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The Role of Forkhead Box Transcription Factors in Zebrafish Ocular Development and the Superior Ocular Fissure
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- Author / Creator
- Stach, Tara R
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Ocular development is a tightly orchestrated process that is dependent upon precise environmental and genetic factors. Disruption of this can result in microphthalmia, anophthalmia and coloboma (MAC), which is a spectrum of congenital ocular disease with lifelong consequences affecting sight. In particular, coding anomalies in a member of the Bone Morphogenetic Protein family, GDF6, results in MAC in patients. Using zebrafish gdf6a-/- mutants, we have found that the microphthalmia present in both patients and model organisms appears to be due the requirement for Gdf6a in the regulation of ocular proliferation, progenitor cell survival, and apoptosis in the developing eye. Additionally, we have found that Gdf6a is located upstream of two forkhead box (FOX) transcription factor genes, foxi1 and foxi2, which provide dorsal-ventral polarity to a region where ocular progenitor cells reside, the ciliary marginal zone (CMZ). This is the first report of its kind regarding spatio-temporal identity of the CMZ, and we show that foxi2 and gdf6a lie on converging genetic pathways that regulate ocular size.
The forkhead box (FOX) proteins are a family of transcription factors with diverse roles in development such as patterning of neural tissue, cell migration, proliferation, differentiation and survival. Mutations in FOX genes are associated with blinding ocular disorders that occur due to malformation of anterior segment components, and the failure of the optic nerve to correctly innervate the brain. Based on expression patterns and a regulation by Gdf6a, we hypothesized that foxi1 and foxi2 have roles in the patterning and growth of the retina. Knockdown of either foxi1 or foxi2 expression using morpholino (MO) technology resulted in aberrant dorsal-ventral ocular patterning and MAC phenotypes. However, foxi1-/- embryos do not have ocular phenotypes similar to those of the morphant, suggesting compensation by redundant genes, an allele that is not fully disruptive to protein function, or off-target MO effects.
The choroid fissure is a well-recognized developmental structure whose failure to close results in coloboma affecting structures of the inferior eye. However, patients with coloboma affecting structures of the superior eye led us to discover the superior ocular fissure, present during zebrafish ocular development and indicative of ~450 million years of evolutionary conservation. Biochemical modeling of transheterozygous CYP1B1 coding anomalies in a proband indicated these might underlie the pathogenicity of superior coloboma, with a requirement for this enzyme to metabolize vitamin A to its active derivative, retinoic acid (RA). Examination of the developing zebrafish retina shows expression of cyp1b1 and other RA metabolism enzymes at the locations of the ocular fissures, and supplementation of vitamin A to a coloboma model during development rescued the phenotype by facilitating closure of the inferior ocular fissure. Finally, I find that antagonism between the FOX genes foxg1 and foxd1 determine the location of fissure formation, with knockdown resulting in superior fissure shift to the nasal axis, or duplication, respectively.
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- Graduation date
- Fall 2014
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- Type of Item
- Thesis
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- Degree
- Master of Science
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- License
- This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.