Fig. 8. uPA and EGF activate N-WASP with divergent requirement for PI3K. (A) Cells were stimulated with 10 ng/ml EGF for 1-10 minutes. EGFR immunoprecipitates were separated on 10% gels and immunoblotted for N-WASP. (B,C) shows N-WASP in Triton X100 soluble cell lysates (cytosolic fraction) and Triton X100 insoluble cell pellets (cytoskeletal fraction). (B) Cells stimulated with 10 ng/ml EGF or 10 nM uPA for 1-30 minutes are shown. (C) Cells stimulated for 10 minutes with 10 nM mut C uPA or 10 ng/ml EGF±LY 294002 are shown (5 µM; pre-treatment 1 hour). The results are representative of three to six experiments.