Chaperoning mitochondrial permeability transition: regulation of transition pore complex by a J-protein, DnaJC15

Mitochondria have a central role in the intrinsic pathway of apoptosis and involve activation of several transmembrane channels leading to release of death factors. Reduced expression of a mitochondrial J-protein DnaJC15 was associated with the development of chemoresistance in ovarian cancer cells. DnaJC15 was found to be a part of mitochondrial protein-transport machinery, though its connection with cell death mechanisms is still unclear. In the present study, we have provided evidence towards a novel function of DnaJC15 in regulation of mitochondrial permeability transition pore (MPTP) complex in normal and cancer cells. Overexpression of DnaJC15 resulted in MPTP opening and induction of apoptosis, whereas reduced amount of protein suppressed MPTP activation, upon cisplatin treatment. DnaJC15 was found to exert its proapoptotic function through the essential component of MPTP, cyclophilin D (CypD). Our results reveal a specific role of DnaJC15 in recruitment and coupling of CypD with mitochondrial permeability transition. In summary, our analysis provides first-time insights on the functional connection between mitochondrial inner membrane protein translocation machinery-associated J-protein DnaJC15 and regulation of cell death pathways.

Microcystin-RR induced apoptosis in tobacco BY-2 suspension cells is mediated by reactive oxygen species and mitochondrial permeability transition pore status

When tobacco BY-2 cells were treated with 60 mu g/mL MC-RR for 5 d, time-dependent effects of MC-RR on the cells were observed. Morphological changes such as abnormal elongation, evident chromatin condensation and margination, fragmentation of nucleus and formation of apoptotic-like bodies suggest that 60 mu g/mL MC-RR induced rapid apoptosis in tobacco BY-2 cells. Moreover, there was a significant and rapid increase of ROS level before the loss of mitochondrial membrane potential (Delta Psi(m)) and the onset of cell apoptosis. Ascorbic acid (AsA), a major primary antioxidant, prevented the increase of ROS generation, blocked the decrease in Delta Psi(m) and subsequent cell apoptosis, indicating a critical role of ROS in serving as an important signaling molecule by causing a reduction of Delta Psi(m) and MC-RR-induced tobacco BY-2 cell apoptosis. In addition, a specific mitochondrial permeability transition pores (PTP) inhibitor, cyclosporin A (CsA), significantly blocked the MC-RR-induced ROS formation, loss of Delta Psi(m), as well as cell apoptosis when the cells were MC-RR stressed for 3 d, suggesting that PTP is involved in 60 mu g/mL MC-RR-induced tobacco cell apoptosis signalling process. Thus, we concluded that the mechanism of MC-RR-induced apoptosis signalling pathways in tobacco BY-2 cells involves not only the excess generation of ROS and oxidative stress, but also the opening of PTP inducing loss of mitochondrial membrane potential. (C) 2007 Published by Elsevier Ltd.

Cytochrome bc1 regulates the mitochondrial permeability transition by two distinct pathways

The mitochondrial permeability transition (MPT) pore is a calcium-sensitive channel in the mitochondrial inner membrane that plays a crucial role in cell death. Here we show that cytochrome bc₁ regulates the MPT in isolated rat liver mitochondria and in CEM and HL60 cells by two independent pathways. Glutathione depletion activated the MPT via increased production of reactive oxygen species (ROS) generated by cytochrome bc₁. The ROS producing mechanism in cytochrome bc₁ involves movement of the "Rieske" iron-sulfur protein subunit of the enzyme complex, because inhibition of cytochrome bc₁ by pharmacologically blocking iron-sulfur protein movement completely abolished ROS production, MPT activation, and cell death. The classical inhibitor of the MPT, cyclosporine A, had no protective effect against MPT activation. In contrast, the calcium-activated, cyclosporine A-regulated MPT in rat liver mitochondria was also blocked with inhibitors of cytochrome bc₁. These results indicate that electron flux through cytochrome bc1 regulates two distinct pathways to the MPT, one unregulated and involving mitochondrial ROS and the other regulated and activated by calcium.

