DNA-DAMAGE RESULTING FROM THE OXIDATION OF HYDROQUINONE BY COPPER - ROLE FOR A CU(II) CU(I) REDOX CYCLE AND REACTIVE OXYGEN GENERATION/

The myelotoxicity, including leukemia, associated with benzene exposur e has been attributed to the further activation of benzene-derived met abolites. In a previous study, we observed that Cu(II) strongly mediat es the oxidation of hydroquinone (HQ) producing benzoquinone (BQ) and H2O2 through Cu(II)/Cu(I) redox mechanism. Since copper exists in the nucleus and is closely associated with chromosomes and DNA, in this st udy we investigated whether this chemical-metal redox system induces s trand breaks in phiX-174 RFI plasmid DNA. In the presence of micromola r concentrations of Cu(II) and HQ, both single and double strand break s were induced, whereas HQ, Cu(II), H2O2 or BQ alone at the employed c oncentrations elicited no significant damage to DNA. The HQ/Cu(II) sys tem was at least twice as efficient as a H2O2/Cu(II) system at inducin g DNA strand breaks. Of Cu(II), Fe(III), Mn(II), Cd(II) and Zn(II), on ly HQ/Cu(II) induced extensive DNA strand breaks. Among HQ, 1,2,4-benz enetriol (BT), catechol and phenol, HQ/Cu(II) and BT/Cu(II) were the t wo most efficient DNA cleaving systems. The presence of bathocuproined isulfonic acid (BCS) or catalase prevented the HQ/Cu(II)-induced DNA s trand breaks. In addition, the HQ/Cu(II)-induced DNA strand breaks cou ld be completely blocked by reduced glutathione and dithiothreitol, bu t not by L-cysteine. The interaction of L-cysteine with copper in the absence of HQ induced significant DNA strand breaks with the same patt ern of DNA strand breaks as that of HQ/Cu(II) plus L-cysteine. In cont rast to the HQ/Cu(II) system, a HQ/myeloperoxidase (MPO)/H2O2 system d id not induce any DNA strand breaks, and furthermore, the presence of MPO inhibited the HQ/Cu(Il)-induced DNA strand breaks. When DNA pretre ated with Cu(II) was exposed to HQ, DNA strand breaks were formed that could be prevented by BCS or catalase, indicating that DNA-bound copp er can undergo redox cycling in the presence of HQ, generating H2O2. S imilar to the H2O2/Cu(II) system, the HQ/Cu(II)-induced DNA strand bre aks could not be efficiently inhibited by hydroxyl radical scavengers but could be protected by singlet oxygen scavengers, indicating that t he localized generation of singlet oxygen or a singlet oxygen-like ent ity, possibly a copper - peroxide complex, rather than free hydroxyl r adical probably plays a role in the HQ/Cu(II)-induced DNA strand break s. The above results suggest that macromolecule-associated copper and reactive oxygen generation may be important factors in the mechanism o f HQ-induced DNA damage in target cells.