The protein phosphatase inhibitor, okadaic acid, inhibits phosphatidylcholine biosynthesis in isolated rat hepatocytes.
There is evidence that phosphatidylcholine (PC) biosynthesis in hepatocytes is regulated by a phosphorylation-dephosphorylation mechanism. The phosphatases involved have not been identified. We, therefore, investigated the effect of okadaic acid, a potent protein phosphatase inhibitor, on PC biosynthesis via the CDP-choline pathway in suspension cultures of isolated rat hepatocytes. Okadaic acid caused a 15% decrease (P less than 0.05) in [Me-3H]choline uptake in continuous-pulse labeling experiments. After 120 min of treatment, the labeling of PC was decreased 46% (P less than 0.05) with a corresponding 20% increase (P less than 0.05) in labeling of phosphocholine. Cells were pulsed with [Me-3H]choline for 30 min and subsequently chased for up to 120 min with choline in the absence or presence of okadaic acid. The labeling of phosphocholine was increased 86% (P less than 0.05) and labeling of PC decreased 29% (P less than 0.05) by 120 min of chase in okadaic acid-treated hepatocytes. The decrease of label in PC was quantitatively accounted for in the phosphocholine fraction. Incubation of hepatocytes with both okadaic acid and CPT-cAMP did not produce an additive inhibition in labeling of PC. Choline kinase and cholinephosphotransferase activities were unaltered by treatment with okadaic acid. Hepatocytes were incubated with digitonin to cause release of cytosolic components. Cell ghost membrane cytidylyltransferase (CT) activity was decreased 37% (P less than 0.005) with a concomitant 33% increase (P less than 0.05) in released cytosolic cytidylyltransferase activity in okadaic acid-treated hepatocytes. We postulate that CT activity and PC biosynthesis are regulated by protein phosphatase activity in isolated rat hepatocytes.
Hatch
G M
GM
Lipid and Lipoprotein Research Group, University of Alberta, Edmonton, Canada.
Tsukitani
Y
Y
Vance
D E
DE
D000818:Animals; D002478:Cells, Cultured; D002797:Choline Kinase / Q000378:metabolism; D002798:Diacylglycerol Cholinephosphotransferase / Q000378:metabolism; D004988:Ethers, Cyclic / Q000494:pharmacology*; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D019319:Okadaic Acid; D010713:Phosphatidylcholines / Q000096:biosynthesis*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D051381:Rats; D011919:Rats, Inbred Strains; D013347:Subcellular Fractions / Q000378:metabolism
D004988:Ethers, Cyclic; D010713:Phosphatidylcholines; D019319:Okadaic Acid; D002797:Choline Kinase; D002798:Diacylglycerol Cholinephosphotransferase; D010749:Phosphoprotein Phosphatases