Figure 4

Inducible overexpression of EpCAM in EpCAM ^low human breast cancer cell lines with mesenchymal phenotype. (A) EpCAM^low cell lines MDA-MB-231 and Hs578t were transfected to express the tetracycline-repressor protein (TetR). A lentiviral construct for tetracycline (T)-inducible expression of EpCAM was generated and tumor cells stably integrating the construct were selected and named Hs578t^{TetR EpCAM} or MDA-MB-231^{TetR EpCAM}. (B) Stimulation with tetracycline for 24 h resulted in a strong induction of EpCAM gene expression as determined by real-time PCR. Mean±SEM of three independent experiments. (C) Moreover, upregulation of EpCAM was observed on the protein level as determined by Western Blot analysis. In addition to the unglycosylated protein, EpCAM was produced as a glycosylated and hyperglycosylated isoform in both cell lines analyzed. (D) Densitometric analysis of EpCAM protein expression of two independent experiments. All means are calculated in comparison to expression of MDA-MB 231 cells before induction with tetracycline.