Effects of corticotropin-(1-24)-tetracosapeptide on polyphosphoinositide metabolism and protein phosphorylation in rabbit iris subcellular fractions.
Effects of the neuropeptide corticotropin-(1-24)-tetracosapeptide (ACTH) on the endogenous and exogenous phosphorylation of lipids and endogenous phosphorylation of proteins were investigated in microsomes and a 110,000 X g supernatant fraction [30-50% (NH4)2SO4 precipitate; ASP 30-50] obtained from rabbit iris smooth muscle. Subcellular distribution studies revealed that both of these fractions are enriched in diphosphoinositide (DPI) kinase. The 32P labeling of lipids and proteins was measured by incubation of the subcellular fractions with [gamma-32P]ATP. The labeled lipids, which consisted of triphosphoinositide (TPI), DPI, and phosphatidic acid (PA) were isolated by TLC. The microsomal and ASP 30-50 fractions were resolved into six and nine labeled phosphoprotein bands, respectively, by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The basal labeling of both lipids and proteins was rapid (30-60 s), and it was dependent on the presence of Mg2+ in the incubation medium; in general it was inhibited by high concentrations (greater than 0.2 mM) of Ca2+. ACTH stimulated the labeling of TPI and inhibited that of PA in a dose-dependent manner, with maximal effect observed at 50-100 microM of the peptide. ACTH appears to increase TPI labeling by stimulating the DPI kinase. Under the same experimental conditions ACTH (100 microM) inhibited significantly the endogenous phosphorylation of six microsomal phosphoproteins (100K, 84K, 65K, 53K, 48K, and 17K). In the ASP 30-50 fraction, ACTH inhibited the phosphorylation of three phosphoproteins (53K, 48K, and 17K) and stimulated the labeling of six phosphoprotein bands (117K, 100K, 84K, 65K, 42K, and 35K).(ABSTRACT TRUNCATED AT 250 WORDS)
D000324:Adrenocorticotropic Hormone / Q000494:pharmacology*; D000818:Animals; D007498:Iris / Q000378:metabolism*; D010712:Phosphatidic Acids / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols / Q000378:metabolism*; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism*; D011817:Rabbits; D013347:Subcellular Fractions / Q000378:metabolism
D010712:Phosphatidic Acids; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols; D011506:Proteins; D000324:Adrenocorticotropic Hormone