Abstract 1843 Metabolic adaptations to cysteine starvation that influence iron-dependent cell death

Cysteine is a thiol-containing amino acid that supports diverse biological functions and prevents the iron-dependent form of cell death known as ferroptosis. To understand how metabolic changes contribute to ferroptosis under cystine starvation, we profiled the ferroptosis sensitivity and metabolism of a panel of non-small cell lung cancer cell cell lines following cystine withdrawal. We found that cystine deprivation induced both GSH depletion and glutamate accumulation to accelerate ferroptosis. Surprisingly, we found that cystine starvation induced the accumulation of γ-glutamyl peptide synthesis in cysteine starved cells, which was mediated by the GSH synthesis machinery. γ-glutamyl peptide production reduced intracellular glutamate accumulation, thereby preventing glutamate-mediated oxidative stress and ferroptosis induction. While rapid cessation of cytosolic glutathione synthesis occurred following cysteine starvation, it was unknown how mitochondrial cysteine metabolism was altered under starvation, and how this contributed to ferroptosis. We found that despite the rapid depletion of cellular cysteine upon cystine starvation, cysteine-dependent synthesis of Fe-S clusters was maintained in the mitochondria of lung cancer cells, resulting in a maintenance of mitochondrial function and redox state. Moreover, we found that mitochondrial glutathione sustained Fe-S synthesis via CHAC1-mediated catabolism of glutathione. Disruption of CHAC1 resulted in a failure to maintain Fe-S protein activity. Surprisingly, disrupting Fe-S cluster synthesis or mitochondrial electron transport protected against the induction of ferroptosis under cysteine starvation, suggesting that the preservation of mitochondrial function is antagonistic to survival under starved conditions. Overall, our demonstrate that mitochondria preserve their function at the expense of overall cellular health, leading which promotes ferroptosis when cysteine is limiting. This work was supported by grants from the NIH/NCI (R37CA230042 and P01CA250984).