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Fig. 6 | BMC Cancer

Fig. 6

From: Intracellular expression of arginine deiminase activates the mitochondrial apoptosis pathway by inhibiting cytosolic ferritin and inducing chromatin autophagy

Fig. 6

Chromatin autophagy assay at the later time point of arginine deprivation. a: GFP-LC3 reporter fluorescence assay for autophagy in MRC5, HepG2 and PC3 cells. Cells were co-transfected with pcDNA4-ADI plasmid and pEGFP-LC3 plasmid. The fluorescence of EGFP protein was detected by OLIMPUS inverted fluorescence microscope SteREO Discovery V12. b: Immunoblot of LC3-I and LC3-II in MRC5, HepG2 and PC3 cells. Cells were treated as the description of Fig. 6a. LC3 antibody was used to detect LC3-I and LC3-II proteins. C-myc-tag antibody was used to detect c-myc-tag-fused ADI. Full-length blots are presented in Supplementary Fig. S6A. c: the relative quantification for protein expressions in MRC5, PC3 and HepG2 cell lines. Grey scales of protein bands from Fig. 6b were detected by ImageJ 1.52. P values were calculated by comparing pcDNA4-ADI plasmids-treated cells with pcDNA4 plasmid-treated cells in the respective cell lines. **P < 0.01; ***P < 0.001. d: Immunoblots of H3 protein expression in HepG2 and PC3 cells. Cells were transfected with pcDNA4-ADI plasmid. Histone H3 antibody (Cat. # 17168–1-AP) were used to detect H3 protein. Full-length blots are presented in Supplementary Fig. S6B. e: the relative quantification for protein expressions in MRC5, PC3 and HepG2 cell lines. Grey scales of protein bands from Fig. 6d were detected by ImageJ 1.52. P values were calculated by comparing other cells with 24 h-treated cells in the respective cell lines. **P < 0.01; ***P < 0.001. f: Immunofluorescence assay for chromatin autophagy. Cells were cultured in DMEM medium with 2% FBS. Histone H3 antibody (Cat. # 17168–1-AP) were used to detect H3 protein in cells. TRITC conjugated goat anti-rabbit antibody (Cat. # AS10–1018) was used to detect H3 antibody and display the immunofluorescence

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