Lineage and induction of ventrolateral mesoderm

描述(申请人提供)：在脊椎动物中，许多组织和器官全部或部分来自中胚层，包括骨骼肌、心脏、骨骼和肾脏。一个复杂的基因网络调节着这些不同的中胚层衍生物的渐进规范。在非洲爪哇，中胚层由早期原肠胚赤道周围的一条细胞带形成。在非洲爪蛙中胚层模式的模型中，Spemann组织器被突出地显示出来，并被认为通过分泌诱导因子来指定中胚层命运。我们最近已经证明，Spemann组织者非依赖的途径通过原肠胚中期沿着中胚层的动植物轴建立了一种模式。在这个轴上可以观察到两个不同的区域。动物区表达brachyury基因，似乎注定成为脊索和体节，而植物区注定成为腹外侧型中胚层。这项提议的目标是进一步阐明沿着动物/植物轴建立图案的机制。特别是，我们将研究结节生长因子在腹侧中胚层诱导中的作用。我们的初步结果表明，非洲爪哇结节相关2(Xnr2)的表达细胞仅限于原肠胚胎中假定的腹中胚层。这里提出的谱系追踪实验旨在严格测试这些细胞的命运。最终确定早期推测中胚层内细胞的命运，并将它们与限制性基因表达模式相关联，是阐明命运指定的机制和时机的关键的第一步。我们还假设结节在腹侧中胚层的诱导中起重要作用，我们将利用显性负结构和抑制结合蛋白来检测原肠胚中结节的活性。最后，我们的初步结果表明，Xnr2信号和低活性的成纤维细胞生长因子结合在一起，例如存在于植物边缘地带，使外胚层外植体分化为血液，一种腹侧中胚层衍生体，对骨形态发生蛋白(BMP)做出反应。我们将使用消减和表达克隆方法来确定Xnr2信号下游负责改变外胚层外植体对BMP信号的能力的基因。在这个筛选中确定的基因将被进一步表征，以将它们置于腹外侧中胚层诱导的途径中。

DESCRIPTION (provided by applicant): In vertebrates, numerous tissues and organs are derived in whole, or in part, from the mesoderm, including the skeletal muscles, heart, skeleton and kidney. A complex network of genes regulates the progressive specification of these various mesoderm derivatives. In Xenopus, the mesoderm forms from a band of cells around the equator of the early gastrula embryo. In models of mesoderm patterning in Xenopus, the Spemann organizer has figured prominently, and is thought to specify mesodermal fates through its secretion of inducing factors. We have recently shown that a Spemann organizer-independent pathway establishes a pattern along the animal-vegetal axis of the mesoderm by the mid-gastrula stage. Two distinct domains can be observed in this axis. The animal domain expresses the gene brachyury and appears to be fated to become notochord and somite, while the vegetal domain is fated to become ventrolateral mesoderm. The goal of this proposal is to further elucidate the mechanisms that act to establish patterning along the animal/vegetal axis. In particular, we will investigate the role of the nodal growth factors in the induction of ventral mesoderm. Our preliminary results indicate that Xenopus nodal-related-2 (Xnr2)-expressing cells are restricted to the presumptive ventral mesoderm in the gastrula embryo. Lineage tracing experiments proposed here are aimed at rigorously testing the fate of these cells. Conclusively establishing the fates of cells within the early presumptive mesoderm, and correlating them with restricted gene expression patterns, is an essential first step in elucidating the mechanism and timing of specification of fates. We have also hypothesized that nodals plays an essential role in the induction of ventral mesoderm, and we will examine the activity of nodals in the gastrula embryo by use of dominant negative constructs and inhibitory binding proteins. Finally, our preliminary results show that the combination of Xnr2 signaling and low FGF activity, such as exists in the vegetal marginal zone, makes ectoderm explants differentiate as blood, a ventral mesoderm derivative, in response to bone morphogenetic proteins (BMP). We will use subtraction and expression cloning approaches to identify genes downstream of Xnr2 signaling that are responsible for altering the competence of ectoderm explants to BMP signaling. Genes that are identified in this screen will be further characterized to place them in the pathway of ventrolateral mesoderm induction.