FIGURE 2.

Grb2 Tyr¹⁶⁰ is phosphorylated in the presence of active NPM-ALK. A, schematic representation of Grb2 structure and tyrosine residues localization. B, HEK-293T cells were co-transfected with Grb2 WT or Grb2 tyrosine mutants together with the active form of NPM-ALK or the NPM-ALK kinase-dead mutant and blotted with the indicated antibodies. m, mobility. C, HEK-293T cells were co-transfected with Grb2^WT, Grb2^Y160F, or Grb2^Y209F mutants and NPM-ALK. Total cells extracts were analyzed by Western blot for Grb2, ALK, and p-Tyr. D, anti-Grb2 immunoprecipitation was performed on HEK-293T cells transiently co-transfected with NPM-ALK or NPM-ALK^K210R and Grb2^WT or Grb2^Y160F. Western blots of total lysates and anti-Grb2-immunoprecipitated (IP) proteins were blotted with the indicated antibodies.