Virus-driven human gene misregulation in disease

病毒驱动的人类疾病基因失调

• 批准号: 10388202
• 负责人: Matthew Tyson Weirauch
• 金额: $ 67.26万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-15 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10388202
• 关键词: ATAC-seq; Algorithms; Alleles; Autoimmune; Autoimmune Diseases; Behavior; Binding; Biological Assay; CRISPR/Cas technology; Cause of Death; Cell Line; Cell Proliferation; Cells; ChIP-seq; Chromatin; Chromatin Loop; Clustered Regularly Interspaced Short Palindromic Repeats; Communities; Computer Models; Data; Data Set; Dependence; Disease; Disease Progression; Environment; Family; Foundations; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Diseases; Genetic Risk; Genetic Transcription; Genome; Growth Factor; HIV; Health; Herpes zoster disease; Herpesviridae; Herpesviridae Infections; Human; Human Genome; Human Herpesvirus 4; Human Herpesvirus 8; Human Papillomavirus; Individual; Libraries; Link; Lupus; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of urinary bladder; Maps; Measures; Mission; Modeling; Molecular; Monitor; Multiple Sclerosis; National Human Genome Research Institute; Nature; Papillomavirus; Pathogenesis; Pathogenicity; Pathologic; Phenotype; Physiological; Play; Process; Regulator Genes; Reporter; Research; Resources; Role; Technology; Testing; Therapeutic; Transfection; Update; Variant; Viral; Virus; Virus Diseases; base; chromatin immunoprecipitation; computational pipelines; computerized tools; cytokine; data tools; disorder risk; functional genomics; genetic variant; genome editing; genome-wide; genomic data; genomic locus; human disease; insight; novel; novel strategies; novel therapeutics; pathogen; release factor; risk variant; tat Protein; therapy development; transcription factor; transcriptome sequencing

ABSTRACT Thousands of genetic variants have been established for hundreds of human diseases. Yet, the vast majority of these diseases remain idiopathic. Interplay between genetics and the environment likely plays a role in many diseases. In particular, hundreds of associations between viral exposure and disease risk have been established. But with rare exceptions, the mechanisms underlying increased disease risk are unknown. We have recently discovered that the Epstein-Barr virus EBNA2 transcriptional co-factor binds up to half of the risk loci associated with seven autoimmune diseases (142 loci in total), with many examples of allele-dependent EBNA2 binding to autoimmune risk variants. We hypothesize that risk allele-dependent binding of viral TFs explains why other viruses cause or influence specific diseases. However, the data required to discover these mechanisms are currently incomplete. Herpesviruses and human papilloma virus play established roles in several human diseases, and their genomes encode many TFs. We will generate the functional genomics datasets needed to discover the roles of these viral TFs (vTFs) in human disease processes. We anticipate discovering multiple causal human disease variants whose mechanisms act in a viral TF and allele- dependent manner, leading to understanding of disease mechanisms and new therapeutic opportunities. Our approach is a generalizable blueprint for global characterization of pathogenic effects on host gene regulation. Aim 1. Create global maps of viral TF-driven human gene regulation. For eight viruses, we will transfect a viral TF into physiologically and pathologically relevant human primary cells and cell lines. We will measure the effect of vTFs on human gene expression by performing RNA-seq in cells with and without vTF transfection. We will monitor the binding of vTFs to the human genome using chromatin immunoprecipitation and calculate the enrichment of each vTF at established risk loci for all human diseases using our RELI algorithm. Aim 2. Uncover the mechanisms and downstream functional impact of viral TF-human genome interactions. We will characterize the mechanisms by which vTFs alter the human regulatory landscape. We will measure the effect of vTFs on human chromatin accessibility (ATAC-seq) and DNA looping (HiChIP-seq). We will use these datasets to construct computational models evaluating disease-relevant mechanisms. We will examine downstream effects of vTF activity on human cell phenotypes by monitoring cell proliferation, cytokine release, and growth factor release subsequent to vTF transfection. Aim 3. Test the allele-dependency of viral TF-provoked human disease mechanisms. We will identify vTF interactions involving human disease risk allele-dependent mechanisms. We will functionally screen for vTF- and disease risk allele-dependent effects on gene regulatory activity using Massively Parallel Reporter Assays. We will interrogate virus-host genomic datasets for allelic behavior using our MARIO computational pipeline. We will validate risk allele-dependent vTF mechanisms using CRISPR-based genome editing.

摘要 已经建立了数百种人类疾病的数千种基因变异。然而，浩瀚的 这些疾病中的大多数仍然是特发性的。遗传和环境之间的相互作用可能起到了一定的作用 在许多疾病中。特别是，病毒暴露和疾病风险之间的数百种关联已经被 已经成立了。但除了极少数例外，疾病风险增加背后的机制尚不清楚。我们有 最近发现，Epstein-Barr病毒EBNA2转录辅助因子可结合多达一半的风险基因 与7种自身免疫性疾病(总共142个基因座)有关，有许多等位基因依赖的EBNA2的例子 与自身免疫风险变异体结合。我们假设病毒转录因子的风险等位基因依赖的结合解释了 为什么其他病毒会引起或影响特定的疾病。然而，发现这些数据所需的数据 机制目前还不完善。疱疹病毒和人乳头瘤病毒在 几种人类疾病，它们的基因组编码了许多TF。我们将产生功能基因组学 需要数据集来发现这些病毒转录因子(VTF)在人类疾病过程中的作用。我们期待着 发现多种导致人类疾病的变种，其机制作用于病毒转铁蛋白和等位基因- 依赖的方式，导致对疾病机制和新的治疗机会的理解。我们的 该方法是病原菌对宿主基因调控影响的全球表征的概括性蓝图。 目的1.创建病毒转铁蛋白驱动的人类基因调控的全球图谱。对于八种病毒，我们将 将病毒转运蛋白导入生理和病理相关的人类原代细胞和细胞系。我们会 用RNA-SEQ技术检测VTFS对人类基因表达的影响 转染法。我们将使用染色质免疫沉淀来监测VTFS与人类基因组的结合 并使用我们的RERI算法计算每个VTF在所有人类疾病的已建立的风险位置的富集度。 目的2.揭示病毒转铁蛋白-人基因组的作用机制及其下游功能影响 互动。我们将描述VTFS改变人类监管格局的机制。我们 将测量VTFS对人类染色质可及性(ATAC-SEQ)和DNA环化(HiChIP-SEQ)的影响。 我们将使用这些数据集来构建评估疾病相关机制的计算模型。我们会 通过监测细胞增殖、细胞因子检测VTF活性对人类细胞表型的下游影响 VTF转染后释放生长因子。 目的3.检测病毒转铁蛋白引起的人类疾病机制的等位基因依赖性。我们会 确定涉及人类疾病风险等位基因依赖机制的VTF相互作用。我们将在功能上 用大规模平行方法筛选VTF和疾病风险等位基因对基因调节活性的依赖效应 记者阿萨德。我们将使用我们的Mario查询病毒宿主基因组数据集的等位基因行为 计算管道。我们将使用基于CRISPR的基因组来验证风险等位基因依赖的VTF机制 正在编辑。