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.Posttranslational modifications of histone tails play an important role in chromatin structure and function (27). While the presence of several modifications correlates with gene activation and transcription, others have opposing repressive functions and are enriched in transcriptionally inactive or heterochromatic regions. A well-studied type of histone modification is methylation of lysine residues, which is conferred by histone methyltransferases (KMTs). There are three possible states of lysine methylation, namely, mono-, di-, and trimethylation. As expected for reversible marks involved in dynamic gene regulation, the methyl groups can be removed by histone demethylases (KDMs) of either the LSD1 or the Jumonji Cterminal-containing (JmjC) family of proteins (43,48).The JmjC family of histone demethylases consists of approximately 30 members in humans (9, 25). Plant homeodomain (PHD) finger-containing proteins 2 and 8 (PHF2 and PHF8, respectively) and KIAA1718 constitute a subgroup containing a single N-terminal PHD followed by the catalytic JmjC domain. Several protein domains (including the PHD; the MBT, WD40, and Tudor domains; and chromodomains) have been shown to bind to peptides containing either an unmodified or a methylated lysine residue, and some PHDs specifically interact with histone H3 trimethylated at lysine 4 (H3K4me3) (42,51,53). H3K4me3 is considered to be an activating chromatin mark because it is enriched at active RNA polymerase II (RNAPII) transcription start sites (TSSs) (4, 17). In mammalian cells, H3K4me3 is mainly established by the SET1/KMT2 family, which includes the mixed-lineage leukemia 1 to 4 (MLL1 to -4) proteins. These chromatin modifiers can be recruited to promoters upon gene activation (16, 52), and certain ...

The X-linked mental retardation gene PHF8 is a histone demethylase involved in neuronal differentiation

“…Based on in vitro demethylation assays Loenarz et al first reported that PHF8 catalyzed demethylation of H3K9me2/1, H3K27me2 and H3K36me2 [28]. Other studies show PHF8 as a predominant H3K9me2/1 demethylase with either minimal H3K27me2 and H3K36me2 demethylase activity [29] or H3K9me2/1 demethylase activity only [30][31][32][33]. In agreement with the latter studies, we showed that in all assays PHF8 exhibited histone demethylase activity with H3K9me2/1 specificity.…”

Systematic knockdown of epigenetic enzymes identifies a novel histone demethylase PHF8 overexpressed in prostate cancer with an impact on cell proliferation, migration and invasion

“…The observed ratio of few upregulated to many downregulated genes was in line with the suggested positive coactivator function of PHF8 (Fortschegger et al, 2010). Functional annotation of the 62 downregulated genes showed significant enrichment of the integrin signaling pathway.…”

“…A more detailed analysis of potential PHF8 target genes involved in PrCa cell migration may be an important area for future investigation, and to elucidate the functional mechanisms PHF8 is involved in. Interestingly, PHF8 was found to bind to the integrin beta-1 promoter, and linked to positive regulation of genes related to focal adhesion in HeLa cells (Fortschegger et al, 2010). PHF8 silencing also reduced proliferation and mildly inhibited G1-S phase transition, which is in line with a suggested role of PHF8 in cell cycle progression (Feng et al, 2010;Liu et al, 2010).…”

“…Inactivating mutations in the PHF8 JmjCdomain are linked to mental retardation syndromes with the cleft lip/palate (Laumonnier et al, 2005;Abidi et al, 2007;Koivisto et al, 2007;Qiao et al, 2008). This may be because of critical transcriptional co-activator functions of PHF8 in the regulation of neuronal and craniofacial development (Fortschegger et al, 2010;Kleine-Kohlbrecher et al, 2010;Qi et al, 2010). Furthermore, we showed that PHF8 mRNA is overexpressed in a large fraction of PrCa and correlated with features of tumor progression such as high Gleason grading.…”

Lactobacillus paracasei and Lactobacillus plantarum strains downregulate proinflammatory genes in an ex vivo system of cultured human colonic mucosa

“…Interestingly, LP strongly upregulated the gene expression of two chromatin modifying enzymes: EHMT2, a histonelysine N-methyltransferase, and PHF8, a histone demethylase, which were shown to act on histone 3 (Fortschegger et al 2010;Fritsch et al 2010). Upregulation of EHMT2 could be observed after treatment with all three strains compared to Inflamed tissue.…”