Communication neighboring ryanodine receptor channels

描述（由申请人提供）：本项目的目的是了解骨骼肌兰尼碱受体通道（RyRs）的偶联功能。RyR是位于细胞内钙库膜上的细胞内钙释放通道，在那里它们形成有序阵列。细胞刺激后，RyR组短暂开放，并迅速释放到胞质溶胶中的大量钙所需的抽搐肌肉收缩。这些钙释放事件的特征强烈表明参与的RyR同步工作，并且它们都在几毫秒内一起激活和失活。目前还不清楚RyRV如何在细胞中同步它们的门控。因此，很难理解在各种肌肉疾病中发现的异常兴奋-收缩偶联的起源，包括已经鉴定出编码RyR 1的基因突变的那些（恶性高热、中央核心疾病）。由于RyR是细胞内通道，电生理学研究需要细胞亚分级、通道分离和重建成人工脂质双层。几乎所有来自这些双层研究的报告都描述了单个（个体）RyR 1或多个RyR的独立行为。只有A医生。Marks的实验室描述了相邻的RyR（2-4通道），如果FK 506结合蛋白（FKBP）分子与通道相关，则这些通道通过同步协调进行门控。这个过程，新的离子通道领域，被命名为“耦合门控”。尽管了解钙释放的重要性，但对这种RyR 1间的通信知之甚少。在过去的几年里，主要研究者已经开发出了方法，并积累了初步的数据，清楚地证明了研究通道与他的双层系统耦合的可行性。因此，本提案的目标是定义RyR通道之间的通信机制，用于耦合门控。具体目标#1：定义RyR 1通道之间的通信机制。该目标细分为三个子目标：A）定义ATP/Mg 2+在RyR 1配位中的作用; B）定义Ca 2+在配位门控中的作用;以及C）定义FKBP 12（钙稳定蛋白1）和其他辅助蛋白的作用。

DESCRIPTION (provided by applicant): The goal of this project is to understand the coupled function of skeletal muscle ryanodine receptor channels (RyRs). The RyRs are intracellular calcium release channels located in the membranes of intracellular calcium stores, where they form orderly arrays. After cell stimuli, groups of RyRs briefly open and rapidly release to the cytosol the bulk of calcium required for the twitch muscle contraction. The characteristics of these calcium release events strongly indicate that the participating RyRs work in synchrony and they all activate and deactivate together within few milliseconds. It is still unknown how RyRVs communicate for synchronizing their gating in cells. Consequently, it is difficult to understand the genesis of anomalous excitation-contraction coupling as found in various muscle diseases, including those (Malignant Hyperthermia, Central Core Disease) where mutations in the genes encoding RyR1 have been identified. As RyRs are intracellular channels, electrophysiological studies require cell subfractionation, channel isolation and reconstitution into artificial lipid bilayers. Nearly all reports from these bilayers studies describe a single (individual) RyR1 or independent behavior of multiple RyRs. Only Dr. A. Marks's laboratory described neighboring RyRs (2-4 channels) that gated with synchronous coordination if FK506 binding protein (FKBP) molecules were associated with the channels. This process, new for the field of ion channels, was named "coupled gating". Despite its importance to understand calcium release, little else is known about this inter-RyR1 communication. During the last years, the principle investigator has developed methodologies and accumulated preliminary data that clearly evidence the feasibility of studying channels coupling with his bilayer system. Consequently, the goal of this proposal is to define the mechanisms of communication between RyR channels for coupled gating. The specific aims of this proposal are: Specific Aim #1: Define the mechanisms of communication between RyR1 channels. This aim is subdivided in three sub-Aims: A) Define the role of ATP/Mg2+ in RyR1 coordination; B) Define the role of Ca2+ in coordinated gating; and C) Define the role of FKBP12 (Calstabilinl) and other ancillary proteins.