Fig. 6

Catalase-deficient mature adult mice induce mTORC1 activation. A HepG2 cells were treated with 1 mM LLOMe for 24 h and co-treated with 5 mM NAC overnight. Representative fluorescence images of HepG2 cells immunostained with Lysotracker (red) and DAPI (blue). Scale bar represents 5 μm. B Representative fluorescence images of DCFH-DA staining of HepG2 cells treated as in a. Scale bar represents 20 μm. C Lysates of HepG2 cells treated as in A were fractioned as explained in the Materials and Methods section. Equivalent volumes of each fraction were subjected to immunoblotting using the indicated antibodies. D–F Quantified protein level of cathD, cathB and LAMP1 normalized with actin. G Representative fluorescence image of HepG2 cells treated as in A, immunostained with anti-TFEB (red) and DAPI (blue). Scale bar represents 20 μm. Quantified bar represents percentage of nuclear localization of TFEB on treated cell as indicated. H Protein was extracted from HepG2 cells as in a. Immunoblot analysis was performed using whole-cell lysates with indicated antibodies. Bar graph represents mean ± SD (n = 3 experiments). *P < 0.05; **P < 0.0001