Mechanisms underlying production of double-strand breaks in plasmid DNA after decay of 125I-Hoechst
Mechanisms underlying production of double-strand breaks in plasmid DNA after decay of 125I-Hoechst
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DOI：
10.1667/rr3591.1
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发表时间：
2006-08-01
影响因子：
3.4
通讯作者：
Kassis, Amin I.
中科院分区：
文献类型：
--
作者：
Balagurumoorthy, Pichumani;Chen, Kai;Kassis, Amin I.
先前，I-125标记的m-碘-对乙氧基Hoechst 33342((IEH)-I-125)在超螺旋(SC)质粒DNA中产生链断裂的动力学已经证明，在有或没有羟基自由基清除剂二甲基亚砜的情况下，每一次I-125都会产生类似于1DSB的产物。在这些实验中，(IEH)-I-125：DNA的摩尔比为42：1。我们现在假设这个DSB产量(但不是SSB产量)可能是一个高估，因为在仍然与镍(N)DNA结合的41(IEH)-I-125分子中的任何一个中都发生了随后的衰变。为了验证我们的假设，(IEH)-I-125与SC pUC19质粒((IEH-I-125：DNA比例类似于3：1)孵育，并在I-125衰变后定量测定SSB和DSB产量。正如预测的那样，每次I-125衰变产生的DSB的数量是以前报道的一半(与类似于1，+/-DMSO相比，类似于0.5)，而SSB的数量(类似于3/I-125衰变)与先前获得的相似(类似于90%由OH自由基产生)。原子力显微镜的直接可视化证实了(IEH)-I-125在SCDNA中衰变后形成了L和NDNA，并支持了所报道的断链产量。这些发现表明，虽然SSB的产量与(IEH)-I-125与DNA结合的数量无关，但N DNA中发生的125个1衰变可以错误地增加DSB产量。进一步的分析表明，17%的SSB和100%的DSB发生在(IEH)-I-125分子衰变的质粒分子内，而83%的SSB形成于相邻的质粒DNA分子中。(C)2006年由辐射研究学会提供
Previously, the kinetics of strand break production by I-125-labeled m-iodo-p-ethoxyHoechst 33342 ((IEH)-I-125) in supercoiled (SC) plasmid DNA had demonstrated that similar to 1 DSB is produced per I-125 decay both in the presence and absence of the hydroxyl radical scavenger DMSO. In these experiments, an (IEH)-I-125:DNA molar ratio of 42:1 was used. We now hypothesize that this DSB yield (but not the SSB yield) may be an overestimate due to subsequent decays occurring in any of the 41 (IEH)-I-125 molecules still bound to nicked (N) DNA. To test our hypothesis, (IEH)-I-125 was incubated with SC pUC19 plasmids ((IEH)-I-125:DNA ratio of similar to 3:1) and the SSB and DSB yields were quantified after the decay of I-125. As predicted, the number of DSBs produced per I-125 decay is one-half that reported previously (similar to 0.5 compared to similar to 1, +/- DMSO) whereas the number of SSBs (similar to 3/I-125 decay) is similar to that obtained previously (similar to 90% are generated by OH radicals). Direct visualization by atomic force microscopy confirms formation of L and N DNA after (IEH)-I-125 decays in SC DNA and supports the strand break yields reported. These findings indicate that although SSB production is independent of the number of (IEH)-I-125 bound to DNA, the DSB yield can be augmented erroneously by 125 1 decays occurring in N DNA. Further analysis indicates that 17% of SSBs and 100% of DSBs take place within the plasmid molecule in which an (IEH)-I-125 molecule decays, whereas 83% of SSBs are formed in neighboring plasmid DNA molecules. (c) 2006 by Radiation Research Society