Developing an ancient epithelial appendage: FGF signalling regulates early tail denticle formation in sharks

Rory L. Cooper, Kyle J. Martin, Liam J. Rasch, Gareth J. Fraser

Vertebrate epithelial appendages constitute a diverse group of organs that includes integumentary structures such as reptilian scales, avian feathers and mammalian hair. Recent studies have provided new evidence for the homology of integumentary organ development throughout amniotes, despite their disparate final morphologies. These structures develop from conserved molecular signalling centres, known as epithelial placodes. It is not yet certain whether this homology extends beyond the integumentary organs of amniotes, as there is a lack of knowledge regarding their development in basal vertebrates. As the ancient sister lineage of bony vertebrates, extant chondrichthyans are well suited to testing the phylogenetic depth of this homology. Elasmobranchs (sharks, skates and rays) possess hard, mineralised epithelial appendages called odontodes, which include teeth and dermal denticles (placoid scales). Odontodes constitute some of the oldest known vertebrate integumentary appendages, predating the origin of gnathostomes. Here, we used an emerging model shark (Scyliorhinus canicula) to test the hypothesis that denticles are homologous to other placode-derived amniote integumentary organs. To examine the conservation of putative gene regulatory network (GRN) member function, we undertook small molecule inhibition of fibroblast growth factor (FGF) signalling during caudal denticle formation. We show that during early caudal denticle morphogenesis, the shark expresses homologues of conserved developmental gene families, known to comprise a core GRN for early placode morphogenesis in amniotes. This includes conserved expression of FGFs, sonic hedgehog (shh) and bone morphogenetic protein 4 (bmp4). Additionally, we reveal that denticle placodes possess columnar epithelial cells with a reduced rate of proliferation, a conserved characteristic of amniote skin appendage development. Small molecule inhibition of FGF signalling revealed placode development is FGF dependent, and inhibiting FGF activity resulted in downregulation of shh and bmp4 expression, consistent with the expectation from comparison to the amniote integumentary appendage GRN. Overall, these findings suggest the core GRN for building vertebrate integumentary epithelial appendages has been highly conserved over 450 million years. This provides evidence for the continuous, historical homology of epithelial appendage placodes throughout jawed vertebrates, from sharks to mammals. Epithelial placodes constitute the shared foundation upon which diverse vertebrate integumentary organs have evolved.