School of Biology, Centre for Biomolecular Sciences, University of St. Andrews, St. Andrews, Fife, United Kingdom.
J Virol. 2013 May;87(9):4798-807. doi: 10.1128/JVI.03383-12. Epub 2013 Feb 28.
Preparations of parainfluenza virus 5 (PIV5) that are potent activators of the interferon (IFN) induction cascade were generated by high-multiplicity passage in order to accumulate defective interfering virus genomes (DIs). Nucleocapsid RNA from these virus preparations was extracted and subjected to deep sequencing. Sequencing data were analyzed using methods designed to detect internal deletion and "copyback" DIs in order to identify and characterize the different DIs present and to approximately quantify the ratio of defective to nondefective genomes. Trailer copybacks dominated the DI populations in IFN-inducing preparations of both the PIV5 wild type (wt) and PIV5-VΔC (a recombinant virus that does not encode a functional V protein). Although the PIV5 V protein is an efficient inhibitor of the IFN induction cascade, we show that nondefective PIV5 wt is unable to prevent activation of the IFN response by coinfecting copyback DIs due to the interfering effects of copyback DIs on nondefective virus protein expression. As a result, copyback DIs are able to very rapidly activate the IFN induction cascade prior to the expression of detectable levels of V protein by coinfecting nondefective virus.
为了积累缺陷干扰病毒基因组（DI），通过高倍数传代制备了能够强烈激活干扰素（IFN）诱导级联反应的副流感病毒 5（PIV5）。从这些病毒制剂中提取核衣壳 RNA，并进行深度测序。使用旨在检测内部缺失和“回文”DI 的方法分析测序数据，以鉴定和表征存在的不同 DI，并大致定量缺陷和非缺陷基因组的比例。在 IFN 诱导制剂中，无论是 PIV5wt（野生型）还是 PIV5-VΔC（一种不编码功能性 V 蛋白的重组病毒），尾部回文都主导着 DI 群体。尽管 PIV5 V 蛋白是 IFN 诱导级联反应的有效抑制剂，但我们表明，由于回文 DI 对非缺陷病毒蛋白表达的干扰作用，非缺陷 PIV5wt 无法阻止复制回文 DI 对 IFN 反应的激活。因此，在非缺陷病毒表达可检测水平的 V 蛋白之前，回文 DI 能够非常迅速地激活 IFN 诱导级联反应。
