Metabolic regulation of Epstein-Barr Virus-infected tonsillar B cells by EBNA-LP

EBNA-LP 对 Epstein-Barr 病毒感染的扁桃体 B 细胞的代谢调节

• 批准号: 10581497
• 负责人: Jana Cable
• 金额: $ 4.09万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-03-01 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10581497
• 关键词: Adult; Alanine; Antigens; B-Lymphocytes; Binding; Binding Proteins; Biological Assay; Cell Maturation; Cell Proliferation; Cell Survival; Cells; Cellular Metabolic Process; Chromatin; Chromatin Remodeling Factor; Co-Immunoprecipitations; Collagen; Complex; Data; Disease; EP300 gene; Electron Microscopy; Enzymes; Epstein-Barr Virus Infections; Epstein-Barr Virus Nuclear Antigens; Epstein-Barr Virus latency; Epstein-Barr Virus-Related Malignant Neoplasm; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Genus Hippocampus; Glucose; Goals; Growth; Herpesviridae; Higher Order Chromatin Structure; Human Herpesvirus 4; Hydroxyproline; Immune; Immune response; Immunofluorescence Immunologic; In Vitro; Individual; Infection; Infectious Mononucleosis; Leucine; Lymphoid Tissue; Lymphoma; Mediating; Membrane; Memory; Memory B-Lymphocyte; Metabolic; Metabolic Activation; Metabolism; Modeling; Modification; Molecular; Mutation; Nuclear; Nuclear Protein; Oral; Oral cavity; Organelles; Outcome; Oxidative Phosphorylation; Oxygen Consumption; Patients; Peptides; Phase; Post-Translational Protein Processing; Procollagen-Proline Dioxygenase; Proline; Protein Family; Proteins; Regulation; Role; Saliva; Site; Structure of germinal center of lymph node; T-Lymphocyte; Tonsil; Transcription Coactivator; Up-Regulation; Viral Proteins; Virus; Virus Diseases; YY1 Transcription Factor; cell growth; cell growth regulation; insight; latent infection; lymphoblastoid cell line; mutant; new therapeutic target; novel; oral cavity epithelium; promoter; protein oligomer; recruit; stable cell line; therapeutic target; transcription factor; transcriptome sequencing; transmission process; tumorigenesis

Epstein-Barr Virus (EBV) infects over 90% of adults worldwide and initially establishes infection in the oral cavity, including tonsillar B cells. Upon infecting naïve B cells, EBV expresses viral proteins including EBV Nuclear Antigens (EBNAs), transcriptional co-activators that promote B cell maturation and establish latently infected memory B cell reservoirs. EBV-associated malignancies include infectious mononucleosis, and, in immune compromised hosts, tumorigenesis including lymphomas. Recently, it has been appreciated that EBV induces metabolic changes upon infection. Our lab has discovered that EBV upregulates oxidative phosphorylation (OXPHOS) in order to promote cell proliferation and avoid arrest. Similarly, naïve B cells also require increased OXPHOS upon activation by antigen in order to undergo germinal center remodeling and produce memory B cells. Therefore, the ultimate goal of this proposal is to elucidate the molecular mechanisms by which EBV alters OXPHOS, which will enhance our understanding of EBV requirements for latency, and the role of metabolism in B cell maturation. Intriguingly, the viral protein EBNA-Leader Protein (EBNA-LP) is required for infection of naïve B cells, but not memory B cells – although the essential role of EBNA-LP is not well characterized. Our preliminary data suggests the viral protein EBNA-Leader Protein (EBNA-LP) may be essential in upregulation OXPHOS through transcriptional co-activation of metabolic genes. Our data suggests that EBNA-LP binds transcription factors that regulate expression of OXPHOS-related genes including NRF1, ERRα, and YY1 and then recruits chromatin remodeling factors such as P300. This mechanism of transcriptional co-activation mimics the PGC family of proteins, which uses leucine-rich motifs to bind the same OXPHOS transcription factors and recruit remodelers. Our preliminary data supports this model in that EBNA-LP contains multiple leucine-rich motifs, which may be required for binding these transcription factors. Additionally, EBNA-LP forms nuclear bodies, or membraneless organelles, that may be essential for EBNA-LP to regulate transcription, including OXPHOS genes. Our preliminary data suggests that EBNA-LP contains a post-translational modification, hydroxyproline, which promotes protein oligomerization and higher order structures in modified substrates such as collagen. Therefore, I propose that hydroxyprolination of EBNA-LP is required to form EBNA-LP nuclear bodies and co-activate transcription. My overall hypothesis is that EBNA-LP induces transcription of OXPHOS genes by using leucine-rich motifs to mimic the cellular PGC family of proteins, and through forming nuclear bodies upon hydroxyprolination. In Aim 1, I will assess the role of leucine-rich motifs in mediating transcription of OXPHOS genes and upregulation of cellular OXPHOS in tonsillar B cells. In Aim 2, I will determine the role of hydroxyprolination of EBNA-LP in formation of nuclear bodies and metabolic regulation in infected cells. These studies will elucidate novel mechanisms by which a viral protein regulates transcription and B cell metabolism and will provide insight towards understanding EBV-related malignancies.

