In response to viral infections, various pattern recognition receptors (PRRs) are activated for the production of type I interferon (IFN I). As a center of these receptor responses, TANK binding kinase-1 (TBK1) activates interferon regulatory factor 3 (IRF3). SRC is a member of Src family kinases (SFK) which participates in TBK1-mediated IFN I signaling pathway. In mammals, the immunological function of SRC is depended on its interaction with TBK1. To date, SRC has not been studied in fish. In this paper, we cloned the ORF of grass carp (Ctenopharyngodon idellus) SRC (CiSRC). CiSRC has a closer relationship with Sinocyclocheilus rhinocerous SRC (SrSRC). The expression level of CiSRC was significantly up-regulated following poly (I:C) stimulation in grass carp tissues and cells. Subcellular localization results showed that CiSRC is located both in the cytoplasm and nucleus, while CiTBK1 is only located in the cytoplasm of CIK cells. When GFP-CiSRC and FLAG-CiTBK1 were co-transfected into CIK cells, we found that they were co-localized in the cytoplasm. GST-pulldown and Co-immunoprecipitation analysis revealed that CiSRC and CiSRC tyrosine kinase domain deletion mutant (SRC-ΔTyrkc) can interact with CiTBK1, respectively. CiSRC promotes the phosphorylation of CiTBK1. Furthermore, the phosphorylation of TBK1 is more strongly under poly (I:C) stimulation. We also demonstrated that SRC can up-regulate IFN I expression. These results above unraveled that CiSRC initiates innate immune response by binding to and then up-regulating the phosphorylation of TBK1.

为应对病毒感染，各种模式识别受体 (PRR) 被激活以产生 I 型干扰素 (IFN I)。作为这些受体反应的中心，TANK 结合激酶-1 (TBK1) 激活干扰素调节因子 3 (IRF3)。SRC 是参与 TBK1 介导的 IFN I 信号通路的 Src 家族激酶 (SFK) 的成员。在哺乳动物中，SRC 的免疫功能取决于其与 TBK1 的相互作用。迄今为止，尚未在鱼类中研究 SRC。在本文中，我们克隆了草鱼（Ctenopharyngodon idellus）SRC（CiSRC）的ORF。CiSRC与中华环螯虾SRC（SrSRC）的关系更为密切。CiSRC的表达水平在草鱼组织和细胞中的聚（I：C）刺激后显着上调。亚细胞定位结果显示，CiSRC同时位于细胞质和细胞核中，而CiTBK1仅位于CIK​​细胞的细胞质中。当 GFP-CiSRC 和 FLAG-CiTBK1 共转染到 CIK 细胞中时，我们发现它们共定位于细胞质中。GST-pulldown 和共免疫沉淀分析显示 CiSRC 和 CiSRC 酪氨酸激酶结构域缺失突变体 (SRC-ΔTyrkc) 可以分别与 CiTBK1 相互作用。CiSRC 促进 CiTBK1 的磷酸化。此外，TBK1 的磷酸化在 poly (I:C) 刺激下更加强烈。我们还证明了 SRC 可以上调 IFN I 的表达。