Guangdong Laboratory for Lingnan Modern Agriculture, College of Marine Sciences, South China Agricultural University, Guangzhou, China.
Department of Biological Sciences, National University of Singapore, Singapore, Singapore.
Front Immunol. 2023 May 8;14:1185907. doi: 10.3389/fimmu.2023.1185907. eCollection 2023.
Receptor interacting protein 2 (RIP2), serves as a vital sensor of cell stress, is able to respond to cell survival or inflammation, and is involved in antiviral pathways. However, studies on the property of RIP2 in viral infections in fish have not been reported.
In this paper, we cloned and characterized RIP2 homolog from orange-spotted grouper (Epinephelus coioides) (EcRIP2) and further discussed the relevance of EcRIP2 to EcASC, comparing the influences of EcRIP2 and EcASC on the modulation of inflammatory factors and the NF-κB activation to reveal the mechanism of EcRIP2 in fish DNA virus infection.
Encoded a 602 amino acid protein, EcRIP2 contained two structural domains: S-TKc and CARD. Subcellular localization signified that EcRIP2 existed in cytoplasmic filaments and dot aggregation patterns. After SGIV infection, the EcRIP2 filaments aggregated into larger clusters near the nucleus. The infection of SGIV could notably up-regulate the transcription level of the EcRIP2 gene compared with lipopolysaccharide (LPS) and red grouper nerve necrosis virus (RGNNV). Overexpression of EcRIP2 impeded SGIV replication. The elevated expression levels of inflammatory cytokines induced by SGIV were remarkably hindered by EcRIP2 treatment in a concentration-dependent manner. In contrast, EcASC treatment could up-regulate SGIV-induced cytokine expression in the presence of EcCaspase-1. Enhancing amounts of EcRIP2 could overcome the down regulatory effect of EcASC on NF-κB. Nevertheless, increasing doses of EcASC failed to restrain the NF-κB activation in the existence of EcRIP2. Subsequently, it was validated by a co-immunoprecipitation assay that EcRIP2 dose-dependently competed with EcASC binding to EcCaspase-1. With increasing time to SGIV infection, EcCaspase-1 gradually combined with more EcRIP2 than EcASC.
Collectively, this paper highlighted that EcRIP2 may impede SGIV-induced hyperinflammation by competing with EcASC for binding EcCaspase-1, thereby suppressing viral replication of SGIV. Our work supplies novel viewpoints into the modulatory mechanism of RIP2-associated pathway and offers a novel view of RIP2-mediated fish diseases.
受体相互作用蛋白 2（RIP2）作为细胞应激的重要传感器，能够响应细胞存活或炎症，并参与抗病毒途径。然而，关于 RIP2 在鱼类病毒感染中的特性的研究尚未报道。
在本文中，我们从橙斑石斑鱼（Epinephelus coioides）（EcRIP2）中克隆和表征了 RIP2 同源物，并进一步讨论了 EcRIP2 与 EcASC 的相关性，比较了 EcRIP2 和 EcASC 对炎症因子调节和 NF-κB 激活的影响，以揭示 EcRIP2 在鱼类 DNA 病毒感染中的机制。
编码 602 个氨基酸的蛋白，EcRIP2 包含两个结构域：S-TKc 和 CARD。亚细胞定位表明 EcRIP2 存在于细胞质纤维和点状聚集模式中。在 SGIV 感染后，EcRIP2 纤维聚集在靠近细胞核的更大簇中。与脂多糖（LPS）和红鳍东方鲀神经坏死病毒（RGNNV）相比，SGIV 的感染可显著上调 EcRIP2 基因的转录水平。EcRIP2 的过表达可抑制 SGIV 的复制。SGIV 诱导的炎症细胞因子的表达水平升高，EcRIP2 处理呈浓度依赖性显著受阻。相反，在存在 EcCaspase-1 的情况下，EcASC 处理可上调 SGIV 诱导的细胞因子表达。增加 EcRIP2 的量可以克服 EcASC 对 NF-κB 的下调作用。然而，增加剂量的 EcASC 未能抑制存在 EcRIP2 时 NF-κB 的激活。随后，通过共免疫沉淀实验验证，EcRIP2 剂量依赖性地与 EcASC 竞争结合 EcCaspase-1。随着 SGIV 感染时间的延长，EcCaspase-1 逐渐与更多的 EcRIP2 结合，而不是 EcASC。
总之，本文强调 EcRIP2 可能通过与 EcASC 竞争结合 EcCaspase-1 来抑制 SGIV 诱导的过度炎症，从而抑制 SGIV 的病毒复制。我们的工作为 RIP2 相关途径的调节机制提供了新的观点，并为 RIP2 介导的鱼类疾病提供了新的视角。
