A role for βA3/A1-crystallin in type 2 EMT of RPE cells occurring in dry age-related Macular degeneration

P_URPOSE. The RPE cells have a major role in the development of dry age-related macular degeneration (AMD). We present novel evidence that bA3/A1-crystallin, encoded by the Cryba1 gene, a protein known to be important for lysosomal clearance in the RPE, also has a role in epithelial-to-mesenchymal transition (EMT) of RPE cells. M_ETHODS. RPE from dry AMD globes, genetically engineered mice lacking Cryba1 globally or specifically in the RPE, spontaneous mutant rats (Nuc1) with a loss-of-function mutation in Cryba1, and the melanoma OCM3 cell line were used. Spatial localization of proteins was demonstrated with immunofluorescence, gene expression levels were determined by quantitative PCR (qPCR), and protein levels by Western blotting. Cell movement was evaluated using wound healing and cell migration assays. Co-immunoprecipitation was used to identify binding partners of bA3/A1-crystallin. R_ESULTS. bA3/A1-crystallin is upregulated in polarized RPE cells compared to undifferentiated cells. Loss of bA3/A1-crystallin in murine and human RPE cells resulted in upregulation of Snail and vimentin, downregulation of E-cadherin, and increased cell migration. bA3/A1-crystallin binds to cortactin, and loss of bA3/A1-crystallin resulted in increased P-cortactin^Y421. The RPE from AMD samples had increased Snail and vimentin, and decreased E-cadherin, compared to age-matched controls. C_ONCLUSIONS. We introduced a novel concept of dry AMD initiation induced by lysosomal clearance defects in the RPE and subsequent attempts by RPE cells to avoid the resulting stress by undergoing EMT. We demonstrate that bA3/A1-crystallin is a potential therapeutic target for AMD through rejuvenation of lysosomal dysfunction and potentially, reversal of EMT.