Zhang Xue-Heng, Shi Yu-Hong, Chen Jiong
Laboratory of Biochemistry and Molecular Biology, School of Marine Sciences, Ningbo University, Ningbo 315211, China.
Laboratory of Biochemistry and Molecular Biology, School of Marine Sciences, Ningbo University, Ningbo 315211, China; Collaborative Innovation Center for Zhejiang Marine High-efficiency and Healthy Aquaculture, Ningbo University, Ningbo 315211, China.
Mol Immunol. 2015 Aug;66(2):439-50. doi: 10.1016/j.molimm.2015.05.009. Epub 2015 May 25.
C-type lectin receptors (CTLRs) play vital roles in immune responses as pattern-recognition receptors (PRRs). In this study, we identified a novel C-type lectin receptor (PaCTLRC) gene from ayu, Plecoglossus altivelis. Predicted PaCTLRC is a single transmembrane receptor with a typical carbohydrate recognition domain (CRD) at its C-terminus. Sequence comparison and phylogenetic tree analysis showed that PaCTLRC was most closely related to Atlantic salmon (Salmo salar) CLRC, but was significantly different from two other ayu CTLRs, aCLR and PaCD209L. PaCTLRC transcript was detected in all tested tissues and cells, with high levels in the liver; and its expression was significantly altered upon Vibrio anguillarum infection. Refolded recombinant PaCTLRC (rPaCTLRC) agglutinated three types of Gram-positive bacteria (Listeria monocytogenes, Staphylococcus aureus and Streptococcus iniae) and four types of Gram-negative bacteria (Aeromonas hydrophila, Escherichia coli, V. anguillarum and Vibrio parahaemolyticus) in a Ca(2+)-dependent manner in vitro, and Gram-positive bacteria were shown to be biologically relevant ligands for PaCTLRC. rPaCTLRC bound to d-mannose, d-galactose, l-fucose, N-acetyl-d-glucosamine (GlcNAc), lipopolysaccharide (LPS) and peptidoglycan (PGN), exhibiting a relative binding strength to d-mannose and PGN. d-Mannose, l-fucose, GlcNAc, LPS and PGN could inhibit the agglutinating activity of rPaCTLRC, while d-galactose did not functioned. PaCTLRC neutralization using anti-PaCTLRC IgG resulted in the inhibition of phagocytosis by ayu monocytes/macrophages (MO/MΦ) of S. aureus but not of E. coli, and produced a consistently higher survival rate of S. aureus than that of E. coli. d-Mannose, LPS and PGN treatment had no significant influence on the phagocytosis of ayu MO/MΦ. These results suggest that PaCTLRC may serve as a Gram-positive bacteria-preferred PRR which is involved in pathogen recognition and signal transduction in ayu MO/MΦ.
C型凝集素受体（CTLRs）作为模式识别受体（PRRs）在免疫反应中发挥着至关重要的作用。在本研究中，我们从香鱼（Plecoglossus altivelis）中鉴定出一个新的C型凝集素受体（PaCTLRC）基因。预测的PaCTLRC是一种单跨膜受体，在其C末端具有典型的碳水化合物识别结构域（CRD）。序列比较和系统发育树分析表明，PaCTLRC与大西洋鲑（Salmo salar）的CLRC关系最为密切，但与香鱼的另外两种CTLRs，即aCLR和PaCD209L有显著差异。在所有测试的组织和细胞中均检测到PaCTLRC转录本，在肝脏中水平较高；鳗弧菌感染后其表达发生显著变化。体外复性的重组PaCTLRC（rPaCTLRC）以Ca(2+)依赖的方式凝集三种革兰氏阳性菌（单核细胞增生李斯特菌、金黄色葡萄球菌和海豚链球菌）和四种革兰氏阴性菌（嗜水气单胞菌、大肠杆菌、鳗弧菌和副溶血性弧菌），并且革兰氏阳性菌被证明是PaCTLRC的生物学相关配体。rPaCTLRC与d-甘露糖、d-半乳糖、l-岩藻糖、N-乙酰-d-葡萄糖胺（GlcNAc）、脂多糖（LPS）和肽聚糖（PGN）结合，对d-甘露糖和PGN表现出相对结合强度。d-甘露糖、l-岩藻糖、GlcNAc、LPS和PGN可抑制rPaCTLRC的凝集活性，而d-半乳糖则无此作用。使用抗PaCTLRC IgG对PaCTLRC进行中和导致香鱼单核细胞/巨噬细胞（MO/MΦ）对金黄色葡萄球菌的吞噬作用受到抑制，但对大肠杆菌则无影响，并且金黄色葡萄球菌的存活率始终高于大肠杆菌。d-甘露糖、LPS和PGN处理对香鱼MO/MΦ的吞噬作用没有显著影响。这些结果表明，PaCTLRC可能作为一种革兰氏阳性菌偏好的PRR，参与香鱼MO/MΦ中的病原体识别和信号转导。
