Kobayashi T, Kamitani W, Zhang G, Watanabe M, Tomonaga K, Ikuta K
Department of Virology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
J Virol. 2001 Apr;75(7):3404-12. doi: 10.1128/JVI.75.7.3404-3412.2001.
Nuclear transport of viral nucleic acids is crucial to the life cycle of many viruses. Borna disease virus (BDV) belongs to the order Mononegavirales and replicates its RNA genome in the nucleus. Previous studies have suggested that BDV nucleoprotein (N) and phosphoprotein (P) have important functions in the nuclear import of the viral ribonucleoprotein (RNP) complexes via their nuclear targeting activity. Here, we showed that BDV N has cytoplasmic localization activity, which is mediated by a nuclear export signal (NES) within the sequence. Our analysis using deletion and substitution mutants of N revealed that NES of BDV N consists of a canonical leucine-rich motif and that the nuclear export activity of the protein is mediated through the chromosome region maintenance protein-dependent pathway. Interspecies heterokaryon assay indicated that BDV N shuttles between the nucleus and cytoplasm as a nucleocytoplasmic shuttling protein. Furthermore, interestingly, the NES region overlaps a binding site to the BDV P protein, and nuclear export of a 38-kDa form of BDV N is prevented by coexpression of P. These results suggested that BDV N has two contrary activities, nuclear localization and export activity, and plays a critical role in the nucleocytoplasmic transport of BDV RNP by interaction with other viral proteins.
病毒核酸的核转运对许多病毒的生命周期至关重要。博尔纳病病毒（BDV）属于单股负链RNA病毒目，在细胞核中复制其RNA基因组。先前的研究表明，BDV核蛋白（N）和磷蛋白（P）通过其核靶向活性在病毒核糖核蛋白（RNP）复合物的核输入中发挥重要作用。在此，我们表明BDV N具有细胞质定位活性，这是由该序列内的核输出信号（NES）介导的。我们使用N的缺失和替代突变体进行的分析表明，BDV N的NES由一个典型的富含亮氨酸基序组成，并且该蛋白的核输出活性是通过染色体区域维持蛋白依赖性途径介导的。种间异核体试验表明，BDV N作为一种核质穿梭蛋白在细胞核和细胞质之间穿梭。此外，有趣的是，NES区域与BDV P蛋白的结合位点重叠，并且P的共表达可阻止38 kDa形式的BDV N的核输出。这些结果表明，BDV N具有两种相反的活性，即核定位和输出活性，并通过与其他病毒蛋白的相互作用在BDV RNP的核质转运中起关键作用。
