Cloning and Expression Analysis of JHEH Gene in Portunus trituberculatus

doi:10.7668/hbnxb.20191281

Abstract

Abstract: To explore the role of JHEH in crustaceans, the full-length cDNA of JHEH from the swimming crab, Portunus trituberculatus (PtJHEH) was cloned and characterized using the RACE technique followed by bioinformatics analysis, and its gene expression levels were detected in different tissues, molting cycles and ovarian development stages using the Real-time quantitative PCR(RT-qPCR). The obtained PtJHEH was 1 803 bp in length, containing an open reading frame(ORF) of 1 374 bp, which encoded 467 amino acids. The deduced amino acid sequence was predicted to have transmembrane domains, signal peptides, and typical epoxide catalytic domain. The qPCR results showed that PtJHEH was expressed extensively in various tissues, but the highest mRNA level was found in the hepatopancreas, which was significantly higher than other tissues(P<0.05). This might be the reflection that the hepatopancreas was the main site of crustacean metabolism. During the molt cycle, the PtJHEH in the hepatopancreas expressed at highest level in the pre-molt D2 stage, while its expression in Y-organs was remarkably high at the post-molt stages(A and B stages), but then decreased significantly. During the ovarian development process, the expression of PtJHEH in hepatopancreas and Y-organs both increased to the highest level at the late-vitellogenic stage(stage Ⅲ). The expression pattern of PtJHEH in the molting cycle and ovarian development showed a correlation to the fluctuation in the hemolymph MF titers previously reported by our group. This indicates that JHEH may have a role in the regulation of MF degradation.

Key words: Portunus trituberculatus, Juvenile hormone epoxide hydrolase(JHEH), Expression analysis, Molting cycle, Ovarian development

CLC Number:

Q78
S968

ZHENG Liang, XIE Xi, ZHENG Hongkun, XU Dongjie, ZHU Dongfa. Cloning and Expression Analysis of JHEH Gene in Portunus trituberculatus[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2020, 35(6): 225-234. doi: 10.7668/hbnxb.20191281.

