A phosphorylation switch on RbBP5 regulates histone H3 Lys4 methylation

Abstract

The methyltransferase activity of the trithorax group (TrxG) protein MLL1 found within its COMPASS (complex associated with SET1)-like complex is allosterically regulated by a four-subunit complex composed of WDR5, RbBP5, Ash2L, and DPY30 (also referred to as WRAD). We report structural evidence showing that in WRAD, a concave surface of the Ash2L SPIa and ryanodine receptor (SPRY) domain binds to a cluster of acidic residues, referred to as the D/E box, in RbBP5. Mutational analysis shows that residues forming the Ash2L/RbBP5 interface are important for heterodimer formation, stimulation of MLL1 catalytic activity, and erythroid cell terminal differentiation. We also demonstrate that a phosphorylation switch on RbBP5 stimulates WRAD complex formation and significantly increases KMT2 (lysine [K] methyltransferase 2) enzyme methylation rates. Overall, our findings provide structural insights into the assembly of the WRAD complex and point to a novel regulatory mechanism controlling the activity of the KMT2/COMPASS family of lysine methyltransferases.Supplemental material is available for this article.Received October 27, 2014; revised version accepted December 15, 2014. The methyltransferase activity of the trithorax group (TrxG) protein MLL1 as well as the other members of the KMT2 (lysine [K] methyltransferase 2) family found within COMPASS (complex associated with SET1) catalyzes the site-specific methylation of the e-amine of Lys4 (K4) of histone H3 (Shilatifard 2012). While these enzymes share the ability to methylate the same residue on histone H3, the catalytic activity of these enzymes is linked to different biological processes. MLL1/MLL2 di/trimethylate H3K4 (H3K4me2/3) and regulate Hox gene expression during embryonic development (Yu et al. 1995;Dou et al. 2006). MLL3/MLL4 regulate adipogenesis ) and primarily monomethylate H3K4 (H3K4me1) at both enhancer (Herz et al. 2012;Hu et al. 2013) and promoter (Cheng et al. 2014) regions, while SET1A/B are the primary H3K4 trimethyltransferases (Wu et al. 2008). However, despite divergence in catalytic activity and functional roles, enzymes of the KMT2/COMPASS family must assemble into multisubunit complexes to carry out their biological functions.Our molecular understanding of the protein complexes involved in H3K4 methylation stems from the isolation of COMPASS from Saccharomyces cerevisiae (Miller et al. 2001;Roguev et al. 2001;Krogan et al. 2002;Dehe et al. 2006). These studies demonstrated that regulatory subunits found within COMPASS and mammalian COMPASS-like complexes play key roles in stabilizing the enzyme and stimulating its methyltransferase activity as well as targeting the protein complex to specific genomic loci (Couture and Skiniotis 2013). While each of these multisubunit protein complexes contains unique subunits, each member of the KMT2 family associates with a common set of four evolutionarily conserved regulatory proteins; namely, WDR5, RbBP5, Ash2L, and DPY30 (WRAD) (Couture and Skiniotis 2013). The foursubunit complex directly binds ...

Structural basis for activity regulation of MLL family methyltransferases

“…Consistent with previous studies 23,24 , the ASH2L C-terminal SPRY (splA and ryanodine receptor) domain is sufficient to form a heterodimer with RBBP5 to stimulate the HKMT activity of MLL proteins (Fig. 1d, compare lanes 1 and 5).…”

The Set1 complex is dimeric and acts with Jhd2 demethylation to convey symmetrical H3K4 trimethylation

“…However Bre2 may still contribute in vivo in the absence of Sdc1 through a loose association that was lost upon our biochemical fractionation. Relevant to this possibility, the human homologue of Bre2, ASH2L, not only associates with DPY30 but also with the human homologue of Swd1, RbBP5 (Avdic et al 2011;Zhang et al 2015b).…”

Targeting DOT1L and HOX gene expression in MLL-rearranged leukemia and beyond

“…The most frequently observed breakpoints in MLL1 locate to the region between the CxxC domain and PHD domains. ( B ) The subunits of the MLL1-containing complex of proteins associated with Set1 (MLL1-COMPASS) include multiple endocrine neoplasia I (MENIN), host cell factor C1 (HCF1), WD-repeat containing protein 5 (WDR5), Rb-binding protein 5 (RBBP5), absent, small, or homeotic 2-like (ASH2L), and Dpy-30-Like Protein (hDPY30) [53, 54, 57]. ( C ) Structure of MLL1-fusion proteins caused by MLL1 gene translocations.…”