Adaptation to chronic MG132 reduces oxidative toxicity by a CuZnSOD‐dependent mechanism

To study whether and how cells adapt to chronic cellular stress, we exposed PC12 cells to the proteasome inhibitor MG132 (0.1 μM) for 2 weeks and longer. This treatment reduced chymotrypsin‐like proteasome activity by 47% and was associated with protection against both 6‐hydroxydopamine (6‐OHDA; 100 μM) and higher dose MG132 (40 μM). Protection developed slowly over the course of the first 2 weeks of exposure and was chronic thereafter. There was no change in total GSH levels after MG132. Buthionine sulfoximine (100 μM) reduced GSH levels by 60%, but exacerbated 6‐OHDA toxicity to the same extent in both MG132‐treated and control cells and failed to reduce MG132‐induced protection. Chronic MG132 resulted in elevated antioxidant proteins CuZn superoxide dismutase (SOD; +55%), MnSOD (+21%), and catalase (+15%), as well as chaperone heat‐shock protein 70 (+42%). Examination of SOD enzyme activity revealed higher levels of CuZnSOD (+40%), with no change in MnSOD. We further assessed the mechanism of protection by reducing CuZnSOD levels with two independent siRNA sequences, both of which successfully attenuated protection against 6‐OHDA. Previous reports suggested that artificial over‐expression of CuZnSOD in dopaminergic cells is protective. Our data complement such observations, revealing that dopaminergic cells are also able to use endogenous CuZnSOD in self‐defensive adaptations to chronic stress, and that they can even do so in the face of extensive GSH loss.