Ascorbic Acid Enhances Tet-Mediated 5‑Methylcytosine Oxidation and Promotes DNA Demethylation in Mammals

• Published in: Journal of the American Chemical Society Vol. 135; no. 28; pp. 10396 - 10403
• Main Authors: Yin, Ruichuan, Mao, Shi-Qing, Zhao, Bailin, Chong, Zechen, Yang, Ying, Zhao, Chao, Zhang, Dapeng, Huang, Hua, Gao, Juan, Li, Zheng, Jiao, Yan, Li, Cuiping, Liu, Shengquan, Wu, Danni, Gu, Weikuan, Yang, Yun-Gui, Xu, Guo-Liang, Wang, Hailin
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 17.07.2013
• Subjects: 5-Methylcytosine - chemistry; 5-Methylcytosine - metabolism; Animals; Ascorbic Acid - chemistry; Ascorbic Acid - metabolism; DNA - chemistry; DNA - metabolism; DNA Methylation; DNA-Binding Proteins - metabolism; Mice; Oxidation-Reduction; Proto-Oncogene Proteins - metabolism
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja4028346

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 Fe2+. 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.