Oxidative stress increases ubiquitin-protein conjugates in synaptosomes

SYNAPTOSOMES were incubated in the presence of FeSO4 to test the hypothesis that iron-catalyzed oxidative damage causes an increase in the ubiquitinat ion of synaptosomal proteins. Incubation with 10 or 50 mu M FeSO4 caused co ncentration-dependent increases in carbonyl groups (an indication of protei n oxidation) and ubiquitinated proteins (determined by probing Western blot s with a monoclonal antibody to ubiquitin). Differences in protein ubiquiti nation occurred within 5 min of incubation, indicating a rapid response to oxidative stress. Results of experiments with MG-132, an inhibitor of the d egradation of ubiquitinated proteins, suggested that oxidative damage stimu lated ubiquitination rather than inhibited degradation of ubiquitinated pro teins. The data are consistent with the hypothesis that synaptic terminals utilize the ubiquitin/ proteasome proteolytic pathway to degrade oxidativel y damaged proteins. (C) 1999 Lippincott Williams & Wilkins.