Dynamics of exocyst recruitment and assembly

外囊招募和组装的动态

• 批准号: 8725193
• 负责人: Derek K. Toomre
• 金额: $ 34.97万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8725193
• 关键词: Address; Affect; Anthron; Binding; Biochemical; Biological Assay; Cell Polarity; Cell division; Cell membrane; Cell physiology; Cell surface; Cilia; Complex; Data; Diabetes Mellitus; Disease; Docking; Exocytosis; Functional disorder; Genetic; Goals; Growth Factor; Hormones; Image; Imagery; In Vitro; Intervention; Knowledge; Lead; Literature; Malignant Neoplasms; Mammalian Cell; Mediating; Modeling; Molecular; Molecular Conformation; Molecular Target; Monitor; Mutation; Neoplasm Metastasis; Phosphorylation; Phosphotransferases; Play; Polycystic Kidney Diseases; Post-Translational Protein Processing; Process; Proteins; Research; Roentgen Rays; Role; SNAP receptor; Seminal; Sensitivity and Specificity; Series; Site; Small Interfering RNA; Structure; Techniques; Testing; Toxin; Vesicle; Work; Yeasts; base; cancer cell; cell motility; ciliopathy; human disease; migration; mimetics; mutant; novel therapeutics; optogenetics; prevent; protein complex; public health relevance; receptor recycling; single molecule; spatial relationship; stoichiometry; treatment strategy

DESCRIPTION (provided by applicant): The long-term objective of these studies is to elucidate the mechanisms by which exocyst recruitment and assembly are involved in vesicle tethering, docking and fusion. The exocyst is an octomeric protein complex (Sec3/5/6/8/10/15/Exo70/84) that spatially targets vesicles to specific sites on the plasma membrane, such as the bud tip in yeast, where it was identified in a series of seminal studies. In mammalian cells it is responsible for a range of polarized cellular processes such as cell division, migration, stabilization of junctions and formation of a primary cilium. Despite the ubiquitous and essential role of the exocyst, there are critical gaps in understanding how the exocyst 'tethering' complex and other putative tethers function to anchor vesicles and coordinate with downstream SNARE-mediated fusion machinery. While tethers are believed to involve long-range interactions, be reversible and regulate the fidelity of vesicle fusion, these concepts are generally untested and there are few assays to monitor the dynamic changes in discrete biophysical and molecular states of the exocyst. Resolving these topics will include addressing the stoichiometry of the mammalian complex, assembly and disassembly of the exocyst holocomplex, and its coordination with the fusion machinery, all of which are key goals in the proposed studies. The specific aims of this study are: 1) to critically test the longstanding "exocyst as a tether" hypothesis and to study the dynamics of its recruitment to vesicles; 2) to address how the exocyst is able to biophysically promote docking and fusion; 3) to address how post-translational modifications of the exocyst affects its stability and regulates its function. These studies will provide new knowledge of how tethers function and establish a new platform to monitor their discrete states. Understanding the basic mechanisms of how the exocyst functions is highly relevant to human diseases, since its dysfunction is associated with metastasis of cancer cells, diabetes, and ciliopathies, and it is pathologically targeted by anthra toxin. A deeper understanding of how the exocyst functions as a tether and how it is regulated may help identify new molecular targets of intervention.

描述（由申请人提供）：这些研究的长期目的是阐明囊泡绑扎，扩展坞和融合的外囊肿募集和组装的机制。外囊肿是一种八体蛋白复合物（Sec3/5/5/6/8/10/15/exo70/84），将囊泡靶向质膜上的特定位点，例如酵母中的芽尖，在酵母中的芽尖一系列开创性研究。在哺乳动物细胞中，它负责一系列极化的细胞过程，例如细胞分裂，迁移，连接稳定和主要纤毛的形成。 尽管外囊肿的无处不在且重要的作用，但在理解外旋转的“束缚”复合物和其他推定的系群如何锚固囊泡并与下游SNARE介导的融合机械坐标方面存在关键差距。虽然据信系群涉及长期相互作用，但是可逆的并调节了囊泡融合的忠诚度，但这些概念通常未经测试，几乎没有测定方法可以监测外囊肿的离散生物物理和分子状态的动态变化。解决这些主题将包括解决哺乳动物络合物的化学计量，外旋蛋白全胶质复合物的组装和拆卸以及与融合机械的协调，所有这些都是拟议的研究中的关键目标。这项研究的具体目的是：1）批判性地测试长期以来的“外囊作为束缚”假设，并研究其募集到囊泡的动力学； 2）解决外囊肿如何能够生物物理促进对接和融合； 3）解决外囊肿的翻译后修饰如何影响其稳定性并调节其功能。这些研究将提供有关Tethers如何运作并建立新平台来监视其离散状态的新知识。 了解外囊肿功能如何与人类疾病高度相关的基本机制，因为它的功能障碍与癌细胞，糖尿病和纤毛病的转移有关，并且由Anthra毒素在病理上靶向。对外囊肿如何充当系绳及其如何调节的更深入了解可能有助于确定新的干预分子靶标。