Objective and Aims for Project 3The internalization and recycling of cell surface receptors, glucose transporters, and other plasmamembrane proteins is regulated by phosphoinositide metabolism as well as GTPases of the Rab and Arffamilies. Defects in these regulatory mechanisms have been implicated in a variety of disease statesincluding cancer and type II diabetes. The long term objective of Project 3 is to characterize poorlyunderstood mechanisms that integrate phosphoinositide signaling and Rab/Arf GTPase activation with thedynamic molecular assemblies that mediate/regulate trafficking events at the plasma membrane and withinthe endosomal system. To achieve this objective, we will combine crystallographic, biochemical,biophysical, and mutational studies on purified complexes with complementary structure-function analyses incollaboration with Projects 1, 2, 4, and the imaging core. Building on the observations of the previousfunding period and extensive new preliminary data, we will:
(Aim 1) examine the structural determinants ofautoregulation and phosphoinositide dependent membrane targeting in the Grp1 family of Arf GTPaseexchange factors;
(Aim 2) investigate the mechanism of autoregulation, membrane targeting, and Rabactivation by the Rabex-5/Rabaptin-5 complex and Rin1;
and (Aim 3) explore the structural bases forregulation of endocytic recycling by phosphoinositide dependent and independent complexes of EHDproteins with Rabenosyn-5, EHBP1, and FIP2.Relevance to Public HealthBoth type II diabetes and cancer involve defects in the mechanisms by which cell surface receptors forinsulin and growth factors communicate with the molecular machinery that controls cell metabolism andgrowth. By studying these mechanisms comprehensively at the structural, molecular, and cellular levelsthrough collaborative projects the combine the expertise of structural, molecular, and cell biologists we hopeto identify novel mechanism-based strategies that can be exploited for therapeutic intervention.
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