文章摘要
李健,孟繁星,黎明,王日昕,石戈.大弹涂鱼白细胞介素-8基因分子进化及不同病原体刺激对其表达的影响[J].水产学报,2019,43(11):2290~2303
大弹涂鱼白细胞介素-8基因分子进化及不同病原体刺激对其表达的影响
Molecular evolution analysis of interleukin-8 in great blue-spotted mudskipper (Boleophthalmus pectinirostris) and the effect of different pathogen stimulus on its expression
投稿时间:2018-11-22  修订日期:2019-03-17
DOI:10.11964/jfc.20181111545
中文关键词: 大弹涂鱼  白细胞介素-8  分子进化分析  表达分析  ELR基序
英文关键词: Boleophthalmus pectinirostris  interleukin-8  molecular evolutionary analysis  ELR motif  expression analysis
基金项目:国家自然科学基金(31872222);宁波市自然科学基金(2016A610083)
作者单位E-mail
李健 浙江海洋大学海洋科学与技术学院, 浙江 舟山 316000  
孟繁星 宁波大学海洋学院, 浙江 宁波 315000  
黎明 宁波大学海洋学院, 浙江 宁波 315000  
王日昕 宁波大学海洋学院, 浙江 宁波 315000  
石戈 浙江海洋大学海洋科学与技术学院, 浙江 舟山 316000 sg2610105@126.com 
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中文摘要:
      实验从大弹涂鱼皮肤组织转录组中筛选出大弹涂鱼IL-8基因,并分析了其序列特征,构建了系统发生树,评估了IL-8基因在健康个体不同组织中的表达差异,以及在不同病原体刺激下IL-8基因在机体主要免疫器官肝脏和脾脏中的表达情况。结果显示,该基因的开放阅读框由306个碱基组成,编码101个氨基酸残基,包含典型的由18个氨基酸残基组成的信号肽,和1个由62个氨基酸残基组成的SCY结构域,该结构域还包含了4个保守的半胱氨酸残基,分别为Cys-30、Cys-32、Cys-57和Cys-73。大弹涂鱼IL-8氨基酸序列中的受体结合位点基序ELR (Glu-Leu-Arg)被Asn-Ser-His (NSH)取代,系统进化分析表明,纯化选择影响了鱼类该基序的多样性,且IL-8基因在大弹涂鱼乃至硬骨鱼中不可取代。荧光定量实验结果显示,IL-8基因在大弹涂鱼的健康组织中广泛表达,在鳃和脑中表达量最高。细菌和poly(I∶C)注射实验表明,在受到感染后,肝、脾和脑组织中IL-8的表达量均上调,说明IL-8基因在大弹涂鱼肝、脾和脑组织的炎性反应和免疫应答中起到了重要作用,为大弹涂鱼免疫基因的后续研究提供了参考。
英文摘要:
      Great blue-spotted mudskippers (Boleophthalmus pectinirostris) have lived in mudflats along the coasts of Japan and South China for a long time. Compared with the fish living in water, B. pectinirostris, as an amphibious fish, is more vulnerable to terrestrial and aquatic pathogenic factors. Interleukin-8 is a CXC chemokine that plays an important role in regulating the inflammatory response. In this study, we screened the interleukin-8 gene from skin transcriptome of B. pectinirostris, then cloned and verified its coding region. The results indicated that the open reading frame(ORF) of B. pectinirostris IL-8 gene was 306 bp, coding 101 amino acids, including a typical signal peptide of 18 amino acids, and a SCY domain of 62 amino acids, which also possessed the four conserved cysteine residues, namely Cys-30, Cys-32, Cys-57 and Cys-73. Like most fish, B. pectinirostris interleukin-8 amino acid sequence lacked the ELR motif which was the receptor binding site and it was replaced by Asn-Ser-His (NSH) in great blue-spotted mudskipper. In all known fish amino acid sequences, rules of this motif were as follows: the first amino acid was never the amino acid with positive charge, the second amino acid was generally those amino acids with no charge, and most of third amino acid were amino acids with positive charge. Phylogenetic analysis showed that purification selection affected the diversity of this motif in fish. Tissue expression analysis by RT-qPCR indicated that interleukin-8 gene was widely expressed in healthy tissues of B. pectinirostris and it had the highest expression in the gill and brain. Bacteria and poly (I∶C) injection experiments showed that expression of IL-8 in liver, spleen and brain tissues was up-regulated after the infection, indicating that IL-8 plays an important role in the inflammatory response and immune response of liver, spleen and brain tissues. This study provided important reference for the research of immune genes of B. pectinirostris, thus having more comprehensive understanding of the evolution process and function of fish immune genes.
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