Figure 2

AEG-1 activated β-catenin, which could reverse AEG-1-siRNA-mediated MET. (A) AEG-1 promoted β-catenin nuclear translocation. Slu-01 cells were transfected with pcDNA3.1-AEG-1. The subcellular localization of β-catenin was visualized through immunofluorescence (magnification × 400) and Western blotting. (B) Total β-catenin mRNA was detected by RT-PCR. (C) AEG-1 increased β-catenin/TCF transcriptional activity. NCI-H226 cells treated with AEG-1-siRNA, and Slu-01 cells treated with pcDNA3.1-AEG-1 were transfected with TCF-responsive promoter reporter (TOP-flash) or nonresponsive control reporter (FOP-flash); then, luciferase activity was measured as the ratio of TOP/FOP. Relative luciferase activity is presented as the mean ± SD. (error bars) from each sample after normalizing to the control. The asterisk indicates statistical significance (p < 0.01). (D) The morphology characteristics of NCI-H226 and Slu-01 cells were observed through phase-contrast microscopy (magnification × 200). (E) and (F) β-catenin overexpression reverses AEG-1-siRNA-mediated MET. An increasing amount of β-catenin was transfected in NCI-H226 (E) and Slu-01 (F) cells for 24 hours. Total cell lysates were probed with antibodies against E-cadherin, Vimentin, and β-catenin.