Fig. 2

Hotspot mutations lead to increased RIT1 protein abundance. (A) Lollipop plot demonstrating mutations detected in RIT1. (B) 3-D view of predicted human RIT1 structure with three hotspot mutations labeled. Structure was rendered using AlphaFold. (C) Representative images of HeLa cells transiently transfected with GFP-tagged wild-type RIT1 (WT) or mutant forms as indicated. (D) Representative immunoblotting image and quantification for RIT1 expression in HBE135-E6E7 stable cell lines as indicated. Tubulin was used as protein loading control. (E) Representative image of RIT1 protein detected by immunoblotting and corresponding quantification in HBE135-E6E7 cells with stable expression of WT or mutated RIT1 as indicated after treatment with 50 µg/mL of cycloheximide (CHX) to stop protein synthesis. RIT1 levels in cells at the time point zero were normalized as 100%. Tubulin was used as protein loading control. n = 3 for each group. Data were presented as mean ± s.e.m. *P < 0.05, ***P < 0.001