PLATELET CYTOSKELETAL ASSEMBLY AND FUNCTION

血小板细胞骨架的组装和功能

• 批准号: 3347343
• 负责人: ROGER C CARROLL
• 金额: $ 7.42万
• 依托单位: UNIVERSITY OF TENNESSEE KNOXVILLE
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-12-01 至 1986-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3347343
• 关键词: crosslink; cytoskeleton; gel electrophoresis; hemostasis; human tissue; membrane activity; phosphorylation; platelet aggregation inhibitors; platelets; polymerization; protein kinase; radiotracer

Our hypothesis is that the platelet response (shape change, aggregation, secretion and procoagulant activity) to activators may primarily involve the regulation of cytoskeletal assembly (actin polymerization and interaction with myosin, actin-binding protein, and Alpha-actinin-like protein) and the transmembrane interaction of these cytoskeletal assemblies with surface membrane proteins. Such transmembrane interaction would control the expression of aggregation, adhesion, and platelet procoagulant activity (via exposure of high affinity factor Va receptors). It is proposed to test this hypothesis by coordinately examining actin polymerization (by a DNAase I inhibition assay for monomeric actin), in situ actin-binding protein and myosin light chain phosphorylation (total platelet sample denatured and resolved on SDS-PAGE), composition on SDS-PAGE of cytoskeletal cores resistant to detergent extraction due to crosslinking by associated cytoskeletal proteins, and the association of radioiodinated surface membrane proteins with such cores. These processes will be modified in situ by specific inhibitors of platelet phosphorylation, (aspirin, PGI2, dibutyrul cAMP, calmodulin in hibitors) and cytoskeletal protein assembly, (cytochalasin B, phorbol myristate acetate). In vitro experiments with isolated actin-binding protein will address the possible role of phosphorylation in regulation of pseudopodal cytoskeletal assembly. Our long term objectives are to understand what regulates platelet cytoskeletal assembly and function, the nature of transmembrane interaction, and how such transmembrane interactions might control critical hemostatic platelet responses.

我们的假设是，血小板反应（形状变化，聚集， 分泌和促凝血活性）对激活剂的作用可能主要涉及 细胞骨架组装（肌动蛋白聚合和 与肌球蛋白、肌动蛋白结合蛋白和α-肌动蛋白样蛋白相互作用 蛋白质）和这些细胞骨架组装体的跨膜相互作用 表面膜蛋白。 这种跨膜相互作用将 控制聚集、粘附和血小板促凝剂的表达 活性（通过暴露高亲和力因子Va受体）。 是 我建议通过协同检查肌动蛋白来检验这一假设 聚合（通过单体肌动蛋白的DNA酶I抑制测定）， 原位肌动蛋白结合蛋白和肌球蛋白轻链磷酸化（总 血小板样品变性并在SDS-PAGE上解析）， 细胞骨架核心的SDS-PAGE，由于 通过相关的细胞骨架蛋白交联，以及 放射性碘标记的具有这种核心的表面膜蛋白。 这些过程 将被血小板特异性抑制剂原位修饰 磷酸化，（阿司匹林、PGI 2、二丁酸cAMP、钙调蛋白抑制剂） 和细胞骨架蛋白组装（细胞松弛素B、佛波醇肉豆蔻酸酯 乙酸盐）。 用分离的肌动蛋白结合蛋白进行的体外实验将 解决磷酸化在伪足调节中的可能作用， 细胞骨架装配 我们的长期目标是了解 调节血小板细胞骨架组装和功能， 跨膜相互作用，以及这种跨膜相互作用如何可能 控制关键的止血血小板反应。