《生化--郑利民课件 信号转导2－生化课08.ppt》由会员分享，可在线阅读，更多相关《生化--郑利民课件 信号转导2－生化课08.ppt（69页珍藏版）》请在三一文库上搜索。
1、1,Signal-Transduction Pathways 信号转导通路 (2) 郑利民 2008 珠海 ,2,Biosignaling: How are they regulated?,3,凋亡细胞,细胞凋亡：细胞死亡的主要方式；受机体严密调控。 凋亡细胞的清除：主要被巨噬细胞（Mf）吞噬； 摄取普通的凋亡细胞能主动抑制Mf活化 安全清除凋亡细胞，维持组织自身稳态的机制,巨噬细胞摄取凋亡细胞后， 通过分泌多种细胞因子（TGFb，IL10，PGE2等）并减少TNFa等，抑制免疫应答,细胞凋亡和免疫应答,4,Apoptotic cell,已知：粒细胞是清除入侵病原的主力军； 多种病原均可引起粒
2、细胞凋亡。 Q：这类诱导凋亡是病原的致病策略 ? 消灭这些关键免疫细胞， 利用它们作为“特洛依木马”来感染并灭活M,5,UV照射或细菌感染诱导人粒细胞凋亡,UV,S. aureus,E. coli,6,7,大肠杆菌或金葡菌诱导的凋亡粒细胞 刺激巨噬细胞产生TNF和上调CD64表达,8,采用多种不同方式诱导粒细胞凋亡，发现： 照射或老化所致的凋亡细胞能抑制巨噬细胞活化 而病原感染所致的凋亡细胞能显著激活巨噬细胞,抗原递呈细胞（APC）,UV或 老化,凋亡,?,凋亡粒细胞,感染,趋炎症反应 抗原递呈,凋亡,J. Immunol. 2002, 168:6358-65； 2004, 173:6319-
3、26,9,这两类凋亡粒细胞对M的活化具截然相反的作用 （细胞固有一死，但死的意义有不同）说明： 粒细胞凋亡是调控免疫和炎症反应的新型防御机制 早期，吞噬受感染的凋亡细胞可激活M，帮助控制病原； 后期，过剩的粒细胞发生凋亡并抑制M活性，炎症消退。,Question： M如何识别这两类凋亡细胞并作出相反应答？,10,抗原递呈细胞（APC）,UV或 老化,凋亡,凋亡,HSP,凋亡粒细胞,感染 或热敏,趋炎症反应 抗原递呈,1. 凋亡细胞中的危险信号, 如HSP，而不是这些细胞所相伴的细菌，决定了APC的免疫应答类型。 2. 它们可作为天然佐剂，来研制新型抗肿瘤和感染疫苗,J. Immunol. 20
4、04, 173:6319-6326,11,病原菌经常是通过干扰细胞的信号通路而致病 致病性沙门氏菌能侵入巨噬细胞并诱导其凋亡； 非致病性沙门氏菌 经调理后也能进入巨噬细胞，但 不能诱导其凋亡,12,SopE,PKB/Akt,Rac, CDC42,PTK,PI-3K,R,调理后吞噬,病原侵袭,死亡信号,生存信号,细胞凋亡和粘膜屏障损坏,细胞存活,细胞凋亡是由： 细胞内“死亡/生存”信号之间的精密平衡来决定 干扰该平衡就可改变病原对细胞凋亡的最终影响,病原侵袭和吞噬对巨噬细胞凋亡的影响及其机制,13,细胞存活,细胞凋亡,The balance between pro- and anti-apopt
5、otic genes/signals determine the cell fate,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,14,PART 1 Basic characteristics of signal transduction 2 Four general types of signal transducers PART 1 Regulatory mechanisms 2 Some diseases caused by defects in the biosignaling pathways,15,Regulatory mechanisms o
6、f Bio-signals,3.1 Phosphorylation as a regulatory mechanism 3.2 Regulation of transcription by steroid hormones 3.3 Regulation of the cell cycle by protein kinases,16,Regulation of Signaling Pathways,External signals,Second messengers,Modulator proteins,Function,Target proteins,Hormones cAMP Odorant
7、s cGMP Drugs Ca2+ Light DAG IP3 Mitogenic hormones Tyrosine Growth kinases factors,protein kinase And phosphatase,structural proteins And enzymes,metabolic or physiological responses,Plasma membrane,17,Protein Kinases 1: GPCRs (5%), 2: Kinases (2.8% genome), 25% of the mammalian intracellular protei
8、ns undergoes reversible phosphorylation,18,Protein kinases (534 human kinases),Ser/thr Kinases comprise 80% of all protein kinases,AGC,19,Phosphorylation & de-phosphorylation are the most common regulatory mechanisms, mediated by protein kinase and phosphatase, respectively,20,Nobel Prize in Physiol
