Ion Channels and Signaling Mechanisms In T Lymphocytes

T 淋巴细胞中的离子通道和信号传导机制

• 批准号: 6825865
• 负责人: RICHARD S LEWIS
• 金额: $ 37.95万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6825865
• 关键词: T cell receptor; T lymphocyte; biological signal transduction; calcium channel; calcium flux; calcium ion; cell membrane; chemical kinetics; clinical research; confocal scanning microscopy; electrophysiology; endocytosis; flow cytometry; fluorescence resonance energy transfer; gene induction /repression; human subject; human tissue; leukocyte activation /transformation; mitochondria; potassium channel; protein kinase; reporter genes; tissue /cell culture; transcription factor; voltage /patch clamp

DESCRIPTION (provided by applicant): The elevation of intracellular free Ca2+ concentration is an essential signal controlling the differentiation and functions of T lymphocytes. The long-term goal of this proposal is to elucidate the molecular and cellular mechanisms that generate and shape Ca2+ signals in T cells. The Ca2+ release activated Ca2+ (CRAC) channel plays a central role in this process. These channels open in response to the depletion of the Ca2+ stored in the endoplasmic reticulum (ER), but the mechanism linking store depletion to channel activation is not well understood. In addition, Ca2+ entering the cell through CRAC channels interacts with mitochondria and Ca2+ pumps in the plasma membrane to create prolonged and complex Ca2+ signals. Recent developments in fluorescent Ca2+ indicators and fluorescence microscopy have made it possible to address these mechanisms at a subcellular level and visualize microscopic sites of CRAC channel activity. We will address functional interactions between CRAC channels and the ER, mitochondria, and pumps using a combination of approaches, including patch-clamp recording, targeted genetically-encoded Ca2+ indicators, and total internal reflection fluorescence microscopy (TIR-FM). We will distinguish between two models of CRAC activation: physical contact with proteins in the ER, and activation via a diffusible messenger released from the ER. We will explore the role of ER fragmentation in the mechanism of CRAC channel inactivation by Ca2+, and the mechanisms for clustering Ca2+ pumps in the plasma membrane and bringing them close to CRAC channels for functional interaction. The significance of this project is two-fold. First, the results will help to clarify fundamental issues related to store-operated channels and calcium signaling in a wide variety of non-excitable cells. Second, the results of these studies may help identify novel targets for the control of the immune response that may be beneficial in treating autoimmune disorders or immunodeficiencies, and they may help explain immune dysfunction resulting from aberrant operation of the Ca signaling machinery.

描述（由申请人提供）：细胞内游离Ca 2+浓度的升高是控制T淋巴细胞分化和功能的重要信号。该提案的长期目标是阐明在T细胞中产生和形成Ca 2+信号的分子和细胞机制。Ca 2+释放激活的Ca 2+（CRAC）通道在这一过程中起着核心作用。这些通道响应于内质网（ER）中储存的Ca 2+的耗尽而打开，但是将储存耗尽与通道激活联系起来的机制还不清楚。此外，通过CRAC通道进入细胞的Ca 2+与线粒体和质膜中的Ca 2+泵相互作用，以产生延长的复杂Ca 2+信号。荧光Ca 2+指示剂和荧光显微镜的最新发展使得在亚细胞水平上解决这些机制并可视化CRAC通道活动的微观位点成为可能。我们将解决CRAC通道和ER，线粒体和泵之间的功能相互作用，使用的方法，包括膜片钳记录，有针对性的遗传编码的Ca 2+指标，和全内反射荧光显微镜（TIR-FM）的组合。我们将区分CRAC激活的两种模型：与ER中蛋白质的物理接触，以及通过从ER释放的可扩散信使激活。我们将探索ER片段化在Ca 2+灭活CRAC通道机制中的作用，以及在质膜中聚集Ca 2+泵并使其靠近CRAC通道进行功能相互作用的机制。这个项目的意义是双重的。首先，这些结果将有助于澄清与各种非兴奋细胞中的钙库操纵通道和钙信号相关的基本问题。其次，这些研究的结果可能有助于确定新的目标，用于控制免疫反应，这可能有利于治疗自身免疫性疾病或免疫缺陷，他们可能有助于解释免疫功能障碍所造成的钙信号传导机制的异常操作。