title: Biochemical reconstitution of TET1-TDG-BER-dependent active DNA demethylation reveals a highly coordinated mechanism.
abstract: Cytosine methylation in CpG dinucleotides is an epigenetic DNA modification dynamically established and maintained by DNA methyltransferases and demethylases. Molecular mechanisms of active DNA demethylation began to surface only recently with the discovery of the 5-methylcytosine (5mC)-directed hydroxylase and base excision activities of ten-eleven translocation (TET) proteins and thymine DNA glycosylase (TDG). This implicated a pathway operating through oxidation of 5mC by TET proteins, which generates substrates for TDG-dependent base excision repair (BER) that then replaces 5mC with C. Yet, direct evidence for a productive coupling of TET with BER has never been presented. Here we show that TET1 and TDG physically interact to oxidize and excise 5mC, and proof by biochemical reconstitution that the TET-TDG-BER system is capable of productive DNA demethylation. We show that the mechanism assures a sequential demethylation of symmetrically methylated CpGs, thereby avoiding DNA double-strand break formation but contributing to the mutability of methylated CpGs.
authors.affiliation: Department of Biomedicine, University of Basel, Mattenstrasse 28, Basel CH-4058, Switzerland.
authors.forename: Alain R
authors.initials: AR
authors.lastname: Weber
authors.affiliation: Department of Biomedicine, University of Basel, Mattenstrasse 28, Basel CH-4058, Switzerland.
authors.forename: Claudia
authors.initials: C
authors.lastname: Krawczyk
authors.affiliation: Department of Molecular Microbiology, Oslo University Hospital, Rikshospitalet, NO-0372 Oslo, Norway.
authors.forename: Adam B
authors.initials: AB
authors.lastname: Robertson
authors.affiliation: Proteomics and Metabolomics Core Facility, PROMEC, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology, NO-7489 Trondheim, Norway.
authors.forename: Anna
authors.initials: A
authors.lastname: Kuśnierczyk
authors.affiliation: Proteomics and Metabolomics Core Facility, PROMEC, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology, NO-7489 Trondheim, Norway.
authors.forename: Cathrine B
authors.initials: CB
authors.lastname: Vågbø
authors.affiliation: Department of Biomedicine, University of Basel, Mattenstrasse 28, Basel CH-4058, Switzerland.
authors.forename: David
authors.initials: D
authors.lastname: Schuermann
authors.affiliation: Department of Molecular Microbiology, Oslo University Hospital, Rikshospitalet, NO-0372 Oslo, Norway.
authors.forename: Arne
authors.initials: A
authors.lastname: Klungland
authors.affiliation: Department of Biomedicine, University of Basel, Mattenstrasse 28, Basel CH-4058, Switzerland.
authors.forename: Primo
authors.initials: P
authors.lastname: Schär
mesh_terms: D018899:CpG Islands; D003596:Cytosine / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D019175:DNA Methylation; D004260:DNA Repair / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D044127:Epigenesis, Genetic; D004926:Escherichia coli / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D044502:Thymine DNA Glycosylase / Q000235:genetics / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D011518:Proto-Oncogene Proteins; D003596:Cytosine; D044502:Thymine DNA Glycosylase