分子生物學教學課件：Chapter 18 Regulatory RNAs

Chapter18RegulatoryRNAs1RegulationbyRNAsinBacteria.2RNAInterferenceIsaMajorRegulatoryMechanisminEukaryotes.3SynthesisandfunctionofmiRNAmolecules.4TheEvolutionandExploitationofRNAi.5RegulatoryRNAsandX-inactivation.PART1RegulationbyRNAsinBacteriaSmallRNAs(sRNA):

regulationbybasepairing.Riboswitches:

regulationbymetabolite-mediatedstructurechangesAttenuation:

Regulationbyribosomestop-mediatedformationofterminatorsSmallRNAs(sRNA)Targets:

translationinitiationandtranscriptionterminationMechanism:

RegulationbybasepairingwiththetargetedsequencesonmRNA2.RiboswitchesTargets:areregulatoryRNAelementsthatactasdirectsensorsofsmallmoleculemetabolitestocontrolgenetranscriptionortranslationRegulatetranslationinitiationandtranscriptionterminationbyalteringtheaccessibilityofRBSandtheformationofterminator,respectively.ResideupstreamofthetargetedmRNA,andformspecificstructuretobinditssmallmoleculeligandActincisbyalterationofitsownstructureuponthebindingofthesmallmetabolites.MechanismFigure

18-2.ThestructureofariboswitchinitsregulatedmRNA代謝物代謝物Figure18-3a.Controloftranslationinitiationbyariboswitch.Figure

18-4.AlterationofofthestructureoftheSAMriboswitchuponthebindingofSAM(S-adenosylmethionine)10The2ndstructuresof7riboswitchesandmetabolitesthattheysense3.Attenuation(衰減作用)Target:AprematuretranscriptionterminationMechanismRequirestheparticipationofribosomesthattranslatealeaderpeptide.Theprematuretranscriptionterminationistriggeredbyformationofanintrinsicterminatorwhenribosomereadthroughcodonsoftheaminoacidthattheoperonsynthesizes.Transcriptionofthetrpoperonisprematurallystoppedifthetryptophanlevelisnotlowenough,whichresultsintheproductionofaleaderRNAof161nt.13LowTrpHighTrpTranscriptionoftheleaderRNA.TranscriptionofthetrpoperonmRNA.RNAPolPART2RNAinterferenceanditsmechanismDouble-strandedRNAinhibitsexpressionofgeneshomologoustothatRNA.2006年的諾貝爾生理學獎獲得者：MechanismClassify:

siRNAmiRNA

ThreewaysoftheRNAi-directedgenesilencingTriggerdestructionofthetargetmRNAInhibittranslationofthetargetmRNAInducechromatinmodificationTheheartoftheRNAimechanismDicer:

anRNaseIII-likemultidomainribonucleasethatfirstprocessesinputdsRNAintosmallfragmentscalledshortinterferingRNAs(siRNAs)ormicroRNAs(miRNA).DicerthenhelpsloaditssmallRNAproductsintoRISC.RISC

(RNAinducedsilencingcomplexes)alargemultiproteincomplexthatdirecttheboundsiRNAormiRNAtoitstargetandinhibitthetargetgeneexpression.18ThecrystalstructureoftheGiardiaintactDicerenzymeshowsthatthePAZdomain,amodulethatbindstheendofdsRNA,isseparatedfromthetwocatalyticRNaseIIIdomainsbyaflat,positivelychargedsurface.

RISC:thekeycomponentisArgonaute(AGO)Argonaute(AGO):AlargeproteinfamilythatconstituteskeycomponentsofRISCs.---AGOproteinsarecharacterizedbytwouniquedomains,PAZandPIWIPAZandPIWIdomainsarebothessentialtoguidetheinteractionbetweenthesiRNAandthetargetmRNAforcleavageortranslationalrepression.DistinctAGOmembershavedistinctfunctions.Forexample,humanAGO2programsRISCstocleavethemRNAtarget,whereasAGO1andAGO3donot.20Topic3:miRNAbiogenesisandregulation1.MicroRNA(miRNA)&itsprocessingMicroRNA(miRNA):

Atypeofnon-codingsmallRNA(~21–23nucleotides)producedbyDicerfromastem-loopstructuredRNAprecursor(~70-90ntsong).miRNAsarewidelyexpressedinanimalandplantcellsandfunctionsintheformofRNA–proteincomplexes,termedmiRISCs.miRNAshavebeenimplicatedinthecontrolofdevelopmentbecausetheyleadtothedestructionortranslationalsuppressionoftargetmRNAswithhomologytothemiRNA23ThemiRNAgenesandStructureofpri-miRNAsPri-miRNAsbearthe5’capand3’poly(A)tails25Hanetal.,Cell125,887–901,June2,200626miRNAprocessingPri-miRNA(miRNA初級轉錄產物)Drosha(1)pre-miRNA(miRNA前體)Dicer(2)miRNAExportin5(Exp5)transportspre-miRNAtothecytoplasm27ThecrystalstructureoftheGiardiaintactDicerenzymeshowsthatthePAZdomain,amodulethatbindstheendofdsRNA,isseparatedfromthetwocatalyticRNaseIIIdomainsbyaflat,positivelychargedsurface.

RISC:

guideRNAArgonautRdRPRISC30Eulalioetal.Ce1l132,2008MechanismsofmiRNA-MediatedTranslationalInhibitionInhibitionoftranslationelongation:miRNAsrepresstranslationoftargetmRNAsbyblockingtranslationelongationorbypromotingprematuredissociationofribosomes(ribosomedrop-off)(B)Co-translationalproteindegradation:Theproteinisnormallytranslatedafterwhichitisimmediatelydegradedproteolytically.(C)Competitionforthecapstructure:ArgonauteproteinscompetewitheIF4Eforbindingtothecapstructure.(D)Inhibitionofribosomalsubunitjoining:ArgonauteproteinsrecruiteIF6,whichpreventsthelargeribosomalsubunitfromjoiningthesmallsubunit.32TranscriptionalGeneSilencingbyDirectingChromatinModificationShortint