Functional characterization of regulatory variants associated with maternal hyperglycemia

与母体高血糖相关的调节变异的功能特征

基本信息

批准号：
9396924
负责人：
Graham Johnson
金额：
$ 5.71万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-07-01 至 2020-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9396924
关键词：
Acute Alleles Alpha Cell Architecture Automobile Driving Biological Assay CRISPR/Cas technology Child Chromatin Clustered Regularly Interspaced Short Palindromic Repeats Collaborations Complex DNA Data Development Diabetes Mellitus Dimensions Disease Disease susceptibility Dissection Distal Enhancers Etiology Exhibits Frequencies Future Gene Expression Gene Expression Regulation Gene Targeting General Population Genes Genetic Variation Genome Genomic approach Genomics Gestational Diabetes Glucose Goals Human In Situ Incidence Individual Insulin Life Maps Measurement Mediating Metabolic Metabolic Diseases Methodology Mothers Nucleic Acid Regulatory Sequences Obesity Outcome Pathology Pathway interactions Patients Phenotype Population Postpartum Period Pregnancy Race Regulator Genes Regulatory Element Reporter Repression Research Research Personnel Risk Sampling Testing Training Universities Validation Variant adverse outcome base blood glucose regulation cell type cohort diabetes risk diagnostic biomarker epigenome exhaustion fetal genetic approach genetic variant genome editing genome wide association study genome-wide analysis hexokinase insight maternal hyperglycemia medical schools metabolic phenotype novel novel therapeutics obesity risk perinatal outcomes predictive marker pregnant promoter risk variant screening tool trait transcriptome sequencing

项目摘要

Project Summary Maternal hyperglycemia and gestational diabetes mellitus (GDM) complicate 17% of pregnancies annually resulting in serious acute and lifelong perinatal outcomes for both mother and child. A genome-wide association study (GWAS) of 4,437 pregnant mothers established loci associated with gestational glycemic traits. Significant associations included those specific to pregnancy as well as several previously implicated with metabolic traits in non-gravid populations. The objective of this proposal is to quantify the effect of thousands of maternal hyperglycemia associated genetic variants on gene regulation. Our approach to meet this objective will use a population-scale high-throughput reporter assay to determine the effect of all variants in five maternal hyperglycemia GWA loci on gene regulation followed by targeted genome editing strategies and additional genomic assays. We have already completed targeted capture and sequencing of these regions for 835 mothers across four ancestries who exhibited metabolic phenotypes in the extreme deciles of their race group. My hypothesis is that both common and low frequency genetic variants in these regions contribute to maternal hyperglycemia and that the functional consequence of the causal variants will be altered gene expression. I will test that hypothesis in three specific aims. In Aim 1, I will use a high-throughput reporter approach developed by our lab to quantify the impact of putative enhancers and causal variants in maternal hyperglycemia loci on gene regulation. The advantage of assaying donor DNA directly is the ability to identify not only common, but also rare regulatory variants and relate them directly with phenotype. In Aim 2, I will map the long-range physical associations of enhancers to confirm their target genes. The outcome will be the first complete dissection of the three-dimensional cis-regulatory architecture of genomic loci implicated in a human metabolic disorder. In Aim 3, I will use CRISPR/Cas9-based genome and epigenome editing tools to functionally validate causal variants expected to perturb gene expression through altering enhancer activity. The expected outcomes are that I will identify the cis-regulatory framework driving maternal hyperglycemia and demonstrate a novel transferable GWAS screening methodology in a patient cohort. This will have immediate positive impacts on identifying mothers at risk for metabolic disease and new pathways for future studies.

项目摘要孕产妇高血糖和妊娠糖尿病（GDM）每年17％的怀孕复杂性导致母亲和儿童的严重急性和终生的围产期结果。全基因组 4,437位怀孕母亲的协会研究（GWAS）建立了与妊娠血糖相关的基因座特质。重要的关联包括特定于妊娠的关联以及几种先前涉及的在非格拉夫人群中具有代谢特征。该提案的目的是量化成千上万的母体高血糖与基因调节相关的遗传变异。我们见面的方法该目标将使用人口规模的高通量记者测定法来确定所有变体的效果在五个母体高血糖中，GWA基因座对基因调节，然后是针对性的基因组编辑策略和其他基因组测定法。我们已经完成了这些区域的目标捕获和测序对于四个祖先的835位母亲，他们在种族的极端十分中表现出代谢表型团体。我的假设是，这些区域中的常见和低频遗传变异都有助于母体高血糖和因果变异的功能后果将改变基因表达。我将以三个具体目标来检验该假设。在AIM 1中，我将使用高通量记者我们的实验室开发的方法是量化推定增强剂和因果变体对孕产妇的影响高血糖基因座基因调节。直接分析供体DNA的优点是识别的能力不仅常见，而且罕见的调节变体，并将它们直接与表型联系起来。在AIM 2中，我将映射增强子的远程物理关联以确认其靶基因。结果将是第一个完全解剖与人类有关的基因组基因局的三维顺式调节结构代谢障碍。在AIM 3中，我将使用基于CRISPR/CAS9的基因组和表观基因组编辑工具来在功能上，通过改变增强子活性来验证预期将扰动基因表达的因果变异。预期的结果是，我将确定驱动孕妇高血糖的顺式调节框架并在患者队列中展示了一种新型的可转移GWAS筛选方法。这将有直接对确定有代谢疾病风险的母亲和未来的新途径产生积极影响研究。