Shandong Provincial Key Laboratory of Animal Resistance Biology, College of Life Sciences, Shandong Normal University, No. 88 East Wenhua Road, Jinan, 250014, China.
College of Fisheries and Life Science, Hainan Tropical Ocean University, No. 1 Yucai Road, Sanya, 572022, China.
BMC Vet Res. 2022 Dec 12;18(1):433. doi: 10.1186/s12917-022-03538-4.
Interferon (IFN) regulatory factors (IRFs) is a kind of transcription factors, which play an important role in regulating the expression of type I IFN and related genes. In mammals, IRF6 is not relevant with IFN expression, while zebrafish IRF6 was reported to be a positive regulator of IFN expression and could be phosphorylated by both MyD88 and TBK1. However, the role of IRF6 in the immune response and IFN transcription of common carp is unknown.
In the present study, the cDNA of IRF6 gene (CcIRF6) was cloned from common carp using RACE technique, with a total length of 1905 bp, encoding 471 amino acid residues, which possesses two functional domains of DBD and IAD. Similarity analysis showed that CcIRF6 had more than 50% similarity with IRFs of other vertebrates, and had the highest similarity with grass carp and zebrafish, among which the DBD domain was much more conserved. The phylogenetic analysis showed that CcIRF6 is in the branch of Osteichthyes and has the closest relationship with grass carp. In healthy common carp, the CcIRF6 was expressed in all the examined tissues, with the highest level in the oral epithelium, and the lowest level in the head kidney. After intraperitoneal injection of poly(I:C) or Aeromonas hydrophila, the expression of CcIRF6 increased in spleen, head kidney, foregut and hindgut of common carp. Moreover, poly(I:C), LPS, PGN and flagellin induced the expression of CcIRF6 in peripheral leukocytes and head kidney leukocytes of common carp in vitro. In EPC cells, CcIRF6 inhibited the expression of some IFN-related genes and pro-inflammatory cytokines, and dual luciferase reporter assay showed that CcIRF6 reduced the activity of IFN and NF-κB reporter genes.
The present study suggests that CcIRF6 is involved in the antiviral and antibacterial immune response of common carp, and negatively regulate the expression of IFN and NF-κB signalling pathways, which provides a theoretical basis for the study and prevention of fish disease pathogenesis.
干扰素（IFN）调节因子（IRF）是一类转录因子，在调节 I 型 IFN 及相关基因的表达中发挥重要作用。在哺乳动物中，IRF6 与 IFN 表达无关，而斑马鱼 IRF6 被报道为 IFN 表达的正调控因子，可被 MyD88 和 TBK1 磷酸化。然而，IRF6 在鲤鱼免疫反应和 IFN 转录中的作用尚不清楚。
本研究采用 RACE 技术从鲤鱼中克隆出 IRF6 基因（CcIRF6）的 cDNA，全长 1905bp，编码 471 个氨基酸残基，具有 DBD 和 IAD 两个功能域。相似性分析表明，CcIRF6 与其他脊椎动物的 IRFs 具有 50%以上的相似性，与草鱼和斑马鱼的相似性最高，其中 DBD 结构域更为保守。系统进化分析表明，CcIRF6 位于硬骨鱼分支，与草鱼亲缘关系最近。在健康鲤鱼中，CcIRF6 在所有检测组织中均有表达，其中口腔上皮组织表达量最高，头肾组织表达量最低。腹腔注射 poly(I:C)或嗜水气单胞菌后，CcIRF6 在鲤鱼脾脏、头肾、前肠和后肠中的表达量增加。此外，poly(I:C)、LPS、PGN 和鞭毛蛋白在体外诱导鲤鱼外周血白细胞和头肾白细胞中 CcIRF6 的表达。在 EPC 细胞中，CcIRF6 抑制了一些 IFN 相关基因和促炎细胞因子的表达，双荧光素酶报告基因检测显示 CcIRF6 降低了 IFN 和 NF-κB 报告基因的活性。
本研究表明，CcIRF6 参与了鲤鱼的抗病毒和抗细菌免疫反应，并负调控 IFN 和 NF-κB 信号通路的表达，为鱼类疾病发病机制的研究和防治提供了理论依据。
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