Key Laboratory of Animal Virology of Ministry of Agriculture, Zhejiang Universitygrid.13402.34 Center for Veterinary Sciences, Hangzhou, China.
J Virol. 2022 Dec 14;96(23):e0152222. doi: 10.1128/jvi.01522-22. Epub 2022 Nov 21.
Nuclear entrance and stability of porcine circovirus type 2 (PCV2), the smallest virus in mammals, are crucial for its infection and replication. However, the mechanisms are not fully understood. Here, we found that the PCV2 virion maintains self-stability via the host importin 5 (IPO5) during infection. Coimmunoprecipitation combined with mass spectrometry and glutathione -transferase pulldown assays showed that the capsid protein (Cap) of PCV2 binds directly to IPO5. Fine identification demonstrated that the N-terminal residue arginine of Cap is the most critical to efficient binding to the proline residue of IPO5. Detection of replication ability further showed that IPO5 supports PCV2 replication by promoting the nuclear import of incoming PCV2 virions. Knockdown of delayed the nuclear transport of incoming PCV2 virions and significantly decreased the intracellular levels of overexpressed PCV2 Cap, which was reversed by treatment with a proteasome inhibitor or by rescuing IPO5 expression. Cycloheximide treatment showed that IPO5 increases the stability of the PCV2 Cap protein. Taken together, our findings demonstrated that during infection, IPO5 facilitates PCV2 replication by directly binding to the nuclear localization signal of Cap to block proteasome degradation. Circovirus is the smallest virus to cause immune suppression in pigs. The capsid protein (Cap) is the only viral structural protein that is closely related to viral infection. The nuclear entry and stability of Cap are necessary for PCV2 replication. However, the molecular mechanism maintaining the stability of Cap during nuclear trafficking of PCV2 is unknown. Here, we report that IPO5 aggregates within the nuclear periphery and combines with incoming PCV2 capsids to promote their nuclear entry. Concurrently, IPO5 inhibits the degradation of newly synthesized Cap protein, which facilitates the synthesis of virus proteins and virus replication. These findings highlight a mechanism whereby IPO5 plays a dual role in PCV2 infection, which not only enriches our understanding of the virus replication cycle but also lays the foundation for the subsequent development of antiviral drugs.
猪圆环病毒 2 型（PCV2）是哺乳动物中最小的病毒，其核进入和稳定性对于其感染和复制至关重要。然而，其机制尚未完全阐明。在这里，我们发现 PCV2 病毒粒子在感染过程中通过宿主输入蛋白 5（IPO5）维持自身稳定性。免疫共沉淀结合质谱和谷胱甘肽转移酶 pulldown 实验表明，PCV2 的衣壳蛋白（Cap）直接与 IPO5 结合。精细鉴定表明，Cap 的 N 端残基精氨酸是与 IPO5 的脯氨酸残基有效结合的最关键残基。复制能力检测进一步表明，IPO5 通过促进进入的 PCV2 病毒粒子的核输入来支持 PCV2 的复制。敲低 IPO5 延迟了进入的 PCV2 病毒粒子的核转运，并显著降低了过表达 PCV2 Cap 的细胞内水平，用蛋白酶体抑制剂处理或挽救 IPO5 表达可逆转这一结果。环已酰亚胺处理表明，IPO5 增加了 PCV2 Cap 蛋白的稳定性。总之，我们的研究结果表明，在感染过程中，IPO5 通过直接与 Cap 的核定位信号结合来阻止蛋白酶体降解，从而促进 PCV2 的复制。圆环病毒是导致猪免疫抑制的最小病毒。衣壳蛋白（Cap）是唯一与病毒感染密切相关的病毒结构蛋白。Cap 的核进入和稳定性是 PCV2 复制所必需的。然而，PCV2 核运输过程中维持 Cap 稳定性的分子机制尚不清楚。在这里，我们报告 IPO5 聚集在核周缘并与进入的 PCV2 衣壳结合，促进其核进入。同时，IPO5 抑制新合成的 Cap 蛋白的降解，从而促进病毒蛋白的合成和病毒复制。这些发现强调了 IPO5 在 PCV2 感染中发挥双重作用的机制，不仅丰富了我们对病毒复制周期的理解，也为随后开发抗病毒药物奠定了基础。
