Molecular mechanisms of atrial development and regeneration

心房发育和再生的分子机制

基本信息

批准号：
9363356
负责人：
Joshua Waxman
金额：
$ 39.75万
依托单位：
CINCINNATI CHILDRENS HOSP MED CTR
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：
2017-07-21 至 2021-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9363356
关键词：
3&apos Untranslated Regions ATAC-seq Ablation Adult Affect Affinity Chromatography Antisense RNA Arrhythmia Biological Assay Birth CRISPR/Cas technology Cardiac Cardiac Myocytes Cardiovascular Diseases Cardiovascular system Cell Lineage Cell Transplantation Cells Cessation of life Child Chromatin Complex Congenital Abnormality Congenital Heart Defects Data Defect Development Diagnosis Disease Embryo Engineering Epigenetic Process Ethylnitrosourea Etiology Foundations Gene Expression Regulation Generations Genetic Complementation Test Genomics Glean Goals Health Heart Heart Atrium Heart Hypertrophy Heart Injuries Human Impairment In Vitro Individual Knockout Mice Knowledge Life Methods MicroRNAs Modeling Molecular Morphology Mus Mutagenesis Mutation Natural regeneration Neonatal Mortality Newborn Infant Nuclear Hormone Receptors Nuclear Orphan Receptor Orphan Pathway interactions Play Positioning Attribute Prevalence Production Proteins RNA Ribosomes Role Signal Transduction Site Stroke Surface Techniques Testing Tissues Transcript Transcriptional Activation Translations Transposase Untranslated RNA Ventricular Vertebrates Zebrafish apoAI regulatory protein-1 atrioventricular septal defect blastocyst cardiac regeneration cardiogenesis genome editing healing human stem cells improved in utero diagnosis in vivo inhibitor/antagonist injured loss of function mutant next generation sequencing novel novel therapeutics premature prevent progenitor promoter regenerative repaired transcription factor transcriptome sequencing

项目摘要

Project Summary/Abstract Congenital heart defects (CHDs) are the most common congenital malformations. 5% of CHDs comprise atrioventricular septal defects (AVSDs). However, the molecular etiology underlying most AVSDs are not understood. Furthermore, CHDs can cause cardiovascular diseases later in life, resulting in arrhythmias, stroke, and premature death. In order to develop novel therapies able to prevent CHDs and heal specific cardiovascular tissues, it is critical to garner understanding of fundamental mechanisms directing normal cardiac chamber development and regeneration. Therefore, long-term goals of our lab are to understand conserved mechanisms that direct the development of the individual cardiac chambers and chamber-specific mechanisms of regeneration in vertebrates. Few signals are known to be required that specifically direct atrial development, with specific regulators of atrial regeneration not being understood. The specific aims of this proposal are to elucidate the mechanisms by which a syntenic long non-coding RNA (lncRNA) as-oca restricts atrial development and regeneration through inhibition of Nr2f1a translation in zebrafish. The studies in this proposal are relevant to human health as recent genomic analysis indicates that mutations in the orphan nuclear receptor Nr2f2 are associated with AVSDs in humans. While Nr2f2 knockout mice and in vitro studies with human stem cells have revealed requirements for both Nr2f1 and Nr2f2 in atrial development, the mechanisms by which Nr2f proteins direct proper atrial development are not completely understood. Importantly, there is currently no understanding of epigenetic lncRNA-dependent mechanisms regulating Nr2f proteins. Our preliminary analysis of the novel zebrafish mutant acorn worm (aco) indicate that excess expression of as-oca specifically restricts the addition of later differentiating second heart field (SHF)-derived atrial cells. In Aim 1, we will use blastula cell transplantation, in vivo cardiomyocyte differentiation assays, and genome editing to determine the cellular requirements underlying the atrial defects and cause of increased as- oca expression in aco mutants. We do not understand how as-oca inhibits Nr2f1a translation in aco mutants. In Aim 2, we will use RNA and ribosomal association techniques and loss of function methods to determine if as- oca inhibits Nr2f1a translation through interactions with the nr2f1a 3' untranslated region. In addition to their requirements during development, we find that as-oca and nr2f1a are specifically expressed in the atria of adult zebrafish. In Aim 3, we will test the effects of as-oca on embryonic and adult models of cardiac regeneration. Because Nr2f transcription factors play conserved roles in atrial development of all vertebrates, these studies will dramatically improve our understanding of posttranscriptional mechanisms regulating normal vertebrate atrial development and the molecular etiology of AVSDs and ASDs in humans. Ultimately, these studies will generate a foundation for improved therapies capable of preventing CHDs in children and efficiently repairing injured hearts in adults.

项目概要/摘要先天性心脏缺陷（CHD）是最常见的先天畸形。 5% 的先心病包括房室间隔缺损（AVSD）。然而，大多数 AVSD 的分子病因学并不明白了。此外，先心病可在以后的生活中引起心血管疾病，导致心律失常，中风和过早死亡。为了开发能够预防冠心病和治愈特定疾病的新疗法心血管组织，了解指导正常的基本机制至关重要心腔的发育和再生。因此，我们实验室的长期目标是了解指导各个心腔和腔室特异性发育的保守机制脊椎动物的再生机制。已知很少有信号需要专门引导心房心房再生的具体调节因素尚不清楚。本次活动的具体目标提案旨在阐明同线性长非编码RNA (lncRNA) as-oca 限制的机制通过抑制斑马鱼 Nr2f1a 翻译来促进心房发育和再生。这方面的研究该提案与人类健康相关，因为最近的基因组分析表明孤儿中的突变核受体 Nr2f2 与人类 AVSD 相关。而Nr2f2敲除小鼠及体外研究与人类干细胞合作揭示了心房发育中 Nr2f1 和 Nr2f2 的需求， Nr2f 蛋白指导心房正常发育的机制尚不完全清楚。重要的是，目前对 Nr2f 的表观遗传 lncRNA 依赖机制尚不了解蛋白质。我们对新型斑马鱼突变橡子虫（aco）的初步分析表明，过量 as-oca 的表达特别限制了后来分化的第二心区 (SHF) 衍生的细胞的添加心房细胞。在目标 1 中，我们将使用囊胚细胞移植、体内心肌细胞分化测定，以及基因组编辑以确定心房缺陷背后的细胞需求以及心房缺陷增加的原因 aco突变体中oca的表达。我们不明白 as-oca 如何抑制 aco 突变体中的 Nr2f1a 翻译。在目标 2，我们将使用 RNA 和核糖体关联技术以及功能丧失方法来确定是否- oca 通过与 nr2f1a 3' 非翻译区相互作用抑制 Nr2f1a 翻译。除了他们的在开发过程中，我们发现as-oca和nr2f1a具体表达在心房成年斑马鱼。在目标 3 中，我们将测试 as-oca 对胚胎和成人心脏病模型的影响再生。由于 Nr2f 转录因子在所有脊椎动物的心房发育中发挥保守作用，这些研究将极大地提高我们对调节正常转录后机制的理解脊椎动物心房发育以及人类 AVSD 和 ASD 的分子病因学。最终，这些研究将为改进能够有效预防儿童先心病的疗法奠定基础修复成人受伤的心脏。