The histone H3-lysine-4 methyltransferase mixed-lineage leukemia 3 (MLL3) and its closest homolog, MLL4 (aka KMT2D), belong to two homologous transcriptional coactivator complexes, named MLL3 and MLL4 complexes, respectively. MLL3 plays crucial roles in multiple metabolic processes. However, the physiological roles of MLL4 in metabolism and the relationship between MLL3 and MLL4 in metabolic gene regulation are unclear. To address these issues, we analyzed the phenotypes of newly generated MLL4 mutant mice, along with MLL3 mutant and MLL3;MLL4 compound mutant mice. We also performed comparative genome-wide transcriptome analyses in livers of MLL3, MLL4, and MLL3;MLL4 mutant mice. These analyses revealed that MLL3 and MLL4 complexes are key epigenetic regulators of common metabolic processes and the hepatic circadian clock. Subsequent mechanistic analyses uncovered that MLL3/4 complexes function as pivotal coactivators of the circadian transcription factors (TFs), retinoid-related orphan receptor (ROR)-α and -γ, in the hepatic circadian clock. Consistent with disturbed hepatic clock gene expression in MLL4 mutant mice, we found that rhythmic fluctuation of hepatic and serum bile acid (BA) levels over the circadian cycle is abolished in MLL4 mutant mice. Our analyses also demonstrate that MLL4 primarily impinges on hepatic BA production among several regulatory pathways to control BA homeostasis. Together, our results provide strong in vivo support for important roles of both MLL3 and MLL4 in similar metabolic pathways. Conclusion: Both MLL3 and MLL4 complexes act as major epigenetic regulators of diverse metabolic processes (including circadian control of bile acid homeostasis) and as critical transcriptional coactivators of the circadian TFs, RORs.

During development, two cell types born from closely related progenitor pools often express identical transcriptional regulators despite their completely distinct characteristics. This phenomenon implies the need for a mechanism that operates to segregate the identities of the two cell types throughout differentiation after initial fate commitment. To understand this mechanism, we investigated the fate specification of spinal V2a interneurons, which share important developmental genes with motor neurons (MNs). We demonstrate that the paired homeodomain factor Chx10 functions as a critical determinant for V2a fate and is required to consolidate V2a identity in postmitotic neurons. Chx10 actively promotes V2a fate, downstream of the LIM-homeodomain factor Lhx3, while concomitantly suppressing the MN developmental program by preventing the MN-specific transcription complex from binding and activating MN genes. This dual activity enables Chx10 to effectively separate the V2a and MN pathways. Our study uncovers a widely applicable gene regulatory principle for segregating related cell fates.