Molecular Mechanisms of Platelet Exocytosis

血小板胞吐作用的分子机制

基本信息

批准号：
7323371
负责人：
SIDNEY Waldo WHITEHEART
金额：
$ 29.3万
依托单位：
UNIVERSITY OF KENTUCKY
依托单位国家：
美国
项目类别：
财政年份：
1996
资助国家：
美国
起止时间：
1996-08-20 至 2011-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7323371
关键词：
ADP-ribosylation factor 6 Actins Address Adhesions Affect Agonist Alpha Granule Basic Science Binding Biochemical Biological Assay Blood Platelets Blood Vessels Cells Cisplatin/Doxorubicin/Melphalan/Teniposide Clinical Coagulation Process Complex Condition Cytoplasmic Granules Cytoskeleton Data Defect Diagnostic Reagent Disease Elements Event Exocytosis Family Funding GTP-Binding Proteins Goals Guanosine Triphosphate Guanosine Triphosphate Phosphohydrolases Hemorrhage Hemostatic Agents Hemostatic function In Vitro Inflammation Knowledge Learning Lysosomes Mediating Membrane Membrane Fusion Membrane Protein Traffic Membrane Proteins Molecular Molecular Chaperones Molecular Target Myocardial Infarction Nucleotides Pathway interactions Physicians Platelet Activation Play Process Protein Secretion Proteins Recombinant Proteins Relative (related person)Research Research Personnel Rest Role SNAP receptor Signal Transduction Specificity Stimulus Stroke Techniques Thrombosis Time VAMP-2 Vesicle Work Wound Healing base cellubrevin extracellular human VAPA protein insight member novel novel therapeutics programs protein protein interaction receptor receptor function research study response rho rho GTP-Binding Proteins soluble NSF attachment protein syntaxin syntaxin-2 therapeutic target

项目摘要

DESCRIPTION (provided by applicant): Platelet exocytosis is integral to hemostasis and vascular integrity. Conversely, inhibiting exocytosis is an important target for clinically managing improper thrombosis. Thus, understanding the molecular events required for platelet exocytosis will broaden our ability to control platelet function. From previously funded work, it is clear that specific Soluble NSF Attachment Protein Receptors (v- and t-SNAREs) are required for platelet secretion. With this established, our goals have now expanded to address how this exocytosis machinery is regulated. We hypothesize that platelet exocytosis is governed by proteins that control SNARE- SNARE interactions. Three specific aims are proposed: Aim 1) To determine which v-SNARE proteins are required for platelet granule release. Our work demonstrates that VAMP-8/endobrevin is required for platelet exocytosis. The goals of Aim 1 are to complete the characterization of VAMP-8's role, to identify which VAMP (2 or 3) serves as the secondary v-SNARE and to determine when this mechanism is used. Aim 2) To define the pathway of interactions required for t-SNARE function in platelet exocytosis. Previous work assigned roles for SNAP-23, syntaxin 2 and 4. This aim focuses on proteins that regulate these t-SNAREs to promote granule release. The syntaxin chaperones of the Munc18 family and their effectors, Munc13S and Doc2s, will be examined using a combination of immunological and biochemical techniques to determine how these proteins interact during exocytosis. These data will expand our understanding of how the syntaxin t-SNAREs are regulated to pair with appropriate SNAREs and to mediate membrane fusion. Aim 3) To determine how the small GTP-binding protein, ADP-ribosylation factor 6 (Arf6), participates in platelet activation. Members of the Arf family are associated with membrane trafficking, specifically exocytosis. Our work shows that Arf6 is critical to platelet activation, secretion, and actin dynamics. This aim will directly address how changes in Arf6-nucleotide state affect actin cytoskeleton and secretion, and will identify the Arf6 effectors required for those functions. These data will yield insight into the central role of this small GTP- binding protein in activated platelets. Overall the information gained from this proposal will help to identify new therapeutic targets that will be useful for modulating hyperactive platelets, and to provide diagnostic reagents for determining how hypoactive platelets are defective.

描述（由申请人提供）：血小板胞吐作用是止血和血管完整性不可或缺的。相反，抑制胞吐作用是临床管理不当血栓形成的重要目标。因此，了解血小板胞吐作用所需的分子事件将扩大我们控制血小板功能的能力。从先前资助的工作中可以明显看出，血小板分泌需要特定的可溶性NSF附着蛋白受体（V-和T-SNARES）。随着这一既定，我们的目标现在已经扩展，以解决如何调节这种胞吐药机制。我们假设血小板胞吐作用受控制圈圈相互作用的蛋白质的控制。提出了三个具体目标：目标1）确定血小板颗粒释放需要哪些V-SNARE蛋白。我们的工作表明，血小板胞吐作用是必需的VAMP-8/内折。 AIM 1的目标是完成VAMP-8角色的表征，以确定哪种VAMP（2或3）用作次级V-SNARE，并确定何时使用了这种机制。目标2）定义血小板胞吐作用中T-SNARE功能所需的相互作用的途径。先前的工作分配了SNAP-23，语法2和4的作用。此目的集中于调节这些T-snares以促进颗粒释放的蛋白质。将使用免疫学和生化技术的组合来检查Munc18家族的语法伴侣及其效应子Munc13s和Doc2s，以确定这些蛋白质在胞吐作用期间如何相互作用。这些数据将扩展我们对如何调节语法t-snares如何与适当的鼻子配对并介导膜融合的理解。目标3）确定小的GTP结合蛋白ADP-核糖基化因子6（ARF6）如何参与血小板激活。 ARF家族的成员与膜运输，特别是胞吞作用有关。我们的工作表明，ARF6对于血小板激活，分泌和肌动蛋白动力学至关重要。该目标将直接解决ARF6-核苷酸状态的变化如何影响肌动蛋白细胞骨架和分泌，并将确定这些功能所需的ARF6效应子。这些数据将洞悉该小型GTP结合蛋白在活化血小板中的核心作用。总体而言，从该提案中获得的信息将有助于确定新的治疗靶标，这些靶标将有助于调节多动态血小板，并提供诊断试剂，以确定低血压血小板的缺陷。