Molecular and Integrative Biosciences Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Helsinki, Finland.
Division of Structural Biology, Wellcome Centre for Human Genetics, University of Oxford, Oxford, United Kingdom.
mBio. 2018 Oct 2;9(5):e01242-18. doi: 10.1128/mBio.01242-18.
Double-stranded RNA (dsRNA) viruses package several RNA-dependent RNA polymerases (RdRp) together with their dsRNA genome into an icosahedral protein capsid known as the polymerase complex. This structure is highly conserved among dsRNA viruses but is not found in any other virus group. RdRp subunits typically interact directly with the main capsid proteins, close to the 5-fold symmetric axes, and perform viral genome replication and transcription within the icosahedral protein shell. In this study, we utilized phage Φ6, a well-established virus self-assembly model, to probe the potential roles of the RdRp in dsRNA virus assembly. We demonstrated that Φ6 RdRp accelerates the polymerase complex self-assembly process and contributes to its conformational stability and integrity. We highlight the role of specific amino acid residues on the surface of the RdRp in its incorporation during the self-assembly reaction. Substitutions of these residues reduce RdRp incorporation into the polymerase complex during the self-assembly reaction. Furthermore, we determined that the overall transcription efficiency of the Φ6 polymerase complex increased when the number of RdRp subunits exceeded the number of genome segments. These results suggest a mechanism for RdRp recruitment in the polymerase complex and highlight its novel role in virion assembly, in addition to the canonical RNA transcription and replication functions. Double-stranded RNA viruses infect a wide spectrum of hosts, including animals, plants, fungi, and bacteria. Yet genome replication mechanisms of these viruses are conserved. During the infection cycle, a proteinaceous capsid, the polymerase complex, is formed. An essential component of this capsid is the viral RNA polymerase that replicates and transcribes the enclosed viral genome. The polymerase complex structure is well characterized for many double-stranded RNA viruses. However, much less is known about the hierarchical molecular interactions that take place in building up such complexes. Using the bacteriophage Φ6 self-assembly system, we obtained novel insights into the processes that mediate polymerase subunit incorporation into the polymerase complex for generation of functional structures. The results presented pave the way for the exploitation and engineering of viral self-assembly processes for biomedical and synthetic biology applications. An understanding of viral assembly processes at the molecular level may also facilitate the development of antivirals that target viral capsid assembly.
双链 RNA (dsRNA) 病毒将几个 RNA 依赖性 RNA 聚合酶 (RdRp) 与其 dsRNA 基因组一起包装到一种称为聚合酶复合物的二十面体蛋白衣壳中。这种结构在 dsRNA 病毒中高度保守，但在任何其他病毒群中都没有发现。RdRp 亚基通常与主要衣壳蛋白直接相互作用，靠近五重对称轴，并在二十面体蛋白壳内进行病毒基因组复制和转录。在这项研究中，我们利用噬菌体 Φ6，一种成熟的病毒自组装模型，来探究 RdRp 在 dsRNA 病毒组装中的潜在作用。我们证明了 Φ6 RdRp 加速了聚合酶复合物的自组装过程，并有助于其构象稳定性和完整性。我们强调了 RdRp 表面特定氨基酸残基在自组装反应中的作用。这些残基的取代会减少 RdRp 在自组装反应中掺入聚合酶复合物。此外，我们确定当 RdRp 亚基数量超过基因组片段数量时，Φ6 聚合酶复合物的整体转录效率增加。这些结果表明了 RdRp 在聚合酶复合物中的募集机制，并强调了它在病毒衣壳组装中的新作用，除了 RNA 转录和复制的典型功能。双链 RNA 病毒感染范围广泛的宿主，包括动物、植物、真菌和细菌。然而，这些病毒的基因组复制机制是保守的。在感染周期中，形成一种蛋白衣壳，即聚合酶复合物。该衣壳的一个重要组成部分是病毒 RNA 聚合酶，它复制和转录封闭的病毒基因组。许多双链 RNA 病毒的聚合酶复合物结构得到了很好的描述。然而，对于构建这种复合物的层次分子相互作用，我们知之甚少。利用噬菌体 Φ6 自组装系统，我们对介导聚合酶亚基掺入聚合酶复合物以生成功能结构的过程有了新的认识。所呈现的结果为开发用于生物医学和合成生物学应用的病毒自组装过程铺平了道路。在分子水平上了解病毒组装过程也可能有助于开发针对病毒衣壳组装的抗病毒药物。
