The binding of benzo(a)pyrene and N‐methyl‐N′‐nitro‐n‐nitrosoguanidine to subnuclear fractions of akr mouse embryo cells in culture

The marked localization of a carcinogenic polycyclic aromatic hydrocarbon, benzo(a)pyrene, and its metabolites and a carcinogenic alkylating agent, N‐methyl‐N'‐nitro‐N‐nitrosoguanidine, to a specific subnuclear fraction (fraction I) from AKR‐2B mouse embryo cells in culture is described. Fraction I is isolated by sucrose gradient centrifugation of sheared nuclei from cells exposed to the carcinogens. The association of tritiated benzo(a)‐pyrene to fraction I consisted of loosely associated radioactivity which is extractable by organic solvents, and of tightly bound (termed “covalently” bound) radioactivity which is not extractable by organic solvents. Increases in the extent of metabolism of benzo(a)pyrene and in the amount of “covalently” bound radioactivity occur with increasing periods of incubation of the cells with the labelled carcinogen. This observation, together with the fact that these increases are dramatically reduced by inhibiting polycyclic aromatic hydrocarbon metabolism (using the inhibitor 7,8‐benzoflavone), suggests that a time‐dependent metabolism of benzo(a)pyrene is required for “covalent” binding to nuclear material. Data are presented suggesting that a two‐step reaction may be involved in the binding of benzo(a)pyrene to subnuclear macromolecules. The fraction I localization of such structurally diverse chemical carcinogens as benzo(a)pyrene and N‐methyl‐N'‐nitro‐N‐nitrosoguanidine suggests that this fraction may localize all species of chemical carcinogens and that this localization may be involved in the chemically induced malignant transformation of cells.