Fig. 4

SHP2 binding to gp130 is required for IL-6-induced phosphorylation of Gab1. a HEK293 and HEK293 Gab1-KO cells were seeded and transfected with expression vectors (0.4 μg) for IL-5Rα/gp130(YYFFFF) and IL-5Rβ/gp130(YYFFFF), for IL-5Rα/gp130(YFFFFF) and IL-5Rβ/gp130(YFFFFF), or for IL-5Rα/gp130(YFFFFF) and IL-5Rβ/gp130(YF)-SHP2 chimeric receptors. After 24 h, cells were serum-starved for 4 h. Cells were treated with IL-5 (100 ng/ml) for 10 min. Subsequently, cell lysates were prepared and proteins separated by SDS-PAGE. After Western blotting, membranes were stained for (p)Y627-Gab1, Gab1, (p)Erk1/2, and Erk1/2. Representative results of n = 5 independent experiments are shown. b For quantification of Erk2 phosphorylation, signals of (p)Erk2 and Erk2 were analysed via densitometry. The diagram shows the ratio of (p)Erk2 and Erk2. Maximal phosphorylation of Erk2 in each experiment was set to 100%. Data are given as mean of five independent experiments ± SD. Student’s t-test: n.s. = non-significant, *p < 0.05