Bistranded clustered damage generated by ionizing radiation (IR), which contains two or more closely opposed base lesions and/or single-strand breaks (SSBs), is assumed to result in error-prone repair or double-strand breaks (DSBs), and hence leads to adverse biological effects. Although the formation and repair mechanisms of direct DSBs by IR, a special form of clustered damage, have been studied extensively, those of clustered base damage (CBD) containing base lesions or base lesions+SSB are less well understood. In the present study, we have quantified the yield of CBD and assessed the repair efficiency of CBD by base excision repair enzyme.
pDEL19 plasmid DNA was irradiated in aqueous solution by gamma-rays, carbon and iron ion beams, treated with Endo III or Fpg, and subjected to agarose gel electrophoresis. The yields of isolated (base damage and SSBs) and clustered (DSBs and CBD) damage were determined from conformational changes of plasmid DNA. The fraction of clustered damage was 2-3% of total damage and independent of the type of radiation. The ratio of DSBs and CBD was 1:1, indicating that CBD accounts for half of clustered damage.
hOGG1 is a major human DNA glycosylase for oxidative purine lesions generated by IR and oxidizing agents. The damage-excising activity of hOGG1 for CBD containing two closely opposed 8-oxoG lesions was analyzed. hOGG1 excised 8-oxoG and induced DSBs even if two 8-oxoG lesions are less than 5 nucleotides apart. However, the efficiency of DSB formation varied dramatically depending on the configuration of 8-oxoG lesions.