FIG. 2.

MEK1, MEK2, and ERK1 coimmunoprecipitate with IQGAP1. (A) MCF-7 cells were transiently transfected with pMCL-MKK1 (MEK1) or pMCL-MKK2 (MEK2). Twenty-four hours later, cells were starved of serum overnight and then incubated with vehicle (−) or 100 ng/ml EGF (+) for 10 min. After lysis, equal amounts of protein were incubated with NIRS or immunoprecipitated (IP) with anti-IQGAP1 polyclonal antibody. Both immune complexes (IP) and unfractionated lysates (Lysate) were resolved by SDS-PAGE. After transfer to PVDF membranes, the blots were probed with monoclonal anti-IQGAP1 and anti-MEK antibodies. Data are representative of two independent experiments for MEK1 and four independent experiments for MEK2. (B) Untransfected MCF-7 cells were starved of serum for 16 h and then incubated with vehicle (−) or 100 ng/ml EGF (+) for 10 min. Cells were exposed to a cross-linker, DSP, before lysis. Equal amounts of protein lysate were immunoprecipitated (IP) as described for panel A. Western blots were probed with anti-IQGAP1 and anti-ERK1 antibodies. Data are representative of three independent experiments.