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An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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The intrinsic and extrinsic factors that regulate vertebrate photoreceptor specification and differentiation are complex, and our understanding of all the players is far from complete. Her9, the zebrafish ortholog of human HES4, is a basic helix-loop-helix-orange transcriptional repressor that regulates neurogenesis in several developmental contexts. We have previously shown that her9 is upregulated during chronic rod photoreceptor degeneration and regeneration in adult zebrafish, but little is known about the role of her9 during retinal development. To better understand the function of Her9 in the retina, we generated zebrafish her9 CRISPR mutants. Her9 homozygous mutants displayed striking retinal phenotypes, including decreased numbers of rods and red/green cones, whereas blue and UV cones were relatively unaffected. The reduction in rods and red/green cones correlated with defects in photoreceptor subtype lineage specification. The remaining rods and double cones displayed abnormal outer segments, and elevated levels of apoptosis. In addition to the photoreceptor defects, her9 mutants also possessed a reduced proliferative ciliary marginal zone, and decreased and disorganized Müller glia. Mutation of her9 was larval lethal, with no mutants surviving past 13 days post fertilization. Our results reveal a previously undescribed role for Her9/Hes4 in photoreceptor differentiation, maintenance, and survival.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/fisiologia , Neurogênese , Células Fotorreceptoras Retinianas Cones/fisiologia , Células Fotorreceptoras Retinianas Bastonetes/fisiologia , Proteínas de Peixe-Zebra/fisiologia , Peixe-Zebra/embriologia , Animais , Animais Geneticamente Modificados , Diferenciação Celular , Proliferação de Células , Células Fotorreceptoras Retinianas Cones/patologia , Células Fotorreceptoras Retinianas Bastonetes/patologiaRESUMO
Purpose: Autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV) is a devastating inherited autoimmune disease of the eye that displays features commonly seen in other eye diseases, such as retinitis pigmentosa and diabetic retinopathy. ADNIV is caused by a gain-of-function mutation in Calpain-5 (CAPN5), a calcium-dependent cysteine protease. Very little is known about the normal function of CAPN5 in the adult retina, and there are conflicting results regarding its role during mammalian embryonic development. The zebrafish (Danio rerio) is an excellent animal model for studying vertebrate development and tissue regeneration, and represents a novel model to explore the function of Capn5 in the eye. Methods: We characterized the expression of Capn5 in the developing zebrafish central nervous system (CNS) and retina, in the adult zebrafish retina, and in response to photoreceptor degeneration and regeneration using whole-mount in situ hybridization, FISH, and immunohistochemistry. Results: In zebrafish, capn5 is strongly expressed in the developing embryonic brain, early optic vesicles, and in newly differentiated retinal photoreceptors. We found that expression of capn5 colocalized with cone-specific markers in the adult zebrafish retina. We observed an increase in expression of Capn5 in a zebrafish model of chronic rod photoreceptor degeneration and regeneration. Acute light damage to the zebrafish retina was accompanied by an increase in expression of Capn5 in the surviving cones and in a subset of Müller glia. Conclusions: These studies suggest that Capn5 may play a role in CNS development, photoreceptor maintenance, and photoreceptor regeneration.