Expression of sonic hedgehog gene in regenerating newt limbs

Abstract

Urodele amphibians such as the newt can regenerate their limbs even as adults. We have focused on the expression of sonic hedgehog, a gene that plays an important role in pattern formation during limb development, in regenerating limbs. Our in situ hybridization experiments showed that the sonic hedgehog gene was expressed in mesenchymal cells of the posterior region of regenerating limb blastemas, strongly suggesting the presence of a zone of polarizing activity (ZPA) in blastemas. Experiments with axially reversed grafts of blastemas further supported this suggestion. Axial reversal of blastemas leads to the regeneration of supernumerary limbs. The sonic hedgehog gene, which was used as a posterior marker, was expressed not only in the original region (new anterior side) of the graft, but also ectopically in the other region (new posterior side) of the same graft. The formation of supernumerary limbs was closely related to the duplication of sonic hedgehog expression on axially reversed grafts of blastemas. This duplication of sonic hedgehog expression could represent the presence of two ZPAs in axially reversed grafts. Thus, we suggest that the limb-regeneration blastema employs a ZPA as the signaling center for anterior-posterior patterning, as is the case in the developing limb bud.