HHV-8 vIRF interactions in the context of infection

HHV-8 vIRF 在感染情况下的相互作用

• 批准号: 9085244
• 负责人: John Nicholas
• 金额: $ 21.14万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9085244
• 关键词: Address; Affinity; Antiviral Agents; Apoptosis; Apoptotic; BCL2 gene; Binding; Biological; Biology; Cell Cycle Arrest; Cell Survival; Cells; Clinical; Cytostatics; Data; Development; EP300 gene; Endothelial Cells; Evaluation; Family; Genes; Genetic Transcription; Goals; Health; Homologous Gene; Human Herpesvirus 8; Immune; Immune response; Individual; Infection; Interferons; Kaposi Sarcoma; Knowledge; Latent Virus; Life Cycle Stages; Lymphoma; Macaca mulatta; Malignant - descriptor; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Multicentric Angiofollicular Lymphoid Hyperplasia; Natural Immunity; Pathogenesis; Pathway interactions; Peptides; Phase; Precipitation; Production; Proteins; Reagent; Reporting; Research; Rhadinovirus; Role; Specific qualifier value; Stress; Surveys; TP53 gene; Therapeutic; Therapeutic Effect; Time; Transcription Coactivator; Tretinoin; Viral; Viral Proteins; Virus; Virus Diseases; Virus Latency; Work; ataxia telangiectasia mutated protein; base; cellular targeting; in vivo; inhibitor/antagonist; lytic replication; member; novel; overexpression; prevent; primary effusion lymphoma; response; viral interferon regulatory factor; viral interferon regulatory factor-1

 DESCRIPTION (provided by applicant): Human herpesvirus 8 (HHV-8) specifies four viral interferon regulatory factor homologues (vIRFs) that represent a novel set of virus-encoded proteins, so-far identified only in HHV-8 and closely related rhesus rhadinovirus. The HHV-8 vIRFs function to inhibit cellular IRF activities and other components of cellular defense pathways that promote cell cycle arrest and apoptosis in response to virus infection. Two of the vIRFs, vIRF-1 and vIRF-3, have been demonstrated to contribute significantly to either virus productive replication (in endothelial cells) or to latently infected cell viability [in primary efusion lymphoma (PEL) cells]; each vIRF is expressed during both latent (PEL) and lytic replication. Both phases of the virus life cycle are important for HHV-8 malignant pathogenesis: Kaposi's sarcoma, PEL, and multicentric Castleman's disease. Known targeted cellular proteins of vIRF-1 include IRFs, p300/CBP transcriptional co-activators required for IRF-mediated responses, IFN/retinoic acid-induced GRIM19, p53, p53-activating ATM kinase, and certain BH3-only (pro-apoptotic) members of the Bcl-2 family; vIRF-3 targets IRFs, p300/CBP, and p53. However, the full repertoire of inhibitory interactions of these pleiotropic proteins is far from clear and the interactions have not been adequately surveyed in the context of latent and lytic replication. Whilst we have determined that vIRF-1 interactions with BH3-only proteins are critical for successful productive replication in endothelial cells, the roles of other vIRF-1 interactions in lytic replication in these and other cells and the contributions of vIRF-1 and its interactions to PEL latency remain unknown. Similarly, the roles of vIRF-3 and its interactions in productive replication and of specific vIRF-3 interactions in latency remain to be determined. To address these issues, we propose to: (1) undertake affinity precipitations of vIRFs 1 and 3 from latently infected PEL and lytically infected PEL and endothelial cells followed by mass spectrometry analysis of co-precipitated cellular proteins; (2) use suppression, overexpression, and binding-competition analyses to identify the contributions of vIRFs 1 and 3, their cellular targets, and particular vIRF-protein interactions to latently infected cell viability and productive replication and to develop inhibitory agents. The project comprises a broadly-visioned but conceptually and experimentally focused analysis of unique viral innate immune evasion proteins whose functional and mechanistic contributions to virus biology are presently unclear. As these proteins are centrally important to both latency and productive replication, information generated from this study could provide the basis for development of novel antiviral and clinical therapies.

 描述（由适用提供）：人疱疹病毒8（HHV-8）指定了四个病毒干扰素调节因子同源物（VIRFS），代表了一组新型病毒编码的蛋白质，仅在HHV-8中鉴定出来，并密切相关的恒河类病毒。 HHV-8 VIRF的功能可抑制细胞IRF活性和细胞防御途径的其他成分，这些途径促进了细胞周期停滞和凋亡，以响应病毒感染。已证明其中两个VIRFS VIRF-1和VIRF-3可以显着促进病毒生产性复制（在内皮细胞中）或潜在感染的细胞活力[在一级效率淋巴瘤（PEL）细胞中]；每个VIRF在潜在（PEL）和裂解复制期间都表达。病毒生命周期的两个阶段对于HHV-8恶性发病机理都很重要：Kaposi的肉瘤，PEL和多中心Castleman病。 VIRF-1的已知靶向细胞蛋白包括IRF，IRF介导的反应所需的P300/CBP转录共激活剂，IFN/视黄酸诱导的GRIM19，p53，p53激活ATM激酶以及某些BH3的（pro-Pro-Propoptotic）成员的p53激活ATM激酶VIRF-3目标IRF，P300/CBP和P53。但是，这些多效蛋白的抑制作用的全部曲目远非清晰，并且在潜在和裂解复制的背景下没有得到充分的调查。尽管我们已经确定与仅BH3蛋白的VIRF-1相互作用对于在内皮细胞中成功的产物复制至关重要，但其他VIRF-1相互作用在这些和其他细胞中的裂解复制中的作用以及VIRF-1及其对PEL潜伏期的相互作用的作用尚不清楚。同样，VIRF-3的作用及其在延迟中的生产性复制和特定的VIRF-3相互作用中的作用尚待确定。为了解决这些问题，我们提出：（1）从潜在感染的PEL和裂解感染的pel和内皮细胞中进行VIRFS 1和3的亲和力沉淀，然后对共沉淀的细胞蛋白进行质谱分析； （2）使用抑制，过表达和结合竞争分析来确定VIRFS 1和3的贡献，其细胞靶标的以及特定的VIRF蛋白相互作用对潜在感染的细胞活力和产物复制 并发展抑制剂。该项目包括对独特的病毒先天免疫抗性蛋白的广泛审视，但在概念和实验方面的分析，其功能和机械贡献对病毒生物学的功能和机械贡献尚不清楚。由于这些蛋白质对潜伏期和生产性复制都非常重要，因此本研究产生的信息可以为开发新型的抗病毒毒和临床疗法提供基础。