热带海洋学报 ›› 2022, Vol. 41 ›› Issue (1): 42-51.doi: 10.11978/2021029

• 海洋生物学 • 上一篇    下一篇

香港牡蛎(Crassostrea hongkongensis)新型凝集素ChPerlucin的基因克隆与功能研究

赵泽慧1,2(), 张爱娇1,2, 杨雨澄1,2, 毛帆1, 肖述1, 李军1, 张扬1, 向志明1(), 喻子牛1()   

  1. 1. 中国科学院热带海洋生物资源与生态重点实验室(中国科学院南海海洋研究所), 广东 广州 510301
    2. 中国科学院大学, 北京100049
  • 收稿日期:2021-03-04 修回日期:2021-04-19 出版日期:2022-01-10 发布日期:2021-04-25
  • 通讯作者: 向志明,喻子牛,姚衍桃
  • 作者简介:赵泽慧(1993—), 女, 河北人, 硕士研究生, 主要从事贝类免疫与抗逆的研究。email: zhaozehui18@mails.ucas.ac.cn
  • 基金资助:
    广东省自然科学基金项目(2021A1515010541)

Molecular cloning and functional studies of ChPerlucin in Crassostrea hongkongensis

ZHAO Zehui1,2(), ZHANG Aijiao1,2, YANG Yucheng1,2, MAO Fan1, XIAO Shu1, LI Jun1, ZHANG Yang1, XIANG Zhiming1(), YU Ziniu1()   

  1. 1. CAS Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Science, Guangzhou 510301, China
    2. University of Chinese Academy of Sciences, Beijing 100049, China
  • Received:2021-03-04 Revised:2021-04-19 Online:2022-01-10 Published:2021-04-25
  • Contact: XIANG Zhiming,YU Ziniu,YAO Yantao
  • Supported by:
    Natural Science Foundation of Guangdong Province, China(2021A1515010541)

摘要:

C-型凝集素(C-type lectin)是一类能识别并结合多糖的免疫蛋白质超家族, 本文首次克隆了一种新型香港牡蛎C-型凝集素基因——甘露聚糖结合凝集素, 并获得了该基因的全长序列, 命名为ChPerlucin。结果显示, ChPerlucin基因cDNA全长577bp, 包括21bp的5′末端非翻译区(untranslated region, UTR)、73bp的 3′UTR, 以及483bp的开放阅读框(open reading frame, ORF), 编码160个氨基酸, ChPerlucin蛋白理论分子量为18kD, 等电点为5.95。ChPerlucin蛋白含有保守的C-型凝集素样结构域(C-type lectin or carbohydrate-recognition domain, CLECT); 邻接法(Neighbor-Joining, NJ)进化树分析表明, ChPerlucin与其他贝类的Perlucin聚为一支, 说明该基因是软体动物Perlucin家族的新成员。qRT-PCR结果显示, ChPerlucin在香港牡蛎所检测组织和各个胚胎发育时期中均有表达; 病原菌刺激后, ChPerlucin表达量显著升高; 利用毕赤酵母真核成功表达了ChPerlucin重组蛋白, 发现ChPerlucin重组蛋白具有抑菌效果。以上结果表明ChPerlucin在香港牡蛎的先天性免疫过程中发挥了重要的作用。

关键词: 香港牡蛎, 先天性免疫, C-型凝集素, ChPerlucin

Abstract:

The C-type lectin superfamily is a class of immune proteins that can recognize and bind to polysaccharides. In this study, a new type C-type lectin gene of Crassostrea hongkongensis was cloned for the first time, and the full-length sequence of the gene was obtained. The results show that the full length of ChPerlucin gene cDNA is 577 bp, including 5′-untranslated region (UTR) in 21 bp, 3′-UTR in 73 bp, and an open reading frame (ORF) in 483 bp open reading that code 160 amino-acid polypeptide. The predicted molecular mass is 18kD, and the isoelectric point is 5.95. Amino acid sequence alignment showed that the Perlucin gene of Crassostrea hongkongensis contains a conserved carbohydrate recognition domain (CLECT). Neighbor-Joining (NJ) evolutionary tree analysis showed that ChPerlucin clustered with Perlucin from other shellfish, indicating that this gene is a new member of the Perlucin family of mollusks. The results of qRT-PCR showed that ChPerlucin was broadly expressed in various tissues and during different stages of the oyster’s embryonic and larval development; the expression of ChPerlucin increased significantly after bacteria stimulation; the recombinant protein ChPerlucin was successfully expressed using Pichia pastoris eukaryotic; and we find that ChPerlucin recombinant protein has antibacterial effect. The above results indicate that ChPerlucin plays an important role in the innate immunity of Crassostrea hongkongensis.

Key words: Crassostrea hongkongensis, innate immunity, C-type lectin, ChPerlucin

中图分类号: 

  • S944.4