Experimental identification of functional GWAS variants linked to COVID-19 severity in immune cells

免疫细胞中与 COVID-19 严重程度相关的功能性 GWAS 变异的实验鉴定

基本信息

批准号：
10741007
负责人：
BENJAMIN JOACHIM SCHMIEDEL
金额：
$ 22.88万
依托单位：
LA JOLLA INSTITUTE FOR IMMUNOLOGY
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：
2023-05-22 至 2025-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10741007
关键词：
Affect Alleles Antiviral Response Automobile Driving Binding Biological Assay COVID-19 COVID-19 impact COVID-19 pathogenesis COVID-19 risk COVID-19 severity COVID-19 susceptibility CRISPR interference Candidate Disease Gene Cells Clinical Code DNA Sequence Data Set Databases Defect Disease Economics Enhancers Ethnic Origin Exploratory/Developmental Grant Future Gene Expression Gene Expression Profile Genes Genetic Genetic Determinism Genetic Markers Genetic Polymorphism Genetic Risk Hospitalization Human IL10RB gene Immune Immune response Immune system Impairment Individual Inherited Interferons Interleukin-10 Link Long COVID Luciferases Maps Mediating Meta-Analysis Modeling Molecular Morbidity - disease rate Natural Killer Cells Pathogenesis Pathway interactions Patients Population Predisposition Protein Family Quantitative Trait Loci Reporter Reporting Respiratory Failure Risk Role SARS-CoV-2 infection Severities Severity of illness Symptoms T-Lymphocyte Testing Untranslated RNA Vaccines Validation Variant Work acute infection breakthrough infection causal variant cell type disorder risk epigenomics exome sequencing experimental study genetic analysis genome wide association study genome-wide genomic locus immunoregulation in vivo insight interleukin-10 receptor model building monocyte mortality novel oligoadenylate pandemic disease promoter response risk variant severe COVID-19 transcription factor transcriptome viral RNA

项目摘要

PROJECT SUMMARY/ABSTRACT The clinical presentation of SARS-CoV-2 infection in humans can range from very mild or no symptoms to severe respiratory failure. Although hyperactivation of various cellular components of the immune system have been observed in patients with severe COVID-19 illness, the host genetic factors that determine susceptibility to severe COVID-19 illness are not well understood. GWAS studies have reported several genetic loci that are significantly associated with severe COVID-19 and defined several target genes based on their proximity to the risk loci, although this approach does not accurately prioritize causal genes. We conducted one of the first transcriptome-wide association study (TWAS) in primary immune cells to define genes that are associated with COVID-19 severity. In this R21 proposal, we will focus on defining functional COVID-19 risk variants linked to top two candidate genes (OAS1 and IL10RB) from our genetic analyses. In Aim 1, we will determine the functional COVID-19 risk eQTLs associated with OAS1 expression in non- classical monocytes (NCM). Our TWAS study showed significant association of COVID-19 severity with reduced expression of interferon-inducible gene (OAS1) specifically in NCM. OAS1 encodes for oligoadenylate synthase family of proteins that degrade viral RNA and activate anti-viral responses, thus variants altering its expression levels, especially in NCM, a cell type which has been implicated in COVID-19 pathogenesis, are likely to have an important role in host immune responses. Here, we will undertake experimental studies to define the functional variant(s) in the dense OAS1 Neanderthal haploblock harboring >100 SNPs. Briefly, we will perform (i) CRISPRi assays to determine functional enhancers that overlap COVID-19 risk SNPs, (ii) luciferase reporter assays to determine the function variants in such enhancers, (iii) ChIP to determine allele-specific binding of the transcription factor RXRα the binding of which is predicted to be disrupted by COVID-19-risk variants, and (iv) HDR-mediated editing of prioritized COVID-19-risk eQTLs to definitively establish functionality. In Aim 2, we will determine the function COVID-19 risk eQTLs associated with IL10RB expression in NK cells and T cells. Our TWAS showed that increased expression of IL10RB in NK cells was significantly associated with COVID-19 severity. IL10RB encodes for IL-10 receptor beta, and given the immunomodulatory role of IL- 10, it is likely that the higher expression on the IL10RB in NK cells and T cells may enhance their responsiveness to IL-10. Here, we will perform experimental studies as described in Aim 1 to define the functional COVID-19- risk variants that are predicted to perturb the binding of the transcription factors TCF12 and GATA-3 and their role in the modulation of IL10RB expression in NK cells in T cells. Overall, functional studies in this R21 program will provide important mechanistic insights into the genetic basis of COVID-19 severity.

项目概要/摘要人类 SARS-CoV-2 感染的临床表现可以从非常轻微或无症状到严重尽管免疫系统的各种细胞成分过度激活。在患有严重 COVID-19 疾病的患者中观察到，决定易感性的宿主遗传因素严重的 COVID-19 疾病尚未得到充分了解。 GWAS 研究报告了几个基因位点。与严重的 COVID-19 显着相关，并根据与严重的 COVID-19 的接近程度定义了几个目标基因风险位点，尽管这种方法不能准确地优先考虑因果基因，我们进行了第一个研究。原代免疫细胞中的全转录组关联研究（TWAS），以确定与在本 R21 提案中，我们将重点定义与 COVID-19 相关的功能性 COVID-19 风险变体。我们的遗传分析显示了前两个候选基因（OAS1 和 IL10RB）。在目标 1 中，我们将确定与非非感染者中 OAS1 表达相关的功能性 COVID-19 风险 eQTL。我们的 TWAS 研究显示，COVID-19 的严重程度与降低的显着相关性。干扰素诱导基因 (OAS1) 在 NCM 中表达，OAS1 编码寡腺苷酸合酶。降解病毒 RNA 并激活抗病毒反应的蛋白质家族，从而改变其表达的变体水平，特别是在 NCM（一种与 COVID-19 发病机制有关的细胞类型）中，可能具有在这里，我们将进行实验研究来定义功能。密集的 OAS1 尼安德特人单倍体中的变体含有 >100 个 SNP。简而言之，我们将执行 (i) CRISPRi。确定与 COVID-19 风险 SNP 重叠的功能增强剂的测定，(ii) 荧光素酶报告基因测定确定此类增强子中的功能变体，(iii) ChIP 以确定增强子的等位基因特异性结合转录因子 RXRα，其结合预计会被 COVID-19 风险变异体破坏，以及 (iv) HDR 介导的优先 COVID-19 风险 eQTL 编辑，以明确建立功能。在目标 2 中，我们将确定与 NK 细胞中 IL10RB 表达相关的 COVID-19 风险 eQTL 功能我们的 TWAS 显示 NK 细胞中 IL10RB 表达的增加显着相关。 IL10RB 编码 IL-10 受体 β，并具有 IL-19 的免疫调节作用。如图 10 所示，NK 细胞和 T 细胞中 IL10RB 的较高表达可能会增强它们的反应性在这里，我们将按照目标 1 中的描述进行实验研究，以定义功能性 COVID-19- 预计会扰乱转录因子 TCF12 和 GATA-3 结合的风险变异及其 T 细胞中 NK 细胞 IL10RB 表达的调节作用。总体而言，R21 计划中的功能研究将为遗传基础提供重要的机制见解 COVID-19 的严重程度。