/ / Recommend / Download Citations

URL: http://www.hbnxb.net/EN/10.7668/hbnxb.20191281

http://www.hbnxb.net/EN/Y2020/V35/I6/225

References

[1] 刘素宁, 任充华, 李胜. 昆虫变态发育的激素和营养调控研究进展与展望[J].应用昆虫学报, 2019, 56(4):639-651.doi:10.7679/j.issn.2095-1353.2019.074.
Liu S N, Ren C H, Li S. Progress in research on hormonal and nutritional regulation in insect metamorphosis[J]. Chinese Journal of Applied Entomology, 2019, 56(4):639-651.
[2] Tusun A, Li M, Liang X Z, Yang T, Yang B, Wang G R. Juvenile hormone epoxide hydrolase:a promising target for hemipteran pest management[J]. Scientific Reports, 2017, 7(1):789.doi:10.1038/s41598-017-00907-0.
[3] Sin Y W, Kenny N J, Qu Z, Chan K W, Chan K W S, Cheong S P S, Leung R W T, Chan T F, Bendena W G, Chu K H, Tobe S S, Hui J H L. Identification of putative ecdysteroid and juvenile hormone pathway genes in the shrimp Neocaridina denticulata[J]. General and Comparative Endocrinology, 2015, 214(4):167-176.doi:10.1016/j.ygcen.2014.07.018.
[4] Kamita S G, Hinton A C, Wheelock C E, Wogulis M D, Wilson D K, Wolf N M, Stok J E, Hock B, Hammock B D. Juvenile hormone(JH) esterase:why are you so JH specific?[J]. Insect Biochemistry and Molecular Biology, 2003, 33(12):1261-1273.doi:10.1016/j.ibmb.2003.08.004.
[5] Lü F G, Fu K Y, Li Q, Guo W C, Ahmat T, Li G Q. Identification of carboxylesterase genes and their expression profiles in the Colorado potato beetle Leptinotarsa decemlineata treated with fipronil and cyhalothrin[J].Pesticide Biochemistry and Physiology, 2015, 122(9):86-95.doi:10.1016/j.pestbp.2014.12.015.
[6] Wen R R, Wang B Y, Wang B W, Ma L. Characterization and expression profiles of juvenile hormone epoxide hydrolase from Lymantria dispar (Lepidoptera:Lymantridae) and RNA interference by ingestion[J]. Journal of Insect Science, 2018, 18(1):13.doi:10.1093/jisesa/iey002.
[7] Zhao J, Zhou Y L, Li X, Cai W L, Hua H X. Silencing of juvenile hormone epoxide hydrolase gene(Nljheh) enhances short wing formation in a macropterous strain of the brown planthopper, Nilaparvata lugens[J]. Journal of Insect Physiology, 2017, 102(6):18-26.doi:10.1016/j. jinsphys.2017.08.012.
[8] Khlebodarova T M, Gruntenko N E, Grenback L G, Sukhanova M Z, Mazurov M M, Rauschenbach I Y, Tomas B A, Hammock B D. A comparative analysis of juvenile hormone metabolyzing enzymes in two species of Drosophila during development[J]. Insect Biochemistry and Molecular Biology, 1996, 26(8-9):829-835.doi:10.1016/s0965-1748(96)00043-4.
[9] Zhang Q R, Xu W H, Chen F S, Li S. Molecular and biochemical characterization of juvenile hormone epoxide hydrolase from the silkworm, Bombyx mori[J]. Insect Biochemistry and Molecular Biology, 2005, 35(2):153-164.doi:10.1016/j.ibmb.2004.10.010.
[10] 吕凤功. 马铃薯甲虫(Leptinotarsa decemlineata)保幼激素降解酶基因的分子克隆及功能分析[D]. 南京:南京农业大学, 2015.
Lü F G. Cloning and characterization of 3 genes involved in juvenile hormone degeneration in Leptinotarsa decemlineata[D]. Nanjing:Nanjing Agricultural University, 2015.
[11] Mackert A, Hartfelder K, Bitondi M M G, Simões Z L P. The juvenile hormone(JH) epoxide hydrolase gene in the honey bee(Apis mellifera) genome encodes a protein which has negligible participation in JH degradation[J]. Journal of Insect Physiology, 2010, 56(9):1139-1146.doi:10.1016/j.jinsphys.2010.03.007.
[12] Kamita S G, Oshita G H, Wang P, Morisseau C, Hammock B D, Nandety R S, Falk B W. Characterization of Hovi-mEH1, a microsomal epoxide hydrolase from the glassy-winged sharpshooter Homalodisca vitripennis[J]. Archives of Insect Biochemistry and Physiology, 2013, 83(4):171-179.doi:10.1002/arch.21100.
[13] 李真真. 青虾保幼激素环氧水解酶基因全长cDNA的克隆及时空表达分析[D]. 南京:南京农业大学, 2012.doi:10.7666/d.Y2361703.
Li Z Z. Full-length cDNA cloning and spatio-temporal expression analysis of juvenile hormone epoxide hydrolase(JHEH) gene in Macrobrachium nipponense[D]. Nanjing:Nanjing Agricultural University, 2012.
[14] 赵明, 张凤英, 蒋科技, 马凌波. 保幼激素环氧水解酶可作为MF调控卵巢发育的潜在靶标-分子表达证据[A]//中国水产学会.2015年中国水产学会学术年会论文摘要集[C]. 杭州:中国水产学会学术年会, 2015:372-372.
Zhao M, Zhang F Y, Jiang K J, Ma L B. JHEH serve as a potential target for the regulation of ovary through MF:evidence from molecular expression analysis[A]//The China Society of Fisheries. Abstract collection of papers of 2015 annual conference of the China society of fisheries[C]. Hangzhou:Academic Annual Conference of the China Society of Fisheries, 2015:372-372.
