Developmental regulation of HBV biosynthesis by Ten-eleven translocation (Tet) methylcytosine dioxygenases

十十一易位 (Tet) 甲基胞嘧啶双加氧酶对 HBV 生物合成的发育调节

基本信息

批准号：
10733902
负责人：
Alan McLachlan
金额：
$ 39.12万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-06-09 至 2028-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10733902
关键词：
Acute Adult Anabolism Antiviral Therapy Biological Models Capsid Cell Nucleus Cell Therapy Cessation of life Chromatin Structure Chromosome Deletion Chronic Chronic Hepatitis B Circular DNA Clinical Cytoplasm DNA DNA Methylation DNA sequencing Development Dioxygenases Epigenetic Process Gene Silencing Genetic Genetic Transcription Genome Genomic DNA Genomics Grant Health Hepatitis B Virus Histones Individual Life Cycle Stages Liver LoxP-flanked allele Mediating Methylation Molecular Molecular Weight Mus Nuclear Nucleotides Physiological Physiology Predisposition Protein translocation Proteins RNA Regulation Resolution Reverse Transcription Role Signal Transduction Suggestion System Tamoxifen Transgenes Transgenic Mice Viral Viral Genome Viral Load result Virus Diseases Virus Inactivation Virus Integration Virus Replication bisulfite cellular targeting chronic liver disease demethylation design effective therapy human pathogen in vivo inhibitor therapy liver development liver function mouse model new therapeutic target novel novel strategies postnatal prevent recombinase targeted treatment therapeutic target

项目摘要

Project Summary and Relevance. Hepatitis B virus (HBV) infection is a worldwide health problem. It is estimated that there are 200 to 500 million HBV chronic carriers in the world for whom, to date, there is no reliable treatment. HBV causes both acute and chronic liver disease and is responsible for an estimated one million deaths annually. Currently available therapies reduce viral loads but fail to resolve chronic HBV infections. Therefore, effective treatments for chronic HBV infection are urgently required. The major obstacle to the resolution of chronic HBV infections is the eradication or inactivation of nuclear HBV covalently closed circular (ccc) DNA which is the template for viral transcription. To this end, we have developed HNF1α-null HBV transgenic mice and liver-specific Tet-deficient HBV transgenic mice. HNF1α-null HBV transgenic mice synthesize nuclear HBV cccDNA, a fraction of which is extensively methylated in adult mice. Liver-specific Tet-deficient HBV transgenic mice are viable, essentially display normal liver physiology, but lack detectable HBV transcription and replication (i.e. they are effectively “cured”) suggesting Tet is also essential for viral biosynthesis. The observation that Tet-deficient HBV transgenic mice fail to support HBV biosynthesis is consistent with the suggestion that Tet is essential for the developmental demethylation of HBV genomic DNA which epigenetically governs HBV transcription by modulating viral chromatin structure in vivo. Defining the precise temporal requirements for Tet expression associated with HBV transcription and replication will indicate the liver developmental stages when viral biosynthesis is susceptible to inhibition by Tet deficiency. This will be achieved by modulating Tet expression using our recently developed tamoxifen-inducible Tet-deficient HBV transgenic mouse model system. Using this system, the developmental control of HBV transcription, viral biosynthesis and HBV DNA methylation by Tet expression will be established and correlated with the epigenetic histone marks and chromatin structure associated with the HBV genome. Similar studies will be performed using the HNF1α-null Tet-deficient HBV transgenic mouse model of chronic viral infection so the developmental control of HBV transcription, viral biosynthesis, HBV DNA methylation, epigenetic histone marks and chromatin structure by Tet expression associated with the HBV genome can be compared between the HBV transgene DNA and the nuclear HBV cccDNA. Finally, the development of HBV transgenic mice supporting viral biosynthesis exclusively from extrachromosomal genomic DNA, a more physiologically relevant mouse model of chronic viral infection, will be developed so the effect of Tet-deficiency on HBV biosynthesis derived solely from HBV cccDNA can be determined. Defining the molecular signals responsible for the loss of HBV biosynthesis due to Tet deficiency may lead to the identification of cellular therapeutic targets, including but not restricted to Tet proteins, that are amenable to the development of novel small molecular weight therapeutic inhibitors to resolve rather than simply treat chronic HBV infection.

项目摘要和相关性。乙型肝炎病毒 (HBV) 感染是一个全球性的健康问题，估计有 200 至 5 亿人感染。迄今为止，世界上的乙型肝炎慢性携带者还没有可靠的治疗方法，乙型肝炎既可以引起急性乙型肝炎，也可以引起慢性乙型肝炎。据估计，慢性肝病每年导致 100 万人死亡。治疗可减少病毒载量，但无法解决慢性乙型肝炎感染，因此是慢性乙型肝炎的有效治疗方法。解决慢性乙型肝炎感染的主要障碍是乙型肝炎病毒感染。消除或灭活作为病毒模板的核 HBV 共价闭合环状 (ccc) DNA 为此，我们开发了 HNF1α 缺失的 HBV 转基因小鼠和肝脏特异性 Tet 缺陷的小鼠。 HNF1α 缺失的 HBV 转基因小鼠合成核 HBV cccDNA，其中一部分是肝脏特异性 Tet 缺陷的 HBV 转基因小鼠本质上是可行的显示正常的肝脏生理机能，但缺乏可检测到的 HBV 转录和复制（即，它们有效地 “治愈”）表明 Tet 对于病毒生物合成也是必需的。观察到 Tet 缺陷的 HBV 转基因。小鼠无法支持 HBV 生物合成，这与 Tet 对于发育至关重要的建议是一致的。 HBV 基因组 DNA 去甲基化，通过调节病毒从表观遗传学上控制 HBV 转录定义与 HBV 相关的 Tet 表达的精确时间要求。转录和复制将表明病毒生物合成易受感染的肝脏发育阶段 Tet 缺乏的抑制将通过使用我们最近开发的调节 Tet 表达来实现。他莫昔芬诱导的 Tet 缺陷 HBV 转基因小鼠模型系统使用该系统。通过 Tet 表达控制 HBV 转录、病毒生物合成和 HBV DNA 甲基化将被建立并与 HBV 基因组相关的表观遗传组蛋白标记和染色质结构相关。类似的研究将使用 HNF1α 缺失 Tet 缺陷的 HBV 转基因小鼠模型进行病毒感染，从而控制 HBV 转录、病毒生物合成、HBV DNA 甲基化、与 HBV 基因组相关的 Tet 表达的表观遗传组蛋白标记和染色质结构可以比较 HBV 转基因 DNA 和核 HBV cccDNA 最后，HBV 的发展。转基因小鼠仅支持从染色体外基因组 DNA 进行病毒生物合成，这是一种更将开发慢性病毒感染的生理相关小鼠模型，以便观察 Tet 缺乏的影响可以确定仅源自 HBV cccDNA 的 HBV 生物合成的分子信号。由于 Tet 缺乏而造成 HBV 生物合成损失的原因可能导致细胞鉴定靶标，包括但不限于治疗性 Tet 蛋白，适合开发新型药物小分子量治疗抑制剂可以解决而不是简单地治疗慢性乙型肝炎病毒感染。