Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, China.
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; College of Marine Sciences, South China Agricultural University, Guangzhou 510642, China; University of Chinese Academy of Sciences, Beijing, China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, China.
Fish Shellfish Immunol. 2017 Nov;70:351-360. doi: 10.1016/j.fsi.2017.09.030. Epub 2017 Sep 12.
Signal Transducer and Activator of Transcription 1 (STAT1) has been demonstrated to function as a critical mediator in multiple cell processes, such as cell proliferation, cell death, and innate immune response. Interestingly, two orthologues of human STAT1, including STAT1a and STAT1b genes have been identified in different fish. However, the detailed roles of fish STAT1a in virus replication still remained largely uncertain. Here, we cloned a STAT1a from orange-spotted grouper Epinephelus coioides (EcSTAT1a) and characterized its roles during fish virus infection. EcSTAT1a encoded a 751-aa peptide which shared 97% and 93% identity to STAT1 from mandarin fish (Siniperca chuatsi) and Malabar grouper (Epinephelus malabaricus), respectively. Amino acid alignment analysis showed that EcSTAT1a contained a STAT-int domain, a STAT-alpha domain, a STAT-bind domain (DNA binding domain), a SH2 domain and a STAT1-TAZ2 bind domain. In examined tissues from healthy grouper, the expression of EcSTAT1a was predominant in intestine, gill and liver. In grouper cells, the relative expression levels of EcSTAT1a was significantly increased during red-spotted grouper nervous necrosis virus (RGNNV) or Singapore grouper iridovirus (SGIV) infection. Under fluorescence microscopy, we found that EcSTAT1a mainly localized in the cytoplasm. The ectopic expression of EcSTAT1a in vitro significantly delayed the cytopathic effect (CPE) progression evoked by RGNNV and SGIV. Further studies showed that the expression levels of viral genes, including SGIV major capsid protein (MCP), VP19, ICP-18, LITAF and RGNNV coat protein (CP), RNA-dependent RNA polymerase (RdRp) were all significantly reduced in EcSTAT1a overexpressing cells compared to the control vector transfected cells, suggested that EcSTAT1a exerted antiviral activity against iridovirus and nodavirus. Furthermore, overexpression of EcSTAT1a significantly increased the expression of interferon related cytokines or effectors and pro-inflammatory factors. Together, our results elucidated that EcSTAT1a might function as a critical antiviral factor by regulating the host interferon immune and inflammation response.
信号转导子和转录激活子 1（STAT1）已被证明在多种细胞过程中发挥关键介导作用，如细胞增殖、细胞死亡和先天免疫反应。有趣的是，在不同鱼类中已经鉴定出人类 STAT1 的两个同源物，包括 STAT1a 和 STAT1b 基因。然而，鱼类 STAT1a 在病毒复制中的详细作用在很大程度上仍然不确定。在这里，我们从橙色斑点石斑鱼（Epinephelus coioides）克隆了 STAT1a（EcSTAT1a），并研究了其在鱼类病毒感染过程中的作用。EcSTAT1a 编码一个 751 个氨基酸的肽，与来自鳜鱼（Siniperca chuatsi）和马拉巴石斑鱼（Epinephelus malabaricus）的 STAT1 分别具有 97%和 93%的同一性。氨基酸比对分析表明，EcSTAT1a 包含一个 STAT-int 结构域、一个 STAT-alpha 结构域、一个 STAT 结合结构域（DNA 结合结构域）、一个 SH2 结构域和一个 STAT1-TAZ2 结合结构域。在健康石斑鱼的检查组织中，EcSTAT1a 的表达主要在肠、鳃和肝脏中。在石斑鱼细胞中，EcSTAT1a 的相对表达水平在红鳍东方鲀神经坏死病毒（RGNNV）或新加坡石斑鱼虹彩病毒（SGIV）感染期间显著增加。在荧光显微镜下，我们发现 EcSTAT1a 主要定位于细胞质中。体外过表达 EcSTAT1a 可显著延迟 RGNNV 和 SGIV 引起的细胞病变效应（CPE）进展。进一步的研究表明，与对照载体转染细胞相比，过表达 EcSTAT1a 的细胞中病毒基因的表达水平，包括 SGIV 主要衣壳蛋白（MCP）、VP19、ICP-18、LITAF 和 RGNNV 衣壳蛋白（CP）、RNA 依赖性 RNA 聚合酶（RdRp）均显著降低，表明 EcSTAT1a 对虹彩病毒和正粘病毒具有抗病毒活性。此外，过表达 EcSTAT1a 显著增加了干扰素相关细胞因子或效应物和促炎因子的表达。综上所述，我们的结果表明，EcSTAT1a 可能通过调节宿主干扰素免疫和炎症反应，发挥抗病毒的关键作用。
