STEROID ACTION AT THE PLASMA-MEMBRANE - PROGESTERONE STIMULATION OF PHOSPHATIDYLCHOLINE-SPECIFIC PHOSPHOLIPASE-C FOLLOWING RELEASE OF THE PROPHASE BLOCK IN AMPHIBIAN OOCYTES

Progesterone, acting at the amphibian oocyte plasma membrane, triggers the progression of the prophase oocyte nucleus through the first meiotic metaphase. We previously reported a transient increase in 1,2-diacylglycerol (1,2-DG) within the first 1-2 min after exposure of Rana pipiens oocytes to progesterone. We have now investigated the source of the 1,2-DG, using this highly synchronous oocyte population. Phospholipid pools of intact prophase-arrested oocytes were labeled with [H-3]glycerol, [methyl-H-3]choline chloride or 1-O-[H-3]octadecyl-sn-glycero-3-phosphocholine (lyso platelet activating factor, lysoPAF). [H-3]LysoPAF is selectively taken up into the plasma membrane of the intact oocyte and esterified to form the [H-3]alkyl-analogue of phosphatidylcholine (PC). Intact oocytes and/or isolated plasma membranes were then stimulated with progesterone and the changes in [H-3]DG, [methyl-H-3]phosphocholine and [H-3]phospholipids were monitored as a function of time. Progesterone induced a transient increase in [H-3]glycerol-derived DG, [methyl-H-3]phosphocholine and [H-3]alkyl-2-acylglycerol from [H-3]alkyl-PC within the first 2 min, indicating activation of a PC-specific phospholipase C. Different pulse-labeling conditions indicate a biphasic rise in [H-3]DG from [H-3]glycerol-labeled oocytes; the first rise (1-2 min) when phospholipid labeling in the plasma membrane is enriched followed by an approximately 3-fold larger rise at 5-15 min when phospholipids of intracellular membranes are preferentially labeled. An early transient increase in [H-3]DG or [H-3]alkyl-2-acylglycerol was also seen when progesterone and/or guanosine 5'-O-(3-thiotriphosphate) (GTP-gamma-S) were added to isolated plasma-vitelline membranes prepared from oocytes prelabeled with either [H-3]glycerol or [H-3]lysoPAF. Progesterone thus appears to activate a G-protein-linked PC-specific phospholipase C in the oocyte plasma membrane which is followed by much larger DG release from intracellular membranes. The transient character of the hydrolysis suggests that this may represent a mechanism for transducing a membrane event into a meiotic signal.