Control of TCF-4 Expression by VDR and Vitamin D in the Mouse Mammary Gland and Colorectal Cancer Cell Lines (original) (raw)
Figure 4
Mouse TCF7L2 promoter reporter constructs respond to VDR or 1,25(OH)2D3.
(A) Two mouse TCF7L2 promoter constructs were cloned upstream of a luciferase reporter. The four putative VDRE locations relative to the translation start site are denoted, and all VDREs are encoded on the non-coding strand. The −177 and −187 VDREs are overlapping and share a half-site between them. The −1037 construct (-1037-luc) contains the region from −1 to −1037 relative to the translation start site and possesses the −177/−187 VDRE, only, while the −2068 reporter (-2068-luc) contains the region between −96 and −2068 relative to the translation start site of the mouse TCF7L2 promoter, and possesses all four putative VDREs. (B) CaCo2 cells were transfected for 24 hours with Renilla and reporters as indicated as well as different amounts of p53. Data are normalized to Renilla and to empty reporter expression. RLU = Relative Light Units. (C) -1038-Luc or empty vector constructs were transfected into CaCo2 cells with Renilla and different amounts of VDR, as indicated. 24 hours after transfection, cells were treated for an additional 24 hours with 10−7 M 1,25(OH)2D3 or EtOH vehicle control. Error bars represent SEM. Statistics were derived from two-way ANOVA for differences between EtOH- and 1,25(OH)2D3-treated samples: * p<.05; **; p<.005; *** p<.0005. (D) CaCo2 cells were transfected, treated and analyzed as in part C using -2068-luc instead of -1038-luc. Statistics were generated by two-way ANOVA for significant differences between the amount of VDR transfected: *** p<.0005. Error bars represent SEM. RLU: Relative Light Units.