Mapping sites of carboxymethyllysine modification on proteins reveals its consequences for proteostasis and cell proliferation

Abstract

SummaryPosttranslational mechanisms play a key role in modifying the abundance and function of cellular proteins. Among these, modification by advanced glycation end products (AGEs) has been shown to accumulate during aging and age-associated diseases but specific protein targets and functional consequences remain largely unexplored. Here, we devised a proteomic strategy to identify specific sites of carboxymethyllysine (CML) modification, one of the most abundant AGEs. We identified over 1000 sites of CML modification in mouse and primary human cells treated with the glycating agent glyoxal. By using quantitative proteomics, we found that protein glycation triggers a proteotoxic response and directly affects the protein degradation machinery. We show that glyoxal induces cell cycle perturbation in primary endothelial cells and that CML modification reduces acetylation of tubulins and impairs microtubule dynamics. Our data demonstrate the relevance of AGE modification for cellular function and pinpoint specific protein networks that might become compromised during aging.HighlightsA peptide enrichment strategy allows mapping of CML modification in cells and tissuesCML modification competes with ubiquitination or acetylation of lysinesGlyoxal treatment destabilizes the 26S proteasomeGlyoxal arrests cell cycle and impairs microtubule dynamics via altering the tubulin code