Project Summary/Abstract Neurodegeneration is ultimately the main driver of vision loss in retinal diseases including Diabetic retinopathy (DR) and glaucoma. Neuronal cell death and glial activation and the associated neuroinflammation are widely recognized as central aspects of the pathophysiology of these diseases; however very few treatment options exists to control these mechanisms involved in early stages of these diseases. While anti-VEGF is a powerful new treatment option for the late stages of DR that are proliferative diabetic retinopathy and macular edema, there are still no therapies available to prevent early alterations of the neuroretina. αA-crystallin is a chaperone protein, whose protective function, is highly regulated by its phosphorylation on the T148 residue. There is a fundamental gap in our knowledge regarding the molecular mechanisms by which it promotes the survival of retinal cells and how those mechanisms are controlled by its phosphorylation on T148. The long-term goal of this research is to determine how the protective potential of αA-crystallin can be harnessed to reduce or prevent retinal neurodegneration. The objective of this project is to identify the molecular mechanisms by which the function of αA-crystallin protein is modulated by its phosphorylation on T148. Emerging from our preliminary data and previously published work, the central hypothesis guiding this project is that enhancement of αA-crystallin function through positive modulation of its phosphorylation on T148 residue can prevent or reduce DR pathophysiology. The rationale for the proposed research is that enhancement of this intrinsic protective pathway has demonstrated promise for the modulation of metabolic stress-induced neuroinflammation and neurodegeneration. This hypothesis will be tested by pursuing 2 specific aims: i) Determine the cell specific mechanisms of the therapeutic potential of the functionally enhanced αA-crystallin T148D against DR, and ii) Determine the mechanisms of regulation of phosphorylation of αA-crystallin on T148. Both aims will be concurrently pursued and will support each other, as aim 1 will establish the therapeutic potential of the functionally enhanced mutated protein and its cell-specific impact, while aim 2 will identify the cell-specific mechanisms of regulation of this phosphorylation and how they are affected by diabetes. Altogether this research project will identify and characterize the mechanisms of regulation of the neuroprotective function of αA-crystallin and how they can be targeted in neurodegenerative diseases, potentially leading to the development of approaches for utilizing this intrinsic protective pathway to promote retinal cell survival and control neuroinflammation in chronic neurodegenerative conditions and, thereby, preserve visual function. Key terms AlphaA-crystallins, diabetic retinopathy, intrinsic protective mechanisms, phosphorylation, kinase