INDENTIFICATION OF SIGNALING MOLECULES THAT INDUCE HEART FORMATION

（摘自申请人摘要）本项目的主要目标是 了解心肌细胞分化程序是如何被激活的 在脊椎动物的发育过程中。在以前的工作中，研究人员 在前外侧板发现了心脏诱发活动 原肠期鸟类胚胎的内胚层，它可以诱导细胞从 后原条（通常会产生额外的- 胚胎组织和血液）产生心肌细胞。 最近，研究人员扩大了这些研究，以证明 除了来自前内胚层的心脏诱导信号之外， 骨形态发生蛋白（BMPs）在内胚层和外胚层均表达 在心脏诱导中也起重要作用。结果提示 脊椎动物心肌细胞的确定至少有两个阶段 从原始条纹中出现的细胞仍然是可塑的， 形成心脏的能力。那些中胚层细胞接触前 内胚层似乎位于发育区域，在该区域中细胞显示出 成为心脏的潜力。BMP-2的发育领域， 显示出成为心脏的潜力然后，BMP-2和/或BMP-4似乎是 能够诱导该区域内的细胞启动心脏 差异化方案因此，两个不同信号的组合是 诱导心肌细胞所必需的：BMP信号存在于 胚胎的侧部区域与一个尚未形成的 鉴定了由前内胚层分泌的诱导心脏收缩的信号 阵这项提案的目标是识别信号 由前内胚层细胞分泌，诱导 心肌细胞与BMP信号的协同作用。 采用消减杂交技术， 最近发现了一种信号分子，称为“新月”，这是一种 假定的Wnt抑制剂，并特异性表达于前额叶， 当鸡的心肌细胞被诱导时， 胚胎在鸡的早期发育过程中，crescent在一个区域中表达， 与内胚层心脏诱发的 活动谎言在这项提案中，调查人员打算首先 研究新月形是心脏在前部的诱导活动 内胚层如果是这样的话，调查人员将重点关注 了解这种情况是如何发生的，调查人员将重点关注 了解这个信号分子如何与 在各种测定系统中用BMP信号诱导心脏形成 他们将采用一种替代方法 分离前内胚层中心脏诱导活动的方法。 特异性目标1将研究crescent，一种假定的抗Wnt 由前内胚层分泌的分子与BMP信号合作 以诱导心肌细胞的形成。如果新月形，特定目标2 可以与BMP协同诱导心肌细胞的形成， 信号，调查人员将确定新月是否显示出 抗Wnt活性。具体目标3将确定是否 缺乏新月体基因工程的小鼠显示心脏缺陷 阵具体目标4，如果调查人员发现新月公司未能 在体外或体内显示任何心脏诱导活性，它们将 开发一个功能筛选，以确定心脏诱导 由前内胚层分泌的分子。

(Adapted from the Applicant's Abstract) The broad aim of this project is to understand how the cardiac myocyte differentiation program is activated during vertebrate development. In previous work the investigators have identified a cardiac-inducing activity in the anterior lateral plate endoderm of the gastrula stage avian embryos, which can induce cells from the posterior primitive streak (which normally gives rise to extra- embryonic tissues and to blood) to give rise to cardiac myocytes. Recently, the investigators have extended these studies to demonstrate that in addition to a cardiac inducing signal from the anterior endoderm, Bone Morphogenetic Proteins (BMPs) expressed in both endoderm and ectoderm also play an important role in heart induction. The result suggest that there are at least two stages in vertebrate cardiac myocyte determination Cells emerging from the primitive streak are still plastic with respect to their ability to form heart. Those mesodermal cells which contact anterior endoderm seem to lie in a developmental field in which cells display a potential to become heart. BMP-2 developmental field in which cells display a potential to become heart. BMP-2 and/or BMP-4 then appear to be capable of inducing cells within this field to initiated the cardiac differentiation program. Thus, a combination of two distinct signals is necessary for the induction of cardiac myocytes: a BMP signal present in the lateral regions of the embryo collaborates with an as yet to be identified signal secreted by the anterior endoderm to induce heart formation. The goal of this proposal will be to identify the signal secreted by cells in the anterior endoderm that induces the formation of cardiac myocytes in collaboration with a BMP signal. By employing subtractive hybridization technology the investigations have recently identified a signaling molecule, termed "crescent", which is a putative Wnt inhibitor, and is specifically expressed in the anterior endoderm at the time when cardiac myocytes are being induced in the chick embryos. During early chick development, crescent is expressed in a domain that precisely overlaps the location where the endodermal heart inducing activity lies. In this proposal the investigators intend to firstly investigate where crescent is the heart inducing activity in the anterior endoderm. If such is the case, the investigators will focus on understanding how this such is the case, the investigators will focus on understanding on understanding how this signaling molecule collaborates with BMP signals to induce heart formation in the various assay systems that the investigators have developed, they will employ an alternative approach to isolate the heart inducing activity in the anterior endoderm. Specific Aim 1 will investigate whether crescent, a putative anti-Wnt molecule secreted by the anterior endoderm collaborates with BMP signals to induce the formation of cardiac myocytes. Specific Aim 2 if crescent can induce the formation of cardiac myocytes in collaboration with a BMP signal, the investigators will determine whether crescent displays an anti-Wnt activity in this induction. Specific Aim 3 will determine whether mice genetically engineered to lack crescent display a defect in heart formation. Specific Aim 4 if the investigators find that crescent fails to display any heart inducing activity either in vitro or in vivo, they will develop develop a functional screen to identify the cardiac inducing molecule secreted by the anterior endoderm.