Post-GWAS Functional Genomics Analysis to Define Pathogenic Mechanisms for Pulmonary Arterial Hypertension

GWAS 后功能基因组学分析确定肺动脉高压的致病机制

基本信息

批准号：
10697364
负责人：
Wei Sun
金额：
$ 16.07万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-15 至 2027-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10697364
关键词：
Affinity Alleles Allelic Imbalance Animal Model Architecture Basic Science Binding Biology Blood Vessels CRISPR/Cas technology Cell Line Clinical Coupled DNA Binding DNA Sequence Alteration Data Dependence Dependovirus Development Diagnostic Disease Disease susceptibility Down-Regulation Endothelium Enterochromaffin Cells Environment Experimental Animal Model Functional disorder Future Gene Expression Genes Genetic Genetic Predisposition to Disease Genomic approach Genomics Genotype Goals Haplotypes Human Hypoxia Intervention Knockout Mice Knowledge Link Linkage Disequilibrium Lung Lung diseases Mediating Mentors Methods Molecular Molecular Target Mus Mutation Names Pathogenesis Pathogenicity Pathway interactions Patients Persons Physicians Predisposition Proteins Pulmonary Hypertension Reporting Research Research Personnel Research Project Grants Research Technics Resources Risk Role SOX17 gene Sampling Scientist Severities Single Nucleotide Polymorphism Structure of parenchyma of lung Techniques Technology Testing Training Translational Research Universities Untranslated RNA Validation Work career career development endothelial dysfunction familial hypertension functional genomics genetic association genome wide association study genomic locus genomic platform human DNA in vivo induced pluripotent stem cell insight knock-down knowledge base lung hypoxia mouse model nanoparticle nanopolymer novel overexpression pulmonary arterial hypertension pulmonary artery endothelial cell risk variant siRNA delivery skills stem cell biology transcription factor vector virulence gene

项目摘要

PROJECT SUMMARY Pulmonary arterial hypertension (PAH) is an enigmatic and morbid disease where insights are emerging regarding genetic susceptibility to disease. Genome-wide Association Studies (GWAS) have identified single nucleotide polymorphisms (SNPs) that are associated with PAH risk and severity. Yet, the GWAS-reported SNPs are only tags of haplotype SNPs in linkage disequilibrium (LD). Thus, the tag SNP may simply be linked to the true disease-causing functional SNP (fSNP). GWAS also only reveal statistical associations, and it has been challenging to define the mechanisms underlying the contribution of the PAH-associated fSNPs, which are mostly located in the non-coding regions, to the pathogenesis of PAH. Using our recently developed post-GWAS functional genomics platform, I identified a non-coding fSNP rs4738801 in the genomic locus of SOX17 gene, a known endothelial effector increasingly being studied in PAH pathogenesis. I identified that the transcription factor FUBP1 binds to the rs4738801 risk allele C in an allele-imbalanced manner, with lower affinity to risk allele C than non-risk allele G, providing an underlying mechanism how this fSNP regulates the PAH pathogenic gene SOX17 and contributes to the PAH risk. FUBP1 controls PAH-associated pathophenotypes in pulmonary arterial endothelial cells (PAECs). Downregulated by the major acquired PAH trigger hypoxia, FUBP1 and its target gene SOX17 are decreased in lungs and isolated pulmonary ECs from PAH patients and mouse models. A 3.77- fold enrichment of fSNP rs4738801 risk allele C was found in patients with PAH induced by hypoxia, supporting a pathogenic mechanism of a hypoxia-sensitive pathway (FUBP1-SOX17) combined with a disease susceptible genotype (risk allele C) for clinical manifestation of this disease. Based on these data, I hypothesize that the allele-imbalanced binding of transcription factor FUBP1 to fSNP rs4738801 defines the genomic architecture contributing to the SOX17-dependent genetic susceptibility of PAH. I further postulate that the downregulation of FUBP1-SOX17 by hypoxia contributes to the acquired pathogenesis of PAH. To test this hypothesis, I propose 2 specific aims: 1) To define the allele-specific role of fSNP rs4738801 in promoting endothelial dysfunction in PAH in gene-edited iPSC-ECs and PAH patient lung tissues; and 2) To determine the role of FUBP1 in controlling SOX17 and PAH in mouse models. Accomplishing these aims will facilitate my enduing career goal of becoming an independent physician-scientist in PAH functional genomics research. Immediate scientific development objectives include: 1) To develop expertise in PAH genetics and functional genomics; 2) To develop expertise in iPSC-EC biology and CRISPR-Cas9 gene-editing techniques; and 3) To develop skills of in vivo gene expression manipulation and become proficient in the assessment of PAH in animal models. The proposed training plan will provide me with the opportunity to expand my knowledge base to include advanced research techniques in PAH pathogenesis. The resources and expertise of my mentors, contributors, and the rich research environment at the University of Pittsburgh will assure my successful transition to an independent investigator.

