SUMMARY: Vision loss from age-related macular degeneration (AMD) is a major, expanding problem due to the aging population. While treatment is available for intermediate AMD and the advanced wet form, no prevention or treatment is available for early AMD. While cellular heterogeneity can help tissue adapt to stress, degenerative heterogeneity from aging is associated with disease. The current perspective is that RPE (retinal pigmented epithelium) degeneration is a central, chronic process that inevitably leads to cell death. Thus, prior studies have focused on the impact of RPE cell death on AMD pathobiology. However, a subset of RPE cells exhibit features of epithelial-mesenchymal transition (EMT), an adaptive process that allows cells to survive a harsh environment. RPE heterogeneity has been recognized for some time, but the impact of disease driving cell subsets on AMD pathogenesis has not been considered. Lysosomal dysfunction is clearly involved in several age-related diseases including AMD, but the impact of heterogeneity of lysosomal impairment, which can cause disease, on RPE cellular function or heterogeneity has not been considered. This proposal is a departure from the norm because it proposes that RPE heterogeneity including a subset of cells with EMT in early AMD is initiated by heterogeneity of lysosomal clearance defects. RPE in EMT are resistant to death, and thus, could drive AMD pathobiology for an extended period. On the other hand, since EMT can be reversed, RPE in EMT are a compelling treatment target because its vision preserving functions can be restored. Lysosomal function and autophagy are modulated by the master regulator transcription factor EB (TFEB). Our laboratory has demonstrated reduced nuclear TFEB staining in the RPE of early AMD globes compared to age-matched controls, thus identifying TFEB as a potential therapeutic target for AMD. Given the possibility that heterogeneity of lysosomal dysfunction drives RPE heterogeneity, from EMT to degeneration, the objective of this proposal is to determine the impact of restoring lysosomal function and autophagy on the AMD-associated RPE heterogeneity in our validated Cryba1 cKO rodent model of lysosomal impairment that develops a dry AMD-like phenotype, and in human-iPSC derived RPE cells. The central hypothesis of “rejuvenating lysosomal and autophagic function by activating TFEB will maintain RPE cell function and prevent pathologic RPE heterogeneity including EMT, in early AMD” will be addressed with the following aims: Aim 1: Test the extent that TFEB activation rejuvenates lysosomal/autophagy function, and modulates RPE heterogeneity and RPE cell health. Aim 2: Test the degree that TFEB activation in the RPE of Cryba1 cKO mice rescues its AMD-like phenotype. Aim 3: Test the degree that lysosomal heterogeneity leads to RPE heterogeneity in AMD. Successful completion of this proposal will establish the novel concept of lysosome/autophagy dysfunction as a key driver of ...