Overall, our long-term goal is to obtain a better understanding of the regulation of renal epoxygenase enzymes and determine the role they play in renal function. The presence of an active cytochrome P450 monooxygenase in renal tissue is well documented. Even though the kidney contains the second highest body concentration of cytochrome P450 enzymes, relatively little is known about the catalytic and molecular properties of the renal hemoproteins. In earlier studies it was shown that, in addition to catalyzing lauric acid omega/omega-1 oxidation, kidney cytochrome P450 was active in the metabolism of several drugs and carcinogens. The inducibility and the segmental distribution of specific cytochrome P450 isoforms along the nephron is well established. The initial demonstration of a role for cytochrome P450 in the omega/omega-1 oxidation of prostanoids and leukotrienes, known urinary metabolites of these functionally important mediators, suggested a physiological role for this enzyme system. More recent studies have documented a function for renal cytochrome P450 as an active catalyst for the metabolism of arachidonic acid to biologically active products.
The specific aim of this project is to isolate and characterize genes encoding cytochrome P450 2C (epoxygenase) family members from rat kidney and determine the molecular basis for the effect of dietary salt loading on this enzyme system. Preliminary evidence indicates that this family of cytochrome P450 isoforms is involved in the metabolism of endogenous substrates (arachidonic acid) to bio-active compounds which affect renal vascular tone, free water and ion transport. The enzymatic activity of one of these isoforms has been shown to be dramatically induced in the kidney after placing rodents on a high salt diet. We propose to clone and sequence the control and salt-regulated renal epoxygenase genes and then study the molecular basis for the increase in the level of epoxygenase activity in animals given a high salt diet. In addition, other preliminary evidence outlined in project 2 demonstrates that in the Dahl+ salt sensitive strain of rats there is a lack of induction of epoxygenase activity following salt loading. We also propose to clone the homologue of the salt regulated epoxygenase (SREP) gene from the Dahl+ rat kidney and determine the molecular basis for the absence of salt regulation of this enzyme in the Dahl+ (salt sensitive) rats.
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