DNA methylation reprogramming (DMR) is believed to be a key process by which mammalian zygotes gain nuclear totipotency through erasing epigenetic modifications acquired during gametogenesis. Nonetheless, DMR patterns do not seem to be conserved among mammals. To identify uniform rules underlying mammalian DMRs, we explored DMRs of diverse mammalian zygotes. Of the zygotes studied, of particular interest was the bovine zygote; the paternal DNA methylation first decreased and was then rapidly restored almost to the maternal methylation level even before the two-cell stage. The 5-azadeoxycytidine treatment led to complete demethylation of the male pronucleus. The unusually dramatic changes in DNA methylation levels indicate that the bovine male pronucleus undergoes active demethylation, which is followed by de novo methylation. Our results show that, in bovine, the compound processes of active DNA demethylation and de novo DNA methylation, along with de novo H3-K9 trimethylation also, take place altogether within this very narrow window of pronucleus development. Developmental Dynamics 236:2523-2533, 2007.

The cytoplasm of a mature oocyte contains many protein complexes that are programmed to restructure incoming sperm chromatins on fertilization. Of the complicated biochemical events that these functional machineries control, the most impressive and important is epigenetic reprogramming. Despite its importance in epigenetic resetting, or "de-differentiation," of gamete genomes back to an incipient status, the mechanisms of epigenetic reprogramming do not seem to be conserved among mammals. Here, we report that, unlike in the mouse, the pig sperm-derived pronucleus is markedly trimethylated at lysine 9 of histone H3 (H3-m 3 K9), which might be associated with preservation of paternally derived cytosine methylation in pig zygotes. The male H3-m 3 K9 pattern is gradually established during pronucleus development, and this process occurs independently of DNA replication. Considering these unique epigenetic features, the pig zygote is, we believe, suited to serve as another model of epigenetic reprogramming that is antithetical to the well-characterized mouse model. Developmental Dynamics 236:1509 -1516, 2007.

Post-translational modifications of histones play important roles in regulating chromatin dynamics and epigenetic inheritance during mitosis. The epigenetic significance and stability of histone H3-lysine 9 (H3K9) modifications have been well studied in interphase cells, whereas not as much in mitotic cells. Here, we inspected mitosis-coupled alterations in the global modifications of H3K9. Signals for H3K9 mono-, di-methylation and acetylation became invisible as cells entered mitosis in contrast to the pattern observed for H3-serine 10 phosphorylation (H3S10ph). Treatment with the aurora-B inhibitor ZM447439 or expression of the dominant negative mutant Aur-B K106R resulted in prometaphase chromosomes that lacked signals for H3S10ph but were positive for H3K9 modifications. Trimethylation was the sole K9 modification that remained consistently detectable throughout the cell cycle. This phenomenon was specific for H3K9-S10, as this pattern was not observed at H3K27-S28. Methylated H3K27 remained detectable throughout the cell cycle, despite phosphorylation of the adjacent H3S28. Contrastingly, our dot-blot experiment using synthetic peptides showed that phosphorylation of serine residue basically kept adjacent lysine from antibody access. Together, these results suggest that phosphorylation of serine residue occurs in a selective manner, being influenced by the types of modifications and the nature of neighboring lysine residues.