FORMATION IN ISOLATED RAT-LIVER MICROSOMES AND NUCLEI OF BENZO(A)PYRENE METABOLITES THAT BIND TO DNA

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  • 1 January 1976
    • journal article
    • research article
    • Vol. 36  (11) , 4107-4113
Abstract
The hepatic nuclear fraction isolated from 3-methylcholanthrene(MC)-treated rats contained enhanced levels of cytochrome P-450 and aryl hydrocarbon hydroxylase [benzo(a)pyrene (BP) monooxygenase], whereas the activities of epoxide hydrase and NADPH-cytochrome c reductase and the concentration of cytochrome b5 were not altered. The metabolite pattern of BP was investigated by using high pressure liquid chromatography and was similar in nuclei and microsomes from MC-treated rats. After incubation of the nuclear fraction with [3H]BP and NADPH, radioactivity was associated with nuclear DNA and the extent of binding was enhanced by pretreatment of the animals with MC. Binding was strongly inhibited by .alpha.-naphthoflavone but was not influenced by 1,1,1-trichloropropene-2,3-oxide, an inhibitor of expoxide hydrase. In the presence of microsomes from MC-treated rats, increased binding of BP to DNA was observed in nuclei from control and MC-treated rat. When the nuclear DNA was replaced by a corresponding amount of calf thymus DNA, the extent of binding was severalfold enhanced. In contrast to nuclei from control rats, the nuclear fraction from MC-treated rats showed an increase in bound radioactivity when incubated with microsome-free supernatant, obtained by incubating microsomes from MC-treated rats with [3H]BP. The increase in extent of binding was eliminated in the presence of menadione or .alpha.-naphthoflavone. Under the conditions used here the following different processes may have contributed to the total incorporation of BP products into nuclear DNA: formation of DNA-binding products derived from BP by nuclear aryl hydrocarbon hydroxylase; formation of DNA-binding products from microsomal BP metabolites by nuclear aryl hydrocarbon hydroxylase and direct transfer of reactive microsomal metabolites to nuclear DNA.