分子生物學(xué)真核轉(zhuǎn)錄

分子生物學(xué)真核轉(zhuǎn)錄第1頁，課件共34頁，創(chuàng)作于2023年2月1.

RNApolymerase:3types

RNApolymeraseI:innucleolus;rRNAtranscription;themostcellularRNAsynthesis.Didnotinhibitedbya-amanitin

RNApolymeraseII:innucleoplasm,heterogeneousnuclearRNA(hnRNA)transcription(theprecursorformRNA).inhibitedbylowconcentrationsofa-amanitin.

RNApolymeraseIII:aminorenzymeactivity.innucleoplasm;tRNAsandothersmallRNAs(5SrRNA)transcription.inhibitedornotbya-amanitin.

AlleukaryoticRNApolymerasesarelargeproteins,>500kD,8-14subunits.

第2頁，課件共34頁，創(chuàng)作于2023年2月RNApolymerasesIIThelargestsubunithasacarboxy-terminaldomain(CTD),whichconsistsofmultiplerepeatsofaconsensussequenceof7aminoacids.ThesequenceisuniquetoRNApolymeraseII.

TheRNApolymeraseactivitiesofmitochondriaandchloroplastsaresmaller,andresemblebacterialRNApolymerase.第3頁，課件共34頁，創(chuàng)作于2023年2月2.Promoter

第4頁，課件共34頁，創(chuàng)作于2023年2月

RNApolymeraseIpromoterhastwosequencecomponentsCorepromoter:~70bp.UBF1:asinglepolypeptide,itbindstobothregions(GC-rich),afterwhichSL1canbind.RNApolIthenbindstothecorepromoter.UBF1:

SL1:4subunits,onesubunit--TBPinteractswithRNApolymerase.SL1resemblessigmafactor.第5頁，課件共34頁，創(chuàng)作于2023年2月Deletionanalysisshowsthatthepromoterfor5SRNAgenesisinternal;initiationoccursafixeddistance(~55bp)upstreamofthepromoter.

RNApolymeraseIIIhasainternalpromoter

第6頁，課件共34頁，創(chuàng)作于2023年2月

RNApolymeraseIIIhasainternalpromoter

PromotersforRNApolymeraseIIIconsistofbipartitesequences:boxAseparatedfromeitherboxCorboxB.Ortheymayconsistofseparatedsequencesupstreamofthestartpoint(Oct,PSE,TATA).第7頁，課件共34頁，創(chuàng)作于2023年2月PromoterstructureofRNApolymeraseIIIThebindingoftheassemblyfactorsTFIIIAandTFIIIC,theinitiationfactorTFIIIB,andRNApolymeraseIII.第8頁，課件共34頁，創(chuàng)作于2023年2月PromoterstructureofRNApolymeraseIIITFIIIA:azincfinger.TFIIIB:threesubunits.TFIIIC:atleast5subunits,>500kD.TFIIIAandTFIIICareassemblyfactors,whoseroleistoassistthebindingofTFIIIBattherightlocation.TFIIIBfunctionsasapositioningfactor,responsibleforlocalizingRNApolymerasecorrectly.LikeSL1atthepolIpromoter,itresemblesasigmafactor.第9頁，課件共34頁，創(chuàng)作于2023年2月

PromoterstructureofRNApolymeraseII

Oneupstreampromoter,withrelativelyfixedlocation.TATAbox:~25bpupstream,8bpconsensussequenceconsistsentirelyofAT.TheTATAboxtendstobesurroundedbyGC-richsequences.Itisalmostidenticalwiththe-10sequencefoundinbacterialpromoters.

TheminorityofpromotersthatdonotcontainaTATAelementarecalled

TATA-lesspromoters.

TBP:TATA-bindingprotein,~30kD.TAFs:forTBP-associatedfactors.

