College of Marine Life and Fisheries, Huaihai Institute of Technology, 59 Cangwu Road, Lianyungang 222005, China; Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Huaihai Institute of Technology, Lianyungang 222005, China.
Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266000, China.
Fish Shellfish Immunol. 2017 Dec;71:43-49. doi: 10.1016/j.fsi.2017.09.065. Epub 2017 Sep 28.
Chitinase, belonging to family 18 glycosyl hydrolase, is a multi-gene family and it has many functions. Generation of loss-of-function mutant targeting an interesting gene is a common way to clarify its function based on reverse genetics. In this study, we first reported the immune defense of a chitinase gene (EcChi4) in Exopalaemon carinicauda using its EcChi4-deletion mutant. EcChi4 was predominantly expressed in hepatopancreas and was upregulated after challenge with Vibrio parahaemolyticus or Aeromonas hydrophila. After knockout EcChi4 gene using CRISPR/Cas9 tool, the prawns in EcChi4-deletion group had significant higher mortality than those in wild-type group when the prawns were challenged with V. parahaemolyticus or A. hydrophila. In conclusion, we first demonstrate the function of a chitinase gene in immune defense of E. carinicauda by performing directed, heritable gene mutagenesis. In the future, CRISPR/Cas9 should be widely applicable as a feasible means for gene editing in E. carinicauda for the study of important biological questions that cannot be easily addressed in other decapods.
几丁质酶属于 18 家族糖苷水解酶，是一个多基因家族，具有多种功能。基于反向遗传学，针对感兴趣的基因生成功能丧失突变体是阐明其功能的常用方法。在这项研究中，我们首次利用对虾几丁质酶基因（EcChi4）的 EcChi4 缺失突变体报道了其在凡纳滨对虾中的免疫防御作用。EcChi4 在肝胰腺中表达丰度最高，并在受到副溶血弧菌或嗜水气单胞菌刺激后上调表达。使用 CRISPR/Cas9 工具敲除 EcChi4 基因后，EcChi4 缺失组虾在受到副溶血弧菌或嗜水气单胞菌刺激时的死亡率显著高于野生型组虾。总之，我们通过定向、可遗传的基因诱变首次证明了 EcChi4 基因在凡纳滨对虾免疫防御中的功能。在未来，CRISPR/Cas9 应该作为一种可行的基因编辑手段在凡纳滨对虾中广泛应用，用于研究在其他十足目动物中难以解决的重要生物学问题。
