PKCε mediates Rac activation and Rac-mediated responses induced by PMA.

<p>A. NSCLC cells were transfected with either PKCε RNAi (ε1 or ε2) or control (<i>CTRL</i>) RNAi and serum starved for 24 h. A representative Western blot for PKCε expression at the time of PMA treatment is shown (n = 3). B. Cells subject to PKCε or control RNAi were stimulated with PMA (100 nM, 2 min) and Rac-GTP levels determined using a pull-down assay.A representative experiment is shown. The fold-induction in Rac-GTP levels normalized to total Rac at time = 0, as determined by densitometry, is represented in the bottom graphs. Data are expressed as mean ± S.D. (n = 3). *, p<0.05; ***, p<0.001. C. A549 cells were treated with PMA (100 nM, 30 min), fixed, and stained with phalloidin-rhodamine. Representative micrographs are shown (n = 3). D. Quantification of A549 cells bearing ruffles, expressed as mean ± S.D. of 3 individual experiments. **, p<0.01; ***, p<0.001. E. Closure of wounds in response to PMA (100 nM, 30 min) was recorded at 9 h (A549 cells). Experiments were carried out in triplicate plates. Results are expressed as percentage of the closure induced by PMA, and expressed as mean ± S.E.M. (n = 3). ***, p<0.001.