Browsing by Subject "Eye development"
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Item Determining the molecular underpinnings of extracellular matrix breakdown during choroid fissure closure(2016-05-20) Williams, Andre Matthew; Gross, Jeffrey Martin; Wallingford, John BProper closure of the choroid fissure (CF), a transient structure present during early eye development, is critical for eye organogenesis. Choroid fissure closure (CFC) is a dynamic process that requires morphogenetic movements, extracellular matrix (ECM) breakdown, and tissue fusion. Defects in this process result in coloboma, which is responsible for 3-11% of childhood blindness worldwide. Little is known about the mechanism of ECM breakdown during CFC, or what tissues are responsible for facilitating it. We hypothesized that podosomes, small actin-based structures that are capable of breaking down ECM in other cellular contexts, are responsible for ECM breakdown during CFC. Using a mutant of the actin-linker protein Talin, we show that Talin is not responsible for ECM breakdown, as normal ECM breakdown occurs in talin [superscript HI3093Tg] mutants. Mutation of tks4, a core podosome component, results in a small coloboma, suggesting a likely role for podosomes in proper retinal fusion. Previous data suggested that the hyaloid vasculature, which passes through the CF to envelop the lens, is involved in degrading the ECM during CFC. Though the hyaloid vasculature is present, Actin within the hyaloid vasculature did not localize to regions that have intact ECM, suggesting that the hyaloid vasculature is not involved in ECM breakdown. Finally, I used chicken as a potential new model organism to investigate mechanisms of breakdown in CFC. However, no ECM breakdown was observed in chicken CFC, and invasion of mesenchymal cells that separates the CF was observed. Further investigation is required to determine the molecular underpinnings of CFC.Item Molecular mechanisms of choroid fissure closure and ventral retina formation in the zebrafish eye(2010-12) Lee, Jiwoon; Gross, Jeffrey MartinDuring optic cup morphogenesis, the neuroectodermal layers of the optic vesicle (OV) invaginate ventrally, and fuse at the choroid fissure (CF) along the proximo-distal axis such that the retina and retinal pigment epithelium (RPE) are confined within the cup. Failure of CF closure results in colobomas, which are characterized by the persistence of a cleft or hole at the back of the eye. While CF closure is a critical aspect of ocular development, the molecular and cellular mechanisms underlying this process are poorly understood. My research examined CF closure and colobomas using zebrafish as a model system. In the first study, I determined that early cell fate changes within the eye field could cause colobomas using the zebrafish mutant blowout. Colobomas in blowout resulted from defects in optic stalk morphogenesis whereby the optic stalk extended into the retina and impeded the edges of the CF from meeting and fusing. Positional cloning of blowout identified a nonsense mutation in patched1, a negative regulator of the Hedgehog pathway. Up-regulation of Hedgehog pathway activity causes disruption in the patterning of the OV into proximal and distal territories, revealing that cell fate determination, mediated by Hedgehog signaling, is intimately involved in regulating CF closure. In the second study, I examined Bcl6 function and regulation during zebrafish eye development. bcl6 encodes a transcriptional repressor expressed in the ventral retina during zebrafish eye development. Loss of Bcl6 function leads to colobomas along with up-regulation of p53, a previously known Bcl6 target, and an increase in the number of apoptotic cells in the retina, demonstrating that Bcl6 plays a critical role in preventing apoptosis in the retina during early eye development. I also showed that Vax1 and Vax2 act upstream of bcl6 in the ventral retina. Furthermore, I identified functional interactions between Bcl6, Bcor and Hdac1 during eye development, demonstrating that Bcl6 functions along with Bcor and Hdac1 to mediate cell survival by regulating p53 expression. Together my studies expand the gene regulatory network involved in cell fate determination and cell survival during CF closure and ventral retina formation, and provide mechanistic insight into coloboma formation.