生化郑利民信号转导－生化课

生化郑利民信号转导－生化课第1页/共74页2Signal-TransductionPathways

信号转导通路(1)

第2页/共74页3干细胞的分化第3页/共74页4植物的生长发育是在环境因子的影响下正确进行时空表达的过程第4页/共74页5信号受体反应:手触摸就是刺激(信号)，小叶合拢就是反应。偶联刺激到反应之间的生化和分子途径,就是这个反应的信号转导通路

触摸含羞草后,小叶合拢第5页/共74页6细胞信号转导网络的简单模式（信号输入）（信号输出）第6页/共74页7ImportantrolesofbiosignalingFunctionalintegrationofdistantorgans,tissuesandcellsrequirescommunication;Signalingisperhapsaprimalrequirementtorespondtoourenvironment;Thefoundationofanycomplexresponsepathwaylieswithcellularbiochemicals.BiosignalingIntercellular（细胞间）&Intracellular（细胞内）第7页/共74页8常见四种类型：Endocrine(内分泌）Paracrine(旁分泌）Autocrine(自分泌）MembraneattachedproteinIntercellularsignaling

（细胞间信号）第8页/共74页9Fourschemesofintercellularsignaling(1)第9页/共74页10Fourschemesofintercellularsignaling(2)第10页/共74页11Intracellularsignaling

（细胞内信号）第11页/共74页12IntracellularReceptor第12页/共74页13Electron-micrographofmacrophage(pink)attacking

Escherichiacoli(green)第13页/共74页14M吞噬处理入侵细菌及提呈抗原的机制第14页/共74页15SignalTransductionPathway:Complicated第15页/共74页16FcR

CR3Ca++srcPI3kPKCMAPKRhoGTPasegelsolinArp2/3PLCPLDActinrearrangementPhagocytosis;OxidativeactivationSignals

forphagocytosisSignalReceptorAmplificationTransductionResponsessecondmessengers第16页/共74页17信号转导要素：信号或配体,受体,信号放大(产生第二信使),应答和反馈调节胞外 质膜 胞内第17页/共74页18PARTⅠ1Basiccharacteristicsofsignaltransduction2FourgeneraltypesofsignaltransducersPARTⅡ1Regulatorymechanisms2Somediseasescausedbydefectsinthebiosignalingpathways第18页/共74页191Fourbasiccharacteristics:1.1Specificity1.2Amplification1.3Desensitization/Adaptation1.4Integration第19页/共74页20SpecificitySignalmoleculefitsbindingsiteonitscomplementaryreceptor;othersignalsdonotfit.thrombin第20页/共74页21Scatchardanalysisquantifiesthereceptor-ligandinteraction第21页/共74页22oftenshort-lived&lowconcentration第22页/共74页23Desensitization/Adaptation

ReceptoractivationtriggersafeedbackcircuitthatshutsoffthereceptororremovesitfromthecellsurfaceProducearapidandmajorcellularresponsetoatransientsignal第23页/共74页24IntegrationWhentwosignalshaveoppositeeffectsonametaboliccharacteristicsuchasconcentrationofasecondmessengerX,orthemembranepotentialVm,theregulatoryoutcomeresultsfromtheintegratedinputfrombothreceptors第24页/共74页25细胞存活细胞凋亡Thebalancebetweenpro-andanti-apoptoticgenes/signalsdeterminethecellfate细胞接受到“死亡信号”，不一定就会死亡若同时也接受到“生存信号”，就可继续存活第25页/共74页26第26页/共74页272.Fourtypesofsignaltransducers2.1GatedIonChannels

