COPPER ION-MEDIATED SENSITIZATION OF NUCLEAR MATRIX ATTACHMENT SITES TO IONIZING-RADIATION

Exposure of mammalian cells to ionizing radiation induces nuclear matr ix proteins and their attached transcribing DNA sequences to form cros s-links. To characterize the cellular and matrix components necessary for DNA-protein crosslink (DPC) formation, DPC yields have been examin ed in isolated nuclear matrices and in the intermediate steps during c ell fractionation. It was found that, in both unirradiated and irradia ted cells, all components of DPC are retained in isolated nuclei, and the formed DPC are retained as well during the cell fractionation proc edure resulting in nuclear matrices. In contrast, nuclear matrices iso lated from unirradiated cells are deficient in the ability to form DPC upon irradiation, indicating that elements necessary for DPC producti on have been disrupted or removed during the isolation procedure. When isolated nuclei were irradiated, the yield of radiation-induced DPC w as about 2-fold higher than that for intact cells, presumably due to t he removal of soluble cellular scavengers during the isolation procedu re. Treatment of nuclei with Cu2+ to stabilize nuclear structural orga nization during the preparation of the nuclear matrix caused additiona l DNA, especially the matrix-associated newly replicated DNA, to becom e bound to protein. Such treatment also enhanced radiation-induced DPC production which was sensitive to OH radical scavengers. Moreover, ra diation-induced DPC production in Cu2+-treated nuclei was more sensiti ve to EDTA and catalase than in untreated nuclei. It is therefore prop osed that excess DPC induction in Cu2+-treated nuclei occurs preferent ially at the sites of Cu2+ binding to chromatin where hydroxyl radical s are produced repeatedly through the Fenton reaction.