Ascorbate induces ten-eleven translocation (Tet) methylcytosine dioxygenase-mediated generation of 5-hydroxymethylcytosine

J Biol Chem. 2013 May 10;288(19):13669-74. doi: 10.1074/jbc.C113.464800. Epub 2013 Apr 2.


Background: Tet methylcytosine dioxygenase converts 5-mC to 5-hmC in DNA.

Results: Ascorbate significantly and specifically enhances Tet-mediated generation of 5-hmC.

Conclusion: Our findings suggest that ascorbate enhances 5-hmC generation, most likely by acting as a co-factor for Tet methylcytosine dioxygenase to generate 5-hmC.

Significance: The availability of ascorbate could have significant consequences for health and diseases by modulating the epigenetic control of genome activity. Ascorbate (vitamin C) is best known for its role in scurvy, in which the hydroxylation of collagen catalyzed by dioxygenases is incomplete due to ascorbate deficiency. Here, we report a novel function of ascorbate in the hydroxylation of 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC) in DNA catalyzed by Tet (ten-eleven translocation) methylcytosine dioxygenase. The content of 5-hmC is extremely low in mouse embryonic fibroblasts cultured in ascorbate-free medium. Additions of ascorbate dose- and time-dependently enhance the generation of 5-hmC, without any effects on the expression of Tet genes. Treatment with another reducer glutathione (GSH) does not change the level of 5-hmC. Further, blocking ascorbate entry into cells by phloretin and knocking down Tet (Tet1, Tet2, and Tet3) expression by short interference RNAs (siRNA) significantly inhibit the effect of ascorbate on 5-hmC. These results suggest that ascorbate enhances 5-hmC generation, most likely by acting as a co-factor for Tet methylcytosine dioxygenase to hydroxylate 5-mC. Thus, we have uncovered a novel role for ascorbate in modulating the epigenetic control of genome activity.

Keywords: 5-Hydroxymethylcytosine; 5-Methylcytosine; DNA Methylation; Enzymes; Epigenetics; Mouse Embryonic Fibroblast; Phloretin; Tet Methylcytosine Dioxygenases; Transport; Vitamin C.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 5-Methylcytosine / analogs & derivatives
  • Animals
  • Ascorbic Acid / pharmacology
  • Ascorbic Acid / physiology*
  • Cells, Cultured
  • Cytosine / analogs & derivatives*
  • Cytosine / biosynthesis
  • DNA Methylation*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Dioxygenases
  • Epigenesis, Genetic
  • Gene Knockdown Techniques
  • Hydroxylation
  • Mice
  • Mice, Inbred C57BL
  • Phloretin / pharmacology
  • Proto-Oncogene Proteins / genetics
  • Proto-Oncogene Proteins / metabolism*
  • RNA, Small Interfering / genetics


  • DNA-Binding Proteins
  • Proto-Oncogene Proteins
  • RNA, Small Interfering
  • TET1 protein, mouse
  • 5-hydroxymethylcytosine
  • 5-Methylcytosine
  • Cytosine
  • Dioxygenases
  • Tet2 protein, mouse
  • Tet3 protein, mouse
  • Ascorbic Acid
  • Phloretin