Limited demethylation leaves mosaic-type methylation states in cloned bovine pre-implantation embryos.
Cloning by nuclear transfer (NT) has been riddled with difficulties: most clones die before birth and survivors frequently display growth abnormalities. The cross-species similarity in abnormalities observed in cloned fetuses/animals leads us to suspect the fidelity of epigenetic reprogramming of the donor genome. Here, we found that single-copy sequences, unlike satellite sequences, are demethylated in pre-implantation NT embryos. The differential demethylation pattern between genomic sequences was confirmed by analyzing single blastocysts. It suggests selective demethylation of other developmentally important genes in NT embryos. We also observed a reverse relationship between methylation levels and inner cell mass versus trophectoderm (ICM/TE) ratios, which was found to be a result of another type of differential demethylation occurring in NT blastocysts where unequal methylation was maintained between ICM and TE regions. TE-localized methylation aberrancy suggests a widespread gene dysregulation in an extra-embryonic region, thereby resulting in placental dysfunction familiar to cloned fetuses/animals. These differential demethylations among genomic sequences and between differently allocated cells produce varied overall, but specified, methylation patterns, demonstrating that epigenetic reprogramming occurs in a limited fashion in NT embryos.
Kang
Yong-Kook
YK
Animal Developmental Biotechnology Laboratory, Korea Research Institute of Bioscience and Biotechnology, PO Box 115, Yusong, Taejon 305-600, South Korea.
Park
Jung Sun
JS
Koo
Deog-Bon
DB
Choi
Young-Hee
YH
Kim
Sun-Uk
SU
Lee
Kyung-Kwang
KK
Han
Yong-Mahn
YM
D000818:Animals; D001755:Blastocyst / Q000378:metabolism*; D002417:Cattle / Q000196:embryology*; D002418:Cattle Diseases / Q000196:embryology / Q000235:genetics; D019070:Cell Lineage; D002467:Cell Nucleus; D019976:Cloning, Organism / Q000662:veterinary*; D000013:Congenital Abnormalities / Q000196:embryology / Q000235:genetics / Q000662:veterinary; D018899:CpG Islands; D019175:DNA Methylation; D004276:DNA, Satellite / Q000235:genetics; D004475:Ectoderm / Q000166:cytology; D005307:Fertilization in Vitro; D018507:Gene Expression Regulation, Developmental; D018390:Gene Targeting; D007633:Keratins / Q000235:genetics; D007782:Lactoglobulins / Q000235:genetics; D008845:Microinjections; D009030:Mosaicism; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D011401:Promoter Regions, Genetic / Q000235:genetics; D014327:Trophoblasts / Q000166:cytology
D004276:DNA, Satellite; D007782:Lactoglobulins; D007633:Keratins