Harmening Sarah, Bogdanow Boris, Wagner Karen, Liu Fan, Messerle Martin, Borst Eva Maria
Institute of Virology, Hannover Medical School, Hannover, Germany.
Research group "Structural Interactomics", Leibniz Forschungsinstitut für Molekulare Pharmakologie (FMP), Berlin, Germany.
J Virol. 2025 Apr 15;99(4):e0220124. doi: 10.1128/jvi.02201-24. Epub 2025 Mar 10.
Cleavage of human cytomegalovirus (HCMV) genomes and their packaging into capsids requires at least seven essential viral proteins, yet it is not completely understood how these proteins cooperate to accomplish this task. Besides the portal protein pUL104 and the terminase subunits pUL51, pUL56, and pUL89, the UL52 protein is also necessary for HCMV genome encapsidation; however, knowledge about pUL52 is scant. In the absence of pUL52, viral concatemers are not cleaved into unit-length genomes and no DNA-filled capsids are observed, yet no viral or cellular proteins interacting with pUL52 have been identified that would explain how pUL52 exerts its essential role in the HCMV infection cycle. In this study, we aimed at a comprehensive definition of pUL52-interacting proteins in infected cells. Using suitable HCMV mutants, we employed three complementary state-of-the-art proteomic approaches, namely biotin ligase-dependent proximity labeling, affinity purification, and cross-linking mass spectrometry. These experiments, combined with thorough validation by immunoblotting, pointed to several viral DNA-associated proteins and key players pivotal for genome encapsidation as interactors of pUL52. The most noticeable direct pUL52 interaction partners were the terminase subunits pUL56 and pUL89 as well as the portal protein pUL104. Hence, we suggest a model of pUL52 function in which pUL52 mediates association of HCMV genomes with the terminase subunits and the capsid portal. Taken together, our data contribute to the understanding of an essential viral process previously recognized as a prominent antiviral target. Disturbing the identified pUL52 interactions may provide a starting point to develop novel antiviral medication.
Human cytomegalovirus (HCMV) can evoke severe disease in immunocompromised patients and, moreover, is the most frequent viral cause of malformations in newborns. The virus-specific process of genome cleavage and packaging into capsids has emerged as an Achilles heel in the HCMV life cycle, which can be targeted by novel antiviral drugs, yet the mechanism of viral DNA encapsidation is only partially understood. Here, we report that the essential viral cleavage-packaging protein pUL52 interacts with several HCMV proteins known to be crucial for genome packaging, with the most prominent ones being the terminase complex and the portal protein. These data provide insight into the role of pUL52 during HCMV infection and may lay the basis for the development of additional antiviral substances tackling viral DNA packaging.
人巨细胞病毒（HCMV）基因组的切割及其包装到衣壳中至少需要七种必需的病毒蛋白，但这些蛋白如何协同完成这项任务尚不完全清楚。除了门户蛋白pUL104和末端酶亚基pUL51、pUL56和pUL89外，UL52蛋白对于HCMV基因组包装也是必需的；然而，关于pUL52的知识却很少。在缺乏pUL52的情况下，病毒串联体不会被切割成单位长度的基因组，也未观察到充满DNA的衣壳，但尚未鉴定出与pUL52相互作用的病毒或细胞蛋白，这无法解释pUL52如何在HCMV感染周期中发挥其重要作用。在本研究中，我们旨在全面定义感染细胞中与pUL52相互作用的蛋白。我们使用合适的HCMV突变体，采用了三种互补的前沿蛋白质组学方法，即生物素连接酶依赖性邻近标记、亲和纯化和交联质谱分析。这些实验，结合免疫印迹的全面验证，指出了几种病毒DNA相关蛋白以及对基因组包装至关重要的关键蛋白作为pUL52的相互作用蛋白。最显著的直接与pUL52相互作用的伙伴是末端酶亚基pUL56和pUL89以及门户蛋白pUL104。因此，我们提出了一个pUL52功能模型，其中pUL52介导HCMV基因组与末端酶亚基和衣壳门户的结合。综上所述，我们的数据有助于理解一个先前被认为是突出抗病毒靶点的重要病毒过程。干扰已鉴定的pUL52相互作用可能为开发新型抗病毒药物提供一个起点。
人巨细胞病毒（HCMV）可在免疫功能低下的患者中引发严重疾病，此外，它还是新生儿畸形最常见的病毒原因。病毒特异性的基因组切割和包装到衣壳中的过程已成为HCMV生命周期中的一个致命弱点，这可以被新型抗病毒药物靶向，但病毒DNA包装的机制仅被部分理解。在这里，我们报告必需的病毒切割包装蛋白pUL52与几种已知对基因组包装至关重要的HCMV蛋白相互作用，其中最突出的是末端酶复合物和门户蛋白。这些数据提供了对pUL52在HCMV感染期间作用的见解，并可能为开发针对病毒DNA包装的其他抗病毒物质奠定基础。
