The yield, spatial distribution, and rejoining of DNA strand breaks produced by sparsely and densely ionizing radiations: Their relationship to radiosensitivity in mammalian cells

A filter elution assay has been developed that measures DNA damage at the subdenaturing pH of 11.1. This assay is able to quantitate DNA double-strand breaks (DSB) produced y as little as 1 Gy. Its specificity for the measurement of DSB is supported by nonresponsiveness of the assay to {dollar}rm Hsb2Osb2{dollar} and by its response to the restriction endonuclease HaeIII, which produces no DNA damage except DSB. This subdenaturing filter elution assay has been applied to the measurement of DSB yields from different radiations and their rates of enzymatic rejoining in both normal and abnormally radiosensitive cultured mammalian cells.; The length distributions of DNA fragments induced by gamma rays, fast neutrons, and accelerated nitrogen ions were measured by a pH-step elution assay. Although gamma rays, fast neutrons, and accelerated nitrogen ions produce similar total yields of DSB per absorbed dose, the more densely ionizing neutrons and nitrogen ions produce more fragments in the shortest size class and fewer in the longer size classes compared to sparsely ionizing gamma radiation. These observations suggest that densely ionizing radiations cluster DSB on the length scale of chromatin loops (10,000-100,000 bp), a phenomenon that has not been previously reported. Clustering of DSB could contribute to the complexity of DNA damage, reducing the fidelity of enzymatic DNA repair and increasing lethality per DSB. Thus, DSB clustering may account in part for the greater biological effectiveness of densely ionizing radiations per physical absorbed dose.; Finally, the normally radiosensitive CHO-K1 and AA8 cell lines were compared with the abnormally radiosensitive line EM9. EM9 cells demonstrate a DNA break rejoining defect for both SSB and DSB. This defect applies equally to both gamma rays and neutrons, but EM9 cells were nearly normal in their rate of rejoining {dollar}rm Hsb2Osb2{dollar} induced single-strand breaks. The survival curve parameters of these cell lines appear to correlate with residual DSB, but with a different quantitative relationship for CHO-K1 than for AA8 and EM9. This difference may be due to a greater fidelity of DNA repair or greater tolerance to unrejoined DSB for CHO-K1.* ftn*Originally published in DAI Vol. 56, No. 11. Reprinted here with corrected author name.