Fig. 5

Brg1 promotes metastasis in gastric cancer by driving an EMT transcriptional program. a IB analysis of WCLs derived from gastric cancer cell lines AGS. AGS cells were infected with the indicated pTRIPZ-Brg1 lentiviral vectors and were selected with 1 μg/ml puromycin for 72 h to eliminate non-infected cells. Afterward, 100, 300, 1000, 3000 ng/ml doxycycline (DOX) was added to the cells for another 24 h before harvesting for IB analysis. b Real-time PCR assays were performed in pTRIPZ-Brg1 AGS cells in the present or absence of DOX (1000 ng/ml) for 24 h. *p < 0.05, **p < 0.01, Student’s t test. c Representative images of the cell morphology in AGS cells. Scale bar, 100 μm. d IB analysis of WCLs derived from MKN45 cells infected with the indicated lentiviral shRNA plasmids. e Real-time PCR assays were performed in MKN45 cells infected with the indicated lentiviral shRNA plasmids.*p < 0.05, **p < 0.01, Student’s t test. f, g ChIP assays with anti-Brg1 antibody were performed in MKN45 cells infected with the indicated lentiviral shRNA plasmids. Mouse IgG was used as negative control. h, i ChIP assays with anti-Flag antibody were performed in AGS cells transfected with the indicated Flag-Brg1 constructs. Mouse IgG was used as antibody control in this experiment. j–l Representative images and statistical analysis of metastases that formed in lungs of each nude mouse (n = 5) at 2 months after tail vein injection of MKN45 cells (3 × 10⁶/each) transfected with the indicated lentiviral shRNA plasmids. One milligrams PFI3 (SIGMA, SML0939) dissolved in dimethyl sulfoxide plus 0.25 ml phosphate-buffered saline was given intraperitoneally 3 times per week. *p < 0.05, **p < 0.01, Student’s t test. Scale bar, 1 mm