The aim of this proposal is to determine the metabolic regulation, function, and significance of the de novo pathway for PAF biosynthesis in the normal kidney and in renal-induced hypertension and chemically-induced renal necrosis. Specific projects center on the following topics: (a) purification of the acetyltransferase and DTT-insensitive cholinephosphotransferase in the de novo PAF pathway for studies of their properties, development of antibodies against the enzymes, and long-range plans for molecular biology experiments, (b) regulatory controls (enzyme synthesis vs. allosteric factors) that will examine the influence of activators such as hormones, phosphorylation-dephosphorylation of proteins, """"""""cross-talk"""""""" between intermediates of the remodeling and de novo routes, metal ions, and other determinants and inhibitors (both physiological and pharmacological agents) on the de novo synthesis of PAF and its subsequent metabolic fate; (c) catabolism of alkylacetylglycerols (the immediate precursor of de novo PAF) and their retroconversion to alkylacylglycerols and alkylacylglycerophosphocholines (PAF precursor in the remodeling route); (d) topography of enzymes, precursors, and products among membrane systems; and (e) metabolism of an acetylated form of an ether-linked triglyceride analog. Animal models for investigating the enzymes of the de novo pathway of PAF synthesis in kidney diseases will include renal medullary necrosis induced by bromoethylamine hydrobromide and renal hypertension in the 1-clip 1-kidney rat model and spontaneous hypertensive [SHR] rats. Methodology involves enzymological procedures, cell culture, and lipid/protein chromatographic and chemical techniques. We anticipate the results of our studies will lead to a full understanding of the metabolism, regulatory controls, and the underlying biochemical mechanisms that explain how the de novo pathway of PAF biosynthesis is involved in both physiological and pathological functions of the kidney. Another expectation will be the gain of new knowledge about the biochemistry of the alkylacetylglycerols (and the long-chain acyl analog) and alkylglycerols formed in the de novo route of PAF synthesis, since the alkylacetylglycerols (and alkylacylglycerols) are an important type of """"""""diglyceride analog"""""""" with biological activities involved in the modulation of protein kinase C and cellular differentiation.
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