Differential regulation of T-box and homeobox transcription factors suggests roles in controlling chick limb-type identity

The wing and the leg of the chick, although homologous structures, have characteristic patterns of skeletal elements, muscles, tendons, featherbuds and scales. Despite recent advances in understanding the common genetic pathways patterning the wing and leg, the molecular nature of the specification of limb-type identity has remained elusive. Embryological experiments have indicated the existence of limb-specific territories in the flank. In the newt, deviation of nerves from the limb into the flank can induce ectopic limbs to form from this tissue. In the chick, Fibroblast growth factor (FGF)-soaked beads applied to the flank can induce ectopic formation of limbs from the surrounding tissue. In both cases, the type of limb that forms, either a wing/forelimb or leg/hindlimb, is dependent upon the location to which the limb-inducing signal is applied. We have isolated and characterised three candidate genes for controlling limb identity in the chick. Two T-box transcription factors, cTbx4 and cTbx5, are expressed in a restricted manner in the leg bud and wing buds, respectively, cPtx1, a member of the Otx-related subclass of paired-type homeodomain proteins, is expressed exclusively in the leg bud. Using FGF to induce ectopic limb buds of wing, leg and intermediate identity, we show that early expression of cTbx5, cTbx4 and cPtx1 in the induced limb buds correlates with later wing- or leg-type identity of ectopic limbs. We observe a general correlation between the location of an ectopic outgrowth induced by FGF and the identity of the resulting limb but, significantly, we report that there is no definitive rostral-caudal level that divides the ectopic wing and leg territories.