Proteasome inhibitors induce mitochondria-independent apoptosis in human glioma cells

• Published in: FEBS letters Vol. 443; no. 2; pp. 181 - 186
• Main Authors: Kitagawa, Hiroyuki, Tani, Eiichi, Ikemoto, Hideyasu, Ozaki, Isao, Nakano, Atsuhisa, Omura, Satoshi
• Format: Journal Article
• Language: English
• Published: England Elsevier B.V 29-01-1999
• Subjects: AcLLNal; AcLLNal, acetyl-leucinyl-leucinyl-norleucinal; Animals; Apoptosis; Apoptosis - drug effects; Brain Neoplasms - pathology; Caspase 3; Caspases - metabolism; Cell Cycle Proteins; Cell Line; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Cyclins - metabolism; Cysteine Endopeptidases - drug effects; Cysteine Proteinase Inhibitors - pharmacology; Cyt c, cytochrome c; Cytochrome c; DiOC6, 3,3′-dihexyloxacarbocyanine iodide; DMEM, Dulbecco's modified Eagle's minimum essential medium; DTT, dithiothreitol; EDTA, ethylenediaminetetraacetic acid; EGTA, ethylene-glycol-bis(β-amino-ethylether)N,N′-tetraacetic acid; Enzyme Activation; FCS, fetal calf serum; Glioma; Glioma - pathology; HEPES, N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid; HRP, horseradish peroxidase; Humans; Lactacystin; mCICCP, carbonyl cyanide m-chlorophenylhydrazone; Microtubule-Associated Proteins - metabolism; Mitochondria - drug effects; Mitochondrial membrane potential; Multienzyme Complexes - drug effects; NP-40, Nonidet P-40; Nuclear Proteins; PARP, poly(ADP-ribose) polymerase; PMA, phorbol myristate acetate; PMSF, phenylmethylsulfonyl fluoride; Proteasome Endopeptidase Complex; Proto-Oncogene Proteins - metabolism; Proto-Oncogene Proteins c-mdm2; PVDF, polyvinylidene difluoride; SDS, sodium dodecylsulfate; TNF, tumor necrosis factor; Tumor Cells, Cultured; Tumor Suppressor Protein p53 - metabolism; Tumor Suppressor Proteins; ΔΨ m, mitochondrial membrane potential; AcLLNal; SDS, sodium dodecylsulfate; EDTA, ethylenediaminetetraacetic acid; Δ Ψ m, mitochondrial membrane potential; AcLLNal, acetyl-leucinyl-leucinyl-norleucinal; EGTA, ethylene-glycol-bis(β-amino-ethylether) N, N′-tetraacetic acid; Caspase-3; NP-40, Nonidet P-40; DTT, dithiothreitol; Cyt c, cytochrome c; HRP, horseradish peroxidase; Lactacystin; PVDF, polyvinylidene difluoride; mCICCP, carbonyl cyanide m-chlorophenylhydrazone; TNF, tumor necrosis factor; HEPES, N-2-hydroxyethylpiperazine- N′-2-ethanesulfonic acid; DMEM, Dulbecco's modified Eagle's minimum essential medium; Cytochrome c; PMSF, phenylmethylsulfonyl fluoride; Glioma; DiOC 6, 3,3′-dihexyloxacarbocyanine iodide; PMA, phorbol myristate acetate; PARP, poly(ADP-ribose) polymerase; Mitochondrial membrane potential; Apoptosis; FCS, fetal calf serum
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/S0014-5793(98)01709-8

The proteasome inhibitors lactacystin and AcLLNal induced p53-independent apoptosis in two human glioma cell lines, and the apoptosis was accompanied by up-regulation of immunoreactive wild-type p53, p21
Waf1, Mdm2, and p27
Kip1. Pretreatment with cycloheximide decreased the induction of cell death independently of p53 protein status, suggesting that the up-regulation of short-lived proteins is associated with proteasome inhibitor-induced apoptosis. Caspase-3-like proteases were activated in the proteasome inhibitor-mediated apoptosis, and the induction of cell death was inhibited more effectively in the presence of z-VAD.fmk than in the presence of Ac-DEVD.fmk, suggesting that caspases other than caspase-3 are involved. Nonetheless, there were no significant alterations in levels of immunoreactive Bcl-2, Bcl-x
L, Bax, Bad, and Bak, nor any evidence of cytochrome
c release into cytosol and dissipation of Δ
Ψ
m. Thus, the proteasome inhibitor-induced apoptosis is mediated by a mitochondria-independent mechanism, and the once activated caspase-3 does not cause the cytochrome
c release and the Δ
Ψ
m disruption.