WNT3 AND AXIS FORMATION AND CNS DEVELOPMENT

在过去的十年中，Wnt基因家族的成员已被证明在脊椎动物胚胎中具有轴诱导活性。不幸的是，没有表达或遗传数据表明这些基因在正常发育中起着这种作用。然而，最近发现Wnt基因Wnt-3在原肠胚形成之前表达，并且在该位点突变的纯合小鼠缺乏体轴。尽管取得了这些期待已久的结果，但仍有许多问题没有得到解答。因此提出以下实验：具体目标1。确定Wnt-3突变小鼠胚胎体轴缺陷的基础。1.1在E5.5-E7.5胚胎组织学水平检测Wnt-3表达。1.2将产生野生型和Wnt-3突变细胞的嵌合胚胎，以确定早期胚胎中哪个Wnt-3表达区对体轴形成至关重要。1.3将Wnt-3 mRNA注入爪蟾胚胎，检测Wnt-3诱导轴的活性。具体目标2。确定Wnt-3在轴形成过程中与轴形成和Wnt信号通路的抑制因子Axin之间的关系。2.1生成Wnt-3/Axin双突变小鼠，以确定Axin-/-小鼠中存在的多余轴是否依赖于Wnt-3信号。具体目标3。确定Wnt-3在胚胎发生过程中的后期功能。3.1使用Wnt-3的条件等位基因（Wnt-3c），在原始条纹中剔除该基因产物，以确定维持原始条纹形成是否需要Wnt-3。3.2使用Wnt-3c等位基因，Wnt-3将从神经管中特异性消除，以确定其在中枢神经系统中的作用。

Within the last decade, members of the Wnt gene family have been shown to possess axis-inducing activity in vertebrate embryos. Unfortunately, there is no expression or genetic data that would suggest that these genes play this role in normal development. Very recently, however, it was found that a Wnt gene, Wnt-3, is expressed prior to gastrulation and that mice homozygous for a mutation at this locus lack a body axis. Notwithstanding these long awaited results, there are many questions that remain unanswered. Therefore the following experiments are proposed: Specific aim number 1. Determine the basis for the body axis defects in Wnt-3 mutant mouse embryos. 1.1 Wnt-3 expression will be examined at the histological level in E5.5-E7.5 embryos. 1.2 Chimeric embryos derived from wildtype and Wnt-3 mutant cells will be generated to determine which Wnt-3 expressing regions from early embryos are essential for body axis formation. 1.3 The axis inducing activity of Wnt-3 will be tested by injected Wnt-3 mRNA into Xenopus embryos. Specific aim number 2. Determine the relationship between Wnt-3 and Axin ( a repressor or axis formation and of the Wnt signaling pathway) in the process of axis formation. 2.1 Wnt-3/Axin double mutant mice will be generated to determine whether the supernumerary axes present in Axin-/- mice is dependent of Wnt-3 signaling. Specific aim number 3. Determine later functions of Wnt-3 during embryogenesis. 3.1 Using a conditional allele of Wnt-3 (Wnt-3c) the gene product will be eliminated in the primitive streak to determine whether Wnt-3 is required for the maintenance of primitive streak formation. 3.2 Using the Wnt-3c allele, Wnt-3 will be eliminated specifically from the neural tube to determine its role in patterning the central nervous system.