探索非洲爪蟾卵母细胞中的RNA病毒复制。

Exploring RNA virus replication in Xenopus oocytes.

作者信息

Gamarnik Andrea V, Andino Raul

机构信息

Fundacion Instituto Leloir, Buenos Aires, Argentina.

出版信息

Methods Mol Biol. 2006;322:367-78. doi: 10.1007/978-1-59745-000-3_26.

DOI:10.1007/978-1-59745-000-3_26

PMID:16739737

Abstract

Microinjection of poliovirus RNA in Xenopus oocytes initiates a complete and authentic viral replication cycle that yields newly synthesized infectious virus. This system can be used to study the molecular mechanism of the different steps involved in virus replication. Interestingly, viral replication only occurs if poliovirus RNA is coinjected with factors present in HeLa extracts. We have determined that two HeLa cell factors are required for viral replication in oocytes, one involved in initiation of translation (polio translation factor) and the other in RNA synthesis. Thus, microinjection in oocytes provides a strategy to identify and further analyze the function of these host cell factors and to biochemically dissect the mechanism of initiation of poliovirus translation and RNA synthesis. Here, we review protocols, approaches, and potential issues that can be addressed using the oocyte system.

摘要

将脊髓灰质炎病毒RNA显微注射到非洲爪蟾卵母细胞中可启动完整且真实的病毒复制周期，产生新合成的感染性病毒。该系统可用于研究病毒复制过程中不同步骤的分子机制。有趣的是，只有当脊髓灰质炎病毒RNA与HeLa细胞提取物中的因子共同注射时，病毒复制才会发生。我们已经确定，卵母细胞中的病毒复制需要两种HeLa细胞因子，一种参与翻译起始（脊髓灰质炎翻译因子），另一种参与RNA合成。因此，在卵母细胞中进行显微注射提供了一种策略，可用于鉴定和进一步分析这些宿主细胞因子的功能，并从生化角度剖析脊髓灰质炎病毒翻译起始和RNA合成的机制。在此，我们综述了可使用卵母细胞系统解决的实验方案、方法及潜在问题。