Immunosubunit β5i Knockout Suppresses Neovascularization and Restores Autophagy in Retinal Neovascularization by Targeting ATG5 for Degradation

This study identifies a functional role for β5i in RNV regulation. β5i deletion ameliorates RNV and restores autophagy by stabilizing ATG5. These results demonstrate the potential of β5i to serve as a bridge linking the immunoproteasome and autophagy.

Oxygen-induced retinopathy (OIR) was induced in wild-type (WT) and β5i knockout (KO) mouse pups on a C57BL/6J background. Proteasome catalytic subunit expression and proteasome activity were evaluated by quantitative real-time PCR (qPCR) and proteasome activity. Retinal vascular anatomy and neovascularization were characterized and quantified by retinal vascular flat-mount staining, fluorescence angiography, platelet endothelial cell adhesion molecule (PECAM) immunostaining, and hematoxylin and eosin staining. Correlation factors, including VEGF and ICAM-1, were detected by qPCR. Autophagy was examined by transmission electron microscopy (TEM). Autophagy biomarkers, including LC3, P62, ATG5, and ATG7, were measured by immunostaining and immunoblotting. The protein interaction between β5i and ATG5 was detected by immunoprecipitation.

To investigate the functional role of immunoproteasome subunit β5i in pathologic retinal neovascularization (RNV) and its ability to link the immunoproteasome and autophagy.

We observed that β5i had the greatest effect in WT OIR mice. Fundus fluorescence angiography, retinal flat-mount staining, and PECAM staining revealed that pathologic RNV decreased in β5i KO OIR mice compared with WT OIR mice. Concurrently, TEM, immunostaining, and immunoblotting showed that autophagy was induced in β5i KO OIR mice compared to WT OIR mice through increases in autophagosome and LC3 expression and a decrease in P62. Mechanistically, β5i interacted with ATG5 and promoted its degradation, leading to autophagy inhibition and pathogenic RNV. Conclusions: This study identifies a functional role for β5i in RNV regulation. β5i deletion ameliorates RNV and restores autophagy by stabilizing ATG5. These results demonstrate the potential of β5i to serve as a bridge linking the immunoproteasome and autophagy.