PCV2 Rep-衣壳共组装体偶联基因组衣壳化与其合成的分子机制

全球一体化及养殖集约化进程日趋加快，使传统疫苗愈发无法满足高突变率猪圆环病毒2型（PCV2）防控的迫切需求。而不受基因组变异影响的衣壳化过程蕴藏着靶向防控的巨大潜在价值。但PCV2复制过程仍诸多不清楚。前期研究打破人为划分复制步骤间的界限，初步发现衣壳通过结合复制酶蛋白（Rep）以特异性包裹基因组。本研究旨在深入探讨新合成的子代基因组单链DNA如何即时衣壳化的问题，拟首先从胞内分离Rep-衣壳免疫沉淀复合物，并纯化预装PCV2，明确Rep与衣壳的结合关系；进而解析其中Rep与衣壳、Rep与基因组的直接结合关系，Rep结合衣壳以特异性包裹基因组的特性，揭示Rep偶联基因组衣壳化与其合成的作用；最后，通过体外组装验证Rep与衣壳独立于核酸的共组装关系，阐明PCV2 Rep-衣壳共组装体偶联基因组衣壳化与其合成的分子机制。这将为其复制周期提供新理论，并为高效防控中保守性靶点的挖掘提供新依据。

Accelerating of globalization and herd breeding intensification have made traditional vaccines increasingly unable to meet the requirements for prevention and control of PCV2 with high mutation rate. The genome encapsidation process, which is not affected by genomic mutation, has huge potential value for targeted prevention and control. However, many steps in PCV2 replication cycle remain unclear. The boundaries between artificially divided steps were broken, and the capsid bound to the replicase protein (Rep) to specifically wrap the genome was initially found in the preliminary study. This study aims to explore how the newly synthesized single-stranded DNA of the progeny viral genome is immediately encapsidated. First, the Rep-capsid immunoprecipitation complex will be isolated, and the pre-assembled PCV2 will be purified from the infected cells, clarifying the binding relationship between Rep and capsid. Furthermore, the direct binding relationship between Rep and capsid, between Rep and genome, the genome encapsidation ability of Rep-capsid will be analyzed, revealing the role of Rep couples genome encapsidation with its synthesis. Finally, the nucleic acid-independent co-assembly relationship between Rep and capsid will be verified through in vitro assembly, demonstrating the molecular mechanism of PCV2 Rep-capsid co-assembly couples genome encapsidation with its synthesis. Taken together, this study will for the first time provide new insights into the replication cycle of PCV2, and provide new bases for the mining of conservative targets to effectively prevent and control PCV2.