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Clusters of DNA Damage Induced by Ionizing Radiation: Formation of Short DNA Fragments. II. Experimental Detection
Vol. 145, No. 2 (Feb., 1996), pp. 200-209
Published by: Radiation Research Society
Stable URL: http://www.jstor.org/stable/3579175
Page Count: 10
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The basic 30-nm chromatin fiber in the mammalian cell consists of an unknown (possibly helical) arrangement of nucleosomes, with about 1.2 kb of DNA per 10-nm length of fiber. Track-structure considerations suggest that interactions of single δ rays or high-LET particles with the chromatin fiber might result in the formation of multiple lesions spread over a few kilo-bases of DNA (see the accompanying paper: W. R. Holley and A. Chatterjee, Radiat. Res. 145, 188-199, 1996). In particular, multiple DNA double-strand breaks and single-strand breaks may form. To test this experimentally, primary human fibroblasts were labeled with [3 H]thymidine and exposed at 0°C to X rays or accelerated nitrogen or iron ions in the LET range of 97-440 keV/μm. DNA was isolated inside agarose plugs and subjected to agarose gel electrophoresis under conditions that allowed good separation of 0.1-2 kb size DNA. The bulk of DNA remained in the well or migrated only a small distance into the gel. It was found that DNA fragments in the expected size range were formed linearly with dose with an efficiency that increased with LET. A comparison of the yield of such fragments with the yield of total DNA double-strand breaks suggests that for the high-LET ions a substantial proportion (20-90%) of DNA double-strand breaks are accompanied within 0.1-2 kb by at least one additional DNA double-strand break. It is shown that these results are in good agreement with theoretical calculations based on treating the 30-nm chromatin fiber as the target for ionizing particles. Theoretical considerations also predict that the clusters will contain numerous single-strand breaks and base damages. It is proposed that such clusters be designated "regionally multiply damaged sites." Postirradiation incubation at 37°C resulted in a decline in the number of short DNA fragments, suggesting a repair activity. The biological significance of regionally multiply damaged sites is presently unknown.
Radiation Research © 1996 Radiation Research Society