Fig. 3

Effect of KMT2D LCD knockout on cell activities in HEK293T and PANC-1 cells. a Cell viability analysis of HEK293T and PANC-1 cells carrying vectors for the vehicle or KMT2D LCD knockout at 0, 24, 72 and 120 h post-transfection. Compared with the control group (vehicle), KMT2D LCD depletion significantly suppressed cell proliferation, while the OE1 cells (LCD1-depleted KMT2D cells re-expressing full-length KMT2D) and OE2 cells (LCD2-depleted KMT2D cells re-expressing full-length KMT2D) showed restored cell proliferation compared to the wild-type cells (error bars denote standard deviation). b Representative flow cytometry of cell apoptosis assays in PANC-1 cells. The apoptotic cell ratios were significantly increased in PANC-1 cells with KMT2D LCD deficiency. c Western blotting analyses of the apoptosis- and metastasis-related proteins after KMT2D LCD knockdown in HEK293T and PANC-1 cells. KMT2D LCD deficiency increased the Bax/Bcl2 ratio and the epithelial marker-to-mesenchymal marker ratio. Bcl2, 26 KDa; Bax, 20 KDa; EpCAM, 40 KDa; Vimentin, 57 KDa; GAPDH, 37 KDa. d RT-PCR assays of metastasis-associated markers. The gene expressions of epithelial markers (CDH1 and EpCAM) had markedly increased, while those of mesenchymal markers (CDH2 and vimentin) had decreased after KMT2D LCD depletion (error bars denote standard deviation). e Scratch wound-healing assays in PANC-1 cells. Knockout of KMT2D LCD significantly attenuated the cell migration ability. f Representative images of cell immunofluorescence determining the CDH2 and EpCAM expression in PANC-1 cells. After LCD knockout, CDH2 expression significantly reduced while that of EpCAM increased. All values are means ± SD. Data are representative of three independent experiments