Project II - Modeling meningomyelocele in frog using human alleles and folic acid exposure

项目 II - 使用人类等位基因和叶酸暴露模拟青蛙的脑膜脊髓膨出

基本信息

批准号：
10300071
负责人：
Christopher Robert Kintner
金额：
$ 29.65万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-12-01 至 2025-11-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10300071
关键词：
22q11 22q11.2 Address Alleles Animals Benign Biological Assay Biological Models Birth CISH gene Candidate Disease Gene Cells Chromatin Chromosome Deletion Clinical Collaborations Complex Congenital Abnormality Coupled DNA Methylation Data Development DiGeorge Syndrome Disease Dose Effectiveness Embryo Environmental Risk Factor Etiology Event Experimental Models Folic Acid Folic Acid Deficiency Foundations Gene Deletion Gene Expression Genes Genetic Goals Human Incidence Individual Injections Knock-out Lead Literature Mammals Measures Meningomyelocele Messenger RNA Modeling Molecular Morphogenesis Movement Mus Mutagenesis Mutate Mutation Nature Neural Tube Closure Neural Tube Defects Neural Tube Development Neural tube Pathway interactions Patients Pharmacology Phenotype Play Process Rana Recurrence Risk Risk Factors Role Spinal Study models Sum Syndrome Testing Tetrapoda Variant Work Xenopus Xenopus laevis base de novo mutation experimental study gene environment interaction gene interaction genetic risk factor genetic testing genetic variant in vivo loss of function mutation mouse model mutant next generation sequencing null mutation planar cell polarity programs transcriptome sequencing xenopus genome

项目摘要

Abstract – Project II: Modeling meningomyelocele in frog using human alleles and FA exposure Neural tube defects (NTDs) are a relatively common birth defect with a complex etiology and gene-environment interactions (GXEs). Genetic factors play an important role, and these gene variants likely interact with one another in gene-gene interactions (GXGs). A major focus of our Program Project is the assessment of de novo mutations that can be assessed in patients with NTDs, and their modulation by environmental risk factors, such as folic acid (FA) accessibility. This complex etiology has made NTDs an extremely challenging syndrome to predict based on genetic testing and to treat clinically. This proposal is a part of a comprehensive Program Project to investigate the genetic basis of the NTD subtype known as meningomyelocele (MM), localized to the spinal neural tube, and occuring in approximately one in every 2,500 births. Project I in the application uses next generation sequencing to identify de novo gene variants that are associated with human MM patients, and assess for recurrence. Even for recurrent mutations, it is critical to functionally evaluate causality in an in vivo setting. This project exploits Xenopus as a high throughput and high content tetrapod experimental model to assess these variants, taking advantage of the fact that the morphogenetic process of neural tube formation and the underlying molecular pathways involved in neurulation are conserved between Xenopus and mammalian embryos. CrispR mutagenesis in F0 Xenopus embryos will be used to test whether genes that lie within deletions of LCR22C-D in the 22q11.2 interval and that substantially increase risk of MM in humans, cause NTDs as null mutations. Gene variants detected in the planar cell polarity pathway that may increase the risk of MM in humans will be tested using rescue experiments in Xenopus, taking advantage of quantitative assays for measuring planar axis formation. Finally, the assessment of MM gene variants will also exploit recent studies in Xenopus, indicating that folate deficiency also causes NTDs as in mammals. Interactions between genetic risk factors and folate deficiency on the incidence of NTDs will be rapidly assessed using Xenopus, as part of the overall goal to determine whether FA alters gene expression and thus the expressivity of critical gene mutants. In sum, project II will use Xenopus as model to prioritize MM gene variants identified in Project I that can be further pursued in experiments in the mouse embryo in project III. Aim 1. Test genes in the minimal 22q11.2 deletion interval for a role in NTDs using Xenopus. Aim 2. Test human MM alleles for impact on neural tube formation in Xenopus embryos. Aim 3. Test GXE by assessing the impact of FA on MM gene phenotypes in Xenopus.

摘要 – 项目 II：使用人类等位基因和 FA 暴露模拟青蛙脑膜脊髓膨出神经管缺陷（NTD）是一种相对常见的出生缺陷，具有复杂的病因和基因环境相互作用（GXE）发挥着重要作用，这些基因变异可能与其中一个相互作用。我们计划项目的另一个重点是从头评估。可以在 NTD 患者中评估的突变及其受环境风险因素的调节，例如由于叶酸 (FA) 的可及性，这种复杂的病因使 NTD 成为一种极具挑战性的综合征。该提案是基于基因检测进行预测并进行临床治疗的综合计划的一部分。项目旨在调查被称为脑膜脊髓膨出 (MM) 的 NTD 亚型的遗传基础，该亚型定位于脊髓神经管，大约每 2,500 名新生儿中就有 1 人发生，应用程序中的项目 I 使用下一个。世代测序以识别与人类 MM 患者相关的从头基因变异，以及即使对于复发性突变，在体内功能性评估因果关系也至关重要。该项目利用非洲爪蟾作为高通量和高含量的四足动物实验模型。利用神经管形成的形态发生过程和爪蟾和哺乳动物之间参与神经溶解的潜在分子途径是保守的 F0 爪蟾胚胎中的 CrispR 诱变将用于测试是否存在缺失的基因。 22q11.2 间隔中的 LCR22C-D 会显着增加人类患 MM 的风险，导致 NTD 无效平面细胞极性通路中检测到的基因变异可能会增加 MM 的风险。人类将通过爪蟾救援实验进行测试，利用定量分析的优势最后，MM 基因变异的评估也将利用最近的研究。非洲爪蟾，表明叶酸缺乏也会导致 NTD，就像哺乳动物遗传风险之间的相互作用一样。作为该项目的一部分，将使用非洲爪蟾快速评估影响 NTD 发病率的因素和叶酸缺乏总体目标是确定 FA 是否改变基因表达，从而改变关键基因突变体的表达能力。总之，项目 II 将使用非洲爪蟾作为模型来优先考虑项目 I 中确定的 MM 基因变体，这些变体可以在项目 III 的小鼠胚胎实验中进一步进行了研究。目标 1. 使用爪蟾测试最小 22q11.2 缺失区间内的基因在 NTD 中的作用。目标 2. 测试人类 MM 等位基因对非洲爪蟾胚胎中神经管形成的影响。目标 3. 通过评估 FA 对非洲爪蟾 MM 基因表型的影响来测试 GXE。