D1 and AT1 Receptor Interaction--Hypertension--mechanism

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The overall goal of Project by Felder is to study the interaction between two renal sodium regulatory pathways that play a key role in blood pressure regulation. Defects in these pathways have been directly linked to the development of hypertension. The renal dopaminergic and angiotensin systems independently and in concert regulate most of renal sodium excretion. We will determine the mechanisms by which the interactions of dopamine D1, and angiotensin AT1 receptors regulate blood pressure and renal sodium transport through common and independent second messenger pathways. Naturally occurring single nucleotide polymorphisms, G448T, C679T, and C1711T that result in amino acid substitutions R65L, A142V, and A486V, respectively in G protein-coupled receptor kinase 4 gamma isoform (GRK4), specifically of the gamma isoform, constitutively increase enzyme activity, resulting in the phosphorylation and uncoupling of the D1 dopamine receptor from its G protein and effector complexes in the human kidney. These GRK4 gene variants, by themselves or via their interactions with the renin-angiotensin system, play a crucial role in the pathogenesis of human (and rodent) genetic hypertension. The over all hypothesis of Project by Felder is that these amino acid changes in GRK4gamma constitutively desensitize D1 receptors but not AT1 receptors, allowing unfettered AT1 receptor action. Three specific aims have been developed to determine the mechanisms of the differential actions of GRK4gamma variants on D1 and AT1 receptors in human proximal tubule cells. 1. We will determine the mechanism of the increased activity of GRK4gamma carrying amino acid substitutions, R65L, A142V, and A486V. This will test the hypothesis that increased binding of GRK4gamma variants to D1 receptors and/or increased targeting of GRK4 variants to the plasma membrane increase GRK4 activity. 2. We will also determine whether GRK4gamma or its variants (R65L, A142V, and A486V) regulate AT1 receptors. This will test the hypothesis that GRK4gamma variants do not desensitize AT1 receptors and allow unimpeded AT1 receptor actions in hypertension. 3. We will study the fundamental mechanisms involved in the acute and chronic transregulation of D1 and AT1 receptors. This will test the hypothesis that D1 receptors negatively regulate the function and expression of AT1 receptors and that this regulation is impaired in genetic hypertension. Studies will be performed in cells endogenously (e.g., renal proximal tubule cells from normotensive and hypertensive humans and compared to rodents), as well as in cells heterologously expressing D1 and AT1 receptors and GRK4gamma variants using pharmacological, cellular and molecular biological tools and end points to study D1 and AT1 receptor function. Based on our studies on the important role of GRK4gamma variants in D1 receptor signaling, we suggest that a single gene that regulates many other genes may be a major contributor to the pathogenesis of essential hypertension.