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Vol. 14, No. 11, pp. 1377-1389, June 1, 2000
1 Department of Cell Biology; 2 Howard Hughes
Medical Institute and Developmental Genetics Program, Skirball
Institute of Biomolecular Medicine; 3 Department of Radiology
and Pathology and Skirball Institute of Biomolecular Medicine, New York
University School of Medicine, New York, New York 10016 USA;
4 Max Planck Institute of Psychiatry, Clinical Neurogenetics,
80804 Munich, Germany; 5 GSF-Research Center, Institute for
Mammalian Genetics, 85764 Oberschleissheim, Germany; 6 Ronald
O. Perelman Department of Dermatology and 7 Department of
Physiology and Neuroscience, New York University School of Medicine,
New York, New York 10016 USA
Proximal-distal outgrowth of the vertebrate limb bud is regulated
by the apical ectodermal ridge (AER), which forms at an invariant
position along the dorsal-ventral (D/V) axis of the embryo. We have studied the genetic and cellular events that regulate AER formation in the mouse. In contrast to implications from previous studies in chick, we identified two distinct lineage boundaries in
mouse ectoderm prior to limb bud outgrowth using a
Cre/loxP-based fate-mapping approach and a novel
retroviral cell-labeling technique. One border is transient and at the
limit of expression of the ventral gene En1, which
corresponds to the D/V midline of the AER, and the second
border corresponds to the dorsal AER margin. Labeling of AER precursors
using an inducible Cre showed that not all cells that initially express
AER genes form the AER, indicating that signaling is required to
maintain an AER phenotype. Misexpression of En1 at moderate
levels specifically in the dorsal AER of transgenic mice was found to
produce dorsally shifted AER fragments, whereas high levels of
En1 abolished AER formation. In both cases, the dorsal gene
Wnt7a was repressed in cells adjacent to the
En1-expressing cells, demonstrating that signaling
regulated by EN1 occurs across the D/V border. Finally,
fate mapping of AER domains in these mutants showed that En1 plays a
part in positioning and maintaining the two lineage borders.
[Key Words: AER; boundary; En1; Engrailed1; limb; mouse]
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