Methylation at position 5 of cytosine (5-methylcytosine, 5mC) in DNA is a major epigenetic modification that regulates gene transcription and other functions of the genome. Active demethylation is required for a rapid and efficient erasure of 5mC. Until now, mechanisms of active DNA demethylation have been intensively studied but remain elusive. Growth arrest and DNA-damage-inducible protein 45 (Gadd45 ) family proteins have been reported to participate in active DNA demethylation. However, the mechanism of how Gadd45 proteins promote DNA demethylation has remained controversial and is still under investigation.
A research team led by Prof. XU Guoliang at the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences of Chinese Academy of Sciences, demonstrated that Gadd45a promotes active DNA demethylation through thymine DNA glycosylase (TDG). Previous work by XU group (Science. 2011,333:1303-1307) and other groups has shown that TDG can excise 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) generated in Ten-eleven-translocation (Tet)––initiated oxidative demethylation.
Their study showed that a methylated reporter gene is activated in HEK293T cells expressing Gadd45a in combination with catalytically active TDG and Tet. Gadd45a interacts with TDG physically and increases the removal of 5fC and 5caC from genomic and transfected plasmid DNA by TDG. Knockout of both Gadd45a and Gadd45b from mouse ES cells leads to hypermethylation of specific genomic loci most of which are also targets of TDG and show 5fC enrichment in TDG-deficient cells. These findings connect Gadd45 proteins with the Tet-TDG axis, functionally integrating the seemingly diverse demethylation pathways.
A paper about this work entitled “Gadd45a promotes DNA demethylation through TDG” was published online in Nucleic Acids Research on April 6th, 2015. This study was done with assistance from Prof. Primo Schär, at the University of Basel, Switzerland, Prof. Tang Fuchou, at the Biodynamic Optical Imaging Center, Peking University, and Prof. Wang Hailin, at the Research Center for Eco-Environmental Sciences, CAS. It was supported by the Breakthrough Project of Strategic Priority Program of Chinese Academy of Sciences, the National Science Foundation of China and the National Science & Technology Major Project ‘Key New Drug Creation and Manufacturing Program’ of China.
AUTHOR CONTACT:
XU Guoliang, Principal Investigator
Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences
Shanghai 200031, China.
Phone: +86-21-54921332
E-mail: glxu@sibcb.ac.cn
Fig. Gadd45a promotes reduction of 5caC through TDG. (A) HPLC analysis of 5caC in the genomic DNA of HEK293T cells overexpressing full-length Tet2 with and without Gadd45a. The y-axis indicates OD280nm of nucleoside preparations from genomic DNA. Defined nucleosides were used as standards (C, 5mC, 5hmC and 5caC). (B) MAB-seq analysis of oxidized reporter plasmid DNA recovered from transfected HEK293T cells. The oxi-5mC plasmid DNA was transfected into WT or TDG knockout cells with Gadd45a, TDG or vector control. Percentages of the oxi-5mC bases (5fC and 5caC) are indicated.
(Image provided by Prof. XU Guoliang’s group)
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