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（由申请人提供）：人类疱疹病毒 8 (HHV-8) 指定了四种病毒干扰素调节因子同源物 (vIRF)，它们代表一组新的病毒编码蛋白，迄今为止仅在 HHV-8 和密切相关的恒河猴病毒中发现。 HHV-8 vIRF 的功能是抑制细胞 IRF 活性和细胞防御途径的其他成分，这些成分可促进细胞周期停滞和细胞凋亡以响应病毒感染。其中两种 vIRF，vIRF-1 和 vIRF-3，已被证明对病毒的有效复制（在内皮细胞中）或潜伏感染的细胞活力（在原发性渗出性淋巴瘤 (PEL) 细胞中）有显着贡献；每个 vIRF 在潜伏 (PEL) 复制和裂解复制过程中均表达。病毒生命周期的两个阶段对于 HHV-8 恶性发病机制都很重要：卡波西肉瘤、PEL 和多中心卡斯尔曼病。 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 转录共激活因子、IFN/视黄酸诱导的 GRIM19、p53、p53 激活 ATM 激酶以及 Bcl-2 家族的某些仅 BH3（促凋亡）成员； vIRF-3 的目标是 IRF、p300/CBP 和 p53。然而，这些多效性蛋白质的抑制相互作用的全部内容尚不清楚，并且在潜伏和裂解复制的背景下尚未充分研究相互作用。虽然我们已经确定 vIRF-1 与 BH3-only 蛋白的相互作用对于内皮细胞中成功的生产性复制至关重要，但其他 vIRF-1 相互作用在这些细胞和其他细胞的裂解复制中的作用以及 vIRF-1 及其相互作用对 PEL 潜伏期的贡献仍然未知。同样，vIRF-3 及其在生产性复制中的相互作用以及特定 vIRF-3 在潜伏期中的相互作用的作用仍有待确定。为了解决这些问题，我们建议：（1）对潜伏感染的 PEL 和裂解感染的 PEL 以及内皮细胞中的 vIRF 1 和 3 进行亲和沉淀，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 vIRF 1 和 3、它们的细胞靶标以及特定 vIRF-蛋白质相互作用对潜伏感染细胞活力和高效复制的贡献 并开发抑制剂。该项目包括对独特的病毒先天免疫逃避蛋白进行广泛但在概念和实验上集中的分析，其对病毒生物学的功能和机制贡献目前尚不清楚。由于这些蛋白质对于潜伏期和生产性复制都至关重要，因此本研究产生的信息可以为开发新型抗病毒和临床疗法提供基础。