Sequence specificity of Cr(III)–DNA adduct formation in the p53 gene: NGG sequences are preferential adduct-forming sites

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Abstract
Hexavalent chromium [Cr(VI)] is a known etiological factor in human lung cancer. Cr(VI) exposure-related lung cancer has a high mutation incidence in the p53 gene. Upon intake in human cells Cr(VI) is reduced to Cr(III), which is able to conjugate with amino acids and consequently form either binary Cr(III)–DNA or ternary Cr(III)-amino acid–DNA adducts. Both binary and ternary Cr(III)–DNA adducts are mutagenic. We have found that the Escherichia coli nucleotide excision enzyme UvrABC nuclease is able to incise Cr(III)- and Cr(III)-histidine-modified plasmid DNA and the extent of incision is proportional to the amount of Cr(III)–DNA adducts in the plasmid. In order to determine the role of Cr(III)–DNA adducts in the mutagenesis of the p53 gene in human cancer using the UvrABC nuclease incision method, we have mapped the Cr(III)–DNA distribution in PCR DNA fragments amplified from exons 5, 7 and 8 of the p53 gene. We have found that the sequence specificities of Cr(III)–DNA and Cr(III)-histidine–DNA adducts in the p53 gene sequence are identical and that both types of adducts are preferentially formed at –NGG- sequences, including codons 245, 248 and 249, the mutational hotspots in human lung cancer. It has been found that Cr(III)–DNA adducts induce mainly G to T mutations. Therefore, these results suggest that Cr(III)–DNA adduct formation contributes to the p53 gene mutations in lung carcinogenesis.