Unlocking ferroptosis heterogeneity: ATF4 versus SREBF transcriptional programs

Abstract Regulated cell death (RCD) encompasses diverse forms induced by common cellular stressors, including oxidative stress and endoplasmic reticulum stress. Cells may display features from multiple RCD types, implying coexistence of “pure” and “mixed” forms of cell death. Ferroptosis is an iron-dependent form of RCD characterized by lipid peroxide accumulation and absence of specific biomarkers. Its regulation is heterogeneous and pathway-dependent, reflecting the broader complexity of the RCD spectrum. This study hypothesized two major regulatory pathways governing ferroptosis induction and functioning. The first pathway operates through GPX4, a central enzyme preventing ferroptosis by reducing lipid peroxides. Genetic GPX4 knockout and chemical GPX4 inhibition with ML210 resulted in significant upregulation of mevalonate pathway regulators SREBF1 and SREBF2, indicating compensatory anti-ferroptotic activation through the mevalonate pathway. The second pathway functions through the Xc⁻/GSH system critical for glutathione synthesis. System Xc⁻ inhibition leads to intracellular cysteine depletion with subsequent glutathione depletion, inducing ferroptotic cell death. SREBF transcription factor cascades were downregulated in system Xc⁻/GSH depletion experiments, contrasting with GPX4 inhibition responses. Instead, the system Xc⁻/GSH pathway is primarily governed by ATF4 to maintain cystine uptake and glutathione biosynthesis under stress conditions. These findings confirm ferroptosis as a heterogeneous spectrum of cell death with multiple interacting yet mechanistically distinct pathways. Moreover, these data provide a dynamic multi-omics resource for further ferroptosis research.