A Genome-Wide Screen for β-Catenin Binding Sites Identifies a Downstream Enhancer Element That Controls c-Myc Gene Expression

Abstract

Mutations in components of the Wnt signaling pathway initiate colorectal carcinogenesis by deregulating the β-catenin transcriptional coactivator. β-Catenin activation of one target in particular, the c-Myc proto-oncogene, is required for colon cancer pathogenesis. β-Catenin is known to regulate c-Myc expression via sequences upstream of the transcription start site. Here, we report that a more robust β-catenin binding region localizes 1.4 kb downstream from the c-Myc transcriptional stop site. This site was discovered using a genome-wide method for identifying transcription factor binding sites termed serial analysis of chromatin occupancy. Chromatin immunoprecipitation-scanning assays demonstrate that the 5′ enhancer and the 3′ binding element are the only β-catenin and TCF4 binding regions across the c-Myc locus. When placed downstream of a simian virus 40-driven promoter-luciferase construct, the 3′ element activated luciferase transcription when introduced into HCT116 cells. c-Myc transcription is negligible in quiescent HCT116 cells but is induced when cells reenter the cell cycle after the addition of mitogens. Using these cells, we found that β-catenin and TCF4 occupancy at the 3′ enhancer precede occupancy at the 5′ enhancer. Association of c-Jun, β-catenin, and TCF4 specifically with the downstream enhancer underlies mitogen stimulation of c-Myc transcription. Our findings indicate that a downstream enhancer element provides the principal regulation of c-Myc expression.