Stem-cell fate can be influenced by metabolite levels in culture, but it is not known whether physiological variations in metabolite levels in normal tissues regulate stem-cell function in vivo. Here we describe a metabolomics method for the analysis of rare cell populations isolated directly from tissues and use it to compare mouse haematopoietic stem cells (HSCs) to restricted haematopoietic progenitors. Each haematopoietic cell type had a distinct metabolic signature. Human and mouse HSCs had unusually high levels of ascorbate, which decreased with differentiation. Systemic ascorbate depletion in mice increased HSC frequency and function, in part by reducing the function of Tet2, a dioxygenase tumour suppressor. Ascorbate depletion cooperated with Flt3 internal tandem duplication (Flt3ITD) leukaemic mutations to accelerate leukaemogenesis, through cell-autonomous and possibly non-cell-autonomous mechanisms, in a manner that was reversed by dietary ascorbate. Ascorbate acted cell-autonomously to negatively regulate HSC function and myelopoiesis through Tet2-dependent and Tet2-independent mechanisms. Ascorbate therefore accumulates within HSCs to promote Tet activity in vivo, limiting HSC frequency and suppressing leukaemogenesis.

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  1. 1.

    et al. Genotoxic consequences of endogenous aldehydes on mouse haematopoietic stem cell function. Nature 489, 571–575 (2012)

  2. 2.

    et al. IDH1 and IDH2 mutations in gliomas. N. Engl. J. Med. 360, 765–773 (2009)

  3. 3.

    , & Dietary and metabolic control of stem cell function in physiology and cancer. Cell Stem Cell 14, 292–305 (2014)

  4. 4.

    et al. NAD+ repletion improves mitochondrial and stem cell function and enhances life span in mice. Science 352, 1436–1443 (2016)

  5. 5.

    et al. Metabolic differentiation in the embryonic retina. Nat. Cell Biol. 14, 859–864 (2012)

  6. 6.

    et al. Dependence of mouse embryonic stem cells on threonine catabolism. Science 325, 435–439 (2009)

  7. 7.

    et al. Vitamin C induces Tet-dependent DNA demethylation and a blastocyst-like state in ES cells. Nature 500, 222–226 (2013)

  8. 8.

    , , , & Intracellular α-ketoglutarate maintains the pluripotency of embryonic stem cells. Nature 518, 413–416 (2015)

  9. 9.

    et al. Dipeptide species regulate p38MAPK–Smad3 signalling to maintain chronic myelogenous leukaemia stem cells. Nat. Commun. 6, 8039 (2015)

  10. 10.

    et al. Regulation of glycolysis by Pdk functions as a metabolic checkpoint for cell cycle quiescence in hematopoietic stem cells. Cell Stem Cell 12, 49–61 (2013)

  11. 11.

    et al. The distinct metabolic profile of hematopoietic stem cells reflects their location in a hypoxic niche. Cell Stem Cell 7, 380–390 (2010)

  12. 12.

    The SLC23 family of ascorbate transporters: ensuring that you get and keep your daily dose of vitamin C. Br. J. Pharmacol. 164, 1793–1801 (2011)

  13. 13.

    et al. Gene Expression Commons: an open platform for absolute gene expression profiling. PLoS ONE 7, e40321 (2012)

  14. 14.

    et al. Vitamin C promotes maturation of T-cells. Antioxid. Redox Signal. 19, 2054–2067 (2013)

  15. 15.

    , & Regulation of the epigenome by vitamin C. Annu. Rev. Nutr. 35, 545–564 (2015)

  16. 16.

    et al. The histone demethylases Jhdm1a/1b enhance somatic cell reprogramming in a vitamin-C-dependent manner. Cell Stem Cell 9, 575–587 (2011)

  17. 17.

    , & Protein hydroxylation: prolyl 4-hydroxylase, an enzyme with four cosubstrates and a multifunctional subunit. FASEB J. 3, 1609–1617 (1989)

  18. 18.

    Ascorbic acid and carnitine biosynthesis. Am. J. Clin. Nutr. 54, 1147S–1152S (1991)

  19. 19.

    , , & Effect of ascorbate on the activity of hypoxia-inducible factor in cancer cells. Cancer Res. 63, 1764–1768 (2003)

  20. 20.

    et al. Adult hematopoietic stem cells lacking Hif-1α self-renew normally. Blood 127, 2841–2846 (2016)

  21. 21.

    et al. Ascorbic acid enhances Tet-mediated 5-methylcytosine oxidation and promotes DNA demethylation in mammals. J. Am. Chem. Soc. 135, 10396–10403 (2013)

  22. 22.

    et al. Vitamin C modulates TET1 function during somatic cell reprogramming. Nat. Genet. 45, 1504–1509 (2013)

  23. 23.