9、ogy and Medicine 1992,Eldwin G. Krebs,Edmond H. Fischer,USA,“Reversible protein phosphorylation as a biological regulatory mechanism“,J. Biol. Chem. 216:121-132, 1955,21,cAMP,PKA,Phosporylating cellular proteins,Response,Activation of G ProteinCoupled Receptors,22,GEF：Guanine nucleotide Exchange Fac
10、tor GAP：GTPase Activating Protein,Activation of Ras protein,23,Ras蛋白的上游和下游信号通路,24,PI3K的活化机制及PIP3磷酸酶 (PI3K is a downstream target of Ras),25,PI3KAKT途径对细胞生存的调控机制,26,Two-component system or signaling,27,28,Most of these complicated signaling pathways are transduced by the phosphorylation and dephosphor
11、alation of signaling protein, regulated by the protein kinases and phosphatses,29,蛋白激酶（protein kinase，PK） 植物和酵母中2%的基因编码蛋白激酶；而在哺乳动物中，高达58。 根据磷酸化靶蛋白的氨基酸残基的种类不同，蛋白激酶有丝氨酸/苏氨酸激酶、酪氨酸激酶和组氨酸激酶等三类。部分蛋白激酶具有双重底物特异性，既可使丝氨酸或苏氨酸残基磷酸化，又可使酪氨酸残基磷酸化,30,蛋白磷酸酶 protein phosphatase，PP 蛋白磷酸酶的分类与蛋白激酶相对应，分为丝氨酸/苏氨酸型蛋白磷酸酶和酪氨酸
12、型蛋白磷酸酶。有些酶具有双重底物特异性 对蛋白磷酸酶的研究还不如蛋白激酶那样深入。但两者的协同作用在细胞信号转导中的作用是不言而喻的。,31,Phosphorylation as a Regulatory Mechanism,Signal transduction,Altered kinase activity,cGMP cAMP Insulin,Ca2+ kinase,There are 120 kinases in yeast i.e. 2% of their entire genome Some kinases are localized to discrete subcellular
13、regions that are in close proximity to the specific target protein;,32,Many proteins act as targeting or anchoring mediators e.g. AKAP associates tightly with the regulatory subunits of PKA Cells contain many different isoforms of PKC, which are localized by anchoring proteins, e.g. RACKs or recepto
14、rs for activated PKC Phosphatases are also often localized closely to ensure that integrated control is possible.,33,Mechanisms used by hormones, retinoids, and Vit. D to regulate gene expression,Hormone (H), carried to the target tissue on serum binding prot, diffuses across the PM and binds to its
15、 specific receptor protein (Rec) in the nucleus.,H binding changes the conformation of Rec; forms homo or heterodimers and binds to specific R regions called hormone response elements (HREs) in the DNA adjacent to specific genes,Binding regulates transcription of the adjacent gene(s), increasing or
16、decreasing the rate of mRNA formation.,Altered levels of the hormone regulated gene product produce the cellular response to the hormone,34,3.3.1 The cell cycle,What is cell cycle? A process of cellular reproduction,35,36,Four phases: G1, S, G2, M Gap (G1 and G2) phases: Respond to proliferative and
17、 anti-proliferative signals, synthesize RNA and protein Synthetic (S) phase: Replication of the chromosomes Mitotic (M) phase: segregation of chromosomes and other cell constituents into two daughter cells,37,Cell cycle transits orderly and irreversibly,38,Cell cycle transits orderly and irreversibl
