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Raju V. S. Rajala, Brandt Wiskur, Masaki Tanito, Michelle Callegan, Ammaji Rajala; Diabetes Reduces Autophosphorylation of Retinal Insulin Receptor and Increases Protein-Tyrosine Phosphatase-1B Activity. Invest. Ophthalmol. Vis. Sci. 2009;50(3):1033-1040. doi: 10.1167/iovs.08-2851.
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purpose. Protein-tyrosine phosphatase-1B (PTP1B) has been implicated in the negative regulation of insulin signaling. The expression, activity, and functional role of PTP1B in the retina are unknown. In this study, the authors examined the relationship between the retinal insulin receptor (IR) and PTP1B in normal and diabetic mouse retinas.
methods. IR and PTP1B localization was examined by immunohistochemistry. The activation of IR was analyzed using specific antibodies against phosphotyrosine. PTP1B activity was determined in anti–PTP1B immunoprecipitates. Glutathione-S-transferase fusion proteins containing wild-type and catalytically inactive mutant PTP1B was used to study the interaction between IR and PTP1B. Anti–IR immunoprecipitates and the cytoplasmic domain of purified IR were incubated in the presence of ATP, and the autophosphorylation of IR with antiphosphotyrosine antibody was analyzed.
results. Immunohistochemical analysis of PTP1B shows that it is predominantly expressed in nonphotoreceptor layers of the retina, though it is clearly expressed in the inner segments of the rod photoreceptors. The IR is predominately expressed in rod inner segments. Biochemical analysis of rod outer segments indicates the presence of IR and PTP1B. Retinal IR exhibits a high level of basal autophosphorylation, and this autophosphorylation is reduced in diabetic mouse retinas. In vitro, PTP1B is able to dephosphorylate the autophosphorylated IR. Substrate mutant-trap results indicate a stable interaction between IR and PTP1B. Further, PTP1B activity was increased in diabetic mouse retinas.
conclusions. These studies indicate that diabetes reduces the autophosphorylation of retinal IR and increased PTP1B activity. Further, PTP1B regulates the state of IR phosphorylation in the retina.
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