Progesterone triggers the rapid activation of phospholipase D in the amphibian oocyte plasma membrane when initiating the G(2)/M transition

Previous reports indicate that, in the Rana pipiens oocyte, progesterone triggers a rapid rise in 1,2-diacylglycerol (DAG) derived from phosphatidylcholine (PC) in the plasma membranes. This DAG transient, which appears and is terminated within 60-90 s, is derived both from a phospholipase which we assumed to be phospholipase C and from sphingomyelin (SM) synthase. We now find that progesterone stimulates PC and DAG turnover primarily via the phospholipase D (PLD) and phosphatidic acid phosphohydrolase (PAP) pathways as well as via the SM-ceramide pathway. Rana oocytes were prelabeled with [H-3]choline chloride under conditions in which about 70% is incorporated into C of the plasma membrane of the intact oocyte or with [H-3]lysoplatelet activating factor (1-O-octadecyl-sn-glycero-3-phosphocholine, lysoPAF) which is selectively incorporated into plasma membrane PC. Progesterone induced the release of [H-3]choline from intact oocytes into the medium within 60-90 s. This choline release was dose-dependent and was not inhibited by a putative PC-specific phospholipase C inhibitor, D609. Progesterone also induced a transient rise in [H-3]lysoPAF-derived [H-3]DAG within 1-2 min followed by a rise in [H-3]PA. In the presence of 20 mM ethanol, progesterone stimulated formation of [H-3]lysoPAF-derived phosphatidylethanol, indicating progesterone activation of PC-specific PLD and concomitant formation of PA. A DGK inhibitor (D102) reduced the level of [H-3]PA, produced a sustained rise in [H-3]DAG and was a weak inducer of meiosis in oocytes not exposed to progesterone. A PA phosphohydrolase inhibitor (propranolol) elevated [H-3]PA and completely inhibited the progesterone-induced rise in DAG. Progesterone thus acts at oocyte plasma membrane receptors to release PC-derived DAG via both SM synthase and PC-PLD. The duration of the DAG signal is regulated by the coordinate action of DGK and PAP.