METABOLISM OF A LONG-CHAIN DIACYLGLYCEROL BY PERMEABILIZED A10 SMOOTH-MUSCLE CELLS

The regulatory effects of diacylglycerol (DAG) second messengers will be terminated by metabolism. A long-chain DAG, 1-palmitoyl-2-[1-C-14]oleoyl-sn-glycerol (2-[C-14]POG), was metabolized by cultured A10 smooth muscle cells after permeabilization by preincubation with 340 U/ml alpha-toxin from Staphylococcus aureus. In contrast to results with the cell-permeable DAG analogue. dioctanoylglycerol ([H-3]diC8), no appreciabie 2-[C-14]POG degradation could be detected in control A10 cells not treated with alpha-toxin. With permeabilized A10 cells, 2-[C-14]POG was mainly converted into lipolytic products of a lipase pathway, monoacylglycerol (MG) and fatty acid (FA); very little radioactivity was incorporated into triacylglycerol (TG) or phospholipid (PL) via reactions catalyzed by either DAG acyltransferase, cholinephosphotransferase, or DAG kinase. Similar results were obtained in experiments with 1-stearoyl-2-[1-C-14]arachidonoyl-sn-glycerol. The conversion of 2-[C-14]POG into PL and TG was not enhanced by the addition of 1 mM ATP-MgCl2, 1 mM CDP-choline, or 1 mM oleoyl-CoA to the alpha-toxin-treated A10 cells. The formation of FA and MG by permeabilized A10 cells was inhibited by DAG lipase inhibitors, U-57,908 (50 muM) and tetrahydrolipstatin (1-25 nM). The predominant contribution of the lipase pathway to the metabolism of a long-chain DAG, 2-[C-14]POG, by alpha-toxin-treated A10 cells is similar to results for the degradation of [H-3]diC8 by intact A10 cells.