Pax-3 is necessary but not sufficient for lbx1 expression in myogenic precursor cells of the limb

In vertebrates all skeletal muscles of trunk and limbs are derived from condensations of the paraxial mesoderm, the somites. Limb muscle precursor cells migrate during embryogenesis from somites to limb buds where migration stops and differentiation occurs. We have characterized lbx1 homeobox genes in chicken and mice and found them to be expressed in migrating limb muscle precursor cells in both species. Analysis of splotch mutant mice showed that lbx1 and c-met are differently affected by the lack of Pax-3. Limb buds of splotch (Pax-3 mutant) mice were devoid of lbx1 transcripts, while expression of c-met was still detectable at a low level. The presence of c-met-positive cells in splotch mice entering the limbs indicates that migration of cells from somites to limbs is not entirely dependent on Pax-3. We show that induction of epithelial to mesenchymal transition of Pax-3-positive cells by SF/HGF was not sufficient to induce ectopic lbx-1 expression at the inter-limb level, while ectopic limb formation was able to activate lbx1 expression. We postulate that Pax-3 is necessary for lbx1 expression in the lateral tips of somites but additional, yet unknown signals derived from limb buds are needed to initiate lbx1 expression. The role of limb bud-derived signals involved in targeted muscle precursor cell migration, and lbx1 activation was further confirmed by analysis of explanted somite/limb bud co-cultures in collagen gels.