Abnormal histone H3K9 dimethylation but normal dimethyltransferase EHMT2 expression in cloned sheep embryos

The objective was to investigate the relationship between histone H3 lysine 9 (H3K9) dimethylation (me2) and the histone methyltransferase EHMT2 (also known as G9A) in ovine embryos cloned by somatic cell nuclear transfer (SCNT). Levels of H3K9me2 or EHMT2 were detected (with immunostaining) and compared between SCNT and IVF-derived preimplantation embryos. In one-cell embryos, SCNT zygotes had significantly higher levels of H3K9me2 and EHMT2 than IVF zygotes. In cloned embryos, H3K9me2 remained hypermethylated relative to IVF embryos at two-cell and late developmental stages (morula and blastocyst), with no difference (P > 0.05) between IVF and SCNT embryos in EHMT2 levels from two-cell to blastocyst stages. The EHMT2-specific inhibitor, BIX01294, reduced global H3K9me2 levels in cultured ovine cells or SCNT embryos, but it was not appropriate for somatic cell nuclear transfer because of its high cellular toxicity. We inferred that abnormal H3K9me2 hypermethylation in SCNT embryos may not completely arise from EHMT2 expression error.