Mitochondrial Membrane Potential Supported by Exogenous CytochromecOxidation Mimics the Early Stages of Apoptosis

Piana, Gianluigi La; Fransvea, Emilia; Marzulli, Domenico; Lofrumento, N.E.

doi:10.1006/bbrc.1998.8664

Cited by 43 publications

(30 citation statements)

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“…The relationship between ⌬ m and cytochrome c in mitochondrial dysfunction is complex and variable. For example, in cerebellar neurons, cytochrome c is released before any detectable changes in ⌬ m (29,41,47,55). Our data contrast with the finding in a previous report that CSA did not protect mitochondria against H. pylori VacA toxin, which causes mitochondrial damage without inducing a permeability transition (56).…”

Section: Discussioncontrasting

confidence: 99%

Dynamin-2-Dependent Targeting ofMannheimia haemolyticaLeukotoxin to Mitochondrial Cyclophilin D in Bovine Lymphoblastoid Cells

et al. 2008

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Exotoxins which belong to the family containing the RTX toxins (repeats in toxin) contribute to a variety of important human and animal diseases. One example of such a toxin is the potent leukotoxin (LKT) produced by the bovine respiratory pathogen Mannheimia haemolytica. LKT binds to CD18, resulting in the death of bovine leukocytes. In this study, we showed that internalized LKT binds to the outer mitochondrial membrane, which results in the release of cytochrome c and collapse of the mitochondrial membrane potential ( M. haemolytica is the principal bacterial pathogen in the bovine respiratory disease complex (19, 58). The most important virulence factor of M. haemolytica is the LKT, which has potent cytotoxic effects on ruminant leukocytes but not on leukocytes from other species (38). M. haemolytica LKT is a member of the RTX toxin (repeats in toxin) family, which includes LKTs and hemolysins produced by a number of gramnegative bacteria. It was originally thought that M. haemolytica LKT damages cells by inserting into the cell membrane, resulting in pore formation and necrosis (53). At lower LKT concentrations, however, susceptible cells die via caspase-dependent apoptotic pathways (8,10,45,46). LKT was first reported to bind to the ␤ 2 integrin CD18/CD11a, also known as leukocyte functional antigen 1 (LFA-1) (1,24,28). Recent studies, however, have shown that Mac-1 (CD18/CD11b) also binds LKT and that CD18 is the functional receptor for LKT (9, 30). After binding to LFA-1, LKT induces apoptosis of bovine leukocytes as a result of activation of caspase 1, caspase 3, and caspase 9 (4, 10, 31, 46). The LFA-1-dependent cytotoxicity of LKT is a conundrum, because signaling through LFA-1 generally results in cell adhesion and promotes cell survival (34,44,51).In this study we hypothesized that mitochondrial damage and caspase 9 activation could be due to direct binding of LKT to the mitochondrial outer membrane (MOM). In this paper we show that LKT directly targets mitochondria in BL-3 cells, creating lesions in the MOM that can lead to release of proapoptotic proteins, culminating in cell death. We also show that LKT targeting to mitochondria is dynamin-2 dependent and that mitochondria appear to be the principal source of dynamin-2 in BL-3 cells. Treating BL-3 cells with cyclosporine (CSA) depleted mitochondrial dynamin-2, thereby inhibiting LKT transport to mitochondria and cell death. MATERIALS AND METHODSLKT production and purification. Crude LKT was prepared and purified as described previously and was stored at Ϫ80°C until it was used in experiments (4). Inactive toxin from an lktC mutant of M. haemolytica strain A1 (SH 1562), which produces an LKT protein with no biological activity (LKT inact ) (generously provided by S. K. Highlander, Baylor College of Medicine, Houston, TX), was prepared in a similar manner.Cell lines, cell cultures, and antibodies. Nonadherent bovine lymphoblastoid cells (BL-3 cells) and adherent murine macrophages (RAW 264.7 cells) (kindly provided by Ronald Schultz, University of...

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Section: Discussioncontrasting

confidence: 99%

Dynamin-2-Dependent Targeting ofMannheimia haemolyticaLeukotoxin to Mitochondrial Cyclophilin D in Bovine Lymphoblastoid Cells

et al. 2008

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“…It was assumed that under some conditions, when the concentration of cytoplasmic cytochrome c is increased, the external NADH oxidation by intact mitochondria might be significantly activated due to a bi-trans-membrane electron transport catalyzed by external cytochrome c (25), and it could play an essential role in mechanisms of apoptosis. However, it cannot be excluded that the stimulation of the external NADH oxidation by exogenous cytochrome c, observed by the authors (24,25), relates to only damaged mitochondria that represent nearly 10% of the total population of isolated mitochondria (11,13,14). Whatever the mechanism of rotenone-insensitive oxidation of cytoplasmic NADH, activation of this process is likely to be important in redox signaling in apoptosis.…”

