Highlights d A genetic, functional, and structural analysis of mammalian Mediator is provided d Contacts between a conserved core and the tail impact mMED-Pol II interaction d Loss of non-essential mMED subunits affects promoters linked to multiple enhancers d Cohesin is required to tether regulatory DNA; mMED and Pol II are not

Trimethylation of histone H3K36 is a chromatin mark associated with active gene expression, which has been implicated in coupling transcription with mRNA splicing and DNA damage response. SETD2 is a major H3K36 trimethyltransferase, which has been implicated as a tumor suppressor in mammals. Here, we report the regulation of SETD2 protein stability by the proteasome system, and the identification of SPOP, a key subunit of the CUL3 ubiquitin E3 ligase complex, as a SETD2-interacting protein. We demonstrate that SPOP is critically involved in SETD2 stability control and that the SPOP/CUL3 complex is responsible for SETD2 polyubiquitination both in vivo and in vitro. ChIP-Seq analysis and biochemical experiments demonstrate that modulation of SPOP expression confers differential H3K36me3 on SETD2 target genes, and induce H3K36me3-coupled alternative splicing events. Together, these findings establish a functional connection between oncogenic SPOP and tumor suppressive SETD2 in the dynamic regulation of gene expression on chromatin.

SummaryGenes of the mixed lineage leukemia (MLL) family regulate transcription by methylating histone H3K4. Six members of the MLL family exist in humans, including SETD1A, SETD1B and MLL1-MLL4. Each of them plays non-redundant roles in development and disease genesis. MLL1 regulates the cell cycle and the oscillation of circadian gene expression. Its fusion proteins are involved in leukemogenesis. Here, we studied the role of MLL1 in innate immunity and found it selectively regulates the activation of genes downstream of NF-kB mediated by tumor necrosis factor (TNFa) and lipopolysaccharide (LPS). Real-time PCR and genome-wide gene expression profile analysis proved that the deficiency of MLL1 reduced the expression of a group of genes downstream of nuclear factor kB (NF-kB). However, the activation of NF-kB itself was not affected. The MLL1 complex is found both in the nucleus and cytoplasm and is associated with NF-kB. CHIP assays proved that the translocation of MLL1 to chromatin was dependent on NF-kB. Our results suggest that MLL1 is recruited to its target genes by activated NF-kB and regulates their transcription.

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