title: Formation of DNA-protein cross-links in cultured mammalian cells upon treatment with iron ions.
abstract: Formation of DNA-protein crosslinks (DPCs) in mammalian cells upon treatment with iron or copper ions was investigated. Cultured murine hybridoma cells were treated with Fe(II) or Cu(II) ions by addition to the culture medium at various concentrations. Subsequently, chromatin samples were isolated from treated and control cells. Analyses of chromatin samples by gas chromatography/mass spectrometry after hydrolysis and derivatization revealed a significant increase over the background amount of 3-[(1,3-dihydrio-2,4-dioxopyrimidin-5-yl)-methyl]- L-tyrosine (Thy-Tyr crosslink) in cells treated with Fe(II) ions in the concentration range of 0.01 to 1 mM. In contrast, Cu(II) ions at the same concentrations did not produce this DPC in cells. No DNA base damage was observed in cells treated with Cu(II) ions, either. Preincubation of cells with ascorbic acid or coincubation with dimethyl sulfoxide did not significantly alleviate the Fe(II) ion-mediated formation of DPCs. In addition, a modified fluorometric analysis of DNA unwinding assay was used to detect DPCs formed in cells. Fe(II) ions caused significant formation of DPCs, but Cu(II) ions did not. The nature of the Fe(II)-mediated DPCs suggests the involvement of the hydroxyl radical in their formation. The Thy-Tyr crosslink may contribute to pathological processes associated with free radical reactions.
authors.affiliation: Department of Chemical and Biochemical Engineering, University of Maryland Baltimore County, USA.
authors.forename: S A
authors.initials: SA
authors.lastname: Altman
authors.forename: T H
authors.initials: TH
authors.lastname: Zastawny
authors.forename: L
authors.initials: L
authors.lastname: Randers-Eichhorn
authors.forename: M A
authors.initials: MA
authors.lastname: Cacciuttolo
authors.forename: S A
authors.initials: SA
authors.lastname: Akman
authors.forename: M
authors.initials: M
authors.lastname: Dizdaroglu
authors.forename: G
authors.initials: G
authors.lastname: Rao
mesh_terms: D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology; D002460:Cell Line; D002843:Chromatin / Q000737:chemistry; D003300:Copper / Q000494:pharmacology; D003432:Cross-Linking Reagents; D003470:Culture Media; D004247:DNA / Q000378:metabolism*; D004249:DNA Damage / Q000187:drug effects; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D005296:Ferrous Compounds / Q000494:pharmacology; D005609:Free Radicals; D008401:Gas Chromatography-Mass Spectrometry; D006825:Hybridomas; D006861:Hydrogen Peroxide / Q000494:pharmacology; D051379:Mice; D011506:Proteins / Q000378:metabolism*; D013941:Thymine / Q000737:chemistry; D014443:Tyrosine / Q000737:chemistry
chemical_list: D002843:Chromatin; D003432:Cross-Linking Reagents; D003470:Culture Media; D005296:Ferrous Compounds; D005609:Free Radicals; D011506:Proteins; D014443:Tyrosine; D003300:Copper; D004247:DNA; D006861:Hydrogen Peroxide; D001205:Ascorbic Acid; D013941:Thymine; D004121:Dimethyl Sulfoxide