Genetic hypertension may be the result of abnormalities in the renal dopaminergic system. The renal autocrine/paracrine natriuretic function of dopamine, via the D1-like receptors (D1 and D5), is impaired in the spontaneously hypertensive rat. The impairment is not the result of a decrease in renal dopamine production nor is it due to abnormalities in the G proteins or effector proteins. We propose that the impairment resides within one or more intermediate components. Genetic changes occurring within the genes that encode D1-like receptor interacting proteins may lead to disruption of D1-like receptor mediated signal transduction and, in turn, result in increased risk for the development of hypertension.
Specific aim 1 will identify and characterize the proteins that interact with the D1-like receptors in the kidney. We will employ the yeast two- hybrid system to identify genes encoding proteins that interact with the D1-like receptors. The D1 and D5 dopamine receptors will be divided into structural domains consisting of the amino terminal, carboxy terminal, extracellular and intracellular loops and each of the seven transmembrane segments. These domains will be used as molecular probes to detect interactions occurring with proteins expressed from a human kidney cDNA library. Protein-protein interactions will be verified using laser confocal microscopy and fluorescence resonance energy transfer techniques.
Specific aim 2 will test the hypothesis that there are differences in the sequences of the D1-like receptor interacting proteins between normotensive and hypertensive individuals and that it is these differences that lead to hypertension. Genetic variants, especially those occurring within regions identified in specific aim 1 as potential sites of interaction with the D1-like receptor proteins or those that map to regions of the human genome associated with hypertension risk, will be studied further. We will screen for polymorphisms in genomic DNAs derived from control and hypertensive individuals representing several different ethnic groups including Asians, Africans, American Caucasians, and Italian Caucasians. Finally, specific aim 3 will determine if there are differences in the expression of D1-like receptor interacting proteins in young and adult kidney during maturation. A greater understanding of the proteins and protein networks associated with the regulation of the D1-like dopamine receptors will likely lead to greater insight into the roles these receptors play in the maintenance of blood pressure.
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