Porcine TRIM35 positively regulate TRAF3-mediated IFN-β production and inhibit Japanese encephalitis virus replication
Porcine TRIM35 positively regulate TRAF3-mediated IFN-β production and inhibit Japanese encephalitis virus replication
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DOI：
10.1016/j.dci.2021.104290
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发表时间：
2021-10-07
影响因子：
2.9
通讯作者：
Li, Yanhua
中科院分区：
文献类型：
--
作者：
Li, Chenxi;Zhou, Yanyang;Li, Yanhua
TRIM 35蛋白是一种泛素E3连接酶，通过调节先天免疫信号通路中多种衔接蛋白的泛素化来介导干扰素-β（IFN-β）的产生。在这里，我们克隆了猪TRIM 35（porTRIM 35）基因，并分析了其参与IFN-β的表达以及抗日本脑炎病毒（JEV）的抗病毒反应。porTRIM 35基因全长编码493个氨基酸的蛋白质，与人、小鼠、猴和兔的同源基因具有79.6%-89.5%的序列相似性。porTRIM 35具有TRIMs的典型结构特征，包括RING结构域、B-box结构域、卷曲螺旋结构域和PRY/SPYR结构域。外源性过表达porTRIM 35显著上调猪睾丸（ST）细胞中IFN-β的mRNA表达水平，以响应poly（I：C）刺激，而敲低内源性表达porTRIM 35导致IFN-β的表达水平降低。在机械上，porTRIM 35直接与猪TNF受体相关因子3（TRAF 3）相互作用并催化其Lys 63连接的多聚泛素化，从而导致IFN-β产生的上调。同时，我们证明了RING和PRY/SPRY结构域对于porTRIM 35的E3连接酶活性是必需的。在对JEV感染的应答中，porTRIM 35的内源性表达在mRNA水平上被显著抑制，而porTRIM 35的外源性表达显著提高了由JEV感染诱导的IFN-β的表达，并降低了ST细胞中的病毒滴度，表明porTRIM 35是JEV复制的负调节剂。这些数据证明了porTRIM 35在IFN-β表达以及针对JEV复制的抗病毒应答中的重要性。
Tripartite motif 35 (TRIM35) protein is a ubiquitin E3 ligase that mediates interferon-beta (IFN-beta) production via regulating ubiquitination of multiple adaptor proteins in innate immune signaling pathways. Here, we cloned the porcine TRIM35 (porTRIM35) gene and analyzed its involvement in IFN-beta expression as well as the antiviral response against Japanese encephalitis virus (JEV). The full-length porTRIM35 gene encoded a 493-amino acid protein and exhibited 79.6%-89.5% sequence similarity with its orthologues in humans, mice, monkeys and rabbits. porTRIM35 possessed typical structural features of TRIMs, including a RING domain, a B-box domain, a coiled-coil domain and a PRY/SPYR domain. Exogenous overexpression of porTRIM35 significantly up-regulated the mRNA expression level of IFN-beta in swine testicular (ST) cell in response to poly(I:C) stimulation, whereas knockdown endogenous expression of porTRIM35 lead to a decrease in the expression level of IFN-beta. Mechanically, porTRIM35 directly interacted with porcine TNF-receptor associated factor 3 (TRAF3) and catalyzed its Lys63-linked polyubiquitination, thereby leading to the up-regulation of IFN-beta production. Meanwhile, we demonstrated that the RING and PRY/SPRY domains were essential for the E3 ligase activity of porTRIM35. In response to JEV infection, the endogenous expression of porTRIM35 was markedly inhibited at the mRNA level, while exogenous expression of porTRIM35 significantly elevated the expression of IFN-beta induced by JEV infection and reduced viral titers in ST cells, suggesting that porTRIM35 is a negative regulator for JEV replication. These data demonstrate the importance of porTRIM35 in IFN-beta expression as well as the antiviral response against JEV replication.