116 ACTIVE METHYLATION AND ACETYLATION OF HISTONE H3-K9 IN MOUSE EMBRYO WITH DIFFERENT PROPORTIONS OF MATERNAL AND PATERNAL GENOME

Epigenetic modification of parental genomes plays a prominent role in regulating genome expression in the early development of embryos. In general, histone H3 of the paternal genome is demethylated at lysine 9 (H3-K9) during the first and second mitotic divisions in fertilized embryos, while the maternal genome is methylated. We investigated the effects of maternal genomes (Mgen) and paternal genomes (Pgen) on H3-K9 methylation and acetylation during the early development of murine embryos. Histone H3-K9 methylation and acetylation were detected by anti-trimethyl histone H3-K9 and anti-triacetyl histone H3-K9 antibodies. The following embryos were used in this study: (1) intracytoplasmic sperm injection (ICSI) embryos (50% Mgen, 50% Pgen); (2) parthenogenetic diploid embryos (100% Mgen, 0% Pgen); (3) somatic nuclear transfer embryos (50% Mgen, 50% Pgen from previous generation); (4) androgenetic diploid embryos (0% Mgen, 100% Pgen); and (5) haploidized somatic nucleus and sperm embryo (about 25% Mgen, about 75% Pgen). Each experiment was repeated five times to obtain more than 120 embryos per group. Our results show that: (1) in the ICSI embryo, histone H3 methylation occurs in Mgen but not in Pgen at the first and second mitotic divisions; (2) in the parthenogenetic embryo, histone H3 methylation occurs in both nuclei at the first and second mitotic divisions; (3) in the somatic nuclear transfer embryo, histone H3 is methylated in all of the nuclei at the first and second mitotic divisions; (4) in the androgenetic embryo, methylated H3-K9 is detected weakly in the heterochromatin enclosed around the nucleolus of the pronuclei of the one-cell embryo, and methylated in the entire nuclei of the two-cell embryo; and (5) in the haploidized somatic and sperm embryo, the pattern of histone H3-L9 methylation resembles that of the ICSI embryo. While histone H3-K9 acetylation occurs in both paternal and maternal genomes during interphase, even when the nuclear membrane is completely degraded and the chromosome is condensed, it disappears rapidly when the chromosome enters the real metaphase, and reappears at the early stage of pronuclear formation in all types of embryo. These results suggest that the absence of maternal genomes results in histone H3-K9 methylation in the paternal genomes during the first and second mitotic divisions of embryos in mice. In addition, histone H3-K9 acetylation is independent of the presence or absence of maternal or paternal genomes during pre-implantation development in mice. This study was supported by grants-in-aid for Creative Scientific Research (13GS0008) and a project for the realization of regenerative medicine (the research field for the technical development of stem cell manipulation) to T.W. from MEXT, Japan.