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Thomas W Gardner, Mandy K Losiewicz, Steven F Abcouwer, Patrice E Fort, RC; Tissue-Specific Regulatory Mechanisms of Retinal Protein Synthesis by Akt and mTOR pathways. Invest. Ophthalmol. Vis. Sci. 2014;55(13):1031.
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To test the hypothesis that Akt and mTOR regulate retinal protein synthesis, with an emphasis on the impact of glycemic control and insulin signaling. We previously demonstrated that diabetes reduces retinal protein synthesis similarly to changes in other insulin-sensitive tissues but the mechanisms involved have remained undefined.
Total mTOR activity was determined by KLISA™ mTOR Activity Assay while specific mTORC1 and mTORC2 activity were assessed by analysis of raptor or rictor associated mTOR levels and phosphorylation on Ser2481. Further analysis of mTORC1 and mTORC2 complex activity was performed by measure of the downstream effectors: 4E-BP1, phospho-4EBP1 (Thr37/46), S6 ribosomal protein, phospho-S6 (Ser235/236), phospho-S6 (Ser240/244) and total and p-PKCα (Ser657). The regulation of retinal protein synthesis was further studied using ex vivo retinas in association with Akt isoform specific and mTORC1 and mTORC2 specific inhibitors. Subconjunctival insulin and systemic phloridzin treatments were used to manipulate retinal growth factor signaling and systemic glycemia, respectively.
Concomitant to the reduction in retinal protein synthesis, diabetes reduced total mTOR activity by 15% compared to controls, and both subconjunctival insulin and glucose reduction with phloridzin partially restored it. Akt 1/3 inhibitors and torin1 but not rapamycin reduced protein synthesis in retinas from control rats, and reduced mTORC2 but not mTORC1 activity. Unlike other insulin-sensitive tissues, diabetes reduced rictor- but not raptor-associated mTOR phosphorylation. Also, PKCα but not 4EBP1 or S6 ribosomal protein phosphorylation was reduced demonstrating that only mTORC2 activity is affected in the retina by diabetes.
These data show the impact of diabetes on retinal protein synthesis is mTORC1 independent but associated with reduced mTORC2, pointing out the uniqueness of retinal metabolism, and may lead to improved means to maintain retinal anabolic activity during diabetes.
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