TY - JOUR
T1 - Maintenance DNA Methyltransferase Activity in the Presence of Oxidized Forms of 5-Methylcytosine
T2 - Structural Basis for Ten Eleven Translocation-Mediated DNA Demethylation
AU - Seiler, Christopher L.
AU - Fernandez, Jenna
AU - Koerperich, Zoe
AU - Andersen, Molly P.
AU - Kotandeniya, Delshanee
AU - Nguyen, Megin E.
AU - Sham, Yuk Y
AU - Tretyakova, Natalia Y
N1 - Funding Information:
Natalia Y. Tretyakova: 0000-0002-0621-6860 Author Contributions C.L.S. and J.F. contributed equally to this work. Funding This work was supported by National Cancer Institute Grant 2R01 CA-095039. Notes The authors declare no competing financial interest.
Publisher Copyright:
Copyright © 2018 American Chemical Society.
PY - 2018/10/23
Y1 - 2018/10/23
N2 - A precise balance of DNA methylation and demethylation is required for epigenetic control of cell identity, development, and growth. DNA methylation marks are introduced by de novo DNA methyltransferases DNMT3a/b and are maintained throughout cell divisions by DNA methyltransferase 1 (DNMT1), which adds methyl groups to hemimethylated CpG dinucleotides generated during DNA replication. Ten eleven translocation (TET) dioxygenases oxidize 5-methylcytosine (mC) to 5-hydroxymethylcytosine (hmC), 5-formylcytosine (fC), and 5-carboxylcytosine (caC), a process known to induce DNA demethylation and gene reactivation. In this study, we investigated the catalytic activity of human DNMT1 in the presence of oxidized forms of mC. A mass spectrometry-based assay was employed to study the kinetics of DNMT1-mediated cytosine methylation in CG dinucleotides containing C, mC, hmC, fC, or caC across from the target cytosine. Homology modeling, coupled with molecular dynamics simulations, was used to explore the structural consequences of mC oxidation with regard to the geometry of protein-DNA complexes. The DNMT1 enzymatic activity was strongly affected by the oxidation status of mC, with the catalytic efficiency decreasing in the following order: mC > hmC > fC > caC. Molecular dynamics simulations revealed that DNMT1 forms an unproductive complex with DNA duplexes containing oxidized forms of mC as a consequence of altered interactions of the target recognition domain of the protein with the C-5 substituent on cytosine. Our results provide new structural and mechanistic insight into TET-mediated DNA demethylation.
AB - A precise balance of DNA methylation and demethylation is required for epigenetic control of cell identity, development, and growth. DNA methylation marks are introduced by de novo DNA methyltransferases DNMT3a/b and are maintained throughout cell divisions by DNA methyltransferase 1 (DNMT1), which adds methyl groups to hemimethylated CpG dinucleotides generated during DNA replication. Ten eleven translocation (TET) dioxygenases oxidize 5-methylcytosine (mC) to 5-hydroxymethylcytosine (hmC), 5-formylcytosine (fC), and 5-carboxylcytosine (caC), a process known to induce DNA demethylation and gene reactivation. In this study, we investigated the catalytic activity of human DNMT1 in the presence of oxidized forms of mC. A mass spectrometry-based assay was employed to study the kinetics of DNMT1-mediated cytosine methylation in CG dinucleotides containing C, mC, hmC, fC, or caC across from the target cytosine. Homology modeling, coupled with molecular dynamics simulations, was used to explore the structural consequences of mC oxidation with regard to the geometry of protein-DNA complexes. The DNMT1 enzymatic activity was strongly affected by the oxidation status of mC, with the catalytic efficiency decreasing in the following order: mC > hmC > fC > caC. Molecular dynamics simulations revealed that DNMT1 forms an unproductive complex with DNA duplexes containing oxidized forms of mC as a consequence of altered interactions of the target recognition domain of the protein with the C-5 substituent on cytosine. Our results provide new structural and mechanistic insight into TET-mediated DNA demethylation.
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U2 - 10.1021/acs.biochem.8b00683
DO - 10.1021/acs.biochem.8b00683
M3 - Article
C2 - 30230311
AN - SCOPUS:85054702354
SN - 0006-2960
VL - 57
SP - 6061
EP - 6069
JO - Biochemistry
JF - Biochemistry
IS - 42
ER -