Analysis of Poison Exon Inclusion in Genes Associated with Neurodevelopmental Disorders and Autism Spectrum Disorder

神经发育障碍和自闭症谱系障碍相关基因中毒外显子包含分析

• 批准号: 10438549
• 负责人: Stephanie Ann Felker
• 金额: $ 4.35万
• 依托单位: HUDSON-ALPHA INSTITUTE FOR BIOTECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10438549
• 关键词: Affect; Alternative Splicing; Automobile Driving; Bioinformatics; Biological; Biological Assay; Biological Factors; Biometry; Biotechnology; Brain; Calcium; Calcium Channel; Catalogs; Cations; Clinical; Code; Communication; DNA; Data; Data Set; Detection; Development; Developmental Delay Disorders; Developmental Gene; Diagnosis; Disease; Elements; Embryo; Environment; Epilepsy; Evolution; Exons; Family; Foundations; Future; Gene Expression; Gene Expression Regulation; Gene Family; Genes; Genetic; Genetic Transcription; Genetic Variation; Genome; Genomics; In Vitro; Individual; Infant; Informatics; Institutes; Intellectual functioning disability; Lead; Life; Linkage Disequilibrium; Machine Learning; Manuals; Messenger RNA; Methods; Molecular Biology; Molecular Diagnosis; Mus; Mutation; Neurodevelopmental Disorder; Neurologic; Nonsense-Mediated Decay; Onset of illness; Oral; Patients; Phenotype; Poison; Population; Protein Binding Domain; Protein Isoforms; Protein Truncation; Proteins; RNA; RNA Splicing; RNA immunoprecipitation sequencing; RNA-Binding Proteins; Reporter; Research; Resources; Role; SCN8A gene; Scientist; Seizures; Sodium; Sodium Channel; Source; Structure; Techniques; Terminator Codon; Tetrapoda; Therapeutic; Time Management; Tissues; Training; Transcript; Untranslated RNA; Variant; autism spectrum disorder; career development; causal variant; cohort; comparative; comparative genomics; disabling symptom; disease phenotype; dravet syndrome; driving force; early onset; exome; experimental study; genome sequencing; genome wide association study; genome-wide analysis; human disease; improved; insight; large scale data; loss of function; mRNA Decay; mRNA Precursor; machine learning method; member; nervous system disorder; neuron development; novel; postnatal; premature; prevent; proband; random forest; responsible research conduct; scale up; skills; therapeutic target; transcriptome sequencing; voltage; voltage gated channel

Project Summary/Abstract Over one in one hundred infants born are affected by neurodevelopmental disorders (NDDs), which can cause a host of debilitating symptoms including early-onset seizures, developmental delay, and intellectual disability. Application of whole exome/genome sequencing to examine NDD has revealed many disease-causing variations, but in a substantial proportion of cases the genetic cause remains undetermined. The analysis of deep-intronic non-coding variants has provided diagnosis when coding variation cannot be implicated in disease. For example, intronic variants in the “20N” poison exon of SCN1A results in loss of function, causing Dravet Syndrome, an NDD causing prolonged seizures in the first year of life. Poison exons are a mechanism of gene regulation whereby a premature termination codon caused by inclusion of the poison exon induces nonsense- mediated decay of the mRNA transcript. Preliminary analysis has shown that poison exon inclusion (PEI) in SCN1A and SCN8A decreases during embryonic brain development, inversely proportional to total RNA expression. The brain is a hotbed of alternative splicing during early neuronal development therefore it is plausible, if not probable that novel poison exons remain uncharacterized. The overall hypothesis of this proposal is that PEI is a mechanism to differentiallyregulate gene expression during earlyneuronal development, and intronic variants affecting PEI in genes of developmental importance contribute to disease phenotype. In Aim 1, the sodium and calcium voltage-gated channel alpha subunit families will be assessed to detect and characterize novel poison exons. Known and novel PEI will then be determined in the developing brain using publicly available RNA-seq experiments. In Aim 2 this analysis will be scaled up over 100-fold to detect and assess PEI in over 3000 genes associated with NDD and Autism Spectrum Disorder (ASD) in the developing brain and the driving forces of PEI evolution will be assessed. Comparative analysis of poison exons will be conducted using k-means hierarchical clustering and unsupervised random forest machine learning methodologies to determine prime biological factors driving PEI utilization. In Aim 3, intronic variants will be analyzed in probands of NDD and ASD in regions of known and hypothetical PEI and the affect these variants have on disease phenotype will be computationally assessed. These experiments will reveal new insights into alternative splicing in the developing brain and provide potential genetic explanation of currently unknown NDD disease phenotype. The research will be conducted at the HudsonAlpha Institute for Biotechnology, which allows developing scientists to hone skills in a dynamic, hands-on environment with supplemental training courses as necessary. Besides the research outlined in the proposal, training will include courses in conduct of responsible research, advanced biostatistics, machine learning, bioinformatics, and wet lab techniques next to personal development to improve oral and written communication and presentation skills as well as time management.

项目总结/摘要 每100名新生儿中就有1名患有神经发育障碍（NDD）， 一系列使人衰弱的症状，包括早发性癫痫、发育迟缓和智力残疾。 应用全外显子组/基因组测序来检测NDD已经揭示了许多致病基因， 变异，但在很大比例的情况下，遗传原因仍然无法确定。分析 当编码变异不涉及疾病时，深度内含子非编码变异提供了诊断。 例如，SCN 1A的“20 N”毒物外显子中的内含子变体导致功能丧失，引起Dravet 综合征，一种NDD，在生命的第一年引起长时间的癫痫发作。毒外显子是基因的一种机制 由包含毒物外显子引起的提前终止密码子诱导无义的调节， 介导的mRNA转录物的衰变。初步分析表明，在大肠杆菌中， SCN 1A和SCN 8A在胚胎脑发育期间减少，与总RNA成反比 表情大脑是早期神经元发育过程中选择性剪接的温床，因此， 新的毒性外显子仍然未被表征，这是合理的，如果不是可能的话。这项提议的总体假设是， PEI是一种在早期神经元发育过程中差异调节基因表达的机制， 影响发育重要基因中PEI的内含子变异导致疾病 表型在目标1中，将评估钠和钙电压门控通道α亚单位家族， 检测和表征新的毒性外显子。然后将在开发过程中确定已知的和新颖的PEI。 使用公开的RNA-seq实验。在目标2中，这一分析将扩大100倍以上， 检测和评估PEI在3000多个基因与NDD和自闭症谱系障碍（ASD）， 将评估大脑发育和PEI进化的驱动力。毒物外显子的比较分析 将使用k-means分层聚类和无监督随机森林机器学习进行 确定驱动PEI利用的主要生物学因素的方法。在Aim 3中，内含子变体将是 在已知和假设PEI区域的NDD和ASD先证者中分析，以及这些变体的影响 将通过计算评估疾病表型。这些实验将揭示新的见解 选择性剪接在发育中的大脑，并提供潜在的遗传解释，目前未知的NDD 疾病表型这项研究将在哈德逊阿尔法生物技术研究所进行， 培养科学家在充满活力的实践环境中磨练技能，并提供补充培训课程， 必要除了建议中概述的研究外，培训还将包括负责任地开展 研究，先进的生物统计学，机器学习，生物信息学和湿实验室技术旁边的个人 发展，以提高口头和书面沟通和演示技巧以及时间管理。