Reactivities of chemical substances with human protooncogene in application to screening of carcinogenicity

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The reactivities of carcinogen with DNA in the presence of metal ions were investigated by a DNA sequencing technique using ^<32>P-labeled DNA fragments obtained from human c-ha-ras-1 protooncogene.(1) 1,2,4-Benzenetriol, a benzene metabolite, caused strong DNA damage even without alkali treatment. Alkali-labile sites induced by 1,2,4-benzenetriol were base residues of guanine and adjacent thymine. Catalase, SOD and methional inhibited the DNA damage completely, but sodium formate did not inhibit it. The DNA damage was inhibited by the addition of Cu(I)-specific chelating agent, and was accelerated by the addition of Cu(II). The results suggest that the DNA damage was caused by unidenitified active species which was produced by the autoxidation of 1,2,4-benzenetriol in the presence of Cu(II), rather than by hydroxyl radicals. Similarly, 2,5-dihydroxybiphenyl, an 0-phenylphenol metabolite, induced site-specific DNA damage in the presene of copper(II).(2) It is calrified that carcinogeni … More c chromium(VI),iron(III)nitrilotriacetate, and cobalt(II)react with hydrogen peroxide leading to the production of active species including hydroxyl radical and singlet oxygen, which cause DNA damage. Carcinogenic nickel(II) binds to DNA and subsequently reacts with hydrogen peroxide to form active species, which induces DNA damage. The active species seems to be a nickel-oxygen complex rather than singlet oxygen or free hydroxyl radical.(3) Sulfite caused DNA damage in the presence of C0^<2+>,Cu^<2+> and Mn^<2+>. The order of inducing effect on sulfite-dependent DNA damage( Co^<2+>>>Cu^<2+>>Mn^<2+>>Fe^<3+>) was consistent with that of accelerating effect on the initial oxygen consumption rate of sulfite autoxidation. Incubation of DNA with sulfite plus Co^<2+> led to the predominant cleavage at the positions of guanine especially located 5' to guanine. Sulfite plus Cu^<2+> gave a DNA cleavage pattern different from that induced by sulfite plus Co^<2+>. The photolysis of peroxydisulfate (S_2o^2-_), which is known to produce SO^-_ radicals, gave a DNA cleavage pattern simiar to that induced by sulfite plus Co^<2+>. The results suggest that sulfite is rapidly autoxidized in the presence of Co^<2+> to produce SO^-_ radicals causing site-specific DNA damage. Less