18、y,The completion of one phase is required for the beginning of the next. Periodic activation of specific cyclin-Cdk complexes A cycle of protein destruction eliminates proteins used in the preceding phase as well as proteins that would inhibit progression into the next phase.,39,Cyclin-dependent kin
19、ases (Cdks),Cdks drive cell cycle progression by phosphorylating a group of substrates,- Cdks present a unique pattern of activity in each phase of the cell cycle Cdk1 (Cdc2) Cdk2 Cdk3 Cdk4 Cdk5 Cdk6 Cdk7,G1,G2, M,Late G1, S,G1, S, G2, M,40,Regulation of Cdks activity Cyclin: Activating subunit.,Cyc
20、lins were named because of their cyclic expression during the cell cycle,41,Four mechanisms regulate CDK activity: 1 phosphorylation or dephosphorylation, 2 controlled degradation of cyclin subunit, 3 periodic synthesis of CDKs and cyclins, 4 action of specific CDK inhibiting proteins.,42,Regulation
21、 of Cdks activity Phosphorylation and dephospho. in Cdk1,Activating phosphorylation site: Thr 161 Inhibitory phosphorylation site: Thr 14, Thr 15,P,P,43,Growth controlling signals are conveyed by stimulatory or inhibitory pathways,44,some target proteins of CDKs: Laminin - Breakdown of nuclear envel
22、ope before segregation of sister chromatids in mitosis is partly due to Pi-laminin by CDK Actin myosin contractile machinery-Involved in pinching a dividing cell into two equal parts during cytokinesis Retinoblastoma protein, pRb-participates in a mechanism that arrests cell division in G1,CDKs Regu
23、late Cell Division by Phosphorylating Critical Proteins,45,Summary,Four basic characters of biosignals Four schemes of intercellular signaling Four general types of signal transducers gated ion channels; receptor enzymes; G-protein coupled receptors；Steroid Receptors G proteins: Heterotrimeric Activ
24、ation and deactivation mechanisms,46,Major second messengers: cAMP, IP3, DAG, Ca2+, cGMP. Using an example to explain the cascade of events: a single molecule of hormone activates catalyst that in turn activates another catalyst, and so on, results in signal amplification and cell response The cell
25、cycle and its regulation,47,PART 1 Basic characteristics of signal transduction 2 Four general types of signal transducers PART 1 Regulatory mechanisms 2 Some diseases caused by defects in the biosignaling pathways,信号转导异常的原因和机制,一、信号转导异常 1.生物学因素： 干扰细胞内信号转导通路:如霍乱毒素 2. 理化因素： 电离辐射或机械刺影响信号转导成分而致癌 3.遗传因素：
