MTCBP-1 overexpression inhibits the invasive properties of pancreatic cancer cells. (A–C) MTCBP-1 overexpression inhibits transwell invasion of PDAC cells. BxPC3 cells (A), Panc-1 cells (B), or DanG cells (C) stably expressing FLAG control or MTCBP-1-FLAG were seeded in transwell invasion assays for 6 (BxPC3), 24 (Panc-1), or 48 h (DanG) and counted by DAPI staining after fixation. A more than threefold reduction in the number of cells migrating through the filters was observed in MTCBP-1–expressing cells. The stable overexpression was confirmed by Western blot. Bars, 50 µm. (D) MTCBP-1 overexpression suppresses the ECM degradation of PDAC cells. DanG cells stably expressing FLAG control or MTCBP-1-FLAG were seeded on green fluorescent gelatin-coated coverslips and matrix degradation was quantified after 8 h. MTCBP-1 overexpression led to a reduction in the number of cells degrading matrix as well as the area of degradation per cell. Graphs represent normalized averages ± SEM from three independent experiments. Bars, 10 µm. (E and F) MTCBP-1 overexpression reduces invadopodia number in PDAC cells. DanG (E) or BxPC3 (F) cells stably expressing FLAG or MTCBP-1-FLAG were seeded on gelatin-coated coverslips and stained for Tks5 to mark invadopodia. MTCBP-1 overexpression reduces the number of Tks5-positive invadopodia in DanG and BxPC3 cells comparing to the controls. Graphs represent normalized averages ± SEM from three independent experiments. *, P < 0.05; **, P < 0.005; ***, P < 0.0005. Bars, 10 µm. All blot measurements in kD.