MLL Methyltransferases Are Activated in a Two-Step Process Free

The mixed lineage leukemia (MLL) proteins are histone lysine methyltransferases that contain a SET domain (MLL_SET) responsible for methylating histone H3 Lys4 (H3K4), and MLLs are mutated or involved in chromosomal translocations in multiple tumor types. MLL1 alone has limited methyltransferase activity, and is activated upon binding WD repeat domain 5 (WDR5), retinoblastoma binding protein 5 (RBBP5), and ash2 (absent, small, or homeotic)-like (ASH2L). However, the mechanism by which these proteins activate MLLs has not been well established, prompting Li and colleagues to determine the structural basis of MLL activation. RBBP5–ASH2L directly interacted with MLL proteins and activated their methyltransferase activity, but bound only weakly to MLL1, which required WDR5 as a bridging protein that bound simultaneously to MLL1 and RBBP5–ASH2L and promoted full MLL1 activation. MLL3_SET was crystallized in complex with the activation segment and the ASH2L-binding motif of RBBP5 and the splA and ryanodine receptor domain of ASH2L, termed the M3RA complex, both with and without the H3 peptide. The H3 binding residues were highly conserved, suggesting that all MLL proteins had the same specificity and recognition mechanism for H3K4. Mutations in residues responsible for electrostatic interactions at the MLL3–RBBP5–ASH2L interface reduced the association with MLL_SET domains and diminished MLL complex activity. Sequence differences between MLL3 and MLL1 indicated that MLL1 would have weaker binding to RBBP5–ASH2L. MLL_SET by itself was very flexible in solution, and RBBP5–ASH2L binding constrained the flexibility of the MLL_SET motif, allowing for cofactor binding, substrate recognition, and catalytic activation. In addition, the H3 peptide further stabilized the active MLL3_SET conformation. Taken together, these results elucidate a two-step activation mechanism of MLL proteins in which binding to RBBP5–ASH2L reduces the flexibility of MLL_SET, allowing a catalytically competent conformation, and H3 binding induces a MLL_SET conformational change to trigger its methyltransferase activity.

Li Y, Han J, Zhang Y, Cao F, Liu Z, Li S, et al. Structural basis for activity regulation of MLL family methyltransferases. Nature 2016 Feb 17 [Epub ahead of print].