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Variant PRC1 complex-dependent H2A ubiquitylation drives PRC2 recruitment and polycomb domain formation.

Blackledge NP, Farcas AM, Kondo T, King HW, McGouran JF, Hanssen LL, Ito S, Cooper S, Kondo K, Koseki Y, Ishikura T, Long HK, Sheahan TW, Brockdorff N, Kessler BM, Koseki H, Klose RJ - Cell (2014)

Bottom Line: Chromatin modifying activities inherent to polycomb repressive complexes PRC1 and PRC2 play an essential role in gene regulation, cellular differentiation, and development.Here, using a de novo targeting assay in mouse embryonic stem cells we unexpectedly discover that PRC1-dependent H2AK119ub1 leads to recruitment of PRC2 and H3K27me3 to effectively initiate a polycomb domain.This activity is restricted to variant PRC1 complexes, and genetic ablation experiments reveal that targeting of the variant PCGF1/PRC1 complex by KDM2B to CpG islands is required for normal polycomb domain formation and mouse development.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Chromatin Biology and Transcription, Department of Biochemistry, University of Oxford, Oxford, OX1 3QU, UK.

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PCGF 1, 3, and 5 Variant PRC1 Complexes Catalyze H2AK119ub1 and Create A Polycomb Domain Containing PRC2 and H3K27me3(A) A schematic illustrating the core components of canonical and variant PRC1 complexes.(B) The TetO array at its integration site on mouse chromosome 8.(C) Targeting of factors to the TetO via the TetR DNA-binding domain. Numbers represent qPCR primer positions (kb) with respect to TetO array.(D) ChIP analysis for fusion protein occupancy (TetR), RING1B, H2AK119ub1, and histone H3 across the TetO containing locus. Fusion protein identity is indicated above each panel.(E) As in (D) ChIP analysis for PRC2 components and H3K27me3. All ChIP experiments were performed at least in biological duplicate with error bars showing standard error of the mean (SEM).See also Figure S1.
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fig1: PCGF 1, 3, and 5 Variant PRC1 Complexes Catalyze H2AK119ub1 and Create A Polycomb Domain Containing PRC2 and H3K27me3(A) A schematic illustrating the core components of canonical and variant PRC1 complexes.(B) The TetO array at its integration site on mouse chromosome 8.(C) Targeting of factors to the TetO via the TetR DNA-binding domain. Numbers represent qPCR primer positions (kb) with respect to TetO array.(D) ChIP analysis for fusion protein occupancy (TetR), RING1B, H2AK119ub1, and histone H3 across the TetO containing locus. Fusion protein identity is indicated above each panel.(E) As in (D) ChIP analysis for PRC2 components and H3K27me3. All ChIP experiments were performed at least in biological duplicate with error bars showing standard error of the mean (SEM).See also Figure S1.

Mentions: Molecular and functional characterization of the polycomb repressive complexes has revealed that they do not function independently (Bracken et al., 2006; Ku et al., 2008; Papp and Müller, 2006; Schwartz et al., 2006). Instead, H3K27me3 placed by PRC2 is recognized by PRC1 complexes that contain chromobox (CBX) proteins (Cao et al., 2002; Min et al., 2003; Wang et al., 2004b). Based on these initial observations, the prevailing view over the past decade has been that PRC1 is recruited in a hierarchical manner to sites with pre-existing PRC2 activity and H3K27me3. However, it has recently emerged that CBX proteins are in direct competition with two additional factors, RYBP/YAF2, for a mutually exclusive binding site on RING1A/B (Wang et al., 2010). Significantly, H3K27me3-binding CBX proteins are limited to canonical PRC1 complexes containing either PCGF2 (MEL18) or PCGF4 (BMI1) and the Polyhomeotic proteins (PHC1/2/3) (Gao et al., 2012; Levine et al., 2002), while all PCGF proteins interact with RYBP/YAF2 to form variant PRC1 complexes lacking CBX proteins (Farcas et al., 2012; Gao et al., 2012; Gearhart et al., 2006; Lagarou et al., 2008; Sánchez et al., 2007; Tavares et al., 2012) (Figure 1A). The identification of variant PRC1 complexes and the observation that RING1B can occupy many of its target sites in the absence of H3K27me3 suggests that the hierarchical recruitment mechanism cannot explain all PRC1 complex targeting (Schoeftner et al., 2006; Tavares et al., 2012). Therefore, the central principles that underpin recognition of polycomb target sites in vivo and the molecular chain of events that leads to the formation of polycomb domains integrating both PRC1 and PRC2 activity remain unclear.


