Characterizing the role of MINK1 in Congenital Heart Disease and mucociliary clearance

表征 MINK1 在先天性心脏病和粘膜纤毛清除中的作用

• 批准号: 9918965
• 负责人: Vaughn Colleluori
• 金额: $ 3.21万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9918965
• 关键词: Affect; Bilateral; Biological Assay; Biological Models; C-terminal; Candidate Disease Gene; Cardiac; Cardiac development; Cells; Child Health; Cilia; Clustered Regularly Interspaced Short Palindromic Repeats; Congenital Abnormality; Data; Defect; Development; Disease; Disease model; Docking; Dominant-Negative Mutation; Drosophila eye; Embryo; Embryonic Development; Epidermis; Etiology; Future; Genes; Genetic Counseling; Genomics; Genotype; Goals; Homodimerization; Infant Mortality; Knock-out; Lateral; Lead; Left; Literature; Live Birth; Lung; MAPK8 gene; Mentorship; Mesoderm; Missense Mutation; Modeling; Molecular; Mucociliary Clearance; Mutation; Newborn Infant; Nodal; Organ; Pathogenesis; Patients; Pattern; Phenotype; Phosphotransferases; Play; Proteins; Rana; Regulation; Research; Research Personnel; Research Training; Respiratory System; Role; Scientist; Serine; Side; Signal Transduction; Situs Inversus; Structure; Surface; Testing; Threonine; Time; Training; Variant; WNT Signaling Pathway; Wing; Writing; Xenopus; blastomere structure; cardiogenesis; career; cell fate specification; cell motility; cilium biogenesis; cilium motility; congenital heart disorder; disease phenotype; epidermis cell; experimental study; extracellular; fluid flow; gain of function; genetic analysis; genetic testing; germinal center kinases; human disease; improved; improved outcome; kinetosome; loss of function; molecular marker; notch protein; novel; overexpression; respiratory; respiratory toxin; skill acquisition; technique development; training opportunity

Project Summary/ Abstract Congenital Heart Disease (CHD) is the most common birth defect affecting approximately 1% of all live births in the US and is one of the leading causes of infant mortality. A severe form of CHD can result from Heterotaxy (Htx), a disorder of the left-right (LR) patterning during embryonic development. A recent genetic analysis of heterotaxy patients identified a novel CHD candidate gene, mink1. Mink1 encodes a serine-threonine germinal-center kinase with known functions in the JNK and PCP/Wnt signaling pathways. However, it has no known role in LR patterning or cardiac development. Using CRISPR knockout strategies in the high-throughput human disease model, Xenopus, loss of mink1 leads to cardiac and LR patterning defects, and defects in motile cilia formation and resulting fluid flow. The overall goal of this proposal is to investigate the molecular mechanism by which mink1 affects LR patterning, heart development, and cilia formation in the Xenopus (frog) model system. The first aim will use loss of function experiments to determine the required role for mink1 during the LR patterning cascade b testing molecular markers. Mechanistic hypotheses will be guided by an analysis of temporal and spatial expression of mink1 in the whole embryo and the left-right organizer. The second aim will use loss of function experiments to determine the requirement for mink1 during formation of motile cilia on the multi-ciliated cells of the Xenopus epidermis, which recapitulates a common mucociliary defect found in CHD patients. Hypotheses will be directed by a review of relevant literature including a role in regulation of multi-ciliated cell fate specification through Notch signaling or a requirement for Mink1 during basal body docking and/or establishment of polarity. The third aim will determine the role of each mink1 domain in the context of cilia formation on multi-ciliated cells and LR patterning/cardiac development. Then it will be determined if the patient mutation in the kinase-encoding domain is determination to function using multiple functional assays in Xenopus. Altogether, these experiments will improve our understanding of cardiac development and the role of mink1 in the pathogenesis of CHD. 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 applicant's 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 the applicant to pursue a career performing patient-driven research as an independent research scientist.

项目总结/摘要 先天性心脏病（CHD）是最常见的出生缺陷，约占所有活产婴儿的1 在美国，它是婴儿死亡的主要原因之一。严重的冠心病可由心脏异位引起 (Htx)胚胎发育过程中左右（LR）模式的紊乱。最近的一项基因分析表明， 异位症患者确定了一个新的CHD候选基因mink 1。Mink 1编码丝氨酸-苏氨酸 在JNK和PCP/Wnt信号通路中具有已知功能的生殖中心激酶。然而，它没有 已知在LR模式或心脏发育中的作用。使用CRISPR敲除策略在高通量 人类疾病模型，非洲爪蟾，水貂1的损失导致心脏和LR图案缺陷， 运动纤毛的形成和由此产生的流体流动。本提案的总体目标是调查 mink 1影响LR模式、心脏发育和纤毛形成的分子机制， 非洲爪蟾（青蛙）模型系统。第一个目标将使用功能丧失实验来确定 mink 1在LR模式级联B测试分子标记物期间所需的作用。机制假说 将通过分析mink 1在整个胚胎和左右胚胎中的时间和空间表达来指导 主办方第二个目标将使用功能丧失实验来确定在 在爪蟾表皮的多纤毛细胞上形成活动纤毛，它概括了一种常见的 冠心病患者存在黏膜纤毛缺陷。假设将通过相关文献的回顾来指导 包括通过Notch信号传导调节多纤毛细胞命运特化的作用或 Mink 1在基体对接和/或极性建立期间。第三个目标将决定每个人的作用 mink 1结构域在多纤毛细胞纤毛形成和LR模式/心脏发育中的作用。 然后将确定患者激酶编码域中的突变是否是功能决定 在非洲爪蟾中进行多功能检测。总之，这些实验将提高我们对 心脏发育和mink 1在冠心病发病机制中的作用。在未来，这将有利于遗传 测试和咨询，以及改善冠心病的结果，因为治疗可以根据基因型 而不仅仅是CHD表型。此外，本申请书亦详列申请人的培训计划，包括 研究指导，高级课程，新技术培训，以及 科学的专业精神，写作和数据的呈现。本报告中概述的研究和培训 申请将准备申请人追求的职业生涯执行病人驱动的研究作为一个独立的 研究科学家