Abstract
Changes in phosphorylation of the carboxy-terminal domain (CTD) of RNA polymerase II (RNAP) are associated with transcription initiation, elongation and termination1,2,3. Sites of active transcription are generally characterized by hyperphosphorylated RNAP, particularly at Ser 2 residues, whereas inactive or poised genes may lack RNAP or may bind Ser 5-phosphorylated RNAP at promoter proximal regions. Recent studies have demonstrated that silent developmental regulator genes have an unusual histone modification profile in ES cells, being simultaneously marked with Polycomb repressor-mediated histone H3K27 methylation, and marks normally associated with gene activity4,5. Contrary to the prevailing view, we show here that this important subset of developmental regulator genes, termed bivalent genes, assemble RNAP complexes phosphorylated on Ser 5 and are transcribed at low levels. We provide evidence that this poised RNAP configuration is enforced by Polycomb Repressor Complex (PRC)-mediated ubiquitination of H2A, as conditional deletion of Ring1A and Ring1B leads to the sequential loss of ubiquitination of H2A, release of poised RNAP, and subsequent gene de-repression. These observations provide an insight into the molecular mechanisms that allow ES cells to self-renew and yet retain the ability to generate multiple lineage outcomes.
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Acknowledgements
We thank Henrietta Szutorisz, Niall Dillon, Miguel R. Branco (CSC) and Stephen Buratowski (Harvard Medical School, Cambridge, MA) for advice establishing the RNAP ChIP, James Briscoe (MRC-NIMR) and Joana Santos (CSC) for help in the design of Nkx2.2 gene analyses and TS cell culture, Austin Smith (Wellcome Trust Centre for Stem Cell Research, Cambridge, UK) for ES-OS25 and ES-ZHBTc4 cells, Janet Rossant (Hospital for Sick Children, Toronto, Canada) for XEN cells, Sanofi-Aventis (Bethesda, MD) for the kind gift of flavopiridol, Francisco Ramirez (London, UK) for the statistical analyses, Zoe Webster (CSC) for assistance with ES cell culture, Matthias Merkenschlager and Sarah Elderkin (CSC) for advice and the Medical Research Council (UK), Genome Network Project and NoE Epigenome for support.
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M.C. and E.B contributed equally to this work.
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Supplementary Figures S1, S2, S3, S4, S5, S6 S7, Supplementry Tables and Methods (PDF 706 kb)
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Stock, J., Giadrossi, S., Casanova, M. et al. Ring1-mediated ubiquitination of H2A restrains poised RNA polymerase II at bivalent genes in mouse ES cells. Nat Cell Biol 9, 1428–1435 (2007). https://doi.org/10.1038/ncb1663
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DOI: https://doi.org/10.1038/ncb1663
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