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Cagney Coomer, Ann C Morris; Loss of Her9/HES4 in zebrafish disrupts photoreceptor development. Invest. Ophthalmol. Vis. Sci. 2019;60(9):6035.
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© ARVO (1962-2015); The Authors (2016-present)
Her9, the zebrafish ortholog of mammalian HES4, is a bHLH-O transcriptional repressor that is regulated by retinoic acid (RA) signaling. Her9 has been shown to inhibit neurogenesis at the boundaries and/or pro-neural stripes of the neural plate. Our lab has previously shown that Her9 is upregulated during chronic rod photoreceptor degeneration and regeneration but little is known about the role of Her9 during retinal development. To elucidate the role of Her9 during retinal development, we generated her9 CRISPR mutants and characterized their retinal phenotypes. We observed that her9 homozygous mutants fail to demonstrate a normal visually mediated background adaptation (VBA) response, suggesting possible visual impairment and abnormal retinal development. Here, we show that loss of Her9 results in photoreceptor subtype-specific abnormalities, including changes in subtype number, morphology, and survival.
Retinal cell type differentiation in wild type and her9 mutants was analyzed by immunohistochemistry and using fluorescent reporter transgenic lines. Photoreceptor number was quantified, and outer segment length was measured, on a minimum of 3 retinal sections from 10 individuals. Apoptosis was detected by TUNEL assay. Retinal progenitor cell proliferation was analyzed using an antibody to PCNA. Gene expression was analyzed by qPCR and by fluorescent in situ hybridization (FISH).
In the retinas of the her9 homozygous mutants we observed a significant reduction in the number rod photoreceptors present at 6 dpf. Green and red double-cones were also reduced, and the remaining red/green cones displayed an abnormal morphology with shorter outer segments. Interestingly, we found that the number of UV cones was significantly increased in her9 mutant retinas compared to wild type and heterozygous siblings, and the number of blue cones was not changed. The reductions in rod and cone photoreceptors were accompanied by an increase in apoptosis in the outer nuclear layer of her9 mutant retinas. Finally we observed that her9 mutant retinas possessed a reduced number of Müller glia, a much smaller ciliary marginal zone.
Taken together, our results indicate that Her9/HES4 is required for proper photoreceptor development and may regulate cone photoreceptor subtype specification. Her9 may also control the proliferation and survival of retinal progenitor cells.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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