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.
Arf, Arl, and Rab GTPases are core control elements of systems that regulate the biogenesis and trafficking of cellular organelles. These GTPases, their effector proteins, and the enzymes that control their activation and deactivation are manipulated by viral, bacterial, and eukaryotic pathogens and dysregulated in complex disease states including diabetes, cancer, and neuropathies. The underlying mechanisms and disease etiologies remain poorly characterized. The research in this application will investigate molecular and structural bases for trafficking control by GTPases and how these fundamental regulatory processes are subverted by intracellular pathogens. The insights derived from these studies may be useful for development of novel mechanism-based therapeutic strategies.
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