Identifying Endogenous Retroviral Factors in Viral Lymphomagenesis

项目总结/摘要 艾滋病是艾滋病毒感染者死亡的主要原因。在这些恶性肿瘤中， 非霍奇金淋巴瘤（NHL）是最常见的血液肿瘤影响PLWH。尽管 在引入高效抗逆转录病毒疗法（ART）后，PLWH发展成NHL的可能性是 没有艾滋病毒的人。爱泼斯坦巴尔病毒（EBV）是一种致癌病毒，与淋巴瘤最相关 在PLWH中常见。活动性艾滋病毒感染可导致获得性免疫缺陷综合症（艾滋病）， 未经治疗的人，众所周知，艾滋病促进EBV+淋巴瘤的发展，称为艾滋病- 定义癌症（ADC），宿主因素，包括内源性逆转录病毒对淋巴瘤发生的影响， 用ART治疗的免疫活性PLWH仍有待探索。在切萨曼和尼克松实验室的工作 显示EBV和HIV在感染后各自劫持宿主细胞信号传导途径以诱导 内源性逆转录病毒元件，包括人内源性逆转录病毒（HERV）。确定 这些病毒对淋巴瘤发生的贡献，我们将从体外淋巴瘤模型中产生数据，以测试 EBV和HIV蛋白对内源性逆转录病毒癌蛋白表达的影响。此外，我们将 对来自PLWH的弥漫性大B细胞淋巴瘤（DLBCL）进行批量和单细胞测序，以确定 肿瘤微环境（TME）中细胞群的变化。我们假设会有独特的 EBV感染的B细胞群体具有与异常调节相关的癌前转录组特征， HERV表达，包括HERVK（HML 2）Np 9癌基因的表达。 在目标1中，我们将确定EBV潜伏期III LMP 2A和细胞外HIV p17变体对细胞凋亡的影响。 Np 9癌基因的表达。我们将进行逐步的分子和细胞研究，以确定 外源性病毒感染对这种内源性逆转录病毒癌基因表达的翻译影响。我们 将另外进行多组学分析，以鉴定与转录相关的病毒和宿主因子， 病毒介导的Np 9癌基因表达机制。 在目标2中，我们将使用我们新建立的单细胞管道来确定基因座特异性内源性逆转录病毒， 来自PLWH的DLBCL样品的转录。我们在尼克松实验室的初步数据强烈表明 我们将能够使用我们已经开发的管道来识别和量化这些HERV转录本 RNA测序数据。我们将评估EBV+样品与EBV+样品的相对细胞群。 是EBV-和决定宿主基因，逆转录病毒和病毒转录的差异。此外，我们打算 用来自更大组群的EBV+/-新鲜冷冻组织的批量RNA测序验证这些单细胞发现 从PLWH的DLBCL。

PROJECT SUMMARY/ABSTRACT Malignancy is the leading cause of mortality amongst people living with HIV (PLWH). Of these malignancies, Non-Hodgkin lymphoma (NHL) is the most common hematological neoplasm affecting PLWH. Despite the introduction of highly active antiretroviral therapy (ART), PLWH are 10-20 times more likely to develop NHL than those without HIV. Epstein Barr Virus (EBV) is an oncogenic virus that is associated with the lymphomas most commonly observed in PLWH. Active HIV infection can lead to Acquired Immunodeficiency Syndrome (AIDS) in untreated people, and it is known that AIDS promotes the development of EBV+ lymphomas, termed AIDS- defining cancers (ADCs), the effect of host factors, including endogenous retroviruses on lymphomagenesis in immunocompetent PLWH treated with ART, remains to be explored. Work in the Cesarman and Nixon labs has shown that EBV and HIV each hijack host cell signaling pathways upon infection to induce the expression of endogenous retroelements, including as human endogenous retroviruses (HERV). To determine the contributions of these viruses on lymphomagenesis, we will generate data from in vitro lymphoma models to test the effect of EBV and HIV proteins on the expression of endogenous retroviral oncoproteins. Additionally, we will perform bulk and single-cell sequencing of Diffuse Large B Cell Lymphoma (DLBCL) from PLWH to determine the changes in cell populations of the tumor microenvironment (TME). We hypothesize that there will be unique populations of EBV infected B cells with pre-malignant transcriptomic signatures that correlate with dysregulated HERV expression, including expression of the HERVK(HML2) Np9 oncogene. In Aim 1, we will determine the impact of EBV latency III LMP2A and extracellular HIV p17 variants on the expression of the Np9 oncogene. We will perform stepwise molecular and cellular studies to determine the translational impact of exogenous viral infection on the expression of this endogenous retroviral oncogene. We will additionally perform multi-omic analysis to identify viral and host factors relevant to the transcriptional machinery of virus-mediated Np9 oncogene expression. In Aim 2, we will use our novel established single cell pipeline to determine locus-specific endogenous retroviral transcription of DLBCL samples from PLWH. Preliminary data from our work in the Nixon lab strongly suggests that we will be able to use the pipeline that we have developed to identify and quantify these HERV transcripts from RNA-sequencing data. We will assess the relative cell populations of samples that are EBV+ to those that are EBV- and determine the differences in host gene, retroviral and viral transcripts. Additionally, we intend to validate these single cell findings with bulk RNA-sequencing from a larger cohort of EBV+/- fresh frozen tissue of DLBCL from PLWH.