中华乌塘鳢(Bostrychus sinensis)白细胞介素8基因的克隆及免疫应答表达分析  

MOLECULAR CLONING AND EXPRESSION ANALYSES OF THE INTERLEUKIN-8 GENE FROM BOSTRYCHUS SINENSIS IN RESPONSE TO IMMUNE STIMULATION

作　　者：陈林杰 沈斌 张建设 CHEN Lin-Jie;SHEN Bin;ZHANG Jian-She(National Engineering Research Center for Marine Aquaculture,Zhejiang Ocean University,Zhoushan 316022,China)

机构地区：[1]浙江海洋大学、国家海洋设施养殖工程技术研究中心,浙江舟山316022

出　　处：《海洋与湖沼》2024年第2期499-508,共10页Oceanologia Et Limnologia Sinica

基　　金：国家自然科学基金项目,42106101号。

摘　　要：由细菌等感染引起的疾病对中华乌塘鳢(Bostrychus sinensis)养殖业的可持续发展造成严重威胁。白细胞介素8(Interleukin-8,IL-8)在调节炎症反应和抗感染免疫方面扮演着重要的作用。利用PCR扩增和克隆测序获得了中华乌塘鳢IL-8(命名为BsIL-8)cDNA序列。BsIL-8 cDNA序列全长1244 bp,包含576 bp的5’端非编码区序列(5’-UTR),312 bp的开放阅读框序列(ORF)和356 bp的3’端非编码区序列(3’-UTR)。BsIL-8共编码103个氨基酸,包含1个由18个氨基酸组成的信号肽和1个由62个氨基酸组成的SCY结构域。其中,SCY结构域包含1个CXC基序(Cys^(30)-Arg^(31)-Cys^(32))以及4个保守的半胱氨酸残基(Cys^(30)、Cys^(32)、Cys^(57)和Cys^(73))。序列分析结果表明,BsIL-8与大弹涂鱼(Boleophthalmus pectinirostris)的IL-8序列相似性和一致性最高(分别为92.4%和86.4%)。基因组织表达分布结果显示,BsIL-8基因在主要组织器官中呈泛在性表达,头肾BsIL-8基础表达量最高,肝脏次之,外周血最低。Poly(I:C)免疫刺激后,BsIL-8在4个重要免疫器官(脾脏、肝脏、头肾和外周血)中的表达量均发生了显著的上调(分别在刺激后3、12、24和24 h时表达量达到最高,P<0.01)。副溶血弧菌(Vibrio parahaemolyticus)感染后,BsIL-8基因在脾脏和外周血中的表达量均显著上调(均在感染后12 h时表达量达到最高,P<0.01),而在肝脏和头肾中的表达量均显著下调(均在感染后12 h时表达量达到最低,P<0.01)。综上所述,BsIL-8基因参与了中华乌塘鳢抗感染免疫应答过程。The sustainable development of Bostrychus sinensis aquaculture has been seriously impacted by bacterial infectious diseases.Interleukin-8(IL-8)plays an important role in anti-infective immunity and regulation of inflammatory responses.The cDNA sequence of B.sinensis IL-8(named BsIL-8)was obtained by PCR amplification and clonal sequencing.The cDNA sequence of BsIL-8 is 1244 bp in length,comprising a 5’-untranslated region(5’-UTR)of 576 bp,an open reading frame(ORF)of 312 bp,and a 3'-UTR of 356 bp.BsIL-8 encodes 103 amino acids,including an 18-amino acid signal peptide and a 62-amino acid SCY domain.The SCY domain contains a CXC motif(Cys^(30)-Arg^(31)-Cys^(32))and four conserved cysteine residues(Cys^(30),Cys^(32),Cys^(57),and Cys^(73)).Sequence analyses showed that BsIL-8 had the highest sequence similarity and identity with Boleophthalmus pectinirostris IL-8(92.4%and 86.4%,respectively).The RT-PCR results revealed that BsIL-8 was constitutively expressed in the 10 organs/tissues examined in B.sinensis.The head kidney exhibited the highest expression,followed by the liver,while the peripheral blood showed the lowest expression.Following immune stimulation with poly(I︰C),the expression of the BsIL-8 gene was significantly upregulated in the spleen,liver,head kidney,and peripheral blood of B.sinensis.The highest expression levels were observed at 3,12,24,and 24 h post-stimulation,respectively(P<0.01).After Vibrio parahaemolyticus infection,the expression of the BsIL-8 gene was significantly upregulated in the spleen and peripheral blood of B.sinensis,with the highest expression levels observed at 12 h post-infection(P<0.01).Conversely,the liver and head kidney exhibited significant downregulation of the gene,with the lowest expression levels detected at 12 h post-infection(P<0.01).In conclusion,the BsIL-8 gene is involved in the anti-infection immune responses of B.sinensis.

关 键 词：中华乌塘鳢(Bostrychus sinensis) 白细胞介素8(IL-8) 基因表达 免疫应答 副溶血弧菌 

分 类 号：S941[农业科学—水产养殖] S965[农业科学—水产科学] Q789[生物学—分子生物学]