Frequency of U.V. induced protein-DNA crosslinks in cell lines of different sensitivities

We have measured U.V.-induced protein-DNA crosslinking in two cultured human cell lines of different sensitivities. Using a previously published method, involving SDS-protein precipitation, we obtained a biphasic response with an initial slope of 0.6 per cent DNA J-1 m2 up to 50 J m-2 and a second-phase slope of 0.12 per cent DNA J-1 m2 with a background of 22 +/- 13 per cent. Rigorous washing of SDS-protein precipitates reduced background binding to about 5 per cent with a linear U.V. effect up to 100 J m-2 of 0.038 per cent DNA J-1 m2. Binding was judged to be covalent on the grounds of stability to boiling and represented 4.1 crosslinks pg DNA-1 J-1 m2 or 60 crosslinks cell-1 J-1 m2. Similar results were obtained for both cell lines. We conclude that the differences in U.V. survival between cell lines is not related to the extent of protein-DNA crosslinking. We have been unable to detect repair of these lesions in either cell line.