Proteasome and Ribosome Ubiquitination in Retinal Pigment Epithelial (RPE) Cells in Response to Oxidized Low-Density Lipoprotein (OxLDL)

<b>Background/Objectives:</b> Oxidative stress plays a significant role in the development and progression of age-related macular degeneration (AMD). Retinal pigment epithelium (RPE) cells are specialized multifunctional cells indispensable for the maintenance of vision. The dysfunction and death of RPE cells in the macula characterize the onset and development of AMD. Of the various toxic agents that impact the health of the RPE, particular focus has been given to various forms of lipoproteins and their cytotoxic derivatives normally present in the retina. Oxidized low-density lipoprotein (OxLDL), derived from LDL in a pro-oxidative environment, is found adjacent to RPE cells as part of drusen, extracellular deposits that are a hallmark feature of AMD. OxLDL is a potent inflammatory agent and it has been implicated in cardiovascular and neurodegenerative conditions. The cellular molecular mechanisms triggered by OxLDL are only partially understood. The focus of this study was to characterize changes in the proteome of RPE cells after exposure to OxLDL, with a focus on the characterization and quantification of ubiquitinated proteins. <b>Methods:</b> Identification and quantification were performed with a high-resolution LC-MS/MS-based proteomics workflow after immune-enrichment for ubiquitinated peptides. <b>Results:</b> In total, out of the more than 1000 RPE ubiquitinated peptides quantified, OxLDL treatment caused a significant increase in ubiquitinated peptides compared to LDL and untreated cells. Principal component analysis (PCA) of the differentially ubiquitinated proteins (265) reduced the data complexity in two main groups of variables (proteins). <b>Conclusions:</b> Gene ontology enrichment analysis of the grouped proteins with the highest loading contribution to principal component 1 (PC1) and principal component 2 (PC2) revealed significant ubiquitination changes upon OxLDL treatment in proteins of the ubiquitin-proteasome system (UPS) responsible for proteasome-mediated catabolic processes and in protein members of the cellular translation machinery.