FIG. 5.

Role for p120-RasGAP in signaling from EphB2 in NG108 cells. (A) NG-EphB2 and NG-EphB2-EE clones were stimulated with aggregated Fc-ephrinB1 ligand for the indicated times, and whole-cell lysates were separated by SDS-PAGE and immunoblotted (WB) for anti-pTyr. Tyrosine-phosphorylated EphB2 and p62dok-1 are indicated by arrows. (B) Cell lysates from (panel A) were immunoblotted for phosphorylated ERK1/2 (upper panel) and reprobed for ERK1 (lower panel). (C) Serum-starved NG-EphB2 and NG-EphB2-EE cells were incubated in the presence or absence of 2 μg of aggregated ephrin-B1 per ml for 20 min and lysed as indicated in Materials and Methods. EphB2 immunoprecipitates were blotted for p120-RasGAP (upper panel) and reprobed with anti-pTyr antibodies (second panel) and anti-EphB2 (third panel). For control, equivalent amounts of whole-cell lysate (WCL) were immunoblotted for p120-RasGAP to indicate equal input (bottom panel).