FIG. 7.

Pyk2 and Erk regulation is impaired by the disruption of the Hck binding site. (A) Baf-B03 cells were cotransfected with the expression vectors for Eg (lanes 3 and 4) or d771-811 (lanes 5 and 6) and Hck. After stimulation with medium or EPO, cells were lysed as described in the text. Lysates were separated on 10% SDS gels. After blotting, the membranes were incubated with antibodies against phosphorylated Pyk2 (upper panels) or with several control antibodies as indicated (lower panels). (B) Baf-B03 cells expressing the indicated proteins were stimulated for the indicated times with 40 U of EPO/ml. Thereafter, membranes were subjected to Western blots by using antibodies specific for activated Erk1/2 (upper panels). Equal loading was confirmed by blotting with normal Erk antibody (lower panels).