Characterization of the Interaction between Hantavirus Nucleocapsid Protein (N) and Ribosomal Protein S19 (RPS19)
Characterization of the Interaction between Hantavirus Nucleocapsid Protein (N) and Ribosomal Protein S19 (RPS19)
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DOI：
10.1074/jbc.m110.210179
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发表时间：
2011-04-01
影响因子：
4.8
通讯作者：
Mir, Mohammad A.
中科院分区：
文献类型：
--
作者：
Cheng, Erdong;Haque, Absarul;Mir, Mohammad A.
汉坦病毒属布尼亚病毒科，是一种新出现的负链RNA病毒和A类病原体，当通过受感染啮齿动物宿主的排泄物雾化传播给人类时，会引起严重疾病。汉坦病毒已经进化出一种新的翻译起始机制，由核衣壳蛋白（N）操作，其优先促进病毒mRNA的翻译。N与核糖体蛋白S19（RPS 19）结合，RPS 19是40 S核糖体亚基的结构组分。此外，N还结合病毒mRNA 5'帽和病毒mRNA 5' UTR的高度保守的三联体重复序列。N在末端帽和5' UTR处的同时结合有利于在翻译起始期间核糖体加载到病毒转录物上。我们表征了N和RPS 19之间的结合，并证明了N-RPS 19相互作用在N介导的翻译起始机制中的作用。我们表明，N特异性结合RPS 19具有高亲和力和1：1的结合化学计量。N-RPS 19相互作用是一个由蛋白质驱动的过程。RPS 19与N.使用T7 RNA聚合酶，我们合成了汉坦病毒S片段mRNA，其与细胞中病毒RNA依赖性RNA聚合酶产生的转录物相匹配。我们发现，N-RPS 19相互作用在细胞和兔网织红细胞裂解物中的这种mRNA的翻译中起着关键作用。我们的研究结果表明，N-介导的翻译起始机制，它引诱宿主翻译机制的病毒转录本的优先翻译，主要取决于N-RPS 19的相互作用。我们认为N-RPS 19相互作用是一个新的靶点，可以关闭N介导的翻译策略，从而关闭细胞中的病毒复制。
Hantaviruses, members of the Bunyaviridae family, are negative-stranded emerging RNA viruses and category A pathogens that cause serious illness when transmitted to humans through aerosolized excreta of infected rodent hosts. Hantaviruses have evolved a novel translation initiation mechanism, operated by nucleocapsid protein (N), which preferentially facilitates the translation of viral mRNAs. N binds to the ribosomal protein S19 (RPS19), a structural component of the 40 S ribosomal subunit. In addition, N also binds to both the viral mRNA 5' cap and a highly conserved triplet repeat sequence of the viral mRNA 5' UTR. The simultaneous binding of N at both the terminal cap and the 5' UTR favors ribosome loading on viral transcripts during translation initiation. We characterized the binding between N and RPS19 and demonstrate the role of the N-RPS19 interaction in N-mediated translation initiation mechanism. We show that N specifically binds to RPS19 with high affinity and a binding stoichiometry of 1:1. The N-RPS19 interaction is an enthalpy-driven process. RPS19 undergoes a conformational change after binding to N. Using T7 RNA polymerase, we synthesized the hantavirus S segment mRNA, which matches the transcript generated by the viral RNA-dependent RNA polymerase in cells. We show that the N-RPS19 interaction plays a critical role in the translation of this mRNA both in cells and rabbit reticulocyte lysates. Our results demonstrate that the N-mediated translation initiation mechanism, which lures the host translation machinery for the preferential translation of viral transcripts, primarily depends on the N-RPS19 interaction. We suggest that the N-RPS19 interaction is a novel target to shut down the N-mediated translation strategy and hence virus replication in cells.