Elucidating the function of the CHD gene, RAPGEF5, in Wnt signaling

阐明 CHD 基因 RAPGEF5 在 Wnt 信号传导中的功能

• 批准号: 9760814
• 负责人: Woong Hwang
• 金额: $ 2.98万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9760814
• 关键词: Adult; Affect; Affinity; Animal Model; Binding; Biological Assay; Candidate Disease Gene; Cardiac; Cardiac development; Cardiovascular Diseases; Cardiovascular Models; Cell Nucleus; Congenital Abnormality; Cytoplasm; Data; Defect; Developmental Biology; Diagnosis; Diffuse; Disease; Embryo; Embryonic Development; Exhibits; Fluorescence; Foundations; Future; GTP Binding; Gene Activation; Genes; Genetic; Genetic Counseling; Genetic Research; Genetic screening method; Genomics; Genotype; Goals; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Health; Heart; Heart Abnormalities; Heart failure; Homeostasis; Human; Human Cell Line; Importins; In Vitro; Infant Mortality; Kinetics; Knowledge; Lead; Left; Ligation; Live Birth; Malignant Neoplasms; Measures; Mediating; Membrane; Mentorship; Modeling; Molecular; Morbidity - disease rate; Newborn Infant; Nuclear; Nuclear Import; Nuclear Localization Signal; Nuclear Translocation; Operative Surgical Procedures; Pathogenesis; Pathway interactions; Patients; Pattern; Phenotype; Physicians; Play; Process; Proteins; Recovery; Research; Research Training; Role; Scientist; Situs Inversus; System; Testing; Tissues; Training; WNT Signaling Pathway; Work; Writing; Xenopus; base; beta catenin; career; congenital heart disorder; disease phenotype; experience; experimental study; genetic approach; improved; improved outcome; infant morbidity; mortality; mutant; novel; nucleocytoplasmic transport; skill acquisition; stem; technique development

Project Summary/ Abstract Congenital heart disease (CHD) is one of the leading causes of infant mortality and morbidity affecting 8 out of every 1000 newborns in the US. Despite this massive health impact, the genetic causes of this disease are not well understood. In order to better understand CHD, we, along with other labs, are analyzing the genomics of CHD patients. In particular, we examined heterotaxy, a disorder of left-right (LR) patterning that can lead to severe form of cardiac malformation due to failure of cardiac looping. From this genetic approach, we identified RAPGEF5 as a novel candidate gene for heterotaxy and congenital heart disease, which encodes a guanine nucleotide exchange factor that activates RAP GTPase(s). We then modeled this cardiovascular disease in Xenopus by depleting Rapgef5, which recapitulated the human heterotaxy phenotype and demonstrated that rapgef5 affects cardiac looping via regulating the nuclear entry of β-catenin in Wnt signaling. The main goal of this proposal is to investigate the molecular mechanism by which rapgef5 controls the nuclear localization of β-catenin. Of note, the mechanism of β-catenin nuclear translocation remains an outstanding question in Wnt signaling, that has implications for developmental biology and cancer. In this proposal, I have two specific aims based on the following hypothesis: Hypothesis: Rapgef5 maintains nuclear Rap protein(s) in an active GTP bound state, which will preferentially import β-catenin into the nucleus Aim 1: Kinetic analysis of β-catenin nucleocytoplasmic shuttling in vitro Aim 2: Identify a RAP GTPase that mediates the β-catenin nuclear transport system Altogether, these experiments will improve our understanding of cardiac development and the role of rapgef5 in the pathogenesis of CHD as well as its role in Wnt signaling. In the future, this will benefit genetic testing and counseling, as well as improve outcomes in CHD because treatments can be tailored to genotype rather than solely on CHD phenotype. In addition, this application details the my training plan including research mentorship, advanced coursework, training in new techniques, and the development of skills in scientific professionalism, writing, and presentation of data. The research and training outlined in this application will prepare me to pursue a career performing patient-driven research as an independent physician-scientist.

项目摘要/摘要 先天性心脏病(CHD)是婴儿死亡和发病率的主要原因之一，影响到八分之一的婴儿 在美国，每1000个新生儿。尽管对健康造成了巨大的影响，但这种疾病的遗传原因并不是 很好理解。为了更好地了解CHD，我们与其他实验室一起分析了 冠心病患者。特别是，我们检查了异位排列，这是一种左右(LR)模式的紊乱，可能导致 由于心脏循环失败而导致的严重心脏畸形。从这种遗传方法中，我们识别出 RAPGEF5基因是一种新的异质性和先天性心脏病候选基因，它编码一种鸟嘌呤 激活RAP GTP酶的核苷酸交换因子(S)。然后我们对这种心血管疾病进行了建模 通过耗尽Rapgef5，它概括了人类的异质性表型，并证明了 Rapgef5通过调节β-catenin在Wnt信号中的核进入而影响心脏环路。的主要目标是 这项提议是为了研究Rapgef5控制核的分子机制。 β-连环蛋白的定位。值得注意的是，β-连环蛋白核转位的机制仍然是一个突出的问题。 Wnt信号中的问题，这与发育生物学和癌症有关。 在这项建议中，基于以下假设，我有两个具体目标： 假设：Rapgef5维持核Rap蛋白(S)处于活跃的GTP结合状态，这将优先 将β-连环蛋白导入细胞核 目的1：β-连环蛋白核质穿梭的体外动力学分析 目的2：确定介导β-连环蛋白核运输系统的RAP GTP酶 总之，这些实验将提高我们对心脏发育和Rapgef5作用的理解。 在CHD发病机制中的作用及其在Wnt信号中的作用。在未来，这将有利于基因检测和 咨询，以及改善CHD的结果，因为治疗可以针对基因而不是 仅限于CHD表型。此外，这份申请详细说明了我的培训计划，包括研究 指导、高级课程、新技术培训和科学技能的发展 数据的专业性、写作和展示。本申请中概述的研究和培训将 让我做好准备，作为一名独立的内科科学家从事以患者为导向的研究。