ROLE OF C-KINASE IN PLATELET ARACHIDONATE RELEASE

C-激酶在血小板花生四烯酸释放中的作用

• 批准号: 3471188
• 负责人: STEPHEN P HALENDA
• 金额: $ 8.31万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-04-01 至 1993-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3471188
• 关键词: G protein; arachidonate; autoradiography; bacterial toxins; biological information processing; biological signal transduction; calcium; calcium flux; cell free system; eicosanoid metabolism; enzyme mechanism; enzyme substrate; gel electrophoresis; hemostasis; human subject; phospholipase A2; phospholipase inhibitor; phosphorylation; platelets; protein kinase C; radiotracer; terpene saponin; thrombin; thrombosis; thromboxanes; tritium

Aside from its crucial role in the normal hemostatic process, the blood platelet is firmly implicated in the pathogenesis of thrombosis and other vascular diseases; thus it is important to understand the cellular mechanisms which regulate platelet function. Formation of the key platelet activator thromboxane A2 during platelet aggregation is limited by the availability of its chemical precursor, arachidonic acid. This project seeks to clarify the poorly understood mechanism by which arachidonic acid mobilization in human platelets is governed. The roles of three major intracellular signals, Ca2+, the Ca2+/phospholipid-dependent protein kinase (C-kinase), and guanine nucleotide-binding proteins (N-proteins) in the cellular mechanism of arachidonic acid release, will be studied in human platelets in experiments involving intact cells, permeabilized platelets, and platelet-derived subcellular preparations. Previous studies in intact platelets which suggest an involvement of C- kinase in Ca2+-dependent arachidonic acid release will be extended by examining the direct effect of purified C-kinase on the activity of phospholipase A2, the major lipolytic enzyme responsible for arachidonic acid liberation in stimulated platelets. Identification of the C-kinase substrate which mediates activation of phospholipase A2 will be sought. The possible involvement of endogenous phospholipase A2 inhibitors (lipocortins) and their regulation by C-kinase will be studied. And finally, the role of N- proteins will be investigated by using guanine nucleotides and pertussis toxin in permeabilized platelets and studying the effect of purified N-protein subunits on platelet phospholipase A2. Possible interactions among the different signal transducers (Ca2+, C-Kinase, N-proteins) for control of arachidonic acid release will be examined. The goals of this project and future studies are: (1) to identify the cellular factors which control the liberation of arachidonic acid, the precursor for prostaglandins, thromboxanes, and other biologically important substances; and (2) more broadly, to define the intra- and intercellular signalling pathways that regulate platelet aggregation and secretion, as these events are centrally involved in hemostasis and thrombosis.

除了其在正常止血过程中的关键作用外， 血小板牢固地与 血栓形成和其他血管疾病；因此，重要的是 了解调节血小板的细胞机制 功能。 钥匙血小板激活剂血栓烷的形成 血小板聚集期间的A2受到其可用性的限制 化学前体，花生四烯酸。 这个项目试图 阐明蛛网膜的理解不足的机制 控制人血小板中的酸动员。 三个主要细胞内信号Ca2+的作用， Ca2+/磷脂依赖性蛋白激酶（C-激酶）和 鸟嘌呤核苷酸结合蛋白（N蛋白）在细胞中 将在人类中研究蛛网膜酸释放的机理 涉及完整细胞的实验中的血小板，透化 血小板和血小板衍生的亚细胞制剂。 以前的 完整血小板的研究表明C-参与 Ca2+依赖性的花生四烯酸释放中的激酶将是 通过检查纯化的C-激酶对 磷脂酶A2的活性，主要脂肪酶 负责刺激血小板中的花生四烯酸释放。 识别介导活化的C-激酶底物 将寻求磷脂酶A2的含量。 可能参与 内源性磷脂酶A2抑制剂（Lipocortins）及其它们的 将研究C-激酶的调节。 最后，n-的作用 将使用鸟嘌呤核苷酸和 透化血小板中的百日咳毒素并研究效果 血小板磷脂酶A2上纯化的N蛋白亚基。 不同信号传感器之间的可能相互作用 （Ca2+，C-激酶，N蛋白）用于控制蛛网酸 将检查发布。 该项目和未来研究的目标是：（1）确定 控制蛛网膜解放的细胞因子 酸，前列腺素，血栓烷和其他的前体 生物学上重要的物质； （2）更广泛地定义 调节的细胞内和细胞间信号通路 血小板聚集和分泌，因为这些事件是集中的 参与止血和血栓形成。