Molecular aspects of CRKL in heart development and human disease

CRKL 在心脏发育和人类疾病中的分子方面

基本信息

批准号：
9197022
负责人：
BERNICE E MORROW
金额：
$ 81万
依托单位：
ALBERT EINSTEIN COLLEGE OF MEDICINE, INC
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-07-15 至 2020-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9197022
关键词：
22q11 Deletion Syndrome 22q11.2 Accounting Adaptor Signaling Protein Affect Anterior Aortic Valve Stenosis Back Base Pairing Biological Assay Boxing CRKL gene Candidate Disease Gene Cardiac Cell physiology Cells Cessation of life Chromosomes Clinical Code Complex Congenital Heart Defects DNA Data Defect Development DiGeorge Syndrome Disease Embryo Endocardium Etiology Evaluation Focal Adhesion Kinase 1 Genes Genetic Genetic Predisposition to Disease Genetic Transcription Heart Heart Abnormalities Heart Valves Human Human Genetics In Vitro Individual Intracardiac abnormalities Investigation MAP Kinase Gene Mediating Mesenchymal Molecular Mus Mutant Strains Mice Mutation Neural Crest Cell Outcome Pathway interactions Patients Persistent Truncus Arteriosus Pharyngeal Apparatus Phenotype Population Process Rare Diseases Receptor Protein-Tyrosine Kinases Regulatory Element Reporting Research Risk Sampling Septate Shprintzen syndrome Signal Pathway Signal Transduction System Testing Tetralogy of Fallot Tissues Untranslated RNA Validation Variant Vascular Endothelial Growth Factors Work base cardiogenesis cohort congenital heart disorder conotruncal anomaly face syndrome differential expression disease heterogeneity embryo culture exome sequencing gene function gene interaction genetic approach genome sequencing human disease in vitro Assay in vivo insight loss of function malformation middle age mouse model mutant novel paracrine proband programs transcription factor transcriptome sequencing whole genome

项目摘要

ABSTRACT One of the greatest challenges in human genetics is to understand the genetic basis of complex disorders, such as congenital heart disease (CHD). Studies of rare diseases with known genetic etiologies such as the 22q11.2 deletion syndrome (DiGeorge/velo-cardio-facial syndrome), can pinpoint disease genes and mechanisms. Approximately 65% of 22q11DS patients have a congenital heart defect, mostly affecting the cardiac outflow tract (OFT). These range from mild to severe and include persistent truncus arteriosus and tetralogy of Fallot, at the severe end. Although TBX1, encoding a T-box transcription factor has been the strongest candidate gene for CHD on chromosome 22q11.2, patients with similar heart defects, but non- overlapping deletions within this region have been identified. In these patients with atypical deletions, CRKL, encoding a cytoplasmic adaptor to receptor tyrosine kinase signaling is deleted, but not TBX1. This implicates CRKL as a true disease gene. Although there have been extensive studies of TBX1, CRKL is understudied with respect to heart development. The neural crest cells are a critical cell population in the pharyngeal apparatus needed for proper alignment of the cardiac OFT. We propose to explore neural crest cell functions of Crkl by evaluating new upstream and downstream pathways in Aim 1. We will then delineate the subset that is critical for OFT development as mediated by Tbx1 and Crkl. Crkl-/- mice die at E16.5, earlier than can be expected based upon only having OFT defects. Our pathological investigation has found that Crkl-/- embryos have OFT valve defects in addition to alignment malformations. OFT valve anomalies are highly relevant to 22q11DS, as 9% of patients have these defects. This is a newly recognized defect for which the molecular mechanisms are not known. Based upon this, and the presence of similar defects in endocardial-specific null mutant embryos, we hypothesize that Crkl is required in this tissue to form derivative OFT valves via VEGF- MAPK and additional signaling pathways. Endocardial functions will be tested in Aim 2. This work is also clinically important because middle-aged 22q11DS patients have now been reported to have sudden unexplained death, unconnected to the well-known intracardiac malformations. This might be related to the presence of aortic valve stenosis, which accounts for 6.6% of 22q11.2 deletion patients. To connect the mouse work in Aims 1 and 2 back to the human patients, we will evaluate existing whole exome sequence (WES) from the current largest cohort that we are aware, of >1,000, 22q11DS patients for mutations in CRKL interaction network genes in Aim 3. Exploratory studies of non-coding sequences in the candidate genes will be performed. Validation will be done using more DNA samples from 500 22q11DS subjects and from emerging WES data on non-syndromic CHD patients. The synergy of mouse and human studies will offer rapid discoveries for a deep understanding of the complex genetic etiology of CHD.

抽象的人类遗传学中最大的挑战之一是了解复杂疾病的遗传基础，例如先天性心脏病（CHD）。对具有已知遗传病因的罕见疾病的研究，例如 22q11.2 缺失综合征（DiGeorge/velo-cardio-facial 综合征），可以查明疾病基因和机制。大约 65% 的 22q11DS 患者患有先天性心脏病，主要影响心脏功能心脏流出道（OFT）。这些范围从轻微到严重，包括持续性动脉干和法洛四联症，处于严重的末期。尽管编码 T 盒转录因子的 TBX1 一直是染色体 22q11.2 上最强的 CHD 候选基因，患有类似心脏缺陷但非已确定该区域内的重叠删除。在这些具有非典型缺失的患者中，CRKL，编码受体酪氨酸激酶信号传导的细胞质接头被删除，但 TBX1 未被删除。这意味着 CRKL作为真正的疾病基因。尽管对 TBX1 进行了广泛的研究，但 CRKL 的研究还不够关于心脏发育。神经嵴细胞是咽部的重要细胞群正确对准心脏 OFT 所需的装置。我们建议探索神经嵴细胞功能通过评估目标 1 中新的上游和下游路径来评估 Crkl。然后我们将描述子集对于由 Tbx1 和 Crkl 介导的 OFT 发育至关重要。 Crkl-/- 小鼠在 E16.5 时死亡，早于可能的死亡时间仅基于 OFT 缺陷进行预期。我们的病理学调查发现Crkl-/-胚胎除对准畸形外，还存在 OFT 瓣膜缺陷。 OFT 阀门异常与 22q11DS，9% 的患者有这些缺陷。这是一种新发现的缺陷，分子机制尚不清楚。基于此，以及心内膜特异性无效中类似缺陷的存在突变胚胎，我们假设该组织需要 Crkl 通过 VEGF-形成衍生的 OFT 瓣膜 MAPK 和其他信号通路。目标2将测试心内膜功能。这项工作也是临床上很重要，因为据报道中年 22q11DS 患者突然出现不明原因死亡，与众所周知的心内畸形无关。这可能与存在主动脉瓣狭窄，占 22q11.2 缺失患者的 6.6%。连接鼠标目标 1 和 2 的工作回到人类患者，我们将评估现有的全外显子序列 (WES) 目前我们所知的最大队列，有超过 1,000 名 22q11DS 患者出现 CRKL 相互作用突变目标3中的网络基因。将对候选基因中的非编码序列进行探索性研究执行。将使用来自 500 名 22q11DS 受试者和新兴市场的更多 DNA 样本进行验证。非综合征性 CHD 患者的 WES 数据。小鼠和人类研究的协同作用将提供快速深入了解冠心病复杂的遗传病因学的发现。