The Role of Rx in Embryonic Retinogenesis: Determining Genetic Influences on Optic Vesicle Formation and Photoreceptor Cell Fate

Eye development is a dynamic and complex process that is controlled by the interactions of transcription factors, signaling pathways, and growth factors. Disruption of the developmental process can result in ocular malformations or retinal diseases, which can cause blindness. Developing tools to study embryonic retinogenesis and understanding the molecular mechanisms involved are important for increasing our understanding of neural development, understanding ocular malformations such as anophthalmia, and may improve or lead to new treatments for eye diseases including blindness. This work aims to develop new tools for studying early eye development and explore genes associated with optic vesicle development and photoreceptor cell fate. In Study 1, we sought to identify and characterize markers of embryonic cone photoreceptors. We identified that two genes involved in phototransduction, phosducin and cone transducin gamma, are expressed in developing cones. We characterized the temporal and spatial profile of both genes and their associated proteins over the developmental timeline of retinogenesis. Further, we determined their colocalization with known cone and photoreceptor markers and thus established them as useful markers for further studies of early cone histogenesis. In Study 2, we assessed the role of the homeobox gene, Rx, in progenitor proliferation and cell fate determination in the mouse retina using a conditional knockout. Deletion of Rx in retinal progenitors led to a loss of retinal lamination, depletion of the retinal progenitors and in the mature retina showed changes in retinal cell types. Late-born cells (rods, bipolar cells, and Muller glia) were absent, likely due to the depleted progenitor pool. Cones (an early-born retinal cell type) were also absent; examination of cone histogenesis showed Rx is necessary for cone photoreceptor generation. Finally, in Study 3 we identified an effective gene knockdown method for 3D optic vesicle organoid culture that is useful for studying gene expression and early retinal development. Using this method, we assessed the roles of three candidate genes in optic vesicle development and identified one gene that warrants further investigation in vivo. Collectively these studies provide new tools for studying early embryogenesis and further our knowledge of the genetics underlying optic vesicle development and cone photoreceptor formation