PR-Set7/Set8/KMT5A is the sole enzyme known to catalyze monomethylation of histone H4 lysine 20 (H4K20) and is present only in multicellular organisms that compact a large fraction of their DNA. We found that mouse embryos that are homozygous null mutants for the gene PR-Set7 display early embryonic lethality prior to the eight-cell stage. Death was due to the absence of PR-Set7 catalytic activity, since microinjection of the wild type, but not a catalytically inactive version, into two-cell embryos rescued the phenotype. A lack of PR-Set7 activity resulted not only in depletion of H4K20me1 but also in reduced levels of the H4K20me2/3 marks catalyzed by the Suv4-20h1/h2 enzymes, implying that H4K20me1 may be essential for the function of these enzymes to ensure the dimethylated and trimethylated states. Embryonic stem cells that were inducibly deleted for PR-Set7 passed through an initial G 2 /M phase, but the progeny were defective at the subsequent S and G 2 /M phases, exhibiting a delay in their cell cycle, accumulation at G 2 /M, massive DNA damage, and improper mitotic chromosome condensation. Cell cycle analysis after synchronization indicated that the defects were a consequence of decreased H4K20me1 due to the absence of PR-Set7. Most importantly, the lack of H4K20me1 also resulted in defects in chromosome condensation in interphase nuclei. These results demonstrate the critical role of H4K20 monomethylation in mammals in a developmental context.Posttranslational modifications (PTMs) on histones influence intra-and internucleosomal interactions and thereby contribute to the diversity in nucleosome and chromatin structure that impacts distinct genomic processes. Among these modifications, histone methylation had been considered to be relatively stable, but recent studies demonstrated that, similar to the other PTMs, it too is subject to regulation. Many methylating and demethylating enzymes that target different lysine and arginine residues have been identified. These enzymes have distinct specificities with respect to the methylation status (monomethyl, dimethyl, and trimethyl) of each residue. Furthermore, increasing numbers of proteins harboring the motifs that specifically recognize various methylated residues have been identified. These proteins, or effectors, mediate/regulate elaborate chromatin-based processes, such as gene expression, which are dictated by the presence of the PTMs. Thus, the catalysis/removal of PTMs and their recognition by effectors constitute an intricately designed system that is key to genomic integrity and function.One of the residues of histone H4 that can be monomethylated, dimethylated, or trimethylated is lysine 20. Recent comprehensive analysis of H4 modifications with top-down mass spectrometry in human and in Drosophila melanogaster cells revealed that the dimethyl group is deposited on the majority of total H4K20, indicative of the wide distribution of H4K20me2 on chromatin (18, 34). On the other hand, H4K20me1 and H4K20me3 are relatively few in abundance. The study a...

Summary Random X inactivation represents a paradigm for monoallelic gene regulation during early ES cell differentiation. In mice, the choice of X chromosome to inactivate in XX cells is ensured by monoallelic regulation of Xist RNA via its antisense transcription unit Tsix/Xite. Homologous pairing events have been proposed to underlie asymmetric Tsix expression, but direct evidence has been lacking owing to their dynamic and transient nature. Here we investigate the live-cell dynamics and outcome of Tsix pairing in differentiating mouse ES cells. We find an overall increase in genome dynamics including the Xics during early differentiation. During pairing, however, Xic loci show markedly reduced movements. Upon separation, Tsix expression becomes transiently monoallelic, providing a window of opportunity for monoallelic Xist upregulation. Our findings reveal the spatiotemporal choreography of the X chromosomes during early differentiation and indicate a direct role for pairing in facilitating symmetry-breaking and monoallelic regulation of Xist during random X inactivation.

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