MLL4 prepares the enhancer landscape for Foxp3 induction via chromatin looping.

MLL4 is an essential subunit of the histone H3 Lys4 (H3K4)-methylation complexes. We found that MLL4 deficiency compromised the development of regulatory T cells (Treg cells) and resulted in a substantial decrease in monomethylated H3K4 (H3K4me1) and chromatin interaction at putative gene enhancers, a considerable portion of which were not direct targets of MLL4 but were enhancers that interacted with MLL4-bound sites. The decrease in H3K4me1 and chromatin interaction at the enhancers not bound by MLL4 correlated with MLL4 binding at distant interacting regions. Deletion of an upstream MLL4-binding site diminished the abundance of H3K4me1 at the regulatory elements of the gene encoding the transcription factor Foxp3 that were looped to the MLL4-binding site and compromised both the thymic differentiation and the inducible differentiation of Treg cells. We found that MLL4 catalyzed methylation of H3K4 at distant unbound enhancers via chromatin looping, which identifies a previously unknown mechanism for regulating the T cell enhancer landscape and affecting Treg cell differentiation.