FIGURE 7.

Fyn and Pyk2 compete for STEP binding. A, constant amounts (1 μg of cDNA) of full-length STEP and Pyk2 constructs were co-transfected with increasing amounts of Fyn construct into HEK293 cells. Thirty-six hours after transfection, cells were lysed before immunoprecipitation (IP) with anti-STEP antibody and immunoblotting (IB) with anti-Pyk2 antibody. Immunoblotting of lysates indicates increasing amounts of Fyn with increasing amounts of Fyn cDNA but unchanged amounts of Pyk2. Immunoblotting of STEP immunoprecipitates indicates unaltered amounts of STEP in all samples. B, constant amounts (1 μg of cDNA) of STEP and Fyn were co-transfected with increasing amounts of Pyk2. Co-immunoprecipitation of Fyn with STEP was measured by immunoblotting with anti-Fyn antibody. C, competitive binding was tested in vitro with all purified proteins. GST-tagged STEP was adsorbed to glutathione beads and incubated with a constant amount of Pyk2 (500 ng) and increasing amounts of Fyn as indicated.