ATF3 contributes to brucine-triggered glioma cell ferroptosis via promotion of hydrogen peroxide and iron

Abstract

Ferroptotic cell death is characterized by iron-dependent lipid peroxidation that is initiated by ferrous iron and H ₂O ₂ via Fenton reaction, in which the role of activating transcription factor 3 (ATF3) remains elusive. Brucine is a weak alkaline indole alkaloid extracted from the seeds of Strychnos nux-vomica, which has shown potent antitumor activity against various tumors, including glioma. In this study, we showed that brucine inhibited glioma cell growth in vitro and in vivo, which was paralleled by nuclear translocation of ATF3, lipid peroxidation, and increases of iron and H ₂O ₂. Furthermore, brucine-induced lipid peroxidation was inhibited or exacerbated when intracellular iron was chelated by deferoxamine (500 μM) or improved by ferric ammonium citrate (500 μM). Suppression of lipid peroxidation with lipophilic antioxidants ferrostatin-1 (50 μM) or liproxstatin-1 (30 μM) rescued brucine-induced glioma cell death. Moreover, knockdown of ATF3 prevented brucine-induced accumulation of iron and H ₂O ₂ and glioma cell death. We revealed that brucine induced ATF3 upregulation and translocation into nuclei via activation of ER stress. ATF3 promoted brucine-induced H ₂O ₂ accumulation via upregulating NOX4 and SOD1 to generate H ₂O ₂ on one hand, and downregulating catalase and xCT to prevent H ₂O ₂ degradation on the other hand. H ₂O ₂ then contributed to brucine-triggered iron increase and transferrin receptor upregulation, as well as lipid peroxidation. This was further verified by treating glioma cells with exogenous H ₂O ₂ alone. Moreover, H ₂O ₂ reversely exacerbated brucine-induced ER stress. Taken together, ATF3 contributes to brucine-induced glioma cell ferroptosis via increasing H ₂O ₂ and iron.