26、信号蛋白数量/功能改变（突变） 4.免疫学因素：刺激型和阻断型抗受体抗体 5.内环境因素,49,二、信号转导异常的发生环节,Gs,ADH,H2O,H2O,配体、受体或受体后信号通路的任何一个环节出现障碍都会影响到最终效应，使细胞增殖、分化、凋亡、代谢或功能失常，并导致疾病。 以尿崩症为例,50,ADH作用的三个环节异常均可导致尿崩症： ADH分泌减少 中枢性尿崩症 ADH-V2受体变异 肾小管上皮细胞水通道蛋白（AQP2）异常,集合管上皮细胞对ADH的反应性降低,家族性尿崩症,不同受体介导的信号转导通路存在cross-talk 并非所有的信号转导蛋白异常都能导致疾病,51,细胞信号转导异常与疾
27、病,受体、信号转导障碍与疾病 受体数量减少 受体亲和力降低 受体阻断型抗体的作用 协同因子或辅助因子缺陷 受体功能缺陷 受体后信号转导蛋白缺陷,特定信号转导过程减弱或中断,激素抵抗征,52,雄激素受体缺陷与雄激素抵抗征,原因和机制： AR减少或失活突变,男性假两性畸形 特发性无精症症 延髓脊髓性肌萎缩,53,胰岛素受体与胰岛素抵抗性糖尿病,1.遗传性胰岛素受体异常，包括受体的 合成减少 与配体的亲和力降低，如Arg735突变为Ser TPK活性降低，如Gly1008 突变等 2.自身免疫性胰岛素受体异常 血液中存在抗胰岛素受体的抗体,54,二、受体、信号转导过度激活与疾病,某些信号转导蛋白过度
28、表达 某些信号转导蛋白组成型激活突变 刺激型抗受体抗体,生长激素（GH）分泌过多的垂体腺瘤中，30%是由于编码Gs的基因突变所致，从而抑制了GTP酶活性，使Gs处于持续激活状态，cAMP含量增多，垂体细胞生长和分泌功能活跃。,通路过度激活,如肢端肥大症和巨人症,55,霍乱毒素选择性的催化Gs亚基上的Arg201核糖化，使GTP酶活性丧失，不能将GTP水解成GDP，从而使Gs处于不可逆激活状态, 不断刺激AC生成cAMP,胞浆中的cAMP含量可增加至正常的100倍以上， 导致小肠上皮细胞膜蛋白构型改变，大量Cl-和水分子持续转运入肠腔，引起严重腹泻和脱水。,56,霍乱 (Cholera),CT：
29、Cholera Toxin,57,三、多环节信号转导异常与疾病, 肿瘤 1. 促细胞增殖的信号转导过强 生长因子产生增多 受体的改变 生长因子受体表达异常增多 突变使受体组成型激活 细胞内信号转导蛋白的改变 如Ras突变,58,2. 抑制细胞增殖的信号转导过弱,生长抑制因子受体减少、丧失 受体后信号转导通路异常,细胞的生长负调控机制减弱或丧失,59,Oncogene-encoded defective EGF receptor,60,GEF：Guanine nucleotide Exchange Factor GAP：GTPase Activating Protein,Activation o
30、f Ras protein,61,GEF：Guanine nucleotide Exchange Factor GAP：GTPase Activating Protein,62,肿瘤形成必须是在单个细胞中发生多重遗传突变 单个癌基因激活和抑癌基因失活不足以引起肿瘤细胞的显著扩增， 因为细胞对凋亡的易感性也同时增加；凋亡抑制是肿瘤细胞能够生长到足以威胁宿主所必须的；例如： Rb的失活仅会促进p53依赖的凋亡，但不会引起肿瘤，除非细胞的凋亡过程被抑制；,Apoptosis and oncogenesis,63,3.3.3 Oncogenes, Tumor Suppressor Genes (TSG
31、s), and Programmed Cell Death,Oncogenes: encode defective signaling proteins ( e.g., growth factors, receptors, G proteins, protein kinases, or transcription regulators), continually giving the signal for cell division; TSG: encode regulatory proteins; normally inhibit cell division; mutations in th
32、ese genes are genetically recessive Cancer: the result of an accumulation of mutations in oncogenes and TSGs.,64,From normal epithelial cell to cancer,65,细胞信号转导调控与疾病防治,Regulation of cellular signals in prevention & treatment of diseases 以信号转导蛋白为靶分子对疾病进行防治,66,STI571 as a paradigm (范例） for cancer ther
33、apy,In 1960, described the presence of a consistent chromosomal abnormality in CML（一种白血病)，so-called Philadelphia chromosome (Ph) A reciprocal translocation between the long arms of chromosomes 9 and 22, t(9:22)(q34;q11), The molecular consequence of this event is the generation of a chimeric bcr-abl gene (22:9),67,The tyrosine kinase activity of Bcr-Abl stimulates a variety of signaling pathways, leading to alterations in survival properties of cells,68,STI571 an Abl-specific tyrosine kinase inhibitor,Before the discovery of this drug, most CML patients died within 6 month,69,THANKS,