[15] 邱锡尔, 朱冬发, 汤洁, 崔晓雨, 周彦琦, 柳志业. 甲基法尼酯在甲壳动物中的生理作用及其机制的研究进展[J].生物学杂志, 2015, 32(1):76-81.doi:10.3969/j.issn.2095-1736.2015.01.076.
Qiu X E, Zhu D F, Tang J, Cui X Y, Zhou Y Q, Liu Z Y. Review of the physiological functions and their mechanisms of methyl farnesoate in crustacean[J]. Journal of Biology, 2015, 32(1):76-81.
[16] 王凤皎, 朱冬发, 邱锡尔, 谭义川, 周彦琦, 柳志业, 谢熙. 三疣梭子蟹AACT基因克隆及其在卵巢发育中的表达分析[J].水产学报, 2015, 39(6):790-798.doi:10.11964/jfc.20141209636.
Wang F J, Zhu D F, Qiu X E, Tan Y C, Zhou Y Q, Liu Z Y, Xie X. Cloning and expression analysis of acetyl-CoA C-acetyltransferase(AACT) in Portunus trituberculatu[J]. Journal of Fisheries of China, 2015, 39(6):790-798.
[17] 朱韬, 朱冬发, 邱锡尔, 周彦琦, 柳志业, 谢熙. 三疣梭子蟹fps克隆及其在卵巢发育中的表达分析[J].生物学杂志, 2016, 33(3):10-14,19.doi:10.3969/j.issn.2095-1736.2016.03.010.
Zhu T, Zhu D F, Qiu X E, Zhou Y Q, Liu Z Y, Xie X. Cloning of farnesyl diphosphate synthase(fps) from Portunus trituberculatus and its expression during the ovarian development[J].Journal of Biology, 2016, 33(3):10-14,19.
[18] Xie X, Zhu D F, Li Y, Qiu X E, Cui X Y, Tang J. Hemolymph levels of methyl farnesoate during ovarian development of the swimming crab Portunus trituberculatus, and its relation to transcript levels of HMG-CoA reductase and farnesoic acid O-methyltransferase[J]. The Biological Bulletin, 2015, 228(2):118-124.doi:10.1086/BBLv228n2p118.
[19] 沈洁, 朱冬发, 胡则辉, 亓一舟, 汪春建. 三疣梭子蟹蜕皮周期的分期[J].水产学报, 2011, 35(10):1481-1487.doi:10.3724/SP.J.1231.2011.17418.
Shen J, Zhu D F, Hu Z H, Qi Y Z, Wang C J. Molt staging in the swimming crab Portunus trituberculatus[J]. Journal of Fisheries of China, 2011, 35(10):1481-1487.
[20] 姚桂桂, 吴旭干, 杨筱珍, 成永旭, 王春琳. 三疣梭子蟹的第二次卵巢发育规律[J].动物学研究, 2007, 28(4):423-429.doi:10.3312/j.issn.0254-5853.2007.04.017.
Yao G G, Wu X G, Yang X Z, Cheng Y X, Wang C L. The second ovarian development of swimming crab, Portunus trituberculatus[J]. Zoological Research, 2007, 28(4):423-429.
[21] Zhou K, Jia N, Hu C, Jiang Y L, Yang J P, Chen Y X, Li S, Li W F, Zhou C Z. Crystal structure of juvenile hormone epoxide hydrolase from the silkworm Bombyx mori[J]. Proteins, 2014, 82(11):3224-3229.doi:10.1002/prot.24676.
[22] Lee S O, Jeon J M, Oh C W, Kim Y M, Kang C K, Lee D S, Mykles D L, Kim H W. Two juvenile hormone esterase-like carboxylesterase cDNAs from a Pandalus shrimp(Pandalopsis japonica):cloning, tissue expression, and effects of eyestalk ablation[J]. Comparative Biochemistry and Physiology Part B:Biochemistry and Molecular Biology, 2011, 159(3):148-156.doi:10.1016/j.cbpb.2011.03.004.
[23] Homola E, Chang E S. Methyl farnesoate:crustacean juvenile hormone in search of functions[J]. Comparative Biochemistry and Physiology Part B:Biochemistry and Molecular Biology, 1997, 117(3):347-356.doi:10.1016/S0305-0491(96)00337-9.
[24] Lima A V B, Guerra A L, Almeida E A, Taddei F G, Castiglioni L. Characterization of esterase patterns in hepatopancreas of three species of macrobrachium(Palaemonidae)[J]. Biochemical Systematics and Ecology, 2013, 47(7):132-138.doi:10.1016/j.bse.2012.10.006.
[25] 谢熙. 三疣梭子蟹甲基法尼酯合成及在卵巢发育和蜕皮中的作用[D]. 宁波:宁波大学, 2016.
Xie X.The methyl fanesoate biosynthesis and its role in ovarian development and molt cycle of Portunus trituberculatus[D]. Ningbo:Ningbo University, 2016.
[26] 邵娟, 郑亮, 郑宏坤, 谢熙, 朱冬发. 三疣梭子蟹蜕皮周期及眼柄切除后血淋巴甲基法尼酯浓度测定及其合成酶基因的表达分析[J].生物学杂志, 2019, 36(2):28-32.doi:10.3969/j.issn.2095-1736.2019.02.028.
Shao J, Zheng L, Zheng H K, Xie X, Zhu D F. Changes in the hemolymph levels of methyl farnesoate and gene expression of enzymes in its biosynthesis during the molt cycle and after eyestalk ablation of the swimming crab, Portunus trituberculatus[J]. Journal of Biology, 2019, 36(2):28-32.
[27] 汪春建, 朱冬发, 亓一舟, 胡则辉, 谢熙, 沈洁. 三疣梭子蟹蜕皮周期中MIH 基因mRNA水平与蜕皮激素浓度变化[J].水生生物学报, 2013, 37(1):22-28.doi:10.7541/2013.22.
Wang C J, Zhu D F, Qi Y Z, Hu Z H, Xie X, Shen J. Molt-inhibiting hormone levels and ecdysteroid titer during a molt cycle of Portunus trituberculatus[J]. Acta Hydrobiologica Sinica, 2013, 37(1):22-28.
[28] Gong J, Ye H H, Xie Y J, Yang Y N, Huang H Y, Li S J, Zeng C S. Ecdysone receptor in the mud crab Scylla paramamosain:a possible role in promoting ovarian development[J]. Journal of Endocrinology, 2015, 224(3):273-287.doi:10.1530/JOE-14-0526.

Please choose a citation manager

Content to export