2.1.1Ligand-gatedionchannels 2.1.2Voltage-gatedionchannels2.2ReceptorEnzymes2.3GProtein–CoupledReceptors andSecondMessengers2.4SteroidReceptors第27页/共74页28Fourgeneraltypesofsignaltransducers第28页/共74页29WhyIonChannels?Restingpotential:Asymmetricion-distributionActingpotentialGatedIonChannels Ligand-gatedionchannels Voltage-gatedionchannels第29页/共74页30Restingpotential第30页/共74页31IonConc.inMammalianCellsandSerum(mM）IonCytoplasmBloodSerumK+1404Na+12145Cl-4116proteincharges1389Mg+20.81.5Ca+2<0.00021.8WhyIonChannels:asymmetricion-distribution第31页/共74页32asymmetricion-distributionRestingpotential第32页/共74页33pumpandionleakchannelsCl

-leakchannel第33页/共74页34Actingpotential第34页/共74页35ActingpotentialVoltage-gatedNa+channels&K+channels第35页/共74页362.1.1Ligand-gatedionchannel:Bindingofsomesmallmoleculeforcesanallosterictransitioninprotein,open/closechannel.

acetylcholine(乙酰胆碱)receptorionchannel2.1.2Voltage-gatedionchannelAchargedproteindomainmovesrelativetothemembraneinresponsetoachangeintransmembraneelectricalpotential.

(voltage-gatedNa+，

Ca2+，K+channels)第36页/共74页37乙酰胆碱受体离子通道1AchACh第37页/共74页38BindingofAChtoRcauseconformationalchange.AsM2helicestwistslightly,theLeuresidues(yellow)rotateawayfromthechannelandarereplacedbysmallerpolarresidues(blue).Thisgatingmechanismopenschannel,allowingpassageofCa,Na,orK乙酰胆碱受体离子通道2Closed Open第38页/共74页39Voltage-gatedNa+channels1第39页/共74页40Voltage-gatedNa+channels2第40页/共74页412.Fourtypesofsignaltransducers2.1GatedIonChannels

2.1.1Ligand-gatedionchannels 2.1.2Voltage-gatedionchannels2.2ReceptorEnzymes2.3GProtein–CoupledReceptors andSecondMessengers2.4SteroidReceptors第41页/共74页42

2.2ReceptorenzymesAligand-bindingdomain(胞外)andanenzymeactivesiteoncytosolicside,connectedbyasingletransmembranesegment.CommonlyakinasethatphosphorylatesTyrresiduesinspecifictargetproteins（insulinreceptor）Other:synthesizethei.c.secondmessengercGMPinresponsetoex.c.signals

(thereceptorforatrialnatriureticfactor)第42页/共74页43Activationofreceptortyrosinekinases第43页/共74页44InsulinreceptortyrosinekinaseInsulinstructure第44页/共74页45Insulinreceptorbindsinsulinandundergoesautophosphorylationonitscarboxyl-terminalTyrresidues.InsulinreceptorphosphorylatesIRS-1onitsTyrresidues.SH2domainofGrb2bindstoP–TyrofIRS-1.SosbindstoGrb2,thentoRas,causingGDPreleaseandGTPbindingtoRas.ActivatedRasbindsandactivatesRaf-1.Raf-1phosphorylatesMEKontwoSerresidues,activatingit.MEKphosphorylatesERKonaThr&aTyrresidue,activatingit.ERKmovesintothenucleusandphosphorylatesNucleartranscriptionfactorssuchasElk1,activatingthem.PhosphorylatedElk1joinsSRFtostimulatethetranscriptionandtranslationofasetofgenesneededforcelldivision.第45页/共74页46Activationofglycogensynthasebyinsulin第46页/共74页47Regulationofbloodglucoselevel第47页/共74页48

ReceptorforatrialnatriureticfactorTwotypes(isozymes)ofguanylylcyclasethatparticipateinsignaltransduction.第48页/共74页492.Fourtypesofsignaltransducers2.1GatedIonChannels

