Diradylglycerols stimulate phospholipase A2 and subsequent exocytosis in ram spermatozoa. Evidence that the effect is not mediated via protein kinase C.
We tested the hypothesis that the role of diacylglycerol (DAG) in sperm acrosomal exocytosis is related to the activation of phospholipase A2, and that this effect is not mediated via protein kinase C. Treatment of [14C]arachidonic acid-labelled ram spermatozoa with Ca2+ and the ionophore A23187 stimulated both liberation of arachidonic acid and acrosomal exocytosis. No changes in [14C]DAG or [14C]monoacylglycerol were found after stimulation of spermatozoa, thus suggesting that arachidonic acid may be released exclusively via phospholipase A2. An increase in the endogenous levels of diradylglycerols (DRGs), resulting from exposure either to the DAG kinase inhibitor R 59022 or to exogenous 1-oleoyl-2-acetyl-sn-glycerol or 1,2-dioctanoyl-sn-glycerol, led to an increase in both phospholipase A2 activity and exocytosis when cells were stimulated with A23187 and Ca2+. Addition of DRGs that do not stimulate protein kinase C(1,3-dioctanoylglycerol, 1-O-hexadecyl-2-acetyl-rac-glycerol) also resulted in an increase in phospholipase A2 activity and exocytosis. On the other hand, phorbol esters (phorbol 12,13-dibutyrate; phorbol 12-myristate 13-acetate) did not enhance enzyme activity or exocytosis. Finally, exposure to 1-O-hexadecyl-2-O-methyl-rac-glycerol, a compound known to inhibit protein kinase C, did not affect phospholipase A2 activity or acrosomal exocytosis. We therefore conclude that in spermatozoa the messenger role of DAG is related to the activation of phospholipase A2, which in turn would generate an array of metabolites directly or indirectly involved in bringing about exocytosis of the acrosome.
Roldan
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ER
Department of Development and Signalling, AFRC Babraham Institute, Cambridge, U.K.
Fragio
C
C
D000177:Acrosome / Q000201:enzymology; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D004075:Diglycerides / Q000378:metabolism / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D005089:Exocytosis / Q000187:drug effects*; D005989:Glycerides / Q000378:metabolism; D008297:Male; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D011493:Protein Kinase C / Q000378:metabolism*; D012756:Sheep; D013094:Spermatozoa / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D004075:Diglycerides; D005989:Glycerides; D016718:Arachidonic Acid; D015240:Phorbol 12,13-Dibutyrate; D000001:Calcimycin; C035561:diarachidonyl diglyceride; D011493:Protein Kinase C; D010741:Phospholipases A; D054467:Phospholipases A2; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium