Repair pathways for processing of 8-oxoguanine in DNA by mammalian cell extracts

The repair pathways involved in the removal of 8-oxo-7,8-dihydroguanine (8-oxoguanine) in DNA by mammalian cell extracts have been examined. Closed circular DNA constructs containing a single 8-oxoguanine at a defined site were used as substrates to determine the patch size generated after in vitro repair by mammalian cell extracts. Restriction analysis of the repair incorporation in the vicinity of the lesion indicated that up to 75% of the 8-oxoguanine was repaired via the single nucleotide replacement mechanism in both human and mouse cell extracts. Approximately 25% of the 8-oxoguanine lesions were repaired by the long patch repair pathway. Repair incorporation 5' to the lesion, characteristic for nucleotide excision repair, was not significant. Elimination of the DNA polymerase beta (pol beta)-dependent single nucleotide base excision repair pathway in extracts prepared from pol beta-deficient mouse cells resulted in extension of the repair gap to 4-5 nucleotides 3' to the lesion in 50% of the repair events, suggesting the increased involvement of the long patch repair pathway. However, about one-half of the 8-oxoguanine repair was still accomplished through replacement of only one nucleotide in the pol beta-deficient cell extracts. These data indicate the existence of an alternative pol beta-independent single nucleotide repair patch pathway for processing of 8-oxoguanine in DNA.