MASTOPARAN-INDUCED PHOSPHATIDYLCHOLINE HYDROLYSIS BY PHOSPHOLIPASE-D ACTIVATION IN HUMAN ASTROCYTOMA-CELLS

1 The effect of mastoparan on phosphatidylcholine hydrolysis was exami ned in 1321N1 human astrocytoma cells. Mastoparan (3-30 mu M) caused a n accumulation of diacylglycerol (DG) and phosphatidic acid (PA) accom panied by choline release in a concentration- and time-dependent manne r. 2 In the presence of 2% n-butanol, mastoparan (3-100 mu M) induced phosphatidylbutanol (PBut) accumulation in a concentration- and time-d ependent manner, suggesting that mastoparan activates phospholipase D (PLD). Propranolol (30-300 mu M), a phosphatidate phosphohydrolase inh ibitor, inhibited DG accumulation induced by mastoparan, supporting th is idea. 3 Depletion of extracellular free calcium ion did not alter t he effect of mastoparan on PLD activity. 4 A protein kinase C (PKC) in hibitor, calphostin C (1 mu M), did not inhibit mastoparan-induced PLD . activation but the ability of mastoparan to stimulate phospholipase D activity was decreased in the PKC down regulated cells. 5 PLD activi ty stimulated by mastoparan was not prevented by pretreatment of the c ells with pertussis toxin (PT) or C3 ADP-ribosyltransferase. Furthermo re, guanine nucleotides did not affect PLD activity stimulation by mas toparan in membrane preparations. 6 Mastoparan stimulated PLD in sever al cell lines such as RBL-2H3, RBL-1, HL-60, P388, endothelial cells, as well as 1321N1 human astrocytoma cells. 7 These results suggest tha t mastoparan induces phosphatidylcholine (PC) hydrolysis by activation of PLD, not by activation of phosphatidylcholine-specific phospholipa se C (PC-PLC); mastoparan-induced PLD activation is not mediated by G proteins.