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PRIME Report Summary

Project ID: 340081
Funded under: FP7-IDEAS-ERC
Country: United Kingdom

Mid-Term Report Summary - PRIME (Polycomb repressor interactions in relation to the mammalian epigenome.)

Polycomb repressor complexes are highly conserved chromatin modifying machines that play a central role in regulating gene expression through development. The targeting of these complexes to specific genes has generally been attributed to sequence specific DNA binding factors and/or non-coding RNAs. However, recent evidence has suggested that targeting may also involve Polycomb complexes reading chromatin states, referring to the combination of modifications (for example methylation or ubiquitylation) of histone proteins that form the nucleosome around which DNA is wrapped. More specifically, histone modifications may either stimulate or inhibit the target gene binding/activity of Polycomb complexes. PRIME (Polycomb Repressor Interactions with the Mammalian Epigenome) set out to explore these interactions in relation to the major Polycomb complexes, PRC1 and PRC2.
We, together with collaborators in Oxford and Japan, made key breakthroughs at the time the PRIME project was initiated, demonstrating that unmethylated DNA (CpG dinucleotide), which is a characteristic of many gene promoters, is a critical determinant of both PRC1 and PRC2 recruitment. Additionally we made the unexpected discovery that histone H2A ubiquitylation (H2AK119ub1), catalysed by PRC1, can be recognised by PRC2. The latter finding, the converse of the known interaction of PRC1 with PRC2 catalysed histone H3 lysine 27 methylation (H3K27me3), provides a rational explanation for Polycomb occupancy at CpG islands of target genes, based on initial targeting of PRC1 complexes via the PRC1 subunit KDM2B, which recognises unmethylated CpG. We have proposed that this pathway, together with antagonism of Polycomb activity conferred by histone modifications linked to gene activity, restrict Polycomb recruitment to CpG island promoters of genes that are not expressed in a given cell type. Key questions that arise from this hypothesis are; can Polycomb recruitment be initiated by PRC1? Do active gene linked histone modifications antagonise Polycomb binding/activity, and what is the mechanism by which PRC2 recognises H2AK119ub1, catalysed by PRC1. Our work within the PRIME project has gone a considerable way towards answering two of these questions. We have now determined unequivocally that PRC1 recruitment precedes that of PRC2 at a well studied model target, the inactive X chromosome (manuscript in preparation). Additionally we have analysed a PRC2 sub-complex which includes the cofactors Aebp2 and Jarid2, which has been implicated in the recognition of H2AK119ub1. We found that Aebp2 functions as an anti-Polycomb factor (Grijzenhout et al, Development 143, p2716, 2016), and as such is unlikely to mediate H2AK119ub1 recognition. Moreover, in a separate study we demonstrated that Jarid2 but not Aebp2 is required for recognition of H2AK119ub1 by PRC2 (manuscript in revision).
Our focus in the time remaining will be to continue analysis of the interaction of both PRC1 and PRC2 with nucleosomes at the atomic scale (Aim 3), with the goal of predicting other potential stimulatory chromatin modifications and/or modifications that antagonise Polycomb activity. Additionally we will use in vitro assays to determine how Polycomb catalysed histone modifications affect the activity of Trithorax complexes that catalyse histone modifications linked to gene activation (Aim 2). Finally, we will further analyse the relationship of chromatin state and Polycomb activity in vivo, using both the X chromosome inactivation and pericentric heterochromatin models.

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United Kingdom
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