Functional architecture of IP3-evoked local Ca2+signals

IP3诱发的局部Ca2信号的功能架构

• 批准号: 7089834
• 负责人: JONATHAN S MARCHANT
• 金额: $ 20.56万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 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/7089834
• 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

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）：由三磷酸肌醇受体激活触发的细胞质Ca 2+增加调节各种生理事件。相同的IP 3刺激的Ca 2+释放途径的功能障碍是许多疾病的发病机制的基础。因此，重要的是要了解IP 3受体如何产生生理相关的Ca 2+信号，以及病理事件如何失调IP 3受体的行为。 我们的理解IP 3诱发的Ca 2+信号在完整的细胞的功能组织已被转化的高分辨率成像方法。共聚焦成像显示，IP 3诱发的Ca 2+信号是通过显微Ca 2+信号的渐进募集产生的，称为“Ca 2+喷流”，其被解析为高亲和力Ca 2+指示染料的荧光的瞬时变化。这些功能信号被解释为反映了间隔在整个内质网中的IP 3受体的小簇的活性，但IP 3受体的潜在分布和结构架构尚未得到解决。由于全细胞Ca 2+信号的空间模式取决于Ca 2+释放通道的功能和细胞分布，因此必须了解控制IP 3受体“功能结构”的机制。该建议的目的是定义IP 3受体分布如何影响细胞Ca 2+信号的模式，以测试IP 3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，可以解析活细胞中的IP 3受体分布。我们将使用这些新的工具，以及生物化学和分子的方法来（1）解决IP 3受体结构的基础上不同的时空模式的1 P3-诱发的Ca 2+信号;（2）定义在早期发育期间IP 3受体信号传导的功能结构中的生理变化的机制，以及（3）阐明丙型肝炎病毒蛋白NS 5A诱发的IP 3受体信号传导的功能结构的病理变化机制。研究结果将有助于我们了解无处不在的IP 3信号通路在健康和疾病中的作用。