Analysis of the protein interaction networks that orchestrate tegument assembly and envelopment of human cytomegalovirus virions and dense bodies

人巨细胞病毒（HCMV）颗粒的形态发生是一个复杂的过程，需要病毒蛋白的多重相互作用来组装衣壳，附着在被膜层上，并在细胞质中进行最终组装。尽管人们已经对衣壳在细胞核中的组装有了很多了解，但驱动细胞贴壁和贴壁的分子机制还很不清楚。位于内皮层的蛋白质被认为提供衣壳和外皮层之间的界面。外皮层结构似乎是重要的颗粒稳定性。此外，外珠被层面向包膜的内层。因此，外被可能参与最终的病毒粒子包装和维持粒子的完整性。最后，位于病毒体外被膜层的几种蛋白质靶向先天限制系统，其被设计为感测进入的病毒体并在非常早期阶段阻断病毒基因表达和复制。因此，外被膜中的蛋白质在病毒的生命周期中起着几种重要的作用。关于建立和维持外皮层结构所需的蛋白质相互作用网络的知识尚不发达。我们已经表明，在蛋白质组学分析，组成的HCMV被膜之间的菌株是非常保守的，表明被膜组装是高度调节。这表明，在病毒上存在选择性压力，以在给定的摩尔浓度下包装某些蛋白质，以达到颗粒包装和启动其复制周期的最佳条件。本研究的目的是探讨HCMV外被膜组装的机制及其在继发性感染中的作用。该项目集中于以下假设：（i）在HCMV感染细胞的细胞核中形成被膜蛋白网络，（ii）这些蛋白质凭借主要基质蛋白pp65的穿梭功能靶向细胞质病毒组装位点，（iii）细胞质前被膜复合物随后形成为外被膜组装的中心，以及（iv）外皮层通过与自噬成分的相互作用驱动次级吞噬。

The morphogenesis of human cytomegalovirus (HCMV) particles is a complicated process that requires multiple interactions of viral proteins to assemble capsids, to attach the tegument layer, and to perform final envelopment in the cytoplasm. Although a lot has been learned about capsid assembly in the nucleus, the molecular mechanisms that drive tegumentation and envelopment are only poorly defined. The proteins located in the inner tegument are thought to provide the interface between capsids and the outer tegument. The outer tegument structure appears to be important for particle stability. In addition, the outer tegument layer faces the inner lamina of the envelope. Thus the outer tegument is likely involved in final virion envelopment and in maintaining the integrity of the particles. Finally, several proteins, located in the outer tegument layer of the virion target innate restriction systems that are designed to sense incoming virions and to block viral gene expression and replication at a very early stage. Thus the proteins in the outer tegument serve several important functions in the virus life cycle. The knowledge about the protein interaction networks that are required to set up and maintain the outer tegument structure is underdeveloped. We have shown in proteomic analyses that the composition of the HCMV tegument is remarkably conserved between strains, indicating that tegument assembly is highly regulated. This suggests that there is selective pressure on the virus to package certain proteins at a given molarity to achieve optimal conditions for particle envelopment and for the initiation of its replication cycle. The work programme devised here aims at investigating the mechanisms that drive outer HCMV tegument assembly and its role in secondary envelopment. The project focuses on the hypotheses that (i) a tegument protein network is formed in the nucleus of HCMV infected cells, that (ii) these proteins are targeted to cytoplasmic viral assembly sites by virtue of the shuttling function of the major matrix protein pp65, that (iii) a cytoplasmic pre-tegument complex is subsequently formed as a hub for outer tegument assembly, and that (iv) the outer tegument then drives secondary envelopment by interaction with components of autophagy.