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 ci rcular DNA constructs containing a single 8-oxoguanine at a defined site we re used as substrates to determine the patch size generated after in vitro repair by mammalian cell extracts. Restriction analysis of the repair incor poration in the vicinity of the lesion indicated that up to 75% of the 8-ox oguanine was repaired via the single nucleotide replacement mechanism in bo th human and mouse cell extracts. Approximately 25% of the 8-oxoguanine les ions were repaired by the long patch repair pathway. Repair incorporation 5 ' to the lesion, characteristic for nucleotide excision repair, was not sig nificant. Elimination of the DNA polymerase beta (pol beta)-dependent singl e nucleotide base excision repair pathway in extracts prepared from pol bet a-deficient mouse cells resulted in extension of the repair gap to 4-5 nucl eotides 3' to the lesion in 50% of the repair events, suggesting the increa sed involvement of the long patch repair pathway. However, about one-half o f the 8-oxoguanine repair was still accomplished through replacement of onl y one nucleotide in the pol beta-deficient cell extracts. These data indica te the existence of an alternative pol beta-independent single nucleotide r epair patch pathway for processing of 8-oxoguanine in DNA.