SET1/MLL complexes control transcription independently of H3K4me3

Histone H3 lysine 4 trimethylation (H3K4me3) at gene promoters is thought to play a central role in gene transcription. H3K4 methylation is deposited by the SET1 (A/B) and MLL (1-4) multi-protein complexes, but discovering how these essential enzymes shape H3K4me3 has been extremely challenging due to their multiplicity. This has also made determining whether SET1/MLL complexes control transcription through H3K4me3, or non-catalytic activities, an impenetrable problem. Here, we overcome these challenges through leveraging genome-engineering and combinatorial SET1/MLL protein depletion, integrated with genomics, proteomics, and live-cell transcription imaging. We uncover a new SET1B complex and reveal that SET1 and MLL1/2 complexes synergise to define H3K4me3 at gene regulatory elements. Unexpectedly, by decoupling SET1/MLL complex occupancy at promoters from H3K4me3, we discover they primarily control transcription independently of H3K4me3 through counteracting promoter-proximal termination and supporting transcription burst size. These discoveries reveal a new H3K4me3-independent logic for SET1/MLL-dependent control of gene transcription.