Abstract
CRB3 (Crumbs homologue 3), a member of the CRB protein family (homologous to the Drosophila Crumbs), is expressed in different epithelium-derived cell types in mammals, where it seems to be involved in regulating the establishment and stability of tight junctions and in ciliogenesis. This protein has been also detected in the retina, but little is known about its localization and function in this tissue. Our goal here was to perform an in-depth study of the presence of CRB3 protein in the mouse retina and to analyze its expression during photoreceptor ciliogenesis and the establishment of the plexiform retinal layers. Double immunofluorescence experiments for CRB3 and well-known markers for the different retinal cell types were performed to study the localization of the CRB3 protein. According to our results, CRB3 is present from postnatal day 0 (P0) until adulthood in the mouse retina. It is localized in the inner segments (IS) of photoreceptor cells, especially concentrated in the area where the connecting cilium is located, in their synaptic terminals in the outer plexiform layer (OPL), and in sub-populations of amacrine and bipolar cells in the inner plexiform layer (IPL).
Highlights
Mutations in the CRB1 gene (Crumbs homologue 1) have been linked to several human retinal dystrophies, including type 12 retinitis pigmentosa (RP12) and Leber congenital amaurosis (LCA) [1,2]
Using the antibodies described in two publications [11,12], the few authors who have addressed the distribution of Crumbs Homologue 3 Protein (CRB3) in the mouse retina have only located the protein at the level of the outer limiting membrane (OLM), in photoreceptors, and in Muller cells [14,16] In the present study, we detected CRB3 in Muller cells after performing double immunolabeling with anti-Cellular retinaldehyde-binding protein (CRALBP), a Muller cell marker (Fig. S2), we have observed that the majority of the localization is in photoreceptor cells
In adult mice we detected CRB3 in the connecting cilium area, where this protein surrounds the staining obtained for the acetylated tubulin protein, which is present in the cilium [35]
Summary
Mutations in the CRB1 gene (Crumbs homologue 1) have been linked to several human retinal dystrophies, including type 12 retinitis pigmentosa (RP12) and Leber congenital amaurosis (LCA) [1,2]. LCA is a rare inherited eye disease that appears at birth or early in life, affecting sight and showing other related clinical signs within the first few years of life [4]. Crumbs is a transmembrane protein that was initially identified in Drosophila. In this organism, Crumbs plays a crucial role in the establishment and maintenance of cell polarity during the development of several types of epithelial cells [5]. CRB1 is only present in the retina and brain [8]; CRB2 mRNA has been found in the retina, RPE/ choroid, brain and in other tissues at very low levels [9]; whereas CRB3 is expressed in different epithelium-derived cell types, including the retina [10,11,12]
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