Transancestral fine mapping and functional dissection of autophagy-related SLE risk loci

自噬相关 SLE 风险位点的跨祖先精细定位和功能解剖

• 批准号: 9898320
• 负责人: Christopher J Lessard
• 金额: $ 70.12万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-04-09 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9898320
• 关键词: Affect; African; African American; Age; Alleles; Amino Acids; Antigen Presentation; Apoptotic; Asians; Autoimmune Diseases; Autoimmunity; Autophagocytosis; B-Lymphocytes; Bayesian Method; Binding Proteins; Bioinformatics; Biological; Biological Assay; Biological Process; CD4 Positive T Lymphocytes; CDKN1B gene; CRISPR/Cas technology; Candidate Disease Gene; Cell Cycle Progression; Cell Line; Cells; Cellular Assay; Chromatin; Chronic; Clathrin; Codon Nucleotides; Complex; Crohn' s disease; Custom; Cytoplasmic Organelle; Data; Development; Disease; Dissection; Endosomes; Etiology; European; Exhibits; Exposure to; Functional disorder; Gender; Gene Expression; Genes; Genetic; Genetic Polymorphism; Genetic Variation; Genetically Engineered Mouse; Genomics; Genotype; Haplotypes; Homeostasis; Homologous Gene; Host Defense; Human; Immune; Immune response; Immune system; Immunophenotyping; Individual; Inflammation; Koreans; Link; Linkage Disequilibrium; Lymphocyte; Lymphoid; Lysosomes; Mediating; Microbe; Minority; Molecular; Molecular Biology; Mus; Myelogenous; NADPH Oxidase; Odds Ratio; Organ; PRDM1 gene; Pathogenesis; Pathway interactions; Patient Care; Patients; Phagocytes; Play; Pluripotent Stem Cells; Population; Population Control; Process; Proteins; Public Health; Publishing; Quantitative Reverse Transcriptase PCR; RNA Splicing; Recycling; Reporting; Risk; Role; Sample Size; Signal Transduction; Sorting - Cell Movement; Spliced Genes; Stimulus; Structure; Susceptibility Gene; System; Systemic Lupus Erythematosus; T-Cell Proliferation; T-Lymphocyte; TLR7 gene; TP53 gene; Testing; Therapeutic; Thymic epithelial cell; Tissue-Specific Gene Expression; Transcript; Transport Process; Tumor Suppressor Proteins; Variant; Woman; autoinflammatory; case control; causal variant; cell type; chromatin protein; clinical care; cytokine; effective therapy; ethnic minority population; experimental study; genetic variant; genome wide association study; genomic locus; in vivo; induced pluripotent stem cell; insight; lupus-like; lysosomal proteins; member; monocyte; neutrophil; new therapeutic target; novel; novel therapeutics; pathogen; plasma cell differentiation; response; risk variant; screening; targeted treatment; tool; transcriptome sequencing

Systemic lupus erythematosus (SLE) is a chronic, heterogeneous autoimmune disease that disproportionally affects women and the disease is often more severe with end-organ damage in non-white populations. Few effective treatments exist for SLE, largely because the etiology is incompletely understood; however, the disease is likely to occur in genetically susceptible individuals upon environmental triggers. Multiple genome wide association studies (GWAS) and replication studies have identified ~100 (80 published and 20 in the publishing process) SLE susceptibility loci (P<5x10-8) using mainly European- and Asian-derived populations. Our recent Korean GWAS and other published studies of Asians have implicated a number of autophagy- related gene loci in SLE, similar to several other autoimmune diseases. Autophagy is an intracellular process that transports damage cytoplasmic organelles, macromolecular aggregates, or intracellular pathogens to the lysosome for degradation and recycling. Autophagy cross talks with the immune system and controls inflammation through effects on various immune cells to regulate immune homeostasis and to modulate host defenses. Fine mapping autophagy-related gene loci associated with SLE is necessary to gain understanding how these genetic variants affect different aspects of autophagy pathways in specific immune cell types, which may serve as a new drug target for SLE treatment. Due to linkage disequilibrium (LD), each risk locus contains multiple variants, and in some cases, multiple genes that could impact biologic functions relating to SLE pathogenesis. In Aim 1, we propose to utilize LD structure differences between Asian and African-derived ancestries to conduct transracially fine-mapping and bioinformatics analysis of seven SLE risk loci related to autophagy (ATG16L2-FCHSD2-P2RY2, PRDM1-ATG5, DRAM1, CDKN1B, CLEC16A, NCF2 and HIP1) using the Global Screening Array (GSA) that contains headroom for custom variants in SLE cases and controls of non-white minorities (4000 Asians and 3000 African Americans [AA]). By comparing haplotype structures and applying Bayesian approaches, we will obtain the best credible set of variants to pursue further. In Aim 2, we will refine credible variant list by using bioinformatics and molecular approaches to identify functional variants. This will include sorting immune cell subsets from 100 AA SLE patients and 100 AA controls (matched for age and gender) for RNA-seq and qRT-PCR to correlate allelic genotypes with differential gene expression and splicing as well as conducting molecular biology assays. The best candidates will be further tested in Aim 3 to conduct immunophenotyping, RNA-seq and autophagy functional assays to assess the consequences of gene- edited immune cell lines and induced pluripotent stem cells (iPSCs), which will determine the pertinent cell subset from SLE patients and controls to conduct autophagy functional assays for confirmation of genotypic effects. Our results will identify genetic variants underlying SLE risk loci and their molecular/cellular mechanisms leading to dysregulation of autophagy pathways contributing to the development of SLE.

