Encapsidation of the poxvirus genome

痘病毒基因组的衣壳化

基本信息

批准号：
8190013
负责人：
Paula Traktman
金额：
$ 22.95万
依托单位：
MEDICAL COLLEGE OF WISCONSIN
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-06-01 至 2013-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8190013
关键词：
ATP phosphohydrolase Antiviral Therapy Binding Cells Characteristics Complex Cytoplasm Cytoskeleton DNA DNA Probes DNA Viruses DNA biosynthesis Development Exposure to Family Figs - dietary Fright Gene Expression Genetic Genome Histones Human In Vitro Infection Investigation Lead Life Cycle Stages Link Lipid Bilayers Mediating Membrane Modeling Monkeypox virus Morphogenesis Oncolytic Organelles Pathway interactions Physical condensation Polyamines Poxviridae Process Protein Binding Proteins Pump Recombinant Vaccines Resolvase Role Smallpox Smallpox Viruses Specificity Spermidine Spermine Testing Vaccination Vaccinia virus Viral Viral Genome Viral Proteins Virion Virus Virus Diseases base genetic analysis human MCAM protein human morbidity human mortality in vivo insight member mutant pathogen telomere therapeutic target tool viral DNA weapons

ATP phosphohydrolase Antiviral Therapy Binding Cells Characteristics Complex Cytoplasm Cytoskeleton DNA DNA Probes DNA Viruses DNA biosynthesis Development Exposure to Family Figs - dietary Fright Gene Expression Genetic Genome Histones Human In Vitro Infection Investigation Lead Life Cycle Stages Link Lipid Bilayers Mediating Membrane Modeling Monkeypox virus Morphogenesis Oncolytic Organelles Pathway interactions Physical condensation Polyamines Poxviridae Process Protein Binding Proteins Pump Recombinant Vaccines Resolvase Role Smallpox Smallpox Viruses Specificity Spermidine Spermine Testing Vaccination Vaccinia virus Viral Viral Genome Viral Proteins Virion Virus Virus Diseases base genetic analysis human MCAM protein human morbidity human mortality in vivo insight member mutant pathogen telomere therapeutic target tool viral DNA weapons

展开

项目摘要

DESCRIPTION (provided by applicant): Poxviruses, such as vaccinia virus, are unique among DNA viruses in replicating solely within the cytoplasm of the infected cell. As a result, the progression of the viral life cycle is intimately linked to interactions with the organelles, cytoskeleton and cytoplasmic milieu of the host cell. Cytoplasmic replication of poxviruses is also associated with their nearly complete genetic autonomy from the host. Among the ~ 200 gene products encoded by the virus are those mediate gene expression, genome replication and maturation, and the complex process of virion assembly. At its essence, a productive viral infection requires that the viral genome be efficiently replicated and packaged into nascent, infectious virions. It is this process of genome encapsidation, which lies at the interface of replication and morphogenesis, that it is the focus of this exploratory application. Viral DNA synthesis, which is accomplished by a repertoire of virally encoded proteins, is followed by the processing of concatemeric intermediates into mature, monomeric genomes by a virally encoded resolvase. These monomeric genomes are then thought to be condensed and translocated into immature virions (IV) prior to the closure of the IV membrane. Immature virions are delimited by a lipid bilayer and surrounded by an external lattice composed of the viral D13 protein. The interior of the immature virion appears to contain an unstructured pool of proteins that will later form the internal core of the mature virion. Genetic analyses of conditionally lethal viral mutants have identified two viral proteins as being directly associated with genome encapsidation: I6 and A32. In vitro, the I6 protein binds with great specificity to DNA probes representing the telomeric hairpins of the viral genome. These ~50 bp telomeres, which are covalently closed due to the presence of a short hairpin loop, are highly A-T rich (>90%) and contain extrahelical bases (EHBs) on both DNA strands. The binding of I6 to telomeric probes is dependent upon the presence of the EHBs. The A32 protein, in contrast, is thought to belong to the FtsK-HerA family of pumping ATPases, as defined by the conservation of characteristic sequence motifs. We are hypothesizing that the genome is condensed by association with polyamines, that I6 marks mature genomes for encapsidation by binding to their telomeres, and that subsequent interactions between I6 and A32 lead to activation of A32's ATPase activity and translocation of the genome into the immature virion. This hypothesis will be tested in two Aims. Aim I will explore the condensation of the viral genome and its association with I6; Aim II will explore the role of A32 as a translocating ATPase. PUBLIC HEALTH RELEVANCE: The fear that smallpox might be used as a bioterrorist weapon, and the recognition that monkeypox virus is a cause of human morbidity and mortality, has reinforced the need to study the poxviral life cycle in depth. Our investigations of genome encapsidation will provide fundamental insights into this poorly understood facet of the viral life cycle and define new targets for the development of rational antiviral therapies.

描述（由申请人提供）：在DNA病毒中，痘病毒，例如疫苗病毒在仅在感染细胞的细胞质内复制时是独一无二的。结果，病毒生命周期的进展与宿主细胞的细胞器，细胞骨架和细胞质环境密切相关。痘病毒的细胞质复制也与宿主的几乎完全遗传自主性有关。在由病毒编码的200个基因产物中，有那些介导基因表达，基因组复制和成熟以及病毒体组装的复杂过程。从本质上讲，生产性病毒感染要求病毒基因组有效地复制并包装成新生的感染性病毒体。正是这种基因组封装的过程在于复制和形态发生的界面，是该探索性应用的重点。病毒DNA合成是由病毒编码蛋白的曲目完成的，随后是通过病毒编码的分解抗体将串联中间体加工成成熟的单体基因组。然后，这些单体基因组被认为是在静脉膜闭合之前凝结并易位到未成熟病毒体（IV）中。未成熟的病毒体是由脂质双层界定的，并被由病毒D13蛋白组成的外部晶格包围。未成熟的病毒体的内部似乎包含一个非结构化的蛋白质池，后来将形成成熟病毒体的内部核心。有条件致命的病毒突变体的遗传分析已确定两个病毒蛋白与基因组封装直接相关：I6和A32。在体外，I6蛋白与代表病毒基因组的端粒发夹的DNA探针具有很高的特异性结合。这些〜50 bp的端粒由于存在短发夹，因此在两个DNA链上都含有高度A-T富含发夹（> 90％），并且在两个DNA链上都包含外毛基碱（EHB）。 I6与端粒探针的结合取决于EHB的存在。相比之下，A32蛋白被认为属于泵送ATPases的FTSK-HERA家族，这是由特征序列基序的保护所定义的。我们假设该基因组是通过与多胺缔合来凝结的，i6标志着成熟的基因组通过与端粒结合结合，并且随后i6和A32之间的相互作用导致A32的ATPase活性激活，并将基因组转移到不成熟的病毒中。该假设将以两个目的进行检验。目的，我将探索病毒基因组及其与i6的关联的凝结； AIM II将探索A32作为转移ATPase的作用。公共卫生相关性：担心天花可能被用作生物恐怖主义武器，并且认识到蒙基托克病毒是人类发病率和死亡率的原因，这加剧了深入研究Poxviral生命周期的必要性。我们对基因组封装的研究将为病毒生命周期的这一知识所理解的方面提供基本见解，并为发展理性抗病毒疗法的发展定义了新目标。