Epstein-Barr Virus nuclear antigen leader protein in transcription regulation

Epstein-Barr病毒核抗原前导蛋白在转录调控中的作用

• 批准号: 10331840
• 负责人: Chong Wang
• 金额: $ 8.69万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10331840
• 关键词: AIDS-Related Lymphoma; Address; Affect; B Cell Proliferation; B lymphoid malignancy; B-Lymphocytes; Binding Proteins; Biological Assay; Biological Models; CRISPR interference; CRISPR screen; Cells; Chromatin; Clustered Regularly Interspaced Short Palindromic Repeats; DNA; DNA-Binding Proteins; Data; Development; Disease; EBV-associated disease; EP300 gene; Elements; Enhancers; Enzymes; Epithelial Cells; Epstein-Barr Virus Infections; Epstein-Barr Virus Nuclear Antigens; Epstein-Barr Virus latency; Epstein-Barr pathogenesis; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Growth; HIV; Hairy Leukoplakia; Health; Histones; Human; Human Herpesvirus 4; Immune; Immune system; Immunoprecipitation; In Vitro; Infectious Mononucleosis; Knock-out; LMP1; Lead; Lentivirus; Libraries; Link; Lymphoma; Lymphoproliferative Disorders; Malignant Neoplasms; Mediating; Membrane Proteins; Mentors; Molecular; Monitor; Mucous Membrane; Mutation; Nasopharynx Carcinoma; Nuclear Antigens; Oral; Oral cavity; Patients; Persons; Phase; Primary Infection; Proliferating; Proteins; Reporter; Research; Rest; Role; Saliva; Signal Transduction; Simian virus 40; Site; Sorting - Cell Movement; Testing; Tongue; Transcription Coactivator; Transcriptional Regulation; Transfection; Viral Proteins; Work; base; chromatin protein; cofactor; deep sequencing; genome-wide; lymphoblastoid cell line; malignant stomach neoplasm; novel therapeutics; oral cavity epithelium; pathogen; post-transplant; programs; promoter; protein activation; protein complex; stable cell line; transcription factor; transforming virus; vector; virus related cancer

Abstract: Epstein-Barr virus (EBV) associated diseases remain a huge burden in human health. As an orally transmitted pathogen, EBV infection causes infectious mononucleosis and ~200,000 cases of various cancers, including nasopharyngeal carcinoma that occurs in a space immediately adjacent to the oral cavity, some B cell malignancies, and ~10% of gastric cancer. In HIV infected people, EBV causes oral hairy leukoplakia of tongue. To understand the molecular mechanisms through which EBV contributes to disease development, EBV-transformed lymphoblastoid cell lines are used as a model system. EBV nuclear antigen leader protein (EBNALP) is essential for EBV to transform naïve B lymphocytes. Most of its known functions are linked to EBV transcription activator EBNA2. However, EBNALP binds to many enhancer and promoter sites independent of EBNA2. Perturbations of these EBNALP sites with CRISPRi significantly decreased these enhancers’ linked gene expression. Little is known about how EBNALP exerts its EBNA2 independent functions. It is also not known how EBNALP is tethered to the enhancer/promoter sites and how they affect transcription. Therefore, we hypothesize that EBNALP exploits host transcription programs to gain access to host enhancers/promoters, and contributes to EBV transformation through EBNA2-independent mechanisms. During my mentored period, I will address the fundamental question of how EBNALP binds to DNA. We will use CRISPR-based assays to identify host proteins essential for EBNALP enhancer activation. I will first focus on sequence-specific transcription factors (TFs). Chromatin immune precipitation (ChIP) based assays will be used to test the effects of knockout on EBNALP DNA binding. During my R00 phase, I will perform research independently and distinguish my work from my mentor’s by studying different aspects of EBNALP. I will focus my studies on characterizing the enhancer protein complexes assembled by EBNALP onto the enhancers to regulate transcription activity. I will focus on transcription cofactors, basal transcription factors, and histone modifying enzymes. Understanding the mechanisms through which EBNALP binds to DNA and regulates gene transcription may provide promising targets for treating EBV-associated diseases.

摘要： EB病毒（EBV）相关疾病仍然是人类健康的巨大负担。作为 经口传播的病原体，EBV感染引起传染性单核细胞增多症， 各种癌症的病例，包括发生在空间中的鼻咽癌 紧邻口腔，一些B细胞恶性肿瘤和~10%的胃癌。 在HIV感染者中，EBV引起舌的口腔毛状白斑。了解 EBV促进疾病发展的分子机制， EBV转化的类淋巴母细胞系用作模型系统。EBV核抗原 前导蛋白（EBNALP）是EBV转化幼稚B淋巴细胞所必需。其大部分 已知的功能与EBV转录激活因子EBNA 2相关。然而，EBNALP结合到 许多独立于EBNA 2的增强子和启动子位点。这些EBNALP的扰动 具有CRISPRi的位点显著降低了这些增强子关联的基因表达。之甚少 已知EBNALP如何发挥其EBNA 2独立功能。也不知道是如何 EBNALP与增强子/启动子位点以及它们如何影响转录有关。 因此，我们假设EBNALP利用宿主转录程序获得 进入宿主增强子/启动子，并通过 EBNA 2独立机制。在我的指导期间，我将解决 EBNALP如何与DNA结合的基本问题。我们将使用基于CRISPR的检测方法， 鉴定EBNALP增强子激活所必需宿主蛋白。我将首先关注 序列特异性转录因子（TF）。基于染色质免疫沉淀（ChIP） 测定将用于测试敲除对EBNALP DNA结合的影响。在R 00期间 在这一阶段，我将独立进行研究，并通过以下方式将我的工作与导师的工作区分开来： 研究EBNALP的不同方面。我会集中精力研究 由EBNALP组装到增强子上的蛋白质复合物以调节转录 活动我将集中在转录辅因子，基础转录因子和组蛋白 修饰酶了解EBNALP与DNA结合的机制 并调节基因转录可能为治疗EBV相关的 疾病