Figure 5.

Rac and cofilin activities are lowered in keratinocytes deficient in BPAG1e. (A) Levels of active Rac in subconfluent HEKs, HEKCn12s and HEKCn2s were determined by G-LISA through binding of GTP bound Rac1 to an affinity plate followed by antibody detection. Activity levels of Rac1 in HEKCn2s and HEKCn12s are plotted relative to that in HEKs and is significantly lower (significance relative to HEKs, * p < 0.05). Data are from three independent trials. (B) Total cell lysates were immunoblotted for total Rac1 and actin. (C) β4 integrin was immunoprecipitated from cell lysates of HEKs (left) and HEKCn2s (right), and precipitates were then immunoblotted for Rac1 (top panel) and β4 integrin (bottom panel). Blots were densitometrically scanned, the level of Rac immunoprecipitated was normalized to β4 integrin in three independent trials, and the results were plotted in the graph below relative to that in HEKs (* p < 0.012). (D) Cell lysates from HEKs, HEKCn2s, and HEKCn12s were prepared for immunoblotting using antibodies specific for Ser-3–phosphorylated cofilin or F-actin. Blots were densitometrically scanned, the levels were normalized to actin, and the fold increase of phosphorylated cofilin, relative to the level in HEKs, was quantified. The graph beneath the blot displays the mean ± SEM in three independent trials (significance relative to HEKs; * p < 0.05).