Abstract
Purpose :
The polarized retinal pigment epithelium (RPE) performs many functions to ensure photoreceptor health and vision including retinoid recycling, delivery of nutrients to the retina, and daily phagocytosis of outer segments. These functions are made possible by a complex, interconnected, and efficient endosomal trafficking system regulated by approximately 60 Ras analog in brain (Rab) proteins. Studies in other organ systems have shown that specific Rab proteins such as Rab11a preserve epithelial polarity by delivering critical cargo to cell-cell junctions. Little is currently understood about how Rab proteins maintain RPE structural and functional integrity. Here, we investigated the role of Rab11a in regulating RPE junctional stability and examined how this is altered in models of macular degeneration.
Methods :
Expression and localization of Rab11 and junctional proteins (connexin 43, P-cadherin, ZO1) were measured by immunostaining and immunobotting in polarized primary porcine RPE cultures and Abca4-/- mice, a model for Stargardt inherited macular dystrophy. Active Rab11 was measured using prenylation assays. Live imaging of Rab11 trafficking was
was performed using spinning disc confocal microscopy. The integrity of RPE cell-cell junctions in healthy and disease models was established using fluorometric dye transfer assays.
Results :
Our data show that a pathological accumulation of lipofuscin bisretinoids in Abca4-/- mice leads to abnormal activation of Rab11a. This not only induces endosomal abnormalities, but also impairs trafficking of connexin 43 to gap junctions. Expansion of Rab11a+ endosomes leads to increased endocytosis of Cx43, E-cadherin, and other junctional proteins, which destabilizes RPE cell-cell junctions. Decreasing Rab prenylation using small molecule drugs restores endosome dynamics and junctional integrity in the RPE.
Conclusions :
Our data show that Rab11a is essential for maintaining the polarity of the RPE through the precise delivery of junctional proteins. Given that RPE dedifferentiation and atrophy are poorly understood features of macular degenerations, these studies suggest that Rab regulatory systems serve as a novel therapeutic target for macular degeneration.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.