Clone 9 liver cells incubated under aerobic conditions with the proteasome inhibitors lactacystin, clastro lactacystin beta-lactone, and tri-leucine vinyl sulfone, in the absence of an obvious oxidative challenge, underwent oxidative protein modifications, such as loss of solubility, formation of aggregates (predominantly by disulfide bridges), and increased carbonyl formation, similar to those seen with hydrogen peroxide treatment. These alterations were accompanied by modification of cell morphology and loss of cell viability. Remarkably, almost all of these modifications were prevented when cell incubation with proteasome inhibitors was performed under a 3% oxygen atmosphere instead of the 21% oxygen routinely used in cell culture experiments. Our results suggest an oxygen-dependent mechanism for the protein oxidation, protein aggregation, cellular dysfunction, and apoptosis induced by proteasome inhibitors.