Evidence for GCKR involvement in Crk-I- and Crk-II-induced SAPK activation.

HEK 293 cells were cotransfected with constructs that direct the expression of HA-SAPK (lanes 1 to 6), Crk-II (lanes 1, 2), Crk-I (lanes 3, 4), or GCKR (lane 5) in the presence (lanes 1, 3) or absence (lanes 2, 4 to 6) of a construct that expressed a GCKR antisense RNA. HA-SAPK immunoprecipitates were subjected to an in vitro kinase assay using GST-jun^1–79 as a substrate. The fold induction compared to HA-SAPK alone is indicated. The levels of HA-SAPK, Crk-II, and Crk-I as detected by immunoblotting are shown below. These experiments were performed twice with similar results.