Podok Patarida, Xu Lijuan, Xu Dan, Lu Liqun
Key Laboratory of Aquatic Genetic Resources of the Ministry of Agriculture, Shanghai Ocean University, 201306, PR China.
Key Laboratory of Aquatic Genetic Resources of the Ministry of Agriculture, Shanghai Ocean University, 201306, PR China.
Fish Shellfish Immunol. 2014 May;38(1):65-73. doi: 10.1016/j.fsi.2014.03.001. Epub 2014 Mar 14.
Interleukin 11 (IL-11), Intelectin (ITLN) and Purine nucleoside phosphorylase 5a (PNP5a) play important roles in innate immunity. In a previous study to identify differentially expressed immune-related genes, suppression subtractive hybridization (SSH) assay was used to characterize differentially expressed genes in crucian carp (Carassius auratus gibelio) infected with Cyprinid herpesvirus 2 (CyHV-2) in which IL-11, ITLN and PNP5a were identified to be the three most significantly up-regulated genes (Xu et al., Archives of Virology, 2014, http://dx.doi.org/10.1007/s00705-014-2011-9). In this study, the complete open reading frames (ORF) of IL-11, ITLN and PNP5a genes were cloned and sequenced. The full-length cDNAs of the three genes contained an ORF of 597, 945 and 882 bp, encoding a polypeptide of 198, 314 and 293 amino acids, respectively. Phylogenetic analysis indicated that the three genes shared high homology to other bony fish species including Zebrafish. Interestingly, the ITLN gene of crucian carp lacked a 10 aa peptide that was found in the C-terminal of other fish species. A real-time RT-PCR assay was developed to quantitatively examine their tissue distribution. We found that IL-11, ITLN and PNP5a were expressed at low levels in all of the tissues examined. To monitor the response of these genes to CyHV- 2 or Aeromonas hydrophila (A. hydrophila) infection, we determined the expression level of IL-11, ITLN and PNP5a at different time points after infection in kidney. Significant up-regulation of IL-11, ITLN and PNP5a was only observed 72 h post-CyHV-2 injection (P < 0.01), whereas significant up-regulation was observed as early as 6 h after infection with A. hydrophila (P < 0.01). Our results demonstrated that host innate immune response to CyHV-2, at least in which IL-11, ITLN and PNP5a were involved, was slow in comparison to that induced by A. hydrophila. It suggested that CyHV-2 might suppress host innate response during early infection. The lack of a C-terminal peptide of crucian carp ITLN gene implied a possible functional difference of this gene during evolution, which merit further investigation.
白细胞介素11（IL-11）、肝凝集素（ITLN）和嘌呤核苷磷酸化酶5a（PNP5a）在固有免疫中发挥重要作用。在之前一项旨在鉴定差异表达的免疫相关基因的研究中，抑制性消减杂交（SSH）分析被用于表征感染鲤疱疹病毒2（CyHV-2）的鲫鱼（Carassius auratus gibelio）中差异表达的基因，其中IL-11、ITLN和PNP5a被鉴定为上调最为显著的三个基因（Xu等人，《病毒学档案》，2014年，http://dx.doi.org/10.1007/s00705-014-2011-9）。在本研究中，克隆并测序了IL-11、ITLN和PNP5a基因的完整开放阅读框（ORF）。这三个基因的全长cDNA分别包含597、945和882 bp的ORF，分别编码198、314和293个氨基酸的多肽。系统发育分析表明，这三个基因与包括斑马鱼在内的其他硬骨鱼物种具有高度同源性。有趣的是，鲫鱼的ITLN基因在C端缺少一段在其他鱼类物种中发现的10个氨基酸的肽段。开发了一种实时RT-PCR分析方法来定量检测它们的组织分布。我们发现IL-11、ITLN和PNP5a在所有检测的组织中表达水平较低。为了监测这些基因对CyHV-2或嗜水气单胞菌（A. hydrophila）感染的反应，我们测定了感染后不同时间点肾脏中IL-11、ITLN和PNP5a的表达水平。仅在注射CyHV-2后72小时观察到IL-11、ITLN和PNP5a显著上调（P < 0.01），而在用嗜水气单胞菌感染后6小时就观察到显著上调（P < 0.01）。我们的结果表明，宿主对CyHV-2的固有免疫反应，至少在涉及IL-11、ITLN和PNP5a的情况下，与嗜水气单胞菌诱导的反应相比是缓慢的。这表明CyHV-2可能在早期感染期间抑制宿主的固有反应。鲫鱼ITLN基因缺少C端肽段暗示了该基因在进化过程中可能存在功能差异，这值得进一步研究。
