scholarly journals Purified F-ATP synthase forms a Ca2+-dependent high-conductance channel matching the mitochondrial permeability transition pore

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Andrea Urbani ◽  
Valentina Giorgio ◽  
Andrea Carrer ◽  
Cinzia Franchin ◽  
Giorgio Arrigoni ◽  
...  
Keyword(s):  
Lipid Bilayers ◽  
Atp Synthase ◽  
Mitochondrial Permeability Transition ◽  
Adenine Nucleotide ◽  
Adenine Nucleotides ◽  
Permeability Transition Pore ◽  
Anion Channel ◽  
Permeability Transition ◽  
Channel Activity ◽  
Voltage Dependent Anion Channel

Abstract The molecular identity of the mitochondrial megachannel (MMC)/permeability transition pore (PTP), a key effector of cell death, remains controversial. By combining highly purified, fully active bovine F-ATP synthase with preformed liposomes we show that Ca2+ dissipates the H+ gradient generated by ATP hydrolysis. After incorporation of the same preparation into planar lipid bilayers Ca2+ elicits currents matching those of the MMC/PTP. Currents were fully reversible, were stabilized by benzodiazepine 423, a ligand of the OSCP subunit of F-ATP synthase that activates the MMC/PTP, and were inhibited by Mg2+ and adenine nucleotides, which also inhibit the PTP. Channel activity was insensitive to inhibitors of the adenine nucleotide translocase (ANT) and of the voltage-dependent anion channel (VDAC). Native gel-purified oligomers and dimers, but not monomers, gave rise to channel activity. These findings resolve the long-standing mystery of the MMC/PTP and demonstrate that Ca2+ can transform the energy-conserving F-ATP synthase into an energy-dissipating device.

The mitochondrial permeability transition pore

1999 ◽  
Vol 66 ◽  
pp. 167-179 ◽  
Author(s):  
Martin Crompton ◽  
Sukaina Virji ◽  
Veronica Doyle ◽  
Nicholas Johnson ◽  
John M. Ward
Keyword(s):  
Mitochondrial Permeability Transition ◽  
Adenine Nucleotide ◽  
Permeability Transition Pore ◽  
Anion Channel ◽  
Permeability Transition ◽  
Adenine Nucleotide Translocase ◽  
Voltage Dependent Anion Channel ◽  
Inner Membrane ◽  
Membrane Pore ◽  
Voltage Dependent

This chapter reviews recent advances in the identification of the structural elements of the permeability transition pore. The discovery that cyclosporin A (CsA) inhibits the pore proved instrumental. Various approaches indicate that CsA blocks the pore by binding to cyclophilin (CyP)-D. In particular, covalent labelling of CyP-D in situ by a photoactive CsA derivative has shown that pore ligands have the same effects on the degree to which CsA both blocks the pore and binds to CyP-D. The recognition that CyP-D is a key component has enabled the other constituents to be resolved. Use of a CyP-D fusion protein as affinity matrix has revealed that CyP-D binds very strongly to 1:1 complexes of the voltage-dependent anion channel (from the outer membrane) and adenine nucleotide translocase (inner membrane). Our current model envisages that the pore arises as a complex between these three components at contact sites between the mitochondrial inner and outer membranes. This is in line with recent reconstitutions of pore activity from protein fractions containing these proteins. The strength of interaction between these proteins suggests that it may be a permanent feature rather than assembled only under pathological conditions. Calcium, the key activator of the pore, does not appear to affect pore assembly; rather, an allosteric action allowing pore flicker into an open state is indicated. CsA inhibits pore flicker and lowers the binding affinity for calcium. Whether adenine nucleotide translocase or the voltage-dependent anion channel (via inner membrane insertion) provides the inner membrane pore has not been settled, and data relevant to this issue are also documented.

Differential susceptibility of subsarcolemmal and intermyofibrillar mitochondria to apoptotic stimuli

2005 ◽  
Vol 289 (4) ◽  
pp. C994-C1001 ◽  
Author(s):  
Peter J. Adhihetty ◽  
Vladimir Ljubicic ◽  
Keir J. Menzies ◽  
David A. Hood
Keyword(s):  
Cytochrome C ◽  
Muscle Fiber ◽  
Mitochondrial Permeability Transition ◽  
Protein Composition ◽  
Permeability Transition Pore ◽  
Anion Channel ◽  
Permeability Transition ◽  
Cyclophilin D ◽  
Voltage Dependent Anion Channel ◽  
Release Channel

