Setdb1/Eset, a histone lysine methyltransferase, is recruited by various transcription factors to modify local chromatin. The observation that Setdb1-null blastocysts fail to produce epiblast-lineage cells suggests a role for Setdb1 in generating mouse embryonic stem cells (mESCs). When examined in mouse zygotes, Setdb1 proteins appeared as dots at perinucleolar rims of pronuclei, with the dot-shaped signals more prominent in male pronuclei. Setdb1 signals were observed diffusely in the nucleus from the two-cell stage onward and, by the blastocyst, took a punctate form, away from nucleolus. Such varying expression patterns suggest its involvement in diverse biological processes at preimplantation stage. Setdb1 appeared in Oct4-positive cells of inner-cell-mass origin but not in trophectoderm-lineage cells in blastocyst outgrowths. Setdb1 co- immunoprecipitated with Oct4 in mESCs, and Setdb1 expression was markedly reduced upon retinoic acid- induced differentiation. These observations suggest that Setdb1 has an important role in maintaining the self-renewal of mESCs through collaboration with Oct4.

Covalent modifications of histone tails have fundamental roles in chromatin structure and transcriptional activity of a target locus. One of such modifications, Methylation at Lysine 9 of histone H3 (H3-K9) causes several epigenetic phenomena including heterochromatin formation, transcriptional regulation and DNA methylation. Setdb1, H3-K9 specific histone methyltransferase, functions in gene silencing, heterochromatin formation and essential role for early development. Here, we demonstrate that Setdb1 associates with promyelocytic leukemia (Pml) protein from the early stage of mouse development and is a constitutive member of PML nuclear bodies (PML-NBs) that have been linked to many cellular processes such as apoptosis, DNA damage responses, and transcriptional regulation. Immunostaining of mouse blastocyst showed that Setdb1 and Pml signals were scattered in nucleus as a few speckles and microinjected Pmlmyc signals colocalize with Setdb1 signals. This colocalization was observed in mEF and the punctate signals of Setdb1 were observed to be present in every nucleus of mEFs and dividing cells with condensed chromosomes. Arsenic treatment, which induces Pml degradation, also caused Setdb1 signals to disappear. Setdb1 knockdown resulted in disassembly of PML-NBs and immunoprecipitation results demonstrated physical interactions between Setdb1 and Pml. These data suggest that Setdb1 was associated in PML-NB and Setdb1 has important function in maintenance of PML-NB structure.