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Regular Article |
a Department of Veterinary Pathobiology
b Department of Animal Science, University of Missouri-Columbia, Columbia, Missouri 65211
c State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100080, People's Republic of China
We used okadaic acid (OA), a potent inhibitor of protein phosphatases 1 and 2A, to study the regulatory effects of protein phosphatases on mitogen-activated protein (MAP) kinase phosphorylation, morphological changes in the nucleus, and microtubule assembly during pig oocyte maturation and fertilization in vitro. When germinal vesicle (GV) stage oocytes were exposed to OA, MAP kinase phosphorylation was greatly accelerated, being fully activated at 10 min. However, MAP kinase was dephosphorylated by long-term (>20 h) exposure to OA. Correspondingly, premature chromosome condensation and GV breakdown were accelerated, whereas meiotic spindle assembly and meiotic progression beyond metaphase I stage were inhibited. OA also quickly reversed the inhibitory effects of butyrolactone I, a specific inhibitor of maturation-promoting factor (MPF), on MAP kinase phosphorylation and meiosis resumption. Treatment of metaphase II oocytes triggered metaphase II spindle elongation and disassembly as well as chromosome alignment disruption. OA treatment of fertilized eggs resulted in prompt phosphorylation of MAP kinase, disassembly of microtubules around the pronuclear area, chromatin condensation, and pronuclear membrane breakdown, but inhibited further cleavage. Our results suggest that inhibition of protein phosphatases promptly phosphorylates MAP kinase, induces premature chromosome condensation and meiosis resumption as well as pronucleus breakdown, but inhibits spindle organization and suppresses microtubule assembly by sperm centrosomes in pig oocytes and fertilized eggs.
1 This study was supported by grants from the University of Missouri-Columbia to H.S. and Food for the 21st Century to R.S.P. Q.Y.S., a research associate working in the laboratory of H.S., is supported by the Special Funds for Major State Basic Research ("973") project (G1999055902) while working at the University of Missouri. This manuscript is a contribution from the Missouri Agriculture Experiment Station, journal series 13151.
2 Correspondence: Heide Schatten, Department of Veterinary Pathobiology, W137 Veterinary Medicine Building, University of Missouri-Columbia, 1600 East Rollins, Columbia, MO 65211. FAX: 573 884 5414; schattenh{at}missouri.edu
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