Genetic and genomic approaches to understand size and proportion

Abstract

The establishment of proper proportions during development is essential for the final form and function of an organism. While much is known concerning the regulation of proliferation and differentiation, how the relative sizes of structures are established during development and are varied during evolution is poorly understood. Genetic screens in zebrafish have recently identified mutations in a series of potassium channels that are sufficient to cause an increase in the size of fins and barbels. How mutations in potassium channels enable the coordinated growth of these complex tissues is unknown. Here, I show that zebrafish with mutations in the potassium channel kcnk5b have a larger fin size through cell proliferation rather than an increase in cell size. This growth phenotype requires potassium conductance, and overexpression of kcnk5b within the fin mesenchyme is sufficient for growth. I then identify regulation of channel activity through an interaction between the Kcnk5b cytoplasmic C-terminus and the protein phosphatase calcineurin. This interaction is sufficient to regulate fin growth rate independently of the underlying genetic cues for absolute size and without re-specifying positional identity of the fin. To assess the potential for potassium channel-mediated scaling of fin size during evolution, I developed a cross-species targeted sequence capture approach for comparative genomic analysis in species lacking prior genetic resources. This technique was tested in a case-study of scale-loss within fish of the genus Phoxinellus. Using this analysis pipeline, I identify patterns of drift within members of the FGF signaling pathway, and show sufficiency of changes within this pathway in zebrafish mutants to phenocopy the evolved scalation pattern. I then applied this approach toward understanding the evolution of fin proportions in the flying fishes (Exocoetidae, Beloniformes). Through clade-wide exome sequencing of Beloniformes, I have generated key resources to parse the genomic changes associated with evolutionary shifts in skeletal proportionality within this group. With experimental models and genetic tools in the zebrafish, genomic signatures within the dataset can be tested. Together, these findings support a role for bioelectric signaling in organ size regulation and provide an avenue for genetic analysis of evolutionary changes in organ size in non-model organisms.