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V. L. Bonilha, K. G. Shadrach, M. E. Rayborn, J. Wu, N. S. Peachey, I. Saotome, A. I. McClatchey, H. Sun, D. Ong, J. G. Hollyfield; Polarity Changes in the Retinal Pigment Epithelium (RPE) and Retina in an RPE-Ezrin Knockout Model. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1860.
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In the eye, ezrin (an actin-binding protein) has been localized to apical microvilli of Müller cells, RPE apical microvilli and basal infoldings. We have previously shown that ezrin plays an important role in establishing and maintaining RPE apical microvilli, basal infoldings, and apical Muller cells microvilli. However, our observations also showed that photoreceptors in ezrin-/- mice develop more slowly than in wild type littermates. To dissect the effects of ezrin absence in the RPE, we generated an RPE-specific ezrin knockout mouse model.
The RPE-ezrin deficient mice were generated by breeding ezrin flox/flox mice with transgenic mice using an RPE specific promoter to direct the expression of cre recombinase. Eyecups were fixed in 2% paraformaldehyde (PF) and 2.5% glutaraldehyde in 0.1M phosphate buffer and analyzed by transmission electron microscopy (TEM). Eyes of young adult RPE-Ez-/- mice and control littermates were also fixed in 4% PF and processed for cryosectioning and labeling with antibodies specific to RPE plasma membrane. Photoreceptor and RPE function were assessed by ERG.
TEM analysis indicates that RPE microvilli and basal infoldings are both formed, but appear abnormal ultrastructurally. The membranes of apical microvilli and basal infoldings, as well as of the microvilli cytoplasm were very electrondense. In addition, RPE microvilli were shorter than normal and disorganized. RPE-ezrin-/- mice expressed low levels of several apical plasma membrane proteins including Glut-1, CRALBP, and peropsin. ERG studies of RPE-ezrin-/- mice noted only minor changes under dark- and light-adapted conditions; all RPE-driven components were present, although at reduced levels.
Analysis of RPE-Ez-/- mice demonstrates that ezrin is required for the formation and maintenance of the RPE polarized phenotype and that ezrin plays a crucial role in the morphogenesis of both apical microvilli and basal infoldings.
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