Going from Genetic Associations to Identification of Causative Genes

从遗传关联到致病基因的识别

• 批准号: 10555812
• 负责人: Allan I Pack
• 金额: $ 66.63万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2028-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10555812
• 关键词: 3-Dimensional; ATAC-seq; Adipocytes; Affect; Anatomy; Animal Model; Animals; Astrocytes; Behavior; Biological; Biological Assay; Biological Models; CRISPR/Cas technology; Candidate Disease Gene; Cells; Chromatin; Chromatin Loop; Chromosome Mapping; Clinical; Communities; Complement; Complex; Data; Disease; Drosophila genus; Drowsiness; Enhancers; Excessive Daytime Sleepiness; Fatty acid glycerol esters; Fishes; Gene Expression; Gene Targeting; Genes; Genetic; Genome; Genomics; Genotype; Goals; Human; Image; Induced pluripotent stem cell derived neurons; Investments; Knock-out; Knowledge; Link; Magnetic Resonance Imaging; Maps; Medicine; Mesenchymal Stem Cells; Methods; Modeling; Mus; Neuroglia; Neurons; Obesity; Obstructive Sleep Apnea; Osteoblasts; Pathway interactions; Patients; Phenotype; Process; Quantitative Trait Loci; RNA Interference; RNA interference screen; Research; Resolution; Resources; Risk; Risk Factors; Sampling; Severities; Shapes; Signal Transduction; Site; Sleep; Sleep Apnea Syndromes; Sleep Fragmentations; Sleep disturbances; Sleeplessness; Testing; Tongue; Transposase; Validation; Variant; Zebrafish; candidate identification; causal variant; cell type; clinical translation; craniofacial; disorder risk; diverse data; fly; follow-up; gene function; genetic association; genetic variant; genome wide association study; genome-wide; genomic locus; in silico; knock-down; knockout gene; multidimensional data; novel; precision medicine; programs; promoter; sleep behavior; trait; transcriptome sequencing; 3-Dimensional; ATAC-seq; Adipocytes; Affect; Anatomy; Animal Model; Animals; Astrocytes; Behavior; Biological; Biological Assay; Biological Models; CRISPR/Cas technology; Candidate Disease Gene; Cells; Chromatin; Chromatin Loop; Chromosome Mapping; Clinical; Communities; Complement; Complex; Data; Disease; Drosophila genus; Drowsiness; Enhancers; Excessive Daytime Sleepiness; Fatty acid glycerol esters; Fishes; Gene Expression; Gene Targeting; Genes; Genetic; Genome; Genomics; Genotype; Goals; Human; Image; Induced pluripotent stem cell derived neurons; Investments; Knock-out; Knowledge; Link; Magnetic Resonance Imaging; Maps; Medicine; Mesenchymal Stem Cells; Methods; Modeling; Mus; Neuroglia; Neurons; Obesity; Obstructive Sleep Apnea; Osteoblasts; Pathway interactions; Patients; Phenotype; Process; Quantitative Trait Loci; RNA Interference; RNA interference screen; Research; Resolution; Resources; Risk; Risk Factors; Sampling; Severities; Shapes; Signal Transduction; Site; Sleep; Sleep Apnea Syndromes; Sleep Fragmentations; Sleep disturbances; Sleeplessness; Testing; Tongue; Transposase; Validation; Variant; Zebrafish; candidate identification; causal variant; cell type; clinical translation; craniofacial; disorder risk; diverse data; fly; follow-up; gene function; genetic association; genetic variant; genome wide association study; genome-wide; genomic locus; in silico; knock-down; knockout gene; multidimensional data; novel; precision medicine; programs; promoter; sleep behavior; trait; transcriptome sequencing

