Picornavirus Molecular Biology

描述（由申请人提供）：本资助申请中提出的工作目标是加强我们对脊髓灰质炎病毒（PV）的机制的理解，PV是一种属于小核糖核酸病毒科的肠道病毒，可封装其基因组。该病毒家族包括大量人类和动物病原体，每年在全世界造成60多亿人感染。这些感染导致从轻微（普通感冒）到严重（小儿麻痹症）的各种疾病。尽管研究了多年，但PV生命周期中大多数步骤的细节仍然未知。然而，PV的扩散仍然是一个重要的医学问题，因为即使在wt PV的全球传播中断后，预计仍会发生流行性PV感染。本提案可分为三个部分。第一个目标是开发新的遗传工具来研究由PV病毒和密切相关的c -簇柯萨奇A病毒构建的嵌合体的衣壳化遗传学。我们期望嵌合病毒的形态发生表型将对分析衣壳化过程中衣壳和非结构蛋白的相互作用非常有用。该提案的第二个目标涉及肠病毒囊化的两种特异性抑制剂，乙酰胆碱和l -丁硫氨酸-亚砜胺（BSO），它们的作用有望在不同阶段阻断囊化。在本研究中，PV、cav和PV/C-CAV嵌合体将用于鉴定和分析参与衣壳化的蛋白质的逃逸突变体。在我们的第三个目标中，我们建议分析非结构蛋白2CATPase， 3CDpro， VPg和的形态发生中的作用，并寻找难以捉摸的RNA封装信号。我们计划利用遗传和生化研究来分析非结构蛋白2CATPase和3CDpro在衣壳化中的作用。VPg的作用将通过对数百个VPg突变体的大规模扫描来测试，目的是找到复制阳性但封装阴性的突变体。最后，我们将使用一种新的策略（密码子对优化）来扫描PV RNA以寻找难以捉摸的封装信号。由于衣壳化是一种独特的病毒过程，了解其机制将有助于开发针对肠病毒生命周期中这一特定步骤的抗病毒药物。2CATPase和3CDpro蛋白在肠病毒中高度保守，因此它们为治疗多种肠病毒疾病的药物开发提供了一个很好的靶点。相信这些研究和结果不仅对那些研究其他肠病毒包衣的研究者，而且对那些对小核糖核酸病毒或RNA病毒感兴趣的人都有意义。

DESCRIPTION (provided by applicant): It is the objective of the work proposed in this grant application to enhance our understanding of the mechanisms by which poliovirus (PV), an enterovirus belonging to the Picornaviridae, encapsidates its genome. This virus family includes a large number of human and animal pathogens that cause more than 6 billion human infections worldwide each year. These infections lead to a variety of diseases ranging from the mild (common cold) to the serious (poliomyelitis). In spite of research for many years the details of most steps in the life cycle of PV remain unknown. However, the proliferation of PV remains an important medical issue because epidemic PV infections are expected to occur even after the circulation of wt PV is interrupted globally. This proposal can be divided into 3 parts. The first aim deals with the development of new genetic tools to study the genetics of encapsidation of chimeras constructed from PV and the closely related C-cluster coxsackie A viruses. We expect that the morphogenesis phenotypes of the chimeric viruses will be very useful in analyzing the interaction of capsid and nonstructural proteins during encapsidation. The second aim of the proposal deals with two specific inhibitors of enterovirus encapsidation, hydantoin and L-buthionine-sulfoximine (BSO), whose effect is expected to block encapsidation at different stages. In this study PV, CAVs and PV/C-CAV chimeras will be used to identify and analyze escape mutants in proteins involved in encapsidation. In our third aim we propose to analyze the role in morphogenesis of non-structural proteins 2CATPase, 3CDpro, VPg, and, and search for an elusive RNA encapsidation signal. We plan to use both genetic and biochemical studies to analyze the role of nonstructural proteins 2CATPase and 3CDpro in encapsidation. The role of VPg will be tested by a large-scale scan of hundreds of VPg mutants with the aim of finding replication positive but encapsidation negative mutants. Finally, we will use a novel strategy (codon-pair optimization) to scan the PV RNA for an elusive encapsidation signal. Since encapsidation is a uniquely viral process an understanding of its mechanism will aid the development of antiviral drugs that target this particular step in the enteroviral life cycle. The 2CATPase and 3CDpro proteins are highly conserved among enteroviruses hence they provide an excellent target for drug development to treat multiple enteroviral diseases. It is believed that these studies and results will be of interest not only to those investigators who study the encapsidation of other enteroviruses but also to those who are interested in picornaviruses or RNA viruses in general.