Cellsignalingcanaffectvirtuallyeveryaspectofcellstructureandfunction:

Activationofenzymeactivity;

Changeincytoskeletalorganization;

Changeinionpermeability;

InitiationofDNAsynthesis;

Activationorrepressionofgeneexpression.Chapter8

CellSignaling1.OverviewofintercellularsignalingGeneralschemesofintercellularsignaling(1)EachcellisprogrammedtorespondtospecificcombinationsofextracellularsignalmoleculesThecellintegratesdifferentinformationandmountsanappropriateandcomprehensiveresponse.Thesesignalmoleculesworkincombinationstoregulatethebehaviorofthecell.CellsrespondtostimuliviacellsignalingA.Someofthebasiccharacteristicsofcellsignaling

(2)SignalingpathwaysconsistofaseriesofstepssignalmagnificationRecognitionofthestimulusbyaspecificplasmamembranereceptor.Transferofasignalacrosstheplasmamembrane.Signalmagnification:Transmissionofthesignaltoeffectormoleculeswithinthecell,whichcausesachangeincellularactivities.Cessationofthecellularresponseduetoinactivationofthesignalmolecule.

(3)Fourfeaturesofsignal-transducingsystems(4)DifferentcellscanresponddifferentlytothesameextracellularsignalmoleculeVariousresponsesinducedbyneurotramsmitteracetylcholine(5)Acellcanremembertheeffectofsomesignals,afterthesignalhasdisappeared.(Ca2+)ProteinkinaseactivitedbyCa2+to

phosphorylateitselfandotherproteins,theautophosphorylationkeepsthekinaseactivelongafterCa2+levelsreturntonormal.Transientextracellularsignalsofteninducemuchlonger-termchangesincellsduringthedevelopmentofamulticellularorganism.Theyusuallydependonself-activatingmemorymechanismsthatoperatefurtherdownstreaminasignalingpathway,atthelevelofgenetranscription.B.SignalMoleculesandReceptors(1)Signalmolecules:(2)Receptorsincludetwoclasses:

Thefunctionsofreceptors:Receptor-mediatedendocytosis;Signaltransduction:1.Theactivationofreceptormakeforthecascades;2.Thedesensitizationofreceptormakeforclosedownofthecascades;3.Thedown-regulationofreceptormakeforthedepressedreaction;

Differentkindsofintracellularsignalingproteinsalongasignalingpathwayfromasurfacereceptortothenucleus.

Relayproteinssimplypassthemessagetothenextsignalingcomponentinthechain.Messengerproteinscarrythesignalfromoneparttoanother(fromcytosoltonucleus).Adaptor

proteins

linkonesignalingproteinstoanotherAmplifier

proteins,(enzymesorionchannels)signalingcascade.Transducer

proteins

convertthesignalintoadfferentform.Bifurcation(分歧)proteinsspreadthesignalfromonesignalingpathwaytoanother.Integrator

proteins

receivesignalsfromtwoormoresignalingpathwaysandintegratethembeforerelayingasignalonward.Latentgeneregulatory

