Wu Yang, Li Mingwei, Tian Jin, Yan Haoxin, Pan Yudi, Shi Hongyan, Shi Da, Chen Jianfei, Guo Longjun, Feng Li
State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, China.
PLoS Pathog. 2023 Oct 6;19(10):e1011702. doi: 10.1371/journal.ppat.1011702. eCollection 2023 Oct.
Coronaviruses (CoVs) are a family of the largest RNA viruses that typically cause respiratory, enteric, and hepatic diseases in animals and humans, imposing great threats to the public safety and animal health. Porcine deltacoronavirus (PDCoV), a newly emerging enteropathogenic coronavirus, causes severe diarrhea in suckling piglets all over the world and poses potential risks of cross-species transmission. Here, we use PDCoV as a model of CoVs to illustrate the reciprocal regulation between CoVs infection and host antiviral responses. In this study, downregulation of DNA polymerase delta interacting protein 3 (POLDIP3) was confirmed in PDCoV infected IPEC-J2 cells by isobaric tags for relative and absolute quantification (iTRAQ) and Western blotting analysis. Overexpression of POLDIP3 inhibits PDCoV infection, whereas POLDIP3 knockout (POLDIP3-/-) by CRISPR-Cas9 editing significantly promotes PDCoV infection, indicating POLDIP3 as a novel antiviral regulator against PDCoV infection. Surprisingly, an antagonistic strategy was revealed that PDCoV encoded nonstructural protein 5 (nsp5) was responsible for POLDIP3 reduction via its 3C-like protease cleavage of POLDIP3 at the glutamine acid 176 (Q176), facilitating PDCoV infection due to the loss of antiviral effects of the cleaved fragments. Consistent with the obtained data in IPEC-J2 cell model in vitro, POLDIP3 reduction by cleavage was also corroborated in PDCoV infected-SPF piglets in vivo. Collectively, we unveiled a new antagonistic strategy evolved by PDCoV to counteract antiviral innate immunity by nsp5-mediated POLDIP3 cleavage, eventually ensuring productive virus replication. Importantly, we further demonstrated that nsp5s from PEDV and TGEV harbor the conserved function to cleave porcine POLDIP3 at the Q176 to despair POLDIP3-mediated antiviral effects. In addition, nsp5 from SARS-CoV-2 also cleaves human POLDIP3. Therefore, we speculate that coronaviruses employ similar POLDIP3 cleavage mechanisms mediated by nsp5 to antagonize the host antiviral responses to sustain efficient virus infection.
冠状病毒（CoVs）是最大的RNA病毒家族之一，通常在动物和人类中引起呼吸道、肠道和肝脏疾病，对公共安全和动物健康构成巨大威胁。猪德尔塔冠状病毒（PDCoV）是一种新出现的肠道致病性冠状病毒，在世界各地的哺乳仔猪中引起严重腹泻，并带来跨物种传播的潜在风险。在此，我们以PDCoV作为冠状病毒模型，来说明冠状病毒感染与宿主抗病毒反应之间的相互调节。在本研究中，通过相对和绝对定量的等压标签（iTRAQ）和蛋白质免疫印迹分析，证实了在PDCoV感染的IPEC-J2细胞中DNA聚合酶δ相互作用蛋白3（POLDIP3）表达下调。过表达POLDIP3可抑制PDCoV感染，而通过CRISPR-Cas9编辑敲除POLDIP3（POLDIP3-/-）则显著促进PDCoV感染，表明POLDIP3是一种针对PDCoV感染的新型抗病毒调节因子。令人惊讶的是，研究揭示了一种拮抗策略，即PDCoV编码的非结构蛋白5（nsp5）通过其3C样蛋白酶在谷氨酰胺176（Q176）处切割POLDIP3，导致POLDIP3减少，由于切割片段抗病毒作用的丧失而促进PDCoV感染。与体外IPEC-J2细胞模型中获得的数据一致，在体内感染PDCoV的无特定病原体（SPF）仔猪中也证实了POLDIP3因切割而减少。总的来说，我们揭示了PDCoV进化出的一种新的拮抗策略，即通过nsp5介导的POLDIP3切割来对抗抗病毒先天免疫，最终确保病毒的有效复制。重要的是，我们进一步证明，来自猪流行性腹泻病毒（PEDV）和传染性胃肠炎病毒（TGEV）的nsp5具有在Q176处切割猪POLDIP3以消除POLDIP3介导的抗病毒作用的保守功能。此外，严重急性呼吸综合征冠状病毒2（SARS-CoV-2）的nsp5也能切割人POLDIP3。因此，我们推测冠状病毒采用由nsp5介导的类似POLDIP3切割机制来拮抗宿主抗病毒反应，以维持有效的病毒感染。
