Mechanisms of Marburg virus gene expression

马尔堡病毒基因表达机制

• 批准号: 9922784
• 负责人: Rachel Fearns
• 金额: $ 53.48万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-08 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9922784
• 关键词: Affect; Africa; Antiviral Agents; Attention; Binding Proteins; Biological Assay; Biology; Category A pathogen; Cells; Characteristics; Chiroptera; Complex; Computer Analysis; Cytoplasm; Cytoplasmic Inclusion; DNA-Directed RNA Polymerase; Data; Development; Disease Outbreaks; ENG gene; Ebola virus; Elements; Ensure; Environment; Family; Family member; Filovirus; Frankfurt-Marburg Syndrome Virus; Future; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Genome; Goals; Heart; Human; Image; In Vitro; Length; Light; Maps; Messenger RNA; Microscopy; Molecular Biology; National Institute of Allergy and Infectious Disease; Pathogenicity; Play; Polyadenylation; Polyadenylation Pathway; Polymerase; Post-Transcriptional Regulation; Process; Promoter Regions; Proteins; RNA Polymerase I; RNA Stability; RNA Viruses; RNA metabolism; RNA-Binding Proteins; RNA-Directed RNA Polymerase; RNA-Protein Interaction; Recombinants; Research; Research Personnel; Respiratory syncytial virus; Ribosomes; Role; Series; Signal Transduction; Structure; Stuttering; System; Techniques; Testing; Transcription Initiation; Transcription Initiation Site; Transferase; Translations; Viral; Viral Genome; Virus; Virus Diseases; Virus Replication; Work; base; cis acting element; drug development; experimental study; innovation; insight; mRNA Stability; mortality; mutant; pathogenic virus; polyadenylated messenger RNA; promoter; reverse genetics; single molecule; stem; tool; trafficking; vaccine development; viral RNA; viral transmission

Marburg virus (MARV) belongs to the filovirus family and is highly pathogenic in humans. Despite being classified as Category A Priority Pathogen by NIAID, and its potential to cause large-scale outbreaks, similar to the recent Ebola virus outbreak, research on MARV lags significantly behind that on other non-segmented negative sense (NNS) RNA viruses. Here, we propose to perform in-depth analyses of MARV transcription and gene expression. Dissecting the mechanisms of MARV gene expression will not only be instrumental for the targeted development of antiviral drugs, it will also reveal unifying paradigms and distinctions between the NNS RNA viruses. The filovirus genome is transcribed by a virally encoded RNA-dependent RNA polymerase complex, which is capable of generating capped and polyadenylated mRNAs. This process occurs in the cell cytoplasm, close to ribosomes and cellular RNA binding proteins. This project will examine three different stages of MARV gene expression. In Aim 1, we will elucidate the mechanism of transcription initiation at the MARV promoter and investigate the role of structural features of the polymerase in this process. Notably, the MARV promoter sequence has some unusual features, and we intend to explore the functional relevance of these characteristics. In Aim 2, we will determine the function of conserved hairpin loops that are formed at the 5´ end of each MARV mRNA. We will explore the effect of these structures on transcription, RNA stability, trafficking and translation. In Aim 3, we will focus on mRNA polyadenylation and release. The mechanism of mRNA release is not well understood for any NNS RNA virus, and the results obtained in this aim will help to broaden our understanding of NNS RNA virus transcription strategies. This proposal brings together expertise in studying NNS RNA polymerases, MARV molecular biology, and mRNA-protein interactions. Together, the research team has established a unique tool set to achieve the goals of this proposal, including a MARV in vitro polymerase assay, various MARV reverse genetics systems, and highly innovative single molecule mRNA-protein binding assays. These studies will shine new light on a crucial aspect of MARV infection and enhance our understanding of NNS RNA virus biology.

Marburg病毒（MARV）属于Filovirus家族，在人类中具有高度致病性。尽管被归类为NIAID的优先病原体及其引起大规模爆发的潜力，类似于最近的埃博拉病毒爆发，但对MARV的研究滞后于其他非细分负面感（NNS）RNA病毒的研究显着落后于此。在这里，我们建议对MARV转录和基因表达进行深入分析。剖析MARV基因表达的机制不仅对抗病毒药的靶向发展具有重要作用，而且还将揭示NNS RNA病毒之间的统一范式和区别。 FILOVIRUS基因组由病毒编码的RNA依赖性RNA聚合酶复合物转录，该复合物能够产生限制和聚腺苷酸化的mRNA。该过程发生在细胞质中，接近核糖体和细胞RNA结合蛋白。该项目将检查MARV基因表达的三个不同阶段。在AIM 1中，我们将阐明MARV启动子的转录倡议机理，并研究聚合酶在此过程中的结构特征的作用。值得注意的是，MARV启动子序列具有一些异常的特征，我们打算探讨这些特征的功能相关性。在AIM 2中，我们将确定在每个MARV mRNA的5端形成的保守发夹环的功能。我们将探讨这些结构对转录，RNA稳定性，运输和翻译的影响。在AIM 3中，我们将重点放在mRNA聚腺苷酸化和释放上。对于任何NNS RNA病毒而言，mRNA释放的机制尚不很好，并且在此目标中获得的结果将有助于扩大我们对NNS RNA病毒转录策略的理解。该建议汇集了研究NNS RNA聚合酶，MARV分子生物学和mRNA-蛋白质相互作用方面的专业知识。研究团队共同建立了一个独特的工具，以实现该提案的目标，包括MARV体外聚合酶测定，各种MARV反向遗传学系统和高度创新的单分子mRNA-蛋白质结合测定。这些研究将为MARV感染的关键方面提供新的启示，并增强我们对NNS RNA病毒生物学的理解。