Methylation and hydroxymethylation of cytosine alter activity and fidelity of translesion DNA polymerases

• Published in: DNA repair 2024-09, Vol.141, p.103712, Article 103712
• Main Authors: Shilkin, Evgeniy S., Petrova, Daria V., Novikova, Anna A., Boldinova, Elizaveta O., Zharkov, Dmitry O., Makarova, Alena V.
• Format: Article
• Language: English
• Subjects: 5-hydroxymethylcytosine; 5-methylcytosine; 5-Methylcytosine - analogs & derivatives; 5-Methylcytosine - metabolism; 8-Hydroxy-2'-Deoxyguanosine - metabolism; 8-oxo-2′-deoxyguanosine; CpG dinucleotides; Cytosine - analogs & derivatives; Cytosine - metabolism; DNA - metabolism; DNA Damage; DNA Methylation; DNA Polymerase iota; DNA polymerases; DNA Repair; DNA Replication; DNA-Directed DNA Polymerase - metabolism; Humans; Multifunctional Enzymes - metabolism; Nucleotidyltransferases - genetics; Nucleotidyltransferases - metabolism
• ISSN: 1568-7864; 1568-7856; 1568-7856
• DOI: 10.1016/j.dnarep.2024.103712

Epigenetic cytosine methylation covers most of genomic CpG dinucleotides in human cells. In addition to common deamination-mediated mutagenesis at CpG sites, an alternative deamination-independent pathway associated with DNA polymerase activity was previously described. This mutagenesis is characterized by the TCG→TTG mutational signature and is believed to arise from dAMP misincorporation opposite 5-methylcytosine (mC) or its oxidized derivative 5-hydroxymethylcytosine (hmC) by B-family replicative DNA polymerases with disrupted proofreading 3→5′-exonuclease activity. In addition to being less stable and pro-mutagenic themselves, cytosine modifications also increase the risk of adjacent nucleotides damage, including the formation of 8-oxo-2'-deoxyguanosine (8-oxoG), a well-known mutagenic lesion. The effect of cytosine methylation on error-prone DNA polymerases lacking proofreading activity and involved in repair and DNA translesion synthesis remains unexplored. Here we analyze the efficiency and fidelity of translesion Y-family polymerases (Pol κ, Pol η, Pol ι and REV1) and primase-polymerase PrimPol opposite mC and hmC as well as opposite 8-oxoG adjacent to mC in the TCG context. We demonstrate that epigenetic cytosine modifications suppress Pol ι and REV1 activities and lead to increasing dAMP misincorporation by PrimPol, Pol κ and Pol ι in vitro. Cytosine methylation also increases misincorporation of dAMP opposite the adjacent 8-oxoG by PrimPol, decreases the TLS activity of Pol η opposite the lesion but increases dCMP incorporation opposite 8-oxoG by REV1. Altogether, these data suggest that methylation and hydroxymethylation of cytosine alter activity and fidelity of translesion DNA polymerases. [Display omitted] •Methylation and hydroxymethylation of cytosine suppress Pol ι and REV1 activities.•Epigenetic cytosine modifications lead to increasing dAMP misincirporation by PrimPol, Pol κ and Pol ι.•Cytosine methylation alters the translesion activities of PrimPol, Pol η and REV1 opposite 8-oxoG adjacent to mC.