Synthesis of polyphosphoinositides in transverse tubule and sarcoplasmic reticulum membranes of frog skeletal muscle.
Synthesis of polyphosphoinositides has been studied in transverse (T-) tubule and sarcoplasmic reticulum (SR) membrane fractions of frog skeletal muscle, following 32P-labeling with [gamma-32P]ATP. Purified SR and T-tubule fractions respectively synthesize 9.4 +/- 0.8 and 71.9 +/- 9.8 pmol PtdInsP/mg per min, indicating nearly 8-fold higher activity of PtdIns kinase in the T-tubules than in the SR. The activity of this enzyme in both membrane systems is maximum at pH 7 and pCa 6. PtdInsP2 is synthesized from the endogenous PtdInsP, only in T-tubule membranes by the action of PtdInsP kinase. This lipid is the most intensely 32P-labeled phosphoinositide (181.7 +/- 9.2 pmol/mg per min) in these membranes. PtdIns kinase in the T-tubule and SR membranes, and PtdInsP kinase in the former are modulated by the free [Mg2+]. Loss of radiolabel from transiently maximal 32P-incorporation in polyphosphoinositides in T-tubule membranes, concomitant with a decrease in the ATP concentration in the incubation buffer, shows the occurrence of phosphoinositidases in these membranes. Under the conditions used, no such activities were evident in SR membranes. Compound 48/80, a mixture of condensation products of N-methyl-p-methoxyphenethylamine with formaldehyde, known to block phosphoinositidase C and phospholipase A2, causes a dose-dependent increase in the 32P-label of PtdInsP, in T-tubule membranes. The synthesis of lyso PtdInsP2, a deacylated form of PtdInsP2 which occurs in nearly equal quantities in both T-tubule and SR membranes, may result from a mechanism independent of phospholipase A2.
Asotra
K
K
Department of Physiology, School of Medicine, University of California, Los Angeles.
Lagos
N
N
Vergara
J
J
D019870:1-Phosphatidylinositol 4-Kinase; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D009132:Muscles / Q000378:metabolism*; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols / Q000096:biosynthesis*; D010741:Phospholipases A / Q000037:antagonists & inhibitors; D054467:Phospholipases A2; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D011892:Rana catesbeiana; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols; D010770:Phosphotransferases; D019870:1-Phosphatidylinositol 4-Kinase; D010741:Phospholipases A; D054467:Phospholipases A2; D008274:Magnesium; D002118:Calcium