Molecular regulation of the CRAC channel signaling pathway

CRAC通道信号通路的分子调控

• 批准号: 10711690
• 负责人: Xiaowei Hou
• 金额: $ 40.5万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-03 至 2028-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10711690
• 关键词: Address; Affect; Basic Science; Biochemical; Biochemistry; Calcium; Calcium Channel; Calcium Signaling; Cell membrane; Cell physiology; Cells; Complex; Cryoelectron Microscopy; Disease; Drosophila genus; Endoplasmic Reticulum; Foundations; Genes; Goals; Health; Human; Ion Channel; Knowledge; Lead; Lipids; Mammals; Mediating; Membrane; Methods; Molecular; Pathologic; Pathway interactions; Pharmaceutical Preparations; Physiological; Play; Principal Investigator; Proteins; Regulation; Research; Role; STIM1 gene; Signal Pathway; Signal Transduction; Structure; Tissues; X-Ray Crystallography; cell type; experience; gain of function; gain of function mutation; human disease; innovation; loss of function; loss of function mutation; novel; pharmacologic; programs; protein protein interaction; response; sensor; small molecule; structural biology; therapeutic development; tool

Project Summary/Abstract The calcium (Ca2+) release-activated Ca2+ (CRAC) channels mediate Ca2+ influx in response to Ca2+ store depletion in the endoplasmic reticulum (ER), generating Ca2+ signals critical for many cellular processes. Activation of CRAC channels requires the interaction of Orai and STIM proteins, which are the plasma membrane Ca2+ channel and the Ca2+ sensor in the ER membrane; however, the underlying molecular mechanism remains incompletely understood. Mammals, including humans, have three Orai channels (Orai1, Orai2, and Orai3) and two STIM proteins (STIM1 and STIM2), whereas Drosophila has only one set of Orai and STIM. Both loss- and gain-of-function mutations that underlie human diseases have been identified in human Orai1 and STIM1. The physiological roles of Orai2 and Orai3 are less well understood, but there is emerging evidence showing that they could form heteromeric channels with Orai1, which might play roles distinct from channels formed by Orai1 and STIM1. The long-term goal of my research program is to study the molecular regulation of the CRAC channel signaling pathway in order to understand the molecular mechanisms of the diverse functions the CRAC channels play in different tissues and cells under physiological and pathological conditions. Initially, we are prioritizing structure-function studies of the Drosophila Orai-STIM complex and human CRAC channels to launch my research program. In this proposed research, we aim to address key knowledge gaps in (1) the biochemistry and structural biology of the Orai-STIM interaction, (2) the structure and function of human CRAC channels, and (3) regulation of CRAC channels by cellular factors and channel modulators. We will bring innovative approaches and apply the Principal Investigator’s experience in structural biology and ion channel structure-function studies to address these questions. Specifically, we will determine the atomic structure of the Drosophila Orai-STIM complex as well as structures of human Orai channels and human Orai-STIM complexes. We will also develop novel functional approaches and tools for structure-guided functional analysis of CRAC channels. In addition, we propose to study the regulation of the channel structure and function by small molecule modulators including lipids and pharmacological drugs. Finally, we will use functional, biochemical, and structural biology methods (including both X-ray crystallography and cryo-electron microscopy) to characterize the protein-protein interaction of Orai or STIM with a multitude of endogenous protein regulators. In the next five years, we aim to gain a deeper mechanistic understanding of the regulation of the CRAC channel signaling pathway and construct the foundation of knowledge and research tools to understand and study the broader signaling network in which the CRAC channels reside.

项目总结/摘要 钙释放激活的钙通道（CRAC）介导钙内流 内质网（ER）中的耗竭，产生对许多细胞过程至关重要的Ca 2+信号。 CRAC通道的激活需要奥赖和STIM蛋白的相互作用，它们是质膜 Ca 2+通道和ER膜中的Ca 2+传感器;然而，潜在的分子机制仍然存在 不完全理解。包括人类在内的哺乳动物具有三个奥赖通道（Orai 1、Orai 2和Orai 3）， 两种STIM蛋白（STIM 1和STIM 2），而果蝇只有一组奥赖和STIM。损失-和 已经在人Orai 1和STIM 1中鉴定了构成人类疾病基础的功能获得性突变。的 Orai 2和Orai 3的生理作用还不太清楚，但有新的证据表明， 它们可以与Orai 1形成异聚体通道，其作用可能与Orai 1形成的通道不同 STIM1。我的研究计划的长期目标是研究CRAC通道的分子调控 为了了解CRAC通道不同功能的分子机制， 在生理和病理条件下在不同组织和细胞中发挥作用。首先，我们将优先考虑 果蝇Orai-STIM复合物和人类CRAC通道的结构-功能研究， 研究计划。在这项拟议的研究中，我们的目标是解决关键的知识差距（1）生物化学 Orai-STIM相互作用的结构生物学，（2）人类CRAC通道的结构和功能，以及 (3)通过细胞因子和通道调节剂调节CRAC通道。我们将带来创新方法 并应用首席研究员在结构生物学和离子通道结构-功能研究方面的经验 来解决这些问题。具体来说，我们将确定果蝇Orai-STIM的原子结构， 复合物以及人奥赖通道和人Orai-STIM复合物的结构。我们还将开发 用于CRAC通道的结构引导功能分析的新功能方法和工具。此外，本发明还提供了一种方法， 我们建议研究小分子调节剂对通道结构和功能的调节，包括 脂质和药理学药物。最后，我们将使用功能，生物化学和结构生物学方法 （包括X射线晶体学和冷冻电子显微镜）来表征蛋白质-蛋白质 奥赖或STIM与多种内源性蛋白质调节剂的相互作用。在未来五年，我们的目标是 获得对CRAC通道信号传导途径的调节的更深的机制理解，并构建 知识和研究工具的基础，以了解和研究更广泛的信令网络，其中 CRAC信道驻留。