The Mutation frequency of 8-oxo-7,8-dihydroguanine when single strand break is closely present on the same or the opposite strand

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

Ionizing radiation induces clustered DNA damage which is defined as two or more lesions induced within 1-2 helical turns (10-20bp) of DNA by a single radiation track. Recent in vitro studies predict that the non-DSB clustered damage shows high biological effects. It is, however, technically difficult to directly detect non-DSB clustered damage site as well as its effect in living cells. In this study, we investigated the potential of single strand break (SSB) to influence the mutagenicity of base lesions in Escherichia coli. We used plasmid based assay to measure the mutation frequency induced by bistranded clustered damage. As models of clustered damage, we used synthesized oligonucleotides carrying a SSB and 8-oxo-7,8-dihydroguanine (8-oxoG) at a restriction enzyme recognition site. Damaged DNA was transfected into wild-type or glycosylase-deficient strains (fpg, mutY, fpg mutY) of E coli and mutation frequency was assessed by the inability to cut by the restriction enzyme. Clustered damage containing 8-oxoG and SSB located on opposite strand raised mutation frequencies. However, clustered damage containing 8-oxoG and SSB positioned in tandem gave lower mutation frequency than single 8-oxoG lesion. We propose that the mutagenic potential of 8-oxoG depends on whether SSB is located on either strand, same or opposite, to 8-oxoG.