Identification of De Novo Enhancers Activated by TGFβ to Drive Expression of CDKN2A and B in HeLa Cells

Disruption of the ( ) and ( ) genes, which encode three function-independent tumor suppressors, is one of the most common events in human cancer. Because their relative importance in tumor prevention appears to be species- and context-specific, studying their regulation can shed light on mechanisms by which they are bypassed in malignant transformation. We previously unveiled a new pathway in which TGFβ selectively induces at mouse in eye development and cultured fibroblasts. As TGFβ signaling is often derailed in cancer development or progression, we investigated its control of in human cancer. Computational analyses of sequencing and array data from nearly 11,000 patients with cancer in TCGA showed discordant expression of and in most cancer subtypes, with gene copy-number loss and promoter methylation involved in only a subset. Using HeLa cells as a model, we found that exogenous TGFβ induced mRNA and protein, and knockdown limited TGFβ-mediated growth suppression. TGFβ-mediated mRNA induction required SMAD2/3, p38MAPK, and SP1, and mRNA was induced without added RNAPII recruitment. Chromatin immunoprecipitation unveiled a remote enhancer element engaged by TGFβ by a mechanism that partially depended on p38MAPK. CRISPR-based editing of this enhancer limited induction of and by TGFβ, but not by oncogenic RAS. IMPLICATIONS: Our findings reveal new molecular mechanisms by which regulation is coupled to external cues, and those findings represent entry points to further explore pharmacologic strategies to restore their expression in cancer.