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Y.-W. Peng, M. Zallocchi, W. Wang, D. Cosgrove; Basement Membrane Usherin May Play Important Roles in Light-Driven Rod Photoreceptor Protein Translocation. Invest. Ophthalmol. Vis. Sci. 2008;49(13):165. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Usher syndrome type IIa is the most common of the Usher syndromes, and thus the leading single genetic cause of combined deafness and blindness in the world. The gene responsible encodes two different protein isoforms called usherin. The long isoform is found in rod photoreceptors. The short isoform is a basement membrane protein. Whether one or both of these isoforms is functionally important in the retina is not known. Here we explore the functional importance of the basement membrane (short) isoform of usherin.
Immunohistochemistry, RT-PCR, immunoblotting and Isolated retinal cultures were used to study the localization of usherin short form and integrin receptors as well as the light-induced protein translocation.
Using a LN domain specific antibody, usherin short form is localized at the basement membrane of retinal pigment epithelium (RPE). The staining of usherin short form on this location is validated by its absence from usherin knock-out mice (Liu et al., PNAS 2007). The presence of usherin short form at the RPE is confirmed by RT-PCR and western blotting from RPE in eye cup and isolated retina and immunogold analysis. Previously, we have reported that the short isoform of usherin binds integrin α1β1 receptors at the RPE. Both usherin hypomorph and integrin α1-null mouse models show late-onset slow progressive retinal degeneration and markedly slowed protein translocation in rods (Peng et al., ARVO Abstract 2007). Here, we further show that usherin short form co-localizes with both integrin receptors at the basement membrane of RPE placing them at the appropriate location for interaction. To study the function of their interaction we have used cultured isolated retinas removed from the RPE. If the cause of delayed protein translocation emanates from defective ciliary function, where the long isoform of usherin is located, retinas isolated from usherin hypomorph without RPE should still show a delayed rod protein translocation. Contrary to this, in cultured retinas from these mutants we observed transducin α and arrestin are translocated at a rate similar to that observed in wild type mice, suggesting there is no defect in protein translocation in isolated usherin hypomorph mouse retina without RPE.
The short isoform of usherin is present in the basement membrane of Bruchs layer, at the basal aspect of the RPE. Protein translocation defects in usherin hypomorph and integrin α1-null rods are likely due to influences coming from the RPE and usherin-integrin α1 interaction on RPE cells may be required for normal functioning of protein translocation in rods.
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