生物学杂志

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半滑舌鳎(Cynoglossus semilaevis)CD80的基因表达以及重组CD80与外周血淋巴细胞的结合分析

  

  1. 1.中国科学院海洋研究所 海洋生物学重点实验室, 青岛 266071;2. 中国科学院大学, 北京 100049
  • 出版日期:2018-04-18 发布日期:2018-04-18
  • 通讯作者: 孙黎, 研究员,主要研究方向为病原性细菌和病毒的致病机制、鱼类免疫与抗感染机制、细菌和病毒性疫病的免疫防控、深海极端环境微生物生命机制与开发利用等,E-mail: lsun@qdio.ac.cn
  • 作者简介:陈晨, 硕士研究生,主要研究方向为鱼类病害防治,E-mail:kaxcc@126.com
  • 基金资助:
    青岛海洋国家实验室鳌山科技创新计划项目课题“近海健康养殖关键技术研究与新生产模式构建”(No.2015ASKJ02)

Analysis of the expression of Cynoglossus semilaevis CD80 and the interaction between recombinant CD80 and peripheral blood lymphocytes

  1. 1. Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences,Qingdao 266071; 2. University of Chinese Academy of Sciences, Beijing 100049, China
  • Online:2018-04-18 Published:2018-04-18

摘要: CD80属于免疫球蛋白超家族,是T细胞增殖、活化过程中一种重要的共刺激信号。从半滑舌鳎(Cynoglossus semilaevis)全基因组数据库中获得CD80基因(命名为CsCD80)序列(GenBank登录号:XR_521517.2),开放阅读框区域含有882个碱基,共编码293个氨基酸,其理论分子质量为32.45 ku,理论等电点为7.97。序列分析显示,CsCD80含有一个N-末端信号肽序列,一个免疫球蛋白(Ig)结构域和跨膜(TM)结构域。序列多重比对结果显示, CsCD80与牙鲆CD80相似性最高,为41.37%。实时荧光定量PCR结果显示,CsCD80基因在半滑舌鳎肝、脾中有高表达量,哈维氏弧菌和细胞肿大病毒刺激后,CsCD80基因在肝、脾和肾中的表达量均有显著上调。使用原核表达系统表达CsCD80重组蛋白(rCsCD80)并与半滑舌鳎外周血淋巴细胞(PBL)孵育,经过免疫荧光检测显示rCsCD80能结合到PBL表面。这些研究结果表明,CsCD80可能在半滑舌鳎抗细菌和病毒感染的免疫进程中发挥作用。

关键词: 半滑舌鳎, CD80, 表达量分析, 外周血淋巴细胞

Abstract: CD80 is a member of immunoglobulin superfamily, which is an important costimulatory signal necessary for T cell proliferation and activation. In this study, CD80 gene sequence of Cynoglossus semilaevis(CsCD80)(GenBank accession number: XR_521517.2) was obtained from its whole genome sequence. The open reading frame contains 882 bp encoding 293 amino acids with an estimated molecular weight of 32.45 ku and a theoretical pI of 7.97. Amino acid sequence alignment indicated that CsCD80 contained a predicted signal peptide, an immunoglobulin(Ig) domain and a transmembrane(TM) domain. Multiple sequence alignment showed that CsCD80 shared the highest amino acid sequence identity(41.37%) with Japanese flounder CD80 . Quantitative real-time PCR analysis showed that CsCD80 had the highest expression level in liver and spleen. When infected with V. harveyi and megalocytivirus, the expression level of CsCD80 significantly incresed in liver, spleen and kidney. Immunofluorescence test was used for the determination of the recombinant CsCD80 protein(rCsCD80) binding to the surface of peripheral blood lymphocytes(PBLs) when incubated with recombinant CsCD80 protein(rCsCD80) expressed by prokaryotic expression system. These results indicated that CsCD80 might play an important role in the immune response against bacteria and virus in C. semilaevis.

Key words: C. semilaevis, CD80, expression analysis, peripheral blood lymphocytes