Mitochondrial One-Carbon Pathway Supports Cytosolic Folate Integrity in Cancer Cells

Mammalian folate metabolism is comprised of cytosolic and mitochondrial pathways with nearly identical core reactions, yet the functional advantages of such an organization are not well understood. Using genome-editing and biochemical approaches, we find that ablating folate metabolism in the mitochondria of mammalian cell lines results in folate degradation in the cytosol. Mechanistically, we show that QDPR, an enzyme in tetrahydrobiopterin metabolism, moonlights to repair oxidative damage to tetrahydrofolate (THF). This repair capacity is overwhelmed when cytosolic THF hyperaccumulates in the absence of mitochondrially produced formate, leading to THF degradation. Unexpectedly, we also find that the classic antifolate methotrexate, by inhibiting its well-known target DHFR, causes even more extensive folate degradation in nearly all tested cancer cell lines. These findings shed light on design features of folate metabolism, provide a biochemical basis for clinically observed folate deficiency in QDPR-deficient patients, and reveal a hitherto unknown and unexplored cellular effect of methotrexate. [Display omitted] •Formate from mitochondrial serine metabolism protects cytosolic THF from degradation•Methotrexate causes degradation of cytosolic folate by a similar pathway•QDPR can reverse THF oxidation and preserve cytosolic folate levels•This role of QDPR can explain the folate deficiency in patients with QDPR mutations Stratification of folate metabolism into distinct cytosolic and mitochondrial compartments enables repair of oxidative-stress-induced damage to folate metabolites while maintaining overall cellular pools of this nutrient.