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Michele DuVal, A. Phillip Oel, W Ted Allison; Gene Interactions in Cone Photoreceptor Differentiation: gdf6a and tbx2b. Invest. Ophthalmol. Vis. Sci. 2013;54(15):3726.
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© ARVO (1962-2015); The Authors (2016-present)
Functional vision restoration is within reach via stem cell therapy, but one of the largest obstacles is achieving cone photoreceptor differentiation. To enhance progress made using murine models of neuroregeneration, we are using the cone-rich zebrafish model. Here we test if gdf6a and tbx2b interact in cone differentiation. Gdf6a is an emerging factor in congenital photoreceptor degeneration. Tbx2b is implicated in rod-vs-cone specification and UV cone fate (homologue to mammalian blue cones). These factors are known to act in the same pathway establishing dorsal identity in early eye field development, but interactions in cone fate hypothesized herein would be surprising considering the discrete phenotypes observed in the respective zebrafish mutants.
We compared phenotypes of zebrafish mutants lacking gdf6a and/or tbx2b (both with recessive inheritance). Photoreceptors’ abundances were assessed using a combination of transgenes and immunohistochemistry in larval fish. Rates of phenotypes in gdf6a-/- (microphthalmia) and tbx2b-/- (the lor phenotype- overabundance of rods and scarcity of UV cones) were measured. Compound heterozygous gdf6a+/- ; tbx2b+/- fish were bred to assess if the genes interact, indicated by synergistic enhancement of phenotypes or non-Mendelian inheritance.
In gdf6a-/- larvae, UV cones were not detectably different in abundance or distribution, contradicting expected results if gdf6a were upstream of tbx2b. However the blue/UV cone abundance ratio was lower in gdf6a-/- larvae compared to siblings (86% lower, p=0.014) (6dpf). Incrosses of gdf6a+/- ; tbx2b+/- mutants yielded rates of the lor phenotype significantly higher than predicted Mendelian ratios of 25% among microphthalmic larvae (58%, χ2 p<0.001) (6dpf).
Here we establish a novel link between gdf6a and tbx2b in specifying photoreceptor fates. Although gdf6a is upstream of tbx2b during early retinal development, UV cones in gdf6a-/- mutants successfully differentiate, suggesting gdf6a does not directly affect tbx2b at this stage, and may instead regulate other cone subtypes. However the elevated rates of phenotypes when both genes are disrupted indicate that cone development is subject to an intricate regulatory pathway involving both these genes. Understanding this interaction is a necessary step in the refinement of stem cell-based restoration of daytime vision in humans.
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