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)：RABS是Ras GTP酶超家族中最大的分支，在酵母中有10个成员，在哺乳动物细胞中有60多个成员。它们是膜运输的主要调节者，每一种都通过招募不同的效应蛋白来控制膜运输的几个不同方面，如细胞骨架马达、囊泡拴系蛋白和SNARE复合体组装的调节因子。反过来，RABS又受到特定的鸟核苷酸交换因子(GEF)的调控，GEF催化GDP的置换和GTP的结合，以及GTP酶激活蛋白(GAP)刺激GTP的缓慢内在水解率。我们已经提出，一条途径上的相邻RAB通过它们的调节器相互联网；具体地说，我们已经证明，以GTP结合的形式的Rab，Ypt32，招募到了Sec2，即激活下游Rab，Sec4的环境基金，以及Gyp1，这个间隙使上游的Rab，Ypt1失活。这些逆流级联的净效应是一系列程序化的RAB转变，导致膜在沿着胞外途径流动时功能特性发生关键变化。肌醇磷脂在膜运输的时空调控中也起着关键作用。高尔基磷脂酰肌醇4-磷酸(PI(4)P)与Ypt32协同工作，最初招募Sec2，但随后PI(4)P水平的下降引导了Sec2功能的调节开关，在正反馈循环中，它与Sec4效应器Sec15结合。PI(4)P的分布和Sec2的磷酸化一起决定了每个调节回路的使用时间。我们提出了三个目标：1.我们将确定两个蛋白激酶在调节Sec2和其他分泌机制组件中的作用。2.我们将测试重新布线RAB调节电路的效果，以评估几个关于RAB在 膜上交通的控制。3.我们将确定分泌囊泡成熟的几个阶段的分子机制。