Induction of direct adducts, apurinic/apyrimidinic sites and oxidized bases in nuclear DNA of human HeLaS3 tumor cells by tetrachlorohydroquinone

DNA damage induced by tetrachlorohydroquinone (Cl(4)HQ), the quinonoid meta bolite of pentachlorophenol (PCP), was investigated in human HeLa S3 tumor cells. Formation of one major and two minor DNA adducts in cells treated wi th Cl(4)HQ (50-300 muM) was detected by P-32-post-labeling assay and the ad ducts accumulated over the course of the experiment (0.5-2 h), with total a dduct levels estimated to be 3-6 per 10(8) nucleotides. These adducts did n ot correspond to those derived from calf thymus DNA treated with tetrachlor o-1,4-benzoquinone. Results from the apurinic/apyrimidinic (AP) sites assay indicated that the number of AP sites was 2-fold greater in cells exposed to Cl(4)HQ (300 muM) than the corresponding control. Further characterizati on of the AP sites confirmed that Cl(4)HQ induced predominantly (75%) putre scine-excisable AP sites in HeLa S3 cells. In parallel, the concentration o f 8-hydroxy-2 ' -deoxyguanosine (8-HO-dG) in cells treated with Cl(4)HQ for 0.5 and 2 h was increased 2- and 5-fold, respectively, compared with the c ontrol. The extent of oxidative DNA damage induced by Cl(4)HQ was approxima tely two orders of magnitude greater than those of direct DNA adducts, Over all, it appears that reactive oxygen species mediate the parallel formation of AP sites and 8-HO-dG in HeLa S3 cells following treatment with Cl(4)HQ and that the contribution of depurination/depyrimidination of direct DNA ad ducts is relatively insignificant compared with the formation of oxidized A P sites. We conclude that putrescine-excisable AP sites represent a major t ype of ROS-mediated oxidative DNA damage in cellular DNA induced by Cl(4)HQ and may play a role in PCP-induced clastogenicity in mammalian cells.