Fig. 2

Immunoblotting analysis of HEK293T cells transfected with mutant MTOR, MAP2K1, and PTPN11 a An in-frame (15-bp) MTOR deletion, c.4339_4353del p.(Ala1447_Glu1451del), detected in our study resulted in significantly higher phospho-S6 expression compared to wild-type (One-way ANOVA followed by Dunnett's post-hoc test; Empty: P = 0.99; Mutant: P =  < 0.0001; PC: P = 0.0079). Positive control (PC), c.4379T > C p.(Leu1460Pro). b The MAP2K1 variant, c.173_187del p.(Gln58_Glu62del), resulted in significantly higher phospho-S6 and phospho-ERK levels compared to the wild-type (Two-tailed paired t-test; pERK/ERK: P = 0.0048; pS6/S6: P ≤ 0.0001). c, d Time course of relative expression of phospho-ERK and phospho-S6 under stimulation with EGF in cells with PTPN11 transfection (c) and co-transfection with PTPN11 and GAB1 (d). Two-way repeated measurement ANOVA in c; pERK/ERK: P = 0.35; pS6/S6: P = 0.0028. Sidak’s post hoc test in pS6/S6 of c; 0 min: P = 0.63; 5 min: P = 0.96; 30 min: P = 0.58; 60 min: P = 0.34; 120 min: P = 0.14. Two-way repeated measurement ANOVA in d; pERK/ERK: P = 0.38; pS6/S6: P = 0.96. Bars in a, b represent the mean, and those in c, d represent SD. **P < 0.01; ****P < 0.0001. N.S. not significant