基因表达调控

RegulationofGeneExpression

Introduction:LivingorganismsrequiresthepreciseandtimelyregulationofgeneexpressionItisthecapacitytoswitchgeneonandoffthatenablescellstorespondefficientlytochangingenvironmentcyclinG0G1SG2MG1SG2MG0DEABInmulticellularorganisms,complicatedprogrammedpatternsofgeneexpressionareresponsibleforcelldifferentiationaswellasintercellularcooperation.I.Theconceptsandprinciplesofgeneregulation1-1conceptofgeneexpression:theprocessesofthegeneticinformationinDNAconvertedintofunctionalproteins.Theinformationflowincludestranscriptionandtranslation.ThebiosynthesisoftRNAandrRNAarealsobelongtotheprocessesofgeneexpression.PathwayforGeneExpression

(theCentralDogma)1-2:

Specificityofgeneexpression

Ofthe3,000geneinthetypicalbacterialgenomeor>100,000genesinthehumangenome,onlyafractionareexpressedatanygiventime.Temporalspecificity:requirementsforagivengeneproductmaychangewithtimeorfollowthetimesequences.Duringdevelopmentinamulticellulareukaryote,thegeneexpressionofsomeproteinsthatinfluencecellulardifferentiationopenorcloseobeydelicatedorder,thistemporalspecificityinmulticellulareukaryotealsowasnamedstagespecificity.Spatialspecificity:Agivengenehasdifferentexpressionlevelsindifferenttissuesoronlyexpressedinspecifictissuesororgans.Thisspecificitywasalsocalledcellortissuespecificity.Forexample,liver-specificgenearenottranscribedbybrainorkidneycells1-3:typesofgeneexpression

Housekeepinggene:somegeneproductsarerequiredallthetimeandtheirgenesareexpressedataconstantlevelinallthecellsofaspeciesororganism.Thesegenesareoftenreferredtobeashousekeepinggenes.Manyofthegenesforenzymesthatcatalyzestepsincentralmetabolicpathwayssuchasthecitricacidcyclearehousekeepinggenes.Constitutivegeneexpression:constant,seeminglyunregulatedexpressionofhousekeepinggeneiscalledConstitutivegeneexpression.Induction:geneproductsriseinresponsetomolecularsignalsarereferredtoasinducible,andtheprocessofincreasingtheexpressionofthegeneiscalledinduction.TheexpressionofmanygenesencodingDNArepairenzymes,forexample,isinducedinresponsetoDNAdamage.Repression:geneproductsthatdecreaseinconcentrationinresponsetoamolecularsignalsarereferredtobeasrepressible,andthedecreaseingeneexpressioniscalledrepression.Forexample,thepresenceofamplesuppliesoftryptophanleadstorepressionofthegenesfortheenzymescatalyzingtryptophanbiosynthesisinbacteria.Coordinateexpression:onegroupgeneproductsinonemetabolicpathwayareexpressedinoptimalratioofconcentrationinresponsetotheenvironmentsignal.Thisco-expressionandcoordinateregulationofmultiplegenesiscalledcoordinateexpression.mRNAshavemanyribosomebindingsitesinprokaryotes1-4:Significanceoftheregulationofgeneexpression

AdaptationofenvironmentchangeMaintainingthegrowthandproliferationofcellsinprokaryotes.Maintainingcellulardifferentiationandindividualdevelopmentineukaryotes.II.TranscriptionregulationofprokaryoticgenesIntroduction:

Prokaryotesinresponsetosuddenenvironmentalchangestakeonlyminutes,becausetranscriptionandtranslationinprokaryotesarecloselycoupled:Ribosomescommencetranslationnearthe5´endofanascentmRNAsoonafteritisextrudedfromRNApolymerase.Prokaryoteshaveasimplegeneralmechanismforcoordinatingtheregulationofgenes:thegenesareclusteredonthechromosomeandtranscribedtogether.MostprokaryoticmRNAsarepolycistronic.mRNAshavemanyribosomebindingsitesinprokaryotesThereareatleastsixpotentialpointsatwhichtheamountofproteincanberegulationbothinprokaryotesandeukaryotes:synthesisoftheprimaryRNAtranscriptposttranscriptionalprocessingofmRNAmRNAdegradationproteinsynthesis(translation)posttranslationalmodificationofproteinsproteindegradationofalltheprocesses,regulationattheleveloftranscriptioninitiationisthebestdocumentedandmaybethemostcommon.Asforallbiosyntheticpathways,themostefficientplaceforregulationisthefirstreactioninthepathway.Unnecessarybiosynthesiscanbehaltedbeforeenergyisinvested.2-1:

