Cell 146, 67–79 (2011)

Thymine DNA glycosylase (TDG) is known to be a base excision repair protein involved in the removal of G·T mismatches, but recent studies have identified TDG as a regulator of transcription and DNA methylation. Cortellino et al. now clarify these roles by showing that TDG helps maintain a proper methylation state at CpG islands and is a component of a DNA demethylation-repair pathway that excises modified cytosine bases—5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC)—from genomic DNA. Using transgenic mouse technology and knockout-derived cell lines, the authors showed that Tdg null mutations produce embryonic lethality and developmental defects similar to retinoic acid signaling deficiencies. In both a transcriptional reporter system and developing embryos, TDG was shown to have an active role in demethylation of developmentally regulated genes, and an inactivating mutation at the glycosylase active site reproduced the embryonic lethality. Coimmunoprecipitation experiments revealed that TDG is associated with AID—a cytosine deaminase recently found to be involved in the conversion of 5hmC to 5-hydroxymethyluracil (5hmU) in genomic DNA—and GADD45, a stress response protein associated with demethylation. TDG's role in demethylation was dependent on its glycosylase activity, but in vitro assays showed that TDG removes T and 5hmU, but not 5mC or 5hmC, from DNA substrates. Taken together, these data argue that TDG functions as a component of a demethylation complex that removes modified cytosine bases by a sequential deamination and base excision repair pathway.