scholarly journals Cyclosporin A blocks 6-hydroxydopamine-induced efflux of Ca2+ from mitochondria without inactivating the mitochondrial inner-membrane pore

1994 ◽  
Vol 297 (1) ◽  
pp. 151-155 ◽  
Author(s):  
N Reichman ◽  
C M Porteous ◽  
M P Murphy
Keyword(s):  
Oxidative Stress ◽  
Free Radicals ◽  
Cyclosporin A ◽  
Mitochondrial Matrix ◽  
Inner Membrane ◽  
Membrane Pore ◽  
Mitochondrial Inner Membrane ◽  
Mitochondrial Pore ◽  
Pore Opening ◽  
6 Hydroxydopamine

Oxidative stress causes Ca(2+)-loaded mitochondria to release Ca2+. The mechanism of this efflux is unclear, but it appears to be associated with the opening of a pore in the mitochondrial inner membrane. Pore opening depolarizes the mitochondria, letting solutes enter the mitochondrial matrix, causing swelling. Cyclosporin A (CsA) prevents opening of this pore. The neurotoxin 6-hydroxydopamine (6HD) autoxidizes, producing free radicals, which cause oxidative stress. In this paper it is shown that 6HD-induced efflux from Ca(2+)-loaded mitochondria was prevented by CsA. The 6HD-induced Ca2+ efflux was not accompanied by mitochondrial swelling, depolarization of the mitochondrial inner membrane or movement of radiolabelled sucrose into the mitochondrial matrix. In agreement with others [Schlegel, Schweizer and Richter (1992) Biochem. J. 285, 65-69], these findings suggest that the mitochondrial pore remained closed during pro-oxidant-induced Ca2+ efflux. However, the implication that CsA blocks pro-oxidant-induced Ca2+ efflux by some mechanism other than inactivating the mitochondrial pore, suggests that the interaction of CsA with mitochondria may be more complex than is currently supposed.

scholarly journals Oxidative damage to mitochondria is mediated by the Ca2+-dependent inner-membrane permeability transition

1993 ◽  
Vol 294 (3) ◽  
pp. 719-725 ◽  
Author(s):  
N Takeyama ◽  
N Matsuo ◽  
T Tanaka
Keyword(s):  
Oxidative Stress ◽  
Free Radicals ◽  
Cyclosporin A ◽  
Nadh Oxidase ◽  
Experimental System ◽  
Permeability Transition ◽  
Ruthenium Red ◽  
Rat Liver Mitochondria ◽  
O2 Consumption ◽  
Inner Membrane

The ability of O2 metabolites derived from the xanthine-xanthine oxidase system to inhibit mitochondrial function was examined using freshly isolated rat liver mitochondria. Under 2,4-dinitrophenol-uncoupled conditions, mitochondria exposed to free radicals exhibited a significant decrease in O2 consumption supported by NAD(+)-linked substrates, but showed almost no change in O2 consumption in the presence of succinate and ascorbate. Oxidative stress caused the loss of intramitochondrial nicotinamide nucleotides, and addition of NAD+ fully prevented any fall in O2 consumption with NAD(+)-linked substrates. The activity of electron-transfer complex I (NADH oxidase and NADH-cytochrome c oxidoreductase) and the energy-dependent reduction of NAD+ by succinate were unaltered by oxidative stress. Exposure to free radicals also had an uncoupling effect at all three coupling sites. The degree of mitochondrial swelling was closely correlated with the inhibition of State-3 oxidation of site-I substrates and with the increase in State-4 oxidation of succinate. The immunosuppressive agent cyclosporin A completely prevented the mitochondrial damage induced by oxygen free radicals (swelling, Ca2+ release, sucrose trapping, uncoupling and selective inhibition of the mitochondrial respiration of site-I substrates). The same protective effect was found when Ca2+ cycling was prevented, either by chelating Ca2+ with EGTA or by inhibiting Ca2+ reuptake with Ruthenium Red. These findings suggest that the deleterious effect of free radicals on mitochondria in the present experimental system was triggered by the cyclosporin A-sensitive and Ca(2+)-dependent membrane transition, and not by direct impairment of the mitochondrial inner-membrane enzymes.

scholarly journals On the interactions of Ca2+ and cyclosporin A with a mitochondrial inner membrane pore: a study using cobaltammine complex inhibitors of the Ca2+ uniporter

1994 ◽  
Vol 302 (1) ◽  
pp. 181-185 ◽  
Author(s):  
M Crompton ◽  
L Andreeva
Keyword(s):  
Membrane Potential ◽  
Cyclosporin A ◽  
Reperfusion Injury ◽  
The State ◽  
Ischaemia Reperfusion Injury ◽  
Inner Membrane ◽  
Membrane Pore ◽  
Mitochondrial Inner Membrane ◽  
Ischaemia Reperfusion

