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1、1,Signal-Transduction Pathways 信号转导通路 (1) 郑利民 2008 珠海 ,2,干细胞的分化,3,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,4,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,5,细胞信号转导网络的简单模式,（信号输入）,（信号输出）,6,Important roles of biosignaling,Functional integration of distant organs, tissues and cells
2、requires communication; Signaling is perhaps a primal requirement to respond to our environment; The foundation of any complex response pathway lies with cellular biochemicals.,Biosignaling Intercellular（细胞间）& Intracellular（细胞内）,7,常见四种类型： Endocrine (内分泌） Paracrine (旁分泌） Autocrine (自分泌） Membrane atta
3、ched protein,Intercellular signaling （细胞间信号）,8,Four schemes of intercellular signaling (1),9,Four schemes of intercellular signaling (2),10,Intracellular signaling （细胞内信号）,11,Intracellular Receptor,12,Electron-micrograph of macrophage (pink) attacking Escherichia coli (green),13,M吞噬处理入侵细菌及提呈抗原的机制,14
4、,Signal Transduction Pathway: Complicated,15,FcR CR3,Ca,+,src,PI3k,PKC,MAPK,RhoGTPase,gelsolin,Arp2/3,PLC,PLD,Actin rearrangement,Phagocytosis; Oxidative activation,Signals for phagocytosis,Signal,Receptor,Amplification,Transduction,Responses,second messengers,16,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
5、反馈调节,胞外 质膜 胞内,17,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,18,1 Four basic characteristics: 1.1 Specificity 1.2 Amplification 1.3 Desensitization/Adaptatio
6、n 1.4 Integration,19,Specificity Signal molecule fits binding site on its complementary receptor; other signals do not fit.,thrombin,20,Scatchard analysis quantifies the receptor-ligand interaction,21,often short-lived & low concentration,22,Desensitization/Adaptation Receptor activation triggers a
7、feedback circuit that shuts off the receptor or removes it from the cell surface,Produce a rapid and major cellular response to a transient signal,23,Integration When two signals have opposite effects on a metabolic characteristic such as concentration of a second messenger X, or the membrane potent
8、ial Vm, the regulatory outcome results from the integrated input from both receptors,24,细胞存活,细胞凋亡,The balance between pro- and anti-apoptotic 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,25,26,2. Four types of signal transducers,2.1 Gated Ion Channels 2.1.1 Ligand-g
9、ated ion channels 2.1.2 Voltage-gated ion channels 2.2 Receptor Enzymes 2.3 G ProteinCoupled Receptors and Second Messengers 2.4 Steroid Receptors,27,Four general types of signal transducers,28,Why Ion Channels ?,Resting potential: Asymmetric ion-distribution Acting potential Gated Ion Channels Liga
10、nd-gated ion channels Voltage-gated ion channels,29,Resting potential,30,Why Ion Channels: asymmetric ion-distribution,31,asymmetric ion-distribution,Resting potential,32,pump and ion leak channels,Cl - leak channel,33,Acting potential,34,Acting potential,Voltage-gated Na+ channels & K+ channels,35,
11、2.1.1 Ligand-gated ion channel: Binding of some small molecule forces an allosteric transition in protein, open/close channel. acetylcholine (乙酰胆碱) receptor ion channel 2.1.2 Voltage-gated ion channel A charged protein domain moves relative to the membrane in response to a change in transmembrane el
12、ectrical potential. (voltage-gated Na+， Ca2+，K+ channels),36,乙酰胆碱受体离子通道 1,Ach,ACh,37,Binding of ACh to R cause conformational change. As M2 helices twist slightly, the Leu residues (yellow) rotate away from the channel and are replaced by smaller polar residues (blue). This gating mechanism opens ch
13、annel, allowing passage of Ca, Na, or K,乙酰胆碱受体离子通道 2,Closed Open,38,Voltage-gated Na+ channels 1,39,Voltage-gated Na+ channels 2,40,2. Four types of signal transducers,2.1 Gated Ion Channels 2.1.1 Ligand-gated ion channels 2.1.2 Voltage-gated ion channels 2.2 Receptor Enzymes 2.3 G ProteinCoupled Re
14、ceptors and Second Messengers 2.4 Steroid Receptors,41,2.2 Receptor enzymes,A ligand-binding domain (胞外) and an enzyme active site on cytosolic side, connected by a single transmembrane segment. Commonly a kinase that phosphorylates Tyr residues in specific target proteins（insulin receptor） Other: s
15、ynthesize the i.c. second messenger cGMP in response to ex.c. signals ( the receptor for atrial natriuretic factor),42,Activation of receptor tyrosine kinases,43,Insulin receptor tyrosine kinase,Insulin structure,44,Insulin receptor binds insulin and undergoes autophosphorylation on its carboxyl-ter
16、minal Tyr residues.,Insulin receptor phosphorylates IRS-1 on its Tyr residues.,SH2 domain of Grb2 binds to PTyr of IRS-1. Sos binds to Grb2, then to Ras, causing GDP release and GTP binding to Ras.,Activated Ras binds and activates Raf-1.,Raf-1 phosphorylates MEK on two Ser residues, activating it.
