G Protein Regulation of Golgi Structure and Function

摘要 异源三聚体G蛋白（αβγ）以其连接G蛋白偶联受体的功能而闻名 （GPCR）对各种细胞内反应产生影响，从而在传递广泛的信号方面发挥重要作用 各种细胞外信号调节无数的生理过程。在教科书中，G 蛋白质在与细胞质膜的细胞质表面结合时发挥其功能。 与质膜限制性G蛋白信号传导的经典观点相反， 认识到G蛋白定位是动态的和受调节的，使得它们可以可逆地从 G蛋白在细胞内的位置，并且G蛋白可以在细胞内具有重要的细胞功能。 胞内位点。本文的研究重点是理解Gβγ的非正则函数 亚基，特别是Gβγ在调节高尔基体信号转导中的作用。我们之前的工作揭示了 高尔基体定位的Gβγ在调节高尔基体细胞质表面信号通路中的重要作用 膜，其控制去往质膜的选择蛋白质货物的高尔基体出口。的 目前的研究将检验Gβγ调节信号通路的假设， 通过调节生理条件下高尔基体的片段化来控制高尔基体的完整性， 病理生理条件。可逆的高尔基体断裂是一种细胞现象， 正常情况下，如有丝分裂期间，以及发生在疾病状态，如感染，癌症和 神经退行性疾病本申请将集中于确定Gβγ在调节细胞凋亡中的新作用。 有丝分裂高尔基体碎片检查点，并通过询问Gβγ在调节微管- 依赖的高尔基体分裂。此外，此应用程序将定义直接 促进高尔基体上Gβγ的信号传导。这些目标将通过各种实验来实现。 方法，包括培养细胞，免疫荧光显微镜，荧光显微镜活 细胞、生物传感器、药理学抑制剂、突变分析和生物化学测定。

Abstract Heterotrimeric G proteins (αβγ) are well known for their function in linking G protein-coupled receptors (GPCRs) to a variety of intracellular responses, and thereby playing essential roles in transmitting a wide variety of extracellular signals into regulation of countless physiological processes. In the textbook view, G proteins carry out their function while associated with the cytoplasmic surface of a cell’s plasma membrane. In contrast to the classical view of plasma membrane-limited G protein signaling, it is becoming increasingly recognized that G protein localization is dynamic and regulated, such that they can reversibly traffic from the plasma membrane to intracellular locations, and that G proteins can have important cellular functions at intracellular sites. The research in this proposal focuses on understanding non-canonical functions of Gβγ subunits, and specifically roles for Gβγ in regulating signaling at the Golgi. Our previous work revealed an important role for Golgi-localized Gβγ in regulating a signaling pathway on the cytoplasmic surface of Golgi membranes that controls the Golgi exit of select protein cargo destined for the plasma membrane. The research in this current proposal will examine the hypothesis that Gβγ regulates signaling pathways that control Golgi integrity by regulating the fragmentation of the Golgi under physiological and pathophysiological conditions. Reversible Golgi fragmentation is a cellular phenomenon that occurs under normal conditions, such as during mitosis, and that occurs in disease states, such as infection, cancer and neurodegenerative disease. This application will focus on defining a novel role for Gβγ in regulating the mitotic Golgi fragmentation checkpoint and by interrogating a novel role for Gβγ in regulating microtubule- dependent Golgi fragmentation. In addition, this application will define upstream mechanisms that directly promote signaling by Gβγ at the Golgi. These objectives will be pursued by a variety of experimental approaches, including cultured cells, immunofluorescence microscopy, fluorescence microscopy of live cells, biosensors, pharmacological inhibitors, mutational analysis, and biochemical assays.