ABSTRACT Many studies, including analyses in this overall Program of research, have conducted genome-wide association studies (GWAS) to identify genes and loci associated with complex disease. While a number of genetic association signals have been uncovered, the challenge of identifying causative genes at these genetic loci remains. As such, the goal of this Project is to move from GWAS association to identification of causal effector genes relevant to obstructive sleep apnea (OSA) and excessive sleepiness, two key traits studied in this Program. To fill this critical gap in evidence this Project combines state-of-the-art approaches in cell-based and animal models. Analyses will begin by interrogating genetic loci from recent GWAS on OSA and sleepiness, and be extended to evaluate loci from ongoing analyses, including those in Projects 01 and 03. In Aim 1, we will conduct in silico physical `variant to gene mapping' based on our established Assay for Transposase Accessible Chromatin sequencing (ATAC-seq) plus genome-wide promotor-focused Capture C data on relevant cell types - osteoblasts and adipocytes (for anatomy; relevant to Project 01) and neurons and primary astrocytes (for sleepiness; relevant to Project 03). These 3D Genomics analyses will identify the most likely causal genes and variants by determining which candidates at implicated loci directly interact with regions of open chromatin. After identifying likely causal genes and variants, follow-up analyses in animal models will be performed to understand if candidate effector genes act by affecting OSA-related anatomy in mice (Aim 2) and sleep behavior reflective of sleepiness and disturbed sleep in Drosophila and zebrafish (Aim 3). Specifically, Aim 2 will utilize the novel multivariate genotype-phenotype mapping pipeline we developed to identify causal genes affecting OSA risk through effects on craniofacial shape and/or tongue fat. In applying this method, this Aim leverages both existing and newly generated data in a large sample of Diversity Outbred mice with genetic data and anatomical traits quantified via imaging. This cutting-edge approach facilitates determination of the effects of multiple genes on multivariate phenotypes using high-dimensional data to compare directions, not just magnitudes, of associations. For sleepiness, Aim 3 will utilize Drosophila RNAi lines to study the impact of knocking-down candidate genes on sleep-related phenotypes (including sleep amounts, sleep fragmentation, and sleep drive). Then, genes shown to affect sleep in Drosophila will be validated in a vertebrate model by utilizing CRISPR-Cas9 methods to knock-out these same genes in zebrafish and studying the effects on similar sleep phenotypes. Taken together, results in this Project and the other projects in this Program will provide essential knowledge about effector genes for OSA and for sleepiness in OSA in humans. This knowledge is crucial for understanding the biological and clinical impact of GWAS associations, and will facilitate more efficient clinical translation and incorporation of genetic evidence into precision medicine approaches to OSA, as well as provide an important resource for the broader scientific community and proof of principle for the field of applied genomics in OSA.

抽象的 许多研究，包括该整体研究计划中的分析，都进行了全基因组 关联研究（GWAS）识别与复杂疾病相关的基因和基因座。虽然有很多 遗传关联信号已经被发现，在这些遗传上鉴定因果基因的挑战 位置仍然存在。因此，该项目的目标是从GWAS关联转变为因果关系的识别 与阻塞性睡眠呼吸暂停（OSA）和过度嗜睡有关的效应基因，其中两个关键特征在此研究 程序。为了填补这一重要差距，该项目结合了基于细胞和细胞的最新方法 动物模型。分析将首先询问最近关于OSA和嗜睡的GWA的遗传基因座，以及 扩展以评估正在进行的分析中的基因座，包括项目01和03中的基因座。在AIM 1中，我们将 基于我们已建立的转座酶可访问的测定法，以硅物理“变体与基因映射”进行进行 染色质测序（ATAC-SEQ）加上全基因组启动子捕获C数据相关细胞类型的数据 - 成骨细胞和脂肪细胞（用于解剖学；与项目01相关）和神经元和主要星形胶质细胞（用于 嗜睡；与项目03有关）。这些3D基因组学分析将确定最可能的因果基因和 通过确定哪些在含义的基因座的候选者直接与开放染色质区域相互作用来变体。后 识别可能的因果基因和变异，将进行动物模型的后续分析以了解 如果候选效应基因通过影响小鼠中与OSA相关的解剖学作用（AIM 2）和反射性睡眠行为 果蝇和斑马鱼中的嗜睡和睡眠干扰（AIM 3）。具体而言，AIM 2将利用小说 多元基因型 - 表型映射管道我们开发了用于识别影响OSA风险的因果基因 通过对颅面形状和/或舌头脂肪的影响。在应用此方法时，此目标利用了两个现有的 以及具有遗传数据和解剖学特征的多样性繁殖小鼠样本中新生成的数据 通过成像量化。这种尖端的方法有助于确定多个基因对 使用高维数据的多元表型比较了关联的方向，而不仅仅是幅度。 对于嗜睡，AIM 3将利用果蝇RNAi线研究敲门候选基因的影响 在与睡眠有关的表型（包括睡眠量，睡眠碎片和睡眠驱动器）上。然后，基因 通过利用CRISPR-CAS9方法来验证果蝇中的睡眠 在斑马鱼中淘汰这些相同的基因，并研究对相似睡眠表型的影响。在一起， 该项目的结果以及该计划中的其他项目将提供有关效应基因的基本知识 对于OSA和人类OSA的嗜睡。这些知识对于理解生物学和 GWAS关联的临床影响，并将促进更有效的临床翻译和融合 遗传证据纳入OSA的精确医学方法，并为 OSA应用基因组学领域的更广泛的科学界和原则证明。