title: Novel regulatory interactions revealed by studies of murine limb pattern in Wnt-7a and En-1 mutants.
abstract: 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.
authors.affiliation: Department of Molecular and Cellular Biology, The Biolabs, Harvard University, Cambridge, Massachusetts 02138, USA.
authors.forename: J A
authors.initials: JA
authors.lastname: Cygan
authors.forename: R L
authors.initials: RL
authors.lastname: Johnson
authors.forename: A P
authors.initials: AP
authors.lastname: McMahon
mesh_terms: D000818:Animals; D030161:Avian Proteins; D019521:Body Patterning / Q000235:genetics*; D005121:Extremities / Q000196:embryology*; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D053823:Hedgehog Proteins; D018398:Homeodomain Proteins / Q000235:genetics / Q000502:physiology*; D060850:LIM-Homeodomain Proteins; D018878:Limb Buds; D008648:Mesoderm / Q000737:chemistry; D051379:Mice; D009154:Mutation; D011506:Proteins / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000235:genetics; D015534:Trans-Activators; D014157:Transcription Factors; D051153:Wnt Proteins
chemical_list: D030161:Avian Proteins; C080330:En1 protein, mouse; D053823:Hedgehog Proteins; D018398:Homeodomain Proteins; D060850:LIM-Homeodomain Proteins; C489692:Lmx1a protein, mouse; D011506:Proteins; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D015534:Trans-Activators; D014157:Transcription Factors; C490509:WNT7A protein, Gallus gallus; D051153:Wnt Proteins; C490508:Wnt7a protein, mouse