    , , & Ascorbate induces ten-eleven translocation (Tet) methylcytosine dioxygenase-mediated generation of 5-hydroxymethylcytosine. J. Biol. Chem. 288, 13669–13674 (2013)

  24. 24.

    et al. Tet proteins can convert 5-methylcytosine to 5-formylcytosine and 5-carboxylcytosine. Science 333, 1300–1303 (2011)

  25. 25.

    et al. Conversion of 5-methylcytosine to 5-hydroxymethylcytosine in mammalian DNA by MLL partner TET1. Science 324, 930–935 (2009)

  26. 26.

    et al. Clonal evolution of preleukemic hematopoietic stem cells precedes human acute myeloid leukemia. Sci. Transl. Med. 4, 149ra118 (2012)

  27. 27.

    et al. Tet2 loss leads to increased hematopoietic stem cell self-renewal and myeloid transformation. Cancer Cell 20, 11–24 (2011)

  28. 28.

    et al. Ten-eleven-translocation 2 (TET2) negatively regulates homeostasis and differentiation of hematopoietic stem cells in mice. Proc. Natl Acad. Sci. USA 108, 14566–14571 (2011)

  29. 29.

    et al. TET2 inactivation results in pleiotropic hematopoietic abnormalities in mouse and is a recurrent event during human lymphomagenesis. Cancer Cell 20, 25–38 (2011)

  30. 30.

    et al. Genetic characterization of TET1, TET2, and TET3 alterations in myeloid malignancies. Blood 114, 144–147 (2009)

  31. 31.

    et al. Mutational cooperativity linked to combinatorial epigenetic gain of function in acute myeloid leukemia. Cancer Cell 27, 502–515 (2015)

  32. 32.

    et al. Mutation in TET2 in myeloid cancers. N. Engl. J. Med. 360, 2289–2301 (2009)

  33. 33.

    , , & Serum vitamin C and the prevalence of vitamin C deficiency in the United States: 2003–2004 National Health and Nutrition Examination Survey (NHANES). Am. J. Clin. Nutr. 90, 1252–1263 (2009)

  34. 34.

    et al. Inhibition of TET2-mediated conversion of 5-methylcytosine to 5-hydroxymethylcytosine disturbs erythroid and granulomonocytic differentiation of human hematopoietic progenitors. Blood 118, 2551–2555 (2011)

  35. 35.

    et al. Prognostic relevance of integrated genetic profiling in acute myeloid leukemia. N. Engl. J. Med. 366, 1079–1089 (2012)

  36. 36.

    et al. TET1 is a tumor suppressor of hematopoietic malignancy. Nat. Immunol. 16, 653–662 (2015)

  37. 37.

    et al. Acute loss of TET function results in aggressive myeloid cancer in mice. Nat. Commun. 6, 10071 (2015)

  38. 38.

    , , & Vitamin C status and mortality in US adults. Am. J. Clin. Nutr. 72, 139–145 (2000)

  39. 39.

    et al. Relation between plasma ascorbic acid and mortality in men and women in EPIC-Norfolk prospective study: a prospective population study. Lancet 357, 657–663 (2001)

  40. 40.

    et al. Vitamin C increases viral mimicry induced by 5-aza-2′-deoxycytidine. Proc. Natl Acad. Sci. USA 113, 10238–10244 (2016)

  41. 41.

    , , , & Ascorbic acid serum levels are reduced in patients with hematological malignancies. Results Immunol. 6, 8–10 (2016)

  42. 42.

    et al. High-dose vitamin C versus placebo in the treatment of patients with advanced cancer who have had no prior chemotherapy—a randomized double-blind comparison. N. Engl. J. Med. 312, 137–141 (1985)

  43. 43.

    et al. Vitamin C selectively kills KRAS and BRAF mutant colorectal cancer cells by targeting GAPDH. Science 350, 1391–1396 (2015)

  44. 44.

    et al. Pharmacologic doses of ascorbate act as a prooxidant and decrease growth of aggressive tumor xenografts in mice. Proc. Natl Acad. Sci. USA 105, 11105–11109 (2008)

  45. 45.

    et al. Recurrent somatic TET2 mutations in normal elderly individuals with clonal hematopoiesis. Nat. Genet. 44, 1179–1181 (2012)

  46. 46.

    et al. Clonal hematopoiesis and blood-cancer risk inferred from blood DNA sequence. N. Engl. J. Med. 371, 2477–2487 (2014)

