Regulation of Platelet Integrin Signaling in Hemostasis and Thrombosis

描述(申请人提供)：整合素对细胞外配体的亲和力在血小板中受到严格调控。当血管完整性被破坏时，血小板整合素发生构象变化，导致与配体的亲和力增加，称为整合素激活，从而支持止血所需的血小板黏附、聚集和血栓形成。未能适当地调节血小板中的整合素活性会促进血栓性疾病。该项目致力于Talin-整合素相互作用的调节，其基本原理是选择性地调节它们的相互作用可以提供一种新的抗血栓治疗策略。这一概念将在Aim 1中通过分析表达talin1突变体的小鼠来测试，这些突变体选择性地破坏talin激活整合素的能力，而不影响Ten与体内整合素或其他已知talin结合蛋白的结合。目标2将调查细胞和 分子机制，主要集中在双向整合素信号，这是目标1中表征的表型的基础。在目标3中，talin和kindlin的细胞分布将是 通过荧光显微镜对血小板进行检查，将剖析调节talin和kindlin重新分布到质膜的信号通路。总之，这些研究将提供关于整合素-talin相互作用在止血和血栓形成中的生物学意义的新信息，并为调节血小板激动剂诱导的控制整合素激活的蛋白质的动态重新分布的信号通路提供见解。我们提议的实验与NHLBI的使命相一致，共同可能指向预防和治疗心脏病发作和中风的新疗法。

DESCRIPTION (provided by applicant): The affinity of integrins for extracellular ligands is tightly regulated in platelets. Upon disruption of vascular integrity, platelet integrins undergo conformational changes that result in increased affinity for ligand, termed integrin activation, thereby supporting platelet adhesion, aggregation, and thrombus formation necessary for hemostasis. Failure to properly regulate integrin activity in platelets can promote thrombotic disease. This project focuses on the regulation of talin-integrin interactions with the rationale that selectively modulating their interactions could provide a novel anti-thrombotic therapeutic strategy. This concept will be tested in Aim 1 by analyzing mice expressing talin1 mutants that selectively disrupt the capacity of talin to activate integrins without affecting the binding of tain to integrins or other known talin-binding proteins in vivo. Aim 2 will investigate the cellular and molecular mechanisms, focused primarily on bidirectional integrin signaling, that underlie the phenotypes characterized in Aim 1. In Aim 3, the cellular distribution of talin and kindlin will be examined in platelets by fluorescence microscopy and signaling pathways that regulate talin and kindlin redistribution to the plasma membrane will be dissected. Together, these studies will provide new information about the biological significance of integrin-talin interactions in hemostasis and thrombosis and provide insights into the signaling pathways that regulate the platelet agonist-induced dynamic redistribution of proteins that control integrin activation. Together our proposed experiments, consistent with the mission of the NHLBI, may point to novel therapies for the prevention and treatment of heart attack and stroke.