Parvovirus-Cell Interactions

细小病毒-细胞相互作用

• 批准号: 8288689
• 负责人: DAVID J. PINTEL
• 金额: $ 29.94万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8288689
• 关键词: Adenoviruses; Animal Viruses; Antiviral Response; Area; Binding; Capsid; Cell Communication; Code; Complex; DNA; DNA Viruses; DNA biosynthesis; Dependovirus; Disease; Double-Stranded RNA; Elements; Epidermal Growth Factor Receptor; Genome; Helper Viruses; Infection; Knowledge; Life Cycle Stages; Ligase; Light; Messenger RNA; Parvovirus; Play; Production; Protein C; Proteins; RNA; Recombinant adeno-associated virus (rAAV); Role; Symbiosis; Translations; Ubiquitination; Viral; Viral Proteins; Virus; Virus Diseases; Virus Latency; Virus Replication; adeno-associated viral vector; gene therapy; improved; inhibitor/antagonist; lytic replication; mutant; novel; protein kinase R; ubiquitin-protein ligase; viral DNA

DESCRIPTION (provided by applicant): Adeno-associated viruses (AAVs) require helper-virus functions from larger DNA viruses for efficient replication. Although the five adenovirus (Ad) helper functions necessary for AAV replication were identified years ago, details of their role in AAV infection are incomplete. Our recent analysis revealed a surprising complexity in the roles that the Ad helper functions play during AAV replication. Ad E4Orf6 was known to function at an early rate-limiting step in AAV second-strand synthesis. Our closer examination revealed that AAV hijacks the Ad E4Orf6/E1B 55kDa E3 ubiquitin ligase complex for its use; it efficiently targets both an enabler of an inhibitor of AAV replication as well as, surprisingly, AAV proteins themselves. Our finding that Ad VA RNA mutants that could no longer inactivate the dsRNA- activated protein kinase R (PKR) could not support AAV replication led to our discovery of an element within the AAV capsid coding mRNA that binds and activates PKR. This was the first PKR-activating element described within an mRNA generated by a DNA virus. Further, our studies demonstrated that AAV engages the host antiviral response, which had not previously been shown for any parvovirus. This application builds on these observations. Our specific objectives are to understand how the Ad E3 Ub-ligase (Specific Aim I) and Ad VA RNA (Specific Aim II) function during AAV/Ad co-infection. However, in addition to helping solve the long standing issue of how helper-functions facilitate AAV replication, our studies will help explain how sharing a viral E3 ubiquitin-ligase can help optimize the symbiosis of two co-infecting DNA viruses, and how a novel PKR- activating element within an mRNA of a DNA virus might function, engage the host antiviral response, and perhaps govern a switch from viral latency in the absence of helper to expression and lytic replication in its presence. Studies in this last area should also shed light on basic differences between the autonomous and helper-requiring groups of parvoviruses, and also help improve recombinant AAV vector production.

描述（由申请人提供）：腺相关病毒（AAV）需要较大 DNA 病毒的辅助病毒功能才能有效复制。尽管 AAV 复制所需的五种腺病毒 (Ad) 辅助功能在多年前就已被确定，但它们在 AAV 感染中的作用细节尚不完整。我们最近的分析揭示了 Ad 辅助函数在 AAV 复制过程中所扮演的角色令人惊讶的复杂性。 Ad E4Orf6 已知在 AAV 第二链合成的早期限速步骤中发挥作用。我们的仔细检查发现，AAV 劫持了 Ad E4Orf6/E1B 55kDa E3 泛素连接酶复合物以供其使用；它有效地靶向 AAV 复制抑制剂的促进剂，以及令人惊讶的 AAV 蛋白本身。我们发现 Ad VA RNA 突变体无法再使 dsRNA 激活的蛋白激酶 R (PKR) 失活，无法支持 AAV 复制，这导致我们在 AAV 衣壳编码 mRNA 中发现了一个结合并激活 PKR 的元件。这是 DNA 病毒产生的 mRNA 中描述的第一个 PKR 激活元件。此外，我们的研究表明，AAV 会参与宿主的抗病毒反应，此前从未在任何细小病毒中证实过这一点。该应用程序建立在这些观察的基础上。 我们的具体目标是了解 Ad E3 Ub 连接酶（Specific Aim I）和 Ad VA RNA（Specific Aim II）在 AAV/Ad 共感染期间如何发挥作用。然而，除了帮助解决辅助功能如何促进 AAV 复制这一长期存在的问题之外，我们的研究还将有助于解释共享病毒 E3 泛素连接酶如何帮助优化两种共感染 DNA 病毒的共生，以及一种新型病毒如何帮助 AAV 复制。 DNA病毒mRNA内的PKR激活元件可能发挥作用，参与宿主的抗病毒反应，并可能控制从没有辅助因子的情况下的病毒潜伏期到辅助因子存在时的表达和裂解复制的转变。最后一个领域的研究还应阐明细小病毒自主群体和辅助病毒群体之间的基本差异，并有助于改善重组 AAV 载体的生产。