In vivo formation of single-strand breaks in DNA by hydrogen peroxide is mediated by the Haber-Weiss reaction

Abstract

Phenanthroline and bipyridine, strong chelators of iron, protect DNA from single-strand break formation by H 2O 2 in human fibroblasts. This fact strongly supports the concept that these DNA single-strand breaks are produced by hydroxyl radicals generated by a Fenton-like reaction between intracellular Fe 2+ and H 2O 2: H 2O 2 + Fe 2+ → Fe 3+ + OH − + OH ·. Corroborating this idea is the fact that thiourea, an effective OH radical scavenger, prevents the formation of DNA single-strand breaks by H 2O 2 in nuclei from human fibroblasts. The copper chelator diethyldithiocarbamate, a strong inhibitor of superoxide dismutase, greatly enhances the in vivo production of DNA single-strand breaks by H 2O in fibroblasts. This supports the idea that Fe 3+ is reduced to Fe 2+ by superoxide ion: O ⨪ 2 + Fe 3+ → O 2 + Fe 2+ ; and therefore that the sum of this reaction and the Fenton reaction, namely the so-called Haber-Weiss reaction, H 2 O 2 + O ⨪ 2 → O 2 + OH − + OH · , represents the mode whereby OH radical is produced from H 2O 2 in the cell. EDTA completely protects DNA from single-strand break formation in nuclei. The chelator therefore removes iron from the chromatin, and although the Fe-EDTA complex formed is capable of reacting with H 2O 2, the OH radical generated under these conditions is not close enough to hit DNA. Therefore iron complexed to chromatin functions as catalyst for the Haber-Weiss reaction in vivo, similarly to the role played by Fe-chelates in vitro.