Suppression of cGAS-STING- and RIG-I-MAVS-mediated innate immune responses by Epstein-Barr virus-encoded tegument protein BPLF1 through de-ubiquitination

Epstein-Barr virus (“EBV”), a γ-herpesvirus, is the first identified human oncogenic virus. EBV infects more than 90% of the adult population around the world and it is etiologically linked with 2% of human malignancies such as Burkitt’s lymphoma, Hodgkin’s lymphoma, gastric carcinoma and nasopharyngeal carcinoma (“NPC”). The prevalence of EBV infection has suggested that the virus may have developed effective mechanism to evade host innate immunity. In this project, we have screened out a conserved herpesvirus de-ubiquitinase (“DUB”) in EBV, BPLF1, which can potently antagonize the type-I interferon production induced by DNA sensors, cGAS and STING, or RNA sensors, RIG-I and MAVS. Both of the two naturally occurring forms of the large tegument protein BPLF1, the small fragment BPLF1 325 and the full length BPLF1, have also shown potent suppression on cGAS-STING-, RIG-I- and TBK1- induced interferon production. The observed suppressions disappear when the cysteine residue of BPLF1 at position 61 is substituted with alanine, which changed its DUB domain into a catalytically inactive form. This indicates that BPLF1 suppresses innate immunity through its DUB activity. The DUB activity of BPLF1 also promoted EBV infection against cGAS-STING- and TBK1-mediated antiviral responses. The BPLF1 is an effective DUB for all lysine 63 (K63)-, lysine 48 (K48)-and lysine (K27)- linked ubiquitin moieties on adaptor molecule STING and K63- and K48-linked ubiquitin moieties on common adaptor protein TBK1. Physical interaction is found between BPLF1 and STING. The DUB activities of BPLF1 would disturb TBK1-induced IRF3 dimerization. These experiments indicate the DUB activity of the large tegument protein BPLF1 has antagonized innate immune responses through its DUB activities on STING and TBK1 in innate immune signaling. <br>

EB病毒（Epstein-Barr virus, EBV）属于γ疱疹病毒（γ-herpesvirus），是首个被鉴定出的人类致癌病毒。该病毒可感染全球超90%的成年人群，且与约2%的人类恶性肿瘤存在病因学关联，包括伯基特淋巴瘤、霍奇金淋巴瘤、胃癌以及鼻咽癌（nasopharyngeal carcinoma, NPC）。EBV感染的高流行率提示，该病毒已进化出有效逃逸宿主固有免疫的机制。本研究中，我们从EBV中筛选得到一种保守的疱疹病毒去泛素化酶（de-ubiquitinase, DUB）BPLF1，其可强效拮抗由DNA传感器cGAS、STING以及RNA传感器RIG-I、MAVS诱导的I型干扰素产生。两种天然存在的BPLF1大被膜蛋白形式——截短片段BPLF1 325与全长BPLF1——均能有效抑制cGAS-STING、RIG-I及TBK1介导的干扰素产生。当BPLF1第61位半胱氨酸残基突变为丙氨酸，使其DUB结构域变为催化失活形式时，上述抑制效应消失，这表明BPLF1通过其DUB活性抑制固有免疫。BPLF1的DUB活性还可促进EBV对抗cGAS-STING及TBK1介导的抗病毒应答。BPLF1可靶向作用于接头分子STING上的赖氨酸63（K63）、赖氨酸48（K48）及赖氨酸27（K27）连接的泛素链，以及衔接蛋白TBK1上的K63、K48连接的泛素链。研究发现BPLF1与STING存在物理相互作用，且BPLF1的DUB活性会干扰TBK1诱导的IRF3二聚化。上述实验结果表明，大被膜蛋白BPLF1的DUB活性可通过作用于固有免疫信号通路中的STING与TBK1，拮抗宿主固有免疫应答。