17、MEK phosphorylates ERK on a Thr & a Tyr residue, activating it.,ERK moves into the nucleus and phosphorylates Nuclear transcription factors such as Elk1, activating them.,Phosphorylated Elk1 joins SRF to stimulate the transcription and translation of a set of genes needed for cell division.,45,Activ
18、ation of glycogen synthase by insulin,46,Regulation of blood glucose level,47,Receptor for atrial natriuretic factor,Two types (isozymes) of guanylyl cyclase that participate in signal transduction.,48,2. Four types of signal transducers,2.1 Gated Ion Channels 2.1.1 Ligand-gated ion channels 2.1.2 V
19、oltage-gated ion channels 2.2 Receptor Enzymes 2.3 G ProteinCoupled Receptors and Second Messengers 2.4 Steroid Receptors,49,2.3 GPCR and Second messengers,Three essential components: 1. a plasma membrane receptor with seven transmembrane helical segments 2. an enzyme in the plasma membrane that gen
20、erates an intracellular 2nd messenger 3. a guanosine nucleotidebinding protein (G protein),50,Three essential components of G ProteinCoupled Receptors,51,52,A protein binds Guanine nucleotides (GDP, GTP); activated in GTP-form, inactivated in GDP-form Integral membrane protein, heterotrimers (); Hav
21、e similar & subunits, but differ in -subunit When G-protein is activated, the subunit dissociates to interact with an enzymes that generate second messengers (e.g. cAMP) Others: small G-proteins (20-25 kDa), e.g. Ras, Rho, Rac, etc, are not membrane bound.,G protein (GTP-binding protein),53,G protei
22、n (discovery),M. Rodbell: a transducer provided the link between receptor and amplifier. A. G. Gilman: identify & purify the G protein. System: Mutated lymphoma cells containing a normal receptor and cAMP-generating enzyme, was yet unable to respond (produce cAMP), since it lacked the transducer,mut
23、ated cell,normal cell,54,Nobel Prize in Physiology and Medicine 1994,“for their discovery of G-proteins and the role of these proteins in signal transduction in cells“,Alfred G. Gilman,1941-,Martin Rodbell,1925-1998,55,“ON-OFF” switch is regulated by GTP or GDP bound form. All G-proteins has intrins
24、ic GTPase activity, release Pi and inactivated. Activation: release of GDP and replaced by GTP,56,Two major systems: 2.3.1 THE PKA SYSTEM (cAMP as the second messenger) The -Adrenergic Receptor System 2.3.2 THE PKC SYSTEM (DAG,IP3 and Ca2+ as the second messengers),57,The association of active Gs wi
25、th adenylyl cyclase stimulates the cyclase to catalyze cAMP synthesis,Adenosine 3,5-cyclic monophosphate (cAMP),58,synthesized in adrenal medulla; belongs to catecholamines (儿茶酚胺）; target cells include liver, skeletal muscle, heart muscle and adipose; released in response to acute stress,Epinephrine
26、 肾上腺素 signal,59,Epinephrine 肾上腺素 signaling pathway,cAMP,60,Epinephrine 肾上腺素 signaling pathway (2),61,Activation of cAMP-dependent protein kinase (PKA),Inactive PKA: Regulatory (R) subunits: auto-inhibitory domains buried catalytic (C) subunits: substrate-binding sites blocked by R subunits,R subunit
27、s: autoinhibitory domains buried,Active PKA C subunits open substrate binding sites,62,A catalytic subunit of PKA,ATP,Potent inhibitor peptide (PKI): Arg-Arg-Gln-Ala-Ile (consensus sequence recognized by PKA except Ala),63,Epinephrine triggers a series of reactions in hepatocytes in which catalysts
28、activate catalysts, resulting in great “amplification” of the signal,x 分子,10 ,000 x 分子,64,PKA regulates a number of enzymes The proteins phosohorylated by PKA share a region of sequence similarity around the Ser or Thr residue that undergoes phosphorylation, a sequence that marks them for regulation
29、 by PKA. The catalytic site of PKA interacts with several residues near the Thr or Ser residue in the target protein, which define the substrate specificity.,65,66,Desensitization of the PKA system,1 desensitizing -Adrenergic Receptor 2 degrading the second messager,67,Gsbg recruits bARK to the memb
30、rane, where it phospho- Ser at the C-terminus of Recpt. barr binds to the pi- C-terminus of Recpt. Receptor-arrestin complex enters the cell by endocytosis.,68,-Arrestin uncouples receptor from its G protein and brings together 3 enzymes of MAPK cascade. (One stimulus triggers two distinct pathways:
31、 the path activated by G protein and MAPK cascade),69,degrading the second messager,70,The Protein kinase C (PKC) System,Two intracellular 2nd messengers are produced in the hormone-sensitive phosphatidylinositol system: Inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). Both contribute to
32、 the activation of PKC. By raising cytosolic Ca2+, IP3 also activates other Ca2-dependent enzymes; thus Ca2+ also acts as a second messenger,71,72,Calcium Is a Second Messenger in Many Signal Transductions,Normally, Ca2i is 100 nM (而细胞外：1 mM) Hormonal, neural, or other stimuli cause either an influx
33、 of Ca2+ into the cell through specific Ca2+ channels in the plasma membrane or the release of Ca2+ from ER or mitochondria, Changes in Ca2i are detected by Ca2+-binding proteins that regulate a variety of Ca2-dependent enzymes-Calmodulin (CaM),73,Regulation of transcription by steroid hormones,Steroid receptor,