系统性红斑狼疮 (SLE) 是一种慢性、异质性自身免疫性疾病， 影响女性，并且在非白人群体中，这种疾病往往更严重，伴有终末器官损伤。很少 SLE 存在有效的治疗方法，主要是因为其病因尚不完全清楚；然而， 在环境触发下，遗传易感个体可能会发生疾病。多基因组 广泛关联研究 (GWAS) 和重复研究已确定约 100 项（其中 80 项已发表，20 项已在 出版过程）SLE 易感位点 (P<5x10-8) 主要使用欧洲和亚洲人群。 我们最近的韩国 GWAS 和其他发表的针对亚洲人的研究表明，许多自噬- SLE 中的相关基因位点，与其他几种自身免疫性疾病类似。自噬是细胞内的过程 将受损的细胞质细胞器、大分子聚集体或细胞内病原体转运至 溶酶体用于降解和回收。自噬与免疫系统和控制相互作用 通过影响各种免疫细胞来调节免疫稳态和调节宿主而产生炎症 防御。精细定位与 SLE 相关的自噬相关基因位点对于获得了解是必要的 这些遗传变异如何影响特定免疫细胞类型中自噬途径的不同方面， 可能作为SLE治疗的新药物靶点。由于连锁不平衡（LD），每个风险位点包含 多种变异，在某些情况下，多种基因可能影响与 SLE 相关的生物功能 发病。在目标 1 中，我们建议利用亚洲和非洲衍生的 LD 结构差异 祖先对与以下疾病相关的七个 SLE 风险位点进行跨种族精细定位和生物信息学分析 使用自噬（ATG16L2-FCHSD2-P2RY2、PRDM1-ATG5、DRAM1、CDKN1B、CLEC16A、NCF2 和 HIP1） 全局筛选阵列 (GSA)，其中包含 SLE 病例和对照中自定义变异的空间 非白人少数族裔（4000 名亚洲人和 3000 名非裔美国人 [AA]）。通过比较单倍型结构和 应用贝叶斯方法，我们将获得最可信的变体集以进行进一步的研究。在目标 2 中，我们 将通过使用生物信息学和分子方法来识别功能变异来完善可靠的变异列表。 这将包括对来自 100 名 AA SLE 患者和 100 名 AA 对照（年龄匹配）的免疫细胞子集进行分类 和性别）用于 RNA-seq 和 qRT-PCR，将等位基因型与差异基因表达相关联 剪接以及进行分子生物学测定。最好的候选人将在目标 3 中接受进一步测试 进行免疫表型分析、RNA-seq 和自噬功能测定，以评估基因- 编辑免疫细胞系和诱导多能干细胞 (iPSC)，这将确定相关细胞 来自 SLE 患者和对照的子集进行自噬功能测定以确认基因型 影响。我们的结果将识别 SLE 风险位点及其分子/细胞的遗传变异 导致自噬途径失调的机制有助于系统性红斑狼疮的发展。