Molecular cloning and characterization of the cathepsin L gene in Pelodiscus sinensis and its expression in response to bacterial challenge
Molecular cloning and characterization of the cathepsin L gene in Pelodiscus sinensis and its expression in response to bacterial challenge
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中华鳖组织蛋白酶 L 基因的分子克隆和表征及其响应细菌攻击的表达
DOI：
10.1111/are.13768
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发表时间：
2018
影响因子：
2
通讯作者：
Ji Xiang
中科院分区：
文献类型：
--
作者：
Chen Lian;Liang Shi Yuan;Nian Rui;Li Hong;Li Peng;Qu Yan Fu;Wu Ting;Meng Qing Guo;Ji Xiang
组织蛋白酶L是启动蛋白质降解的最重要的溶酶体半胱氨酸蛋白酶之一，它参与许多脊椎动物的免疫反应。然而，它在海龟先天免疫系统中的作用仍然知之甚少。本研究从中国鳖中克隆了组织蛋白酶L基因。然后，我们检测了嗜水气单胞菌和副溶血性弧菌感染后不同组织和不同时间点的mRNA表达。全长cDNA序列克隆自mp。sinensis全长1805 bp，开放阅读框全长1071 bp，编码356个氨基酸的40.39 kDa多肽。推断出的蛋白序列包含一个活跃的Cys、His和Asn三联体，以及保守的ERWNIN和GNFD基序。同源性分析表明，推导出的氨基酸序列与其他已知物种的同源性为77% ~ 96%。序列比对、系统发育分析和结构比较表明，组织蛋白酶L是组织蛋白酶家族的成员。Real - time PCR分析显示，海龟组织蛋白酶L mRNA在各种组织中普遍表达，且在肝脏中的表达水平高于其他组织。肝内表达量在a后24小时达到峰值。2次（12和72小时）。parahemolyticusinfection。这些结果表明，p的组织蛋白酶L基因。中华念珠菌可能参与了对a的抗菌反应过程。hydrophilaandV。parahemolyticus。该研究对进一步探索p抗菌免疫反应的分子机制具有重要意义。制成。
Cathepsin L is one of the most important lysosomal cysteine proteases for the initiation of protein degradation, and it is involved in the immune response in many vertebrates. However, its function in the innate immune system of turtles remains poorly understood. Here, we cloned the cathepsin L gene from the Chinese soft‐shelled turtlePelodiscus sinensis. We then examined the mRNA expression in different tissues and at different time points after infection byAeromonas hydrophilaorVibrio parahemolyticus. The full‐length cDNA sequence cloned fromP. sinensiswas 1,805 bp with a 1,071 bp open reading frame, which encodes a 40.39 kDa polypeptide of 356 amino acids. The deduced protein sequence contained an active triad of Cys, His and Asn, and conserved ERWNIN and GNFD motifs. Homology analysis showed that the deduced amino acid sequence shared 77%–96% identity with other known species. Sequence alignment, phylogenetic analysis and structural comparisons revealed that cathepsin L is a member of the cathepsin family. Real‐time PCR analyses showed that turtle cathepsin L mRNA is ubiquitously expressed in various tissues, and the expression level is higher in the liver than in other tissues. The expression level in the liver peaks 24 hr afterA. hydrophilainfection and at two times (12 and 72 hr) afterV. parahemolyticusinfection. These results suggest that the cathepsin L gene ofP. sinensisis likely involved in the process of the antibacterial response toA. hydrophilaandV. parahemolyticus. This study is of great importance for future work exploring the molecular mechanism of antibacterial immune responses inP. sinensis.