Figure 8. GCN2 participates in the eIF2α-ATF4-xCT pathway and cisplatin resistance in response to mitochondrial dysfunction and in cisplatin-resistant cells.
A. Western blot analysis of GCN2 activation in SC-M1 cells under oligomycin treatments for 24 h. B. Western blot analysis of GCN2 activation between the parental and the cisplatin-resistant gastric cancer cells. The activation of GCN2 was determined by Western blot with specific antibodies against phosphorylated GCN2 (p-GCN2) and GCN2. The immunoblot values were normalized to α-tubulin. C. Western blot analysis of the GCN2-eIF2α-ATF4-xCT pathway in the GCN2-silenced SC-M1 cells (siGCN2) and the control cells (siRNA for non-target sequence, siScramble, siScr) under oligomycin treatments for 24 h. The immunoblot values were normalized to α-tubulin. D. The specific siRNA (60 pmol for 4 × 105 cells in a 6-cm dish) against GCN2 was used to knock down GCN2 in the SC-M1 cells. The cells were treated with oligomycin and cisplatin for 48 h. The cell viability was determined by SRB assay. E. The GCN2 siRNA (60 pmol for 4 × 105 cells in a 6-cm dish) was used to knock down GCN2 in the SC-M1CisR cells, and the GCN2-eIF2α-ATF4-xCT pathway was analyzed by Western blot analysis. F. The GCN2-silenced SC-M1CisR cells (siGCN2) and the control SC-M1CisR cells were treated with cisplatin for 48 h. The cell viability was determined by SRB assay. Data represent the mean ± SEM of three independent experiments. *p < 0.05, compared to the control group or parental cells; & p < 0.05, compared to the individual siScr group.