Regulation of Mesodermal Progenitors in Transgenic Zebrafish

描述（由申请人提供）：早期脊椎动物发育的一个标志是胚胎体从前部（A）到后部（P）的渐进生长，这依赖于位于胚胎最后端称为尾芽的区域中的多能干细胞样祖细胞群。这个祖细胞群逐渐释放中胚层细胞，这些细胞填充体节（主要是肌肉），以及形成脊髓的神经细胞，直到建立完整的A-P轴。为了使身体正常形成，必须仔细控制从尾芽释放的速率和细胞增殖，以便沿着整个A-P轴产生正确比例的细胞。人们对这些过程是如何调节和整合的知之甚少。本研究的第一个目的是研究细胞离开尾芽进入体节的机制。基于最近的初步结果与一个新的转基因株系，Wnt信号在调节这一过程中的作用将阐明通过测试Wnt功能的两个假设。此外，分析tbx 16/spadetail启动子内的一个小的独特元件，该元件在细胞决定离开祖细胞群体时被激活，这将为了解调节身体伸长时中胚层命运承诺的初始步骤的分子机制提供关键见解。确定控制细胞分配的机制将是理解脊椎动物胚胎如何精确调节尾芽细胞释放的重要一步。第二个目标将确定为什么细胞增殖在原肠胚后胚胎中受到严格调控，使得祖细胞是静止的，并且仅在它们第一次开始分化时才分裂。这一目标将测试的假设，这种调节是必不可少的，让正常的信号和/或形态发生的中胚层祖细胞及其衍生物，使用一种新的转基因系，我们最近生产的。确定为什么脊椎动物胚胎严格控制增殖是必不可少的了解胚胎如何调节竞争的需要，以增加细胞数量，但保持复杂的信号和形态发生过程，建立胚胎体计划。特别是在哺乳动物中的研究表明，后祖细胞是干细胞样群体，其有助于形成多种细胞类型。由于能够产生表达信号传导和细胞增殖的时间控制调节因子的转基因系，以及能够容易地敲除基因功能，斑马鱼为理解脊椎动物干细胞如何在体内调节提供了一个极好的模型系统。由于干细胞在治疗许多疾病方面具有巨大的前景，本文所述的研究将提供有关控制干细胞维持和组织形成的信号网络和调节因子的有价值的信息。

DESCRIPTION (provided by applicant): A hallmark of early vertebrate development is the progressive growth of the embryonic body from the anterior (A) to the posterior (P), which relies on a multipotent stem-like progenitor population located at the most posterior end of the embryo, in a region called the tailbud. This progenitor population gradually releases mesodermal cells that populate the somites (primarily muscle), as well as neural cells that form the spinal cord, until the complete A-P axis has been established. For the body to form normally, both the rate of release from the tailbud and cell proliferation must be carefully controlled so tht the correct proportion of cells is produced along the entire A-P axis. How these processes are regulated and integrated is poorly understood. The first aim of this proposal will examine the mechanisms by which cells leave the tailbud and enter the somites. Based on recent preliminary results with a new transgenic line, the role of Wnt signaling in regulating this process will be elucidated by testing two hypotheses for Wnt function. In addition, analysis of a small unique element within the tbx16/spadetail promoter, which is activated just as cells make the decision to leave the progenitor population, will provide key insight into the molecular mechanism that regulates the initial step in the commitment to the mesodermal fate as the body elongates. Identifying the mechanisms that control cell allocation will be a major step forward in understanding how the vertebrate embryo precisely regulates the release of cells from the tailbud. The second aim will determine why cell proliferation is tightly regulated in the post-gastrula embryo such that the progenitors are quiescent, and only divide when they first begin to differentiate. This aim will test the hypotheses that this regulation is essential to allow normal signaling and/or morphogenesis of the mesodermal progenitors and their derivatives using a novel transgenic line we have recently produced. Determining why the vertebrate embryo strictly controls proliferation is essential for understanding how the embryo regulates the competing needs to increase cell number yet maintain the complex signaling and morphogenetic processes necessary to establish the embryonic body plan. Studies, particularly in mammals, show that the posterior progenitors are a stem-cell like population, which contribute to a variety of cell types. With the ability to produce transgenic lines expressing temporally controlled regulators of signaling and cell proliferation, as well as the ability to easily knock down gene function, zebrafish provides an excellent model system for understanding how vertebrate stem cells are regulated in vivo. As stem cells have great promise for the treatment of many diseases, the studies described here will provide valuable information about the signaling networks and regulatory factors that control stem cell maintenance and tissue formation.