1、Signal-Transduction Pathways 郑利民 200512 珠海,细胞信号转导网络的简单模式,（信号输入）,（信号输出）,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,信号受体反应 触摸含羞草后小叶合拢 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,树突状细胞和T淋巴细胞(红色)相互作用,Important roles of biosignaling,Functional integration of distant organs, tissues and cells requires communi
2、cation;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）,Four schemes of intercellular signaling (1),Four schemes of intercellular signaling (2),I
3、ntracellular Receptor,Electron-micrograph of macrophage (pink) attacking Escherichia coli (green),M吞噬处理入侵细菌及提呈抗原的机制,FcR CR3,Ca,+,src,PI3k,PKC,MAPK,RhoGTPase,gelsolin,Arp2/3,PLC,PLD,Actin rearrangement,Phagocytosis; Oxidative activation,Signals for phagocytosis,Signal,Receptor,Amplification,Transduct
4、ion,Responses,second messengers,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,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,1 Four basic characteristics: 1
5、.1 Specificity 1.2 Amplification 1.3 Desensitization/Adaptation 1.4 Integration,Specificity Signal molecule fits binding site on its complementary receptor; other signals do not fit.,Scatchard analysis quantifies the receptor-ligand interaction,often short-lived & low concentration,Desensitization/A
6、daptation Receptor activation triggers a 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,Integration When two signals have opposite effects on a metabolic characteristic such as concentration of a seco
7、nd messenger X, or the membrane potential Vm, the regulatory outcome results from the integrated input from both receptors,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,细胞存活,细胞凋亡,The balance between
8、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,2. Four general types of signal transducers,2.1 Gated Ion Channels2.1.1 Ligand-gated ion channels2.1.2 Voltage-gated ion channels 2.2 Receptor Enzymes 2.3 G ProteinCoupled Receptors and Second Mess
9、engers 2.4 Steroid Receptors,Four general types of signal transducers,Why Ion Channels ?,Resting potential: Asymmetric ion-distributionActing potentialGated Ion ChannelsLigand-gated ion channelsVoltage-gated ion channels,Resting potential,Why Ion Channels: asymmetric ion-distribution,asymmetric ion-
10、distribution,Resting potential,pump and ion leak channels,Cl - leak channel,Acting potential,Voltage-gated Na+ channels & K+ channels,Resting state,Acting potential 1-3,Acting potential 3-4,Acting potential 4,Acting potential 5-6,Acting potential,Acting potential,2.1.1 Ligand-gated ion channel: Bind
11、ing 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 electrical potential. (voltage-gated N
12、a+， Ca2+，K+ channels),乙酰胆碱受体离子通道 1,Ach,ACh,Binding of ACh to receptor 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 the channel, allowing the passage of
13、Ca, Na, or K,乙酰胆碱受体离子通道 2,Closed Open,Voltage-gated Na+ channels 1,Voltage-gated Na+ channels 2,2. Four general types of signal transducers,2.1 Gated Ion Channels2.1.1 Ligand-gated ion channels2.1.2 Voltage-gated ion channels 2.2 Receptor Enzymes 2.3 G ProteinCoupled Receptors and Second Messengers
14、2.4 Steroid Receptors,2.2 Receptor enzymes,A ligand-binding domain (ex.c. membrane surface) and an enzyme active site on the cytosolic side, connected by a single transmembrane segment. Commonly a kinase that phosphorylates Tyr residues in specific target proteins（insulin receptor） Other: synthesize
15、 the i.c. second messenger cGMP in response to ex.c. signals ( the receptor for atrial natriuretic factor),Activation of receptor tyrosine kinases,Insulin receptor tyrosine kinase,Insulin structure,Insulin Receptor Is a Tyr-Specific Protein Kinase 1,Activation of IR Tyr kinase by autophosphorylation
16、,IR Is a Tyrosine-Specific Protein Kinase 2,Insulin receptor binds insulin and undergoes autophosphorylation on its carboxyl-terminal 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 releas
17、e and GTP binding to Ras.,Activated Ras binds and activates Raf-1.,Raf-1 phosphorylates MEK on two Ser residues, activating it. 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.,P
18、hosphorylated Elk1 joins SRF to stimulate the transcription and translation of a set of genes needed for cell division.,Activation of glycogen synthase by insulin,Regulation of blood glucose level,Receptor for atrial natriuretic factor,Two types (isozymes) of guanylyl cyclase that participate in sig
19、nal transduction.,When activated, a guanylyl cyclase produces guanosine 3,5-cyclic monophosphate (cGMP) from GTP,2. Four general types of signal transducers,2.1 Gated Ion Channels2.1.1 Ligand-gated ion channels2.1.2 Voltage-gated ion channels 2.2 Receptor Enzymes 2.3 G ProteinCoupled Receptors and S
