Quantitative flux analysis reveals folate-dependent NADPH production

By Jing Fan, Jiangbin Ye, Jurre J. Kamphorst, Tomer Shlomi, Craig B. Thompson, Joshua D. Rabinowitz. Nature, 2014

1,044 citations recorded by the source corpus.

ATP is the dominant energy source in animals for mechanical and electrical work (for example, muscle contraction or neuronal firing). For chemical work, there is an equally important role for NADPH, which powers redox defence and reductive biosynthesis. The most direct route to produce NADPH from glucose is the oxidative pentose phosphate pathway, with malic enzyme sometimes also important. Although the relative contribution of glycolysis and oxidative phosphorylation to ATP production has been extensively analysed, similar analysis of NADPH metabolism has been lacking. Here we demonstrate the ability to directly track, by liquid chromatography–mass spectrometry, the passage of deuterium from labelled substrates into NADPH, and combine this approach with carbon labelling and mathematical modelling to measure NADPH fluxes. In proliferating cells, the largest contributor to cytosolic NADPH is the oxidative pentose phosphate pathway. Surprisingly, a nearly comparable contribution comes from serine-driven one-carbon metabolism, in which oxidation of methylene tetrahydrofolate to 10-formyl-tetrahydrofolate is coupled to reduction of NADP+ to NADPH. Moreover, tracing of mitochondrial one-carbon metabolism revealed complete oxidation of 10-formyl-tetrahydrofolate to make NADPH. As folate metabolism has not previously been considered an NADPH producer, confirmation of its functional significance was undertaken through knockdown of methylenetetrahydrofolate dehydrogenase (MTHFD) genes. Depletion of either the cytosolic or mitochondrial MTHFD isozyme resulted in decreased cellular NADPH/NADP+ and reduced/oxidized glutathione ratios (GSH/GSSG) and increased cell sensitivity to oxidative stress. Thus, although the importance of folate metabolism for proliferating cells has been long recognized and attributed to its function of producing one-carbon units for nucleic acid synthesis, another crucial function of this pathway is generating reducing power.

Subfields: Cancer Research, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/980872385c227588a566b91d6a09ab042a04119b

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Citations in the corpus

Regulation of mammalian nucleotide metabolism and biosynthesis (2015, methodology): “The source of this NADPH is variable according to cell type (98), but the oxidative branch of the pentose phosphate pathway (PPP) is an efficient cytoplasmic source, which also links glucose metabolism to ribose production.”
Targeting Metabolism for Cancer Therapy (2017, background): “Serine can also contribute to NADPH production via the folate cycle, which serves to maintain redox homeostasis and support anabolic reactions (Fan et al., 2014; Lewis et al., 2014; Ye et al., 2014).”
Understanding the intersections between metabolism and cancer biology (2017, neutral): “A uthor M anuscript A uthor M anuscript A uthor M anuscript A uthor M anuscript source of cytosolic NAPDH (Fan et al., 2014; Lewis et al., 2014) (Figure 2).”
Lifestyle, Oxidative Stress, and Antioxidants: Back and Forth in the Pathophysiology of Chronic Diseases (2020, background): “40) and a recently identified folate-dependent mechanism (Fan et al., 2014).”
The Roles of NRF2 in Modulating Cellular Iron Homeostasis (2017, background): “While NADPH abundance has been determined as a biomarker for ferroptosis sensitivity (139), no investigations to date have determined the role of NRF2 in enhancing or ameliorating ferroptosis by mediating NADPH levels.”
MYC, Metabolism, and Cancer. (2015, background): “In addition, genetic reduction of MTHFD2 leads to oxidative stress, illustrating the importance of this pathway in a model cell culture system (131).”
ROS homeostasis and metabolism: a dangerous liason in cancer cells (2016, background): “The PPP is a major catabolic pathway of glucose through which cancer cells produce large amounts of ribose-5 phosphate, a precursor of nucleotide synthesis and NADPH, a key molecule that is used to drive anabolic processes and to detoxify harmful ROS.90 Activation of the PPP represents a key hallmar”