Apoptosis can be evoked by reactive oxygen species (ROS)-induced mitochondrial release of the proapoptotic factors cytochrome c and apoptosis-inducing factor (AIF). Because skeletal muscle is composed of two mitochondrial subfractions that reside in distinct subcellular regions, we investigated the apoptotic susceptibility of subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondria. SS and IMF mitochondria exhibited a dose-dependent release of protein in response to H2O2 (0, 25, 50, and 100 μM). However, IMF mitochondria were more sensitive to H2O2 and released a 2.5-fold and 10-fold greater amount of cytochrome c and AIF, respectively, compared with SS mitochondria. This finding coincided with a 44% ( P < 0.05) greater rate of opening (maximum rate of absorbance decrease, Vmax) of the protein release channel, the mitochondrial permeability transition pore (mtPTP), in IMF mitochondria. IMF mitochondria also exhibited a 47% ( P < 0.05) and 60% (0.05 < P < 0.1) greater expression of the key mtPTP component voltage-dependent anion channel and cyclophilin D, respectively, along with a threefold greater cytochrome c content, but similar levels of AIF compared with SS mitochondria. Despite a lower susceptibility to H2O2-induced release, SS mitochondria possessed a 10-fold greater Bax-to-Bcl-2 ratio ( P < 0.05), a 2.7-fold greater rate of ROS production, and an approximately twofold greater membrane potential compared with IMF mitochondria. The expression of the antioxidant enzyme Mn2+-superoxide dismutase was similar between subfractions. Thus the divergent protein composition and function of the mtPTP between SS and IMF mitochondria contributes to a differential release of cytochrome c and AIF in response to ROS. Given the relatively high proportion of IMF mitochondria within a muscle fiber, this subfraction is likely most important in inducing apoptosis when presented with apoptotic stimuli, ultimately leading to myonuclear decay and muscle fiber atrophy.

scholarly journals The HIV-1 Viral Protein R Induces Apoptosis via a Direct Effect on the Mitochondrial Permeability Transition Pore

2000 ◽  
Vol 191 (1) ◽  
pp. 33-46 ◽  
Author(s):  
Etienne Jacotot ◽  
Luigi Ravagnan ◽  
Markus Loeffler ◽  
Karine F. Ferri ◽  
Helena L.A. Vieira ◽  
...  
Keyword(s):  
Direct Effect ◽  
Viral Protein ◽  
Mitochondrial Permeability Transition ◽  
Adenine Nucleotide ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Voltage Dependent Anion Channel ◽  
Intact Cells ◽  
Viral Protein R ◽  
Hiv 1

Viral protein R (Vpr) encoded by HIV-1 is a facultative inducer of apoptosis. When added to intact cells or purified mitochondria, micromolar and submicromolar doses of synthetic Vpr cause a rapid dissipation of the mitochondrial transmembrane potential (ΔΨm), as well as the mitochondrial release of apoptogenic proteins such as cytochrome c or apoptosis inducing factor. The same structural motifs relevant for cell killing are responsible for the mitochondriotoxic effects of Vpr. Both mitochondrial and cytotoxic Vpr effects are prevented by Bcl-2, an inhibitor of the permeability transition pore complex (PTPC). Coincubation of purified organelles revealed that nuclear apoptosis is only induced by Vpr when mitochondria are present yet can be abolished by PTPC inhibitors. Vpr favors the permeabilization of artificial membranes containing the purified PTPC or defined PTPC components such as the adenine nucleotide translocator (ANT) combined with Bax. Again, this effect is prevented by addition of recombinant Bcl-2. The Vpr COOH terminus binds purified ANT, as well as a molecular complex containing ANT and the voltage-dependent anion channel (VDAC), another PTPC component. Yeast strains lacking ANT or VDAC are less susceptible to Vpr-induced killing than control cells yet recover Vpr sensitivity when retransfected with yeast ANT or human VDAC. Hence, Vpr induces apoptosis via a direct effect on the mitochondrial PTPC.

scholarly journals Formation of High-Conductive C Subunit Channels upon Interaction with Cyclophilin D

2021 ◽  
Vol 22 (20) ◽  
pp. 11022
Author(s):  
Giuseppe Federico Amodeo ◽  
Natalya Krilyuk ◽  
Evgeny V. Pavlov
Keyword(s):  
Lipid Bilayers ◽  
Atp Synthase ◽  
Mitochondrial Permeability Transition ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Cyclophilin D ◽  
Inner Mitochondrial Membrane ◽  
C Subunit ◽  
High Concentrations ◽  
Main Component

The c subunit of the ATP synthase is an inner mitochondrial membrane (IMM) protein. Besides its role as the main component of the rotor of the ATP synthase, c subunit from mammalian mitochondria exhibits ion channel activity. In particular, c subunit may be involved in one of the pathways leading to the formation of the permeability transition pore (PTP) during mitochondrial permeability transition (PT), a phenomenon consisting of the permeabilization of the IMM due to high levels of calcium. Our previous study on the synthetic c subunit showed that high concentrations of calcium induce misfolding into cross-β oligomers that form low-conductance channels in model lipid bilayers of about 400 pS. Here, we studied the effect of cyclophilin D (CypD), a mitochondrial chaperone and major regulator of PTP, on the electrophysiological activity of the c subunit to evaluate its role in the functional properties of c subunit. Our study shows that in presence of CypD, c subunit exhibits a larger conductance, up to 4 nS, that could be related to its potential role in mitochondrial toxicity. Further, our results suggest that CypD is necessary for the formation of c subunit induced PTP but may not be an integral part of the pore.

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