Induction of ectopic external gills and tetrapodomorph-like skeletal elements through homeotic transformations in the salamander branchial region

Homeotic transformations are morphological changes associated with alterations of identities of segments in serially repeated systems and these changes may be a source of modifications in body plan evolution. Retinoic acid (RA) signaling has previously been shown to induce homeotic transformations in the vertebral column, although its role in other vertebrate segmented systems remains unexplored. Here, we use pharmacological inhibition of RA receptors to study homeotic transformations in the larval salamander branchial region. This region normally contains three pairs of external gills supported by the underlying skeleton, however upon treatments we observe induction of ectopic outgrowths in the posterior portion of the apparatus. We characterize these outgrowths as ectopically induced fourth external gills on the position of the otherwise gill-free segment. This induction is further associated with transformations and appearance of cartilaginous elements that phenocopy similar elements in fossil stem amphibians and tetrapodomorphs. These experimentally instigated morphological changes qualify as homeotic transformations in the branchial region and present re-emergence of features that were lost prior to the origin of modern tetrapods. More broadly, our results point to RA signaling as a potent driver regulating the number and composition of pharyngeal segments and thus controlling the evolution of the vertebrate pharyngeal apparatus.