Interplay of maturation-promoting factor and mitogen-activated protein kinase inactivation during metaphase-to-interphase transition of activated bovine oocytes

The objective of the present study was to examine the activity changes in h istone H1 kinase (also known as maturation-promoting factor [MPF]) and mito gen-activated protein kinase (MAPK) and their constituent proteins in in vi tro-matured bovine oocytes after in vitro fertilization (IVF) or after part henogenetic activation induced by calcium ionophore A23187 alone or by the ionophore followed by either 6-dimethylaminopurine (6-DMAP) or cycloheximid e (CHX), Inactivation of both H1 kinase and MAPK occurred after both A23187 +6-DMAP treatment and IVF; inactivation of H1 kinase preceded inactivation of MAPK, However, MAPK was inactivated much earlier in 6-DMAP-treated oocyt es, Further analysis of constituent cell cycle proteins of these kinases by Western blot showed that A23187 alone could not induce changes in cdc2, cd c25, or ERK2 but induced reduction of cyclin B1, IVF and A23187+CHX induced similar changes: cyclin B1 was destroyed shortly after activation followed by accumulation of cyclin B1, phosphorylation of cdc2, and dephosphorylati on of ERK2 at pronuclear formation 15 h after activation. No change in cdc2 5 was observed at this time. In contrast, A23187+6-DMAP treatment resulted in earlier phosphorylation of cdc2 and dephosphorylation of ERK2 at 4 h aft er treatment when the pronucleus formed. Moreover, accumulation of both cdc 25 and cyclin B1 was detected at 15 h, Microinjection of ERK2 antibody into A23187-treated oocytes resulted in pronuclear formation. In conclusion, ac tivation of bovine oocytes with 6-DMAP led to earlier inactivation of MAPK, while CHX induced inactivation of MAPK parallel to that following sperm-in duced oocyte activation. Destruction of cyclin B is responsible for inactiv ation of MPF, while phosphorylation of cdc2 is likely responsible for maint aining its low activity, Inactivation of MAPK is closely associated with pr onuclear development regardless of the activation protocol used.