Characterization of Toll‐like receptor 22 in turbot (Scophthalmus maximus)
Characterization of Toll‐like receptor 22 in turbot (Scophthalmus maximus)
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DOI：
10.1016/j.fsi.2017.05.025
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发表时间：
2017-07
影响因子：
4.7
通讯作者：
J. Xing;Xiujuan Zhou;Xiaoqian Tang;X. Sheng;W. Zhan
中科院分区：
文献类型：
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作者：
J. Xing;Xiujuan Zhou;Xiaoqian Tang;X. Sheng;W. Zhan
toll样受体（TLRs）对激活先天免疫系统以应对入侵的病原体至关重要。本文采用RT-PCR和原位杂交技术对大菱鲆TLR22基因（tbTLR22）的表达谱进行了分析。然后用配体或病原体模拟其表达模式。大比目鱼体内注射了iniptococcus， wardsiella tarda, Hirame rhabdovirus virus (HIRRV), polyinosinic: polycytidylic acid (Poly I:C), peptide - glycan (PGN), lipopolaccharides (LPS)；在培养的外周血白细胞（PBL）中加入poly I:C、PGN或LPS；然后用定量PCR法检测肝脏、脾脏、鳃、肾脏和培养PBL中tbTLR22的表达。制备tbTLR22重组蛋白（rp-tbTLR22）及其抗体，Western-blotting检测抗体与组织的反应，ELISA检测rp-tbTLR22与所有刺激物的结合情况。结果显示，PolyI: C显著上调tbTLR22的表达，而PGN和LPS组tbTLR22的表达变化不显著；肝脏和脾脏tbTLR22显著升高。感染最多3.6次，感染3.3次；在肝脏和肾脏。病毒感染最多3.4次，最多4.1次；其次是鳃部和肾脏，分别是hrv感染后的4.8倍和4.1倍。Rp-tbTLR22抗体能分别识别40 kDa、90 kDa和120 kDa的肝脏、肾脏、鳃和脾脏总蛋白。rp-tbTLR22在体外可与3种配体和病原体结合。表达和反应数据为tbTLR22建立了清晰的识别模型。
Toll-like receptors (TLRs) are essential for activation of the innate immune system in response to invading pathogens. In this paper, expression profiles of the turbot (Scophthalmus maximus) TLR22 gene (tbTLR22) were analyzed with RT-PCR andin situhybridization. Then its expression patterns simulated with ligands or pathogens were investigated. Streptococcus iniae, Edwardsiella tarda, Hirame rhabdovirus virus (HIRRV), polyinosinic: polycytidylic acid (Poly I:C), peptidoglycan (PGN), or lipopolysaccharides (LPS) was injected to turbot; poly I:C, PGN, or LPS was added into cultured peripheral blood leukocytes (PBL); and then the tbTLR22 in liver, spleen, gill, kidney and cultured PBL was measured using Quantitative PCR. The recombinant protein of tbTLR22 (rp-tbTLR22) and its antibody were produced, then the reactions of antibody to tissues were detected by Western-blotting, and the binding of rp-tbTLR22 to all the stimulants was detected using ELISA. The results showed tbTLR22 expression was significantly up-regulated by PolyI: C, but no significant change in PGN and LPS groups; tbTLR22 significantly increased in liver and spleen afterS.iniaeinfection with the maximum of 3.6 times and 3.3 times; in liver and kidney afterE. tardainfection with the maximum of 3.4 times and 4.1 times; and then in gill and kidney after HIRRV infection by 4.8 and 4.1 times. Rp-tbTLR22 antibody could recognize the total protein from liver, kidney, gill and spleen at 40 kDa, 90 kDa and 120 kDa, respectively. The rp-tbTLR22 could bind to three ligands and pathogensin vitro. The expression and reaction data gave a clear recognization model of tbTLR22.