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Dynamic Events Are Differently Mediated by Microfilaments, Microtubules, and Mitogen-Activated Protein Kinase During Porcine Oocyte Maturation and Fertilization In Vitro¹

^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, China

ABSTRACT

The role of microfilaments, microtubules, and mitogen-activated protein (MAP) kinase in regulation of several important dynamic events of porcine oocyte maturation and fertilization is described. Fluorescently labeled microfilaments, microtubules, and cortical granules were visualized using either epifluorescence microscopy or laser scanning confocal microscopy. Mitogen-activated protein kinase phosphorylation was revealed by Western immunoblotting. We showed that 1) microfilament disruption did not affect meiosis resumption and metaphase I meiotic apparatus formation but inhibited further cell cycle progression (chromosome separation) even though MAP kinase was phosphorylated; 2) cortical granule (CG) migration was driven by microfilaments (but not microtubules), and once the chromosomes and CGs were localized beneath the oolemma their anchorage to the cortex was independent of either microfilaments or microtubules; 3) neither microfilaments nor microtubules were involved in CG exocytosis during oocyte activation; 4) sperm incorporation was mediated by microfilaments, while pronuclear (PN) syngamy was controlled by microtubules rather than microfilaments; 5) spindle microtubule organization was temporally correlated with MAP kinase phosphorylation, while the extensive microtubule organization in the sperm aster that is required for PN apposition and syngamy occurred in the absence of MAP kinase activation; and 6) MAP kinase phosphorylation did not change either when microtubules were disrupted by nocodazole or when cytoplasmic microtubule asters were induced by taxol. The present study suggests that the role of the cytoskeleton during porcine oocyte maturation is similar to that of rodents, while the mechanisms of fertilization in pig resemble those of lower vertebrates.

FOOTNOTES

First decision: 6 September 2000.

¹ This study was supported by grants from the University of Missouri-Columbia to H.S., Food for the 21st Century Program to R.S.P. Q.Y.S., a research associate working in H.S.'s lab, is supported by the "973" project (G1999055902) while working at the University of Missouri. This manuscript is a contribution from the Missouri Agriculture Experiment Station, journal series no. 13051.

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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