Single strand break affects mutagenic potential of clustered DNA damage containing 8-oxoG

Noguchi, Miho; Urushibara, Ayumi*; Yokoya, Akinari; Shikazono, Naoya

Isolated DNA damages such as single strand break (SSB) and base damage are restored efficiently in cells. It is possible that the non-DSB clustered damage including SSB or base damage contribute to high biological effects, such as mutation. In this study, we investigated the mutagenicity of clustered DNA damage containing SSB and base damage in Escherichia coli. We used plasmid based assay in E. coli to measure the mutation frequency induced by bistranded clustered damage. As a model of clustered damage, we used synthesized oligonucleotides carrying an SSB and/or 8-oxoGs at a restriction enzyme recognition site. Damaged DNA was transfected into E coli and mutation frequency was assessed by the inability to cut by the restriction enzyme. The mutation frequency of clustered damage carrying a 8-oxoG opposite to a second 8-oxoG was the highest in all types of clustered damage tested. However, when a SSB was added to bistranded 8-oxoGs, the mutation frequency became lower in every E. coli strain. These results suggest that SSB located on the same strand to 8-oxoG reduces mutagenic potential of 8-oxoG and that the repair of the SSB or replication is involved in the process. Our studies demonstrate that the mutagenic potential of clustered damage including 8-oxoG depends on existence and position of SSB in clustered damage.