Spatial Regulation of RGS and G Protein Signaling

描述（由申请人提供）：在此资助下，我们发现了许多G蛋白结合蛋白，建立了它们的特性，并揭示了异源三聚体G蛋白在细胞中的新作用。在目前的资助期间，我们表征了GIV，一种激活G蛋白的非受体GEF。GIV作为一种多模块信号转导器，组装蛋白质复合物并将G亚基与其他信号传导途径（生长因子和PI 3 K/Akt信号传导）和几种重要的细胞功能联系起来，包括细胞迁移，自噬，分泌蛋白的运输和生长因子受体的内吞作用。GIV还结合G，促进EGF受体下调，并关闭来自内体的增殖信号。提出的工作的总体目标是利用我们已经开发的工具和见解来阐明G蛋白的一些众所周知但知之甚少的功能，特别是在细胞内膜上。我们将专注于两个具体目标：具体目标#1：查明GIV及其与G β和G β相互作用调节EGFR贩运和生长因子信号传导的机制。我们基于初步数据的工作模型是，GIV在EGF刺激后依次结合G蛋白和G蛋白，GIV的磷酸化调节与两种G蛋白的相对结合，并决定细胞是否迁移或增殖。 具体目标#2：确定G β和GIV在高尔基体功能中的作用，并定义相关机制。我们的数据表明，GIV的耗尽或GIV突变体未能激活G β，减缓了新合成的蛋白质通过高尔基体的转运。我们基于初步数据的工作假设是，GIV通过激活G β，下调Arf 1活性和促进囊泡脱壳来实现这一点。 这些研究有望为GIV如何结合和调节两种不同的G蛋白以及影响生长因子信号传导、高尔基体功能、细胞迁移和细胞增殖提供新的见解。基于GIV在调节细胞行为中的重要作用，明确GIV的功能和相互作用是非常必要的。我们的研究不仅将提供对G蛋白在细胞功能中的基本作用的见解，而且还可能建立新的范例，这将有助于确定新的治疗靶点，用于开发药理学和抗肿瘤药物。通过利用我们掌握的新工具、专业知识和信息，我们处于独特的位置来解决这些问题。

DESCRIPTION (provided by applicant): Under this grant we have discovered a number of G protein binding proteins, established their properties and revealed new roles for heterotrimeric G proteins in cells. During the current funding period we characterized GIV, a non-receptor GEF that activates G�proteins. GIV serves as a multimodular signal transducer that assembles protein complexes and links G�ubunits to other signaling pathways (growth factor and PI3K/Akt signaling) and to several important cell functions, including cell migration, autophagy, transport of secretory proteins, and endocytosis of growth factor receptors. GIV also binds G� promotes EGF receptor downregulation, and shuts down proliferative signaling from endosomes. The overall goal of the work proposed is to take advantage of the tools and insights we have developed to shed light on some well-known but poorly understood functions of G proteins especially on intracellular membranes. We will focus on two specific aims: Specific Aim #1: To pinpoint the mechanisms by which GIV and its interactions with G�and G�regulate EGFR trafficking and growth factor signaling. Our working model based on preliminary data is that GIV sequentially binds G� and G�after EGF stimulation and that phosphomodification of GIV regulates the relative binding to the two G proteins and determines whether cells migrate or proliferate. Specific Aim #2: To determine the role of G� and GIV in Golgi functions and to define the mechanisms involved. Our data indicate that depletion of GIV or a GIV mutant that fails to activate G� slows transport of newly synthesized proteins through the Golgi. Our working hypothesis based on preliminary data is that GIV does so by activating G�, down-regulating Arf1 activity, and facilitating vesicle uncoating. These studies can be expected to provide novel insights into how GIV binds and regulates two different G proteins and influences growth factor signaling, Golgi functions, cell migration and cell proliferation. Based on its crucil functions in regulating cell behavior, it is essential to define GIV's functions and interactions. Our studies will not only provide insights into fundamental roles of G proteins in cell functions, but also it may establish new paradigms that will help to identify new therapeutic targets for development of pharmacologic and anti-tumor agents. We are uniquely positioned to tackle these problems by taking advantage of the new tools, expertise, and information we have in hand.