conditional gene targeting in mice by means of cre-loxp strategy represents a powerful approach to study mammalian gene function. this approach is however dependent on the availability of suitable strains of mice with a tissue or time restricted activity of the cre recombinase. Here we describe Aldh3-Cre transgenic mice as a useful tool to conditionally delete genes in cornea, a specialized transparent tissue found on the anterior-most part of the eye, which acts as a protective barrier and contributes to the refractive power. Using a set of floxed alleles we demonstrate high Aldh3-Cre activity in corneal epithelial cells, corneal stroma and conjunctival epithelial cells at postnatal stages. Aldh3-Cre will thus be particularly beneficial for functional analysis of genes which are vital for postnatal development of cornea and conjunctiva.

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Chromatin remodeling complexes are required for many distinct nuclear processes such as transcription, DNA replication and DNA repair. However, how these complexes contribute to the development of complex tissues within an organism is poorly characterized. Imitation switch (ISWI) proteins are among the most evolutionarily conserved ATP-dependent chromatin remodeling factors and are represented by yeast Isw1/Isw2, and their vertebrate counterparts Snf2h (Smarca5) and Snf2l (Smarca1). In this study, we focused on the role of the Snf2h gene during development of the mammalian retina. We show that Snf2h is expressed in both retinal progenitors and post-mitotic retinal cells. Using Snf2h conditional knockout mice (Snf2h cKO), we found that when Snf2h is deleted the laminar structure of the adult retina is not retained, the overall thickness of the retina is significantly reduced compared with controls, and the outer nuclear layer (ONL) is completely missing. Depletion of Snf2h did not influence the ability of retinal progenitors to generate all of the differentiated retinal cell types. Instead, Snf2h function is critical for proliferation of retinal progenitor cells. Cells lacking Snf2h have a defective S-phase, leading to the entire cell division process impairments. Although, all retinal cell types appear to be specified in the absence of Snf2h function, cell cycle defects and concomitantly increased apoptosis in Snf2h cKO result in abnormal retina lamination, complete destruction of the photoreceptor layer and, consequently, in a physiologically non-functional retina.

Chromatin remodeling complexes are required for many distinct nuclear processes such as transcription, DNA replication, and DNA repair. However, the contribution of these complexes to the development of complex tissues within an organism is poorly characterized. Imitation switch (ISWI) proteins are among the most evolutionarily conserved ATP-dependent chromatin remodeling factors and are represented by yeast Isw1/Isw2, and their vertebrate counterparts Snf2h (Smarca5) and Snf2l (Smarca1). In this study, we focused on the role of the Snf2h gene during the development of the mammalian retina. We show that Snf2h is expressed in both retinal progenitors and post-mitotic retinal cells. Using Snf2h conditional knockout mice (Snf2h cKO), we found that when Snf2h is deleted, the laminar structure of the adult retina is not retained, the overall thickness of the retina is significantly reduced compared with controls, and the outer nuclear layer (ONL) is completely missing. The depletion of Snf2h did not influence the ability of retinal progenitors to generate all the differentiated retinal cell types. Instead, the Snf2h function is critical for the proliferation of retinal progenitor cells. Cells lacking Snf2h have a defective S-phase, leading to the entire cell division process impairments. Although all retinal cell types appear to be specified in the absence of the Snf2h function, cell-cycle defects and concomitantly increased apoptosis in Snf2h cKO result in abnormal retina lamination, complete destruction of the photoreceptor layer, and consequently, a physiologically non-functional retina.

Mammalian corneal development is a multistep process, including formation of corneal epithelium (CE), endothelium and stroma during embryogenesis followed by postnatal stratification of the epithelial layers, and continuous renewal of the epithelium to replace the most outer corneal cells. Herein we employed Cre-loxP system to conditionally deplete Pax6 proteins in two domains of ocular cells, including the ocular surface epithelium (cornea, limbus and conjunctiva) or postnatal CE, via K14-cre or Aldh3-cre, respectively. Earlier and broader inactivation of Pax6 in the OSE resulted in thickened OSE with CE and limbal cells adopting the conjunctival keratin expression pattern. More restricted depletion of Pax6 in postnatal CE resulted in the abnormal cornea marked by reduced epithelial thickness despite of increased epithelial cell proliferation. Immunofluorescence studies showed loss of Keratin 12, an intermediate filament and diffused expression of adherens junction components, together with reduced tight junction protein, Zonula occludens-1 (ZO-1). Furthermore, expression of Keratin 14, basal cell marker in apical layers, indicates impaired differentiation of corneal epithelial cells. Collectively, our data demonstrate that Pax6 is essential for maintaining proper differentiation and strong intercellular adhesion in postnatal corneal epithelial cells, whereas limbal Pax6 is required for preventing the outgrowth of conjunctival cells to the cornea.