Guanidine hydrochloride inhibits mammalian orthoreovirus growth by reversibly blocking the synthesis of double-stranded RNA
Guanidine hydrochloride inhibits mammalian orthoreovirus growth by reversibly blocking the synthesis of double-stranded RNA
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DOI：
10.1128/jvi.02106-06
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发表时间：
2007-05-01
影响因子：
5.4
通讯作者：
Nibert, Max L.
中科院分区：
文献类型：
--
作者：
Murray, Kenneth E.;Nibert, Max L.
已知毫摩尔浓度的盐酸胍（GuHCl）抑制许多具有正义RNA基因组的植物和动物病毒的复制。例如，GuHCl与脊髓灰质炎病毒蛋白2C（ATP酶）的核苷酸结合区可逆地相互作用，导致对病毒负义RNA合成的特异性抑制。因此，GuHCl的使用允许脊髓灰质炎病毒基因表达和RNA复制的时空分离，并提供了在体外复制反应期间同步启动负义RNA合成的有力工具。在本研究中，我们研究了盐酸胍对哺乳动物正呼肠孤病毒（MRV），一种来自呼肠孤病毒科的双链RNA（dsRNA）病毒的影响。15 mM盐酸胍可逆性抑制小鼠L929细胞中的MRV生长。此外，15 mM GuHCl提供了病毒dsRNA合成的特异性抑制，同时保留了正义RNA合成和病毒mRNA翻译。通过使用GuHCl提供MRV基因表达和基因组复制的时间分离，我们获得的证据表明，MRV初级转录支持足够的蛋白质合成组装形态正常的病毒工厂含有功能性复制酶复合物。此外，GuHCl和放线菌酮的协调使用使我们能够证明MRV dsRNA合成可以在不存在正在进行的蛋白质合成的情况下发生，尽管程度有限。利用可逆抑制MRV dsRNA合成的未来研究将集中于阐明GuHCl的靶标以及MRV复制酶复合物的组分。
Millimolar concentrations of guanidine hydrochloride (GuHCl) are known to inhibit the replication of many plant and animal viruses having positive-sense RNA genomes. For example, GuHCl reversibly interacts with the nucleotide-binding region of poliovirus protein 2C(ATPase), resulting in a specific inhibition of viral negative-sense RNA synthesis. The use of GuHCl thereby allows for the spatiotemporal separation of poliovirus gene expression and RNA replication and provides a powerful tool to synchronize the initiation of negative-sense RNA synthesis during in vitro replication reactions. In the present study, we examined the effect of GuHCl on mammalian orthoreovirus (MRV), a double-stranded RNA (dsRNA) virus from the family Reoviridae. MRV growth in murine L929 cells was reversibly inhibited by 15 mM GuHCl. Furthermore, 15 mM GuHCl provided specific inhibition of viral dsRNA synthesis while sparing both positive-sense RNA synthesis and viral mRNA translation. By using GuHCl to provide temporal separation of MRV gene expression and genome replication, we obtained evidence that MRV primary transcripts support sufficient protein synthesis to assemble morphologically normal viral factories containing functional replicase complexes. In addition, the coordinated use of GuHCl and cycloheximide allowed us to demonstrate that MRV dsRNA synthesis can occur in the absence of ongoing protein synthesis, although to only a limited extent. Future studies utilizing the reversible inhibition of MRV dsRNA synthesis will focus on elucidating the target of GuHCl, as well as the components of the MRV replicase complexes.