Antiviral Activity of ISG15 Against Porcine Deltacoronavirus Replication in IPEC-J2 by ISGylating PCNA

In this study, we identified known and novel candidate ISG15-binding proteins and analyzed the protein complexes by affinity pulldown and quantitative LC-MS/MS analyses in an IPEC-J2 cell line stably expressing ISG15 during PDCoV infection. We demonstrated that ISG15 conjugated to PCNA via specific Lys residues, which is the critical host factor for PDCoV replication, thereby suppressing viral propagation. We obtained evidence suggesting that HERC5 E3 ligase specifically promotes PCNA ISGylation and USP18 served as an ISG15-de-conjugating enzyme. We believe that our study makes a significant contribution to the literature because it is in advances our understanding of the function of ISG15 with expanding the physical and functional candidate protein interaction networks of ISG15 and suggesting ISGylation of PCNA plays a crucial role in PDCoV replication.本研究在稳定表达干扰素刺激基因15（ISG15）的IPEC-J2细胞系中，于猪德尔塔冠状病毒（PDCoV）感染期间，通过亲和pull-down实验与定量液相色谱-串联质谱（LC-MS/MS）分析，鉴定了已知及新型候选ISG15结合蛋白，并对蛋白复合物进行了系统表征。本研究证实，ISG15可通过特定赖氨酸残基与增殖细胞核抗原（PCNA）结合——PCNA作为PDCoV复制的关键宿主因子，该ISG化修饰（ISGylation）可有效抑制病毒增殖。本研究获得的证据表明，E3泛素连接酶HERC5可特异性促进PCNA的ISG化修饰，而泛素特异性蛋白酶18（USP18）则作为ISG15去结合酶发挥功能。本研究拓展了ISG15的物理与功能候选蛋白互作网络，加深了学界对ISG15功能的认知，并证实PCNA的ISG化修饰在PDCoV复制过程中发挥关键作用，因此本研究为相关领域的研究提供了重要学术参考。