项目概要肺动脉高压（PAH）是一种神秘而致病的疾病，人们的见解正在不断涌现关于疾病的遗传易感性。全基因组关联研究（GWAS）已经确定了单一与 PAH 风险和严重程度相关的核苷酸多态性 (SNP)。然而，GWAS 报告的 SNP 仅是连锁不平衡 (LD) 中单倍型 SNP 的标签。因此，标签 SNP 可以简单地链接到真正的致病功能性 SNP (fSNP)。 GWAS 也仅揭示统计关联，并且已被定义 PAH 相关 fSNP 的潜在机制具有挑战性，这些 fSNP 是大多位于非编码区，与PAH发病有关。使用我们最近开发的后 GWAS 通过功能基因组学平台，我在 SOX17 基因的基因组位点中发现了一个非编码 fSNP rs4738801，已知的内皮效应器越来越多地在 PAH 发病机制中得到研究。我发现转录因子 FUBP1 以等位基因不平衡的方式与 rs4738801 风险等位基因 C 结合，与风险等位基因的亲和力较低 C 比非风险等位基因 G 更重要，提供了该 fSNP 如何调节 PAH 致病基因的潜在机制 SOX17 会增加 PAH 风险。 FUBP1 控制肺动脉中 PAH 相关的病理表型内皮细胞（PAEC）。由主要获得性 PAH 引发缺氧的 FUBP1 及其靶点下调基因 SOX17 在肺中减少，并从 PAH 患者和小鼠模型中分离出肺 EC。 3.77- 在缺氧引起的 PAH 患者中发现了 fSNP rs4738801 风险等位基因 C 的倍数富集，支持缺氧敏感通路（FUBP1-SOX17）与疾病易感基因相结合的致病机制该疾病临床表现的基因型（风险等位基因 C）。根据这些数据，我假设转录因子 FUBP1 与 fSNP rs4738801 的等位基因不平衡结合定义了基因组结构导致 PAH 的 SOX17 依赖性遗传易感性。我进一步假设下调缺氧导致的 FUBP1-SOX17 的表达导致 PAH 的获得性发病机制。为了检验这个假设，我建议 2 个具体目标： 1) 明确 fSNP rs4738801 在促进内皮功能障碍中的等位基因特异性作用基因编辑 iPSC-EC 和 PAH 患者肺组织中的 PAH； 2) 确定 FUBP1 在控制中的作用小鼠模型中的 SOX17 和 PAH。实现这些目标将有助于我成为最终的职业目标多环芳烃功能基因组学研究的独立医师科学家。立即科学发展目标包括： 1) 发展 PAH 遗传学和功能基因组学方面的专业知识； 2) 发展专业知识 iPSC-EC 生物学和 CRISPR-Cas9 基因编辑技术； 3) 培养体内基因表达技能操作并精通动物模型中 PAH 的评估。拟议的培训计划将为我提供了扩展知识库的机会，包括 PAH 的先进研究技术发病。我的导师、贡献者的资源和专业知识以及丰富的研究环境匹兹堡大学将确保我成功过渡为独立调查员。