Latent Developmental Potential to Form Limb-Like Structures in Fish Fins Revealed by Mutations in the Vav2/N-WASP Pathway

Abstract

Despite descending from a common antecedent structure, fish fins and tetrapod limbs have evolved drastically divergent skeletal patterns. These changes have been instrumental to the success of vertebrate lineages both on land and in water. A key feature of limb evolution is the elaboration of the endoskeleton along the proximodistal axis. Teleost fins, on the other hand, have no such articulations in their diminutive endoskeleton. I investigated the genetic basis of fin patterning using a forward genetic approach in the zebrafish (Danio rerio) and identified two mutants that develop novel long bones along the proximodistal axis of the pectoral fin. The new bones are well patterned, form joints, and connect with fin musculature. Formation of the new bones requires Hox11 paralogs. Loss of Hox13 paralogs results in an enhancement of the mutant phenotype, with additional new bones formed distally. These results suggest that the new bones are patterned with similar mechanisms used to specify the middle aspect of tetrapod limbs. Removal of Wasl, the mouse paralog of the affected gene in fish mutants, from limb mesenchyme results in a phenotype similar to Hoxa-11 mutants. My data reveal that zebrafish have the developmental potential to form articulated fin endoskeletons using limb-like patterning cues. As this potential was likely present in the bony fish ancestor, zebrafish retain this potential in a latent state, and it can be induced by a simple mutation. Wasl was not known to have roles in patterning or skeletal development. By generating zebrafish mutants and mouse conditional mutants, I found that this gene is required for dermal and endochondral bone patterning in the axial and appendicular skeletons. The observed phenotypes are similar to Hox mutants, suggesting that Wasl interacts with Hox in skeletal patterning globally. I then analyzed genetic regulation of fin development in a basally branching teleost relative, the bowfin (Amia calva), which retains ancestral proximodistal elaboration of the pectoral fin. Using RNAseq analysis, I identify developmental genetic features shared between fins and limbs, as well as Holostean-specific changes. These data act as a touchstone linking disparate appendages to identify common mechanisms and causes of morphological change.