Ascorbic acid enhances Tet-mediated 5-methylcytosine oxidation and promotes DNA demethylation in mammals

J Am Chem Soc. 2013 Jul 17;135(28):10396-403. doi: 10.1021/ja4028346. Epub 2013 Jul 1.


DNA hydroxymethylation and its mediated DNA demethylation are critical for multiple cellular processes, for example, nuclear reprogramming, embryonic development, and many diseases. Here, we demonstrate that a vital nutrient ascorbic acid (AA), or vitamin C (Vc), can directly enhance the catalytic activity of Tet dioxygenases for the oxidation of 5-methylcytosine (5mC). As evidenced by changes in intrinsic fluorescence and catalytic activity of Tet2 protein caused by AA and its oxidation-resistant derivatives, we further show that AA can uniquely interact with the C-terminal catalytic domain of Tet enzymes, which probably promotes their folding and/or recycling of the cofactor Fe(2+). Other strong reducing chemicals do not have a similar effect. These results suggest that AA also acts as a cofactor of Tet enzymes. In mouse embryonic stem cells, AA significantly increases the levels of all 5mC oxidation products, particularly 5-formylcytosine and 5-carboxylcytosine (by more than an order of magnitude), leading to a global loss of 5mC (∼40%). In cells deleted of the Tet1 and Tet2 genes, AA alters neither 5mC oxidation nor the overall level of 5mC. The AA effects are however restored when Tet2 is re-expressed in the Tet-deficient cells. The enhancing effects of AA on 5mC oxidation and DNA demethylation are also observed in a mouse model deficient in AA synthesis. Our data establish a direct link among AA, Tet, and DNA methylation, thus revealing a role of AA in the regulation of DNA modifications.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 5-Methylcytosine / chemistry
  • 5-Methylcytosine / metabolism*
  • Animals
  • Ascorbic Acid / chemistry
  • Ascorbic Acid / metabolism*
  • DNA / chemistry
  • DNA / metabolism*
  • DNA Methylation
  • DNA-Binding Proteins / metabolism*
  • Dioxygenases
  • Mice
  • Oxidation-Reduction
  • Proto-Oncogene Proteins / metabolism*


  • DNA-Binding Proteins
  • Proto-Oncogene Proteins
  • TET1 protein, mouse
  • 5-Methylcytosine
  • DNA
  • Dioxygenases
  • Tet2 protein, mouse
  • Ascorbic Acid