Quantitative Dissection and Stoichiometry Determination of the Human SET1/MLL Histone Methyltransferase Complexes

Abstract

Methylation of lysine 4 on histone H3 (H3K4) at promoters is tightly linked to transcriptional regulation in human cells. At least six different COMPASS-like multisubunit (SET1/MLL) complexes that contain methyltransferase activity for H3K4 have been described, but a comprehensive and quantitative analysis of these SET1/MLL complexes is lacking. We applied label-free quantitative mass spectrometry to determine the subunit composition and stoichiometry of the human SET1/MLL complexes. We identified both known and novel, unique and shared interactors and determined their distribution and stoichiometry over the different SET1/MLL complexes. In addition to being a core COMPASS subunit, the Dpy30 protein is a genuine subunit of the NURF chromatin remodeling complex. Furthermore, we identified the Bod1 protein as a discriminator between the SET1B and SET1A complexes, and we show that the H3K36me-interactor Psip1 preferentially binds to the MLL2 complex. Finally, absolute protein quantification in crude lysates mirrors many of the observed SET1/MLL complex stoichiometries. Our findings provide a molecular framework for understanding the diversity and abundance of the different SET1/MLL complexes, which together establish the H3K4 methylation landscape in human cells.

The H3K4 methyltransferase Setd1a is first required at the epiblast stage, whereas Setd1b becomes essential after gastrulation

“…Possibly, the discrepancy with the Wdr5 knockdown phenotype is due to the presence of Wdr5 in several other protein complexes [e.g. ATAC (Nagy and Tora, 2007;van Nuland et al, 2013)]. Embryos homozygous for an Ash2l hypomorphic allele showed a more severe phenotype than that of the Setd1a knockout, consistent with the inclusion of Ash2l in other H3K4 methyltransferase complexes.…”

“…S2). Whether the two proteins are expressed in different patterns, or at very different levels as has been recently suggested (van Nuland et al, 2013), remains to be determined. At least in ESCs, overexpression of Setd1b cannot rescue the proliferation defect caused by loss of Setd1a.…”

“…2A), a recent proteomic analysis of ESCs suggests that Setd1a is much more highly expressed than Setd1b (van Nuland et al, 2013). Possibly, therefore, the proliferation defects caused by loss of Setd1a can be compensated by overexpressing Setd1b.…”

Multiple cellular proteins interact with LEDGF/p75 through a conserved unstructured consensus motif

“…In addition to MLL1, JPO2 and PogZ, we found the motif in the known LEDGF/p75 interaction partners MLL2 (ref. 41) and ASK (DBF4) from the CDC7-ASK complex 32 . Moreover, IBM and IBM-like motifs were found in the mediator complex subunit 1 (Med1; residues 891-898) and IWS1 (residues 484-491; Fig.…”

Activin/Nodal signaling and NANOG orchestrate human embryonic stem cell fate decisions by controlling the H3K4me3 chromatin mark

“…This observation suggests that the effects of DPY30 knockdown are the results of impaired H3K4me3 activity of COMPASS complexes rather than their impaired recruitment onto the chromatin, in agreement with what was reported in previous studies (Jiang et al 2011;Ernst and Vakoc 2012;Yang et al 2014). Finally, it was recently reported that DPY30 can also be found in the NURF nucleosome remodeling complex (van Nuland et al 2013), and thus we decided to investigate the binding profile of the NURF catalytic subunit BPTF on Activin/Nodal target genes. In contrast to the COM-PASS complexes, BPTF enrichment on S/N/D-bound regions was limited and independent from the presence of Activin/Nodal signaling (Fig.…”