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黄颡鱼中鹅型溶菌酶(LysG)基因的分子鉴定与表达分析

Molecular identification and expression analysis of a goose-type lysozyme (LysG) gene in yellow catfish Pelteobagrus fulvidraco.

作者信息

Liu Qiu-Ning, Xin Zhao-Zhe, Zhang Dai-Zhen, Jiang Sen-Hao, Chai Xin-Yue, Li Chao-Feng, Zhou Chun-Lin, Tang Bo-Ping

机构信息

Jiangsu Key Laboratory for Bioresources of Saline Soils, Jiangsu Synthetic Innovation Center for Coastal Bio-agriculture, Jiangsu Provincial Key Laboratory of Coastal Wetland Bioresources and Environmental Protection, School of Ocean and Biological Engineering, Yancheng Teachers University, Yancheng 224051, PR China.

Jiangsu Key Laboratory for Bioresources of Saline Soils, Jiangsu Synthetic Innovation Center for Coastal Bio-agriculture, Jiangsu Provincial Key Laboratory of Coastal Wetland Bioresources and Environmental Protection, School of Ocean and Biological Engineering, Yancheng Teachers University, Yancheng 224051, PR China.

出版信息

Fish Shellfish Immunol. 2016 Nov;58:423-428. doi: 10.1016/j.fsi.2016.09.034. Epub 2016 Sep 16.

Abstract

Lysozymes, innate immunity molecules, play a vital role in immune response to pathogens. The yellow catfish Pelteobagrus fulvidraco (Siluriformes: Bagridae) is an economically important fish in China. The aim of this study was to quantify expression of the P. fulvidraco LysG gene (a g-type lysozyme) in response to pathogen-associated molecular patterns (PAMP) challenge. First, the P. fulvidraco LysG gene (PfLysG) was cloned and characterized. The full-length cDNA of PfLysG is 1323 bp, including a 5'-untranslated region (UTR) of 131 bp, a 3'-UTR of 634 bp, and an open reading frame of 558 bp encoding a polypeptide of 185 amino acids, which contains a transglycosylase SLT domain (Pfam01464). The predicted molecular weight of the protein is 20.52 kDa with a pI of 9.08. Two catalytic residues and seven N-acetyl-D-glucosamine binding sites are conserved in the sequence and there is no predicted signal peptide. The deduced PfLysG protein sequence has 84%, 76% and 69% percent identity with the LysGs from Ictalurus furcatus, Danio rerio, and Salmo salar, respectively. The predicted tertiary structure of PfLysG is very similar to that from other animals. Phylogenetic analysis showed that PfLysG is closely related to those from Teleostei. Real-time quantitative reverse transcription-PCR (qPCR) analysis showed that PfLysG was expressed in all examined tissues and most highly expressed in head kidney, spleen, and intestine. After simulated pathogen challenge with lipopolysaccharide and polyriboinosinic polyribocytidylic acid, respectively, the mRNA expression of PfLysG was upregulated significantly at different time points. The results suggest that the identified g-type lysozyme of P. fulvidraco is involved in innate immune responses.

摘要

溶菌酶作为天然免疫分子,在对病原体的免疫反应中发挥着至关重要的作用。黄颡鱼(鲇形目:鲿科)是中国一种具有重要经济价值的鱼类。本研究的目的是量化黄颡鱼LysG基因(一种g型溶菌酶)在病原体相关分子模式(PAMP)刺激下的表达情况。首先,对黄颡鱼LysG基因(PfLysG)进行了克隆和表征。PfLysG的全长cDNA为1323 bp,包括131 bp的5'非翻译区(UTR)、634 bp的3'UTR和558 bp的开放阅读框,编码一个由185个氨基酸组成的多肽,该多肽包含一个转糖基酶SLT结构域(Pfam01464)。预测该蛋白质的分子量为20.52 kDa,pI为9.08。序列中两个催化残基和七个N - 乙酰 - D - 葡萄糖胺结合位点保守,且无预测的信号肽。推导的PfLysG蛋白序列与斑点叉尾鮰、斑马鱼和大西洋鲑的LysG分别具有84%、76%和69%的同一性。PfLysG的预测三级结构与其他动物的非常相似。系统发育分析表明,PfLysG与硬骨鱼纲的关系密切。实时定量逆转录PCR(qPCR)分析表明,PfLysG在所有检测组织中均有表达,并在头肾、脾脏和肠道中表达最高。分别用脂多糖和聚肌苷酸 - 聚胞苷酸进行模拟病原体刺激后,PfLysG的mRNA表达在不同时间点均显著上调。结果表明,所鉴定的黄颡鱼g型溶菌酶参与天然免疫反应。

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