Institute of Animal Husbandry and Veterinary Medicine, Zhejiang Academy of Agricultural Sciences, Hangzhou, 310021, China.
Institute of Animal Husbandry and Veterinary Medicine, Zhejiang Academy of Agricultural Sciences, Hangzhou, 310021, China.
J Proteomics. 2021 Aug 15;245:104281. doi: 10.1016/j.jprot.2021.104281. Epub 2021 Jun 5.
Duck enteritis virus (DEV), the causative agent of duck viral enteritis, causes a contagious, lethal viral disease in Anseriformes (waterfowls). In virus infection, host-virus interaction plays a crucial role in virus replication and pathogenesis. In our previous study, mRFP was fused with the C-terminus of DEV glycoprotein C (gC) to construct a fluorescent-tag DEV virus rgCRFP. In the current study, fluorescent fusion protein (gC-mRFP) was used as the proteomic probe. Co-immunoprecipitation and mass spectrometric analysis of proteins from rgCRFP-infected chicken embryo fibroblasts using commercial anti-RFP antibody led to the identification of a total of 21 gC interacting host proteins. Out of these 21 proteins, the interaction of seven host proteins (GNG2, AR1H1, PPP2CA, UBE2I, MCM5, NUBP1, HN1) with DEV gC protein was validated using membrane-bound split-ubiquitin yeast two-hybrid system (MbYTH) and bimolecular fluorescence complementation (BiFC) analyses. It indicated direct interaction between these proteins with DEV gC protein. This study has furthered the current understanding of DEV virus infection and pathogenesis. SIGNIFICANCE: gC is an crucial glycoprotein of duck enteritis virus that plays an important role in the viral life cycle. Uncovering the interaction between virus-host is very important to elucidate the pathogenic mechanism of the virus. In this study, host factors interacting with DEV gC have been discerned. And seven host proteins (GNG2, AR1H1, PPP2CA, UBE2I, MCM5, NUBP1, HN1) have been further validated to interact with DEV gC using MbYTH and BiFC analyses. These outcomes could shed light on how DEV manipulates the cellular machinery, which could further our understanding of DEV pathogenesis.
鸭肠炎病毒（DEV）是鸭病毒性肠炎的病原体，可引起雁形目（水禽）的传染性、致死性病毒性疾病。在病毒感染中，宿主-病毒相互作用在病毒复制和发病机制中起着至关重要的作用。在我们之前的研究中，mRFP 与 DEV 糖蛋白 C（gC）的 C 末端融合，构建了荧光标记的 DEV 病毒 rgCRFP。在本研究中，荧光融合蛋白（gC-mRFP）被用作蛋白质组学探针。使用商业抗 RFP 抗体对 rgCRFP 感染的鸡胚成纤维细胞中的蛋白质进行共免疫沉淀和质谱分析，共鉴定出 21 种与 DEV gC 相互作用的宿主蛋白。在这 21 种蛋白质中，使用膜结合的分裂泛素酵母双杂交系统（MbYTH）和双分子荧光互补（BiFC）分析验证了 7 种宿主蛋白（GNG2、AR1H1、PPP2CA、UBE2I、MCM5、NUBP1、HN1）与 DEV gC 蛋白的相互作用。结果表明这些蛋白与 DEV gC 蛋白直接相互作用。本研究进一步加深了对 DEV 病毒感染和发病机制的认识。意义：gC 是鸭肠炎病毒的一种重要糖蛋白，在病毒生命周期中起着重要作用。揭示病毒-宿主之间的相互作用对于阐明病毒的致病机制非常重要。在本研究中，已经发现与 DEV gC 相互作用的宿主因子。并且使用 MbYTH 和 BiFC 分析进一步验证了七种宿主蛋白（GNG2、AR1H1、PPP2CA、UBE2I、MCM5、NUBP1、HN1）与 DEV gC 相互作用。这些结果可以揭示 DEV 如何操纵细胞机制，这可以进一步加深我们对 DEV 发病机制的理解。
