Epitranscriptomic regulation of HBV gene expression

HBV基因表达的表观转录组调控

• 批准号: 10569036
• 负责人: ALEEM SIDDIQUI
• 金额: $ 39.5万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-03-01 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10569036
• 关键词: 3' Untranslated Regions; 5' Untranslated Regions; Adenosine; Affinity; Antiviral Agents; Applications Grants; Binding; Bioinformatics; Biological Assay; Biological Process; Capsid; Cell Nucleus; Cells; Chronic; Chronic Hepatitis; Chronic Hepatitis B; Circular DNA; Code; Core Protein; Cytoplasm; DNA; DNA biosynthesis; Data; Development; Enzymes; Gene Expression; Genetic Transcription; Genome; Genomic DNA; Health; Hepatitis B Virus; Homeostasis; Human; Immune response; Immunoprecipitation; Infection; Inflammation; Length; Life Cycle Stages; Liver; Liver diseases; Maps; Messenger RNA; Methods; Methylation; Methyltransferase; Modification; Monitor; Mutate; Mutation; Nucleosome Core Particle; Nucleotides; Obesity; Pathogenesis; Patients; Play; Poly(A)+ RNA; Positioning Attribute; Primary carcinoma of the liver cells; Process; Proteins; RNA; RNA Viruses; RNA methylation; Regulation; Reporting; Research; Resolution; Reverse Transcription; Risk Factors; Role; Scheme; Site; Stress; Structure; System; Techniques; Terminator Codon; Tissues; Transcript; Translations; Vaccines; Viral; Viral Genome; Viral Proteins; Virus; Virus Diseases; Virus Replication; Withdrawal; base; design; epitranscriptomics; liver biopsy; posttranscriptional; therapeutically effective; transcriptome; viral DNA; viral RNA; virus identification

Project Summary/Abstract Human hepatitis B virus (HBV causes chronic hepatitis. Chronic hepatitis B is a major risk factor in the progression to the onset of hepatocellular carcinoma (HCC). In spite of an effective vaccine and currently used antivirals, there are estimated 350 million infected carriers worldwide. Antivirals do not eliminate HBV DNA in the nucleus. A better understanding of the role of host and viral factors contributing to chronic hepatitis is needed. Epitranscriptomic regulation of HBV gene expression provides another layer of regulatory schemes of gene expression. In this study, we propose to characterize the posttranscriptional modification of viral RNAs at the N6 position of the adenosine base termed m6A. Our extensive preliminary studies described in the grant application show that HBV RNAs are methylated as m6A. Using the strategy of silencing of host methylases and demethylase enzymes involved in epitranscriptomic homeostasis, we present evidence that HBV RNAs are m6A modified. There was clearly an effect on translation of viral transcripts in the absence of enzyme that catalyze m6A modification. Our results with MeRIP immunoprecipitation followed by Tru-seq analysis identified a potential range of sequences enriched in a peak of m6A residues. We mutated about 13 `A' residues in this region that spans from nucleotides 1700 to 2000 within the HBV genome. These were examined for translation of HBV proteins and reverse transcription function. Results were striking, in that, the translation was negatively regulated but reverse transcription was positively regulated by m6A modification. Based on these exciting data, we propose to continue characterization of m6A modification of HBV RNAs and identify the functional relevance of this modification in the HBV infection. This study opens a new avenue of HBV research to investigate epitranscriptomic regulation of the unique HBV life cycle and possible contribution to the progression to chronicity associated with infection.

项目总结/摘要 人类B型肝炎病毒（HBV）引起慢性肝炎。慢性B型肝炎是一个主要的危险因素， 进展至肝细胞癌（HCC）发作。尽管有一种有效的疫苗， 据估计，全世界有3.5亿感染者。抗病毒药物不能消除HBV DNA， 原子核更好地理解宿主和病毒因素在慢性肝炎中的作用， needed. HBV基因表达的表位调控提供了另一层调控机制， 基因表达。在这项研究中，我们提出了病毒RNA的转录后修饰的特点， 腺苷碱基的N6位称为m6 A。我们在拨款中描述的广泛的初步研究 应用表明HBV RNA被甲基化为m6 A。利用宿主甲基化酶沉默策略 和脱甲基酶参与表转录组的稳态，我们提出的证据表明， m6 a修改在缺乏酶的情况下， 催化m6 A修饰。我们用MeRIP免疫沉淀法和Tru-seq分析的结果鉴定了 富含m6 A残基峰的潜在序列范围。我们突变了大约13个'A'残基， HBV基因组中从1700到2000个核苷酸的区域。这些被检查， HBV蛋白的翻译和逆转录功能。结果是惊人的，因为，翻译是 m6 A的修饰负调控逆转录，而m6 A修饰正调控逆转录。基于这些 令人兴奋的数据，我们建议继续表征HBV RNA的m6 A修饰，并确定 这种修饰在HBV感染中的功能相关性。本研究为HBV的研究开辟了一条新的途径 研究独特的HBV生命周期的表观转录调控以及可能的作用， 与感染相关的慢性化进展。