Characterization of DNA topoisomerase-1 in Spodoptera exigua for toxicity evaluation of camptothecin and hydoxy-camptothecin.
Characterization of DNA topoisomerase-1 in Spodoptera exigua for toxicity evaluation of camptothecin and hydoxy-camptothecin.
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甜菜夜蛾 DNA 拓扑异构酶 1 的表征用于喜树碱和羟基喜树碱的毒性评价
DOI：
10.1371/journal.pone.0056458
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Jiang H
中科院分区：
文献类型：
--
作者：
Zhang L;Ma D;Zhang Y;He W;Yang J;Li C;Jiang H
喜树碱（Camptothecin，CPT）是一种从喜树中分离得到的生物碱，其作用靶点是真核生物DNA拓扑异构酶1（DNA Topo 1）。除了作为有效的抗癌剂外，CPT及其衍生物现在正在探索作为潜在的杀虫剂用于昆虫控制。在这项研究中，我们评估了他们的毒性昆虫同源物，拓扑1蛋白甜菜夜蛾（甜菜夜蛾Hübner），许多重要作物的世界性害虫。色葡萄甜菜Topo 1基因的开放阅读框为2790个碱基对，预测编码930个氨基酸的多肽。推测的多肽在昆虫物种中的残基位点V420、L530、A653和T729（根据人类Topo 1编号）具有多态性，预测这些残基位点赋予CPT敏感性。随后使用含有残基337-930并在细菌BL 21细胞中表达的截短形式检测该蛋白质的DNA松弛活性。纯化的蛋白质保留了松弛双链DNA的能力，并以剂量依赖的方式对CPT及其衍生物羟基喜树碱（HCPT）敏感。从IOZCAS-Spex-II细胞中提取的天然Topo 1也具有相同的抑制作用，IOZCAS-Spex-II细胞是从甜菜夜蛾建立的细胞系。此外，CPT和HCPT处理降低了Topo 1蛋白的稳定积累，尽管mRNA表达增加。我们的研究提供了S.甜菜Topo 1基因及其氨基酸多态性的研究，为深入研究喜树碱对害虫的潜在毒力机制提供线索。这些结果也是有用的，在未来开发更有效的Topo 1靶向CPT杀虫剂。
Camptothecin (CPT), a plant alkaloid originally isolated from the native Chinese tree, Camptotheca acuminate, exerts the toxic effect by targeting eukaryotic DNA topoisomerase 1 (DNA Topo1). Besides as potent anti-cancer agents, CPT and its derivatives are now being explored as potential pesticides for insect control. In this study, we assessed their toxicity to an insect homolog, the Topo1 protein from beet armyworms (Spodoptera exigua Hübner), a worldwide pest of many important crops. The S. exigua Topo1 gene contains an ORF of 2790 base pairs that is predicted to encode a polypeptide of 930 amino acids. The deduced polypeptide exhibits polymorphism at residue sites V420, L530, A653 and T729 (numbered according to human Topo1) among insect species, which are predicted to confer sensitivity to CPT. The DNA relaxation activity of this protein was subsequently examined using a truncated form that contained the residues 337–930 and was expressed in bacteria BL21 cells. The purified protein retained the ability to relax double-stranded DNA and was susceptible to CPT and its derivative hydroxy-camptothecin (HCPT) in a dose-dependent manner. The same inhibitory effect was also found on the native Topo1 extracted from IOZCAS-Spex-II cells, a cell line established from beet armyworms. Additionally, CPT and HCPT treatment reduced the steady accumulation of Topo1 protein despite the increased mRNA expression in response to the treatment. Our studies provide information of the S. exigua Topo1 gene and its amino acid polymorphism in insects and uncover some clues about potential mechanisms of CPT toxicity against insect pests. These results also are useful for development of more effective Topo1-targeted CPT insecticides in the future.