NADPH production by the oxidative pentose-phosphate pathway supports folate metabolism

NADPH donates high-energy electrons for antioxidant defence and reductive biosynthesis. Cytosolic NADP is recycled to NADPH by the oxidative pentose-phosphate pathway (oxPPP), malic enzyme 1 (ME1) and isocitrate dehydrogenase 1 (IDH1). Here we show that any one of these routes can support cell growth, but the oxPPP is uniquely required to maintain a normal NADPH/NADP ratio, mammalian dihydrofolate reductase (DHFR) activity and folate metabolism. These findings are based on CRISPR deletions of glucose-6-phosphate dehydrogenase (G6PD, the committed oxPPP enzyme), ME1, IDH1 and combinations thereof in HCT116 colon cancer cells. Loss of G6PD results in high NADP, which induces compensatory increases in ME1 and IDH1 flux. But the high NADP inhibits DHFR, resulting in impaired folate-mediated biosynthesis, which is reversed by recombinant expression of Escherichia coli DHFR. Across different cancer cell lines, G6PD deletion produced consistent changes in folate-related metabolites, suggesting a general requirement for the oxPPP to support folate metabolism. The oxidative pentose-phosphate pathway (oxPPP) is a major NADPH producer. Here the authors show that malic enzyme or isocitrate dehydrogenase can support the growth of cells lacking the oxPPP, but the oxPPP is necessary to maintain a normal NADPH/NADP ratio, DHFR activity and folate metabolism.