Hu Guo-Bin, Zhang Shou-Feng, Yang Xi, Liu Da-Hai, Liu Qiu-Ming, Zhang Shi-Cui
College of Marine Life Sciences, Ocean University of China, Qingdao 266003, China; Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao 266003, China.
College of Marine Life Sciences, Ocean University of China, Qingdao 266003, China.
Fish Shellfish Immunol. 2015 Jun;44(2):399-409. doi: 10.1016/j.fsi.2015.03.001. Epub 2015 Mar 11.
Toll-like receptor 22 (TLR22) exists exclusively in aquatic animals and recognizes double stranded RNA (dsRNA). In the present study, a tlr22 gene and its 5'-flanking sequence were cloned from turbot, Scophthalmus maximus, its immune responsive expression was subsequently studied in vivo. The turbot (sm)tlr22 gene spans over 5.6 kb with a structure of 4 exon-3 intron and encodes 962 amino acids. The deduced protein shows the highest sequence identity (76.7%) to Japanese flounder Tlr22 and possesses a signal peptide sequence, a leucine-rich repeat (LRR) domain composed of 27 LRR motifs, a transmembrane region and a Toll/interleukin-1 receptor (TIR) domain. Phylogenetic analysis grouped it with other teleost Tlr22s. The interferon-stimulated response element (ISRE) and signal transducer and activator of transcription (STAT) binding site important for the basal transcriptional activity of TLR3 were predicted in the 5'-flanking sequence of smtlr22 gene. Quantitative real-time PCR (qPCR) analysis demonstrated the constitutive expression of smtlr22 mRNA in all examined tissues with higher levels in the head kidney, kidney and spleen. Further, smtlr22 expression was significantly up-regulated following challenge with polyinosinic: polycytidylic acid (poly I:C), lipopolysaccharide (LPS) or turbot reddish body iridovirus (TRBIV) in the gills, head kidney, spleen and muscle, with maximum increases ranging from 2.56 to 6.24 fold upon different immunostimulants and organs. These findings suggest a possible role of Smtlr22 in the immune responses to the infections of a broad range of pathogens that include DNA and RNA viruses and Gram-negative bacteria.
Toll样受体22（TLR22）仅存在于水生动物中，可识别双链RNA（dsRNA）。在本研究中，从大菱鲆（Scophthalmus maximus）中克隆了tlr22基因及其5'侧翼序列，随后在体内研究了其免疫反应性表达。大菱鲆（sm）tlr22基因全长超过5.6 kb，具有4个外显子-3个内含子的结构，编码962个氨基酸。推导的蛋白质与牙鲆Tlr22的序列同一性最高（76.7%），并具有信号肽序列、由27个富含亮氨酸重复（LRR）基序组成的LRR结构域、跨膜区和Toll/白细胞介素-1受体（TIR）结构域。系统发育分析将其与其他硬骨鱼Tlr22归为一类。在smtlr22基因的5'侧翼序列中预测到了对TLR3基础转录活性重要的干扰素刺激反应元件（ISRE）和信号转导及转录激活因子（STAT）结合位点。定量实时PCR（qPCR）分析表明，smtlr22 mRNA在所有检测组织中均有组成性表达，在头肾、肾脏和脾脏中表达水平较高。此外，在用聚肌苷酸：聚胞苷酸（poly I:C）、脂多糖（LPS）或大菱鲆红体虹彩病毒（TRBIV）攻击鳃、头肾、脾脏和肌肉后，smtlr22表达显著上调，不同免疫刺激剂和器官的最大增幅在2.56至6.24倍之间。这些发现表明Smtlr22在对包括DNA和RNA病毒以及革兰氏阴性菌在内的多种病原体感染的免疫反应中可能发挥作用。
