Signaling through Rho GTP/GDP Exchange Factors

通过 Rho GTP/GDP 交换因子发出信号

• 批准号: 7212276
• 负责人: PHILIP B WEDEGAERTNER
• 金额: $ 27.92万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-04-01 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7212276
• 关键词: Actins; Address; Agonist; Binding; Biochemical; Biological Assay; Biological Models; Cell Fractionation; Cell Nucleus; Cell Surface Receptors; Cell membrane; Cells; Cellular biology; Cultured Cells; Cytoplasm; Cytoskeleton; Disease; F-Actin; Family; Feedback; Funding; GTP-Binding Proteins; Grant; Guanine Nucleotide Exchange Factors; Heterotrimeric GTP-Binding Proteins; Immunofluorescence Microscopy; Intracellular Signaling Proteins; Lead; Life; Link; Localized; Location; Mammalian Cell; Mediating; Molecular; Monomeric GTP-Binding Proteins; N-terminal; Play; Protein Isoforms; Proteins; RGS Domain; Research; Research Personnel; Role; Serum; Signal Pathway; Signal Transduction; Signaling Protein; Site; Small Interfering RNA; Specificity; Stimulus; Techniques; Testing; Thromboxane A2 Receptor; Time; Transfection; Transferase; cell growth; cellular imaging; design; extracellular; insight; member; mutant; novel; prevent; programs; research study; response; rho; uncontrolled cell growth

DESCRIPTION (provided by applicant): Intracellular signaling pathways depend upon appropriate and unique subcellular locations of their constituent proteins. Frequently, key intracellular signaling proteins move from one subcellular location to another (e.g., from cytoplasm to plasma membranes or from cytoplasm to nucleus) in response to extracellular stimuli. Incorrectly localized proteins can prevent completion of a particular signaling pathway or can cause unregulated signaling, such as uncontrolled cell growth. Understanding how cellular proteins come together at specific times and subcellular sites will lead to insight into ways to block inappropriate signaling in disease states. The heterotrimeric (alphabetagamma) G proteins act as molecular switches to relay information from activated cell-surface receptors to appropriate intracellular effectors. One family of G protein a subunits, consisting of alpha12 and alpha13, can activate the Rho family of small GTPases. Rho, in turn, activates signaling pathways that stimulate cell growth and morphological changes. A large family of Rho guanine nucleotide exchange factors (GEF) activates Rho, and a sub-family, termed RGS-RhoGEFs, and consisting of p115-RhoGEF, PDZ-RhoGEF, and LARG, function as direct links between alpha12/13 and Rho. However, the mechanisms responsible for activation of the RGS-RhoGEFs by alpha12 and alpha13 are poorly understood. The main objectives of this application are to elucidate the mechanisms that control subcellular localizations of the RGS-RhoGEFs, determine how differential and regulated subcellular localization of the RGS-RhoGEFs contribute to signaling function, and define the interactions between alpha12/13 and the RGS-RhoGEFs. These objectives will be addressed through the following specific aims: (1) Define subcellular localization of LARG, and determine changes in localization in response to activated Ga and GPCRs; (2) Define the functional significance of the actin-binding domain of PDZ-RhoGEF; (3) Define the involvement of Rho in alpha13- dependent PM recruitment of p115-RhoGEF; (4) Define interactions between RGS-RhoGEFs and alpha13-alpha12/13 or alphaq, and determine signaling functions of RhoGEF binding-defective mutants of alpha12/13. This research will utilize cultured mammalian cells as the model systems and will employ cell biology and biochemical techniques to examine the central hypothesis that subcellular localization plays a critical role in understanding RGS-RhoGEF function and how alpha12/13 activate Rho signaling pathways through p115-RhoGEF, PDZ-RhoGEF and LARG.

描述（由申请人提供）：细胞内信号通路取决于其组成蛋白的适当和独特的亚细胞位置。通常，关键的细胞内信号蛋白从一个亚细胞位置转移到另一个亚细胞膜（例如，从细胞质到质膜或从细胞质到核）移动，以响应细胞外刺激。错误的局部蛋白质可以防止完成特定的信号通路或可能导致不受控制的信号传导，例如不受控制的细胞生长。了解细胞蛋白如何在特定时间和亚细胞位点聚集在一起，将洞悉阻止疾病状态中不适当信号传导的方法。 异三聚体（Alphabetagamma）G蛋白充当分子开关，从活化的细胞表面受体传递到适当的细胞内效应子。一个由α12和α13组成的G蛋白A亚基一个家族可以激活小GTPases的Rho家族。 RHO反过来激活了刺激细胞生长和形态变化的信号通路。大量的Rho鸟嘌呤核苷酸交换因子（GEF）激活了Rho，一个子家庭称为RGS-RHOGEFS，由P115-Rhogef，PDZ-RHOGEF和LARG组成，并且在Alpha12/13和Rho之间的直接联系起作用。但是，对Alpha12和alpha13的RGS-RHOGEF激活的机制知之甚少。本应用程序的主要目标是阐明控制RGS-RHOGEFS亚细胞定位的机制，确定RGS-RHOGEF的差异和调节的亚细胞定位如何促进信号功能，并定义Alpha12/13和RGS-RHOGEFS之间的相互作用。这些目标将通过以下特定目的解决：（1）定义LARG的亚细胞定位，并确定对激活的GA和GPCR的响应定位变化； （2）定义PDZ-RHOGEF的肌动蛋白结合结构域的功能意义； （3）定义RHO参与α13-依赖性PM募集P115-Rhogef； （4）定义RGS-RHOGEFS与Alpha13-Alpha12/13或Alphaq之间的相互作用，并确定Alpha12/13的Rhogef结合缺陷突变体的信号传导功能。这项研究将利用培养的哺乳动物细胞作为模型系统，并将采用细胞生物学和生化技术来研究中心假设，即亚细胞定位在理解RGS-RHOGEF功能以及Alpha12/13如何通过P115-RHOGEF，PDZ-RGEF，PDZ-RGEF，PDZ-RGF和LARG和LARG激活Rho信号通路。