Li Chaozheng, Li Haoyang, Xiao Bang, Chen Yonggui, Wang Sheng, Lǚ Kai, Yin Bin, Li Sedong, He Jianguo
State Key Laboratory for Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou, PR China; Institute of Aquatic Economic Animals and Guangdong Province Key Laboratory for Aquatic Economic Animals, Sun Yat-sen University, PR China; Guangdong Provincial Key Laboratory of Marine Resources and Coastal Engineering, PR China; School of Marine Sciences, Sun Yat-sen University, Guangzhou, PR China; South China Sea Resource Exploitation and Protection Collaborative Innovation Center (SCS-REPIC), PR China.
State Key Laboratory for Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou, PR China; Institute of Aquatic Economic Animals and Guangdong Province Key Laboratory for Aquatic Economic Animals, Sun Yat-sen University, PR China; Guangdong Provincial Key Laboratory of Marine Resources and Coastal Engineering, PR China.
Dev Comp Immunol. 2017 May;70:45-58. doi: 10.1016/j.dci.2017.01.005. Epub 2017 Jan 6.
Thioester-containing proteins (TEPs) are present in a wide range of species from deuterostomes to protostomes and are thought to be involved in innate immunity. In the current study, a TEP gene homologous to insect TEPs (iTEP) from the crustacean Litopenaeus vannamei, named LvTEP1, is cloned and functionally characterized. The open reading frame (ORF) of LvTEP1 is 4383 bp in length, encoding a polypeptide of 1460 amino acids with a calculated molecular weight of 161.1 kDa LvTEP1, which is most similar to other TEPs from insects, contains some conserved sequence features, including a N-terminal signal peptide, a canonical thioester (TE) motif, and a C-terminal distinctive cysteine signature. LvTEP1 is expressed in most immune-related tissues, such as intestine, epithelium, and hemocytes, and the mRNA level of LvTEP1 is upregulated in hemocytes after bacterial and viral challenges, indicating its involvement in the shrimp innate immune response. An expression assay in Drosophila S2 cells shows LvTEP1 to be a full-length secretory protein, and processed forms are present in the supernatant. Of note, only the processed form of LvTEP1 protein can bind to both the gram-negative bacterium Vibrio parahaemolyticus and the gram-positive bacterium Staphylococcus aureus in vitro, and its abundance can be induced after bacterial treatment. Moreover, knockdown of LvTEP1 renders shrimps more susceptible to both V. parahaemolyticus and S. aureus, as well as white spot syndrome virus (WSSV) infection, suggesting its essential defensive role against these invading microbes. We also observe that the expression of LvTEP1 is regulated in a manner dependent on both NF-κB and AP-1 transcription factors in naive shrimps and in vitro, suggesting that LvTEP1 could be poised in the body cavity prior to infection and thus play an important role in basal immunity. Taken together, our findings provide some in vitro and in vivo evidence for the involvement of LvTEP1 in shrimp innate immunity and provide some insight into its expression regulation mediated by multiple transcription factors or signaling pathways.
含硫酯蛋白（TEPs）存在于从后口动物到原口动物的广泛物种中，被认为参与先天免疫。在本研究中，从甲壳类凡纳滨对虾中克隆了一个与昆虫TEPs（iTEPs）同源的TEP基因，命名为LvTEP1，并对其进行了功能表征。LvTEP1的开放阅读框（ORF）长度为4383 bp，编码一个1460个氨基酸的多肽，计算分子量为161.1 kDa。LvTEP1与昆虫的其他TEPs最为相似，包含一些保守的序列特征，包括一个N端信号肽、一个典型的硫酯（TE）基序和一个C端独特的半胱氨酸特征。LvTEP1在大多数免疫相关组织中表达，如肠道、上皮组织和血细胞，在细菌和病毒攻击后，血细胞中LvTEP1的mRNA水平上调，表明其参与对虾先天免疫反应。在果蝇S2细胞中的表达分析表明，LvTEP1是一种全长分泌蛋白，其加工形式存在于上清液中。值得注意的是，只有加工形式的LvTEP1蛋白在体外能与革兰氏阴性菌副溶血性弧菌和革兰氏阳性菌金黄色葡萄球菌结合，并且在细菌处理后其丰度可以被诱导。此外，敲低LvTEP1使对虾对副溶血性弧菌、金黄色葡萄球菌以及白斑综合征病毒（WSSV）感染更敏感，表明其对这些入侵微生物具有重要的防御作用。我们还观察到，在未感染的对虾体内和体外，LvTEP1的表达以一种依赖于NF-κB和AP-1转录因子的方式受到调控，这表明LvTEP1可能在感染前在体腔中待命，从而在基础免疫中发挥重要作用。综上所述，我们的研究结果为LvTEP1参与对虾先天免疫提供了一些体外和体内证据，并为其由多种转录因子或信号通路介导的表达调控提供了一些见解。
