The Permeability Transition Pore Complex: A Target for Apoptosis Regulation by Caspases and Bcl-2–related Proteins

Marzo, Isabel; Brenner, Catherine; Zamzami, Naoufal; Susin, Santos A.; Beutner, Gisela; Brdiczka, Dieter; Rémy, R.; Xie, Zhihua; Reed, John C.; Kroemer, Guido

doi:10.1084/jem.187.8.1261

Cited by 641 publications

(525 citation statements)

References 58 publications

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“…Interestingly, mitochondrial permeability transition can lead to both apoptosis and necrosis (Lemasters et al, 1998). CKMT1 is located in the intermembrane space and also interacts with VDAC -ANT complexes (Marzo et al, 1998). We suggest that CKMT1 might induce apoptosis through specialised systems such as the PTP in OSCCs.…”

Section: Discussionmentioning

confidence: 85%

Ubiquitous mitochondrial creatine kinase downregulated in oral squamous cell carcinoma

et al. 2006

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In this study, we performed two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionisation time of fly mass spectrometry to identify the protein(s) associated with the development of oral squamous cell carcinomas (OSCCs) by comparing patterns of OSCC-derived cell lines with normal oral keratinocytes (NOKs), and found that downregulation of ubiquitous mitochondrial creatine kinase (CKMT1) could be a good candidate. Decreased levels of CKMT1 mRNA and protein were detected in all OSCC-derived cell lines examined (n ¼ 9) when compared to those in primary normal oral keratinocytes. Although no sequence variation in the coding region of the CKMT1 gene with the exception of a nonsense mutation in exon 8 was identified in these cell lines, we found a frequent hypermethylation in the CpG island region. CKMT1 expression was restored by experimental demethylation. In addition, when we transfected CKMT1 into the cell lines, they showed an apoptotic phenotype but no invasiveness. In clinical samples, high frequencies of CKMT1 downregulation were detected by immunohistochemistry (19 of 52 (37%)) and quantitative real-time RT -PCR (21 of 50 (42%)). Furthermore, the CKMT1 expression status was significantly correlated with tumour differentiation (Po0.0001). These results suggest that the CKMT1 gene is frequently inactivated during oral carcinogenesis and that an epigenetic mechanism may regulate loss of expression, which may lead to block apoptosis.

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Section: Discussionmentioning

confidence: 85%

Ubiquitous mitochondrial creatine kinase downregulated in oral squamous cell carcinoma

et al. 2006

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“…SUDHL4 cells exhibited a relatively higher GSH concentration compared with HL60 cells and could account for the greater intensity of PK11195-induced CMH 2 DCF fluorescence in HL60 cells (approximately one log fold). Bcl-2 transfection was not found to attenuate the generation of ROS in K562 cells, nor modulate cellular GSH content; however, Bcl-2 could conceivably antagonize ROS-induced permeability transition via its well described stabilizing effect on the permeability transition pore complex, the gating of which is regulated by the redox state of critical vicinal thiols (Costantini et al, 1996(Costantini et al, , 2000Marzo et al, 1998). This would be consistent with our finding that PK11195 can induce mitochondrial swelling in HL60 mitochondria, but not in SUDHL4 mitochondria which hyperexpress Bcl-2 due to t(14;18).…”

Section: Discussionmentioning

confidence: 96%

Bcl-2 resistant mitochondrial toxicity mediated by the isoquinoline carboxamide PK11195 involves de novo generation of reactive oxygen species

