Abstract
DNA methylation imprints that are established in spermatogenesis and oogenesis are essential for functional gametes. However, the mechanisms underlying gamete-specific imprinting remain unclear. In this study, we investigated whether male and female gametes derived from newborn mice are epigenetically plastic and whether DNA methylation imprints are influenced by the niche surrounding the nuclei of the gametes. When prospermatogonia possessing sperm-specific DNA methylation imprints were fused with enucleated fully grown oocytes and exposed to the ooplasm for 5–6 days, the DNA methylation status of the reconstituted oocytes remained identical to that of prospermatogonia for all the imprinted regions analysed. These results suggest that the imprinting status of prospermatogonia is stable and that the epigenome of prospermatogonia loses sexual plasticity. By contrast, when non-growing oocytes lacking oocyte-specific DNA methylation imprints were fused with enucleated fully grown oocytes and the reconstituted oocytes were then cultured for 5–6 days, the Igf2r, Kcnq1ot1 and, unexpectedly, H19/Igf2 differentially methylated regions (DMRs) were methylated. Methylation imprints were entirely absent in oocytes derived from 5-day-old mice, and H19/Igf2 DMR is usually methylated only in spermatogenesis. These findings indicate that in the nuclei of non-growing oocytes the chromatin conformation changes and becomes permissive to DNA methyltransferases in some DMRs and that mechanisms for maintaining non-methylated status at the H19/Igf2 DMR are lost upon long exposure to mature ooplasm.
Highlights
DNA methylation imprints that are established in spermatogenesis and oogenesis are maintained after fertilisation, which results in parental-origin-specific gene expression in the somatic cell lineage
It has been reported that the maintenance of allelespecific DNA methylation is required for protection against DNA demethylation by pluripotency-associated protein 3 (DPPA3; Nakamura et al 2007, 2012), maintenance of DNA methylation by DNA methyltransferase 1 (DNMT1; Li et al 1993), and maintenance of the unmethylated status of differentially methylated regions (DMRs) by CTCF (Schoenherr et al 2003, Szabo et al 2004)
Sperm-specific DNA methylation imprints are established during the mitotic arrest of prospermatogonia in foetal testes (Davis et al 2000, Hiura et al 2007), whereas oocyte-specific DNA methylation imprints are established during oocyte growth, and oocytes possessing all imprints first appear in the peri-pubertal stage (Lucifero et al 2004, Hiura et al 2006)
Summary
DNA methylation imprints that are established in spermatogenesis and oogenesis are maintained after fertilisation, which results in parental-origin-specific gene expression in the somatic cell lineage. To determine whether DNA methylation imprints are influenced by the niche surrounding the nuclei of the gametes, we conducted fusion experiments using nuclear transfer technique and COC reconstitution for long exposure of nuclei derived from gametes in the cytoplasm of fully grown oocytes (Fig. 1). Imprinting status of nuclei derived from prospermatogonia after long exposure to ooplasm As has been reported previously (Davis et al 2000, Hiura et al 2007), H19/Igf2 and Dlk1/Gtl2 intergenic (IG) DMRs were fully methylated, whereas Igf2r, Kcnq1ot1 and Mest DMRs were not methylated in prospermatogonia derived from newborn mice (Fig. 2 and Table 1).
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