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. 2016 Sep 27;7(43):70881–70897. doi: 10.18632/oncotarget.12282

Figure 3. Depletion of PAK protein expression reduces invadopodia formation.

Figure 3

A-B. Transient depletion of PAK1 (A) and PAK4 (B) expression in the WM-115 and A-375M2 cell lines at 4 days post siRNA transfection. A double knockdown was performed using PAK1 Oligo 1 and PAK4 Oligo 2 oligonucleotides. Control cells were transfected with a control non targeting siRNA. Significance was calculated for protein depleted cell lines compared to wildtype. over 3 independent experiments; *= P 00< 0.05. Densitometric data were normalized to GAPDH, which was used as a loading control. C. Representative images of the 3D spheroid invasion assay Examples of invading cells are indicated by black arrows. Scale bar = 100μm. D. Quantification of the number of cells that had invaded surrounding matrix E. the number of cells that invaded further than 100μm from the spheroid mass. Significance was calculated to wildtype. 9 spheroids over 3 independent experiments; * = P 00< 0.05. F. Representative invadopodia assay images of WM-115 cells in which PAK1 and PAK4 proteins are depleted. The degradation was measured using ImageJ software. Scale bars = 10μm. The percentage of cells with invadopodia G. and the area of degradation from invadopodia per cell H.. Significance was calculated to wildtype cells. 150 (Percentage) or 90 (Degradation) cells, over 3 independent experiments; * = P 00< 0.05. In all cases data are the mean values ± S.E.M.