Effects of isocoumarins isolated from Paepalanthus bromelioides on mitochondria: Uncoupling, and induction/inhibition of mitochondrial permeability transition

Isolated mitochondria may undergo uncoupling, and in presence of Ca2+ at different conditions, a mitochondrial permeability transition (MPT) linked to protein,thiol oxidation, and demonstrated by CsA-sensitive mitochondrial swelling; these processes may cause cell death either by necrosis or by apoptosis. Isocoumarins isolated from the Brazilian plant Paepalanthus bromelioides (Eriocaulaceae) paepalantine (9,10-dihydroxy-5,7-dimethoxy-1H-naptho(2,3c)pyran-1-one), 8,8'-paepalantine dimer, and vioxanthin were assayed at 1-50 mu M on isolated rat liver mitochondria, for respiration, MPT, protein thiol oxidation, and interaction with the mitochondrial membrane using 1,6-diphenyl-1,3,5-hexatriene (DPH). The isocoumarins did not significantly affect state 3 respiration of succinate-energized mitochondria; they did however, stimulate 4 respiration, indicating mitochondrial uncoupling. Induction of MPT and protein thiol oxidation were assessed in succinate-energized mitochondria exposed to 10 mu M Ca2+; inhibition of these processes was assessed in non-energized organelles in the presence of 300 mu M t-butyl hydroperoxide plus 500 mu M Ca2+. Only paepalantine was an effective MPT/protein thiol oxidation inducer, also releasing cytochrome c from mitochondria; the protein thiol oxidation, unlike mitochondrial swelling, was neither inhibited by CsA nor dependent on the presence of Ca2+. Vioxanthin was an effective inhibitor of MPT/protein thiol oxidation. All isocoumarins inserted deeply into the mitochondrial membrane, but only paepalantine dimer and vioxantin decreased the membrane's fluidity. A direct reaction with mitochondrial membrane protein thiols, involving an oxidation of these groups, is proposed to account for MPT induction by paepalantine, while a restriction of oxidation of these same thiol groups imposed by the decrease of membrane fluidity, is proposed to account for MPT inhibition by vioxanthin. (c) 2006 Published by Elsevier B.V..

Dehydromonocrotaline induces cyclosporine A-insensitive mitochondrial permeability transition/cytochrome c release

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The cardioprotective effect of a short-term aerobic exercise program and the mitochondrial permeability transition pore of the Rat

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Skeletal muscle disuse atrophy : implications on intracellular signaling pathways and mitochondrial permeability transition pore function

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

The cardioprotective effect of a short-term aerobic exercise program and the mitochondrial permeability transition pore of the Rat

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Skeletal muscle disuse atrophy : implications on intracellular signaling pathways and mitochondrial permeability transition pore function

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

Nicorandil protects cardiac mitochondria against permeability transition induced by ischemia-reperfusion

Ischemia followed by reperfusion is known to negatively affect mitochondrial function by inducing a deleterious condition termed mitochondrial permeability transition. Mitochondrial permeability transition is triggered by oxidative stress, which occurs in mitochondria during ischemia-reperfusion as a result of lower antioxidant defenses and increased oxidant production. Permeability transition causes mitochondrial dysfunction and can ultimately lead to cell death. A drug able to minimize mitochondrial damage induced by ischemia-reperfusion may prove to be clinically effective. We aimed to analyze the effects of nicorandil, an ATP-sensitive potassium channel agonist and vasodilator, on mitochondrial function of rat hearts and cardiac HL-1 cells submitted to ischemia-reperfusion. Nicorandil decreased mitochondrial swelling and calcium uptake. It also decreased reactive oxygen species formation and thiobarbituric acid reactive substances levels, a lipid peroxidation biomarker. We thus confirm previous reports that nicorandil inhibits mitochondrial permeability transition and demonstrate that nicorandil inhibits this process by preventing oxidative damage and mitochondrial calcium overload induced by ischemia-reperfusion, resulting in improved cardiomyocyte viability. These results may explain the good clinical results obtained when using nicorandil in the treatment of ischemic heart disease.

Bax interacts with the permeability transition pore to induce permeability transition and cytochrome c release in isolated mitochondria

Cytochrome c release and the mitochondrial permeability transition (PT), including loss of the transmembrane potential (Δψ), play an important role in apoptosis. Using isolated mitochondria, we found that recombinant Bax and Bak, proapoptotic members of the Bcl-2 family, induced mitochondrial Δψ loss, swelling, and cytochrome c release. All of these changes were dependent on Ca2+ and were prevented by cyclosporin A (CsA) and bongkrekic acid, both of which close the PT pores (megachannels), indicating that Bax- and Bak-induced mitochondrial changes were mediated through the opening of these pores. Bax-induced mitochondrial changes were inhibited by recombinant Bcl-xL and transgene-derived Bcl-2, antiapoptotic members of the Bcl-2 family, as well as by oligomycin, suggesting a possible regulatory effect of F0F1-ATPase on Bax-induced mitochondrial changes. Proapoptotic Bax- and Bak-BH3 (Bcl-2 homology) peptides, but not a mutant BH3 peptide nor a mutant Bak lacking BH3, induced the mitochondrial changes, indicating an essential role of the BH3 region. A coimmunoprecipitation study revealed that Bax and Bak interacted with the voltage-dependent anion channel, which is a component of PT pores. Taken together, these findings suggest that proapoptotic Bcl-2 family proteins, including Bax and Bak, induce the mitochondrial PT and cytochrome c release by interacting with the PT pores.

