FIGURE 5.

GPX4 regulated the level of GSTM1 by protein stabilization. (A) Representative Western blot image showing the GPX4 and GSTM1 expression in GPX4 knockdown PC9‐BrM3 cells. Ctrl, control PC9‐BrM3 cells; shNC, PC9‐BrM3 cells transfected with negative control shRNA; sh1, PC9‐BrM3 cells transfected with GPX4‐targeted shRNA vector 1; sh2, PC9‐BrM3 cells transfected with GPX4‐targeted shRNA vector 2. The bar graphs are summarized results from three independent experiments. **P < 0.01 versus shNC group. (B) Representative Western blot image showing the GPX4 and GSTM1 expression in PC9 cells with GPX4 overexpression. Ctrl, control PC9 cells; NC, PC9 transfected with negative control plasmid; OE, PC9 transfected with GPX4 plasmid. The bar graphs are summarized results from three independent experiments. **P < 0.01 versus NC group. (C) Immunofluorescent analysis of GPX4 (green) and GSTM1 (red) expression and distribution in PC9 cells with GPX4 overexpression. Cell nuclei are stained with DAPI (blue). Scale bar, 25 μm. NC, PC9 transfected with negative control plasmid; OE, PC9 transfected with GPX4 plasmid. (D) The potential interaction between GPX4 and GSTM1 was determined by immunoprecipitation using anti‐GPX4 and anti‐GSTM1 in PC9 cells with GPX4 overexpression. GPX4 was pulled down by anti‐GPX4, and then GPX4 and GSTM1 were detected by Western blot. On the contrary, GSTM1 was pulled down by anti‐GSTM1, and then GPX4 and GSTM1 were detected by Western blot. NC, PC9 transfected with negative control plasmid; OE, PC9 transfected with GPX4 plasmid. (E) Results of Western blot analysis showing protein expressions of GSTM1 in individual groups of cells following treatment with CHX (50 μM). The standardized quantitative curves show the protein degradation rate in the indicated groups. NC, PC9 transfected with negative control plasmid; OE, PC9 transfected with GPX4 plasmid (**P < 0.01). Representative images of each group from three independent experiments are presented