Fig. 2: CUR5g inhibits autophagosome degradation by blocking autophagosome-lysosome fusion. | Cell Death Discovery

Fig. 2: CUR5g inhibits autophagosome degradation by blocking autophagosome-lysosome fusion.

From: CUR5g, a novel autophagy inhibitor, exhibits potent synergistic anticancer effects with cisplatin against non-small-cell lung cancer

Fig. 2

A Western blot analysis of LC3B-II and SQSTM1 levels in A549 cells treated with DMSO or 10 μM CUR5g in the absence or presence of 3-MA (10 mM) for 24 h. GAPDH was used as a loading control. (n = 3; *p < 0.05, **p < 0.01 vs. Control). B Western blot analysis of LC3B-II and SQSTM1 levels in A549 cells treated with DMSO or 10 μM CUR5g in the absence or presence of CQ (30 μM) for 24 h. GAPDH was used as a loading control. (n = 3; *p < 0.05, **p < 0.01 vs. 0). C Western blot analysis of LC3B-II and SQSTM1 levels in A549 cells cultured in complete medium or EBSS in the absence or presence of CUR5g (10 μM) for 24 h. GAPDH was used as a loading control. (n = 3; *p < 0.05, **p < 0.01 vs. 0, #p < 0.05 vs. CUR5g). D Representative fluorescence photographs of HEK293T cells stably expressing RFP-GFP-LC3B reporter. Cells were treated with DMSO or CUR5g (10 μM) in complete medium for 24 h, treated with EBSS for 6 h. Bafilomycin A1 (100 nM)-treated cells were used as positive controls. Nuclei were stained with DAPI. Scale bar = 10 μm. Histogram shows average number of autophagosomes (yellow) and autolysosomes (red) per cell. (n = 3; *p < 0.05; **p < 0.01 vs. control). E Western blot analysis of LC3B-II and SQSTM1 levels in A549 cells treated with 10 μM CUR5g for 0–24 h. GAPDH was used as a loading control. (n = 3; *p < 0.05 vs. 0). F Representative fluorescence photographs of the colocalization of GFP-LC3B and LysoTracker Red in U87 cells treated with DMSO, CUR5g (10 μM) for 24 h, treated with CQ (30 μM), or incubated with EBSS for 6 h. Nuclei were stained with DAPI. Scale bar = 10 μm. Histogram shows the percentage of LC3B+LysoTracker Red+ puncta (yellow) relative to LC3B+ puncta (green). (n = 3; **p < 0.01 vs. Control). G Representative fluorescence images of the colocalization of GFP-LC3B and LAMP1 in U87 cells treated with DMSO, CUR5g (10 μM) for 24 h, treated with CQ (30 μM), or incubated with EBSS for 6 h. Nuclei were stained with DAPI. Scale bar = 10 μm. Histogram shows the percentage of LAMP1+LC3B+ puncta (yellow) relative to LC3B+ puncta (green). (n = 3; **p < 0.01 vs. Control).

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