Role of the PB2 627 Domain in Influenza A Virus Polymerase Function.
Role of the PB2 627 Domain in Influenza A Virus Polymerase Function.
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DOI：
10.1128/jvi.02467-16
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发表时间：
2017-04-01
影响因子：
5.4
通讯作者：
Fodor E
中科院分区：
文献类型：
--
作者：
Nilsson BE;Te Velthuis AJW;Fodor E
甲型流感病毒的RNA基因组由病毒RNA依赖的RNA聚合酶转录和复制，该聚合酶由亚基PA、PB1和PB2组成。高分辨率结构数据显示，聚合酶组装成一个中心聚合酶核心和几个辅助的高度灵活的突出结构域。辅助性PB2帽结合和PA核酸内切酶结构域都参与帽抢夺，但辅助性PB2 627结构域在甲型流感病毒宿主范围限制中的作用仍然知之甚少。在这项研究中，我们使用结构导向截断PB2亚基，以证明缺乏627结构域的PB2亚基在细胞核中积累，并与PB1和PA组装成异三聚体聚合酶。此外，我们发现缺乏PB2 627结构域的重组病毒聚合酶能够在体外进行帽抓取、帽依赖性转录起始和帽非依赖性ApG二核苷酸延伸，这表明流感病毒RNA聚合酶的PB2 627结构域不参与聚合酶的核心催化功能。然而，在细胞环境中，627结构域对转录和复制都是必不可少的。特别是，我们发现PB2 627结构域对于感染细胞中cRNA复制中间体的积累至关重要。总之，这些结果进一步加深了我们对PB2 627结构域在流感病毒RNA基因组转录和复制中的作用的理解。甲型流感病毒是一个重大的全球健康威胁，不仅在人类和鸟类中引起疾病，而且给世界各地的经济造成重大压力。甲型禽流感病毒聚合酶通常不能在哺乳动物宿主中有效地发挥作用，需要适应性突变来恢复聚合酶的活性。这些适应包括病毒聚合酶PB2亚基627结构域的突变，但这些突变如何在分子水平上使宿主适应仍有待确定。在本报告中，我们描述了627结构域在聚合酶功能中的作用，并为甲型流感病毒的复制机制提供了见解。
The RNA genome of influenza A viruses is transcribed and replicated by the viral RNA-dependent RNA polymerase, composed of the subunits PA, PB1, and PB2. High-resolution structural data revealed that the polymerase assembles into a central polymerase core and several auxiliary highly flexible, protruding domains. The auxiliary PB2 cap-binding and the PA endonuclease domains are both involved in cap snatching, but the role of the auxiliary PB2 627 domain, implicated in host range restriction of influenza A viruses, is still poorly understood. In this study, we used structure-guided truncations of the PB2 subunit to show that a PB2 subunit lacking the 627 domain accumulates in the cell nucleus and assembles into a heterotrimeric polymerase with PB1 and PA. Furthermore, we showed that a recombinant viral polymerase lacking the PB2 627 domain is able to carry out cap snatching, cap-dependent transcription initiation, and cap-independent ApG dinucleotide extension in vitro, indicating that the PB2 627 domain of the influenza virus RNA polymerase is not involved in core catalytic functions of the polymerase. However, in a cellular context, the 627 domain is essential for both transcription and replication. In particular, we showed that the PB2 627 domain is essential for the accumulation of the cRNA replicative intermediate in infected cells. Together, these results further our understanding of the role of the PB2 627 domain in transcription and replication of the influenza virus RNA genome. IMPORTANCE Influenza A viruses are a major global health threat, not only causing disease in both humans and birds but also placing significant strains on economies worldwide. Avian influenza A virus polymerases typically do not function efficiently in mammalian hosts and require adaptive mutations to restore polymerase activity. These adaptations include mutations in the 627 domain of the PB2 subunit of the viral polymerase, but it still remains to be established how these mutations enable host adaptation on a molecular level. In this report, we characterize the role of the 627 domain in polymerase function and offer insights into the replication mechanism of influenza A viruses.