Figure 4.

Time course of receptor-mediated tyrosine phosphorylation of the β chain, SHP-2, and Jak3. Cytokine-deprived parental CTLL-2 cells or cells coexpressing the chimeric receptor chains αγ-PROX + ββ were stimulated at room temperature with either IL-2 (200 units/ml) or GM-CSF (200 ng/ml). At the indicated time points [expressed in seconds (s) or minutes], cells were added directly to lysis buffer and subjected to immunoprecipitation (IP) followed by immunoblotting with antibodies to murine IL-2Rβ (Left), human IL-2Rβ (Right), SHP-2, or phosphotyrosine. Equivalent loading of samples across lanes was confirmed by reprobing with the same antibody used for immunoprecipitation (data not shown). These second blots also were used to confirm the phosphotyrosine content of murine IL-2Rβ, chimeric ββ, and SHP-2.