Coordination of membrane traffic through rab GEF and GAP cascades

通过 rab GEF 和 GAP 级联协调膜运输

• 批准号: 9094212
• 负责人: PETER Jay NOVICK
• 金额: $ 34.5万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9094212
• 关键词: Address; Alleles; Appearance; Binding; Biochemical; Biological Assay; Cell Fractionation; Cell physiology; Cell-Free System; Collection; Complex; Coupled; Defect; Diabetes Mellitus; Disease; Drops; Event; Exocytosis; Feedback; Fluorescence Microscopy; GTP Binding; GTPase-Activating Proteins; Glass; Glucose; Goals; Golgi Apparatus; Growth; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Host Defense; Human; Hydrolysis; Lead; Leg; Link; Location; Malignant Neoplasms; Mammalian Cell; Membrane; Membrane Protein Traffic; Modeling; Modification; Molecular; Motor; Myosin Type V; Nerve Degeneration; Nutrient; Pathway interactions; Phosphatidylinositols; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Play; Protein Kinase; Proteins; Recruitment Activity; Regulation; Regulatory Pathway; Role; SERPINA4 gene; SNAP receptor; Secretory Component; Secretory Vesicles; Series; Signal Transduction; Site; Staging; Structure; System; Testing; Time; Total Internal Reflection Fluorescent; Toxic effect; Vesicle; Work; Yeasts; casein kinase; casein kinase II; flexibility; human disease; in vivo; member; mutant; novel; pathogen; phosphatidylinositol 4-phosphate; preference; programs; public health relevance; rab GTP-Binding Proteins; trafficking

 DESCRIPTION (provided by applicant): Rabs constitute the largest branch of the Ras GTPase superfamily, with ten members in yeast and more than 60 in mammalian cells. They serve as master regulators of membrane traffic, each typically controlling several different aspects of a specific stage of membrane traffic by recruiting diverse effector proteins such as cytoskeletal motors, vesicle tethering proteins and regulators of SNARE complex assembly. Rabs, in turn, are regulated by specific guanine nucleotide exchange factors (GEFs) that catalyze the displacement of GDP and binding of GTP as well as GTPase activating proteins (GAPs) that stimulate the slow intrinsic rate of GTP hydrolysis. We have proposed that adjacent Rabs on a pathway are networked to one another through their regulators; specifically we have shown that the Rab, Ypt32, in its GTP-bound form recruits Sec2, the GEF that activates the downstream Rab, Sec4, as well as Gyp1, the GAP that inactivates the upstream Rab, Ypt1. The net effect of these counter-current cascades is a programmed series of Rab transitions that lead to critical changes in the functional identity of the membrane as it flows along the exocytic pathway. Phosphoinositide also play key roles in the temporal and spatial regulation of membrane traffic. The Golgi pool of phosphatidylinositol 4-phosphate (PI(4)P) works in concert with Ypt32 to initially recruit Sec2, yet a subsequent drop in PI(4)P levels directs a regulatory switch in Sec2 function in which it binds to the Sec4 effector Sec15 in a positive feedback loop. PI (4) P distribution together with Sec2 phosphorylation determine when each regulatory circuit is used. We propose three aims: 1. we will define the role of two protein kinases in the regulation of Sec2 and other components of the secretory machinery. 2. We will test the effects of "rewiring" the rab regulatory circuits to evaluate several models concerning the role of rabs in the control of membrane traffic. 3. We will determine the molecular mechanisms underlying several stages of secretory vesicle maturation.

 描述（申请人提供）：Rab 构成 Ras GTPase 超家族的最大分支，在酵母中有 10 个成员，在哺乳动物细胞中有 60 多个成员。它们作为膜运输的主调节器，通常通过招募不同的效应蛋白（例如细胞骨架马达、囊泡束缚蛋白和 SNARE 复合体组装的调节器）来控制膜运输特定阶段的几个不同方面。反过来，Rab 受到特定鸟嘌呤核苷酸交换因子 (GEF) 的调节，GEF 催化 GDP 的置换和 GTP 的结合，以及 GTP 酶激活蛋白 (GAP) 刺激 GTP 水解的缓慢内在速率。我们提出，一条通路上的相邻 Rab 通过其监管者相互联网；具体来说，我们已经表明，Rab Ypt32以其GTP结合形式招募Sec2（激活下游Rab Sec4的GEF）以及Gyp1（失活上游Rab Ypt1的GAP）。这些逆流级联的净效应是一系列程序化的 Rab 转变，当膜沿着胞吐途径流动时，会导致膜的功能特性发生关键变化。磷酸肌醇在膜交通的时间和空间调节中也发挥着关键作用。高尔基体的磷脂酰肌醇 4-磷酸 (PI(4)P) 库与 Ypt32 协同工作，最初招募 Sec2，但随后 PI(4)P 水平的下降会引导 Sec2 功能的调节开关，在正反馈环中与 Sec4 效应器 Sec15 结合。 PI (4) P 分布与 Sec2 磷酸化一起决定何时使用每个调节电路。我们提出三个目标： 1. 我们将定义两种蛋白激酶在调节 Sec2 和分泌机制的其他组件中的作用。 2. 我们将测试“重新布线” rab 调节回路的效果，以评估有关 rabs 在 膜流量的控制。 3. 我们将确定分泌囊泡成熟几个阶段的分子机制。