MLL3 ChIP sequencing in murine and human HCC cells

<p>Mutations in genes encoding components of chromatin modifying and remodeling complexes are among the most frequently observed somatic events in human cancers. For example, missense and nonsense mutations targeting the mixed lineage leukemia family member 3 (<em>MLL3</em>/<em>KMT2C</em>) histone methyltransferase occur in a range of solid tumors and heterozygous deletions encompassing <em>MLL3</em> occur in a subset of aggressive leukemias. Although <em>MLL3</em> loss can promote tumorigenesis in mice, the molecular targets and biological processes by which MLL3 suppresses tumorigenesis remain poorly characterized. Here we combined genetic, epigenomic, and animal modeling approaches to demonstrate that one of the mechanisms by which MLL3 links chromatin remodeling to tumor suppression is by co-activating the <em>Cdkn2a</em> tumor suppressor locus. Disruption of <em>Mll3</em> cooperates with <em>Myc</em> overexpression in the development of murine hepatocellular carcinoma (HCC), in which MLL3 binding to the <em>Cdkn2a</em> locus is blunted, resulting in reduced H3K4 methylation and low expression levels of the locus-encoded genes, <em>Ink4a</em> and <em>Arf</em>. Conversely, elevated <em>MLL3</em> expression increases its binding to the CDKN2A locus and co-activates gene transcription. Endogenous Mll3 restoration reverses these chromatin and transcriptional effects and triggers <em>Ink4a</em>/<em>Arf</em>-dependent apoptosis. Underscoring the human relevance of this epistasis, we found that genomic alterations in <em>MLL3</em> and <em>CDKN2A</em> display mutual exclusivity in human HCC samples. These results collectively point to a new mechanism for disrupting CDKN2A activity during cancer development and, in doing so, link MLL3 to an established tumor suppressor network.</p>