EBV reactivation causes widespread host de novo promoter transcription and transcriptional interference

EBV 重新激活导致广泛的宿主从头启动子转录和转录干扰

• 批准号: 10647826
• 负责人: ERIK K FLEMINGTON
• 金额: $ 41.28万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10647826
• 关键词: AIDS population; AIDS related cancer; Architecture; Automobile Driving; Binding; Binding Sites; Biological Models; Cell Nucleus; Cells; Chromatin; Complex; Cues; Cytoplasm; DNA Polymerase II; DNA Tumor Viruses; Data; Dedications; Direct Lytic Factors; Down-Regulation; Environment; Epstein-Barr Virus-Related Lymphoma; Epstein-Barr Virus-Related Malignant Neoplasm; Fostering; Frequencies; Gene Expression; Genes; Genetic; Genetic Transcription; Genome; Genomics; HIV; HIV/AIDS; Herpesviridae; Human Herpesvirus 4; Human Herpesvirus 8; Incidence; Individual; Link; Lymphoma; Lymphomagenesis; Lytic; Mediating; Messenger RNA; Metabolism; Modeling; Mus; Nature; Pathway interactions; Population; Positioning Attribute; Predisposition; Process; Production; Proteins; RNA; RNA-Binding Proteins; Resources; Role; Site; Specific qualifier value; Stimulus; Structural Protein; Trans-Activators; Transcript; Transcription Initiation; Translations; Viral; Viral Packaging; Viral Proteins; Viral Structural Proteins; Virion; Virus; Virus Diseases; Virus Latency; Virus Replication; Virus-Cell Membrane Interaction; cell growth regulation; co-infection; gammaherpesvirus; high volume manufacturing; humanized mouse; insight; lytic replication; mRNA Expression; mRNA Transcript Degradation; mouse model; particle; programs; promoter; response; therapeutic target; transcription factor; transcriptome; virus host interaction

Summary The Epstein Barr virus (EBV) is a DNA tumor virus that causes an elevated incidence of lymphoma in the HIV/AIDS population. While more than 90% of the world's population carries EBV, the virus typically exists in a “latent” state with little impact on the host. In response to certain stimuli or local microenvironmental cues, however, EBV enters the lytic viral replication program, leading to viral spread both between and within hosts. In addition to the known role of viral latency proteins in EBV associated cancers, there are well-established links between lytic replication and EBV associated cancers; and elevated EBV lytic replication in HIV co- infection (+ or – ART) likely contributes to the increased susceptibility of HIV infected individuals to EBV associated lymphomas. With minimal genetic content, viruses are inexorably dependent on host cell resources for their replication and they evolve mechanisms to modulate host cell metabolic processes to facilitate efficient virus production. One of the most conserved virus-host interactions in herpesvirus replication is “host shut off” where virus encoded factors degrade host cell mRNAs, freeing up translation resources for dedicated production of viral structural proteins. Recently, the Glaunsinger lab showed that the murine γ-herpesvirus, MHV68 also inhibits Pol II loading on cell genes. This illustrates the utilization of layered host shut off mechanisms that together overcome the rate-limiting step of viral structural protein production. Using EBV reactivation models that facilitate assessment of transcriptome changes in pure reactivating cell populations, we gained new and unexpected insights into EBV's interactions with the host cell transcriptome. Among a number of observed transcriptome alterations is our finding that EBV causes transcription initiation at more than 25,000 cell genomic sites that have no previous evidence of promoter activity. These “de novo” promoters are unusually simple, being composed of only one or two short viral transcription factor binding motif(s) that due to their short nature, occur randomly at a high frequency across the cell genome. Our preliminary data suggests that one of these motifs is recognized by the viral preinitiation complex, vPIC, which binds a “TATA”-like motif (TATT(TA)AA) and has been shown in some cases to activate viral late genes without apparent support from other transcription factors. We hypothesize that EBV utilizes the ability of vPIC to single-handedly activate transcription through these simple motifs to spawn thousands of host de novo promoters and substantially alter the cell transcriptome and chromatin architecture. Our preliminary studies also show that the most highly active de novo promoters have enriched localization within 1.5 kb of canonical cell promoters, with transcriptional orientations toward the respective canonical promoters. We hypothesize that while EBV utilizes the primal nature of late viral promoter motifs to target thousands of sites throughout the cell genome, it leverages chromatin context near canonical promoters to drive high-level de novo promoter transcription, leading to targeted transcriptional interference (TI) of host promoters. This represents a new mechanistic concept for virus-host interactions that contributes to disrupted cell gene expression and dedicated production of viral structural proteins. In this proposal, we will 1) investigate the underlying mechanisms of de novo promoter specification and local chromatin context driving high level transcription near existing canonical promoters, and 2) investigate the impact of de novo transcription and transcriptional interference in downregulating cell mRNA expression.

