Proteasome inhibition bidirectionally regulates ferroptosis sensitivity

Ferroptosis is a non-apoptotic cell death mechanism whose fundamental regulation continues to be elucidated. The proteasome is an essential multiprotein complex important for protein turnover. How proteasome function regulates ferroptosis is difficult to study given that proteasome inhibition causes apoptosis. Here, we isolated the effects of proteasome inhibition on ferroptosis by combining small molecule cell death inducers, pathway-specific cell death inhibitors, and direct cell death imaging. We find that proteasome inhibition enhances sensitivity to ferroptosis induced by glutathione peroxidase 4 (GPX4) inhibition while simultaneously promoting resistance to ferroptosis induced by system x_c^- inhibition. Sensitization to GPX4 inhibition requires protein synthesis but not the apoptosis execution machinery and is opposed by the activating transcription factor 4 (ATF4) stress response pathway. These findings establish proteasome-mediated protein turnover as a context-dependent regulator of ferroptosis and identify protein synthesis and ATF4 as key determinants of GPX4 inhibitor sensitivity.