DNA damage resulting from the oxidation of hydroquinone by copper: role for a Cu(II)/Cu(I) redox cycle and reactive oxygen generation

Abstract

The myelotoxicity, including leukemia, associated with benzene exposure has been attributed to the further activation of benzene-derived metabolites. In a previous study, we observed that (Cu(II) strongly mediates 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 study we investigated whether this chemical--metal redox system induces strand breaks in phi X-174 RFI plasmid DNA. In the presence of micromolar concentrations of Cu(II) and HQ, both single and double strand breaks were induced, whereas HQ, Cu(II), H2O2 or BQ alone at the employed concentrations elicited no significant damage to DNA. The HQ/Cu(II) system was at least twice as efficient as a H2O2/Cu(II) system at inducing DNA strand breaks. Of Cu(II), Fe(III), Mn(II), Cd(II) and Zn(II), only HQ/Cu(II) induced extensive DNA strand breaks. Among HQ, 1,2,4-benzenetriol (BT), catechol and phenol, HQ/Cu(II) and BT/Cu(II) were the two most efficient DNA cleaving systems. The presence of bathocuproinedisulfonic acid (BCS) or catalase prevented the HQ/Cu(II)-induced DNA strand breaks. In addition, the HQ/Cu(II)-induced DNA strand breaks could be completely blocked by reduced glutathione and dithiothreitol, but not by L-cysteine. The interaction of L-cysteine with copper in the absence of HQ induced significant DNA strand breaks with the same pattern of DNA strand breaks as that of HQ/Cu(II) plus L-cysteine. In contrast to the HQ/Cu(II) system, a HQ/myeloperoxidase (MPO)/H2O2 system did not induce any DNA strand breaks, and furthermore, the presence of MPO inhibited the HQ/Cu(II)-induced DNA strand breaks. When DNA pretreated with Cu(II) was exposed to HQ, DNA strand breaks were formed that could be prevented by BCS or catalase, indicating that DNA-bound copper can undergo redox cycling in the presence of HQ, generating H2O2. Similar to the H2O2/Cu(II) system, the HQ/Cu(II)-induced DNA strand breaks could not be efficiently inhibited by hydroxyl radical scavengers but could be protected by singlet oxygen scavengers, indicating that the localized generation of singlet oxygen or a singlet oxygen-like entity, possibly a copper-peroxide complex, rather than free hydroxyl radical probably plays a role in the HQ/Cu(II)-induced DNA strand breaks. The above results suggest that macromolecule-associated copper and reactive oxygen generation may be important factors in the mechanism of HQ-induced DNA damage in target cells.

Oxidative mechanisms in the toxicity of metal ions

“…42 The presence of singlet oxygen scavengers but not hydroxyl radical scavengers provided partial protection, suggesting that singlet oxygen rather than hydroxyl radical may play a role in the induction of DNA strand breaks. However, these scavengers are lacking in absolute specificity, 7 and the involvement of singlet oxygen may be small because there is currently no known mechanism by which it can be formed from oxyradicals, although Steinbeck et al 43 have reported that phagocytosing neutrophils can generate singlet oxygen intracellularly.…”

Effect of lycopene and β-carotene on peroxynitrite-mediated cellular modifications

“…The method is as described by Li and Trush (1993) with some modifications. Briefly, 0.1μg of DNA was incubated with various chemicals in KPB, pH 7.4 at 37 o C for 30 min (final volume 10 μl).…”

Oxidised guanidinohydantoin (Ghox) and spiroiminodihydantoin (Sp) are major products of iron- and copper-mediated 8-oxo-7,8-dihydroguanine and 8-oxo-7,8-dihydro-2′-deoxyguanosine oxidation

“…80 µg of copper, with the total 130 blood concentration of copper about 16 µM. 41 Copper can become available in vivo for Fenton reactions to occur, as it is present in blood plasma as metalloproteins and as a number of transport and storage complexes. Copper also exists, however, in the cell nucleus where it may be involved in the 135 condensation of DNA-histone fibres into higher order chromatin structures.…”