The cellular biology of turnip mosaic virus

萝卜花叶病毒的细胞生物学

• 批准号: RGPIN-2014-06193
• 负责人: Laliberté, JeanFrançois
• 金额: $ 3.42万
• 依托单位: Institut national de la recherche scientifique
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2016
• 资助国家: 加拿大
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=612083
• 关键词: cellular; biology; turnip; mosaic; virus

Genome replication of positive-strand RNA viruses is closely associated with membranes of the infected cell. For instance, turnip mosaic virus (TuMV) infection induces in the infected cells the formation of elaborate membranous organelle-like structures that sustain viral RNA synthesis and virus intercellular movement. The TuMV genome consists of a single RNA molecule of ~10 000 nucleotides of positive polarity. This RNA codes for a single polyprotein that is processed by three viral proteinases to yield eleven mature proteins. One protein, namely the 6K2 protein, is responsible for membrane recruitment. Two 6K2-induced structures are observed in infected cells: a large perinuclear globular structure that leads to the production of several small motile peripheral vesicles. These vesicles contain viral RNA, viral replication proteins and host factors, indicating that 6K2 vesicles harbour viral replication complexes. 6K2 vesicles are also found in association with plasmodesmata and can move directly into adjoining cells. Symplasmic movement of TuMV is thus achieved as a membrane-associated virus replication complex associated with 6K2. The intercellular movement of these vesicles depends on the secretory pathway and on myosin motors. My research objectives for the next 5 years will be to further elucidate the architecture of these membrane rearrangements and to identify some of the molecular events that lead to these changes. More precisely, my research team will use transmission electron microscopy (TEM) and tomography to get a 3-dimensional view of some of the cellular remodelling that are observed in infected cells and in vascular tissues, particularly in xylem vessels. We will also investigate the role of the tonoplast in virion assembly. My group will also map the molecular determinants of 6K2 that are involved in the formation of the vesicles. Preliminary experiments have indicated that the 6K2 protein has a tryptophan-based endoplasmic reticulum export signal that interacts with COPII coatomer components. We will confirm the involvement of these host proteins as well as the SNARE-like vesicle-associated protein (VAP) AtPva11, AtPva12 , and AtPva22 in 6K2 vesicle biogenesis. Finally, we will screen for host factors that are part of 6K2-associated viral replication complexes using a proteomic approach and validate their involvement in TuMV infection.

正链RNA病毒的基因组复制与感染细胞的膜密切相关。例如，芜菁花叶病毒（TuMV）感染诱导受感染细胞形成复杂的膜质细胞器样结构，维持病毒RNA合成和病毒细胞间运动。TuMV基因组由一个约10000个核苷酸的正极性RNA分子组成。这种RNA编码一种多蛋白，这种多蛋白经过三种病毒蛋白酶的加工，产生11种成熟蛋白。一种蛋白质，即6K2蛋白，负责膜的募集。在感染细胞中观察到两种6k2诱导的结构：一个大的核周球形结构，导致几个小的活动外周囊泡的产生。这些囊泡含有病毒RNA、病毒复制蛋白和宿主因子，表明6K2囊泡含有病毒复制复合体。6K2囊泡也与胞间连丝有关，可直接进入相邻细胞。因此，TuMV的同质运动是通过与6K2相关的膜相关病毒复制复合体实现的。这些囊泡的细胞间运动依赖于分泌途径和肌球蛋白运动。我未来5年的研究目标将是进一步阐明这些膜重排的结构，并确定导致这些变化的一些分子事件。更准确地说，我的研究小组将使用透射电子显微镜（TEM）和断层扫描来获得在感染细胞和维管组织，特别是木质部血管中观察到的一些细胞重构的三维视图。我们还将探讨舌质体在病毒粒子组装中的作用。我的小组还将绘制参与囊泡形成的6K2的分子决定因素。初步实验表明，6K2蛋白具有一个以色氨酸为基础的内质网输出信号，该信号与COPII涂层组分相互作用。我们将证实这些宿主蛋白以及snare样囊泡相关蛋白（VAP） AtPva11、AtPva12和AtPva22参与6K2囊泡的生物发生。最后，我们将使用蛋白质组学方法筛选作为6k2相关病毒复制复合体一部分的宿主因子，并验证它们在TuMV感染中的作用。