Molecular anatomy of Sendai virus

仙台病毒（SeV）是最近重新命名的副粘病毒科呼吸道病毒属的一员。除了磷酸蛋白外，SeV P基因还使用两种辅助蛋白，V和C。虽然P蛋白已被确定为病毒RNA合成所必需的调节蛋白，但V和C蛋白在病毒复制和发病机制中的作用仍不清楚。甚至还没有确定这些蛋白质对于病毒复制是必需的还是仅仅是辅助的。这些问题现在正通过SeV反向遗传学得到澄清。当P基因的精确拷贝编码P蛋白时，共转录编辑的mRNA指导V蛋白。这个编辑事件包括假模板将一个G残基插入到模板中特定位置的新生链上，从而将阅读框寄存器从P ORF移到编码富含半胱氨酸蛋白的-1 ORF。通过破坏编辑位点或在下游引入无义突变，在cDNA质粒中更多的编辑位点产生SeV抗基因组的全长拷贝，我们创建了V基因表达完全缺陷（V(-)）或死亡c端半截短（VDELTAC）的突变病毒。他们的特征得出的结论是，尽管V蛋白在组织培养细胞中的病毒复制是完全没有必要的，但它对于维持小鼠体内的高病毒载量以引起肺炎和其他疾病是必不可少的。这种“奢侈”的体内功能主要由富含半胱氨酸的c端一半编码。V蛋白似乎是SeV应对一些早期宿主清除病毒反应所必需的。仙台C蛋白在未编辑的P mRNA和编辑的V mRNA中以嵌套的C'、C、Y1和Y2蛋白表达，使用相对于P ORF的+1框架，并从不同的起始密码子开始。其中，C蛋白是感染细胞中主要的表达种，其摩尔比是其他3种的数倍。我们首先沉默C‘和C蛋白，发现恢复的C/C’（-）病毒在组织培养细胞中的RNA合成受损，复制严重减弱。更值得注意的是，C/C'（-）病毒几乎完全不能在小鼠体内生长，因此对小鼠无致病性。沉默所有四种C蛋白也是可能的，从而产生一种更受损但可存活的克隆，即4C（-）病毒。由此可见，SeV C蛋白是一类非必需基因产物，但在体外完全复制能力中起着重要作用，在体内增殖和发病中是必不可少的。少

Sendai virus (SeV) is a member of the recently renamed Respirovirus genus of the Paramyxovirinae. SeV P gene gives use to two accessory proteins, V and C, in addition to the phospho (P) protein. While the P protein has been identified to be a modulator protein essential for viral RNA synthesis, the roles of the V and C proteins in viral replication and pathogenesis have remained unclear. It has not even been established whether these proteins are essential or simply auxiliary for viral replication. These issues are now being clarified by SeV reverse genetics.While the exact copy of the P gene encodes the P protein, the cotranscriptionally edited mRNA directs the V protein. This editing event involves pseudotemplated insertion of one G residue to the nascent chain at a specific site in the template, which shifts the reading frame register form the P ORF to access the -1 ORF encoding a cysteine-rich protein. By disrupting the editing site or introducing nonsense mutations just downstream … More of the editing site in a cDNA plasmid generating a full-length copy of the SeV antigenome, we created mutant viruses totally defective in V gene expression (V(-)) or with truncation of die C-terminal half (VDELTAC). Their characterization led to the conclusions that, although completely dispensable for viral replication in tissue cultured cells, the V protein was essential for maintaining a high viral load in mice to cause pneumonia and that. this "luxury" iii vivo function was encoded primarily by the cysteine-rich C-terminal half. The V protein appeared to be essential for SeV to cope with some early host response(s) recruited to clear the virus.The Sendai C protein is expressed as a nested set of proteins, C', C, Y1 and Y2, from both the unedited P mRNA and the edited V mRNA using the +1 frame relative to the P ORF and starting at different initiation codons. Among them, C protein is the major species expressed in infected cells at molar ratio several-fold higher than the other three. We first silenced C' and C proteins and found that the recovered C/C'(-) viruses were impaired in RNA synthesis and severely attenuated in replication in tissue cultured cells. More notably, the C/C'(-) viruses were almost totally incapable of growing productively in mice and, hence, nonpathogenic for the mice. Silencing all four C proteins was also possible, giving rise to a still more impaired but viable clone, the 4C(-) virus. Thus, it was concluded that SeV C proteins are categorically nonessential gene products but greatly contribute to full replication capability in vitro and are indispensable for in vivo multiplication and pathogenesis. Less