TRIC, a Novel Modulator of Intracellular Ca Homeostasis

描述（由申请人提供）：细胞信号传导的一个基本方面是从肌浆网（SR）或内质网（ER）中隔离的细胞内储存中释放Ca。为了在肌肉细胞中的兴奋-收缩偶联或在非肌肉细胞中的兴奋-分泌偶联的有效操作，穿过SR或ER膜的反离子运动必须伴随Ca的快速流出以中和在Ca释放期间产生的瞬时负电位。尽管在SR或ER膜中已经报道了对单价阳离子具有选择性的通道，但是还没有鉴定出编码SR或ER定位的阳离子选择性通道的基因。我们最近发现了一个新的基因编码的三聚体细胞内阳离子选择性通道（TRIC），其中包含一个细胞内膜保留序列的羧基端和一个保守的亮氨酸拉链基序的氨基端。纯化的TRIC蛋白可以在脂质双层膜中形成阳离子选择性通道，这提供了TRIC可能在Ca释放的急性期提供反离子运动机制的某些方面的可能性。生物化学研究确定TRIC和SERCA之间的功能相互作用，这可能会影响SR内的Ca稳态。本项目的重点是定义TRIC的细胞和分子功能，作为一个反离子通道，调节SR膜对一价阳离子的渗透性和/或作为跨SR膜的Ca转运的调节剂。我们将首先使用脂质双层重构系统建立TRIC的单通道特性。通过生物化学和分子检测，我们将确定TRIC的相互作用伙伴，可能参与调节TRIC的通道活性，或在调节TRIC和骨骼肌中的SERCA之间的功能相互作用。通过野生型对照和缺乏TRIC的突变肌纤维之间的比较研究，我们将确定TRIC在介导SR的Ca释放的急性期和SR的活性Ca摄取中的生理作用。由于TRIC代表一类靶向细胞内细胞器的新型阳离子通道，了解这种蛋白质在肌肉生理学中的作用将为我们提供对肌肉相关疾病中Ca信号传导调节的有价值的见解，以及在功能失调的Ca调节导致病理学的其它疾病中，例如神经变性疾病。

DESCRIPTION (provided by applicant): A fundamental aspect of cellular signaling is the release of Ca from sequestered intracellular stores in the sarcoplasmic reticulum (SR) or endoplasmic reticulum (ER). For efficient operation of excitation-contraction coupling in muscle cells, or excitation-secretion coupling in non-muscle cells, counter ion movement across the SR or ER membrane must accompany the rapid efflux of Ca to neutralize the transient negative potential produced during Ca release. Although channels selective for monovalent cations have been reported in SR or ER membranes, no gene has been identified that encodes a SR or ER localized cation selective channel. We have recently discovered a novel gene encoding a trimeric intracellular cation-selective channel (TRIC) that contains an intracellular membrane-retention sequence at the carboxyl-terminus and a conserved leucine-zipper motif at the amino-terminus. Purified TRIC protein can form a cation-selective channel in lipid bilayer membrane, providing the possibility that TRIC may provide certain aspects of the counter-ion movement mechanism during the acute phase of Ca release. Biochemical studies identify a functional interaction between TRIC and SERCA, which may influence Ca homeostasis inside the SR. The focus of this project is to define the cellular and molecular functions of TRIC as a counter-ion channel in regulating the permeability of SR membrane to monovalent cations and/or as a modulator of Ca transport across the SR membrane. We will first establish the single channel properties of TRIC using the lipid bilayer reconstitution system. Through biochemical and molecular assays, we will identify the interacting partners for TRIC that may participate in regulating the channel activity of TRIC, or in modulating the functional interaction between TRIC and SERCA in skeletal muscle. Through comparative studies between wild type control and mutant muscle fibers lacking TRIC, we will define the physiological role of TRIC in mediating both the acute phase of Ca release from the SR and active Ca uptake into the SR. As TRIC represents a novel class of cation channels targeted to an intracellular organelle, knowledge of the role of this protein in muscle physiology will provide us valuable insights into the regulation of Ca signaling in muscle-related diseases, and in other diseases where dysfunctional Ca regulation results in pathology, such as neurodegenerative disorders.