Li Ling, Qiu Limei, Song Linsheng, Song Xiaoyan, Zhao Jianmin, Wang Lingling, Mu Changkao, Zhang Huan
The Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Rd, Qingdao 266071, China.
Fish Shellfish Immunol. 2009 Nov;27(5):625-32. doi: 10.1016/j.fsi.2009.07.009. Epub 2009 Jul 25.
Tumor necrosis factor receptors (TNFRs) are a superfamily of proteins characterized by the unique cysteine-rich domain (CRD) and their important roles in diverse physiological and pathological events such as inflammation, apoptosis, autoimmunity and organogenesis. The first member of the molluscan TNFR family, designated as CfTNFR, was identified from Zhikong scallop Chlamys farreri by expressed sequence tag (EST) and rapid amplification of cDNA ends (RACE) approaches. The full-length cDNA of CfTNFR was of 1334 bp, consisting of a 5' UTR of 17 bp, a 3' UTR of 69 bp with a poly (A) tail, and an open reading frame (ORF) of 1248 bp encoding a polypeptide of 415 amino acids with a theoretical isoelectric point of 8.33 and predicted molecular weight of 47.07 kDa. There were a signal peptide, a CRD, a transmembrane region and a death domain in the deduced amino acid sequence of CfTNFR, suggesting that it was a typical type I membrane protein. The high identities (22-40%) of CfTNFR with other TNFR superfamily members indicated that CfTNFR should be a member of TNFR superfamily, and moreover, it should be the first death domain-containing TNFR found in invertebrates. Phylogenetic analysis revealed that CfTNFR was closely related to TNFR-like proteins from Strongylocentrotus purpuratus, Drosophila melanogaster and Ciona intestinalis, and they formed a separate branch apart from vertebrate TNFRs. The spatial expression of CfTNFR transcripts in healthy and bacteria challenged scallops was examined by quantitative real-time PCR. CfTNFR transcripts could be detected in all tested tissues, including haemocytes, gonad, gill, mantle and hepatopancreas, and significantly up-regulated in the tissues of gonad, gill, mantle and hepatopancreas after Listonella anguillarum challenge, indicating that CfTNFR was constitutive and inducible acute-phase protein involved in immune defence. The present results suggested the existence of the TNFR-like molecules and TNF-TNFR system in low invertebrates, and provided new insights into the role of CfTNFR in scallop innate immune responses to invading microorganisms.
肿瘤坏死因子受体（TNFRs）是一类蛋白质超家族，其特征在于独特的富含半胱氨酸结构域（CRD），并且在多种生理和病理过程中发挥重要作用，如炎症、细胞凋亡、自身免疫和器官发生。通过表达序列标签（EST）和cDNA末端快速扩增（RACE）方法，从栉孔扇贝（Chlamys farreri）中鉴定出软体动物TNFR家族的首个成员，命名为CfTNFR。CfTNFR的全长cDNA为1334 bp，由17 bp的5'非翻译区（UTR）、69 bp的3'UTR和一个带有poly（A）尾的开放阅读框（ORF）组成，该ORF为1248 bp，编码一个由415个氨基酸组成的多肽，理论等电点为8.33，预测分子量为47.07 kDa。在CfTNFR的推导氨基酸序列中有一个信号肽、一个CRD、一个跨膜区和一个死亡结构域，表明它是一种典型的I型膜蛋白。CfTNFR与其他TNFR超家族成员具有较高的同源性（22 - 40%），这表明CfTNFR应该是TNFR超家族的成员，而且它应该是在无脊椎动物中发现的首个含有死亡结构域的TNFR。系统发育分析表明，CfTNFR与紫海胆（Strongylocentrotus purpuratus）、黑腹果蝇（Drosophila melanogaster）和玻璃海鞘（Ciona intestinalis）的TNFR样蛋白密切相关，它们形成了一个独立于脊椎动物TNFRs的分支。通过定量实时PCR检测了CfTNFR转录本在健康和受细菌攻击的扇贝中的空间表达情况。在所有测试组织中都能检测到CfTNFR转录本，包括血细胞、性腺、鳃、外套膜和肝胰腺，鳗弧菌（Listonella anguillarum）攻击后，性腺、鳃、外套膜和肝胰腺组织中的CfTNFR转录本显著上调，表明CfTNFR是参与免疫防御的组成型和诱导型急性期蛋白。目前的结果表明低等无脊椎动物中存在TNFR样分子和TNF - TNFR系统，并为CfTNFR在扇贝对入侵微生物的固有免疫反应中的作用提供了新的见解。
