Radiolysis of Chromatin Extracted from Cultured Mammalian Cells: Formation of DNA–protein Cross Links

Summary

Chromatin extracted from Chinese hamster lung fibroblasts has been examined for the formation of radiation-induced DNA-protein cross links, using a membrane filter assay. The relative efficiencies of the aqueous radical intermediates, OH, e⁻_aq and O⁻₂, were investigated. Cross links were found in gamma-irradiated isolated chromatin and in chromatin irradiated in the cell before isolation. When isolated chromatin was irradiated under conditions in which the chromosomal proteins were dissociated from the DNA, no cross links were detectable. The most efficient radical for the production of cross links in irradiated, isolated chromatin was found to be the hydroxyl radical, whereas, the superoxide radical was essentially ineffective. For chromatin irradiated in the cell before isolation, the greatest effect was seen for cells irradiated in an atmosphere of nitrous oxide, suggesting the hydroxyl radical may be involved in the formation of cross links in intact cells also. The formation of cross links in chromatin irradiated in cells before isolation was considerably less efficient than in irradiated, isolated chromatin.