Figure 3

WT1 represses E-cadherin proximal promoter. (A) Schematic diagram of the E-cadherin proximal promoter with 3 potential WT1 binding sites. (B) Effect of WT1 on E-cadherin promoter was tested in a dose response in PC3 cells. 250 ng of the E-cadherin proximal promoter reporter construct was cotransfected either with pCMV4 or GFP/WT1 expression construct at 0, 250, 500 and 750 ng concentrations. DNA concentrations were held constant by adding increasing amounts of pCMV4 and reporter activity analyzed as described in methods. (C and D) E-cadherin proximal promoter reporter construct was cotransfected either with pCMV4 or GFP/WT1 expression construct in PC3 (C) and DU145 (D) cells. Luciferase activity was measured and normalized to protein concentration. Data are reported relative to luciferase activity of pCMV4 transfected cells. Experiments were repeated three times in triplicate. Significance was determined by student’s t-test comparing GFP/WT1 transfected cells to pCMV4 transfected (*p ≤ 0.05, ***p ≤ 0.001).