Sendai virus C proteins are categorically nonessential gene products but silencing their expression severely impairs viral replication and pathogenesis
Sendai virus C proteins are categorically nonessential gene products but silencing their expression severely impairs viral replication and pathogenesis
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DOI：
10.1046/j.1365-2443.1998.00170.x
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发表时间：
1998-02-01
期刊：
影响因子：
2.1
通讯作者：
Nagai, Y
中科院分区：
文献类型：
--
作者：
Kurotani, A;Kiyotani, K;Nagai, Y
背景资料：仙台病毒（SeV）的P/C mRNA编码一套嵌套的辅助蛋白C ′、C、Y1和Y2，统称为C蛋白，分别起始于ACG/81和AUG/114，183，201在相对于磷酸（P）蛋白的ORF的+1框中，磷酸（P）蛋白是RNA聚合酶的较小亚基。其中，C是在感染细胞中表达的主要种类，其摩尔比是其他三种的几倍。然而，它们的功能仍然是一个谜。甚至还没有确定C蛋白是否是病毒复制所必需的。许多其他病毒在单负病毒目编码C-样蛋白，但它们的作用也仍然有待defined.Results：通过利用最近开发的反向遗传学系统，以恢复感染性SeV的cDNA，我们创建了突变体，其中C蛋白的框架是不同的沉默。不表达C和C '，但表达Y1和Y2的C/C'（-）病毒在各种组织和组织的组织培养细胞以及鸡胚中的复制中严重减毒。更值得注意的是，它们几乎完全不能在自然宿主中高效生长，因此对小鼠无致病性。在C/C '（-）病毒中，基因表达和基因组复制似乎都受损。此外，沉默的Y1和Y2的表达也是可能的，和严重受损，但可行的克隆，4C（-）病毒，被分离出来，不表达的四个C蛋白。结论：SeV的C蛋白是绝对非必需的基因产物，但大大有助于在体外的完整复制能力，是不可或缺的在体内增殖和发病。这项研究代表了第一个全面的功能评估的辅助C蛋白的单负性病毒。
Background: The P/C mRNA of Sendai virus (SeV), a prototypic member of the family Paramyxoviridae in the Mononegavirales superfamily comprising a large number of nonsegmented negative strand RNA viruses, encodes a nested set of accessory proteins, C', C, Y1 and Y2, referred to collectively as C proteins, initiating, respectively, at ACG/81 and AUGs/114, 183, 201 in the +1 frame relative to the ORF of phospho (P) protein, the smaller subunit of RNA polymerase. Among them, C is the major species expressed in infected cells at a molar ratio which is several-fold higher than the other three. However, their function has remained an enigma. It has not even been established whether or not the C proteins are essential for viral replication. Many other viruses in Mononegavirales encode C-like proteins, but their roles also remain to be defined.Results: By taking advantage of a recently developed reverse genetics system to recover infectious SeV from cDNA, we created mutants in which C protein frames were variously silenced. C/C'(-) viruses which did riot express C and C', but did express Y1 and Y2, were severely attenuated in replication in tissue culture cells of various species and tissues, as well as in embryonated chicken eggs. More notably, they were almost totally incapable of growing productively in-and hence nonpathogenic for mice-the natural host. Both gene expression and genome replication appeared to be impaired in C/C'(-) viruses. Additionally silencing the Y1 and Y2 expression was also possible, and a critically impaired but viable clone, the 4C(-) virus, was isolated which expressed none of the four C proteins.Conclusion: SeV C proteins are categorically nonessential gene products, but greatly contribute to full replication capability in vitro and are indispensable for in viva multiplication and pathogenesis. This study represents the first comprehensive functional assessment of the accessary C protein for Mononegavirales.