DNA methylation and Cloning

Yong-Mahn Han

Although animal clones derived from somatic cells have been successfully produced in several mammalian species, developmental potential of the reconstructed embryos after nuclear transfer is still low. Developmental aberrancies in the present cloning system include a high abortion rate in early gestation and increased perinatal death. We may address these developmental failures in two ways. One is the reprogramming of the donor genome in the recipient oocytes during early embryonic development, and the other is the cloning procedure itself. We analyzed methylation patterns in cloned bovine embryos to monitor the epigenetic reprogramming process of donor genomic DNA. Epigenetic modification such as DNA methylation has been considered to be one of the putative mechanisms regulating nuclear reprogramming. In mammals, DNA methylation is an essential process in the regulation of transcription during embryonic development and is associated with gene silencing. Genome-wide demethylation may be a prerequisite for the formation of pluripotent stem cells that are important for later development. In this study, we have hypothesized that the anomalies in cloned animals may be due to incomplete epigenetic reprogramming of donor genomic DNA. Our findings demonstrated that aberrant methylation patterns, except single-copy sequences, were observed in various genomic regions of cloned bovine embryos. The overall genomic methylation status of cloned embryos was quite different from that of normal embryos produced in vitro or in vivo, but it closely resembled that of donor cells. There were also significant variations in the degree of methylation among individual cloned embryos. These results suggest that the developmental failures of cloned embryos may be due to incomplete epigenetic reprogramming of donor genomic DNA. In addition, we found a reverse relationship between methylation levels and inner cell mass versus trophectoderm ratios in cloned blastocysts. Trophectoderm-localized methylation aberrancy suggests a widespread gene dysregulation in the extraembryonic region, thereby resulting in placental dysfunction familiar to cloned fetuses and animals.