Uncovering the Role of GPR39 in the kidney

PROJECT SUMMARY G-protein coupled receptors (GPCRs) are seven transmembrane domain receptors that make up the largest class of proteins in the mammalian genome. When activated by an appropriate ligand, these receptors elicit specific and coordinated cellular responses that dictate a wide variety of physiological processes. One such receptor that I have begun to investigate is GPR39. While initial studies reported that zinc is the endogenous ligand, more recent reports demonstrated that zinc acts as an allosteric potentiator of GPR39 signaling. Although the endogenous ligand for GPR39 is still unclear, synthetic ligands have been key in understanding GPR39 signaling and have implicated functions for GPR39 in the heart, bone, skin, pancreas, and gastrointestinal tract. However, despite its well-recorded abundance in the kidney, there have been no published studies into its role in renal physiology. In preliminary data, I find that GPR39 localizes to principal cells (AQP2-positive) in the inner medullary collecting duct, and data from our collaborators indicates that GPR39 activation influences renal water handling. I therefore hypothesize that GPR39 activation inhibits water reabsorption by sequestering AQP2 in sub-cellular compartments. In agreement with this hypothesis, I find that treatment of collecting duct cells in vitro with a GPR39-specific agonist, cpd1324, altered dDAVP-induced AQP2 localization. Here, I propose two Specific Aims to uncover the mechanistic role that GPR39 plays in renal water handling. In Aim 1, I will examine three potential mechanisms by which GPR39 activation influences AQP2 trafficking in vitro. Aim 1A will test whether GPR39 activation inhibits AVPR2 expression and activity, and Aims 1B/C will test whether GPR39 activation inhibits forward trafficking (Aim 1B) or promotes reverse trafficking of AQP2 (Aim 1C). In Aim 2, I will determine the physiologic function of GPR39 in wild-type (WT) and knockout (KO) animals under conditions of water restriction and rehydration after water restriction. To do this, I will quantify changes in parameters including drinking volume, plasma sodium, urinary osmolality, and urinary output in WT and KO animals at baseline and after intervention. In addition, I will perform molecular analyses on renal tissues to monitor changes in AQP2 expression, phosphorylation status, and localization. Together, the experiments in this proposal will provide key insight into both the mechanism (Aim 1) and the physiological role (Aim 2) that GPR39 plays in kidney physiology.