Figure 6.

Stable cell lines expressing Stx4 N-terminal peptide have reduced invadopodium formation and gelatin degradation. A, parental MDA-MB-231 cells and stable cell lines expressing GFP, GFP-Stx4-FL, or GFP–Stx4–N-term were seeded onto Alexa Fluor 594–labeled gelatin and incubated for 4 h prior to fixation, staining for F-actin, and analysis by confocal microscopy. Scale bars = 10 μm. B, quantification of invadopodium formation. Cells with F-actin puncta overlying dark spots of gelatin degradation were counted as cells forming invadopodia. Percentages of cells forming invadopodia are shown from three independent experiments in which 100 cells/sample were counted and normalized to parental MDA-MB-231 cells. C, quantification of gelatin degradation. Parental, GFP, GFP-Stx4-FL, and GFP–Stx4–N-term stable cells were plated onto fluorescent gelatin for 24 h and then fixed. Cells were analyzed for dark areas of degradation and scored as described under “Experimental Procedures.” Percentages of cells degrading gelatin are shown from three independent experiments in which 50 cells/sample were analyzed. All data are presented as percent of control ± S.D. Asterisks denote values significantly different from control (*, p < 0.05). All data represent three or more biological replicates with at least three technical replicates.