation of Gene Expression in Eukaryotes

ExpressioninEukaryotesRegulationofGeneExpressioninEukaryotesEukaryoteHasaVeryHugeGenomeHumangenomehas3.3billionbp.HumanGeneshave1-178exonsAverageexoncodesfor48aminoacidsAveragepolypeptidehasalengthof430aminoacids.EukaryoticDNAhasManyTandemRepeatsChromatinActivationChromatinisthecombinationofDNAandproteinsthatmakesupchromosomes.

Euchromatinhasaloosestructure,whereasheterochromatinismoretightlycondensed.Genesareactivelytranscribedineuchromatinbutarerepressedinheterochromatin.EukaryoticDNAIsTightlyPackagedInARepeatingArrayofNucleosomesThemajorstructuresinDNAcompactionDNaseIHypersensitiveSites

WithinChromatinRegulationofGeneExpressioninEukaryotesisVeryComplexRNApolymeraseIsynthesizesapre-rRNA45S,whichmaturesinto28S,18Sand5.8SrRNAswhichwillformthemajorRNAsectionsoftheribosome.RNApolymeraseIIsynthesizesprecursorsofmRNAsandmostsnRNAandmicroRNAs.RNApolymeraseIIIsynthesizestRNAs,rRNA5SandothersmallRNAs.RNAPolymeraseIIandTypeIIPromotorexon1intron1exonnintronnUpstreamDownstreamInitiationenhancerSilencerPromoterTATAboxGCboxCAATboxenhancerTATAbox(TATAAAA)isat-25andisinvolvedinpositioningtheenzymeforcorrectinitiation.GCboxisat-90containsthesequenceGGGCGGandisrecognizedbythefactorSP1.

CAATbox(CCAATCT)isat–75andisrecognizedbyalargegroupoftranscriptionfactorsandplaysastrongroleindeterminingtheefficiencyofthepromoter.RNAPolyIITranscriptionInitiationMachineryRNApolyIIholoenzymeisrecruitedtothepromotersofprotein-codinggenes.ItconsistsofRNApolyII,asubsetofgeneraltranscriptionfactors,andregulatoryproteins.Partoftheassemblyoftheholoenzymeisreferredtoasthepreinitiationcomplex,becauseitsassemblytakesplaceonthegenepromoterbeforetheinitiationoftranscription.TranscriptionunitsforRNApolymeraseIhaveacorepromoterseparatedby-70bpfromtheupstreampromoterelement.UBFbindingtotheUCEincreasestheabilityofcore-bindingfactortobindtothecorepromoter.Core-bindingfactor(SL1)positionsRNApolymeraseIatthestartpoint.RNAPolymeraseITranscriptionUnitsforRNAPolymeraseIIIPromotersforRNApolymeraseIIImayconsistofbipartitesequencesdownstreamofthestartpoint.withboxAseparatedfromeitherboxCorboxB,ortheymayconsistofseparatedsequencesupstreamofthestartpoint(Oct,P5E,TATA).tRNAEpigeneticsEpigeneticsareinheritedandreversiblemodificationstonucleotidesorchromosomesthatdonotchangetheDNAsequence.butcanaltergeneexpression.DNAMethylation,HistoneModification,noncodingRNAWhatIsDNAMethylation?DNAmethylationisthecovalentaddition

ofa

methylgroup

(CH3)tocytosine

withinthecontextofCpGdinucleotide

.WhatIsCpGIsland?CpGislandsoftenarehighlyenrichedatgenepromoters.About70%ofhumanpromotershaveahighCpGcontent.

