title: Protein carbonylation and aggregation precede neuronal apoptosis induced by partial glutathione depletion.
abstract: While the build-up of oxidized proteins within cells is believed to be toxic, there is currently no evidence linking protein carbonylation and cell death. In the present study, we show that incubation of nPC12 (neuron-like PC12) cells with 50 μM DEM (diethyl maleate) leads to a partial and transient depletion of glutathione (GSH). Concomitant with GSH disappearance there is increased accumulation of PCOs (protein carbonyls) and cell death (both by necrosis and apoptosis). Immunocytochemical studies also revealed a temporal/spatial relationship between carbonylation and cellular apoptosis. In addition, the extent of all three, PCO accumulation, protein aggregation and cell death, augments if oxidized proteins are not removed by proteasomal degradation. Furthermore, the effectiveness of the carbonyl scavengers hydralazine, histidine hydrazide and methoxylamine at preventing cell death identifies PCOs as the toxic species. Experiments using well-characterized apoptosis inhibitors place protein carbonylation downstream of the mitochondrial transition pore opening and upstream of caspase activation. While the study focused mostly on nPC12 cells, experiments in primary neuronal cultures yielded the same results. The findings are also not restricted to DEM-induced cell death, since a similar relationship between carbonylation and apoptosis was found in staurosporine- and buthionine sulfoximine-treated nPC12 cells. In sum, the above results show for the first time a causal relationship between carbonylation, protein aggregation and apoptosis of neurons undergoing oxidative damage. To the best of our knowledge, this is the first study to place direct (oxidative) protein carbonylation within the apoptotic pathway.
authors.affiliation: Department of Cell Biology and Physiology, University of New Mexico-Health Sciences Center, Albuquerque, NM, USA.
authors.forename: Anushka
authors.initials: A
authors.lastname: Dasgupta
authors.forename: Jianzheng
authors.initials: J
authors.lastname: Zheng
authors.forename: Oscar A
authors.initials: OA
authors.lastname: Bizzozero
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D005978:Glutathione / Q000172:deficiency* / Q000378:metabolism; D002479:Inclusion Bodies / Q000378:metabolism*; D051379:Mice; D009410:Nerve Degeneration / Q000378:metabolism / Q000473:pathology; D016716:PC12 Cells; D061251:Primary Cell Culture; D050050:Protein Carbonylation / Q000502:physiology*; D051381:Rats
chemical_list: D005978:Glutathione