信号通路和肿瘤.ppt

信号通路和肿瘤信号通路和肿瘤引言：细胞信号转导与生命过程引言：细胞信号转导与生命过程问题提问题提出和理论产生出和理论产生细胞信号转导理论概述细胞信号转导理论概述信号转导研究中重大理论问题及热点领域信号转导研究中重大理论问题及热点领域信号转导研究方法与工具信号转导研究方法与工具信号转导理论研究及应用举例：在肿瘤发生信号转导理论研究及应用举例：在肿瘤发生发展中信号转导意义发展中信号转导意义信号转导与肿瘤临床：诊疗和治疗信号转导与肿瘤临床：诊疗和治疗细胞信号转导经典文件举例细胞信号转导经典文件举例信号通路和肿瘤2/48引言引言信号转导与生命过程信号转导与生命过程问题提出和理论产生问题提出和理论产生信号通路和肿瘤3/48细胞信号转导理论建立以前细细胞信号转导理论建立以前细胞生物学胞生物学细胞显微结构（胞膜、胞浆、胞核）细胞生理功效（生存、“活性”、分裂增殖、胞间连接、吞饮、分泌、迁移、死亡)细胞组分生物化学（脂、糖、核酸、蛋白）细胞超微结构和亚细胞结构（脂质双层膜结构、细胞器)信号通路和肿瘤4/48组织生长需要组织生长需要细胞分裂增殖细胞分裂增殖细胞细胞生长因子生长因子细胞周期细胞周期蛋白表示蛋白表示病原体侵入病原体侵入抗感染状态抗感染状态细胞细胞抗原抗原细胞因子细胞因子表示分泌表示分泌细胞过分生长细胞过分生长细胞死亡细胞死亡细胞细胞死亡因子死亡因子胞内致死胞内致死分子表示分子表示信号通路和肿瘤5/48细胞骨架蛋细胞骨架蛋白表示、激活白表示、激活牵动细胞移动牵动细胞移动(Cellmovement)趋化因子趋化因子细胞粘附细胞粘附细胞存活细胞存活(Survival)抗凋亡因子抗凋亡因子表示、激活表示、激活胞外信号胞外信号信号作用信号作用于细胞于细胞基因表基因表达改变达改变细胞表细胞表型改变型改变信号通路和肿瘤6/48信号通路和肿瘤7/48信号通路和肿瘤8/48细胞信号转导理论概述细胞信号转导理论概述信号通路和肿瘤9/48信号通路和肿瘤10/48胞外信号分子胞外信号分子(可溶性分子、细胞表面分子、组织基质分子可溶性分子、细胞表面分子、组织基质分子)靶细胞跨膜分子靶细胞跨膜分子(狭义受体如狭义受体如EGFR或广义受体如或广义受体如Integrin)靶细胞受体（胞内段）化学改变（如磷酸化、二聚体形成）靶细胞受体（胞内段）化学改变（如磷酸化、二聚体形成）靶细胞内信号转导分子化学改变与激活靶细胞内信号转导分子化学改变与激活（如磷酸化、去磷酸化、聚体形成）（如磷酸化、去磷酸化、聚体形成）激活信号转导分子进入胞核激活信号转导分子进入胞核进入胞核转导分子作用于基因转录调控区进入胞核转导分子作用于基因转录调控区基因表示改变基因表示改变信号通路和肿瘤11/48ExtracellularSignalMolecules1.GrowthFactorsPDGF(Platelet-DerivedGrowthFactor),EGF(EpidermalGrowthFactor),TGF-(TransformingGrowthFactor-),EPO(Erythropoietin),NGF(NerveGrowthFactor),IGF(Insulin-likeGrowthFactor),TPO(Thrombopoietin)2.CytokinesIFN-(Interferon-),IFN-(Interferon-),TNF(TumorNecrosisFactor),Interleukins(1,2,3,4)3.DeathmoleculesFas4.AdhesionmoleculesCadherins,Adhesin5.HormoneInsulin6.Stress信号通路和肿瘤12/48Signal Transducing Receptors1.Transmembranereceptorsthathaveintrinsicenzymaticactivity.AutophosphorylationPhosphorylationofothersubstratesA)Tyrosinekinases:PDGF-R,insulin-R,EGFRandFGF-RB)Tyrosinephosphatases:e.g.CD45C)Guanylatecyclases:e.g.natriureticpeptidereceptors)D)Serine/Threoninekinases:activinandTGF-receptors信号通路和肿瘤13/482.Receptorsthatarecoupled,insidethecell,toGTP-bindingandhydrolyzingproteins(G-proteins).e.g.,adrenergicreceptors,odorantreceptors,andcertainhormonereceptors(e.g.glucagon,angiotensin,vasopressinandbradykinin).3.Receptorsthatarefoundintracellularlyanduponligandbindingmigratetothenucleuswheretheligand-receptorcomplexdirectlyaffectsgenetranscriptione.g.,STAT1,3,4,5,6(Signaltransducerandactivatoroftranscription)4.Simplereceptors:e.g.,ion-channelsthatleadtochangesinmembraneelectricpotential信号通路和肿瘤14/48信号转导过程中生物化学信号转导过程中生物化学磷酸化反应（酪氨酸激酶、丝磷酸化反应（酪氨酸激酶、丝/苏氨苏氨酸激酶）酸激酶）蛋白质构象改变蛋白质构象改变去磷酸化反应（磷酸酶）去磷酸化反应（磷酸酶）受体或其它信号转导分子聚体化受体或其它信号转导分子聚体化信号通路和肿瘤15/48SignalTransducersReceptor