Molecular Mechanisms of Gonadotropin-Induced Oocyte Maturation
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Date
2004-01-10
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Abstract
In vitro maturation of oocytes is routinely utilized for the production of embryos for commercial as well as research purposes. The objectives of the research described in this dissertation were: 1) to examine the molecular mechanism involved in gonadotropin-induced resumption of meiosis in cultured bovine and murine cumulus oocyte complexes (COC); 2) to determine the developmental potential of bovine oocytes maintained in meiotic arrest by inhibition of transcription; and 3) to analyze patterns of gene expression in bovine COC during the onset of gonadotropin-induced oocyte maturation.
Murine COC were maintained in meiotic arrest by culture in the presence of either of the transcriptional inhibitors, 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole (DRB) or α-amanitin. For either transcriptional inhibitor to effectively block maturation, FSH but not hCG, was required in the culture medium. Transcriptional inhibition of oocyte maturation was ineffective if denuded oocytes were utilized. Differential activation of Type I and Type II PKA was performed in murine COC. Activation of Type I PKA resulted in the inhibition of maturation whereas activation of Type II did not. Activation of Type II PKA resulted in a transcriptional event required for maturation of murine and bovine COC and mimicked the action of FSH. The developmental competency of bovine COC maintained in meiotic arrest for 20 h by DRB was not different from control COC.
Comparison of the patterns of mRNA for oocytes matured for 0h, 4h or 4h+DRB resulted in the isolation of 4 amplicons that were expressed at 4h but not in 0h or 4h+DRB groups. This result was reconfirmed by semi-quantitative PCR. No homology to known sequences was found suggesting that they may represent novel transcripts.
The following model for FSH induced oocyte maturation is proposed: FSH binds to its cumulus cell receptor and increases cAMP. The elevation of cAMP results in activation of Type I and Type II PKA. Activation of Type I PKA inhibits oocyte maturation whereas activation of Type II PKA induces gene transcription that subsequently leads to the resumption of meiosis.
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Keywords
oocyte, transcription, gonadotropins, bovine, murine
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Degree
PhD
Discipline
Physiology