Epstein-Barr病毒（EBV）全球超过90％的成年人感染，最初在口腔中建立感染， 包括扁桃体B细胞。感染B细胞后，EBV表达包括EBV核的病毒蛋白 抗原（EBNA），转录共激活剂，促进B细胞成熟并建立了潜在感染 内存B单元库。与EBV相关的恶性肿瘤包括传染性单核细胞增多症，并在免疫中 受损的宿主，包括淋巴瘤在内的肿瘤发生。最近，人们对EBV的影响很感激 感染后代谢变化。我们的实验室发现EBV上调氧化物磷酸化 （Oxphos）为了促进细胞增殖并避免停滞。同样，幼稚的B细胞也需要增加 抗原激活后的oxphos，以进行生发中心重塑并产生记忆b 细胞。因此，该提案的最终目标是阐明EBV改变的分子机制 Oxphos，这将增强我们对EBV对延迟的要求的理解，以及代谢在 B细胞成熟。有趣的是，病毒蛋白EBNA-LEADER蛋白（EBNA-LP）是幼稚的 B细胞，但不是记忆B细胞 - 尽管EBNA-LP的基本作用没有很好地表征。我们的 初步数据表明，病毒蛋白EBNA-LEADER蛋白（EBNA-LP）在上调可能是必不可少的 通过代谢基因的转录共激活Oxphos。我们的数据表明EBNA-LP结合 调节OXPHOS相关基因表达的转录因子，包括NRF1，ERRα和YY1和 然后募集染色质重塑因子，例如p300。转录共激活模拟的这种机制 PGC的蛋白质家族，它使用富含亮氨酸的基序来结合相同的Oxphos转录因子和 招募远程。我们的初步数据支持该模型，因为EBNA-LP包含多个亮氨酸丰富的 基序，这可能是结合这些转录因子所必需的。此外，EBNA-LP形成核体， 或无膜细胞器，这对于EBNA-LP调节转录可能至关重要，包括Oxphos 基因。我们的初步数据表明，EBNA-LP包含翻译后修饰，羟基， 促进蛋白质的低聚和高阶结构（例如胶原蛋白）中的高阶结构。 因此，我建议需要EBNA-LP的羟基化来形成EBNA-LP核体并共同激活转录。我的总体假设是EBNA-LP通过使用诱导OXPHOS基因的转录 富含亮氨酸的基序以模仿细胞PGC蛋白质家族，并通过形成核体 羟基化。在AIM 1中，我将评估富含亮氨酸基序在介导OXPHOS转录中的作用 扁桃体B细胞中细胞Oxphos的基因和上调。在AIM 2中，我将确定 EBNA-LP在受感染细胞中核体和代谢调节的形成中的羟基化。这些 研究将阐明病毒蛋白调节转录和B细胞代谢的新型机制 并将为了解与EBV相关的恶性肿瘤提供见识。