Section: Resultsmentioning

confidence: 97%

“…According to some data, the rate of external NADH oxidation by rat liver mitochondria may be stimulated by extramitochondrial cytochrome c (22). The concept of "bi-trans-membrane" electron transport was developed to explain this phenomenon (22)(23)(24), assuming its possible relation to the mechanisms of apoptosis (25). These authors suggested that extramitochondrial cytochrome c, reduced on the OMM by external NADH, might be directly oxidized by cytochrome c oxidase of mitochondria, transferring electrons through the intact OMM.…”

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confidence: 99%

Cytochrome c Sorption-Desorption Effects on the External NADH Oxidation by Mitochondria

Lemeshko

2002

Journal of Biological Chemistry

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The rupture of the outer mitochondrial membrane is known to be critical for cell death, but the mechanism, specifically its redox-signaling aspects, still needs to be studied in more detail. In this work, the external NADH oxidation by rat liver mitochondria was studied under the outer membrane rupture induced by the mitochondria hypotonic treatment or the inner membrane permeability transition. The saturation of the oxidation rate was observed as a function of mitochondrial protein concentration. This effect was shown to result from cytochrome c binding to the mitochondrial membranes. At a relatively high concentration of mitochondria, the oxidation rate was strongly activated by 4 mM Mg 2؉ due to cytochrome c desorption from the membranes. A minimal kinetic model was developed to explain the main phenomena of the external NADH oxidation modulated by cytochrome c and Mg 2؉ in mitochondria with the ruptured outer membrane. The computational behavior of the model closely agreed with the experimental data. We suggest that the redox state of the released cytochrome c, considered by other authors to be important for apoptosis, may strongly depend on its oxidation by the fraction of mitochondria with the ruptured outer membrane and on the cytoplasmic cytochrome c reductase activity.

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“…We are interested in characterizing and delineating, in dissimilar metabolic states, the functional activity of the cytosolic NADH/cyto-c electron transport pathway, a system involved in the direct oxidation of NADH present outside the mitochondria [11][12][13][14]. This is a reducing equivalent transport system independent and distinct from both the shuttle system and the respiratory chain.…”

Section: Introductionmentioning

confidence: 99%

Valinomycin induced energy-dependent mitochondrial swelling, cytochrome c release, cytosolic NADH/cytochrome c oxidation and apoptosis

et al. 2011

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In valinomycin induced stimulation of mitochondrial energy dependent reversible swelling, supported by succinate oxidation, cytochrome c (cyto-c) and sulfite oxidase (Sox) [both present in the mitochondrial intermembrane space (MIS)] are released outside. This effect can be observed at a valinomycin concentration as low as 1 nM. The rate of cytosolic NADH/cyto-c electron transport pathway is also greatly stimulated. The test on the permeability of mitochondrial outer membrane to exogenous cyto-c rules out the possibility that the increased rate of exogenous NADH oxidation could be ascribed either to extensively damaged or broken mitochondria. Accumulation of potassium inside the mitochondria, mediated by the highly specific ionophore valinomycin, promotes an increase in the volume of matrix (evidenced by swelling) and the interaction points between the two mitochondrial membranes are expected to increase. The data reported and those previously published are consistent with the view that "respiratory contact sites" are involved in the transfer of reducing equivalents from cytosol to inside the mitochondria both in the absence and the presence of valinomycin. Magnesium ions prevent at least in part the valinomycin effects. Rather than to the dissipation of membrane potential, the pro-apoptotic property of valinomycin can be ascribed to both the release of cyto-c from mitochondria to cytosol and the increased rate of cytosolic NADH coupled with an increased availability of energy in the form of glycolytic ATP, useful for the correct execution of apoptotic program.

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Mitochondrial Membrane Potential Supported by Exogenous CytochromecOxidation Mimics the Early Stages of Apoptosis

Cited by 43 publications

References 34 publications

Dynamin-2-Dependent Targeting ofMannheimia haemolyticaLeukotoxin to Mitochondrial Cyclophilin D in Bovine Lymphoblastoid Cells

Dynamin-2-Dependent Targeting ofMannheimia haemolyticaLeukotoxin to Mitochondrial Cyclophilin D in Bovine Lymphoblastoid Cells

Cytochrome c Sorption-Desorption Effects on the External NADH Oxidation by Mitochondria

Valinomycin induced energy-dependent mitochondrial swelling, cytochrome c release, cytosolic NADH/cytochrome c oxidation and apoptosis

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