Functional G Protein pathways in platelet activation

血小板激活中的功能性 G 蛋白途径

• 批准号: 7201656
• 负责人: Satya P. Kunapuli
• 金额: $ 49.48万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-01 至 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7201656
• 关键词: ADP Receptors; Agonist; Applications Grants; Aspirin; Biochemical; Bleeding time procedure; Blood Platelets; Complement; Cytoplasmic Granules; Data; EEF1A2 gene; Event; Fibrinogen; G-Protein Signaling Pathway; GTP-Binding Proteins; Generations; Hemostatic function; Human; Injury; Integrins; Knockout Mice; Mediating; Modeling; Molecular Genetics; Mus; Pathway interactions; Phospholipase A2; Phospholipase C; Phosphorylation; Phosphotransferases; Physiological; Platelet Activation; Play; Protein Isoforms; Protein Tyrosine Kinase; Proteinase-Activated Receptors; Reaction; Receptor Activation; Relative (related person); Research Personnel; Role; Second Messenger Systems; Shapes; Signal Pathway; Signal Transduction; Signaling Molecule; Testing; Thrombin; Thrombosis; Thromboxane A2; Thromboxane Receptor; Thromboxanes; Thrombus; Transgenes; Transgenic Mice; Tyrosine; Tyrosine Phosphorylation; Work; YWHAQ gene; ferric chloride; in vivo; knockout gene; programs; protein kinase C-delta; second messenger; src-Family Kinases; thrombin receptor peptide (42-47)

DESCRIPTION (provided by applicant): Platelet activation plays a major role in hemostasis and thrombosis. Platelet agonists cause shape change, fibrinogen receptor activation, dense granule release, and thromboxane A2 (TXA2) generation, leading to the activation of other platelets. The mechanisms regulating these platelet physiological events have not been completely understood. All the platelet agonists, either directly or indirectly, depend on G protein pathways to cause platelet activation. We propose to further understand the downstream events in the G protein pathways and their second messengers in platelet activation using complementary biochemical, pharmacological, and gene knockout approaches. ADP and thrombin differ in their ability to activate platelets. ADP fails to cause dense granule release in aspirin-treated platelets, whereas thrombin can. ADP depends on integrin signaling to activate phospholipase A2, whereas thrombin does not. Whereas both these agonists activate Gq-phospholipase C pathways, only thrombin stimulates G12/13 pathways. This grant application is built upon our recent studies demonstrating important roles for a) G12/13 pathways in platelet fibrinogen receptor activation, b) protein kinase C delta isoform in thromboxane generation, and c) Gi pathways in Akt phosphorylation in platelets. We will test the hypothesis that G12/13 pathways contribute to dense granule release, TXA2 generation, and Akt phosphorylation and activation, using pharmacological approaches complemented with platelets from Galpha12 and Galpha13 gene knockout mice and constitutively active Galpha12 and Galpha13 transgenic mice. We will evaluate the relative contributions of Gq/PLC pathways and G12/13 pathways to agonist-induced dense granule release, TXA2 generation, and Akt phosphorylation. We also hypothesize that HAX-1, HS-1, and Src family kinases are activated downstream of G12/13 pathways, which play an important role in platelet activation. We will delineate some of these signaling molecules downstream of the G12/13 pathways, as this pathway is the least understood in platelets. Finally, we will evaluate the functional role of HS1 in platelets using mice-deficient in HS1 in ex vivo platelet functional studies and in vivo thrombosis models. We have strong preliminary data supporting each of the above specific aims. These studies will enhance our understanding of the signaling pathways and their role in platelet activation, and might identify potential newer targets for the treatment of thrombosis.

描述（由申请方提供）：血小板活化在止血和血栓形成中起主要作用。血小板激动剂引起形状改变、纤维蛋白原受体活化、致密颗粒释放和血栓素A2（TXA 2）生成，从而导致其它血小板的活化。调节这些血小板生理事件的机制尚未完全了解。所有的血小板激动剂，直接或间接地，依赖于G蛋白途径引起血小板活化。我们建议进一步了解下游事件中的G蛋白通路和它们的第二信使血小板活化使用互补的生化，药理学和基因敲除的方法。ADP和凝血酶激活血小板的能力不同。ADP不能引起阿司匹林处理的血小板释放致密颗粒，而凝血酶可以。ADP依赖于整合素信号传导来激活磷脂酶A2，而凝血酶不依赖于整合素信号传导来激活磷脂酶A2。尽管这两种激动剂都激活Gq-磷脂酶C途径，但只有凝血酶刺激G12/13途径。该资助申请建立在我们最近的研究基础上，这些研究证明了a）G12/13途径在血小板纤维蛋白原受体活化中的重要作用，B）蛋白激酶C δ亚型在血栓素生成中的重要作用，以及c）Gi途径在血小板Akt磷酸化中的重要作用。我们将测试的假设，即G12/13途径有助于致密颗粒释放，TXA 2的产生，Akt磷酸化和激活，使用药理学方法补充血小板从Galpha 12和Galpha 13基因敲除小鼠和组成性活跃的Galpha 12和Galpha 13转基因小鼠。我们将评估Gq/PLC途径和G12/13途径对激动剂诱导的致密颗粒释放、TXA 2产生和Akt磷酸化的相对贡献。我们还假设HAX-1、HS-1和Src家族激酶在G12/13通路的下游被激活，这在血小板活化中起重要作用。我们将描述G12/13通路下游的一些信号分子，因为这种通路在血小板中是最不了解的。最后，我们将评估HS 1在血小板中的功能作用，使用小鼠缺乏HS 1在体外血小板功能研究和体内血栓形成模型。我们有强有力的初步数据支持上述每一个具体目标。这些研究将增强我们对信号通路及其在血小板活化中的作用的理解，并可能确定治疗血栓形成的潜在新靶点。