5'-nicked apurinic/apyrimidinic sites are resistant to beta-elimination by beta-polymerase and are persistent in human cultured cells after oxidative stress.
Genomic DNA is continuously exposed to oxidative stress. Whereas reactive oxygen species (ROS) preferentially react with bases in DNA, free radicals also abstract hydrogen atoms from deoxyribose, resulting in the formation of apurinic/apyrimidinic (AP) sites and strand breaks. We recently reported high steady-state levels of AP sites in rat tissues and human liver DNA (Nakamura, J., and Swenberg, J. A. (1999) Cancer Res. 59, 2522-2526). These AP sites were predominantly cleaved 5' to the lesion. We hypothesized that these endogenous AP sites were derived from oxidative stress. In this investigation, AP sites induced by ROS were quantitated and characterized. A combination of H(2)O(2) and FeSO(4) induced significant numbers of AP sites in calf thymus DNA, which were predominantly cleaved 5' to the AP sites (75% of total aldehydic AP sites). An increase in the number of 5'-AP sites was also detected in human cultured cells exposed to H(2)O(2), and these 5'-AP sites were persistent during the post-exposure period. beta-Elimination by DNA beta-polymerase efficiently excised 5'-regular AP sites, but not 5'-AP sites, in DNA from cells exposed to H(2)O(2). These results suggest that 5'-oxidized AP sites induced by ROS are not efficiently repaired by the mammalian short patch base excision repair pathway.
Nakamura
J
J
Department of Environmental Sciences, University of North Carolina, Chapel Hill, North Carolina 27599, USA.
La
D K
DK
Swenberg
J A
JA
D000080242:8-Hydroxy-2'-Deoxyguanosine / Q000031:analogs & derivatives; D000818:Animals; D002417:Cattle; D004247:DNA / Q000187:drug effects; D004249:DNA Damage; D019951:DNA Polymerase beta / Q000378:metabolism*; D004260:DNA Repair; D043224:Deoxyribonuclease (Pyrimidine Dimer); D004305:Dose-Response Relationship, Drug; D004706:Endodeoxyribonucleases / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D006147:Guanine / Q000031:analogs & derivatives / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D017665:Hydroxyl Radical / Q000378:metabolism; D018384:Oxidative Stress; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013950:Thymus Gland / Q000187:drug effects; D013997:Time Factors
D029968:Escherichia coli Proteins; D017382:Reactive Oxygen Species; D017665:Hydroxyl Radical; D006147:Guanine; C078307:8-oxo-7,8-dihydrodeoxyguanine; D000080242:8-Hydroxy-2'-Deoxyguanosine; D004247:DNA; D006861:Hydrogen Peroxide; D019951:DNA Polymerase beta; D004706:Endodeoxyribonucleases; D043224:Deoxyribonuclease (Pyrimidine Dimer); C106403:NTH protein, E coli