proteins

areactivatedatthecellsurfacebyactivatedreceptorsandthenmigratetothenucleustostimulategenetrascription.Mostactivatedcell-surfacereceptorsrelaysignalsviasmallmolecules(secondmessengers)andanetworkofintracellularsignalingproteins.Twotypesofintracellularsignalingcomplexes,enhancethespeed,efficiency,andspecificityoftheresponse.InteractionsbetweenintracellularsignalingproteinsaremediatedbymodularbindingdomainsPHdomain:Pleckstrin:血小板、白細胞C激酶底物同源域Bcellactivation(3)FourgeneraltypesofsignaltransducersC.TwotypesofintracellularsignalingproteinsthatactasMolecularSwitches.Phosphorylationanddephosphorylationviaproteinkinasesandphosphatases.TherebystimulatingorinhibitingtheactivitiesGAPsinactivateG-protein;GEFsactivatesG-protein;GDIs(guaninenucleotide-dissociationinhibitors)maintaintheG-proteininactive.GEF:guanine-nucleotide-exchangefactorGRF:guanine-nucleotidereleasefactor,GAP:GTPase-activatingproteinTheNobelPrizeinPhysiologyorMedicine1992“fortheirdiscoveriesconcerningreversibleproteinphosphorylationasabiologicalregulatorymechanism”Signaltransductionpathwaysconsistprimarilyofproteinkinasesandproteinphosphatases.ThealterationsintheconformationofsignalingprteinsareusuallyaccomplishedbyproteinkinasesandproteinphosphatasesThehumangenomeencodesasmanyas2000differentproteinkinasesandmorethan300differentproteinphosphatases.TheNobelPrizeinPhysiologyorMedicine1994“fortheirdiscoveriesofG-proteinsandtheroleoftheseproteinsinsignaltransductionincells”G-proteinactivationandinactivationcycle(NIH.1970s)TheaccessoryproteinsforcyclingofG-proteinsGTPase-activatingproteins(GAPs):MostGproteinspossesssomecapabilitytohydrolyzeaGTP,butthiscapabilityisgreatlyacceleratedbyinteractionwithspecificGAPs.TherebyGAPsinactivateG-proteinbystimulatingittohydrolyzeitsboundGTP.Guaninenucleotide-exchangefactors(GEFs):AninactiveGproteinisconvertedtotheactiveformwhentheboundGDPisreplacedwithaGTP.GEFsareproteinsthatbindtoaninactiveGproteinandstimulatedissociationoftheboundGDP.TherebyGEFsactivatesG-protein.Guaninenucleotide-dissociationinhibitors(GDIs):GDIsareproteinsthatinhibitthereleaseofaboundGDPfromaG-protein,thusmaintainingG-proteinintheinactive,GDP-boundstate.2.SignaltransdutionmediatedbythereceptorswithincellsA.Nuclearreceptorsforsteroid,retinoicacid,vitaminDandthyroidhormones

B.NitricoxidegassignalsbybindingdirectlytoanenzymeinsidethetargetcellTwodiscovers:Duringthe1980s,bacteria-killingactivityofmacrophagesincellculturedependsonthepresenceofarginineinthemedium.arg.isasubstratefornitricoxidesynthasesNO(importantbiologicalmolecule).Ithasbeenknownformanyyearsthatacetylcholinedilatebloodvesselsbycausingtheirsmoothmusclestorelax.In1980,R.Furchgott(NYSMC)concludedthatbloodvesselsaredilatedbecausetheendothelialcellsproduceasignalmoleculethatmakessmoothmusclecellsrelax.In1986workbyFurchgottandparallelworkbyLouisIgnarro(UCLA)identifiedNOasthesignalthatcauserelaxationofthevascularsmoothmuscle.1998,ReceivedNobelPrize(R.Furchgott,L.Ignarro,F.Murad)TheactionofNitricoxideonbloodvesselsThemechanismbywhichacetylcholinestimulationoftheendothelialcellsleadstosmoothmusclerelaxationalsoexplainsthemechanismofactionofthechemicalnitroglycerin.Thedrugsildenafil,soldunderthetradenameViagra,isaninhibitorofacyclicGMP-specificphosphodiesterasethatnormallycatalyzesthebreakdownofcyclicGMP.3.Signaltransductionmediatedby

thereceptorsonthecellsurfaceA.MediatedbytheIon-LinkedReceptorswhichconvertchemicalsignalsintoelectricalones4or6-helixtransmembranereceptorThreestatesoftheacetylcholinereceptorB.SignaltransductionmediatedbyG

protein-linkedreceptorsThestructureofGprotein-linkedreceptors:Seven-helixtransmembrane;C-terminal:Ser-andThr-rich---thesitesofphosphorylationmakeforthedesensitizationofGPCRbyGRKGPCRsuperfamily:C.elegansgenome19000genes,encodesapproximately1000differentGPCRs..

