DNA compaction plays a key role in radioprotection against double-strand breaks as revealed by single-molecule observation

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  • 単分子観察法によるＤＮＡの放射線切断に関する研究

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Abstract

Introduction Naturally occurring polyamines such as spermine and spermidine have been shown to protect DNA against radiation-induced single- and double-strand breaks. Several possible mechanisms for the radioprotective effects of polyamines have been proposed, including (1) direct scavenging of radiation-generated hydroxyl radical, and (2) the induction of DNA compaction or aggregation that would reduce susceptibility to radiation damage. However, most previous studies have been carried out without clearly distinguishing between the compaction of single DNA molecules and the aggregation of multiple DNA molecules. There has been no direct evidence that the conformational change of individual DNA molecules is a crucial factor in radioprotection. This may be due to the lack of a suitable experimental methodology. Further studies are needed to clarify which mechanism, i.e., the induction of compaction or radical scavenging, is the primary factor in the radioprotective effect of polyamines. Recently, we directly observed the conformational changes of single giant DNA molecules larger than 100 kbp in solution using fluorescence microscopy. We found that giant DNA molecules undergo a large discrete transition from an elongated state to a compact state upon the addition of various condensing agents including spermidine. In addition, it became clear that such a highly compacted DNA is unfolded by the addition of salt. Thus, in the present study we applied the above-mentioned experimental conditions to investigate the effect of g-ray irradiation on different DNA conformations, either an elongated coil state (treatment with spermidine in the presence of salt) or a folded compact state (treatment with spermidine in the absence of salt). Results and Discussion We performed single-DNA observation to measure double-strand breaks caused by g-ray irradiation. To analyze the efficiency of the breakage reaction in a semi-quantitative manner, we used T4 DNA, 166 kbp. The efficiency of g-ray-induced breakage for compact DNA in the presence of spermidine (3+) was ca 1/25 of that for elongated DNA. This finding suggests that the primary protective effect of polyamines is due to the compaction of DNA. References Y. Yoshiakwa, T. Mori, N. Magone, K. Hibino, and K. Yoshikawa, Chem. Phys. Lett., 456 (2008) 80-83.

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