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Mol. Hum. Reprod. Advance Access originally published online on May 20, 2005
Molecular Human Reproduction 2005 11(6):397-404; doi:10.1093/molehr/gah184
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© The Author 2005. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions{at}oupjournals.org

RNA interference in meiosis I human oocytes: towards an understanding of human aneuploidy

Hayden A. Homer1,2,5, Alex McDougall2,4, Mark Levasseur2, Alison P. Murdoch1,3 and Mary Herbert1,3,5

1Newcastle Fertility Centre at Life, International Centre for Life, Times Square, Newcastle upon Tyne NE1 4EP, 2School of Cell and Molecular Biosciences and 3School of Surgical and Reproductive Sciences, The Medical School, Framlington Place, University of Newcastle, Newcastle upon Tyne NE2 4HH, UK4Present address: UMR 7009 CNRS/Université Pierre et Marie Curie (Paris VI), Observetoire Océanologique, 06230 Villefranche-sur-Mer, France

5 To whom correspondence should be addressed at: Wansbeck General Hospital, Woodhorn Lane, Ashington, Northumberland NE63 9JJ, UK. Email: h.a.homer{at}ncl.ac.uk

Although female meiosis I errors account for the majority of human aneuploidy, their molecular basis is largely unknown. By elucidating gene function, gene knockdown using RNA interference (RNAi) could shed light on this enigmatic process. In practice, however, the extreme paucity of immature human oocytes makes the evaluation of gene-targeting tools difficult. Here, we undertake RNAi in human oocytes and describe an approach employing mouse oocytes which could overcome the problem of limited biological material. We designed a short interfering RNA (siRNA) designated si539 to target the human mitotic arrest deficient 2 (hMad2) spindle checkpoint component. In human oocytes microinjected with si539, the hMad2 signal detected by Western blotting was 85–92% less intense than in oocytes injected with control siRNA indicating efficient silencing. Further examination of si539's targeting efficiency was undertaken using a green fluorescent protein (GFP)-tagged hMad2 mRNA construct in mouse oocytes. Consistent with Western blot analysis, si539 reduced hMad2-GFP expression in mouse oocytes by ~94% and relieved the meiosis I arrest otherwise induced by hMad2-GFP in mouse oocytes. By facilitating the investigation of candidate genes involved in regulating human female meiosis I, this approach can bring us closer to understanding the origins of aneuploidies such as Down's syndrome.

Key words: green fluorescent protein/Mad2/meiosis I/oocytes/RNA interference


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H. A Homer, A. McDougall, M. Levasseur, A. P Murdoch, and M. Herbert
Mad2 is required for inhibiting securin and cyclin B degradation following spindle depolymerisation in meiosis I mouse oocytes
Reproduction, December 1, 2005; 130(6): 829 - 843.
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