Tyrosine Kinases(RTKs)contains:An extracellular ligand binding domain.An intracellular tyrosine kinase domain.An intracellular regulatory domain.A transmembrane domain.TyrosinephosphorylationInteractwithandphosphorylateSrchomologydomain2(SH2)-containingproteins(e.g.,PLC-,Ras,PI-3K,etc)Phosphorylateotherkinasesphosphorylateproteins,whichuponphosphorylated,canenterthenuclearandbindDNAregulatoryregions.信号通路和肿瘤16/48ClassExamplesStructural Features of ClassIEGF receptor,NEU/HER2,HER3cysteine-rich sequencesIIinsulin receptor,IGF-1 receptorcysteine-rich sequences;characterized by disulfide-linked heterotetramersIIIPDGF receptors,c-Kitcontain 5 immunoglobulin-like domains;contain the kinase insertIVFGF receptorscontain 3 immunoglobulin-like domains as well as the kinase insert;acidic domainVvascular endothelial growth factor(VEGF)receptorcontain 7 immunoglobulin-like domains as well as the kinase insert domainVIhepatocyte growth factor(HGF)and scatter factor(SC)receptorsheterodimeric like the class II receptors except that one of the two protein subunits is completely extracellular.The HGF receptor is a proto-oncogene that was originally identified as the Met oncogeneVIIneurotrophin receptor family(trkA,trkB,trkC)and NGF receptorcontain no or few cysteine-rich domains;NGFR has leucine rich domainCharacteristics of the Common Classes of RTKs 信号通路和肿瘤17/48Non-ReceptorProteinTyrosineKinases(PTKs)Twonon-receptorPTKfamilies:1)ThearchetypaplPTKfamilty:Src-relatedproteins2)Januskinase(Jak)familyMostnon-receptorPTKscoupletocellularreceptorsthatlackenzymaticactivitythemselves(e.g.,CD4,CD8,TCRandallcytokinereceptorssuchasIL-2R信号通路和肿瘤18/48信号通路和肿瘤19/48信号通路和肿瘤20/48信号通路和肿瘤21/48ReceptorSerine/ThreonineKinases(RSTKs)Typicalexample:ReceptorsfortheTGF-superfamilyofligandsTheTGF-superfamilyinclude30multifunctionalproteins,e.g.,activins,inhibinsandthebonemorphogeneticproteins(BMPs).17RSTKsisolatedarein2subfamilies:typeIandtypeIIreceptors.NuclearproteinsrespondingtoTGF-activationincludec-MycandSmadLigandsbindtothetypeIIreceptorsComplexedwithtypeIreceptorsTypeIIRphosphorylatestypeIreceptorInitiationofsignalingcascade信号通路和肿瘤22/48信号通路和肿瘤23/48Non