FIG. 5.

Coimmunoprecipitation of Gab1 with wild-type but not mutant Shp-2. (A) Anti-Gab1 antiserum (2 μl) was added to 1 ml (1 mg of total protein) of cell lysates from Shp-2^+/+ or Shp-2^−/− cells stimulated by EGF at the indicated times and immunoprecipitated (IP). The precipitates were immunoblotted (IB) with an anti-Shp-2 antibody that recognizes both wild-type and Shp-2^Δ46-110 proteins. The same membrane was then reprobed with an anti-Gab1 antiserum. (B) Gab1 protein was immunoprecipitated from EGF-stimulated wild-type (+/+), heterozygous (+/−), homozygous mutant (−/−), and two rescue cell lines, R2 and R4, in which the wild-type Shp-2 cDNA was reintroduced (55). Each immunoprecipitation was performed with 800 μg of total protein except in lane 5, which had 3 mg of total protein. In the lower panel, equal amounts (40 μg) of cell lysates were directly immunoblotted with the anti-Shp-2 antibody.