Volume 33 Issue 5
Sep.  2012
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SHI Yu-Hong, CHEN Jiong, GAO Shan-Shan, SHEN Guang-Qiang, LU Xin-Jiang, LI Ming-Yun. Cloning, physical and chemical property analysis of the Japanese sea bass Wap65-2 gene and its expression following Vibrio harveyi infection. Zoological Research, 2012, 33(5): 481-486. doi: 10.3724/SP.J.1141.2012.05481
Citation: SHI Yu-Hong, CHEN Jiong, GAO Shan-Shan, SHEN Guang-Qiang, LU Xin-Jiang, LI Ming-Yun. Cloning, physical and chemical property analysis of the Japanese sea bass Wap65-2 gene and its expression following Vibrio harveyi infection. Zoological Research, 2012, 33(5): 481-486. doi: 10.3724/SP.J.1141.2012.05481

Cloning, physical and chemical property analysis of the Japanese sea bass Wap65-2 gene and its expression following Vibrio harveyi infection

doi: 10.3724/SP.J.1141.2012.05481
  • Received Date: 2012-04-19
  • Rev Recd Date: 2012-08-21
  • Publish Date: 2012-10-10
  • The warm temperature acclimation related 65 kDa protein-2 (Wap65-2), a teleost plasma glycoprotein, plays an important role in immune regulation against bacterial infection. Here, for the first time we determined the full length cDNA sequence of the Japanese sea bass Wap65-2 gene (1 601 bp in length excluding the 3'-polyA tail). The sequence contains an open reading frame that encodes a protein of 436 amino acids with a molecular weight of 4.87?104. The predicted protein had a signal peptide in the N-terminal domain containing 19 residues. Sequence comparison and phylogenetic tree analysis showed that the Japanese sea bass Wap65-2 has a relatively high similarity to the Dicentrarchus labrax Wap65-2. In the healthy Japanese sea bass, Wap65-2 mRNA was expressed mainly in the liver and weakly in the heart and muscle. qRT-PCR results revealed that liver Wap65-2 transcripts were significantly increased after a Vibrio harveyi infection, and peaked 24 hour post injection (6.89 fold increase). The Japanese sea bass Wap65-2 protein was expressed in Escherichia coli and subsequently used for antiserum preparation. Western blot analysis showed that Wap65-2 was significantly increased in V. harveyi infected Japanese sea bass and reached a maximum of 5.33-fold increase at 36 h. In conclusion, the alteration of Japanese sea bass Wap65-2 expression was tightly associated with the progression of the V. harveyi bacterial infection.
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  • [1]
    Aliza D, Ismail IS, Kuah MK, Shu-Chien AC, Muhammad TST. 2008. Identification of Wap65, a human homologue of hemopexin as a copper-inducible gene in swordtail fish, Xiphophorus helleri [J]. Fish Physiol Biochem, 34(2): 129-138.
    [2]
    Cho YS, Kim BS, Kim DS, Nam YK. 2012. Modulation of warm-temperature-acclimation-associated 65-kDa protein genes (Wap65-1 and Wap65-2) in mud loach (Misgurnus mizolepis, Cypriniformes) liver in response to different stimulatory treatments [J]. Fish Shellfish Immunol, 32(5): 662-669.
    [3]
    Clark MS, Burns G. 2008. Characterisation of the warm acclimated protein gene (wap65) in the Antarctic plunderfish (Harpagifer antarcticus) [J]. DNA Seq, 19(1): 50-55.
    [4]
    Hirayama M, Kobiyama A, Kinoshita S, Watabe S. 2004. The occurrence of two types of hemopexin-like protein in medaka and differences in their affinity to heme [J]. J Exp Biol, 207(8): 1387-1398.
    [5]
    Hirayama M, Nakaniwa M, Ikeda D, Hirazawa N, Otaka T, Mitsuboshi T, Shirasu K, Watabe S. 2003. Primary structures and gene organizations of two types of Wap65 from the pufferfish Takifugu rubripes [J]. Fish Physiol Biochem, 29(3): 211-224.