2.1.1Ligand-gatedionchannels 2.1.2Voltage-gatedionchannels2.2ReceptorEnzymes2.3GProtein–CoupledReceptors andSecondMessengers2.4SteroidReceptors第49页/共74页502.3GPCRandSecondmessengersThreeessentialcomponents:1.aplasmamembranereceptorwithseventransmembranehelicalsegments2.anenzymeintheplasmamembranethatgeneratesanintracellular2ndmessenger3.aguanosinenucleotide–bindingprotein(Gprotein)第50页/共74页51ThreeessentialcomponentsofGProtein–CoupledReceptors第51页/共74页52第52页/共74页53AproteinbindsGuaninenucleotides(GDP,GTP);activatedinGTP-form,inactivatedinGDP-formIntegralmembraneprotein,heterotrimers();Havesimilar&subunits,butdifferin-subunitWhenG-proteinisactivated,thesubunitdissociatestointeractwithanenzymesthatgeneratesecondmessengers(e.g.cAMP)Others:smallG-proteins(~20-25kDa),e.g.Ras,Rho,Rac,etc,arenotmembranebound.Gprotein(GTP-bindingprotein)第53页/共74页54Gprotein(discovery)M.Rodbell:atransducerprovidedthelinkbetweenreceptorandamplifier.A.G.Gilman:identify&purifytheGprotein.System:MutatedlymphomacellscontaininganormalreceptorandcAMP-generatingenzyme,wasyetunabletorespond(producecAMP),sinceitlackedthetransducermutatedcellnormalcell第54页/共74页55NobelPrizeinPhysiologyandMedicine1994"fortheirdiscoveryofG-proteinsandtheroleoftheseproteinsinsignaltransductionincells"AlfredG.Gilman

1941-MartinRodbell

1925-1998第55页/共74页56“ON-OFF”switchisregulatedbyGTPorGDPboundform.AllG-proteinshasintrinsicGTPaseactivity,releasePiandinactivated.Activation:releaseofGDPandreplacedbyGTP第56页/共74页57Twomajorsystems:2.3.1THEPKASYSTEM (cAMPasthesecondmessenger)

The-AdrenergicReceptorSystem2.3.2THEPKCSYSTEM

(DAG,IP3andCa2+asthesecondmessengers)第57页/共74页58TheassociationofactiveGs

withadenylylcyclasestimulatesthecyclasetocatalyzecAMPsynthesisAdenosine3’,5’-cyclicmonophosphate(cAMP)第58页/共74页59synthesizedinadrenalmedulla;belongstocatecholamines(儿茶酚胺）;targetcellsincludeliver,skeletalmuscle,heartmuscleandadipose;releasedinresponsetoacutestressEpinephrine肾上腺素signal第59页/共74页60Epinephrine肾上腺素signalingpathwaycAMP第60页/共74页61Epinephrine肾上腺素signalingpathway(2)第61页/共74页62ActivationofcAMP-dependentproteinkinase(PKA)InactivePKA:Regulatory(R)subunits:auto-inhibitorydomainsburiedcatalytic(C)subunits:substrate-bindingsitesblockedbyRsubunitsRsubunits:autoinhibitorydomainsburiedActivePKACsubunitsopensubstratebindingsites第62页/共74页63AcatalyticsubunitofPKAATPPotentinhibitorpeptide(PKI):Arg-Arg-Gln-Ala-Ile(consensussequencerecognizedbyPKAexceptAla)第63页/共74页64Epinephrinetriggersaseriesofreactionsinhepatocytesinwhichcatalystsactivatecatalysts,resultingingreat“amplification”ofthesignalx分子10,000x分子第64页/共74页65PKAregulatesanumberofenzymesTheproteinsphosohorylatedbyPKAsharearegionofsequencesimilarityaroundtheSerorThrresiduethatundergoesphosphorylation,asequencethatmarksthemforregulationbyPKA.ThecatalyticsiteofPKAinteractswithseveralresiduesneartheThrorSerresidueinthetargetprotein,whichdefinethesubstratespecificity.第65页/共74页66第66页/共74页67DesensitizationofthePKAsystem1desensitizingβ-AdrenergicReceptor2degradingthesecondmessager第67页/共74页68GsbgrecruitsbARKtothemembrane,whereitphospho-

SerattheC-terminusofRecpt.barrbindstothepi-C-terminusofRecpt.Receptor-arrestincomplex