The mitochondrial inner membrane contains a Ca(2+)-activated pore of possible relevance to the pathogenesis of ischaemia/reperfusion injury which is inhibited by the immunosuppressant cyclosporin A (CSA). The present study employs a number of novel cobaltammine complex inhibitors of the Ca2+ uniporter (mediating Ca2+ uptake) to examine whether intramitochondrial Ca2+ influences the capacity of CSA to block the pore. Using dissipation of the inner membrane potential as a means of monitoring the state of the pore, it is shown that CSA blockade is facilitated as Ca2+ uptake is restricted. Ca2+ also depresses and reverses the binding of [3H]CSA to mitochondria, but Ca2+ is ineffective when its uptake is prevented. It is concluded that a high intramitochondrial Ca2+ concentration antagonizes pore inhibition by CSA. The significance of this is discussed.

scholarly journals Recruitment of mitochondrial cyclophilin to the mitochondrial inner membrane under conditions of oxidative stress that enhance the opening of a calcium-sensitive non-specific channel

1994 ◽  
Vol 302 (2) ◽  
pp. 321-324 ◽  
Author(s):  
C P Connern ◽  
A P Halestrap
Keyword(s):  
Oxidative Stress ◽  
Adenine Nucleotide ◽  
Adenine Nucleotides ◽  
Mitochondrial Matrix ◽  
Mitochondrial Membranes ◽  
Isomerase Activity ◽  
Mitochondrial Inner Membrane ◽  
Cyclosporin Treatment ◽  
Sds Page ◽  
Mitochondrial Pore

Binding of mitochondrial matrix cyclophilin (CyP) to the rat liver mitochondrial membranes was detected by SDS/PAGE and Western blotting with suitable antipeptide antibodies. Binding was not affected by prior exposure of mitochondria to Ca2+, adenine nucleotides or inhibitors of the adenine nucleotide translocase, but was greatly increased by t-butyl hydroperoxide (tBH), phenylarsine oxide or diamide. These all sensitized the opening of the non-specific mitochondrial pore to [Ca2+], and the effect of tBH was shown to be maintained after washing away the tBH, consistent with it being caused by the enhanced CyP binding. The bound CyP did not demonstrate peptidyl-prolyl cis-trans isomerase activity. CyP-binding was prevented by 5 microM cyclosporin A, but not reversed by cyclosporin treatment of the membranes. The effect of tBH on binding was concentration-dependent and maximal within 30 s.

scholarly journals Two forms of Opa1 cooperate to complete fusion of the mitochondrial inner-membrane

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Yifan Ge ◽  
Xiaojun Shi ◽  
Sivakumar Boopathy ◽  
Julie McDonald ◽  
Adam W Smith ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Short Form ◽  
Membrane Dynamics ◽  
Complete Fusion ◽  
Inner Membrane ◽  
Membrane Pore ◽  
Mitochondrial Inner Membrane ◽  
Pore Opening ◽  
Organelle Morphology

Mitochondrial membrane dynamics is a cellular rheostat that relates metabolic function and organelle morphology. Using an in vitro reconstitution system, we describe a mechanism for how mitochondrial inner-membrane fusion is regulated by the ratio of two forms of Opa1. We found that the long-form of Opa1 (l-Opa1) is sufficient for membrane docking, hemifusion and low levels of content release. However, stoichiometric levels of the processed, short form of Opa1 (s-Opa1) work together with l-Opa1 to mediate efficient and fast membrane pore opening. Additionally, we found that excess levels of s-Opa1 inhibit fusion activity, as seen under conditions of altered proteostasis. These observations describe a mechanism for gating membrane fusion.

scholarly journals Two forms of Opa1 cooperate to complete fusion of the mitochondrial inner-membrane

2019 ◽  
Author(s):  
Yifan Ge ◽  
Xiaojun Shi ◽  
Sivakumar Boopathy ◽  
Julie McDonald ◽  
Adam W. Smith ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Short Form ◽  
Membrane Dynamics ◽  
Complete Fusion ◽  
Inner Membrane ◽  
Membrane Pore ◽  
Mitochondrial Inner Membrane ◽  
Pore Opening ◽  
Organelle Morphology

AbstractMitochondrial membrane dynamics is a cellular rheostat that relates metabolic function and organelle morphology. Using an in vitro reconstitution system, we describe a mechanism for how mitochondrial inner-membrane fusion is regulated by the ratio of two forms of Opa1. We found that the long-form of Opa1 (l-Opa1) is sufficient for membrane docking, hemifusion and low levels of content release. However, stoichiometric levels of the processed, short form of Opa1 (s-Opa1) work together with l-Opa1 to mediate efficient and fast membrane pore opening. Additionally, we found that excess levels of s-Opa1 inhibit fusion activity, as seen under conditions of altered proteostasis. These observations describe a mechanism for gating membrane fusion.