  47. 47.

    et al. Age-related clonal hematopoiesis associated with adverse outcomes. N. Engl. J. Med. 371, 2488–2498 (2014)

  48. 48.

    et al. Aortic wall damage in mice unable to synthesize ascorbic acid. Proc. Natl Acad. Sci. USA 97, 841–846 (2000)

  49. 49.

    et al. FLT3 mutations confer enhanced proliferation and survival properties to multipotent progenitors in a murine model of chronic myelomonocytic leukemia. Cancer Cell 12, 367–380 (2007)

  50. 50.

    et al. Ascorbic-acid transporter Slc23a1 is essential for vitamin C transport into the brain and for perinatal survival. Nat. Med. 8, 514–517 (2002)

  51. 51.

    , , & Inducible gene targeting in mice. Science 269, 1427–1429 (1995)

  52. 52.

    , , , & Ascorbic acid and dehydroascorbic acid analyses in biological samples. Anal. Biochem. 204, 1–14 (1992)

  53. 53.

    et al. Oxidation of alpha-ketoglutarate is required for reductive carboxylation in cancer cells with mitochondrial defects. Cell Reports 7, 1679–1690 (2014)

  54. 54.

    , , & MetaboAnalyst 3.0—making metabolomics more meaningful. Nucleic Acids Res. 43, W251–W257 (2015)

  55. 55.

    et al. Revised map of the human progenitor hierarchy shows the origin of macrophages and dendritic cells in early lymphoid development. Nat. Immunol. 11, 585–593 (2010)

  56. 56.

    et al. Bethesda proposals for classification of nonlymphoid hematopoietic neoplasms in mice. Blood 100, 238–245 (2002)

  57. 57.

    , , & NparLD: an R software package for the nonparametric analysis of longitudinal data in factorial experiments. J. Stat. Softw. 50, 1–23 (2012)

  58. 58.

    et al. Deep imaging of bone marrow shows non-dividing stem cells are mainly perisinusoidal. Nature 526, 126–130 (2015)

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S.J.M. is a Howard Hughes Medical Institute (HHMI) Investigator, the Mary McDermott Cook Chair in Pediatric Genetics, the Kathryn and Gene Bishop Distinguished Chair in Pediatric Research, the director of the Hamon Laboratory for Stem Cells and Cancer, and a Cancer Prevention and Research Institute of Texas Scholar. M.A. was a Royal Commission for the Exhibition of 1851 Research Fellow. We thank F. Harrison for sharing the Slc23a2/ mice, N. Loof and the Moody Foundation Flow Cytometry Facility for flow cytometry, K. Correll, A. Leach and A. Gross for mouse colony management and BioHPC at UT Southwestern for providing high-performance computing. This work was supported by the Cancer Prevention and Research Institute of Texas and the National Institutes of Health (R37 AG024945 and R01 DK100848).

Author information


  1. Children’s Research Institute and the Department of Pediatrics, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA

    • Michalis Agathocleous
    • , Corbin E. Meacham
    • , Rebecca J. Burgess
    • , Elena Piskounova
    • , Zhiyu Zhao
    • , Genevieve M. Crane
    • , Brianna L. Cowin
    • , Emily Bruner
    • , Malea M. Murphy
    • , Zeping Hu
    • , Ralph J. DeBerardinis
    •  & Sean J. Morrison
  2. Department of Pathology, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA

    • Weina Chen
  3. Department of Medicine, University of Utah, Salt Lake City, Utah, USA

    • Gerald J. Spangrude
  4. Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA

    • Ralph J. DeBerardinis
    •  & Sean J. Morrison


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M.A. conceived and performed most experiments. C.E.M. performed experiments with Tet2fl and Tet2fl;Flt3ITD mice in Extended Data Figs 5, 8. R.J.B. and E.P. performed the histone methylation analysis. E.P. and Z.Z. performed RNA-sequencing analysis. Z.Z. performed the statistical analyses. G.M.C. assessed haematopathology in Figs 4, 5. E.B. and B.L.C. provided technical assistance. M.M.M. performed collagen staining. W.C. provided human bone marrow specimens. G.J.S. provided some of the data from Slc23a2/ mice. Z.H., M.A., and R.J.D. developed the metabolomics methods and R.J.D. helped to interpret metabolomics results. M.A. and S.J.M. designed experiments, interpreted results and wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Sean J. Morrison.

Reviewer Information Nature thanks H. Christofk, R. Levine and the other anonymous reviewer(s) for their contribution to the peer review of this work.

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