Functional architecture of IP3-evoked local Ca2+signals

IP3诱发的局部Ca2信号的功能架构

基本信息

批准号：
6823620
负责人：
JONATHAN S MARCHANT
金额：
$ 20.34万
依托单位：
UNIVERSITY OF MINNESOTA
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-07-01 至 2008-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6823620
关键词：
Xenopus oocyte calcium flux calcium indicator carbohydrate receptor cellular pathology chimeric proteins endoplasmic reticulum fluorescent dye /probe hepatitis C virus inositol phosphates nonmammalian vertebrate embryology protein localization protein structure function virus protein

项目摘要

DESCRIPTION (provided by applicant): Cytoplasmic Ca2+ increases triggered by activation of inositol trisphosphate receptors regulate a huge variety of physiological events. Dysfunction of the same IP3-stimulated Ca2+ release pathway underlies the pathogenesis of many disorders. Therefore, it is important to understand how IP3 receptors generate physiologically relevant Ca2+ signals, and how pathological events dysregulate IP3 receptor behavior. Our understanding of the functional organization of IP3-evoked Ca2+ signals in intact cells has been transformed by high resolution imaging methods. Confocal imaging has shown that IP3-evoked Ca2+ signals are generated by progressive recruitment of microscopic Ca2+ signals, known as 'Ca2+ puffs', which are resolved as transient changes in fluorescence of high affinity Ca2+ indicator dyes. These functional signals are interpreted as reflecting the activity of small clusters of IP3 receptors spaced throughout the endoplasmic reticulum, but the underlying distribution, and structural architecture of IP3 receptors is unresolved. As spatial patterning of whole cell Ca2+ signals depends on both the function and the cellular distribution of Ca2+ release channels, it is imperative to understand the mechanisms that control the 'functional architecture' of IP3 receptors. The aim of this proposal is to define how IP3 receptor distribution impacts the patterning of cellular Ca2+ signals to test the central hypothesis that IP3 receptor architecture is modulated by physiological and pathological cues. To achieve this goal, we have optimized fluorescent protein-based tools that resolve IP3 receptor distribution in live cells. We will use these novel tools, as well as biochemical and molecular approaches to (1) resolve IP3 receptor architecture underlying different spatiotemporal patterns of 1P3-evoked Ca2+ signaling; (2) define the mechanisms of physiological change in the functional architecture of lP3 receptor signaling during early development and (3) delineate the mechanisms of pathological change in the functional architecture of lP3 receptor signaling evoked by the hepatitis C viral protein NS5A. Results will aid our understanding of the role of the ubiquitous IP3 signaling pathway in both health and disease.

描述（由申请人提供）：由肌醇三磷酸受体激活引发的细胞质Ca2+增加调节多种生理事件。 IP3 刺激的 Ca2+ 释放途径的功能障碍是许多疾病的发病机制的基础。因此，了解 IP3 受体如何产生生理相关的 Ca2+ 信号，以及病理事件如何失调 IP3 受体行为非常重要。高分辨率成像方法改变了我们对完整细胞中 IP3 诱发的 Ca2+ 信号功能组织的理解。共聚焦成像表明，IP3 诱发的 Ca2+ 信号是通过逐步募集微观 Ca2+ 信号（称为“Ca2+ 泡芙”）产生的，这些信号被解析为高亲和力 Ca2+ 指示剂染料荧光的瞬时变化。这些功能信号被解释为反映了遍布内质网的 IP3 受体小簇的活性，但 IP3 受体的潜在分布和结构体系尚未解决。由于全细胞 Ca2+ 信号的空间模式取决于 Ca2+ 释放通道的功能和细胞分布，因此必须了解控制 IP3 受体“功能结构”的机制。该提案的目的是定义 IP3 受体分布如何影响细胞 Ca2+ 信号的模式，以测试 IP3 受体结构受生理和病理线索调节的中心假设。为了实现这一目标，我们优化了基于荧光蛋白的工具，可解析活细胞中 IP3 受体的分布。我们将使用这些新工具以及生化和分子方法来（1）解析 1P3 诱发的 Ca2+ 信号传导的不同时空模式下的 IP3 受体结构； (2) 定义早期发育过程中lP3受体信号传导功能结构的生理变化机制；(3) 描述由丙型肝炎病毒蛋白NS5A引起的lP3受体信号传导功能结构的病理变化机制。结果将有助于我们了解普遍存在的 IP3 信号通路在健康和疾病中的作用。