Ligands:hormones,neurotransmitters,chemoattractants,odorantsandtastants,photonsThestructureandactivationofGproteinsActivationofGproteinbythereceptor.RelayofthesignalfromG-proteintoeffectorCyclicAMPsignalingpathwaySecondmessengers(cAMP),aneffector,amplifytheresponsetoasingleextracellularligandbycAMPtotriggerareactioncascade.ThecascadestartswiththebindingofcAMPtocAMP-dependentproteinkinaseA.PKAinhibitsglycogensynthaseandactivatesphosphorylasekinase.ThespecificityofGprotein-coupledresponsesDifferentligandssimilarmechanismtriggeringdifferentcellularresponses.Thereceptorforagivenligandcanexistinseveraldifferentisoforms,thathavedifferentaffinitiesfortheligandandforaparticularGprotein.9differentisoformsoftheadrenergicreceptor;15isoformsofthereceptorforserotonin(5-HT)Atleast20differentG,5differentGand12differentGsubunits;9isoformsoftheeffectoradenylylcyclase.DifferentcombinationsofspecificsubunitsconstrucctGproteinshavingdifferentcapabilitiesofreactingwithspecificisoformsofbothreceptorsandeffectors.ActivationofcAMP-dependentproteinkinase(PKA)bycAMPIP3-Ca2+pathwayandDG-PKCpathwayThefirstindicationthatphospholipidsmightbeinvolvedincellularresponsesemergedfromstudiescarriedoutintheearly1950sbyLowellandMabelHokinofMontrealGeneralHospitalandMcGrillUniversity.---TosdugytheeffectsofacetylcholineonRNAsynthesisinthepancreas.PHdomainshavebeenidentifiedinover150differentproteins.BindingofaproteintoaPIP2orPIP3messenger(remaininthelipidbilayer),recruitstheproteintothecytosolicfaceofthemembranes.ThepathwaythroughphospholipaseCresultsinariseinintracellularCa+

SignalsGPLRGPPLCIP3andDAG(twinsignals).IP3IP3receptor(Ca2+channel,locatedatthesurfaceofsER)ElevationofcytosolicCa2+;DAGactivatesPKCtophosphoralateSerandThrontargetproteins.Calciumbindstocalcium-bindingproteins(CaM)whichaffectsotherproteins.Cytosoliccalciumlevelsaredeterminedbyeventswithinamembrane.[Ca2+]oscillationsByPhorbolesterTwotypesof[Ca2+]channelsinsER:

IP3receptors(IP3-gatedCa2+-releasechannels);Ryanodinereceptors(RyRs),

RyRsarefoundprimarilyinexcitablecellsandbeststudiedinskeletalmusclecells.OneoftheeffectsofCaffeineistolowerthesensitivityofRyRstoCa2+,therebyincreasingthelikelihoodthattheseionchannelswillbeopenedbyCICR,makingcellsmoreexcitable.

咖啡因的作用之一就是降低ryanodine受體對Ca2+的敏感性，因而增加這些離子通道被CICR開通的可能性，使細胞更容易興奮。calcium-inducedcalciumrelease(CICR)Structureofcalmodulin,acytosolicproteinof148aminoacidsthatbindCa2+ionsCaMisfounduniversallyinplants,animals,andeukaryoticmicroorganisms.Ca2+/CaMteinkinase(CaM-kinase)mediatemanyoftheactionsofCa2+inanimalcells.Thefunctionsofincreasethelevelsofcytosoliccalcium-CaM:start-upembryodevelopmentafterthefecundation.excitatingcontractofmusclecells;excitatingsecretionofendocrineandnervecells.

