title: Oxidation of alpha-ketoglutarate is required for reductive carboxylation in cancer cells with mitochondrial defects.
abstract: Mammalian cells generate citrate by decarboxylating pyruvate in the mitochondria to supply the tricarboxylic acid (TCA) cycle. In contrast, hypoxia and other impairments of mitochondrial function induce an alternative pathway that produces citrate by reductively carboxylating α-ketoglutarate (AKG) via NADPH-dependent isocitrate dehydrogenase (IDH). It is unknown how cells generate reducing equivalents necessary to supply reductive carboxylation in the setting of mitochondrial impairment. Here, we identified shared metabolic features in cells using reductive carboxylation. Paradoxically, reductive carboxylation was accompanied by concomitant AKG oxidation in the TCA cycle. Inhibiting AKG oxidation decreased reducing equivalent availability and suppressed reductive carboxylation. Interrupting transfer of reducing equivalents from NADH to NADPH by nicotinamide nucleotide transhydrogenase increased NADH abundance and decreased NADPH abundance while suppressing reductive carboxylation. The data demonstrate that reductive carboxylation requires bidirectional AKG metabolism along oxidative and reductive pathways, with the oxidative pathway producing reducing equivalents used to operate IDH in reverse.
authors.affiliation: Children's Medical Center Research Institute, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA.
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Mullen
authors.affiliation: Children's Medical Center Research Institute, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA.
authors.forename: Zeping
authors.initials: Z
authors.lastname: Hu
authors.affiliation: Children's Medical Center Research Institute, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA.
authors.forename: Xiaolei
authors.initials: X
authors.lastname: Shi
authors.affiliation: Children's Medical Center Research Institute, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA.
authors.forename: Lei
authors.initials: L
authors.lastname: Jiang
authors.affiliation: Children's Medical Center Research Institute, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA.
authors.forename: Lindsey K
authors.initials: LK
authors.lastname: Boroughs
authors.affiliation: Advanced Imaging Research Center, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA.
authors.forename: Zoltan
authors.initials: Z
authors.lastname: Kovacs
authors.affiliation: Department of Pathology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA.
authors.forename: Richard
authors.initials: R
authors.lastname: Boriack
authors.affiliation: Department of Pathology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA.
authors.forename: Dinesh
authors.initials: D
authors.lastname: Rakheja
authors.affiliation: Department of Medicine, Northwestern University, Chicago, IL 60611-3008, USA; Department of Cell and Molecular Biology, Northwestern University, Chicago, IL 60611-3008, USA.
authors.forename: Lucas B
authors.initials: LB
authors.lastname: Sullivan
authors.affiliation: Urological Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: W Marston
authors.initials: WM
authors.lastname: Linehan
authors.affiliation: Department of Medicine, Northwestern University, Chicago, IL 60611-3008, USA; Department of Cell and Molecular Biology, Northwestern University, Chicago, IL 60611-3008, USA.
authors.forename: Navdeep S
authors.initials: NS
authors.lastname: Chandel
authors.affiliation: Children's Medical Center Research Institute, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA; McDermott Center for Human Growth and Development, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8502, USA. Electronic address: ralph.deberardinis@utsouthwestern.edu.
authors.forename: Ralph J
authors.initials: RJ
authors.lastname: DeBerardinis
mesh_terms: D000818:Animals; D002460:Cell Line; D045744:Cell Line, Tumor; D002952:Citric Acid Cycle; D006224:Cricetinae; D003412:Cricetulus; D005649:Fumarate Hydratase / Q000235:genetics; D006801:Humans; D007521:Isocitrate Dehydrogenase / Q000378:metabolism; D007656:Ketoglutaric Acids / Q000378:metabolism*; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D009243:NAD / Q000378:metabolism; D061087:NADP Transhydrogenase, AB-Specific / Q000378:metabolism; D009369:Neoplasms / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction
chemical_list: D007656:Ketoglutaric Acids; D009243:NAD; D007521:Isocitrate Dehydrogenase; D061087:NADP Transhydrogenase, AB-Specific; D005649:Fumarate Hydratase