Fig. 4: Ets-1 regulates epithelial–mesenchymal transition (EMT) in soraR cells.

A Huh7 naive and B Hep3B naive cells stably overexpressing Ets1-WT (Huh7-Ets-1 WT and Hep3B-Ets-1 WT respectively) were treated with (+) or without (−) 1 µg/ml Doxycycline (DOX) to induce ectopic Ets-1 expression in low serum (1% FBS) media. The cells were harvested at various time points and analyzed by western blots with the indicated antibodies. C Subconfluent Hep3B naive cells were transiently transfected with pLIX-403-Ets-1 and treated without DOX (lane 2) or with DOX (lane 3) for 48 h followed by RNA extraction and qPCR analysis of the indicated EMT genes. UN- indicates untransfected control and OE- indicates overexpression. D Hep3B-soraR cells transiently transfected with control- or Ets-1-siRNA for 48 and 72 h were analyzed by western blots. E Hep3B-soraR cells transfected as in D for 72 h were analyzed by qPCR for EMT genes. The data in C, E represent the mean ± S.D. of three independent PCR reactions. F Hep3B-soraR cells transfected with control- or Ets-1-siRNA were subjected to transwell migration assay for 48 h as in Fig. 1E. Scale bar, 100 µm. For quantitation, cells were counted in four different fields and plotted as bar graphs (G). The data represent the mean ± S.D. of two independent experiments. Significant differences were determined by t test and indicated as: *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001.