Wilhelm bernhard workshop on the cell nucleus

Lack of epigenetic reprogramming in early sheep embryos produced by somatic cell nuclear transfer

Beaujean N.¹, Taylor J.², Gardner J.², Wilmut I.², Meehan R.³ and Young L.<sup>4

1</sup>Biologie du Développement et Reproduction, INRA Jouy-en-Josas, France, ²Department of Gene Expression and Development, Roslin Institute, Midlothian, UK, ³Department of Biomedical Sciences, University of Edinburgh, UK ⁴School of Human Development, Queens Medical Centre, University of Nottingham, UK
Low success rates have been universally reported for cloning by nuclear transfer, with most of the resulting animals presenting abnormalities. Cloning by nuclear transfer usually involves taking the nucleus from a somatic cell and transferring this into an egg previously stripped of its genetic content. The simplest explanation for the problems encountered with nuclear transfer is that they arise from incomplete reprogramming of the transferred nucleus. When the somatic nucleus is transferred into the enucleated egg, it needs to switch from its ‘somatic’ status to an ‘embryonic’ status and start the development program of an embryo. Addition of a methyl group to DNA (methylation) or removal (demethylation) are key regulators of gene expression. If somatic methylation marks are not removed and replaced by embryonic marks, this could interfere with embryonic gene expression and successful development. In this work we investigated DNA methylation in early sheep embryos. From examination of normal sheep embryos between zygote (Day 1) and blastocyst (Day 7), we found few changes in DNA methylation patterns. This is in contrast to the mouse where total demethylation is observed. However, in cloned sheep embryos global methylation was even higher than in normal embryos, suggesting that some of the somatic methylation marks had been retained. By the blastocyst stage they also showed very intense methylation in the trophectoderm cells, the first differentiated cells found in embryos which go on to form extra-embryonic tissues. This may be one of the causes of the abnormal placental formation which is frequently observed in cloned animals. These results match recent investigations of mouse and cattle cloned embryos suggesting abnormal reprogramming of DNA methylation is widespread in cloned animals, although the exact nature of the disturbance varies between species.