Vincent Stephane D, Dunn N Ray, Hayashi Shigemi, Norris Dominic P, Robertson Elizabeth J
Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts 02138, USA.
Genes Dev. 2003 Jul 1;17(13):1646-62. doi: 10.1101/gad.1100503.
It is well known that cell fate decisions in the mouse organizer region during gastrulation ultimately govern gut formation and patterning, left-right axis determination, and development of the central nervous system. Previous studies suggest that signaling pathways activated by Nodal, bone morphogenetic protein (BMP), and Wnt ligands coordinately regulate patterning of the streak and the formation of midline organizing tissues, but the specific contributions of these molecules within discrete cell lineages are poorly defined. Here we removed Smad2 activity in the epiblast, using a conditional inactivation strategy. Abrogation of Smad2 does not compromise primitive streak (PS) formation or gastrulation movements, but rather results in a failure to correctly specify the anterior definitive endoderm (ADE) and prechordal plate (PCP) progenitors. To selectively lower Nodal activity in the posterior epiblast, we generated a novel allele lacking the proximal epiblast enhancer (PEE) governing Nodal expression in the PS. As for conditional inactivation of Smad2, germ-line deletion of the PEE selectively disrupts development of the anterior streak. In striking contrast, the node and its midline derivatives, the notochord and floor plate, develop normally in both categories of mutant embryos. Finally, we show that removal of one copy of Smad3 in the context of a Smad2-deficient epiblast results in a failure to specify all axial midline tissues. These findings conclusively demonstrate that graded Nodal/Smad2 signals govern allocation of the axial mesendoderm precursors that selectively give rise to the ADE and PCP mesoderm.
众所周知，原肠胚形成期间小鼠组织者区域的细胞命运决定最终控制着肠道的形成和模式、左右轴的确定以及中枢神经系统的发育。先前的研究表明，由Nodal、骨形态发生蛋白（BMP）和Wnt配体激活的信号通路协同调节原条的模式和中线组织的形成，但这些分子在离散细胞谱系中的具体作用尚不清楚。在这里，我们使用条件性失活策略去除了上胚层中的Smad2活性。Smad2的缺失并不影响原条（PS）的形成或原肠胚运动，而是导致无法正确指定前确定内胚层（ADE）和脊索前板（PCP）祖细胞。为了选择性降低后上胚层中的Nodal活性，我们生成了一个新的等位基因，该等位基因缺乏控制PS中Nodal表达的近端上胚层增强子（PEE）。与Smad2的条件性失活一样，PEE的种系缺失选择性地破坏了前原条的发育。与之形成鲜明对比的是，在这两类突变胚胎中，节点及其中线衍生物脊索和底板均正常发育。最后，我们表明，在Smad2缺陷的上胚层背景下去除一个Smad3拷贝会导致无法指定所有轴向中线组织。这些发现确凿地证明，分级的Nodal/Smad2信号控制着轴向中内胚层前体的分配，这些前体选择性地产生ADE和PCP中胚层。
