PHF8 Targets Histone Methylation and RNA Polymerase II To Activate Transcription

Abstract

Mutations in PHF8 are associated with X-linked mental retardation and cleft lip/cleft palate. PHF8 contains a plant homeodomain (PHD) in its N terminus and is a member of a family of JmjC domain-containing proteins. While PHDs can act as methyl lysine recognition motifs, JmjC domains can catalyze lysine demethylation. Here, we show that PHF8 is a histone demethylase that removes repressive histone H3 dimethyl lysine 9 marks. Our biochemical analysis revealed specific association of the PHF8 PHD with histone H3 trimethylated at lysine 4 (H3K4me3). Chromatin immunoprecipitation followed by high-throughput sequencing indicated that PHF8 is enriched at the transcription start sites of many active or poised genes, mirroring the presence of RNA polymerase II (RNAPII) and of H3K4me3-bearing nucleosomes. We show that PHF8 can act as a transcriptional coactivator and that its activation function largely depends on binding of the PHD to H3K4me3. Furthermore, we present evidence for direct interaction of PHF8 with the C-terminal domain of RNAPII. Importantly, a PHF8 disease mutant was defective in demethylation and in coactivation. This is the first demonstration of a chromatin-modifying enzyme that is globally recruited to promoters through its association with H3K4me3 and RNAPII.

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Supplemental material for this article can be found at http://mcb.asm.org/.

We thank Kristian Helin, Min Gyu Lee, and Clément Carré for providing reagents and the Ultrasequencing and Microarrays Units at the CRG for performing Solexa and microarray analyses. We are particularly grateful to Marc Vigneron (ESBS, Strasbourg, France) for providing unpublished RNAPII antibodies and the GST-CTD construct. We thank Harm-Jan Vos for help in peptide synthesis, Hetty van Teeffelen for technical assistance, and Michiel Vermeulen for sharing unpublished results.

K.F. received an Erwin-Schrödinger fellowship from the Austrian Science Fund FWF (J2728-B12). P.D.G., N.S.O., and H.T.M.T. were supported by grants from the Netherlands Organization for Scientific Research (NWO-CW TOP 700.57.302), the European Union (EUTRACC LSHG-CT-2006-037445), and the Netherlands Proteomics Center. R.S. was supported by a grant from the NIH (CA090758).

We declare that we have no competing financial interest.