The Matrix Protein of Vesicular Stomatitis Virus Binds Dynamin for Efficient Viral Assembly
The Matrix Protein of Vesicular Stomatitis Virus Binds Dynamin for Efficient Viral Assembly
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DOI：
10.1128/jvi.01400-10
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发表时间：
2010-12-01
影响因子：
5.4
通讯作者：
Gaudin, Yves
中科院分区：
文献类型：
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作者：
Raux, Helene;Obiang, Linda;Gaudin, Yves
基质蛋白(M)指导单病毒目病毒的组装和出芽过程。利用双杂交系统，研究了水疱性口炎病毒（VSV） M的氨基末端与动力蛋白pleckstrin同源结构域相互作用。这两种蛋白在被编码c-myc标记动力蛋白的质粒转染并被VSV感染的细胞中共免疫沉淀证实了这种相互作用。动力蛋白在病毒周期中的作用（除了它在病毒粒子内吞作用中的作用）是由病毒周期的后期对动力蛋白敏感这一事实提出的。通过丙氨酸扫描，我们发现了M蛋白的一个突变，该突变消除了这种相互作用并降低了病毒产量。突变病毒（M.L4A）的适应发生得很快，从而分离出了逆转录病毒，其中M蛋白尽管氨基酸序列与野生型不同，但恢复了与动力蛋白的显著相互作用。这证明突变表型是由于M和动力蛋白之间的相互作用丧失。突变病毒M.L4A的感染周期在后期被阻断，导致在细胞膜出芽过程中几乎没有子弹状病毒。这与细胞外围核衣壳的积累和VSV糖蛋白定位的不同模式有关。最后，我们发现m -动力蛋白相互作用影响网格蛋白依赖的内吞作用。我们的研究表明劫持内吞途径可能是包膜病毒组装和在质膜上出芽的一个重要特征。
Matrix proteins (M) direct the process of assembly and budding of viruses belonging to the Mononegavirales order. Using the two-hybrid system, the amino-terminal part of vesicular stomatitis virus (VSV) M was shown to interact with dynamin pleckstrin homology domain. This interaction was confirmed by coimmunoprecipitation of both proteins in cells transfected by a plasmid encoding a c-myc-tagged dynamin and infected by VSV. A role for dynamin in the viral cycle (in addition to its role in virion endocytosis) was suggested by the fact that a late stage of the viral cycle was sensitive to dynasore. By alanine scanning, we identified a single mutation of M protein that abolished this interaction and reduced virus yield. The adaptation of mutant virus (M.L4A) occurred rapidly, allowing the isolation of revertants, among which the M protein, despite having an amino acid sequence distinct from that of the wild type, recovered a significant level of interaction with dynamin. This proved that the mutant phenotype was due to the loss of interaction between M and dynamin. The infectious cycle of the mutant virus M.L4A was blocked at a late stage, resulting in a quasi-absence of bullet-shaped viruses in the process of budding at the cell membrane. This was associated with an accumulation of nucleocapsids at the periphery of the cell and a different pattern of VSV glycoprotein localization. Finally, we showed that M-dynamin interaction affects clathrin-dependent endocytosis. Our study suggests that hijacking the endocytic pathway might be an important feature for enveloped virus assembly and budding at the plasma membrane.