Structural Basis for Rab and Arf GTPase Regulated Membrane Trafficking

 描述（由申请人提供）：Rab、Arf和Arl GTP酶作为控制膜运输以及膜界定细胞器的生物发生、身份和成熟的系统的核心调节组分发挥作用。这些基本控制机制的缺陷与复杂的疾病状态和遗传相关疾病有关。运输GTP酶的功能循环和输出，包括通过鸟嘌呤核苷酸交换因子（GEF）的激活、通过GTP酶激活蛋白（GAP）的失活以及与效应物的相互作用，也被病毒、细菌和真核病原体操纵以逃避宿主应答并支持复制。了解GTP酶的运输调节机制以及这些机制如何被病原体操纵和在疾病条件下失调是一个至关重要的挑战。膜募集和GEF和GAP的自动调节是控制运输GTP酶的功能输出的主要范例。本申请的中心目标是通过结合多种生物化学，生物物理和结构方法，从宿主和病原体的角度研究分子和结构机制，识别GEF和GAP催化活性时空控制的新原则。特别是，我们将研究结构机制：Arf GEF膜招聘和变构激活（目的1）;膜靶向嗜肺军团菌GEF DrrA和质膜突触融合蛋白（目的2）的相互作用;和Rab灭活GAP和操纵病原体。这些研究将为宿主生物体中的贩运控制机制以及这些过程如何被细胞内病原体破坏提供新的见解。这些结果可能有助于设计基于机制的治疗策略，靶向GT3依赖性运输过程。

 DESCRIPTION (provided by applicant): Rab, Arf and Arl GTPases function as core regulatory components of systems that control membrane trafficking as well as the biogenesis, identity, and maturation of membrane delimited organelles. Defects in these fundamental control mechanisms are implicated in complex disease states and genetically-linked disorders. The functional cycle and outputs of trafficking GTPases, including activation by guanine nucleotide exchange factors (GEFs), deactivation by GTPase activating proteins (GAPs), and interactions with effectors, are also manipulated by viral, bacterial, and eukaryotic pathogens to evade host responses and support replication. Understanding the mechanisms underlying trafficking regulation by GTPases and how these mechanisms are manipulated by pathogens and dysregulated in disease conditions represents a critically important challenge. Membrane recruitment and autoregulation of GEFs and GAPs are major paradigms for controlling the functional outputs of trafficking GTPases. The central objective of this application is to discern new principles underlying spatiotemporal control of GEF and GAP catalytic activities by combining multiple biochemical, biophysical, and structural approaches to investigate the molecular and structural mechanisms from host and pathogen perspectives. Specially, we will investigate structural mechanisms for: Arf GEF membrane recruitment and allosteric activation (Aim 1); membrane targeting of the Legionella pneumophila GEF DrrA and interaction with plasma membrane syntaxins (Aim 2); and Rab deactivation by GAPs and manipulation by pathogens. These studies will provide new insights into trafficking control mechanisms in host organisms and how these processes are subverted by intracellular pathogens. The results may be useful for the design of mechanism- based therapeutic strategies targeting GTPase dependent trafficking processes.