Genetic Engineering Research Centre, College of Life Science, Chongqing University, Chongqing 400030, China; Institute of Plant Physiology and Ecology, Chinese Academy of Sciences Key Laboratory of Insect Developmental and Evolutionary Biology, CAS, Shanghai 200032, China.
Genetic Engineering Research Centre, College of Life Science, Chongqing University, Chongqing 400030, China.
Dev Comp Immunol. 2014 May;44(1):76-85. doi: 10.1016/j.dci.2013.11.012. Epub 2013 Dec 1.
Thioredoxin peroxidases (Tpxs) are a ubiquitous family of antioxidant enzymes that play important roles in protecting organisms against oxidative stress. Here, one Tpx was cloned from Spodoptera litura named as SlTpx. The full-length cDNA consists of 1165 bp with 588 bp open reading frame, encoding 195 amino acids. The putative amino acid sequence shared >70% identity with Tpxs from other insects. Phylogenetic analysis revealed that SlTpx is closely related to other available lepidopteran Tpxs. Real-time PCR analysis showed that SlTpx can be induced by Nomuraea rileyi infection in some detected tissues at the mRNA level. The strongest expression was found in hemocytes of unchallenged and N. rileyi-challenged S. litura. Western blotting showed SlTpx protein in the hemocytes, head and cuticle from normal S. litura. However, when N. rileyi was inoculated into the body cavity of S. litura larvae, SlTpx protein was detected in head, hemocytes, fatbody, midgut, malpighian tubule, but not in the hemolymph and cuticle. Moreover, time-course analysis showed that SlTpx mRNA/protein expression levels were up-regulated in the hemocytes, when S. litura were infected by N. rileyi or injected with H2O2. The levels of N. rileyi-induced reactive oxygen species (ROS) in hemocytes were evaluated, and revealed that N. rileyi infection caused generation of ROS, and induced changes in expression of SlTpx. In addition, the heterologously expressed protein of this gene in Escherichia coli showed antioxidant activity; it removed H2O2 and protected DNA. Knocking down SlTpx transcripts by dsRNA interference resulted in accelerated insect death with N. rileyi infection. This is believed to be the first report showing that SlTpx has a significant role in resisting oxidative stress caused by N. rileyi infection.
硫氧还蛋白过氧化物酶（Tpxs）是一种普遍存在的抗氧化酶家族，在保护生物免受氧化应激方面发挥着重要作用。在这里，我们从斜纹夜蛾（Spodoptera litura）克隆了一种 Tpx，命名为 SlTpx。全长 cDNA 由 1165bp 组成，其中 588bp 为开放阅读框，编码 195 个氨基酸。推测的氨基酸序列与其他昆虫的 Tpxs 具有>70%的同一性。系统进化分析表明，SlTpx 与其他鳞翅目 Tpxs 密切相关。实时 PCR 分析显示，SlTpx 在一些检测组织中可以被杆状病毒感染诱导，在未受挑战和感染杆状病毒的斜纹夜蛾中，mRNA 水平表达最强。Western blot 显示 SlTpx 蛋白存在于正常斜纹夜蛾的血细胞、头部和表皮中。然而，当杆状病毒接种到斜纹夜蛾幼虫的体腔时，SlTpx 蛋白在头部、血细胞、脂肪体、中肠、马氏管中被检测到，但在血淋巴和表皮中未被检测到。此外，时间进程分析表明，当斜纹夜蛾被杆状病毒或 H2O2 感染时，SlTpx 的 mRNA/蛋白表达水平在血细胞中上调。评估了杆状病毒感染引起的血细胞中活性氧（ROS）的水平，结果表明杆状病毒感染引起了 ROS 的产生，并诱导了 SlTpx 的表达变化。此外，该基因在大肠杆菌中的异源表达蛋白具有抗氧化活性；它清除了 H2O2 并保护了 DNA。用 dsRNA 干扰敲低 SlTpx 转录本会导致斜纹夜蛾在感染杆状病毒时死亡加速。这是第一个表明 SlTpx 在抵抗杆状病毒感染引起的氧化应激方面具有重要作用的报告。
