Hainan Provincial Key Laboratory for Functional Components Research and Utilization of Marine Bio-resources, Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences, Haikou, 571101, People's Republic of China; Guangdong Provincial Key Laboratory for Healthy and Safe Aquaculture, School of Life Sciences, South China Normal University, Guangzhou, 510631, People's Republic of China.
Institute for Brain Research and Rehabilitation, South China Normal University, Guangzhou, 510631, People's Republic of China.
Fish Shellfish Immunol. 2019 Mar;86:662-671. doi: 10.1016/j.fsi.2018.12.011. Epub 2018 Dec 8.
Glutaredoxin (Grx) is a class molecule oxidoreductase, which can regulate the redox state of proteins and plays a key role in antioxidant defense. However, the informations of Grx cDNA sequences and their functions are lack in decapod crustacea. In the present study, the cDNA of LvGrx 2 was cloned from the Pacific white shrimp, Litopenaeus vannamei. The open reading frame (ORF) of LvGrx 2 was 360 bp, which encoded a polypeptide of 119 amino acids. The molecular mass of the predicted protein is 12.87 kDa with an estimated pI of 8.22. Sequence alignment showed that the amino acid sequence of LvGrx 2 shares 59%, 59% and 58% identity with that of the coelacanth Latimeria chalumnae, the plateau frog Nanorana parkeri and the half-smooth tongue sole Cynoglossus semilaevis, respectively. Quantitative real-time PCR analysis revealed that LvGrx 2 were detected in a wide range of tissues, with highest expression in gill, hepatopancrea and intestine, and weakest expression in muscle. The expression responses of LvGrx 2 were analyzed in hepatopancrea and gill after ammonia-N stress or lipopolysaccharide (LPS) injection. During ammonia-N exposure, the LvGrx 2 transcriptions in hepatopancrea and gill significantly up-regulated, and the peak value appeared after 12 h and 24 h exposure respectively. After LPS injection, expression levels of LvGrx 2 in hepatopancrea obviously increased in the early and late stages, while LvGrx 2 transcription in gill sharply up-regulated in the middle period. These results suggest that LvGrx 2 may play a vital role in shrimp defense system against environmental stress and pathogen infection. RNA interference experiment was designed to further probe roles of LvGrx 2 during ammonia-N exposure. Ammonia-N induced obvious improvement in expression levels of LvGrx 2, LvGrx 3, GPx, GST and Trx, accompanied by increases of protein carbonyl and malondialdehyde (MDA) contents. However, transcription of GPx and GST were much weaker in LvGrx 2 interfered-shrimp, and oxidative damage in both lipid and protein were more serious. These results further suggest that LvGrx 2 in shrimp participates in oxidative defence and regulation of antioxidant system.
谷胱甘肽还原酶（Grx）是一类分子氧化还原酶，可调节蛋白质的氧化还原状态，在抗氧化防御中发挥关键作用。然而，甲壳动物的 Grx cDNA 序列及其功能信息仍然缺乏。本研究从凡纳滨对虾（Litopenaeus vannamei）中克隆了 LvGrx 2 的 cDNA。LvGrx 2 的开放阅读框（ORF）为 360bp，编码 119 个氨基酸的多肽。预测蛋白的分子量为 12.87 kDa，等电点（pI）为 8.22。序列比对表明，LvGrx 2 的氨基酸序列与腔棘鱼（Latimeria chalumnae）、高原蛙（Nanorana parkeri）和半滑舌鳎（Cynoglossus semilaevis）的分别具有 59%、59%和 58%的同源性。实时定量 PCR 分析显示，LvGrx 2 在广泛的组织中均有表达，其中在鳃、肝胰腺和肠中表达量最高，在肌肉中表达量最低。分析了氨氮胁迫或脂多糖（LPS）注射后肝胰腺和鳃中 LvGrx 2 的表达反应。在氨氮暴露期间，肝胰腺和鳃中的 LvGrx 2 转录物明显上调，分别在暴露 12 和 24 小时后达到峰值。注射 LPS 后，肝胰腺中 LvGrx 2 的表达水平在早期和晚期明显增加，而鳃中 LvGrx 2 的转录在中期急剧上调。这些结果表明，LvGrx 2 在虾的防御系统中可能在应对环境胁迫和病原体感染方面发挥重要作用。设计 RNA 干扰实验进一步探讨 LvGrx 2 在氨氮暴露期间的作用。氨氮诱导 LvGrx 2、LvGrx 3、GPx、GST 和 Trx 的表达水平明显提高，同时伴随着蛋白质羰基和丙二醛（MDA）含量的增加。然而，在 LvGrx 2 干扰虾中，GPx 和 GST 的转录水平要弱得多，脂质和蛋白质的氧化损伤也更为严重。这些结果进一步表明，虾中的 LvGrx 2 参与了氧化防御和抗氧化系统的调节。
