Weber F, Dunn E F, Bridgen A, Elliott R M
Division of Virology, Institute of Biomedical and Life Sciences, University of Glasgow, Church Street, Glasgow G11 5JR, Scotland, United Kingdom.
Virology. 2001 Mar 1;281(1):67-74. doi: 10.1006/viro.2000.0774.
The small (S) genomic segment of Bunyamwera virus (family Bunyaviridae, genus Bunyavirus) encodes the nucleocapsid protein, N, and a nonstructural protein, NSs, in overlapping reading frames. In order to elucidate the function of NSs, we established a plasmid-based minireplicon system using mammalian cells that express large amounts of T7 RNA polymerase. Expression of N, the viral polymerase protein (L), and a minireplicon containing a reporter gene was sufficient to reconstitute functional virus nucleocapsids. Coexpression of NSs, however, led to a dose-dependent decrease in reporter activity without affecting expression of controls. The inhibition could not be reversed by overexpression of N, L or the minireplicon, indicating that the NSs effect was not caused by a reduction in virus gene expression. The NSs proteins of two other members of the Bunyavirus genus, Guaroa virus and Lumbo virus, were also inhibitory in our system. The intracellular localisation of Bunyamwera virus NSs was investigated and found to be predominantly cytoplasmic, but intranuclear inclusion was also detected. Taken together, these data suggest that, in mammalian cells, the bunyavirus NSs protein controls the activity of the viral polymerase by a highly conserved mechanism.
布尼亚病毒科布尼亚病毒属的布尼亚姆韦拉病毒的小（S）基因组片段在重叠阅读框中编码核衣壳蛋白N和一种非结构蛋白NSs。为了阐明NSs的功能，我们利用表达大量T7 RNA聚合酶的哺乳动物细胞建立了基于质粒的微型复制子系统。N、病毒聚合酶蛋白（L）以及含有报告基因的微型复制子的表达足以重建功能性病毒核衣壳。然而，NSs的共表达导致报告基因活性呈剂量依赖性下降，而不影响对照的表达。N、L或微型复制子的过表达无法逆转这种抑制作用，这表明NSs的作用不是由病毒基因表达的降低引起的。布尼亚病毒属的另外两个成员瓜罗阿病毒和伦波病毒的NSs蛋白在我们的系统中也具有抑制作用。对布尼亚姆韦拉病毒NSs的细胞内定位进行了研究，发现其主要位于细胞质中，但也检测到核内包涵体。综上所述，这些数据表明，在哺乳动物细胞中，布尼亚病毒NSs蛋白通过一种高度保守的机制控制病毒聚合酶的活性。
