Regulation of Nuclear Signaling Pathways by the Adenovirus E4-ORF3 Protein

描述(由申请人提供)：具有线性dsDNA基因组的病毒，如腺病毒(Ad)和疱疹病毒，会遇到许多宿主细胞反应，可能严重抑制病毒复制。这项建议的重点是腺病毒E4-ORF3蛋白，它通过对抗细胞抗病毒反应来促进病毒复制。近年来，腺病毒已被认为是免疫功能低下患者的重要病原体。目前尚无针对Ad感染的病毒特异性治疗方法。因此，为了促进抗病毒治疗的发展，充分了解宿主对Ad感染的反应以及用来抑制这些反应的病毒策略变得越来越重要。AdE4-ORF3蛋白通过将多种细胞蛋白重新定位到称为Track的核结构中来抑制它们的活性。E4-ORF3在感染期间抑制DNA损伤反应，并抵消干扰素(干扰素)反应。E4-ORF3蛋白诱导两种细胞蛋白Nbs1和Mre11的总合作用，这两种蛋白是细胞DNA损伤反应中至关重要的效应因子。E4-ORF3还与相扑结合，并使用相扑结合基序将细胞蛋白重新定位到核轨道上。这些结果将E4-ORF3放在细胞相加系统的结点上。相扑蛋白翻译后修饰调控多种细胞过程，包括转录、DNA复制、DNA修复、亚细胞定位和泛素化。E4-ORF3诱导的细胞蛋白总甲基化可能被用来抑制宿主的抗病毒活性。有人建议研究E4-ORF3诱导Nbs1和Mre11相加作用的机制，这些修饰的功能后果，并鉴定和研究其他由E4-ORF3诱导相加作用的细胞蛋白。E4-ORF3蛋白与相扑的相互作用可能代表了E4-ORF3招募将细胞蛋白相加到核轨道上的基础，并提出了实验来验证这一想法。E4-ORF3蛋白通过隔离两种主要的抗病毒效应蛋白PML和Daxx来抑制Ad感染期间的干扰素反应。在干扰素诱导的抗病毒状态下，PML和Daxx抑制Ad复制的机制(S)将被研究。E4-ORF3的另外两个靶标，转录抑制因子TIF1？和TIF1？，在干扰素反应中的作用将被确定。E4-ORF3蛋白将大量与各种功能相关的不同细胞蛋白招募到核轨道上。 这种小病毒蛋白是如何做到这一点的尚不清楚。将对野生型和突变型E4-ORF3蛋白进行物理分析，目的是研究E4-ORF3蛋白的结构及其与功能的关系。总的来说，这些研究将为干扰病毒复制的细胞机制和抵消这些影响的病毒反应提供独特的见解。

DESCRIPTION (provided by applicant): Viruses with linear, dsDNA genomes, such as the adenoviruses (Ad) and herpesviruses, encounter a number of host cell responses that may severely inhibit virus replication. This proposal focuses on the adenovirus E4-ORF3 protein which promotes virus replication by counteracting cellular antiviral responses. Adenoviruses have been recognized in recent years as significant pathogens in immunocompromised patients. There is no virus-specific therapy for Ad infection. It has become increasingly important, therefore, to fully understand host responses to Ad infection and viral strategies used to inhibit these responses in order to foster the development of antiviral therapies. The Ad E4-ORF3 protein functions by relocalizing a variety of cellular proteins into nuclear structures, referred to as tracks, to inhibit their activities. E4-ORF3 inhibits a DNA damage response and counteracts an interferon (IFN) response during infection. The E4-ORF3 protein induces the sumoylation of two cellular proteins, Nbs1 and Mre11, which are critically important effectors in a cellular DNA damage response. E4-ORF3 also binds SUMO and uses SUMO binding motifs to relocalize cellular proteins into nuclear tracks. These results place E4-ORF3 at the nexus of the cellular sumoylation system. Post-translational protein modification by SUMO regulates diverse cellular processes including transcription, DNA replication, DNA repair, subcellular localization, and ubiquitination. The induction of cellular protein sumoylation by E4-ORF3 likely is used to inhibit host antiviral activities. Studies are proposed to investigate the mechanism by which E4-ORF3 induces Nbs1 and Mre11 sumoylation, the functional consequences of these modifications, and identify and investigate other cellular proteins whose sumoylation is induced by E4-ORF3. The interaction of the E4-ORF3 protein with SUMO may represent the foundation by which E4-ORF3 recruits sumoylated cellular proteins into nuclear tracks and experiments are proposed to test this idea. The E4-ORF3 protein inhibits an IFN response during Ad infection by sequestering the proteins PML and Daxx, two prominent antiviral effectors. The mechanism(s) by which PML and Daxx inhibit Ad replication during the IFN-induced antiviral state will be investigated. The contribution of two other targets of E4-ORF3, transcriptional repressors TIF1¿ and TIF1¿, during an IFN response will be determined. The E4- ORF3 protein recruits a large number of different cellular proteins, associated with a variety of functions, into nuclear tracks. How this small viral protein accomplishes this is unknown. Physical analyses of wild type and mutant E4-ORF3 proteins will be conducted with the goal to study E4-ORF3 protein structure as it relates to function. Collectively, these studies will provide unique insight into cellular mechanisms that interfere with virus replication and viral responses that counteract these effects.