-ReceptorSerine/ThreonineKinases1)cAMP-dependentproteinkinase(PKA)2)ProteinkinaseC(PKC)3)Mitogenactivatedproteinkinases(MAPKorERK)(requiringphosphorylationofbothtyrosineandthreonine)G-ProteinCoupledReceptors1.1000GPCRs,mostofwhichareorphanreceptors)2.ThreedifferentclassesofGPCR:1)GPCRsthatmodulateadenylatecyclaseactivityandproducecAMP2)GPCRsthatactivatePLC-gleadingtohydrolysisofpolyphosphoinositides:angiotensin,bradykininandvasopressinreceptors.3)Photoreceptor信号通路和肿瘤24/48IntracellularHormoneReceptors1.Residingwithinthecytoplasm.2.Thesteroid/thyroidhormonereceptorsuperfamily(e.g.glucocorticoid,vitaminD,retinoicacidandthyroidhormonereceptors):bindsteroid/thyroidhormone,translocatetonuclearandbindspecificDNAsequenceshormoneresponseelements(HREs).*PhosphatasesinSignalTransduction1.TransmembranePTPs:e.g.,CD45.2.IntracellularPTPs.信号通路和肿瘤25/48胞外信号分子胞外信号分子(可溶性分子、细胞表面分子、组织基质分子可溶性分子、细胞表面分子、组织基质分子)靶细胞跨膜分子靶细胞跨膜分子(狭义受体如狭义受体如EGFR或广义受体如或广义受体如Integrin)靶细胞受体（胞内段）化学改变（如磷酸化、二聚体形成）靶细胞受体（胞内段）化学改变（如磷酸化、二聚体形成）靶细胞内信号转导分子化学改变与激活靶细胞内信号转导分子化学改变与激活（如磷酸化、去磷酸化、聚体形成）（如磷酸化、去磷酸化、聚体形成）激活信号转导分子进入胞核激活信号转导分子进入胞核进入胞核转导分子作用于基因转录调控区进入胞核转导分子作用于基因转录调控区基因表示改变基因表示改变信号通路和肿瘤26/48信号转导研究中重大理信号转导研究中重大理论问题及热点领域论问题及热点领域信号通路和肿瘤27/48信号转导通路调控信号转导通路调控磷酸化磷酸化去磷酸化调控去磷酸化调控信号转导分子消长调控（分子半衰期）信号转导分子消长调控（分子半衰期）不一样通路之间效应调控不一样通路之间效应调控胞内内源性抑制物调控功效胞内内源性抑制物调控功效信号通路和肿瘤28/48Cross-Talk信号通路和肿瘤29/48信号转导效应特异性信号转导效应特异性WhenandWhere?Cooperationwithothersignalingpathways?Pre-existingtranscriptionco-factorsdifferentiallyexpressedandactivatedindifferentcelltypes?Pre-existingco-activatorsoftargetproteins?Subcellularlocalizationoftransducers?Optimallevel(orathreshold)ofphosphorylation/dephosphorylation?信号通路和肿瘤30/48替换通路替换通路（AlternativePathways)信号通路和肿瘤31/48信号转导研究方法与信号转导研究方法与工具工具信号通路和肿瘤32/48一、蛋白质磷酸化状态检测一、蛋白质磷酸化状态检测1、免疫印迹(phospho-proteinspecificantibodies)2、免疫沉淀(protein-specificantibody+phospho-AAantibody3、流式细胞仪分析4、Luminex分析二、信号转导分子过分表示或过分激活二、信号转导分子过分表示或过分激活1、Overexpressionbygenetransduction2、Constitutivelyactivatedmutants三、基因转录活性测定三、基因转录活性测定1、Electrophoreticmobilityshiftanalysis(EMSA)、Reportergeneexpressiondetection信号通路和肿瘤33/48四、信号转导分子表示或活性抑制四、信号转导分子表示或活性抑制1、Anti-sense2、RNAi3、Geneknock-out4、Dominantnegativemutants(1)Ligand-bindingsite(2)Phosphorylationsite(3)Dockingsite(4)Protein-proteinbindingsite(5)DNAbindingsite5、Small-moleculeinhibitors：e.g.,tyrosinekinaseinhibitor(TKi)6、Inhibitoryoligopeptides信号通路和肿瘤34/48信号转导在肿瘤发生信号转导在肿瘤发生发展中意义发展中意义信号通路和肿瘤35/48Signalingmoleculesinvolvedincancerdevelopment/progressionReceptors1)Growthfactorreceptors:EGFR2)Hormonereceptor:ER,AR3)Angiogenicreceptros:VEGF,PDGF,IGF4)Deathreceptors5)TheIntegrinsystem信号通路和肿瘤36/48Transducers1)Ras2)Raf3)Rhofamily4)PI-3K/Akt5)Deathtransducers6)STAT-37)Transcriptionfactors1)c-Myc2)c-Junandc-fos3)STAT-34)信号通路和肿瘤37/48BiologicalEffectsofSignalingRelatedtoCancerDevelopment/ProgressionCellimmobilizationAbrogationofapoptosisActivationofcellcycleandremovalofcellcyclecheckpointsAngiogenesisCellinvasionMetastasisDrugresistance信号通路和肿瘤38/48信号通路和肿瘤39/48PhosphorylationtargetsofPI-3KAktForkhead-relatedtranscriptionfactor1(FKHR-L1)14-3-3bindingFKHR-L1retainingincytosolabrogationofgeneactivationbyFKHR-L1AktBad14-3-3bindingReleaseofBcl-2andBcl-XCellsurvivalAktGSK3GSK3catalyticactivityturnedoffPermittingactivationofc-MycandcyclinDPDK1phosphorylationofotherkinases(p70S6-kinasse,CISK,PKC)Cellgrowthandsurvival信号通路和肿瘤40/48信号转导与肿瘤临床信号转导与肿瘤临床诊疗、预防与治疗诊疗、预防与治疗Expressionlevel,mutationsandantibodiesofsignalingmoleculesincancerdiagnosis1)EGFR:lung,H&N,glioma2)TGF-receptor(typeII):lung,H&N,3)Ras4)Androgenreceptoranddownstreammolecules5)Estrogenreceptoranddownstreammolecules6)信号通路和肿瘤41/48Proteinphosphorylationandcancerdiagnosis1)Determinationofsinglephosphorylatedsignalingmolecules:EGFR2)Phosphoproteinprofiling:proteomics3)Phospho-proteinbasedimagingtechnologySignalinginhibitorandcancerprevention/therapy1)EGFR-selectiveTKi:Iressa2)EGFRantibodies3)FarnesylationorGeranylgeranylationinhibitorstargetingRasandRho4)VEGFRantibody(andTKi):Avastin信号通路和肿瘤42/48经典文件举例经典文件举例信号通路和肿瘤43/48Stat3asanOncogeneJacquelineF.Bromberg1,MelissaH.Wrzeszczynska1,GeetaDevgan1,YanxiangZhao2,RichardG.Pestell3,ChrisAlbanese3andJamesE.Darnell,Jr.,1,1LaboratoryofMolecularCellBiology,TheRockefellerUniversity,NewYork,NewYork10021-6399,USA2LaboratoryofMolecularBiophysics,TheRockefellerUniversity,NewYork,NewYork10021-6399,USA3TheAlbertEinsteinCancerCenter,DepartmentofDevelopmentalandMolecularBiology,andDepartmentofMedicine,AlbertEinsteinCollegeofMedicine,Bronx,NewYork10461,USAReceived30March1999;Revised23June1999.Availableonline27SeptemberCell Volume 98,Issue 3,295-303 信号通路和肿瘤44/48信号通路和肿瘤45/48信号通路和肿瘤46/48信号通路和肿瘤47/48信号转导理论在各生命科学领信号转导理论在各生命科学领域中普遍意义域中普遍意义以本课程中各讲为例：以本课程中各讲为例：干细胞蛋白质组学细胞凋亡肿瘤转移血管增生信号通路和肿瘤48/48