[F31] Mechanisms of Median Appendage Development

Description

Project Summary: Limb differences are relatively common congenital birth anomalies, yet many patients lack a definitive diagnosis. Known developmental pathways do not account for all instances of limb differences, highlighting a need to study limb development from unique perspectives. Due to their homology to tetrapod limbs, paired appendages of aquatic species known as pectoral and pelvic fins have been extensively studied. Conversely, the developmental origins of median appendages such as the dorsal, anal, and caudal fins, are severely understudied. Median appendages arose prior to paired appendages evolutionarily, and it is believed that the developmental mechanisms that build median appendages were redeployed to build paired appendages. By studying the mechanisms underlying median appendage development in zebrafish, I can inform fundamental mechanisms of appendage development, potentially providing new avenues for diagnoses and treatment of limb differences. Median fins are derived from the sclerotome, a somitic subcompartment comprised of mesenchymal stem cells that differentiate into a multitude of structures including vertebrae and ribs. The goal of this project is to elucidate the mechanisms underlying sclerotome contribution to median fins. How axial and dorsal-ventral positioning of sclerotome cells contributes to median fin progenitor fate remains unknown. Further, we have opposing loss-of-fin (eomesa) and gain-of-fin (pax9) transcription factor mutants that will allow us to elucidate the genetics of median fin development. Epistasis experiments indicate that pax9 is epistatic to eomesa. Single- cell RNA sequencing of pax9 mutants suggest that pax9 biases sclerotome cells away from fin progenitor fate and towards another sclerotome derivative fate. Two aims test the overarching hypothesis that sclerotome cells at specific axial and dorsal-ventral positions face a binary fate choice to become median fin progenitors. In Aim 1, I will use focal Cre activation, the Zebrabow reporter system, and somite transplantations to determine the location, clonality, and commitment of fin progenitor cells. In Aim 2, I will use in situ hybridization expression analyses, single-cell RNA sequencing, and chromatin immunoprecipitation sequencing to determine when a genetic interaction between eomesa and pax9 occurs, whether it is a direct interaction, and if eomesa shifts sclerotome cell fate towards fin progenitor fate at the expense of another sclerotome derivative fate. During this fellowship, I will have support from my mentor Dr. James Nichols, co-sponsor Dr. Linda Barlow, my dissertation committee, and many individuals ranging in expertise from bioinformatics to microscopy. I will seek formal and informal training opportunities to further build my mentorship, science communication, and technical skills. Between the vibrant environment in the Nichols Lab and the many resources available to me as a Cells, Stem Cells and Development program graduate s Istituzione: UNIVERSITY OF COLORADO DENVER PI: Margaret Kay Keating Progetto: 1F31HD122320-01

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Source

Source: reporter.nih.gov