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 microM) 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 microM) and higher dose MG132 (40 microM). 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 microM) 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.
Leak
Rehana K
RK
Department of Neurology, Pittsburgh Institute of Neurodegenerative Diseases, University of Pittsburgh, Pittsburgh, Pennsylvania, USA. leakrk@upmc.edu
Zigmond
Michael J
MJ
Liou
Anthony K F
AK
D000220:Adaptation, Biological / Q000187:drug effects*; D018663:Adrenergic Agents / Q000494:pharmacology; D000818:Animals; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D002374:Catalase / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D004298:Dopamine / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects* / Q000502:physiology*; D016627:Oxidopamine / Q000494:pharmacology; D016716:PC12 Cells / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D051381:Rats; D013482:Superoxide Dismutase / Q000502:physiology*; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism
D018663:Adrenergic Agents; D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; D007976:Leupeptins; D034741:RNA, Small Interfering; D019328:Buthionine Sulfoximine; D016627:Oxidopamine; D002374:Catalase; D014446:Tyrosine 3-Monooxygenase; D013482:Superoxide Dismutase; D046988:Proteasome Endopeptidase Complex; D005978:Glutathione; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; D004298:Dopamine