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R. Rajala, M.D. Chan, H. Tomita, M.E. McClellan; Adapter Grb14 in Retinal Insulin Receptor Signaling . Invest. Ophthalmol. Vis. Sci. 2005;46(13):5129.
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Purpose: Cells of bovine and rat retina contain high affinity receptors for insulin. However, little research has been done on these receptors. We have previously demonstrated the light–induced tyrosine phosphorylation of retinal insulin receptor (IRß) and localized the light effect to the photoreceptor neurons independent of insulin secretion. The purpose of this study is to identify binding partners to retinal IRß, since mutation in either IRß autophosphorylation sites or its binding partner Dock in Drosophila has a severe phenotype of photoreceptor axonal misguidance. Methods: We used yeast two–hybrid assay of protein–protein interaction in the yeast Saccharomyces cerevisiae to identify binding partners to IRß from bovine retinal cDNA library, and identified Grb14, a novel member of the Grb7 family. Anti–peptide antibodies were generated to Grb14 and used to study its expression in various fractions of the retina. Rod outer segments (ROS) were prepared from the light–and dark–adapted rats. Immunohistochemistry was performed on light–and dark–adapted rat retina sections. Results: Yeast two–hybrid screening identified Grb14 as an IRß binding partner. Western blot analysis indicated the presence of Grb14 in ROS, cytosol, and nucleus. We have found more Grb14 bound to light–adapted ROS membranes than to dark–adapted membranes. Immunohistochemical studies indicate that Grb14 is localized in the outer nuclear layer in the dark and in ROS in the light. Conclusions: These findings suggest that Grb14 is a major binding partner of IRß in the retina and that its intracellular localization is light–dependent.
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