Regulation of cardiac morphogenesis by Nkx genes in the zebrafish embryo

描述（由申请人提供）： 主要研究者寻求在指导环境中进行高级培训，以研究斑马鱼胚胎中Nkx基因对心脏形态发生的调节。在Deborah Yelon博士的实验室中，在哥伦比亚大学儿科心脏病学分部的支持下，主要目标是为主要研究者提供一个环境，以进行正式课程和实验室研究的培训，这将促进她发展成为一名独立的医生-科学家。主要研究者将使用斑马鱼模型研究心腔形成缺陷，作为了解先天性心脏病（CHD）遗传调控的一种手段。NKX 2 -5是人类CHD的关键致病基因，在小鼠正常心室形成中起关键作用。然而，对NKX 2 -5调控的潜在细胞和分子机制的理解是非常有限的。我们的初步研究表明，斑马鱼NKX 2 -5同源物nkx2.5和nkx2.7的功能丧失会损害心室和心房心腔的形态发生。此外，我们发现这些胚胎中异常心脏形态发生的第一个迹象出现在心脏的初始组装过程中：具体地说，nkx2.5和nkx2.7基因功能的抑制导致心管异常短而宽的心室部分的形成。我们假设nkx基因是驱动心管组装早期阶段的细胞过程的关键调节因子。为了验证这一假设，我们提出了三个具体目标：1。描述nkx基因调控发育中心室延长的细胞机制。通过nkx缺陷胚胎的表型特征，我们将测试的假设，即nkx基因指导特定方面的增殖，细胞形状的变化，细胞运动，和/或细胞极性在心室组装的过程中。2.明确nkx基因在心房形态发生中的作用。在确定了nkx基因对心室形态发生的影响后，我们将使用类似的实验策略来检验nkx基因在心房组装过程中发挥平行作用的假设。3.确定nkx基因调控心管组装的下游效应基因。使用微阵列分析和随后的验证，注释和靶基因的功能丧失分析，我们将确定在心脏的初始组装过程中发挥重要作用的nkx通路的组成部分。总之，这些研究将为CHD的细胞过程和分子机制提供新的线索。从长远来看，这一认识可能为我们提供机会，以产生更好的预后信息，改善复发风险的预测，并可能在冠心病家庭中预防疾病。 (End摘要）

DESCRIPTION (provided by applicant): The Principal Investigator seeks advanced training in a mentored environment to study the regulation of cardiac morphogenesis by Nkx genes in the zebrafish embryo. In the laboratory of Dr. Deborah Yelon and with the support of the Division of Pediatric Cardiology at Columbia University, the primary objective is to provide the Principal Investigator with an environment to pursue training in formal courses and laboratory research that will enhance her development into an independent physician-scientist. The Principal Investigator will use the zebrafish model to study defects in cardiac chamber formation as a means to understand the genetic regulation of congenital heart disease (CHD). NKX2-5 is a key causative gene in human CHD and plays a critical role in normal chamber formation in mice. However, comprehension of the underlying cellular and molecular mechanisms regulated by NKX2-5 is extremely limited. Our preliminary studies suggest that loss-of-function of the zebrafish NKX2-5 homologs, nkx2.5 and nkx2.7, impairs morphogenesis of the ventricular and atrial cardiac chambers. Furthermore, we find that the first sign of abnormal cardiac morphogenesis in these embryos appears during the initial assembly of the heart: specifically, the inhibition of nkx2.5 and nkx2.7 gene function leads to the formation of an abnormally short and wide ventricular portion of the heart tube. We hypothesize that nkx genes are critical regulators of the cellular processes that drive the early stages of heart tube assembly. To test this hypothesis, we propose three specific aims: 1. To characterize the cellular mechanisms by which nkx genes regulate elongation of the developing ventricle. Through phenotypic characterization of nkx-deficient embryos, we will test the hypothesis that nkx genes guide specific aspects of proliferation, cell shape change, cell movement, and/or cell polarity during the process of ventricle assembly. 2. To define the cellular role that nkx genes play in atrial morphogenesis. Having determined the impact of nkx genes on ventricular morphogenesis, we will use similar experimental strategies to test the hypothesis that nkx genes play a parallel role during atrial assembly. 3. To identify the downstream effector genes through which nkx genes regulate heart tube assembly. Using microarray analysis and subsequent validation, annotation, and loss-of-function analysis of target genes, we will identify the components of the nkx pathway that play essential roles during the initial assembly of the heart. Together, these studies will shed new light on the cellular processes and molecular mechanisms underlying CHD. In the long term, this insight may provide us with the opportunity to generate better prognostic information, improved prediction of recurrence risk, and possible disease prevention within families with CHD. (End of Abstract)