characteristicsofprokaryotictranscriptionThe

subunitoftheE.coliRNApolymeraseholoenzymeisaprokaryoticspecificfactorthatmediatesspecificpromoterrecognitionandbinding.

coreenzyme

holoenzyme

70:recognizetheinitiationsiteinmanygenes

32

:recognizeandinitiatethetranscriptionofheatshockproteingenesBoth

70

and

32belongtoheatshockprotein(Hsp)Different

isresponsibleforitsspecifictranscriptioninitiation.Themodelofoperonisthecommonmechanismofprokaryotictranscriptionregulation(detailsseebelow)

Repressorsandrepressor-operatorbindingarethecommonpatternsofregulationopenorcloseoftranscriptioninprokaryotes.Relatedconcepts:Repressorsbindtoapromoter,blockingaccessofRNApolymerasetothepromoterActivatorsbindnearapromoter,enhancingtheRNApolymerase-promoterinteractionThebindingsitesforrepressorsarecalledoperators.Operatorsitesaregenerallynearandoftenoverlapthepromoter.RNApolymerasesbindtoDNAandinitiatetranscriptionatspecificsitesintheDNAcalledpromoters.TheregulationoftranscriptioninitiationistheregulationofinteractionofRNApolymerasewithitspromoter.E.colipromotershaveaconsensussequence.(seebelow)Thegeneareclusteronthechromosomeandtranscribedtogether.Thesinglepromoterinitiatetranscriptionofthecluster.Operon:Thegenecluster,thepromoter,andtheadditionalsequencesthatfunctioninregulationaretogethercalledanoperon;asegmentofDNAcontainingadjacentgenesincludingstructuralgenesandoperatorgeneandregulatorygene2-1:theLacoperon

In1960,FrancoisJacobandJacquesMonoddemonstratedthattwogenesinvolvedinlactosemetabolismwerecoordinatelyregulatedbyageneticelementlocatedadjacenttothem.Thetermsoperonandoperatorwerefirstintroducedinthispaper.2-2-1：Thestructureoflacoperon

PI—thepromoterfortheIgeneI—IgeneencodesaLacrepressorthatcausegenerepression.P—promoterO—operatorPI

Zgeneencodesβ-galactosidase,whichcleaveslactosetogalactoseandglucose

Ygeneencodesgalactosidepermease,whichtransportslactoseintothecell.

Ageneencodestransacetylase,whosephysiologicalfunctionisunknown.PI2-2-3：PositiveregulationofCAP

(catabolitegeneactivationprotein)

CAPisadimericprotein,whichhavetwobindingsite,oneforcAMP,anotherforCAP-cAMPbindingsiteinthepromoterofLacoperon.CAP-cAMPcomplex,butnotCAPitself,bindstotheLacoperonandstimulatestranscription.2-2-4：ThecoordinateregulationofrepressorandCAP2-3:otherregulationmechanism

2-3-1:trpoperonandtranscriptionattenuationThestructureandregulationoftrpoperon：tryptophanactsasacorepressor.

Attenuatorandattenuation

whentryptophanisabundant,theavailabilityoftryptophanresultsintheprematureterminationoftrpoperontranscription.Thecontrolelementresponsibleforthiseffectistermedanattenuator.Thistranscriptioncontrolmechanismnamedattenuation.Mechanismofattenuation:

Theattenuatortranscriptcontainsfourcomplementarysegmentsthatcanformoneoftwosetsofhairpins.segment3and4togethercompriseanormaltranscriptionterminator;segment2and3togethercompriseanti-terminator.

