Figure 3

PAK4 K473me decreases adhesion of the cells and modulates cell morphology. (A) Cell adhesion to fibronectin. MDA-MB-231 cells stably expressing empty plasmid, Flag PAK4 wild-type or Flag PAK4 K473R were serum starved over-night. Cells were then plated on a fibronectin pre-coated 96-well plate, followed by crystal violet staining. Crystal violet staining was solubilized and quantified. Error bars are s.d. Statistical analysis was performed for 5 experimental repeats using one-way ANOVA. ***p < 0.001. (B) Filopodia and cell actin structures staining. MDA-MB-231 cells stably expressing empty plasmid, Flag PAK4 wild-type or Flag PAK4 K473R were fixed and stained with Vybrant™ DiI Cell-Labeling Solution (on the left) or phalloidin (on the right) and Hoechst or DAPI. Cells were then visualized by confocal microscopy (63×). Scale bar 10 µm. Graph on the bottom shows percent cells with filopodia manually counted. Error bars are s.d. Statistical analysis was performed for n > 32 cells per condition using one-way ANOVA. ***p < 0.001.