Figure 2.

aPKCζ is required for proper cell adhesion. (A) aPKCζ^−/− and control cells were subjected to wound scratch assay and immunostained with anti-paxillin antibody. Bars are 20 µm. (B) Quantification of the amount of pY118-paxillin. Lysates from aPKCζ^−/− and control cells were subjected to Western blot analysis using anti-paxillin and anti-pY118-paxillin antibodies. The amounts of paxillin were normalized relative to actin. Bands were analyzed by densitometry using ImageJ. Paxillin phosphorylation was calculated as the ratio of phosphorylated paxillin to total paxillin. Values represent the mean ± SEM for 4 independent experiments, * p < 0.05, values are aPKCζ^−/− cells compared with control cells. Pax, paxillin and pY118-pax, pY118-paxillin. (C) aPKCζ^−/− and control cells were seeded on dishes, and after 2.5 h phase-contrast images of the cells were taken by confocal microscopy. Arrows indicate rounded detached cells and arrowheads indicate adhered cells. Bars are 50 µm. (D) Quantification of adhered cells. Adhered and rounded detached cells in 15 randomly-chosen fields were counted and the percentage of adhered cells relative to the total number of cells in each field was calculated. Values represent the mean ± SEM for 4 independent experiments subjected to 2-tailed, 2-sampled unequal variance Student's t test, *** p < 0.001, values are aPKCζ^−/− cells compared with control cells.