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Concepcion Lillo, Saúl Herranz, Antonio E. Paniagua, Almudena Velasco, Juan M. Lara, José Aijón; Characterization Of The Crb3 Protein In The Mouse Retina. Invest. Ophthalmol. Vis. Sci. 2011;52(14):923.
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The CRB protein family (Crumbs like proteins: CRB1, CRB2 and CRB3) are transmembrane proteins implicated in the maintenance of the apico-basal polarity in epithelial cells, and they have been found in the retina of mammals. CRB1 is localized in the subapical region of the outer limiting membrane (OLM), and its mutations are associated to human retinal dystrophies such as Retinitis Pigmentosa and Leber Congenital Amaurosis. CRB2 and CRB3 seem to be also present in the OLM, but there are only scarce studies which analyse its role in the retina of mammals or in degenerative diseases. Some authors have proposed that CRB3 and/or CRB2 might be localized in the OPL, and that CRB3 could be in the photoreceptors’ connecting cilium, since it is known that this protein is involved in the ciliogenesis of some epithelial cells. Our purpose is to deeply analyse the precise localization of CRB3 in the mouse retina during development and in adults as a first approximation to study its role in the retina.
Retinas from P0, P4 and adult (P90) wild type mice were used for western blot and double immunofluorescence analyses for CRB3 and a batch of antibodies to label different retinal cell types, such as photoreceptor cells (opsin, recoverin and PNA), bipolar cells (PKCalpha), horizontal cells (CB), amacrine cells (CB, CR, TH and GABA) or to label synaptic profiles (VGLUT1, synaptophysin and bassoon). We have also used an antibody that labels acetylated tubulin to identify the connecting cilium of the photoreceptor cells and beta-catenin to label the adherens junctions.
CRB3 is present in the mouse retina at all studied stages. During development, the labelling for CRB3 appears when the synaptic layers and rods’ segments start to develop and it is co-localized with beta-catenin, a protein present in the adherens junctions. In adult mice, CRB3 is located all along the rods’ inner segments, especially in the connecting cilium area, and in the rods’ synaptic terminals. It is also detected in the synaptic terminals of some subpopulations of amacrine cells and in rods bipolar cells, but it does not co-localize with beta-catenin in any case.
Since CRB3 expression coincides with the development of the plexiform layers and the photoreceptors’ inner segments, CRB3 might be implicated in the correct formation of the synaptic contacts and the inner segments, where it could be necessary for the ciliogenesis of the connecting cilium. In the adult retina, CRB3 could play a role in the maintenance of the connecting cilium and of the synapses in both retinal plexiform layers.
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