Figure 3

Quantification of focal adhesion localization. A) CHO-K1 cells were transfected with the empty EGFP vector or GFP N-terminally tagged versions of wild type or FAK mutants. Stably expressing populations were selected and lysed. Twenty-five μg of lysate was immunoblotted to examine FAK expression. Endogenous FAK and GFP-FAK are indicated. B) CHO-K1 cells stably expressing the EGFP vector alone, or GFP-FAK fusion proteins were plated on fibronectin coated coverslips overnight, then fixed and used for immunofluorescent staining with a GFP antibody. The cells were counterstained with a paxillin antibody to locate focal adhesions. GFP-positive cells were scored for GFP localization at focal adhesions. The data was analyzed using a one-way analysis of variance and the Dunnett post test (* denotes p < 0.01). C) CE cells transfected with GFP-FAK fusion proteins were plated on fibronectin-coated coverslips overnight, then fixed and used for immunofluorescent staining with a GFP antibody. The cells were also stained with a paxillin antibody to facilitate identification of focal adhesions. A second example of GFP-E997A localized to focal adhesion is also shown (lower right panel).