Research Training in Virus RNA Localization

描述（由申请方提供）：呼肠孤病毒是分段dsRNA病毒呼肠孤病毒科的原型成员，包括重要的人类（轮状病毒）和兽医（蓝舌病）病原体。10种独特的正义病毒RNA定位于病毒工厂以包装成子代核心颗粒的机制尚不清楚，将通过本提案中的实验进行解决。将通过原位杂交和感染细胞中溴尿苷标记的病毒正义RNA的免疫染色来检查呼肠孤病毒正义RNA转录后所采取的路径。将通过将病毒或对照RNA显微注射到呼肠孤病毒感染的细胞或表达呼肠孤病毒RNA结合蛋白的转染细胞中的研究来检查病毒RNA定位于病毒工厂的机制。如果病毒RNA特异性定位于病毒工厂，则将鉴定参与定位的反式和顺式作用因子。还将研究RNA到工厂的非特异性定位机制。了解RNA定位过程可能会导致新的抗病毒策略的发展，并将有助于发展一个强大的反向遗传学系统的dsRNA病毒。

DESCRIPTION (provided by applicant): Reoviruses are prototypical members of the Reoviridae family of segmented dsRNA viruses that include important human (rotavirus) and veterinary (blue tongue disease) pathogens. The mechanisms by which 10 unique positive-sense viral RNAs are localized to viral factories for packaging into progeny core particles is poorly understood and will be addressed by the experiments in this proposal. The paths taken by reovirus positive-sense RNAs after they are transcribed will be examined by both in situ hybridization and immunostaining of bromouridine labeled viral positive-sense RNAs in infected cells. The mechanism for localization of viral RNAs to viral factories will be examined by microinjection studies of viral or control RNAs into reovirus infected cells, or transfected cells expressing reovirus RNA-binding proteins. If viral RNAs are specifically localized to viral factories, trans- and cis-acting factors involved in localization will be identified. Mechanisms for non-specific localization of RNA to factories will also be examined. Understanding the RNA localization process may lead to the development of new anti-viral strategies, and will aid in the development of a robust reverse genetics system for dsRNA viruses.