Cygan J A, Johnson R L, McMahon A P
Department of Molecular and Cellular Biology, The Biolabs, Harvard University, Cambridge, Massachusetts 02138, USA.
Development. 1997 Dec;124(24):5021-32. doi: 10.1242/dev.124.24.5021.
Classical embryological experiments have demonstrated that dorsal-ventral patterning of the vertebrate limb is dependent upon ectodermal signals. One such factor is Wnt-7a, a member of the Wnt family of secreted proteins, which is expressed in the dorsal ectoderm. Loss of Wnt-7a results in the appearance of ventral characteristics in the dorsal half of the distal limb. Conversely, En-1, a homeodomain transcription factor, is expressed exclusively in the ventral ectoderm, where it represses Wnt-7a. En-1 mutants have dorsal characteristics in the ventral half of the distal limb. Experiments in the chick suggest that the dorsalizing activity of Wnt-7a in the mesenchyme is mediated through the regulation of the LIM-homeodomain transcription factor Lmx-1. Here we have examined the relationship between Wnt-7a, En-1 and Lmx-1b, a mouse homolog of chick Lmx-1, in patterning the mammalian limb. We find that Wnt-7a is required for Lmx-1b expression in distal limb mesenchyme, and that Lmx-1b activation in the ventral mesenchyme of En-1 mutants requires Wnt-7a. Consistent with Lmx-1b playing a primary role in dorsalization of the limb, we find a direct correlation between regions of the anterior distal limb in which Lmx-lb is misregulated during limb development and the localization of dorsal-ventral patterning defects in Wnt-7a and En-1 mutant adults. Thus, ectopic Wnt-7a expression and Lmx-1b activation underlie the dorsalized En-1 phenotype, although our analysis also reveals a Wnt-7a-independent activity for En-1 in the repression of pigmentation in the ventral epidermis. Finally, we demonstrate that ectopic expression of Wnt-7a in the ventral limb ectoderm of En-1 mutants results in the formation of a second, ventral apical ectodermal ridge (AER) at the junction between Wnt-7a-expressing and nonexpressing ectoderm. Unlike the normal AER, ectopic AER formation is dependent upon Wnt-7a activity, indicating that distinct genetic mechanisms may be involved in primary and secondary AER formation.
经典胚胎学实验表明，脊椎动物肢体的背腹模式形成依赖于外胚层信号。其中一个这样的因子是Wnt-7a，它是分泌蛋白Wnt家族的成员，在背侧外胚层表达。Wnt-7a的缺失会导致远端肢体背侧半部出现腹侧特征。相反，同源异型域转录因子En-1仅在腹侧外胚层表达，在那里它抑制Wnt-7a。En-1突变体在远端肢体腹侧半部具有背侧特征。鸡的实验表明，Wnt-7a在间充质中的背化活性是通过对LIM-同源异型域转录因子Lmx-1的调控介导的。在这里，我们研究了Wnt-7a、En-1和Lmx-1b（鸡Lmx-1的小鼠同源物）在哺乳动物肢体模式形成中的关系。我们发现Wnt-7a是远端肢体间充质中Lmx-1b表达所必需的，并且En-1突变体腹侧间充质中Lmx-1b的激活需要Wnt-7a。与Lmx-1b在肢体背化中起主要作用一致，我们发现在肢体发育过程中Lmx-1b表达失调的前远端肢体区域与Wnt-7a和En-1突变体成年动物背腹模式形成缺陷的定位之间存在直接相关性。因此，异位Wnt-7a表达和Lmx-1b激活是背化En-1表型的基础，尽管我们的分析还揭示了En-1在抑制腹侧表皮色素沉着方面具有不依赖Wnt-7a的活性。最后，我们证明在En-1突变体的腹侧肢体外胚层中异位表达Wnt-7a会导致在表达Wnt-7a和不表达Wnt-7a的外胚层交界处形成第二个腹侧顶端外胚层嵴（AER）。与正常AER不同，异位AER的形成依赖于Wnt-7a活性，这表明初级和次级AER形成可能涉及不同的遗传机制。