MostofthemethylationoccursashortdistancefromtheCpGislands(CpGislandshores)ratherthanintheislandsthemselves.MechanismofDNAMethylationandDemethylationDNAcytosinemethylation,hydroxylation,anddemethylation.(A)ThequestionmarkindicatespossibleactivityofDNAdemethylases.(B)Conversionof5mCto5hmCinmammalianDNAbytheMLLfusionpartnerTET1.(C)Itiscurrentlyunknownwhether5hmCisanendproductoranintermediateinactiveDNAdemethylation.ThequestionmarkindicatesapossibleMTase-assistedremovaloftheC5-boundhydroxymethylgroup.Tet：Ten-eleventranslocation5mC:5-methylcytosine5hmC:5-hydroxymethylcytosineDNAmethylationinvolvestheadditionofamethylgrouptothe5positionofthecytosinepyrimidinering.DNAMethyltransferasesDnmt3b,ofwhichexpressionisstage-andcelltype-specific,canmethylatenucleosomecoreregion,andmaycontributetoglobalmethylation.ChromatinremodelingthatpromotesthecreationofnakedDNAmayregulateDnmt3a-dependentmethylation.Dnmt3L,whichhasnoDNAmethylationactivity,isshowntobeindispensablefortheglobalmethylationingermcells.Dnmt3LdirectlyinteractswithDnmt3aandDnmt3btofacilitatetheiractivity.WhatAretheResultsofDNAMethylation?DNAmethylationcancausegenesilencingbyinterferencewithrecognitionandbindingofpositivelyactingtranscriptionfactorsPromotionofinteractionandrecruitmentofnegativefactors.DNAMethylationAndDiseasesCREB:cAMP-responseelement-bindingproteinWSTF:WilliamssyndrometranscriptionfactorSmallncRNAs:~18–31nt,siRNAs,miRNAs,piRNAs,inthecellcytoplasm.MediumncRNAs:~31–2000nt,snRNAs,snoRNAs,resideinthecellnucleus.LongncRNAs:from200ntuptoseveralhundredkbHousekeepingncRNAs:ribosomal,transfer,snRNAsandsnoRNAs,playingcrucialrolesinmanycellularprocesses,RegulatoryncRNAs:miRNAs,piRNAs,siRNAs,lncRNAs,promoter-associatedRNAs(PAARs),enhancerRNAs(eRNAs).Non-CodingRNAmiRNAsmiRNAregulategenesviaimperfectcomplementarityto3’UTRofgeneCleavedbyDicerfrom70-100ntpre-miRNAhairpinincytosolCanbeexpressedinatissuespecificfashion~1/3Resideinsideintrons~2/3independenttranscriptionunitsOfteninclusters.Manytimesnearthegenestheyregulateorinsidethem.ThebiogenesisandfunctionofmiRNAsmiRNAsInvolvedInCellDeterminationlncRNAs’sCharacteristicsResideinthenucleusLowlevelofsequenceconservationIntronicorintergenicGuidecomplexestotherightspotsinthegenomeModelingofnucleararchitectureJunk:somesequencesaretranscribedunnecessarilyScaffold:keepstheproteinstogetherByproductRNAInterference,siRNARNAiisanRNA-dependentgenesilencingprocessthatiscontrolledbytheRNA-inducedsilencingcomplex(RISC)andisinitiatedbyshortdouble-strandedRNAmoleculesinacell'scytoplasm,wheretheyinteractwiththecatalyticRISCcomponentargonaute.TwotypesofsmallRNAmolecules–microRNAandsmallinterferingRNA–arecentraltoRNAinterference.WhenthedsRNAisexogenousorendogenous,theRNAisimporteddirectlyintothecytoplasmandcleavedtoshortfragmentsbytheenzymeDicer.HistonesaresubjecttoposttranslationalmodificationbyenzymesprimarilyontheirN-terminaltails,butalsointheirglobulardomains.Suchmodificationsincludemethylation,acetylation,phosphorylation,ubiquitination,SUMOylation,andADP-ribosylation.Thisaffectstheirfunctionofgeneregulation.HistoneModificationThenucleosomecoreparticleconsistsofapproximately147basepairsofDNAwrappedin1.67left-handedsuperhelicalturnsaroundahistoneoctamerconsistingof2copieseachofthecorehistonesH2A,H2B,H3,andH4.HistoneAcetylationdescribesareactionthatintroducesan

acetyl

functionalgroup

intoahistonebyhistoneacetyltransferase

(HAT),andHistone

Deacetylation

istheremovaloftheacetylgroupby

histonedeacetylase

(HDAC).

Histoneacetylationislinkedto

transcriptional

activationandassociatedwith

euchromatin.Histonemethylation

isaprocessbywhichmethylgroupsaretransferredto

aminoacids

of

histone

byHistonemethyltransferases.Methylatedhistonescaneitherrepressoractivatetranscription.HistoneDemethylationisdonebyhistonedemethylase.HistonePhosphorylation

istheadditionofa

phosphate

(PO43?)grouptoa

histonebykinaseandHistoneDephosphortlationisdonebyphosphatases.UbiquitilytionUbiquitinationisanenzymaticprocessinwhichtheCterminalglycineintheactivatedubiquitinformsanamidebondtothelysineinthemodifiedprotein.E1:ubiquitin-activatingenzymeE2:ubiquitin-conjugatingenzymeE3:ubiquitin-proteinligases

DegradationoftargetproteinChangesofstructureandactivityoftargetproteinAntigenprocessing,Apoptosis,Biogenesisoforganelles,Cellcycleanddivision,DNAtranscriptionandrepair,Differentiationanddevelopment,Immuneresponseandinflammation,Neuralandmusculardegeneration,Morphogenesisofneuralnetworks,Modulationofcellsurfacereceptors,Responsetostressandextracellularmodulators,Ribosomebiogenesis,Viralinfection.HistoneCodeThehistonecodeisahypothesis