Protein synthesis is required for the transition to Ca(2+)-dependent regulated secretion in progesterone-matured Xenopus oocytes.
Calcium (Ca) ionophores trigger cortical granule exocytosis in progesterone-matured Xenopus oocytes (eggs), but not in immature oocytes. Prior work suggested that this secretory transition involved a Ca-dependent isoform of protein kinase C (PKC). To address this possibility, we treated eggs with several different inhibitors of Ca-dependent PKCs. Although these agents (eg., staurosporine, Ro31-8220) completely blocked cortical granule exocytosis that is triggered in eggs by phorbol esters, they had no impact on ionomycin-evoked secretion of cortical granule lectin. These data suggest that Ca-dependent PKCs do not mediate secretory triggering in eggs. Instead, further investigation revealed that protein synthesis (but not RNA synthesis) was required for eggs to secrete in response to ionomycin. Moreover, we observed that when oocytes were matured by injection of maturation promoting factor (MPF), they failed to secrete in response to ionomycin. Collectively, these results suggest that the progesterone-dependent maturation pathway induces these cells either to synthesize de novo, a protein that mediates Ca-dependent secretory triggering, or that intrinsic Ca-sensing machinery is modified in a protein-synthesis-dependent fashion. Initial efforts to distinguish between these possibilities (using Ca overlay, pharmacological and immunoblot strategies) revealed that such Ca-binding proteins as calmodulin, synaptotagmin1, CAPS, rabphilin-3A and calcineurin were unlikely to transduce the secretory effects of ionomycin in eggs. Thus, the cortical reaction in these cells may rely on a novel mechanism for initiating Ca-dependent exocytosis.
Kohan
Sirus A
SA
Department of Molecular & Medical Pharmacology, UCLA School of Medicine Los Angeles, CA 90095, USA.
Gundersen
Cameron B
CB
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D005089:Exocytosis / Q000502:physiology; D015151:Immunoblotting; D015759:Ionomycin / Q000378:metabolism; D007476:Ionophores / Q000378:metabolism; D016200:Maturation-Promoting Factor / Q000494:pharmacology; D009865:Oocytes / Q000187:drug effects* / Q000378:metabolism* / Q000502:physiology; D011374:Progesterone / Q000494:pharmacology*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D014981:Xenopus / Q000378:metabolism*
D007476:Ionophores; D011500:Protein Synthesis Inhibitors; D011374:Progesterone; D015759:Ionomycin; D011493:Protein Kinase C; D016200:Maturation-Promoting Factor; D002118:Calcium