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Lakshmi S. Pillai, Wen Wen, Ann C. Morris; Sox11 is Required for Proper Optic Fissure Closure and Photoreceptor Development. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1317.
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The SRY-Box transcription factor Sox11 is expressed in the developing central and peripheral nervous system, and regulates the development and differentiation of several organ systems. Sox11-/- mice are microphthalmic and exhibit Peter's anomaly and anterior segment dysgenesis, suggesting that Sox11 is required for proper ocular morphogenesis. Here, we have investigated the role of Sox11 in ocular development in the zebrafish, using an in vivo knockdown approach.
Translation blocking morpholinos (MO) were injected into 1-cell stage wild type and transgenic zebrafish embryos. The embryos were collected at 24, 48 and 72 hours post fertilization and processed for whole-mount in situ hybridization, histological and immunohistochemical examination. Digoxigenin labeled riboprobes were used to identify expression patterns of sox11a, sox11b, pax6a, and crx. Apoptosis was analyzed via TUNEL labeling, and cell proliferation and cellular differentiation were analyzed via immunohistochemistry for PH3, BrdU, and cell type specific markers. Morpholino-injected embryos were examined by bright field and fluorescence microscopy; area measurements of the eye were taken by outlining the entire eye and the lens using Nikon Elements software.
Whole-mount in situ hybridization showed that zebrafish sox11 paralogs, sox11a, and sox11b, have overlapping yet distinct expression patterns in the eye during early development. Furthermore, both transcripts are present in mitotic retinal progenitor cells as well as post-mitotic precursor cells. Microscopic and immunohistological examination showed that knockdown of both Sox11a and Sox11b resulted in micropthalmia, delayed lens development, coloboma, and specific reduction of photoreceptor cells. These phenotypes were rescued by co-injection of mRNA specific for sox11a and sox11b. Additionally, Sox11 knockdown resulted in a decrease in the expression levels of pax6a, crx, and Nr2e3 along with an increase in the number of mitoses. No significant difference in apoptosis was observed between control and Sox11 morphant eyes.
Our results show that reduction in the levels of Sox11 causes abnormal ocular morphogenesis as well as coloboma. This phenotype is accompanied by a specific deficit in photoreceptor cells in the dorsal retina. Such results are consistent with the hypothesis that Sox11 regulates early ocular morphogenesis as well as photoreceptor differentiation in the developing retina, possibly by regulating expression of genes necessary for photoreceptor cell fate specification. Studies are ongoing to characterize the mechanism of optic fissure closure defect in Sox11 deficient eyes.
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