Contributions of two nuclear localization signals of influenza A virus nucleoprotein to viral replication
Contributions of two nuclear localization signals of influenza A virus nucleoprotein to viral replication
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DOI：
10.1128/jvi.01434-06
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发表时间：
2007-01-01
影响因子：
5.4
通讯作者：
Kawaoka, Yoshihiro
中科院分区：
文献类型：
--
作者：
Ozawa, Makoto;Fujii, Ken;Kawaoka, Yoshihiro
甲型流感病毒的RNA基因组在细胞核内转录和复制，与病毒聚合酶亚单位蛋白(PA、PB1和PB2)和核蛋白(NP)形成病毒核糖核蛋白复合体(VRNPs)。NP是vRNPs的主要组成部分，至少有两个氨基酸序列作为核定位信号(NLSS)：非常规NLS(残基3至13；NLS1)和二分NLS(残基198至216；NLS2)。虽然已知这两种NLS都在核运输中发挥作用，但它们对病毒复制的相对贡献却知之甚少。因此，我们研究了它们在NP亚细胞/硫核定位、病毒RNA(VRNA)转录和病毒复制中的贡献。取消非常规NLS导致NP主要定位于细胞质，并影响其在vRNA转录中的活性。然而，我们能够创造出一种病毒，其NP在NLS1中包含已知的氨基酸替换，从而取消其核定位功能，尽管该病毒被高度减毒。这些结果表明，虽然非常规NLS对病毒复制不是必需的，但对于有效的病毒mRNA合成是必要的。另一方面，二聚体NLS对NP的核转运的贡献有限，对vRNA转录和NP的核仁积累是必不可少的。无法生成NLS2不起作用的病毒。因此，NP的两段NLS对甲型流感病毒的复制是必不可少的，而不是非常规的NLS。
The RNA genome of influenza A virus, which forms viral ribonucleoprotein complexes (vRNPs) with viral polymerase subunit proteins (PA, PB1, and PB2) and nucleoprotein (NP), is transcribed and replicated in the nucleus. NP, the major component of vRNPs, has at least two amino acid sequences that serve as nuclear localization signals (NLSs): an unconventional NLS (residues 3 to 13; NLS1) and a bipartite NLS (residues 198 to 216; NLS2). Although both NLSs are known to play a role in nuclear transport, their relative contributions to viral replication are poorly understood. We therefore investigated their contributions to NP subcellular/sulmuclear localization, viral RNA (vRNA) transcription, and viral replication. Abolishing the unconventional NLS caused NP to localize predominantly to the cytoplasm and affected its activity in vRNA transcription. However, we were able to create a virus whose NP contained amino acid substitutions in NLS1 known to abolish its nuclear localization function, although this virus was highly attenuated. These results indicate that while the unconventional NLS is not essential for viral replication, it is necessary for efficient viral mRNA synthesis. On the other hand, the bipartite NLS, whose contribution to the nuclear transport of NP is limited, was essential for vRNA transcription and NP's nucleolar accumulation. A virus with nonfunctional NLS2 could not be generated. Thus, the bipartite NLS, but not the unconventional NLS, of NP is essential for influenza A virus replication.