Characterization of hantavirus N protein-mediated translation mechanism

性状（由申请方提供）：汉坦病毒属布尼亚病毒科成员，是一种有包膜的负链新兴RNA病毒和A类病原体，通过感染啮齿动物的排泄物雾化传播给人类。汉坦病毒已经进化出一种独特的翻译机制，由病毒核衣壳蛋白（N）操作，其优先促进病毒mRNA在宿主细胞中的翻译，其中细胞转录物竞争相同的翻译机制。这种翻译机制在概念上不同于迄今为止已知的所有翻译机制，并且促进加帽mRNA的翻译而不需要eIF 4F帽结合复合物。汉坦病毒mRNA含有独特的42-52个核苷酸长的5'非翻译区（UTR），在5'末端具有高度保守的三联体重复序列，其特异性结合三聚体N蛋白。此外，N还通过核糖体蛋白S19（RPS 19）与40 S核糖体亚基结合，该蛋白位于40 S亚基的头部区域。N优先将40 S核糖体亚基装载到病毒mRNA 5' UTR上，并促进它们在体外的翻译。我们将进一步表征5' UTR在细胞中通过N介导的翻译机制翻译病毒mRNA中的作用。我们将确定汉他病毒mRNA是否特异性地依赖于N-介导的翻译机制进行翻译。我们还将鉴定和表征N蛋白中的RPS 19结合结构域，并证明N-RPS 19相互作用在N介导的翻译策略中的作用。我们将确定N蛋白是否仍然与多核糖体相关mRNA中80 S核糖体的40 S亚基相关。这些研究将揭示一种新的翻译起始机制，这种机制可能是大多数负链RNA病毒在感染过程中用于促进其mRNA在宿主细胞中的翻译。拟议的研究还将揭示未来抗病毒药物设计的新靶点。

DESCRIPTION (provided by applicant): Hantaviruses, members of the Bunyaviridae family are enveloped negative stranded emerging RNA viruses and category A pathogens that are transmitted to humans through aerosolized excreta of infected rodents. Hantaviruses have evolved a unique translation mechanism, operated by viral nucleocapsid protein (N), which preferentially facilitates the translation of viral mRNAs in host cells where cellular transcripts are competing for the same translation machinery. This translation mechanism is conceptually different from all translation mechanisms known to date, and facilitates the translation of capped mRNAs without the requirement of eIF4F cap binding complex. Hantaviral mRNAs contain a unique 42-52 nucleotide long 5' untranslated region (UTR) harboring a highly conserved triplet repeat sequence at the 5' terminus that specifically binds to the trimeric N protein. In addition, N also binds to the 40S ribosomal subunit via the ribosomal protein S19 (RPS19), located at the head region of 40S subunit. N preferentially loads 40S ribosomal subunits onto the viral mRNA 5' UTR and facilitates their translation in vitro. We will further characterize the role of 5' UTR in the translation of viral mRNAs by N-mediated translation mechanism in cells. We will determine whether hantaviral mRNAs specifically depend on N-mediated translation mechanism for their translation. We will also identify and characterize the RPS19 binding domain in N protein and demonstrate the role of N-RPS19 interaction in N-mediated translation strategy. We will determine whether N protein remains associated with the 40S subunit of 80S ribosome in polysome associated mRNAs. These studies will shed light on a novel mechanism of translation initiation that likely most negative stranded RNA viruses use to favor the translation of their mRNAs in host cells during infection. The proposed studies will also reveal novel targets for antiviral drug design in future.