EpCAM-overexpression inhibits cell spreading and epithelial sheet migration.
(A) SDS extracts from MDCK cells Mhe16 and Mhe33 lines were immunoblotted for EpCAM and GAPDH. MDCK cells express endogenous EpCAM (ME) whereas MhE lines express both ME and untagged human EpCAM (HE). Graph shows quantification of EpCAM protein levels normalized to GAPDH protein levels in the same samples. For total EpCAM protein levels in Mhe16 and MhE33, intensities for MDCK-endogenous EpCAM (ME) and human EpCAM (HE) were combined. Error bars: S.E.M. of three samples for each cell line. Compared to parental MDCK cells, the EpCAM level was 11x higher in MhE16 and 10x higher in MhE33 cells. (B) Phase images (top) and quantification of area/cell (graph) of individual colonies of indicated cell lines. Scale bars: 20 μm. Area per cell was significantly decreased in MhE16 and MhE33 cells with ***p values < 0.0001 (two-tailed Mann-Whitney) compared to MDCK cells. Error bars: S.E.M. of 90 MDCK, 268 MhE16 and 284 MhE33 small colonies of 5 to 200 cells. (C) Phase images of indicated cell lines at 0 hrs migration (top row) and at 6 hrs migration (middle row). The bottom row shows an overlay of “6 hrs” images onto “0 hrs” images. Scale bars: 100 μm. Graph in (C) shows quantification of average change of area/image after 6 hrs migration for each cell line. Bars: S.E.M. of nine (MDCK), four (MhE16) or five (MhE33) experiments. MDCK values are the same as shown in the graph of Fig 1C. Stars correspond to p values derived from unpaired Student’s t-test. For MhE16 **p = 0.0063 and for MhE33 **p = 0.0074 compared to MDCK.