Oxidation of HRas cysteine thiols by metabolic stress prevents palmitoylation in vivo and contributes to endothelial cell apoptosis

Here we demonstrate a new paradigm in redox signaling, whereby oxidants resulting from metabolic stress directly alter protein palmitoylation by oxidizing reactive cysteine thiolates. In mice fed a high‐fat, high‐sucrose diet and in cultured endothelial cells (ECs) treated with high palmitate and high glucose (HPHG), there was decreased HRas palmitoylation on Cys181/184 (61±24% decrease for cardiac tissue and 38±7.0% in ECs). This was due to oxidation of Cys181/184, detected using matrix‐assisted laser desorption/ionization time of flight (MALDI TOF)‐TOF. Decrease in HRas palmitoylation affected its compartmentalization and Ras binding domain binding activity, with a shift from plasma membrane tethering to Golgi localization. Loss of plasma membrane‐bound HRas decreased growth factor‐stimulated ERK phosphorylation (84±8.6% decrease) and increased apoptotic signaling (24±6.5‐fold increase) after HPHG treatment that was prevented by overexpressing wild‐type but not C181/184S HRas. The essential role of HRas in metabolic stress was made evident by the similar effects of expressing an inactive dominant negative N17‐HRas or a MEK inhibitor. Furthermore, the relevance of thiol oxidation was demonstrated by overexpressing manganese superoxide dismutase, which improved HRas palmitoylation and ERK phosphorylation, while lessening apoptosis in HPHG treated ECs.—Burgoyne, J. R., Haeussler, D. J., Kumar, V., Ji, Y., Pimental, D. R., Zee, R. S., Costello, C. E., Lin, C., McComb, M. E., Cohen, R. A., Bachschmid, M. M. Oxidation of HRas cysteine thiols by metabolic stress prevents palmitoylation in vivo and contributes to endothelial cell apoptosis. FASEB J. 26, 832–841 (2012). www.fasebj.org