总结 爱泼斯坦巴尔病毒（EBV）是一种DNA肿瘤病毒，其导致淋巴瘤的发病率升高。 艾滋病毒/艾滋病人口。虽然世界上超过90%的人口携带EBV，但该病毒通常存在于一个 “潜伏”状态，对宿主影响不大。对某些刺激或局部微环境线索的反应， 然而，EBV进入裂解性病毒复制程序，导致病毒在宿主之间和宿主内传播。 除了已知病毒潜伏蛋白在EBV相关癌症中的作用外， 裂解性复制和EBV相关癌症之间的联系;以及HIV共感染者中EBV裂解性复制的升高。 感染（+或- ART）可能导致HIV感染个体对EBV的易感性增加 相关淋巴瘤 由于具有最小的遗传内容，病毒不可阻挡地依赖于宿主细胞资源进行复制， 它们进化出调节宿主细胞代谢过程的机制以促进有效的病毒生产。一 疱疹病毒复制中最保守的病毒-宿主相互作用是“宿主关闭”， 因子降解宿主细胞mRNA，释放翻译资源用于专门生产病毒结构蛋白， proteins.最近，Glaunsinger实验室发现鼠γ-疱疹病毒MHV 68也抑制Pol II， 加载细胞基因。这说明了分层主机关闭机制的利用， 克服了病毒结构蛋白产生的限速步骤。 使用EBV再活化模型促进纯再活化细胞中转录组变化的评估 通过对人群的研究，我们获得了关于EBV与宿主细胞转录组相互作用的新的和意想不到的见解。 在许多观察到的转录组改变中，我们发现EBV在转录水平引起转录起始， 超过25，000个细胞基因组位点，以前没有证据表明启动子活性。这些“重新” 启动子非常简单，仅由一个或两个短的病毒转录因子结合组成， 基序，由于其短的性质，在整个细胞基因组中以高频率随机出现。我们 初步数据表明，这些基序之一被病毒前起始复合物vPIC识别， 结合“TATA”样基序（TATT（TA）AA），并在某些情况下显示激活病毒晚期基因 而没有其他转录因子的明显支持。我们假设EBV利用vPIC的能力 通过这些简单的基序单独激活转录， 启动子，并基本上改变细胞转录组和染色质结构。 我们的初步研究还表明，活性最高的从头启动子具有丰富的定位 在典型细胞启动子的1.5kb内，转录方向朝向相应的典型启动子。 发起人。我们假设，虽然EBV利用晚期病毒启动子基序的原始性质靶向 在整个细胞基因组的数千个位点中，它利用典型启动子附近的染色质背景， 驱动高水平的从头启动子转录，导致宿主的靶向转录干扰（TI） 发起人。这代表了病毒-宿主相互作用的一种新的机制概念， 细胞基因表达和专门生产病毒结构蛋白。 在这个提议中，我们将1）调查从头启动子特化和局部启动子特化的潜在机制， 在现有的典型启动子附近驱动高水平转录的染色质背景，以及2）研究 从头转录和转录干扰对下调细胞mRNA表达影响。