Fig. 6
From: Missense variant in TTBK2 kinase domain causes loss of function and impaired protein phosphorylation
The expression of several autophagy-related proteins is increased in FLAG-TTBK2-L209F cells but the autophagic flux seems unaffected. (a) Protein expression analysis of p62, LC3B-I, LC3B-II and LC3B-II/LC3B-I ratio in non-treated FLAG-TTBK2-WT (WT) and FLAG-TTBK2-L209F (L209F) cells (incubated with the control vehicle DMEM) and in cells incubated with 10 µM chloroquine (CQ) for autophagy blockage. Immunoblotting was performed using specific antibodies and Ponceau S staining was used as loading control. (b) Protein expression analysis of the upstream autophagy-related phosphoproteins raptor, ULK1 and beclin-1 in WT and L209F cells by immunoblotting. Raptor Ser792, ULK1 Ser555 and Beclin-1 Ser93 phosphorylated levels, as well as total raptor, ULK1 and beclin-1 levels were normalized to Ponceau S staining (loading control). Phosphorylated levels of raptor, ULK1 and beclin-1 were also normalized to the respective total protein levels. Quantification data are presented in percentage as mean ± SD of at least three independent experiments; non-significant (ns), *P ≤ 0.05, **P ≤ 0.01 and ***P ≤ 0.001 compared with the WT cell line and with non-treated cells in panel (a) (one-way ANOVA/Tukey); non-significant (ns), *P ≤ 0.05 and **P ≤ 0.01 compared with the WT cell line in panel (b) (t-test).
