Muramoto Yukiko, Takada Ayato, Fujii Ken, Noda Takeshi, Iwatsuki-Horimoto Kiyoko, Watanabe Shinji, Horimoto Taisuke, Kida Hiroshi, Kawaoka Yoshihiro
Laboratory of Microbiology, Department of Disease Control, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
J Virol. 2006 Mar;80(5):2318-25. doi: 10.1128/JVI.80.5.2318-2325.2006.
The genome of influenza A viruses comprises eight negative-strand RNA segments. Although all eight segments must be present in cells for efficient viral replication, the mechanism(s) by which these viral RNA (vRNA) segments are incorporated into virions is not fully understood. We recently found that sequences at both ends of the coding regions of the HA, NA, and NS vRNA segments of A/WSN/33 play important roles in the incorporation of these vRNAs into virions. In order to similarly identify the regions of the PB2, PB1, and PA vRNAs of this strain that are critical for their incorporation, we generated a series of mutant vRNAs that possessed the green fluorescent protein gene flanked by portions of the coding and noncoding regions of the respective segments. For all three polymerase segments, deletions at the ends of their coding regions decreased their virion incorporation efficiencies. More importantly, these regions not only affected the incorporation of the segment in which they reside, but were also important for the incorporation of other segments. This effect was most prominent with the PB2 vRNA. These findings suggest a hierarchy among vRNA segments for virion incorporation and may imply intersegment association of vRNAs during virus assembly.
甲型流感病毒的基因组由八个负链RNA片段组成。尽管所有八个片段都必须存在于细胞中才能实现高效的病毒复制，但这些病毒RNA（vRNA）片段被纳入病毒粒子的机制尚未完全了解。我们最近发现，A/WSN/33的HA、NA和NS vRNA片段编码区两端的序列在这些vRNAs纳入病毒粒子过程中发挥重要作用。为了同样鉴定该毒株PB2、PB1和PA vRNAs中对其纳入至关重要的区域，我们构建了一系列突变vRNAs，这些vRNAs拥有绿色荧光蛋白基因，两侧分别是各片段编码区和非编码区的部分序列。对于所有三个聚合酶片段，其编码区末端的缺失降低了它们纳入病毒粒子的效率。更重要的是，这些区域不仅影响它们所在片段的纳入，对其他片段的纳入也很重要。这种效应在PB2 vRNA中最为显著。这些发现表明vRNA片段在纳入病毒粒子方面存在层级关系，并且可能意味着在病毒组装过程中vRNAs之间存在片段间关联。
