RYBP-PRC1 Complexes Mediate H2A Ubiquitylation at Polycomb Target Sites Independently of PRC2 and H3K27me3

Abstract

SummaryPolycomb-repressive complex 1 (PRC1) has a central role in the regulation of heritable gene silencing during differentiation and development. PRC1 recruitment is generally attributed to interaction of the chromodomain of the core protein Polycomb with trimethyl histone H3K27 (H3K27me3), catalyzed by a second complex, PRC2. Unexpectedly we find that RING1B, the catalytic subunit of PRC1, and associated monoubiquitylation of histone H2A are targeted to closely overlapping sites in wild-type and PRC2-deficient mouse embryonic stem cells (mESCs), demonstrating an H3K27me3-independent pathway for recruitment of PRC1 activity. We show that this pathway is mediated by RYBP-PRC1, a complex comprising catalytic subunits of PRC1 and the protein RYBP. RYBP-PRC1 is recruited to target loci in mESCs and is also involved in Xist RNA-mediated silencing, the latter suggesting a wider role in Polycomb silencing. We discuss the implications of these findings for understanding recruitment and function of Polycomb repressors.

Characterization of non-canonical Polycomb Repressive Complex 1 subunits during early mouse embryogenesis

“…This observation was surprising given the expected nuclear localization of RYBP based on findings in other cell types. 15,37 However, immediately after fertilization, we did not detect RYBP at the zygote stage, neither in the cytoplasm nor in the nucleoplasm of any of the 2 pronuclei ( Fig. 2A).…”

“…Subsequent recruitment of PRC1 then leads to H2AK119ub on those domains through the enzymatic activity of RING1A/RING1B, and to increased chromatin compaction. However, recent evidence supports an emerging view whereby PRC1 is indeed recruited to genomic regions occupied by PRC2 and in a manner dependent on H3K27, but additionally, PRC1 can be recruited onto chromatin independently of H3K27me3, presumably through alternative protein-protein interactions, via the Mel18 subunit for example 37 , or through ncRNAs recruitment, such as the interaction between CBX7 and ANRIL. 7,42 Thus, both PRC2 and PRC1 can mediate repression independently of each other.…”

“…The primary antibodies were: H2AK119ub (Cell Signaling D27C4, rabbit monoclonal; characterized in 13 , YAF-2 (ThermoFisher Scientific PA5-30359, rabbit polyclonal, http:// www.thermofisher.com/YAF2-Antibody-Polyclonal/PA5-30359. html), RYBP (Abcam ab5976, rabbit polyclonal, characterized in 37 , L3MBTL1 and L3MBTL2 (from D. Reinberg, rabbit polyclonal; characterized in 40 and 39 ). A dilution of 1/250 was used for all the primary antibodies.…”

“…When assembled with RYBP, non-canonical PRC1 mediates H2AK119ub without the need of pre-existing H3K27me3. 37 We first examined the localization of RING1 and YY1 Binding Protein (RYBP). Albeit weakly, in the oocyte RYBP was distributed both in the nucleus and the cytoplasm.…”

Advances in understanding chromosome silencing by the long non-coding RNA Xist

“…However, it has been shown that PRC1 complexes and associated H2AK119u1 are enriched on the inactive X in the absence of PRC2 and H3K27me3 [55]. In a recent study, H3K27me3 independent recruitment of PRC1 has been shown to be attributable to distinct PRC1 like complexes, RYBP-PRC1, in which the protein RYBP is included [56]. Both CBX family proteins and another core PRC1 subunit, MPH1/2/3, are excluded from RYBP-PRC1.…”