HEART DEVELOPMENT AND GENETICS

这项建议是为了继续我们对心血管发育的研究。 这项工作的长期目标是确定与 心血管系统的形成，这些基因对 它的形式和功能。我们试图通过基因解剖来做到这一点，在其他方面 用突变来定义发展中的重要步骤 心脏，并最终在分子上定义负责基因 水平。我们选择了斑马鱼作为合适的模型系统。 这种小型淡水硬骨鱼很容易饲养，可以产生数百只 每天的大卵子，所有的发育都发生在外部 透明的胚胎。它特别容易受到基因筛选的影响。我们的 重点是心血管系统。在3天内 受精我们已经发现斑马鱼的心脏形成和循环， 变得细分为由阀门隔开的腔，以及 开始产生有效的循环。通过单细胞注射 我们已经开始追踪心脏前体细胞，因为它们出现在 囊胚，并追踪它们在新生心脏内的迁徙和命运 管子。此外，我们还发现了影响心脏的突变。 例如，我们正在研究纯合子缺乏的鱼类品系 任何心跳。这项建议的具体目的是：(1) 描绘心脏的谱系，并决定细胞的命运决定 这导致了它的各种成分。例如，初步的 结果表明，分离的前体细胞的后代填充在 心内膜而不是心肌，以及心房而不是 脑室；(2)产生单克隆标记，除了 我们目前有特定于小室的表位，以确定表位 提供图案化信息，以及识别 心脏祖细胞亚群；(3)继续分子 分析影响心脏的斑马鱼突变，特别关注 无声的心脏突变。我们的初步证据确凿 这表明这种疾病的细胞生物学存在于 细胞膜上的电活动和细胞膜上的 收缩(和赤字是一种“机电” 我们的初步证据表明，我们可能有 确定了分子缺陷，我们认为这是肌钙蛋白T的缺陷， 肌动球蛋白调节复合体的一种成分。我们试图测试这一点 通过分子分析明确提出假说。 心血管疾病的遗传学甚至还知之甚少。 尽管许多心脏疾病都有一个重要的遗传因素。 斑马鱼系统为分子生物学研究提供了一个潜在的模型系统 对这种疾病的解剖。

This proposal is to continue our studies of cardiovascular development. The long-term goal of this work is to identify the genes responsible for fashioning of the cardiovascular system, genes that are critical both to its form and function. We seek to do so by genetic dissection, in other words by using mutations to define the important steps in development of the heart and ultimately to define the responsible genes at the molecular level. We have selected the zebrafish as the appropriate model system. This small fresh water teleost is easily raised, and produces hundreds of large eggs daily, with all development occurring externally in a transparent embryo. It is particularly amenable to genetic screens. Our focus is specifically upon the cardiovascular system. Within 3 days of fertilization we have found that the zebrafish heart forms and loops, becomes subdivided into chambers which are separated by valves, and begins to generate an effective circulation. By single cell injection we have begun to track the cardiac progenitor cells as they arise in the blastula, and to track their migration and fate within the nascent heart tube. In addition, we have identified mutations that affect the heart. For example, we are studying fish strains in which the homozygote lacks any heart beat. The specific aims of this proposal are: (1) To delineate the lineage of the heart, and determine the cell fate decisions which give rise to its various components. For example, preliminary results suggest that the progeny of separate precursor cells populate the endocardium as opposed to myocardium, and the atrium rather than the ventricle; (2) To generate monoclonal markers, in addition to the chamber-specific ones we have currently, in order to identify epitopes that provide patterning information, as well as ones that identify subpopulations of cardiac progenitor cells; (3) To continue the molecular analysis of zebrafish mutations that affect the heart, with special focus upon the silent heart mutation. Our preliminary evidence strongly suggests that the cell biology of this disorder resides at the coupling interface between electrical activity at the cell membrane and contraction (and that the deficit is one of "electromechanical dissociation"). Our preliminary evidence suggests that we may have identified the molecular defect, which we believe to be in troponin T, a component of the actomyosin regulatory complex. We seek to test this hypothesis specifically by molecular analysis. The genetics of cardiovascular disease is only poorly understood, even though many cardiac disorders have an important hereditary component. The zebrafish system provides a potential model system for molecular dissection of such disorders.