Variant PRC1 complex-dependent H2A ubiquitylation drives PRC2 recruitment and polycomb domain formation.

Blackledge NP, Farcas AM, Kondo T, King HW, McGouran JF, Hanssen LL, Ito S, Cooper S, Kondo K, Koseki Y, Ishikura T, Long HK, Sheahan TW, Brockdorff N, Kessler BM, Koseki H, Klose RJ - Cell (2014)

PCGF 1, 3, and 5 Variant PRC1 Complexes Catalyze H2AK119ub1 and Create A Polycomb Domain Containing PRC2 and H3K27me3(A) A schematic illustrating the core components of canonical and variant PRC1 complexes.(B) The TetO array at its integration site on mouse chromosome 8.(C) Targeting of factors to the TetO via the TetR DNA-binding domain. Numbers represent qPCR primer positions (kb) with respect to TetO array.(D) ChIP analysis for fusion protein occupancy (TetR), RING1B, H2AK119ub1, and histone H3 across the TetO containing locus. Fusion protein identity is indicated above each panel.(E) As in (D) ChIP analysis for PRC2 components and H3K27me3. All ChIP experiments were performed at least in biological duplicate with error bars showing standard error of the mean (SEM).See also Figure S1.
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Related In: Results  -  Collection

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fig1: PCGF 1, 3, and 5 Variant PRC1 Complexes Catalyze H2AK119ub1 and Create A Polycomb Domain Containing PRC2 and H3K27me3(A) A schematic illustrating the core components of canonical and variant PRC1 complexes.(B) The TetO array at its integration site on mouse chromosome 8.(C) Targeting of factors to the TetO via the TetR DNA-binding domain. Numbers represent qPCR primer positions (kb) with respect to TetO array.(D) ChIP analysis for fusion protein occupancy (TetR), RING1B, H2AK119ub1, and histone H3 across the TetO containing locus. Fusion protein identity is indicated above each panel.(E) As in (D) ChIP analysis for PRC2 components and H3K27me3. All ChIP experiments were performed at least in biological duplicate with error bars showing standard error of the mean (SEM).See also Figure S1.
Mentions: Molecular and functional characterization of the polycomb repressive complexes has revealed that they do not function independently (Bracken et al., 2006; Ku et al., 2008; Papp and Müller, 2006; Schwartz et al., 2006). Instead, H3K27me3 placed by PRC2 is recognized by PRC1 complexes that contain chromobox (CBX) proteins (Cao et al., 2002; Min et al., 2003; Wang et al., 2004b). Based on these initial observations, the prevailing view over the past decade has been that PRC1 is recruited in a hierarchical manner to sites with pre-existing PRC2 activity and H3K27me3. However, it has recently emerged that CBX proteins are in direct competition with two additional factors, RYBP/YAF2, for a mutually exclusive binding site on RING1A/B (Wang et al., 2010). Significantly, H3K27me3-binding CBX proteins are limited to canonical PRC1 complexes containing either PCGF2 (MEL18) or PCGF4 (BMI1) and the Polyhomeotic proteins (PHC1/2/3) (Gao et al., 2012; Levine et al., 2002), while all PCGF proteins interact with RYBP/YAF2 to form variant PRC1 complexes lacking CBX proteins (Farcas et al., 2012; Gao et al., 2012; Gearhart et al., 2006; Lagarou et al., 2008; Sánchez et al., 2007; Tavares et al., 2012) (Figure 1A). The identification of variant PRC1 complexes and the observation that RING1B can occupy many of its target sites in the absence of H3K27me3 suggests that the hierarchical recruitment mechanism cannot explain all PRC1 complex targeting (Schoeftner et al., 2006; Tavares et al., 2012). Therefore, the central principles that underpin recognition of polycomb target sites in vivo and the molecular chain of events that leads to the formation of polycomb domains integrating both PRC1 and PRC2 activity remain unclear.

Bottom Line: Chromatin modifying activities inherent to polycomb repressive complexes PRC1 and PRC2 play an essential role in gene regulation, cellular differentiation, and development.Here, using a de novo targeting assay in mouse embryonic stem cells we unexpectedly discover that PRC1-dependent H2AK119ub1 leads to recruitment of PRC2 and H3K27me3 to effectively initiate a polycomb domain.This activity is restricted to variant PRC1 complexes, and genetic ablation experiments reveal that targeting of the variant PCGF1/PRC1 complex by KDM2B to CpG islands is required for normal polycomb domain formation and mouse development.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Chromatin Biology and Transcription, Department of Biochemistry, University of Oxford, Oxford, OX1 3QU, UK.

Show MeSH