G protein-coupled receptor kinase 4 (GRK4) regulates the phosphorylation and function of the dopamine D3 receptor

Abstract

During conditions of moderate sodium excess, the dopaminergic system regulates blood pressure and water and electrolyte balance by engendering natriuresis. Dopamine exerts its effects on dopamine receptors, including the dopamine D3 receptor. G protein-coupled receptor kinase 4 (GRK4), whose gene locus (4p16.3) is linked to essential hypertension, desensitizes the D1 receptor, another dopamine receptor. This study evaluated the role of GRK4 on D3 receptor function in human proximal tubule cells. D3 receptor co-segregated in lipid rafts and co-immuno-precipitated and co-localized in human proximal tubule cells and in proximal and distal tubules and glomeruli of kidneys of Wistar Kyoto rats. Bimolecular fluorescence complementation and confocal microscopy revealed that agonist activation of the receptor initiated the interaction between D3 receptor and GRK4 at the cell membrane and promoted it intracellularly, presumably en route to endosomal trafficking. Of the four GRK4 splice variants, GRK4-γ and GRK4-α mediated a 3- and 2-fold increase in the phosphorylation of agonist-activated D3 receptor, respectively. Inhibition of GRK activity with heparin or knockdown of GRK4 expression via RNA interference completely abolished p44/42 phosphorylation and mitogenesis induced by D3 receptor stimulation. These data demonstrate that GRK4, specifically the GRK4-γ and GRK4-α isoforms, phosphorylates the D3 receptor and is crucial for its signaling in human proximal tubule cells. © 2009 by The American Society for Biochemistry and Molecular Biology, Inc.