Young D F, Chatziandreou N, He B, Goodbourn S, Lamb R A, Randall R E
School of Biomedical Sciences, University of St. Andrews, Fife, Scotland KY16 9TS, United Kingdom.
J Virol. 2001 Apr;75(7):3363-70. doi: 10.1128/JVI.75.7.3363-3370.2001.
Previous work has demonstrated that the V protein of simian virus 5 (SV5) targets STAT1 for proteasome-mediated degradation (thereby blocking interferon [IFN] signaling) in human but not in murine cells. In murine BF cells, SV5 establishes a low-grade persistent infection in which the virus fluxes between active and repressed states in response to local production of IFN. Upon passage of persistently infected BF cells, virus mutants were selected that were better able to replicate in murine cells than the parental W3 strain of SV5 (wild type [wt]). Viruses with mutations in the Pk region of the N-terminal domain of the V protein came to predominate the population of viruses carried in the persistently infected cell cultures. One of these mutant viruses, termed SV5 mci-2, was isolated. Sequence analysis of the V/P gene of SV5 mci-2 revealed two nucleotide differences compared to wt SV5, only one of which resulted in an amino acid substitution (asparagine [N], residue 100, to aspartic acid [D]) in V. Unlike the protein of wt SV5, the V protein of SV5 mci-2 blocked IFN signaling in murine cells. Since the SV5 mci-2 virus had additional mutations in genes other than the V/P gene, a recombinant virus (termed rSV5-V/P N(100)D) was constructed that contained this substitution alone within the wt SV5 backbone to evaluate what effect the asparagine-to-aspartic-acid substitution in V had on the virus phenotype. In contrast to wt SV5, rSV5-V/P N(100)D blocked IFN signaling in murine cells. Furthermore, rSV5-V/P N(100)D virus protein synthesis in BF cells continued for significantly longer periods than that for wt SV5. However, even in cells infected with rSV5-V/P N(100)D, there was a late, but significant, inhibition in virus protein synthesis. Nevertheless, there was an increase in virus yield from BF cells infected with rSV5-V/P N(100)D compared to wt SV5, demonstrating a clear selective advantage to SV5 in being able to block IFN signaling in these cells.
先前的研究表明，猿猴病毒5（SV5）的V蛋白在人类细胞而非鼠细胞中靶向信号转导和转录激活因子1（STAT1）进行蛋白酶体介导的降解（从而阻断干扰素[IFN]信号传导）。在鼠BF细胞中，SV5建立了一种低度持续性感染，其中病毒会根据局部IFN的产生在活跃状态和抑制状态之间波动。在传代持续感染的BF细胞后，筛选出了比SV5野生型（wt）亲本W3株更能在鼠细胞中复制的病毒突变体。V蛋白N端结构域的Pk区域发生突变的病毒在持续感染细胞培养物中携带的病毒群体中占主导地位。其中一种突变病毒被分离出来，称为SV5 mci-2。与wt SV5相比，SV5 mci-2的V/P基因序列分析显示有两个核苷酸差异，其中只有一个导致V蛋白中的氨基酸替换（天冬酰胺[N]，第100位残基，替换为天冬氨酸[D]）。与wt SV5的蛋白不同，SV5 mci-2的V蛋白在鼠细胞中阻断IFN信号传导。由于SV5 mci-2病毒在V/P基因以外的基因中还有其他突变，构建了一种重组病毒（称为rSV5-V/P N(100)D），其在wt SV5骨架内仅包含此替换，以评估V蛋白中天冬酰胺到天冬氨酸的替换对病毒表型有何影响。与wt SV5相比，rSV5-V/P N(100)D在鼠细胞中阻断IFN信号传导。此外，BF细胞中rSV5-V/P N(100)D病毒蛋白的合成持续时间比wt SV5长得多。然而，即使在感染rSV5-V/P N(100)D的细胞中，病毒蛋白合成也有后期但显著的抑制。尽管如此，与wt SV5相比，感染rSV5-V/P N(100)D的BF细胞的病毒产量有所增加，这表明SV5能够在这些细胞中阻断IFN信号传导具有明显的选择优势。
