Figure 3: TEAD4-S reduces cancer cell proliferation and EMT.

(a) RNA levels (upper panel) and protein levels (lower panel) of TEAD4-S in various NSCLC lines and normal bronchial cells were measured by semi-quantitative RT–PCR and western blot. The mean±s.d. for the percentage of TEAD4-S mRNA was plotted from three experiments. Each cancer cell line was compared with the two normal cell lines to calculate P values using Student’s t-test. (b) H157 or A549 cells stably expressing YAP, YAP/TEAD4-FL, YAP/TEAD4-S, YAP/RBM4 or vector control were analysed by colony formation assay. All experiments were performed in triplicates, with mean±s.d. of relative colony numbers plotted (P values from t-test). Images of the whole plate were shown. (c) H157 or A549 cells stably expressing YAP, YAP/TEAD4-FL, YAP/TEAD4-S, YAP/RBM4 or vector control were analysed by soft agar assay. All experiments were performed in triplicates, with mean±s.d. of relative colony numbers plotted (P values from t-test). (d) Western blot of epithelial and mesenchymal markers was performed using lysates from indicated H157 and A549 stable cells. (e) The subcellular localizations of TEAD4-FL or TEAD4-S with YAP. Cells were co-transfected with Flag-tagged YAP (red) and GFP-fused TEAD4 (green), and visualized with immunofluorescence assay. Scale bar, 25 μm. The localization of TEAD4-FL and TAD4-S was quantified in transfected cells, and the per cent of cells with nuclear or cytoplasmic TEAD4 were plotted (>50 cells were captured and quantified in both samples).