Ontogeny of the Anuran Urostyle: The Developmental Context of Evolutionary Novelty

The urostyle, a composite skeletal structure formed at the onset of metamorphosis in frogs, is considered a structural novelty. Despite its evolutionary importance, major aspects of its development and evolution remain enigmatic. The urostyle forms by fusion of the mesoderm-derived coccyx with the endoderm- or superficial mesoderm-derived hypochord when the tadpoles metamorphose into frogs. Frog metamorphosis, where tissue coordination is mediated by thyroid hormone (TH), involves dramatic changes in locomotion. During this period, a remarkable transition from swimming using fins/tails (in tadpoles) to jumping, walking, digging, climbing, and swimming using limbs (in adults) occurs (an axial-driven to a limb-driven transition). Here, I focus on the ontogenetic and genetic underpinnings of development of the urostyle. Using a nearly complete developmental series from free-living larvae to metamorphs, I studied the ontogeny of the urostyle in Xenopus tropicalis, adapting clearing and staining techniques, histology, immunohistology, cell proliferation, and cell death (Chapter 2). Following documentation of morphological changes, I identified genes and gene-regulatory pathways responsible for formation of the urostyle (a structure that is conserved across all extant anurans) (Chapter 3), using RNA-seq and ATAC-seq approaches. Since formation of the urostyle coincides with metamorphosis, I also looked at the possible role of TH in controlling formation of this structure, both morphologically (Chapter 2) and genetically (Chapter 3), which helped me narrow down the genes directly regulated by TH. Finally, I tested the function of two candidate genes in development of the urostyle by using a functional assay (creating frameshift mutations and a conditional knockout system) (Appendix). Through my dissertation work, I highlight how the changes to an underlying ancestral gene regulatory network would eventually alter the phenotype and genotype, giving rise to evolutionary novelties