UnlikecAMP,whoseactionismediatedbystimulationofaPK,Calciumcanaffectanumberofdifferenttypesofcellulareffectors,includingproteinkinase.Dependingonthecelltype,calciumionscanactivateorinhibitvariousenzymeandtransportsystems,changetheionicpermeabilityofmembranes,inducemembranefusion,oraltercytoskeletalstructureandfunction.RegulatingcalciumconcentrationsinplantcellsCytosoliccalciumchangesinresponsetoseveralstimuli,includinglight,pressure,gravity,andhormones.Calciumsignalingaidsindecreasingturgorpressureinguardcells.脫落酸

（a）氣孔的照片，每一個氣孔兩側并列著一對保衛細胞。當保衛細胞中維持高膨壓時，細胞如在圖上所見的向外膨出，氣孔因而保持開放。（b）控制氣孔大小的一個因子是脫落酸(ABA)，當ABA濃度增加時，質膜上的鈣離子通道打開，致使Ca2+流入（步驟1），這導致Ca2+從胞內貯存場所釋放出來（步驟2）。胞內Ca2+的再度上升使K+流入通道關閉而K+流出通道開啟（步驟3），因而導致胞內溶質濃度降低和水分喪失（步驟4）。

G-protein-linkedreceptordesensitizationdependsonreceptorphosphorylationbyPKA,PKC,CaMK2orG-protein-linkedreceptorkinases(GRKs)Thetargetcellscanbecomedesensitizedtoasignalmoleculebyfiveways.

Threegeneralwaysofthedesensitization:1.Receptorinactivationbyalteration;2.Receptorsequestrationbyinternalization;3.Receptordown-regulationbydestroyinginLs.

Sequestration;down-regulation;inactivation;inactivation;inhibitoryproteinThemechanismsthatshutoffasignalareasimportantasthemechanismsthatturniton.Toxinsproducedbybacteriathatcausecholera(affectingGsa)andwhoopingcough(pertussis)(affectingGia)Gs和Gi兩者都是細菌毒素的靶位點霍亂毒素(Choleratoxin),是由霍亂致病菌產生的一種酶,其能催化ADP核糖從胞內NAD+

轉移到Gs的α亞基.ADP核糖化(ribosylation)改變了α亞基的性質,使其不能將結合的GTP水解從而一直保持活化狀態.并持續激活腺苷酸環化酶,結果延長了高水平cAMP的維持,引起小腸上皮細胞的Cl-

和H2O進入腸道,因此造成腹瀉.百日咳毒素(Pertussistoxin),是由百日咳(whoopingcough)細菌產生,其催化Giα亞基的ADP核糖化,阻止了亞基和受體相互作用,結果使α亞基一直和GDP結合,使其不能作用靶蛋白.theophylline

D.J.[?θi??fili:n]

K.K.[θi?ɑf?l?n,?θio?f?l?in]

n.茶堿C.Receptortyrosinekinase(RTK)and

RTK-RassignalingpathwayRTKarethesecondmajortypeofcell-surfacereceptors

(over50kindshavebeenidentified,thatareinvolvedprimarilyinthecontrolofcellgrowthanddifferentiationratherthanintermediarymetabolism.)

SignalingligandsofRTKs:1.Nervegrowthfactor(NGF)2.Platelet-derivedgrowthfactor(PDGF)3.Fibroblastgrowthfactor(FGF)4.Epidermalgrowthfactor(EGF)5.insulinandinsulin-likeGF(IGF-1)6.Ephrins(Eph)7.vascularendothelialfactor(VEGF)Sixclassesofenzyme-linkedreceptorshavethusfarbeenidentified:Receptortyrosinekinase(RTK);Tyrosine-kinase-associatedreceptor;Receptorliketyrosinephosphatases;Receptorserine/threoninekinase;Receptorguanylylcyclases;Histidine-kinase-associatedreceptorStructureandactivationofreceptortyrosinekinasesRTKactivitystimulatedbycross-phosphorylation.PhsphorylatedTyrosineServeasdockingsitesforproteinwithSH2domains

