Three-dimensional skeletal patterning during sea urchin embryogenesis

Multi-tissue pattern formation during development is a complex process in which extracellular communication events specify distinct cell types and regulate exquisite embryonic morphogenesis. The sea urchin embryo provides an excellent model for studying pattern formation due to relative genetic and morphological simplicity. The larval skeleton is secreted via biomineralization by the skeletogenic primary mesenchyme cells (PMCs). The PMCs undergo an epithelial-mesenchymal transition and migrate as individual cells within the blastocoel into stereotypic positions; this regulated PMC migration and positioning is required for normal skeletal patterning. Elegant PMC transplant experiments have demonstrated that PMC positioning, and thus skeletal patterning, is directed by ectodermal cues, and not by cues internal to the PMCs. In recent years, new efforts have been made to identify the ectodermal gene products that regulate skeletal development. The transcription factors Otp, Pax2/5/8 and Strim1, signaling by FGF, VEGF, and Wnt5a ligands have all been implicated in skeletal development in the sea urchin embryo; however, loss-of-function analysis for most of these gene products results in inhibition of skeletogenesis, suggesting that they regulate biomineralization and not PMC positioning and patterning. Notably, no skeletal patterning genes have previously been identified that pattern specific parts of the larval skeleton. This dissertation takes candidate-based and systems-level approaches to identify novel skeletal patterning genes that pattern distinct parts of the larval skeleton. We find that activation of the Alk4/5/7 receptor is required during gastrulation for anterior PMC positioning and skeletal patterning. We next test the function of the TGF-ß ligand Univin and find that it is necessary and sufficient for secondary skeletal patterning, indicating that Univin is a likely signaling ligand in anterior skeletal patterning. We also report a ventral accumulation of sulfated proteoglycans that requires function of the sulfate transporter, SLC26a2/7. This SLC-dependent sulfated proteoglycan accumulation is necessary and sufficient to attract PMCs to the ventral territory, and thereby pattern the ventral transverse skeletal elements. Finally, BMP5-8 function is required for left-side skeletal and serotonergic neuron development. Together, these studies reveal novel ectodermal genes that specifically regulate skeletal patterning across the anterior-posterior, dorsal-ventral, and left-right axes in Lytechinus variegatus embryos.2017-01-01T00:00:00