A mitochondrial perspective on striated muscle physiopathology: insights from sepsis, denervation, and dystrophinopathies.

La mitochondrie est de plus en plus reconnue pour sa contribution à la dégénerescence musculaire. Les dysfonctions mitochondriales, en plus de causer une défaillance énergétique, contribuent à la signalisation apoptotique, stimule la production de ROS et peuvent induire une surcharge calcique. Ces caractéristiques sont tous reliées à certains types de myopathies. Cette thèse met en lumières comment certaines dysfonctions mitochondriales peuvent intervenir dans la pathogenèse de diverses myopathies. Nous démontrons que les dysfonctions mitochondriales sont impliqués dans l’atrophie dû à la perte d’innervation. Par contre, la désensabilisation de l’ouverture du pore mitochondrial de transition de perméabilité, via ablation génétique de cyclophiline-D, ne prévient ni la signalisation apoptotique mitochondrial ni l’atrophie. Nous avons aussi observé des dysfonctions mitochondriales dans le muscle atteint de dystrophie musculaire de Duchenne qui furent améliorés suite à une transfection de PGC1-α, laquelle résulta aussi en une amélioration de la pathologie. Finalement, nous démontrons que le recyclage de mitochondrie par les voies de mitophagies et de contrôles de la qualité impliquant Parkin et possiblement d’autres voies de signalisation inconnues sont cruciales au recouvrement cardiaqe lors d’un choc septique.

Mitochondrial Swelling and Incipient Outer Membrane Rupture in Preapoptotic and Apoptotic Cells

Outer mitochondrial membrane (OMM) rupture was first noted in isolated mitochondria in which the inner mitochondrial membrane (IMM) had lost its selective permeability. This phenomenon referred to as mitochondrial permeability transition (MPT) refers to a permeabilized inner membrane that originates a large swelling in the mitochondrial matrix, which distends the outer membrane until it ruptures. Here, we have expanded previous electron microscopic observations that in apoptotic cells, OMM rupture is not caused by a membrane stretching promoted by a markedly swollen matrix. It is shown that the widths of the ruptured regions of the OMM vary from 6 to 250 nm. Independent of the perforation size, herniation of the mitochondrial matrix appeared to have resulted in pushing the IMM through the perforation. A large, long focal herniation of the mitochondrial matrix, covered with the IMM, was associated with a rupture of the OMM that was as small as 6 nm. Contextually, the collapse of the selective permeability of the IMM may precede or follow the release of the mitochondrial proteins of the intermembrane space into the cytoplasm. When the MPT is a late event, exit of the intermembrane space proteins to the cytoplasm is unimpeded and occurs through channels that transverse the outer membrane, because so far, the inner membrane is impermeable. No channel within the outer membrane can expose to the cytoplasm a permeable inner membrane, because it would serve as a conduit for local herniation of the mitochondrial matrix. Anat Rec, 2012. (c) 2012 Wiley Periodicals, Inc.

Release of NO from a nitrosyl ruthenium complex through oxidation of mitochondrial NADH and effects on mitochondria

Nitrosyl ruthenium complexes are promising NO donor agents with numerous advantages for the biologic applications of NO. We have characterized the NO release from the nitrosyl ruthenium complex [Ru(NO2)(bpy)(2)(4-pic)](+) (I) and the reactive oxygen/nitrogen species (ROS/RNS)-mediated NO actions on isolated rat liver mitochondria. The results indicated that oxidation of mitochondrial NADH promotes NO release from (I) in a manner mediated by NO2 formation (at neutral pH) as in mammalian cells, followed by an oxygen atom transfer mechanism (OAT). The NO released from (I) uncoupled mitochondria at low concentrations/incubation times and inhibited the respiratory chain at high concentrations/incubation times. In the presence of ROS generated by mitochondria NO gave rise to peroxynitrite, which, in turn, inhibited the respiratory chain and oxidized membrane protein-thiols to elicit a Ca2+-independent mitochondrial permeability transition; this process was only partially inhibited by cyclosporine-A, almost fully inhibited by the thiol reagent N-ethylmaleimide (NEM) and fully inhibited by the NO scavenger 2-(4-carboxyphenyl)-4,45,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO). These actions correlated with the release of cytochrome c from isolated mitochondria as detected by Western blotting analysis. These events, typically involved in cell necrosis and/or apoptosis denote a potential specific action of (I) and analogs against tumor cells via mitochondria-mediated processes. (C) 2012 Elsevier Inc. All rights reserved.