第10頁，課件共34頁，創(chuàng)作于2023年2月Theassemblesatthepromoter

第11頁，課件共34頁，創(chuàng)作于2023年2月PromotersforRNApolymeraseIIhaveshortsequenceelements

CAATbox:-75,CAAT.acommonelements,functionsineitherorientation.Toincreasespromoterstrength.

GCbox:-90,GGGCGG.Oftenmultiplecopies.ineitherorientation.arelativelycommonpromotercomponent.

第12頁，課件共34頁，創(chuàng)作于2023年2月第13頁，課件共34頁，創(chuàng)作于2023年2月PromotersforRNApolymeraseIIhaveshortsequenceelements

第14頁，課件共34頁，創(chuàng)作于2023年2月PromotersforRNApolymeraseIIhaveshortsequenceelements

第15頁，課件共34頁，創(chuàng)作于2023年2月3.Enhancerbidirectionalelementsthatassistinitiationfunctionineitherorientationandinanylocation(upstreamordownstream)relativetothepromoter

AnenhancerinthevirusSV40

Itcontainstwoidenticalsequencesof72bpeach,repeatedintandem~200bpupstreamofthestartpointofatranscriptionunit.Each72bprepeatcontainsacopyoftheenhancer

第16頁，課件共34頁，創(chuàng)作于2023年2月4.RNAprocessing第17頁，課件共34頁，創(chuàng)作于2023年2月

mRNAprocessing5’Capping3’CleavageandpolyadenylationSplicingPre-mRNAmethylationmRNAediting第18頁，課件共34頁，創(chuàng)作于2023年2月Splicing

第19頁，課件共34頁，創(chuàng)作于2023年2月Splicing

TheendsofnuclearintronsaredefinedbytheGT-AGrule.(GT-AGrule).第20頁，課件共34頁，創(chuàng)作于2023年2月第21頁，課件共34頁，創(chuàng)作于2023年2月Splicingoccursintwostages,inwhichthe5’exonisseparatedandthenisjoinedtothe3’exon.

第22頁，課件共34頁，創(chuàng)作于2023年2月第23頁，課件共34頁，創(chuàng)作于2023年2月5’Cappingcap0:inalleukaryotes;guanine-7-methyltransferase.

cap1:thepredominanttypeofcapinalleukaryotesexceptunicellularorganisms;2’-O-methyl-transferase.cap2:<10-15%ofthetotalcappedpopulation.

第24頁，課件共34頁，創(chuàng)作于2023年2月Poly(A)tialadds~200Aresiduestothefree3′-OHendofthemRNA;catalyzedbytheenzymepoly(A)polymerasethepoly(A)-bindingproteins(PABP)bindsonpoly(A)tail,OnePABPmonomerof~70kDisboundevery10-20basesofthepoly(A)tail.Removalofthepoly(A)tailprecedesthedegradationofcertainmRNAs;Theabilityofthepoly(A)toprotectmRNAagainstdegradationrequiresbindingofthePABP.Removalofpoly(A)inhibitstheinitiationoftranslation.However,therearemanyexamplesinearlyembryonicdevelopmentwherepolyadenylationofaparticularmRNAiscorrelatedwithitstranslation.第25頁，課件共34頁，創(chuàng)作于2023年2月snRNA

(smallnuclearRNA)isanyoneofmanysmallRNAspeciesconfinedtothenucleus;severalofthesnRNAsareinvolvedinsplicingorotherRNAprocessingreactions.(100-300basesand105-106moleculespercell.Innaturalstate,theyexistasribonucleoproteinparticles(snRNP).ThespliceosomecontainssnRNA

Spliceosome:proteinsandRNAsexistedasribonucleoproteinparticles.ThesnRNPsinvolvedinsplicingareU1,U2,U5,U4,andU6.EachsnRNPcontainsasinglesnRNAandseveral(<20)proteins.TheU4andU6snRNPsareusuallyfoundasasingle(U4/U6)particle.ThesnRNPstogethercontain~40individualproteins.

第26頁，課件共34頁，創(chuàng)作于2023年2月第27頁，課