(Srchomologyregion).OtherproteinmodulessuchasSH3bindstoproline-richmotifsinintracellularproteins.dimerizationStepsinactivationofRasbyRTKsPhosphotyrosinesofRTKactasbindingsitesforaspecificSH2proteincalledGRB2(Growthfactorreceptorbindingproteininmammalian).GRB2isnotaproteinwithcatalyticactivity,butonethatfunctionssolelyasanadaptermoleculethatlinksotherproteinsintoacomplex.Sos(sonofsevenless)isaguaninenucleotideexchangefactorforRas(Ras-GEF)WhenaligandbindstotheRTKandrecruitstheGrb2-Sostotheinnersurfaceofthemembrane,theSosproteinbindstoRascausingGDP/GTPexchange,thusactivatingRas.RTK-RassignalingpathwayInsulinreceptorTheactivatedRTKtransfersphosphategroupsto(1)autophosphorylation,(2)phosphorylationofIRSsTargetprotein:asmallfamilyofsolubleproteinsubsrtates.insulinreceptorsubstrates(IRSs)PhosphorylatedIRSactsasa“dockingstation”thatbindsproteinspossessingSH2domains.酪氨酸-磷酸化的IRS在激活各種信號傳導途徑中的作用—結合并活化各種效應器蛋白。(a)IRS被活化的胰鳥素受體磷酸化據信可以激活PI3K(phosphatidylinositol3-hydroxykinase)，PI-PLC

和Ras途徑，尚有另外一些不很確定的途徑也能為IRS所激活。(b)PI3K的活化導致膜結合的磷酸肌醇PIP2和PIP3生成。若干信號傳導途徑中的一個關鍵酶是PKB，它通過其蛋白質上的PH結構域與PIP2相互作用。這一反應改變PKB分子的構象，使之成為另一激酶（PDK1）的底物，PDK1磷酸化PKB，使之完全活化。一旦PKB被激活，便成為若干條的不同信號途徑中的一個重要組分，介導胰島素應答反應。這些途徑導致葡萄糖轉運蛋白轉位到質膜、糖原合成和細胞中某些新蛋白質的合成。PKB的失活是通過蛋白磷酸酶PP2A將其去磷酸化而完成。

Regulationofgeneexpressionbyinsulin

ActivationofglycogensynthesisbyinsulinRegulationofbloodglucoselevelGTP-GDPcyclingofRasproteinRasgenemutationsisfoundin30%ofallhumantumors.Ras(21Kda)consistsofasinglesmallsubunit.RashasaveryweakGTPaseactivityandwouldremainintheactive,GTP-boundstatefor30mins.Inthecell,RasactivityisregulatedbyGAPthatstimulatetheRasGTPaseabout105-fold.MutationsinoneoftheRas-GAPgenes(NF1)causeneurofibro-matosis1.Thedrugsinhibittheenzymefarnesyltransferase,thatpreventattachmentofRas,havebeendeveloped.MAPkinase=mitogen-activatedproteinkinase;MAP-KKK=Raf(Ser/Thr-PK)Todate,14MAPKKKs,7MAPKKs,and13MAPKshavebeenidentifiedinmammals.AdaptingtheMAPkinasecascadetotransmitdifferenttypesofinformation.yeastmatingpheromones;Fruitfliesdifferentiationofthephotoreceptors;Floweringplantsdefenseagainstpathogens.mammalscellproliferation.MAP-kinaseserine/threoninephosphorylationPathwayactivatedbyRasMEKD.Jak-STATsignalingpathwayJanuskinases(Jaks)andSTATs:Jaks:Aclassofcytoplasmictyrosinekinases,includingJak1,Jak2,Jak3,andTyk2,andeachisassociatedwithparticularcytokinereceptors;STATs:JaksthenphosphorylateandactivateasetoflatentgeneregulatoryproteinscalledSTATs(signaltransducersandactivatorsoftranscription,whichhaveanSH2domain),whichmoveintothenucleusandstimulatethetranscriptionofspecificgenes.SignalingligandsandCytokinereceptors:Ligands:morethan30cytokinesandhormonesactivatetheJak-STATpathwaybybindingtocytokinereceptors.Cytokinereceptors:theyarecomposedoftwoormorepolypeptidechains.