DNA methylation at a bovine alpha satellite I repeat CpG site during development following fertilization and somatic cell nuclear transfer

• Published in: PloS one Vol. 8; no. 2; p. e55153
• Main Authors: Couldrey, Christine, Wells, David N
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 01.02.2013; Public Library of Science (PLoS)
• Subjects: Adrenal Glands - metabolism; Analysis of Variance; Animal tissues; Animals; Artificial insemination; Biology; Blastocyst - metabolism; Blastocysts; Bovidae; Cattle; Cell division; Cloning; CpG islands; CpG Islands - genetics; CpG Islands - physiology; Deoxyribonucleic acid; DNA; DNA methylation; DNA Methylation - genetics; DNA Primers - genetics; DNA, Satellite - genetics; Embryos; Epigenesis, Genetic - physiology; Epigenetic inheritance; Epigenetics; Female; Fertilization; Fertilization in Vitro; Fetuses; Gene expression; Implantation; In vitro fertilization; Insemination, Artificial; Kidney - metabolism; Kidneys; Male; Methylation; Muscle, Skeletal - metabolism; Muscles; Nuclear transfer; Nuclear transfer (Cloning); Nuclear Transfer Techniques; Oocytes; Placenta; Placenta - metabolism; Pregnancy; Reproduction (biology); Semen - chemistry; Somatic cell nuclear transfer; Sperm
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0055153

Incomplete epigenetic reprogramming is postulated to contribute to the low developmental success following somatic cell nuclear transfer (SCNT). Here, we describe the epigenetic reprogramming of DNA methylation at an alpha satellite I CpG site (αsatI-5) during development of cattle generated either by artificial insemination (AI) or in vitro fertilization (IVF) and SCNT. Quantitative methylation analysis identified that SCNT donor cells were highly methylated at αsatI-5 and resulting SCNT blastocysts showed significantly more methylation than IVF blastocysts. At implantation, no difference in methylation was observed between SCNT and AI in trophoblast tissue at αsatI-5, however, SCNT embryos were significantly hyper-methylated compared to AI controls at this time point. Following implantation, DNA methylation at αsatI-5 decreased in AI but not SCNT placental tissues. In contrast to placenta, the proportion of methylation at αsatI-5 remained high in adrenal, kidney and muscle tissues during development. Differences in the average proportion of methylation were smaller in somatic tissues than placental tissues but, on average, SCNT somatic tissues were hyper-methylated at αsatI-5. Although sperm from all bulls was less methylated than somatic tissues at αsatI-5, on average this site remained hyper-methylated in sperm from cloned bulls compared with control bulls. This developmental time course confirms that epigenetic reprogramming does occur, at least to some extent, following SCNT. However, the elevated methylation levels observed in SCNT blastocysts and cellular derivatives implies that there is either insufficient time or abundance of appropriate reprogramming factors in oocytes to ensure complete reprogramming. Incomplete reprogramming at this CpG site may be a contributing factor to low SCNT success rates, but more likely represents the tip of the iceberg in terms of incompletely reprogramming. Until protocols ensure the epigenetic signature of a differentiated somatic cell is reset to a state resembling totipotency, the efficiency of SCNT is likely to remain low.