Removal of a hydrophobic domain within the mature portion of a mitochondrial inner membrane protein causes its mislocalization to the matrix

1990 ◽  
Vol 10 (5) ◽  
pp. 1873-1881
Author(s):  
S M Glaser ◽  
B R Miller ◽  
M G Cumsky
Keyword(s):  
Amino Acids ◽  
Mitochondrial Matrix ◽  
Mature Protein ◽  
Wild Type ◽  
Inner Membrane ◽  
Hydrophobic Domain ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Inner Membrane Protein ◽  
Cognate Protein

We have examined the import and intramitochondrial localization of the precursor to yeast cytochrome c oxidase subunit Va, a protein of the mitochondrial inner membrane. The results of studies on the import of subunit Va derivatives carrying altered presequences suggest that the uptake of this protein is highly efficient. We found that a presequence of only 5 amino acids (Met-Leu-Ser-Leu-Arg) could direct the import and localization of subunit Va with wild-type efficiency, as judged by several different assays. We also found that subunit Va could be effectively targeted to the mitochondrial inner membrane with a heterologous presequence that failed to direct import of its cognate protein. The results presented here confirmed those of an earlier study and showed clearly that the information required to "sort" subunit Va to the inner membrane resides in the mature protein sequence, not within the presequence per se. We present additional evidence that the aforementioned sorting information is contained, at least in part, in a hydrophobic stretch of 22 amino acids residing within the C-terminal third of the protein. Removal of this domain caused subunit Va to be mislocalized to the mitochondrial matrix.

scholarly journals The translocator maintenance protein Tam41 is required for mitochondrial cardiolipin biosynthesis

2008 ◽  
Vol 183 (7) ◽  
pp. 1213-1221 ◽  
Author(s):  
Stephan Kutik ◽  
Michael Rissler ◽  
Xue Li Guan ◽  
Bernard Guiard ◽  
Guanghou Shui ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Respiratory Chain ◽  
Matrix Protein ◽  
Mitochondrial Protein ◽  
Pleiotropic Effects ◽  
Mitochondrial Matrix ◽  
Carrier Proteins ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane

The mitochondrial inner membrane contains different translocator systems for the import of presequence-carrying proteins and carrier proteins. The translocator assembly and maintenance protein 41 (Tam41/mitochondrial matrix protein 37) was identified as a new member of the mitochondrial protein translocator systems by its role in maintaining the integrity and activity of the presequence translocase of the inner membrane (TIM23 complex). Here we demonstrate that the assembly of proteins imported by the carrier translocase, TIM22 complex, is even more strongly affected by the lack of Tam41. Moreover, respiratory chain supercomplexes and the inner membrane potential are impaired by lack of Tam41. The phenotype of Tam41-deficient mitochondria thus resembles that of mitochondria lacking cardiolipin. Indeed, we found that Tam41 is required for the biosynthesis of the dimeric phospholipid cardiolipin. The pleiotropic effects of the translocator maintenance protein on preprotein import and respiratory chain can be attributed to its role in biosynthesis of mitochondrial cardiolipin.

scholarly journals Removal of a hydrophobic domain within the mature portion of a mitochondrial inner membrane protein causes its mislocalization to the matrix.

1990 ◽  
Vol 10 (5) ◽  
pp. 1873-1881 ◽  
Author(s):  
S M Glaser ◽  
B R Miller ◽  
M G Cumsky
Keyword(s):  
Amino Acids ◽  
Mitochondrial Matrix ◽  
Mature Protein ◽  
Wild Type ◽  
Inner Membrane ◽  
Hydrophobic Domain ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Inner Membrane Protein ◽  
Cognate Protein

We have examined the import and intramitochondrial localization of the precursor to yeast cytochrome c oxidase subunit Va, a protein of the mitochondrial inner membrane. The results of studies on the import of subunit Va derivatives carrying altered presequences suggest that the uptake of this protein is highly efficient. We found that a presequence of only 5 amino acids (Met-Leu-Ser-Leu-Arg) could direct the import and localization of subunit Va with wild-type efficiency, as judged by several different assays. We also found that subunit Va could be effectively targeted to the mitochondrial inner membrane with a heterologous presequence that failed to direct import of its cognate protein. The results presented here confirmed those of an earlier study and showed clearly that the information required to "sort" subunit Va to the inner membrane resides in the mature protein sequence, not within the presequence per se. We present additional evidence that the aforementioned sorting information is contained, at least in part, in a hydrophobic stretch of 22 amino acids residing within the C-terminal third of the protein. Removal of this domain caused subunit Va to be mislocalized to the mitochondrial matrix.

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