Phosphorylation of Gap Junction Proteins

描述：(由申请人提供)Gap Junction通信非常关键 在许多细胞过程中很重要，包括控制细胞增殖， 胚胎发育和细胞分化。这些途径提供了 包括离子在内的小分子(~lt；1000 Da)的细胞间扩散， 代谢物和第二信使。在脊椎动物中，缝隙连接由 连接蛋白家族的蛋白质，该家族包含十几个成员。 由不同连接蛋白组成的通道可以根据不同的情况而打开或关闭 通过改变跨连接电压、pH和磷酸化程度。这 提案关注连接蛋白43(Cx43)和连接蛋白45有几个原因；两者都是 已知在多个丝氨酸上被磷酸化，两者都是正/负的 然而，受TPA等激酶效应物的调控，它们具有非常不同的 胞质中的传导特性和序列同源性很低 尾部区域除了一系列的双丝氨酸重复序列。这项建议 研究了磷酸化在这些缝隙连接调节中的作用。 蛋白质及其形成的通道。我们假设除了 对门控的影响，磷酸化对于调节其他 Cx43‘S“生命周期”的几个方面(即连接蛋白寡聚、连接蛋白 贩运、缝隙连接组装和蛋白质周转)。我们的具体目标 目的是(1)确定Cx43和Cx45内的磷酸化事件，这 调控缝隙连接蛋白的运输、缝隙连接组装、门控和周转 动态平衡细胞。(2)表征与连接蛋白结合并调节连接蛋白的激酶 功能。(3)确定细胞周期依赖性连接蛋白的变化 在有丝分裂过程中允许连接蛋白重新分布的磷酸化。我们计划 应用我们对连接蛋白磷酸化和磷酸化特异性的知识 皮肤损伤后缝隙连接蛋白状态分析及组织改建的探讨 心脏病时的缝隙连接。

DESCRIPTION: (provided by applicant) Gap junctional communication is critically important in many cell processes including control of cell proliferation, embryonic development, and cell differentiation. These pathways provide for cell-to-cell diffusion of small molecules (<1000 Da) including ions, metabolites, and second messengers. In vertebrates, gap junctions are composed of proteins from the connexin family, which contains over a dozen members. Channels composed of different connexins can be opened or closed to varying degrees by changes in transjunctional voltage, pH, and phosphorylation. This proposal is focused on connexins 43 (Cx43) and 45 for several reasons; both are known to be phosphorylated on multiple serines, both are positively/negatively regulated by kinase effectors such as TPA, however, they have very different conducting properties and share little sequence homology in the cytoplasmic tail region aside from a series of double serine repeats. This proposal examines the role of phosphorylation in the regulation of these gap junction proteins and the channels that they form. We hypothesize that in addition to effects on gating, phosphorylation is important for the regulation of other aspects of Cx43's "life cycle" (i.e., connexin oligomerization, connexin trafficking, gap junction assembly, and protein turnover). Our specific aims are to (1) determine the phosphorylation events within Cx43 and Cx45, which regulate connexin trafficking, gap junction assembly, gating and turnover in homeostatic cells. (2) Characterize kinases that bind to and regulate connexin function. (3) Determine the cell cycle dependent changes in connexin phosphorylation that allow connexin redistribution during mitosis. We plan to apply our knowledge of connexin phosphorylation and phosphorylation specific probes to the analysis of connexin status in wounded skin and the remodeling of gap junctions during heart disease.