    [6]
    Huang ZA, Chen J, Lu XJ, Shi YH, Li MY. 2011. Alteration on the expression of ayu coagulation factor X gene upon Listonella anguillarum infection [J]. Zool Res, 32(5): 492-498. [黄左安, 陈炯, 陆新江, 史雨红, 李明云. 2011. 香鱼凝血因子X 基因表达与鳗利斯顿氏菌感染的相关性. 动物学研究, 32(5): 492-498.]
    [7]
    Kong CJ, Huang ZA, Chen J, Shi YH, Lu XJ. 2012. Molecular cloning, sequence analysis and expression of ayu complement component C9 gene [J]. Zool Res, 33(2): 151-157. [孔铖将, 黄左安, 陈炯, 史雨红, 陆新江. 2012. 香鱼补体成分C9基因的克隆、序列分析及表达. 动物学研究, 33(2): 151-157.]
    [8]
    Li CH, Chen J, Shi YH, Li MY. 2009. Characterization of Listonella anguillarum as the aetiological agent of vibriosis occurred in cultured ayu (Plecoglossus altivelis) in Ninghai country, China [J]. Acta Microbiol Sin, 49(7): 931-937. [李长红, 陈炯, 史雨红, 李明云. 2009. 宁海地区香鱼弧菌病病原菌鉴定. 微生物学报, 49(7): 931-937.]
    [9]
    Li CH, Chen J, Shi YH, Lu XJ. 2011. Use of suppressive subtractive hybridization to identify differentially expressed genes in ayu (Plecoglossus altivelis) associated with Listonella anguillarum infection [J]. Fish Shellfish Immunol, 31(3): 500-506.
    [10]
    Livak KJ, Schmittgen TD. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method [J]. Methods, 25(4): 402-408.
    [11]
    Peatman E, Terhune J, Baoprasertkul P, Xu P, Nandi S, Wang SL, Somridhivej B, Kucuktas H, Li P, Dunham R, Liu ZJ. 2008. Microarray analysis of gene expression in the blue catfish liver reveals early activation of the MHC class I pathway after infection with Edwardsiella ictaluri [J]. Mol Immunol, 45(2): 553-566.
    [12]
    Sarropoulou E, Fernandes JM, Mitter K, Magoulas A, Mulero V, Sepulcre MP, Figueras A, Novoa B, Kotoulas G. 2010. Evolution of a multifunctional gene: the warm temperature acclimation protein Wap65 in the European seabass Dicentrarchus labrax [J]. Mol Phylogenet Evol, 55(2): 640-649.
    [13]
    Sha ZX, Xu P, Takano T, Liu H, Terhune J, Liu ZJ. 2008. The warm temperature acclimation protein Wap65 as an immune response gene: its duplicates are differentially regulated by temperature and bacteria infections [J]. Mol Immunol, 45(5): 1458-1469.
    [14]
    Shi YH, Chen J, Li CH, Li MY. 2010. Molecular cloning of liver Wap65 cDNA in ayu (Plecoglossus altivelis) and mRNA expression changes following Listonella anguillarum infection [J]. Mol Biol Rep, 37(3): 1523-1529.
    [15]
    Shi YH, Chen J, Li CH, Yang HY, Lu XJ. 2011. The establishment of a library screening method based on yeast two-hybrid system and its use to determine the potential interactions of liver proteins in ayu, Plecoglossus altivelis [J]. Fish Shellfish Immunol, 30(4-5): 1184-1187.
    [16]
    Shi YH, Chen J, Chang HL, Lu XJ, Zhang DM, Li HY, Zhao ZX, Li MY. 2012. Detection of bacterial pathogens in aquaculture samples by DNA microarray analysis [J]. Aquaculture, 338-341: 29-35.
    [17]
    Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. 2011. MEGA5: Molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods [J]. Mol Bio Evol, 28(10): 2731-2739.
    [18]
    Wang BK, Yu JH, Li Y, Ji SW, Xu HS. 2002. Isolation and identification of pathogen (Vibrio harveyi) from sea perch, Lateolabrax japonicus [J]. J Fish Sci Chn, 9(1): 52-55. [王保坤, 余俊红, 李筠, 纪伟尚, 徐怀恕. 2002. 花鲈弧菌病病原菌(哈维氏弧菌)的分离与鉴定. 中国水产科学, 9(1): 52-55.]
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