Abstract
Many studies have revealed pathways of epigenetic gene silencing by Polycomb repressive complex 2 (PRC2) in vivo, but understanding the underlying molecular mechanisms requires biochemistry. Here we analyze interactions of reconstituted human PRC2 with nucleosome complexes. Histone modifications, the H3K27M cancer mutation, and inclusion of JARID2 or EZH1 in the PRC2 complex have unexpectedly minor effects on PRC2–nucleosome binding. Instead, protein-free linker DNA dominates the PRC2–nucleosome interaction. Specificity for CG-rich sequences is consistent with PRC2 occupying CG-rich DNA in vivo. PRC2 preferentially binds methylated DNA regulated by its AEBP2 subunit, suggesting how DNA and histone methylation collaborate to repress chromatin. We find that RNA, known to inhibit PRC2 activity, is not a methyltransferase inhibitor per se. Instead, RNA sequesters PRC2 from nucleosome substrates, because PRC2 binding requires linker DNA, and RNA and DNA binding are mutually exclusive. Together, we provide a model for PRC2 recruitment and an explanation for how actively transcribed genomic regions bind PRC2 but escape silencing.
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Acknowledgements
We thank C. J. Lim and other members of the Cech lab for useful conversations. We thank C. Davidovich (Monash University, Clayton, Australia) for initial trials of expression and purification of JARID2 and for stimulating discussion. We thank K. Luger (University of Colorado Boulder) and members of her laboratory—especially U. M. Muthurajan and P. Dyer—for providing plasmids and helpful discussion of reconstituting nucleosomes. We thank J. Müller and his group (MPI Martiensreid, Germany) and Beat Fierz (EPFL, Lausanne, Switzerland) for sharing unpublished data and discussions. T.W.M. is supported by National Institutes of Health grants R37-GM086868 and PO1-CA196539. T.R.C. is supported by the Howard Hughes Medical Institute.
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X.W., R.D.P., and T.R.C. designed the experiments. X.W. and R.D.P. carried out experiments. A.R.G. carried out protein purification. Z.Z.B., E.J.G. and T.W.M. carried out the synthesis of modified histone H3. X.W., R.D.P., and T.R.C. wrote the manuscript.
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T.R.C. is on the board of directors of Merck, Inc., which provides no funding for his research.
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Wang, X., Paucek, R., Gooding, A. et al. Molecular analysis of PRC2 recruitment to DNA in chromatin and its inhibition by RNA. Nat Struct Mol Biol 24, 1028–1038 (2017). https://doi.org/10.1038/nsmb.3487
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DOI: https://doi.org/10.1038/nsmb.3487
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