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Biol Reprod. 2014 Nov;91(5):120. doi: 10.1095/biolreprod.114.120451. Epub 2014 Sep 17.

Establishment of paternal genomic imprinting in mouse prospermatogonia analyzed by nuclear transfer.

Author information

1
RIKEN BioResource Center, Tsukuba, Ibaraki, Japan Graduate School of Life and Environmental Science, University of Tsukuba, Ibaraki, Japan.
2
RIKEN BioResource Center, Tsukuba, Ibaraki, Japan.
3
Department of Epigenetics, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan.
4
RIKEN BioResource Center, Tsukuba, Ibaraki, Japan Department of Epigenetics, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan.
5
Department of Epigenetics, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan Global Center of Excellence Program for International Research Center for Molecular Science in Tooth and Bone Diseases, Tokyo Medical and Dental University, Tokyo, Japan PRESTO, Japan Science and Technology Agency, Saitama, Japan.
6
RIKEN BioResource Center, Tsukuba, Ibaraki, Japan Graduate School of Life and Environmental Science, University of Tsukuba, Ibaraki, Japan Center for Disease Biology and Integrative Medicine, Faculty of Medicine, University of Tokyo, Tokyo, Japan ogura@rtc.riken.go.jp.

Abstract

In mice, the establishment of paternal genomic imprinting in male germ cells starts at midgestation, as suggested by DNA methylation analyses of differentially methylated regions (DMRs). However, this information is based on averages from mixed populations of germ cells, and the DNA methylation pattern might not always provide a full representation of imprinting status. To obtain more detailed information on the establishment of paternal imprinting, single prospermatogonia at Embryonic Days 15.5 (E15.5), E16.5, and E17.5 and at Day 0.5 after birth were cloned using nuclear transfer; previous reports suggested that cloned embryos reflected the donor's genomic imprinting status. Then, the resultant fetuses (E9.5) were analyzed for the DNA methylation pattern of three paternal DMRs (IG-DMR, H19 DMR, and Rasgrf1 DMR) and the expression pattern of imprinted genes therein. The overall data indicated that establishment of genomic imprinting in all paternally imprinted regions was completed by E17.5, following a short intermediate period at E16.5. Furthermore, comparison between the methylation status of DMRs and the expression profiles of imprinted genes suggested that methylation of the IG-DMR, but not the H19 DMR, solely governed the control of its imprinted gene cluster. The Rasgrf1 DMR seemed to be imprinted later than the other two genes. We also found that the methylation status of the Gtl2 DMR, the secondary DMR that acquires DNA methylation after fertilization, was likely to follow the methylation status of the upstream IG-DMR. Thus, the systematic analyses of prospermatogonium-derived embryos provided additional important information on the establishment of paternal imprinting.

KEYWORDS:

DNA methylation; genomic imprinting; mouse; nuclear transfer; prospermatogonium

PMID:
25232016
DOI:
10.1095/biolreprod.114.120451
[Indexed for MEDLINE]

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