Fig 5.

The Ras/MAPK pathway modulates Shp2 recruitment in an RSK-dependent manner. (A) HEK293 cells were transfected with Myc-Gab2, serum starved overnight, pretreated with PD184352 (10 μM), and stimulated with EGF (25 ng/ml) over a time course. Associated endogenous Grb2 and Shp2 within Myc-Gab2 immunoprecipitates were assayed by immunoblotting. (B) HEK293 cells were transfected with Myc-tagged Gab2, serum starved overnight, and pretreated with PD184352 (10 μM) and/or PMA (100 ng/ml) for 30 min, prior to treatment with EGF (25 ng/ml) for 5 or 10 min. Associated Shp2 was assayed as described for panel A. (C) As for panel A, except that cells were cotransfected with MEK-DD, serum starved overnight, and stimulated with EGF over a time course. (D) As for panel A, except that cells were pretreated with the RSK inhibitor BI-D1870 (10 μM) for 30 min prior to EGF stimulation.