BETA CELL INHIBITORS ACT BY DISTRUPTING SNARE COMPLEX

β 细胞抑制剂通过破坏圈套复合体发挥作用

• 批准号: 6624814
• 负责人: THOMAS SCHERMERHORN
• 金额: $ 10.83万
• 依托单位: KANSAS STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-03-15 至 2003-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6624814
• 关键词: G protein; biological signal transduction; guanine nucleotide binding protein; immunoprecipitation; insulin inhibitor; intermolecular interaction; membrane proteins; pancreatic islet function; syntaxin

DESCRIPTION (taken from the application) The candidate is a veterinarian who has completed three years of postgraduate clinical training and will soon complete a Ph.D. degree. The candidate's short-term goal is to obtain additional research training in an environment that will foster the skills needed to become an independent research scientist. During that time the candidate seeks a limited clinical commitment in order to maintain the clinical skills and focus that will be needed to achieve his long-term objective. Cornell's College of Veterinary Medicine is an ideal site for the candidate to pursue his immediate goals as the training environment in the Department of Molecular Medicine is superb and the clinical facilities are state-of-the-art. The candidate's long-term goal is to obtain a faculty position at a veterinary college and to develop a comprehensive research program that encompasses both basic and clinical research. In this application, the candidate proposes to investigate the effects of inhibitors of pancreatic beta cell secretion on the proteins directly involved in insulin exocytosis. Specifically, the hypothesis that inhibition of exocytosis in beta cells is due to disruption of the core complex will be tested. This unique signaling pathway will be studied using a variety of techniques to identify and to characterize the molecular interactions that produce the powerful inhibitory effect. A likely target for inhibitor action is the "core complex" of proteins, composed of synaptobrevin, syntaxin, and SNAP 25. Together these proteins form a tightly bound helical structure that tethers insulin-containing granules to the islet cell membrane. The core complex is an essential intermediate in exocytosis and is regulated by a large number of cellular proteins. Inhibitor-induced changes in the core complex composition will be detected using a novel coimmunoprecipitation assay (the "core complex assay"). The cellular mechanisms responsible for producing the core complex effects of the inhibitors will also be characterized. First, the inhibitory GTP-binding protein involved will be identified using co-immunoprecipitation, antisense, and permeabilization techniques. Second, the core complex target for the activated G protein pathway will be determined using co-immunoprecipitation and pull-down assays, clostridial neurotoxin cleavage experiments, and over-expression studies. Finally, characterization of the essential molecular features of target proteins will be done to elucidate the direct molecular interactions.

描述(取自应用程序) 应聘者是一名完成了三年研究生课程的兽医 他将接受临床培训，并将很快完成博士学位。候选人的 短期目标是在一个环境中获得额外的研究培训 这将培养成为一名独立研究科学家所需的技能。 在此期间，应聘者寻求有限的临床承诺，以便 保持实现HIS所需的临床技能和专注 长期目标。康奈尔大学兽医学院是一个理想的选址 对于应聘者来说，在培训环境中追求他的近期目标 分子医学系是一流的，临床设施是 最先进的。这位候选人的长期目标是获得一名教师 在兽医学院的职位，并开展一项全面的研究 包括基础研究和临床研究的计划。在此应用程序中， 候选人建议研究胰腺抑制物的作用。 β细胞分泌的蛋白质直接参与胰岛素的胞吐作用。 具体地说，认为抑制β细胞胞吐作用是由于 对核心复合体的破坏将进行测试。这一独特的信号通路 将使用各种技术进行研究，以识别和表征 产生强大抑制作用的分子相互作用。一个 抑制剂作用的可能靶点是蛋白质的“核心复合体”，由 Synaptobrevin、Synaxin和SNAP 25。这些蛋白质结合在一起形成了一个紧密的 将含胰岛素颗粒捆绑在胰岛上的螺旋结构 细胞膜。核心复合体是胞吐作用的重要中间体， 是由大量细胞蛋白质调控的。抑制物引起的变化 在核心，复杂的成分将被检测到使用一种新的 免疫共沉淀试验(“核心复合体试验”)。细胞机制 负责产生核心复合作用的抑制剂也会 被刻画出来。首先，涉及的抑制性GTP结合蛋白将是 免疫共沉淀、反义和渗透鉴定 技巧。第二，激活的G蛋白途径的核心复合体靶点 将使用免疫共沉淀法和下拉试验进行测定， 梭状芽胞杆菌神经毒素裂解实验和过度表达研究。 最后，对靶标的基本分子特征进行表征 蛋白质将被用来阐明直接的分子相互作用。