Whentryptophanisabundant,theribosomefollowscloselybehindtheRNApolymeraseandfastpassesthroughsegment1andpresentsonsegment2.Theformationofthebasepaired2·3hairpinwasblocked.The3·4hairpin,antranscriptionalterminator,cantherebyformthusabortingtranscription.

Viceversa,whentryptophanisinadequate,theribosomestallsonthetandemTrpcodonsofsegment1.Thissituationpermitstheformationofthe2·3hairpinwhich,inturn,precludestheformationofthe3·4hairpin.RNApolymerasethereforetranscribesthroughthisunformedterminatorandcontinuestrpoperontranscription.AttenuationinthetrpoperonEffectivelyaddsafinetuningtotheregulationofthetrpoperon.Severalkeypoints:Transcription&translationaretightlycoupledinbacteria(attenuationrequiresthis).If[Trp]isadequate，transcriptionisterminatedbeforethetrpoperon.If[Trp]isinadequate，transcriptioniscompleted.

Fiveotheraminoacids-biosynthesizingoperonsareknowntoberegulatedbyattenuation,suchasPhe,Leu,His,ThrThetandemaminoacidcodonsintheleadersequencearerichintheircorrespondingaminoacidresidues.Forexample,hisoperon,whichspecifiesenzymessynthesizinghistidine,hasseventandemHisresiduesinitsleaderpeptide.2-3-2:SOSresponse

AgentsthatdamageDNA,suchasUVradiation,alkylationagents,induceacomplicatedsystemofcellularchangesinE.coliknownastheSOSresponse.E.coliincreasetheircapacitytorepairdamageDNA,andceasedividing.Duringnormalgrowth,LexAlargelyrepressesSOSgeneexpression,whenDNAdamagehasbeenextensive,RecAisactivatedandstimulateLexAcleavage.TheLexA-repressiblegeneareconsequentlyreleasedfromrepressionanddirectthesynthesisofSOSproteins.III.Transcriptionregulationofeukaryoticgenomes

3-1.Thepropertiesofeukaryoticgenomes

EukaryoticgenomesareenormouslylargerandmorecomplicatethanthoseofprokaryotesE.coli:4×106bpcodefor~3,000genesHuman:2.9×109bpcodefor30,000~40,000unexpressedDNAinhumangenomeaccountsforabout90%~95%,encodedgenesonlyaccountfor5%.MonocistronThetranscriptofeukaryoticgeneismonocistron.OnegenetranscribeonemRNAandonemoleculeofmRNAtranslateonemoleculeofpolypeptide.ineukaryotes,mRNAshavesuchsequences

RepetitivesequencesViralandprokaryoticDNA(s)havefew,ifany,repeatedsequences.EukaryoteshavefourclassesofDNAs:uniquesequences(~1copy/haploidgenome)moderatelyrepetitivesequences(<106copies/haploidgenome)highlyrepeatssequences(>106copies/haploidgenome)invertedrepeats(plimentary)DNAG-quadruplexstructuresNatChem.2013Mar;5(3):182-6.

Eukaryoticgenesareinterspersedwithunexpressedregions.Pre-mRNAcontainsthetranscriptionofanentirestructuregene,includingitsintrons.Then,followingcappingandpolyadenylation,theintronsareexcisedandtheirflankingexonsareconnected,aprocesscalledgenesplicing,toyieldthematuremRNA.3-2.ThecharacteristicsofeukaryotictranscriptionGeneticexpressionineukaryotesismainlyregulatedthroughthecontroloftranscriptionalinitiation.

