Activity of vesicular stomatitis virus M protein mutants in cell rounding is correlated with the ability to inhibit host gene expression and is not correlated with virus assembly function
Activity of vesicular stomatitis virus M protein mutants in cell rounding is correlated with the ability to inhibit host gene expression and is not correlated with virus assembly function
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DOI：
10.1006/viro.1996.8415
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发表时间：
1997-03-03
期刊：
影响因子：
3.7
通讯作者：
McKenzie, MO
中科院分区：
文献类型：
--
作者：
Lyles, DS;McKenzie, MO
水疱性口炎病毒（VSV）的基质(M)蛋白除了在病毒组装中起作用外，还参与病毒诱导的细胞围合和宿主定向基因表达的抑制。先前的实验表明，两种M蛋白突变体在基因上将M蛋白抑制宿主定向基因表达的能力与其在病毒组装中的功能分离：来自tsO82病毒的M蛋白在病毒组装中具有完全功能，但在抑制宿主定向基因表达方面存在缺陷，而缺乏氨基酸4-21的MN1缺失突变体抑制宿主定向基因表达，但在病毒组装中不起作用。本实验将这两种突变M蛋白与wt M蛋白诱导的细胞圆缩进行了比较。用体外转录的M蛋白mRNA转染BHK细胞，并在转染后24小时评估细胞圆切程度。MN1蛋白在诱导细胞圆缩方面几乎与wt M蛋白一样有效，而从转染RNA中表达的tsO82 M蛋白不能诱导细胞圆缩，尽管它确实使BHK细胞具有较少的细长形状。这些结果表明，MN1和tsO82 M蛋白诱导细胞圆缩的能力与其病毒组装功能无关。相反，这些突变体的细胞围合活性与它们抑制宿主定向基因表达的能力有关。先前的数据表明，这两种细胞病变活动可以分离，可以很容易地通过所需的M蛋白表达的定量差异来解释。tsO82病毒感染BHK细胞或L细胞均可诱导细胞圆缩，但与感染wt VSV后相比，细胞圆缩延迟，这表明tsO82 M蛋白保留了一定的细胞病变活性。荧光显微镜观察转染细胞中肌动蛋白、波形蛋白和微管蛋白的分布。在转染了tso82mmrna的细胞中，这些细胞骨架元件与阴性对照转染的细胞无法区分。在转染wt M或MN1 mRNA后变圆的细胞中，含有肌动蛋白的丝被重组成一个厚的核周环，但没有解聚。相反，微管蛋白和波形蛋白似乎弥散分布在圆形细胞的细胞质中。这些结果支持了M蛋白诱导的细胞圆缩是由微管和/或中间细丝的解聚引起的。(C) 1997学术出版社。
In addition to its role in virus assembly, the matrix (M) protein of vesicular stomatitis virus (VSV) is involved in virus-induced cell rounding and inhibition of host-directed gene expression. Previous experiments have shown that two M protein mutants genetically dissociate the ability of M protein to inhibit host-directed gene expression from its function in virus assembly: M protein from tsO82 virus is fully functional in virus assembly but defective in the inhibition of host-directed gene expression, while the MN1 deletion mutant, which lacks amino acids 4-21, inhibits host-directed gene expression but cannot function in virus assembly. Experiments presented here compared cell rounding induced by these two mutant M proteins to that of wt M protein. BHK cells were transfected with M protein mRNA transcribed in vitro, and the extent of cell rounding was evaluated at 24 hr posttransfection. The MN1 protein was nearly as effective as wt M protein in the induction of cell rounding, while tsO82 M protein expressed from transfected RNA was not able to induce cell rounding above that observed in negative controls without M protein, although it did cause BHK cells to have a less elongated shape. These results indicate that the ability of MN1 and tsO82 M proteins to induce cell rounding is not correlated with their virus assembly function. Instead the cell rounding activity of these mutants is correlated with their ability to inhibit host-directed gene expression. Previous data suggesting that these two cytopathic activities could be dissociated can be readily accounted for by quantitative differences in M protein expression required. Infection of either BHK cells or L cells with tsO82 virus induced cell rounding, although cell rounding was delayed relative to that following infection with wt VSV, suggesting that tsO82 M protein retains some cytopathic activity. The distribution of actin, vimentin, and tubulin in transfected cells was determined by fluorescence microscopy. In cells transfected with tsO82 M mRNA, these cytoskeletal elements were indistinguishable from those of negative control transfected cells. In cells rounded as a result of transfection with wt M or MN1 mRNA, actin-containing filaments were reorganized into a thick perinuclear ring but were not depolymerized. In contrast, tubulin and vimentin appeared to be diffusely distributed throughout the cytoplasm of rounded cells. These results support the idea that cell rounding induced by M protein results from the depolymerization of microtubules and/or intermediate filaments. (C) 1997 Academic Press.