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: 6228873
• 负责人: THOMAS SCHERMERHORN
• 金额: $ 2.25万
• 依托单位: CORNELL 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 2001-06-10
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6228873
• 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） 该候选人是一名兽医，已经完成了三年的研究生学习 临床培训，并将很快完成博士学位。℃下候选人的 短期目标是在一个环境中获得额外的研究培训， 这将培养成为独立研究科学家所需的技能。 在此期间，候选人寻求有限的临床承诺，以便 保持临床技能和重点，将需要实现他的 长期目标。康奈尔大学兽医学院是一个理想的地点 为了让候选人追求他的直接目标， 分子医学系是一流的，临床设施是 候选人的长期目标是获得一个教师 在兽医学院的职位，并制定一个全面的研究 该计划包括基础和临床研究。在本申请中， 候选人建议研究胰腺癌抑制剂的作用， β细胞分泌的蛋白质直接参与胰岛素胞吐。 具体来说，抑制β细胞胞吐作用是由于 核心复合体的破坏将受到考验。这种独特的信号通路 将使用各种技术进行研究， 产生强大抑制作用的分子相互作用。一 抑制剂作用的可能靶点是蛋白质的“核心复合物”， synaptobrevin，syntaxin，和SNAP 25。这些蛋白质一起形成一个紧密的 将含胰岛素的颗粒束缚在胰岛上的束缚螺旋结构 细胞膜核心复合物是胞吐作用的重要中间体， 受到大量细胞蛋白的调控。抑制剂诱导的变化 在核心复杂的组成将被检测到使用一种新的 免疫共沉淀测定（“核心复合物测定”）。的细胞机制 负责产生抑制剂的核心复合物效应的分子也将 被定性。首先，所涉及的抑制性GTP结合蛋白将被 使用免疫共沉淀、反义和透化鉴定 技术.第二，核心复合物靶点为活化的G蛋白通路 将使用免疫共沉淀和下拉测定来确定， 梭菌神经毒素切割实验和过表达研究。 最后，对靶分子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。