APC is required for mammary tumor cell migration and mesenchymal morphology. (A) Wound-filling assays were performed by scratching confluent cultures of APC–knockdown and control 4T07 cells in the presence of 10 μg/ml mitomycin c and capturing phase-contrast images at 0, 12, 24, and 36 h post-wounding. Representative images from 0 and 36 h are shown; dashed white lines represent location of the wound. APC-knockdown cells were impaired in their ability to fill the wound at 36 h. 100X. (B) Quantification of the wound area at all time points shows that APC knockdown significantly decreased the migration of 4T07 cells at all time points. *p < 0.05 compared to pLKO.1 cells. (C) The morphology of APC-knockdown cells was quantified on phalloidin-stained cells grown on coverslips and categorized as having two protrusions (bipolar) or a rounded morphology. APC-knockdown cells have a higher percentage of cells with a rounded morphology compared to controls (p < 0.05 for APC-knockdown cells vs. pLKO.1 cells by Fisher’s exact test). (D) Representative images of the cells whose morphology was quantified in (C) and stained with phalloidin (red) and Hoechst (blue). Arrows illustrate cell-cell interactions, which are more prominent in APC-knockdown cells. 630X.