5-methylcytosine (5mC) is an epigenetic DNA modification that represses transcription when placed into CpG-rich promoter sequences. The removal of 5mC from DNA occurs through stepwise oxidation of the methyl group by 5mC oxidases, the so-called Ten-Eleven-Translocation (TET) enzymes. These oxidases sequentially produce 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). Whereas 5hmC occurs at substantial levels in several mammalian cell and tissue types, the other two oxidation products are often described as intermediates in the DNA demethylation pathway, which is completed by base excision repair following excision of 5fC and 5caC by thymine DNA glycosylase (TDG). 5caC is a block to polymerases and may elicit a DNA damage response. Mammalian genomes encode three TET enzymes, TET1, TET2, and TET3. TET3 exists as three major isoforms including an oocyte-specific form, likely responsible for the preferential oxidation of 5mC in the paternal genome of fertilized oocytes, as well as two somatic isoform that differ with regards to presence or absence of an N-terminal zinc finger-type CXXC domain. We recently characterized the full-length TET3 isoform containing the CXXC domain (TET3FL). This CXXC domain binds to unmethylated CpG sequences but, unexpectedly, its highest affinity is towards 5-carboxylcytosine (5caC). We determined the crystal structure of the CXXC domain - 5caC-DNA complex revealing the structural basis of the binding specificity of this domain as a reader of 5CcaCG sequences. We propose that binding of TET3 to its reaction product 5caC through its CXXC domain enforces the genomic specificity of this protein. Mapping of TET3FL in neuronal cell populations showed that Tet3FL is localized at the transcription start sites (TSS) of genes involved in lysosome function, mRNA processing and key genes of the base excision repair pathway. Thus, Tet3FL may function as a regulator of 5caC removal by base excision repair. TET3FL peaks coincided with binding sites for the master regulator of the lysosome and autophagy pathways, TFEB. Active removal of accumulating 5mC from the TSS of genes coding for essential proteins by TET3FL may be important for preventing age-associated diseases including neurodegeneration and cancer.
Citation Format: Gerd Peter Pfeifer. Oxidized 5-methylcytosine: At the interface of epigenetics and DNA repair [abstract]. In: Proceedings of the AACR International Conference: New Frontiers in Cancer Research; 2017 Jan 18-22; Cape Town, South Africa. Philadelphia (PA): AACR; Cancer Res 2017;77(22 Suppl):Abstract nr IA11.