Fig. 3

CircEIF4G3 destabilizes δ-catenin protein and inactivates β-catenin signaling. (A) TRAP experiment and LC-MS/MS analysis. GST protein expression was detected by western blot. δ-catenin protein (encoded by CTNND1 gene) was identified and indicated. (B) The binding of circEIF4G3 to δ-catenin protein was validated by TRAP assay followed by western blot. (C) The association of circEIF4G3 to δ-catenin protein was determined by RIP assay followed by qRT-PCR. (D) The relative expression levels of δ-catenin gene in GC cells with or without circEIF4G3 overexpression. (E–F) Western blot analyses of the indicated proteins in circEIF4G3 overexpressing GC cells. (G) TOP/FOP flash luciferase reporter assays for β-catenin activity in control and circEIF4G3 overexpressing GC cells. Results were normalized to the Renilla internal control. (H) Immunofluorescent staining of β-catenin (red) in control and circEIF4G3 overexpressing GC cells with or without LiCl treatment. Scale bar = 25 μm. (I) QRT-PCR analysis of circEIF4G3 expression and western blot assay for δ-catenin protein expression in paired tumor and non-tumor tissues. Data are shown as means ± SD. *P < 0.05, **P < 0.01, ***P < 0.001