Abstract
Ionizing radiation generates various genomic lesions such as base damage, strand breaks, and DNA-protein crosslinks (DPCs). The formation of base danmage and DNA double-strand breaks (DSBs) is promoted by the presence of oxygen, whereas that of DPCs is inhibited, showing an opposite trend of oxygen dependence. Base damage is repaired by base excision repair, and DSBs are repaired by homologous recombination (HR) and nonhomologous end joining. However, the principal repair mechanism and biological effect of DPCs remain largely elusive. We irradiated transplanted mouse tumors under normoxic and hypoxic conditions and analyzed DSBs and DPCs. The yields of DSBs were greater under normoxic than hypoxic conditions, and those of DPCs were greater under hypoxic than normoxic conditions. It was also suggested that DPCs induced by ionizing radiation are stable relative to those induced by aldehydes. The roles of nucleotide excision repair (NER) and HR in the repair of DPCs were investigated using mammalian cells and their cell-free extracts. The results showed that the maximum size of DPCs amenable to NER was 8 kDa, eliminating its repair role for DPCs. It was also suggested that the replication fork stalled by remaining DPCs is reactivated by HR. In in vitro transcription reactions with T7 RNA polymerase, DPCs in the transcribed, but not nontranscribed, strand strongly inhibited transcription. DPCs significantly reduced the transcriptional fidelity.
