Xin Lusheng, Zhang Huan, Zhang Ran, Li Hui, Wang Weilin, Wang Lingling, Wang Hao, Qiu Limei, Song Linsheng
Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; University of Chinese Academy of Sciences, Beijing 100049, China.
Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China.
Dev Comp Immunol. 2015 Dec;53(2):339-48. doi: 10.1016/j.dci.2015.08.002. Epub 2015 Aug 6.
Interleukin 17 (IL17) is a proinflammatory cytokine that plays an important role in immune response. Recently, five novel IL17 homologs have been identified by screening and analyzing the genome of pacific oyster Crassostrea gigas. In the present study, the functions of CgIL17-5 were investigated by examining the distribution of its mRNA and protein, ligands binding and modulation in immune response. The mRNA expression levels of CgIL17-5 in hemocytes of oysters post twice challenges of Vibrio splendidus were all significantly up-regulated (P < 0.01), while the secondary pathogen infection attenuated the expression level of CgIL17-5 mRNA compared with the primary challenge. CgIL17-5 was found to be located on oyster hemocyte membranes through fluorescence confocal assay. The luciferase reporter assays showed that CgIL17-5 could activate the transfactors NF-κB, CREB and ATF-1, and involve in their signal pathways in HEK293T cells. Meanwhile, CgIL17-5 could augment the IL6 synthesis in HuVEC cells, playing the similar roles as human IL17 in inflammatory response. Additionally, the recombinant CgIL17-5 (rCgIL17-5) could directly bind peptidoglycan (PGN), lipopolysaccharide (LPS), poly (I:C) and β-1,3-glucan, with the highest affinity to PGN, and significantly inhibit the growth of Micrococcus luteus and Escherichia coli. All the results collectively suggested that CgIL17-5, as an ancient inflammatory cytokine, could not only activate signal transduction for the release of other cytokines, but also mediate the clearance of extracellular bacteria in oysters.
白细胞介素17(IL17)是一种促炎细胞因子,在免疫反应中起重要作用。最近,通过筛选和分析太平洋牡蛎(Crassostrea gigas)的基因组,鉴定出了五种新的IL17同源物。在本研究中,通过检测其mRNA和蛋白质的分布、配体结合以及在免疫反应中的调节作用,对CgIL17-5的功能进行了研究。在受到两次灿烂弧菌攻击后,牡蛎血细胞中CgIL17-5的mRNA表达水平均显著上调(P < 0.01),而与初次攻击相比,继发病原体感染使CgIL17-5 mRNA的表达水平有所下降。通过荧光共聚焦分析发现CgIL17-5位于牡蛎血细胞的细胞膜上。荧光素酶报告基因检测表明,CgIL17-5可以激活转录因子NF-κB、CREB和ATF-1,并在HEK293T细胞中参与它们的信号通路。同时,CgIL17-5可以增加人脐静脉内皮细胞(HuVEC)中IL6的合成,在炎症反应中发挥与人IL17类似的作用。此外,重组CgIL17-5(rCgIL17-5)可以直接结合肽聚糖(PGN)、脂多糖(LPS)、聚肌苷酸胞苷酸(poly (I:C))和β-1,3-葡聚糖,对PGN的亲和力最高,并能显著抑制藤黄微球菌和大肠杆菌的生长。所有结果共同表明,CgIL17-5作为一种古老的炎症细胞因子,不仅可以激活其他细胞因子释放的信号转导,还可以介导牡蛎中细胞外细菌的清除。