Improvement in the in vitro development of cloned pig embryos after kdm4a overexpression and an H3K9me3 methyltransferase inhibitor treatment

Abstract

Aberrant epigenetic reprogramming is a major cause of the developmental failure of embryos after somatic cell nuclear transfer (SCNT). Histone H3 lysine 9 trimethylation (H3K9me3), a histone marker of transcriptional repression, is considered a key barrier to the development of cloned embryos. In the present study, H3K9me3 levels were much higher in SCNT embryos than IVF embryos at the 4-cell and 2-cell stages. The microinjection of the kdm4a mRNA encoding an H3K9me3 demethylase significantly increased the developmental efficiency of cloned porcine embryos. Moreover, we evaluated the effect of chaetocin, an inhibitor of histone methyltransferases suv39h1/2, on SCNT embryo development. Chaetocin did not suppress the H3K9me3 modification in porcine embryonic fibroblast (PEF) but downregulated the expression of suv39h1, suv39h2, and kdm4d. However, 10 nM chaetocin treatment efficiently decreased the H3K9me3 level in cloned embryos. Importantly, a chaetocin treatment at the 4-cell stage for 6 h significantly increased the blastocyst rate and total cell numbers. Furthermore, the inhibitor treatment upregulated the expression of related developmental genes. In summary, both overexpression of kdm4a and treatment with a suv39h1/2 inhibitor improve the epigenetic reprogramming of cloned embryos and further improve the developmental competence in vitro.

Keywords: Chaetocin; H3K9me3; Kdm4a; Pig; Somatic cell nuclear transfer.