Genotoxicity and mutagenicity of chromium(VI)/ascorbate-generated DNA adducts in human and bacterial cells.
Reduction of carcinogenic Cr(VI) by vitamin C generates ascorbate-Cr(III)-DNA cross-links, binary Cr(III)-DNA adducts, and can potentially cause oxidative DNA damage by intermediate reaction products. Here, we examined the mutational spectrum and the importance of different forms of DNA damage in genotoxicity and mutagenicity of Cr(VI) activated by physiological concentrations of ascorbate. Reduction of Cr(VI) led to a dose-dependent formation of both mutagenic and replication-blocking DNA lesions as detected by propagation of the pSP189 plasmids in human fibroblasts. Disruption of Cr-DNA binding abolished mutagenic responses and normalized the yield of replicated plasmids, indicating that Cr-DNA adducts were responsible for both mutagenicity and genotoxicity of Cr(VI). The absence of DNA breaks and abasic sites confirmed the lack of a significant production of hydroxyl radicals and Cr(V)-peroxo complexes in Cr(VI)-ascorbate reactions. Ascorbate-Cr(III)-DNA cross-links were much more mutagenic than smaller Cr(III)-DNA adducts and accounted for more than 90% of Cr(VI) mutagenicity. Ternary adducts were also several times more potent in the inhibition of replication than binary complexes. The Cr(VI)-induced mutational spectrum consisted of an approximately equal number of deletions and G/C-targeted point mutations (51% G/C --> T/A and 30% G/C --> A/T). In Escherichia coli cells, Cr(VI)-induced DNA adducts were only highly genotoxic but not mutagenic under either normal or SOS-induced conditions. Lower toxicity and high mutagenicity of ascorbate-Cr(III)-DNA adducts in human cells may result from the recruitment of an error-prone bypass DNA polymerase(s) to the stalled replication forks. Our results suggest that phosphotriester-type DNA adducts could play a more important role in human than bacterial mutagenesis.
Quievryn
George
G
Brown University, Department of Pathology and Laboratory Medicine, Providence, Rhode Island 02912, USA.
Peterson
Elizabeth
E
Messer
Joseph
J
Zhitkovich
Anatoly
A
D001205:Ascorbic Acid / Q000737:chemistry / Q000633:toxicity*; D001483:Base Sequence; D002461:Cell Line, Transformed; D002857:Chromium / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity*; D018736:DNA Adducts / Q000737:chemistry / Q000633:toxicity*; D004249:DNA Damage / Q000235:genetics; D004252:DNA Mutational Analysis; D004261:DNA Replication / Q000187:drug effects / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000187:drug effects* / Q000378:metabolism*; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism*; D016153:Genes, Suppressor; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009153:Mutagens / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity*; D010084:Oxidation-Reduction; D012343:RNA, Transfer / Q000235:genetics
D018736:DNA Adducts; D009153:Mutagens; C096275:supF tRNA; D002857:Chromium; C074702:chromium hexavalent ion; D012343:RNA, Transfer; D001205:Ascorbic Acid