et al. 2001

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SummaryResistance to apoptosis is a major obstacle preventing effective therapy for malignancy. Mitochondria localized anti-death proteins of the Bcl-2 family play a central role in inhibiting apoptosis and therefore present valid targets for novel therapy. The peripheral benzodiazepine receptor (PBR) shares a close physical association with the permeability transition pore complex (PTPC), a pivotal regulator of cell death located at mitochondrial contact sites. In this study we investigated the cytotoxicity of the PBR ligand, PK11195, in the micromolar concentration range. PK11195 induced antioxidant inhibitable collapse of the inner mitochondrial membrane potential (∆Ψ m ) and mitochondrial swelling in HL60 human leukaemia cells, but not in SUDHL4 lymphoma cells (which exhibited a higher level of reduced glutathione and relative tolerance to chemotherapy or pro-oxidant induced ∆Ψ m dissipation). PK11195 induced the production of hydrogen peroxide that was not inhibited by Bcl-2 transfection, nor depletion of mitochondrial DNA. ROS production was however blocked by protonophore, implicating a requirement for ∆Ψ m . Our findings suggest that PK11195-induced cytotoxicity relies upon Bcl-2 resistant generation of oxidative stress; a process only observed at concentrations several orders of magnitude higher that required to saturate its receptor. 1397-1404 © 2001 Cancer Research Campaign doi: 10.1054/ bjoc.2001.1788, available online at http://www.idealibrary.com on http://www.bjcancer.com methyl-N-methyl-N-(1-methylpropyl)-isoquinoline carboxamide (PK11195), carbonylcyanide m-chlorophenylhdrazone (CCCP), propidium iodide and all cell culture reagents were purchased from Sigma-Aldrich Ltd, UK. FITC-conjugated mouse antihuman Bcl-2 monoclonal antibody, and the Dako intrastain fixation/permeabilization kit were purchased from Dako, UK. Cell culture and treatmentsHL60, KG1A, K652 cells stably transfected with the Bcl-2 gene (K562 B4) or vector only (K562 N2) (kindly provided by Dr DE Banker and Dr F Applebaum, The Fred Hutchinson Cancer Research Center, USA), and SUDHL4 lymphoma cell lines, were maintained in exponential suspension cultures in RPMI 1640 medium supplemented with 10% fetal calf serum, 5 mM glutamine, 100 µg ml -1 streptomycin and 100 U ml -1 penicillin. Cells were grown in a humidified atmosphere of 5% CO 2 /95% air at 37˚C. PK11195 was dissolved in ethanol at a stock concentration of 8.7 mg ml -1 , and added to cells at a 75 µM final concentration for 4 hours; vehicle alone was also used in medium. To test the effect of an antioxidant on PK11195 activity, cells were treated with 10 mM N-acetylcysteine for 45 minutes prior to treatment with PK11195.Cells were incubated with VP16, or ara-C at 10 µM final concentration for 24 hours, or with the thiol crosslinking prooxidant, diamide, at 100 µM concentration for 24 hours. The protonophore CCCP was used to dissipate ∆Ψ m by treating cells at a concentration of 10 µM for 15 minutes prior to treatment with PK11195. Depletion of mitochondrial DNAKG1A cells ...

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“…(2) Direct prevention of cytochrome c release and subsequent inhibition of an increased production of oxygen radicals by the respiratory chain [41]. (3) Stabilization of the mitochondria by preventing dissipation of the mitochondrial membrane potential ⌬⌿ m , e.g., by facilitating proton exchange or by blocking permeability transition [42,43]. (4) The inhibition of the release of additional mitochondrial proapoptotic factors, such as AIF, to prevent alternative death pathways [9].…”

Section: Discussionmentioning

confidence: 99%

Differential Effects of Bcl-2 on Cell Death Triggered under ATP-Depleting Conditions

Single

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Nicotera

2001

Experimental Cell Research

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The intracellular ATP concentration decides on the onset of either apoptosis or necrosis in Jurkat cells exposed to death stimuli. Bcl-2 can block apoptotic demise, which occurs preferably under conditions of high cellular ATP levels. Here, we investigated the effects of Bcl-2 on the necrotic type of cell demise that prevails under conditions of energy loss. ATP levels were modulated by using mitochondrial inhibitors, such as rotenone or S-nitrosoglutathione, in medium either lacking glucose or supplemented with glucose to stimulate glycolytic ATP generation. Under conditions of ATP depletion, staurosporine (STS) induced >90% necrosis in vector control-transfected cells, whereas bcl-2-transfected cells were protected. Thus, the antiapoptotic protein Bcl-2 can reduce the overall amount of cell death in ATP-depleted cells regardless whether it occurs by apoptosis or necrosis. Cytochrome c release, normally preceding STS-induced necrosis, was also inhibited by Bcl-2. However, Bcl-2 did not prevent an initial STS-induced drop of the mitochondrial membrane potential (⌬⌿ m ). Therefore, the mechanisms whereby Bcl-2 prevents cell death and favors retention of cytochrome c in the mitochondria require neither the maintenance of mitochondrial ⌬⌿ nor the maintenance of normal ATP levels.

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The Permeability Transition Pore Complex: A Target for Apoptosis Regulation by Caspases and Bcl-2–related Proteins

Cited by 641 publications

References 58 publications

Ubiquitous mitochondrial creatine kinase downregulated in oral squamous cell carcinoma

Ubiquitous mitochondrial creatine kinase downregulated in oral squamous cell carcinoma

Bcl-2 resistant mitochondrial toxicity mediated by the isoquinoline carboxamide PK11195 involves de novo generation of reactive oxygen species

Differential Effects of Bcl-2 on Cell Death Triggered under ATP-Depleting Conditions

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