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东北七鳃鳗(Lethenteron morii)MyD88:鉴定、表达及功能特征分析。

Northeast Chinese lamprey (Lethenteron morii) MyD88: Identification, expression, and functional characterization.

机构信息

Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources, Ministry of Education, Shanghai Ocean University, Shanghai, 201306, China; International Research Center for Marine Biosciences, Ministry of Science and Technology, Shanghai Ocean University, Shanghai, 201306, China; Key Laboratory of Freshwater Aquatic Genetic Resources, Ministry of Agriculture, Shanghai Ocean University, Shanghai, 201306, China.

Department of Fisheries and Wildlife, Michigan State University, East Lansing, MI, 48824, USA.

出版信息

Fish Shellfish Immunol. 2019 Nov;94:539-547. doi: 10.1016/j.fsi.2019.09.035. Epub 2019 Sep 15.

Abstract

Myeloid differentiation factor 88 (MyD88) is a key adaptor of Toll-like receptors (TLR), an important pattern recognition receptor of the innate immune system. To study the origin and evolution of the vertebrate TLR signaling pathway in innate immune systems, we analyzed the biological characteristics and functions of the MyD88 gene in Northeast Chinese lamprey (Lethenteron morii) using PCR amplification, real-time PCR analysis, dual luciferase reporter gene assay, immunofluorescence assay, and other methods. Bioinformatics analysis showed that LmMyD88 has a modular structure consisting of Toll/IL-1R domain (TIR) and death domain (DD), which is typical of the MyD88 family. A phylogenetic tree showed that the homology of LmMyD88 was consistent with the phylogenetic status of lampreys. Tissue expression analysis indicated that the mRNA expression was expressed in some normal tissues of larval and adult L. morii. Real-time PCR analysis showed that the expression of LmMyD88 in tissues, such as gill and kidney, of the adult increased significantly after infection by Pseudomonas aeruginosa. Subcellular localization results showed that LmMyD88 was expressed in the nucleus, cytoplasm, and other parts. A dual luciferase reporter assay indicated that LmMyD88 activated nuclear factor kappa B downstream of the TLR signaling pathway. This study suggested that LmMyD88 might play an important role in the innate immune signal transduction process of L. morii.

摘要

髓样分化因子 88(MyD88)是 Toll 样受体(TLR)的关键衔接子,是先天免疫系统中重要的模式识别受体。为了研究脊椎动物 TLR 信号通路在先天免疫系统中的起源和进化,我们采用 PCR 扩增、实时荧光定量 PCR 分析、双荧光素酶报告基因检测、免疫荧光检测等方法,分析了东北七鳃鳗(Lethenteron morii)MyD88 基因的生物学特性和功能。生物信息学分析表明,LmMyD88 具有由 Toll/IL-1R 结构域(TIR)和死亡结构域(DD)组成的模块化结构,这是典型的 MyD88 家族结构。系统进化树分析表明,LmMyD88 与七鳃鳗的系统进化地位一致。组织表达分析显示,LmMyD88 在幼鳗和成鳗的一些正常组织中均有表达。实时荧光定量 PCR 分析表明,在成鳗的鳃和肾脏等组织中,感染铜绿假单胞菌后 LmMyD88 的表达显著增加。亚细胞定位结果表明,LmMyD88 表达于细胞核、细胞质等部位。双荧光素酶报告基因检测表明,LmMyD88 可激活 TLR 信号通路下游的核因子 κB。本研究表明,LmMyD88 可能在七鳃鳗的先天免疫信号转导过程中发挥重要作用。

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