CA INFLUX FACTOR-- STRUCTURE/FUNCTION & ROLE IN DIABETES

CA 流入因子——结构/功能

• 批准号: 6517584
• 负责人: Richard Banfield Marchase
• 金额: $ 32.29万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-06-01 至 2004-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6517584
• 关键词: T lymphocyte; Xenopus oocyte; bioassay; biological information processing; biological signal transduction; calcium channel; calcium flux; carbohydrate biosynthesis; endoplasmic reticulum; hexosamines; human tissue; hyperglycemia; insulin dependent diabetes mellitus; interleukin 2; leukocyte activation /transformation; mass spectrometry; membrane transport proteins; nuclear magnetic resonance spectroscopy; protein biosynthesis; protein kinase C; protein structure function; tissue /cell culture; voltage /patch clamp

Numerous signal transduction pathways are dependent upon agonist-induced increases in cytoplasmic free Ca2+. The initial phase of this response is often due to the generation of inositol 1,4,5-trisphosphate and a subsequent release of Ca2+ from the endoplasmic reticulum (ER). The depletion of ER Ca2+ stores results in an influx of extracellular Ca2+ into the cytoplasm, a process termed capacitative or store-operated Ca2+ entry (SOCE). SOCE is implicated in a remarkable array of biological processes, including T cell activation, the secretion of insulin, vasoconstriction, and heart development. One hypothesis that has emerged to explain the transmission of information regarding ER Ca2+ depletion to the plasma membrane proposes a critical role for a novel diffusible messenger molecule, Ca2+ influx factor (ClF). In the last year data that have been gathered by this group of investigators provide strong support for the conclusion that CIF exists, that it is synthesized upon ER Ca2+ store depletion, and that it is responsible for activation of two distinct store-operated plasma membrane Ca2+ channels. Imaging and patch-clamp techniques for CIF's function have provided unique and independently verified data on mammalian CIF. In addition, this group has determined that Saccharomyces cerevisiae that are genetically deficient in an organellar Ca2+ ATPase also make a CIF that to date is indistinguishable from its mammalian counterpart. This inexpensive and plentiful source of material, the purification schemes for CIF that are well underway, and the assays that have been developed place this group in a unique position to purify and structurally characterize this novel signaling molecule. Experiments also will be performed to define the biosynthetic pathway for generating CIF upon ER Ca2+ store depletion, and to define the mechanism by which CIF activates the two plasma membrane Ca2+ + channels that contribute to SOCE. The information that we gain will be applied to a major health problem, diabetes, which mischaracterized by an underlying impairment of SOCE. The hyperglycemia-induced activation of the hexosamine biosynthetic pathway and/or protein kinase C will be investigated as possible regulators of SOCE. The described approach will involve a cross-disciplinary, multi-investigator initiative that applies biologic, biochemical, and structural procedures to this problem. Successful completion of these studies will provide pivotal information on SOCE and may define novel therapeutic targets for diabetes.

许多信号转导途径依赖于 激动剂诱导细胞质游离 Ca2+ 增加。本次活动的初始阶段 反应通常是由于肌醇 1,4,5-三磷酸的生成和 随后从内质网 (ER) 释放 Ca2+。耗尽 ER Ca2+ 储存导致细胞外 Ca2+ 流入 细胞质，这一过程称为电容性或钙库操作的 Ca2+ 进入 (SOCE)。 SOCE 涉及一系列显着的生物过程，包括 T 细胞活化、胰岛素分泌、血管收缩和心脏 发展。为解释病毒传播而出现的一种假设 有关质膜 ER Ca2+ 耗竭的信息提出了 新型可扩散信使分子 Ca2+ 流入因子的关键作用 (ClF)。该小组去年收集的数据 调查人员对 CIF 存在的结论提供了强有力的支持，即 它是在 ER Ca2+ 储备耗尽时合成的，并且负责 激活两个不同的钙池操纵的质膜 Ca2+ 通道。 成像和膜片钳技术为 CIF 功能提供了独特且 独立验证的哺乳动物 CIF 数据。此外，本组还有 确定酿酒酵母在遗传上有缺陷 细胞器 Ca2+ ATP 酶也产生 CIF，迄今为止与 它的哺乳动物对应物。这种廉价且丰富的材料来源， 正在进行中的 CIF 纯化方案，以及 已开发出使该组处于净化和纯化的独特位置 结构表征了这种新型信号分子。实验也会 进行以确定在 ER Ca2+ 上生成 CIF 的生物合成途径 存储耗尽，并定义 CIF 激活这两个机制的机制 质膜 Ca2+ + 通道有助于 SOCE。该信息表明 我们的成果将应用于解决一个主要的健康问题——糖尿病， 由于 SOCE 的潜在损伤而被错误表征。高血糖引起的 己糖胺生物合成途径和/或蛋白激酶 C 的激活将 作为 SOCE 的可能监管者进行调查。所描述的方法将 涉及跨学科、多研究者的倡议，适用 该问题的生物学、生化和结构程序。成功的 这些研究的完成将为 SOCE 提供关键信息，并可能 确定糖尿病的新治疗靶点。