Abstract
Clustered DNA damage, defined as two or more lesions within one to two helical turns of DNA induced by a single radiation track, is a unique feature of ionizing radiation. We have studied the biological consequences of bi-stranded clustered damage sites which consist of a combination of DNA lesions, such as a single strand break (SSB), an apurinic/apyrimidinic (AP) site, and an 8-oxo-7,8-dihydroguanine (8-oxoG), using a bacterial plasmid-based assay. We found significantly lower transformation frequencies for the clustered SSB + AP lesions than that for either a single SSB or a single AP site. We suggest that a double strand break (DSB) or a replication block is formed during the processing of the SSB + AP cluster. When the two lesions are placed farther apart (10-20bp), the transformation efficiencies are comparable to those of the single lesions. This recovery of transformation efficiency for separated lesions requires PolI activity. Similarly, the mutation frequency depends on the separation of the clustered SSB + 8-oxoG, although the SSB + 8-oxoG cluster, in contrast to the SSB + AP cluster, transforms at a comparable efficiency to the efficiencies of single lesions. PolI also seems to play an important role in avoiding mutations, as the lack of PolI enhances the mutation frequency of the separated lesions to the level of that of the closely spaced lesions. These results indicate that the biological consequences of clustered DNA damage strongly depend on the repair synthesis of the comprised lesion(s).