EukaryoticnucleicontainthreedistincttypesofRNApolymerasethatdifferintheRNAssynthesis:RNApolymeraseIsynthesizesprecursorsofmostribosomalRNAs(45S),whichisprocessedbyaseriesofcleavagestepstoproducethematurerRNA'sRNApolymeraseIIsynthesizesmRNAprecursors(hnRNA)RNApolymeraseIIIsynthesizesprecursorsof5srRNA,thetRNAsandsmallnuclearRNAsRNApolymerase RNAsynthesisedI rRNA45SII hnRNAIII tRNA,5SrRNA,snRNAInprokaryotes,

OneRNApolymerasecatalysesallRNAsynthesisTypesofRNA-polineukaryotesEukaryoticRNApolymerase(500~700kD)arecharacterizedbysubunitcompositionofcomplexity.Eachenzymecontainstwononidentical“large”(>100kD)subunitandanarrayofupto12differentsmall(<50kD)subunits.RNApolymeraseIIlacksabilitytobindtoitspromoters.Transcriptionalinitiationismediatedbycell-specificfactorsGeneraltranscriptionfactorsassociatedwithRNAPolIIinhumancells

*TATABindingProtein=TBP33helicaseTATAboxTranscriptionalactivechromatinisstructurallydistinctanincreasedsensitivityoftheDNAtonuclease-mediateddegradation.ManyhypersensitivesitescorrespondtobindingsitesforknownregulatoryproteinstheDNAintranscriptionallyactivechromatintendstobeundermethylated.transcriptionallyactivechromatintendstobedeficientinhistoneH1;othercorehistoneshaveatendencytobemodifiedbyacetylation.Topologicalstatechange:thenegativesupercoilingofnaturallyoccurringDNAsresultsinatorsionalstrainthatpromotestopologicalstatechangefromnegativetopositivesupercoiling.Mosteukaryoticpromotersarepositivelyregulated.onewaytoimprovespecificityistousemultipleregulatoryprotein,ifseveralpositiveregulatoryproteinsmusteachbindspecificDNAsequence.OneofnegativebindingissufficienttoadequatelyblockRNApolymeraseaction.Positiveregulationinalargegenomeissimplythatitismoreefficient.Eukaryoteshaveanuclearmembranethatseparatestheirchromosomefromtheircytoplasm,therebyphysicallydivorcingthetranscriptionalprocessfromthattranslation.

posttranscriptionalprocessesexonaresplicedmRNAsarecappedmRNAshavepoly(A)tailsIV.RegulatoryfactorsforeukaryotictranscriptioninitiationactivatorsandrepressorsofeukaryoticgenetranscriptionrecognizespecificcontrolsequenceintheDNAtowhichtheybindcalledcis-actingelements.Theproteintranscriptionfactorsthatbindtothesecis-elementsareknowastrans-actingfactor.PairsofDNAsequences,whichareonthesameDNAmolecule,aresaidtobeincis(Latin:ontheside)whilethoseondifferentDNAmoleculesaresaidtobeintrans(Latin:across).4-1.Cis-actingelements

promoterEukaryoticpromoterarefarmorecomplexthanthoseforprokaryotes.TATAbox(consensussequenceTATAAAA):located25to30basepairsfromthemRNAinitiationsite.ThesesequenceappeartobebindingsitesforatranscriptionfactorTFIIDthatisrequiredforRNApolymeraseIIbinding.Itcontrolstheveracityandfrequencyoftranscriptionalinitiation

GCbox:Sequence:GGGCGGLocation:-30~-110bpFunction:ItisthebindingsiteforaproteincalledSp1.housekeepinggenehaveoneormorecopiesofthesequencelocatedupstreamfromtheirtranscriptionstartsites.CCAATboxSequence:GCCAATLocation:–50~–110Function:ItisthebindingsiteforCTF1(CAAT-bindingtranscriptionfactor)andC/EBP(enhancerbindingprotein).Enhancersaretranscriptionalactivatorsthatcanhavevariablepositionandorientations.Enhancer:itisaDNAsequencethatcandeterminethetemporalandspatialspecificitiesofexpressionandincreasethepromoteractivity.Enhancersmediatemuchoftheselectivegeneexpressionineukaryotes.Silencer:Itisanegativeregulationelement.Itwillrepressthetranscriptiononceinteractedwithspecificproteins.4-2.Trans-actingfactors

Inmostcases,thetranscriptionregulatoryproteins(transcriptionfactor,TF)ineukaryotesthatbindtoenhancerorpromoterareactivatorproteins.TherearetwotypesofTF:Generaltranscriptionfactor:Thetaskperformedby