20、econd Messengers 2.4 Steroid Receptors,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 generates an intracellular second messenger 3. a guanosine nucleotidebinding protein (G p
21、rotein),Three essential components of G ProteinCoupled Receptors,A protein binds Guanine nucleotides (GDP, GTP); activated in GTP-form, inactivated in GDP-form Integral membrane protein, heterotrimers (); Have similar & subunits, but differ in -subunit When G-protein is activated, the subunit dissoc
22、iates 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),G protein Signal affected enzyme Effect,Gs epinephrine AC stimulatoryglucagonGi catecholamines AC inhib
23、itoryGq Ach, catecholamines PLC stimulatoryGt photons cGMP-PDE stimulatory,Heterotrimeric G protein ( - subunits),AC: adenylyl cyclase; PLC: phospholipase C,G protein (discovery),M. Rodbell: a transducer provided the link between the receptor and the amplifier. A.G. Gilman: identify & purify the G p
24、rotein. System: Mutated lymphoma cells containing a normal receptor and cAMP-generating enzyme, was yet unable to respond (produce cAMP), since it lacked the transducer,mutated cell,normal cell,The Nobel Prize in Physiology and Medicine 1994,“for their discovery of G-proteins and the role of these p
25、roteins in signal transduction in cells“,Alfred G. Gilman,1941-,Martin Rodbell,1925-1998,“ON-OFF” switch is regulated by GTP or GDP bound form. All G-proteins has intrinsic GTPase activity, release Pi and inactivated. Activation: release of GDP and replaced by GTP,Two major systems: 2.3.1 THE PKA SY
26、STEM (cAMP as the second messenger)The -Adrenergic Receptor System 2.3.2 THE PKC SYSTEM (DAG,IP3 and Ca2+ as the second messengers),The association of active Gs with adenylyl cyclase stimulates the cyclase to catalyze cAMP synthesis,Adenosine 3,5-cyclic monophosphate (cAMP),synthesized in adrenal me
27、dulla;belongs to catecholamines (儿茶酚胺）;target cells include liver, skeletal muscle, heart muscle and adipose;released in response to acute stress,Epinephrine 肾上腺素 signal,Epinephrine 肾上腺素 signaling pathway,cAMP,Epinephrine 肾上腺素 signaling pathway (2),Activation of cAMP-dependent protein kinase (PKA),I
28、nactive PKA: Regulatory (R) subunits: auto-inhibitory domains buried catalytic (C) subunits: substrate-binding sites blocked by R subunits,R subunits: autoinhibitory domains buried,Active PKA C subunits open substrate binding sites,A catalytic subunit of PKA,ATP,Potent inhibitor peptide (PKI): Arg-A
29、rg-Gln-Ala-Ile (consensus sequencerecognized by PKAexcept Ala),Epinephrine triggers a series of reactions in hepatocytes in which catalysts activate catalysts, resulting in great “amplification” of the signal,x 分子,10 ,000 x 分子,PKA regulates a number of enzymes The proteins phosohorylated by PKA shar
30、e a region of sequence similarity around the Ser or Thr residue that undergoes phosphorylation, a sequence that marks them for regulation 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.,Desen
31、sitization of the PKA system,1 desensitizing -Adrenergic Receptor2 degrading the second messager,Gsbg recruits bARK to the membrane, 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.,-Arrestin uncoup
32、les the serpentine receptor from its G protein and brings together the three enzymes of the MAPK cascade. (One stimulus triggers two distinct pathways: the path activated by G protein and MAPK cascade),The Protein kinase C (PKC) System,Two intracellular second messengers are produced in the hormone-
33、sensitive phosphatidylinositol system: Inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). Both contribute to the activation of PKC. By raising cytosolic Ca2+, IP3 also activates other Ca2-dependent enzymes; thus Ca2+ also acts as a second messenger,IP3 and DAG,Calcium Is a Second Messenger
34、 in Many Signal Transductions,Normally, Ca2i is 100 nM (而细胞外：1 mM) Hormonal, neural, or other stimuli cause either an influx 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 protei
35、ns that regulate a variety of Ca2-dependent enzymes-Calmodulin (CaM),Some proteins regulated by Calmodulin,The action of a group of compounds known as tumor promoters is attributable to their effects on PKC. E.g., phorbol esters, a potent synthetic PKC activator; apparently mimic cellular DAG as second messengers, but unlike naturally occurring DAGs. they are not rapidly metabolized. By continuously activating PKC, these synthetic tumor promoters interfere with the normal regulation of cell growth and division,Regulation of transcription by steroid hormones,Steroid receptor,
