State Key Laboratory of Developmental Biology of Freshwater Fish, College of Life Science, Hunan Normal University, Changsha, 410081, China.
State Key Laboratory of Developmental Biology of Freshwater Fish, College of Life Science, Hunan Normal University, Changsha, 410081, China.
Fish Shellfish Immunol. 2023 Nov;142:109095. doi: 10.1016/j.fsi.2023.109095. Epub 2023 Sep 18.
IKKε plays an important role in the activation of IRF3/IRF7 and the production of interferon (IFN), however, its regulation remains obscure in human. E3 ligase TRIM25 has been reported to manipulate the K63-linked ubiquitination of RIG-I, leading to the activation of RIG-I/IFN signaling. To elucidate the role of TRIM25 in teleost, a TRIM25 homolog (bcTRIM25) was cloned and characterized from black carp (Mylopharyngodon piceus). bcTRIM25 contains 653 amino acids, possessing conservative RING, B-box and SPRY domain, which is highly expressed in muscle, spleen and skin. bcTRIM25 knock-down enhanced the antiviral ability of host cells. bcTRIM25 over-expression alone in EPC cells attenuated bcIFNa promoter transcription in the reporter assays and impeded PKR and MX1 expression in qRT-PCR. Interestingly, co-IP assays indicated that bcTRIM25 interacted with bcIKKε and the induced bcIFNa promoter transcription by bcIKKε was notably hindered by bcTRIM25. Furthermore, bcIKKε-induced expression of interferon stimulated genes (ISGs) and antiviral activity were dampened by bcTRIM25. Further exploration showed that bcTRIM25 visibly enhanced the ubiquitination of bcIKKε but significantly attenuated the phosphorylation of bcIKKε. Thus, our data demonstrate for the first time in vertebrate that TRIM25 negatively regulates IKKε through enhancing its ubiquitination, which sheds a light on the regulation of IKKε/IFN signaling.
IKKε 在 IRF3/IRF7 的激活和干扰素（IFN）的产生中发挥重要作用，但其在人类中的调节仍不清楚。E3 连接酶 TRIM25 已被报道操纵 RIG-I 的 K63 连接泛素化，导致 RIG-I/IFN 信号的激活。为了阐明 TRIM25 在硬骨鱼中的作用，从草鱼（Mylopharyngodon piceus）中克隆并鉴定了一个 TRIM25 同源物（bcTRIM25）。bcTRIM25 包含 653 个氨基酸，具有保守的 RING、B-box 和 SPRY 结构域，在肌肉、脾脏和皮肤中高度表达。bcTRIM25 敲低增强了宿主细胞的抗病毒能力。单独在 EPC 细胞中转染 bcTRIM25 增强了 bcIFNa 启动子的转录活性，并通过 qRT-PCR 抑制了 PKR 和 MX1 的表达。有趣的是，co-IP 实验表明，bcTRIM25 与 bcIKKε 相互作用，bcIKKε 诱导的 bcIFNa 启动子转录明显受到 bcTRIM25 的阻碍。此外，bcTRIM25 显著降低了 bcIKKε 诱导的干扰素刺激基因（ISGs）的表达和抗病毒活性。进一步探索表明，bcTRIM25 显著增强了 bcIKKε 的泛素化，但显著降低了 bcIKKε 的磷酸化。因此，我们的数据首次在脊椎动物中表明，TRIM25 通过增强其泛素化来负调控 IKKε，这为 IKKε/IFN 信号的调节提供了新的视角。
