Increased phosphorylation and dimethylation of XY body histones in the Hr6b-knockout mouse is associated with derepression of the X chromosome

Mono-ubiquitylated H2A marks the transcriptionally silenced XY body during male meiotic prophase. Concomitant with H2AK119ub1, the ubiquitin-conjugating enzyme HR6B is also enriched on the XY body. We analyzed H2A and H2B ubiquitylation in Hr6b-knockout mouse spermatocytes, but no global changes were detected. Next, we analyzed phosphorylation of the threonine residues T120 and T119 that are adjacent to the K119 and K120 target sites for ubiquitylation in H2A and H2B, respectively. In wild-type cells, H2AT120phand H2BT119phmark meiotically impaired and silenced chromatin, including the XY body. In Hr6b-knockout spermatocytes, the H2BT119phsignal was unchanged, but H2AT120phwas enhanced from late pachytene until metaphase I. Furthermore, we found increased H3K4dimethylation on the X and Y chromosomes of diplotene Hr6b-knockout spermatocytes, persisting into postmeiotic round spermatids. In these cells, the X and Y chromosomes maintained an unchanged H3K9m2level, even when this modification was lost from centromeric heterochromatin. Analysis of gene expression showed derepression of X chromosome genes in postmeiotic Hr6b-knockout spermatids. We conclude that HR6B exerts control over different histone modifications in spermatocytes and spermatids, and that this function contributes to the postmeiotic maintenance of X chromosome silencing.

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