FIG. 3.

The c-Myc 3′ β-catenin/TCF4 binding region enhances SV40 promoter-driven luciferase gene expression. (A) Diagram of reporter constructs with the SV40 promoter represented as green rectangles, and the firefly luciferase gene is shown as yellow rectangles. The rectangles labeled c-Myc 5′ EN correspond to the 588 bp β-catenin/TCF enhancer (21). The rectangles labeled c-Myc 3′ EN correspond to a 615-bp fragment that begins 1,410 bp downstream from the c-Myc transcription stop site and binds β-catenin and TCF4 in ChIP assays. An “X” indicates that a TCF consensus motif was mutated. (B) The firefly luciferase reporter plasmids, a control plasmid expressing Renilla luciferase, and plasmids encoding β-catenin S45F and Lef-1 were transfected into HEK293 cells. After 24 h, firefly luciferase levels were assayed and normalized to Renilla luciferase levels. The activities of the respective reporters are aligned with their schematic representations. Activity is represented as relative change in the presence of β-catenin and Lef-1 and is normalized to background β-catenin/Lef-1 activation of the pGL3-promoter vector. (C) HCT116 cells were transfected with the indicated firefly luciferase reporters and a control Renilla luciferase plasmid. Luciferase activities were measured as in HEK293 cells, and data are represented as the change in expression relative to levels obtained with the pGL3-promoter vector alone. Error bars indicate standard deviations.