Inhibitors of the Quinone-binding Site Allow Rapid Superoxide Production from Mitochondrial NADH:Ubiquinone Oxidoreductase (Complex I)

Lambert, Adrian J.; Brand, Martin D.

doi:10.1074/jbc.m406576200

Cited by 430 publications

(373 citation statements)

References 33 publications

(38 reference statements)

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“…Subsequent inhibition of the complex I of the respiratory chain by rotenone, excessive JC-1 aggregation disappeared, indicating an interference of NAC with the reentry of protons into the mitochondrial matrix inner space while the respiratory complex remains intact. In luteal cells from early phase corpus luteum, excessive JC-1 aggregation after exposure to NAC and the protonophore was not observed and rotenone decreased Dw as expected from results of other cell types (21,22,59,61). Therefore, considering the NAC-provoked reduction of the viability of luteal cells from midphase corpus luteum, our data suggest that proton leakage seems to be beneficial for the mitochondrial function.…”

Section: Discussionsupporting

confidence: 77%

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N‐acetylcysteine impairs survival of luteal cells through mitochondrial dysfunction

Löhrke

Weitzel

et al. 2010

Cytometry Pt A

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N-acetylcysteine (NAC) is known as an antioxidant and used for mucus viscosity reduction. However, this drug prevents or induces cell death depending on the cell type. The response of steroidogenic luteal cells to NAC is unknown. Our data shows that NAC can behave as an antioxidant or prooxidant in dependency on the concentration and mitochondrial energization. NAC elevated the flowcytometric-measured portion of hypodiploid (dying) cells. This rise was completely abolished by aurintricarboxylic acid, an inhibitor of topoisomerase II. NAC increased the secretion of nitric oxide and cellular nitrotyrosine. An image analysis indicated that cells pretreated with NAC and loaded with DHR showed a fluorescent structure probably elicited by the oxidative product of DHR, rhodamine 123 that sequesters mitochondrially. Pretreating luteal cells with NAC or adding NAC directly to mitochondrial fractions followed by assessing the mitochondrial transmembrane potential difference (Dw) by the JC-1 technique demonstrated a marked decrease in Dw. A protonophore restored Dw and rotenone (an inhibitor of respiratory chain complex I) inhibited mitochondrial recovering. Thus, in steroidogenic luteal cells from healthy mature corpus luteum, NAC impairs cellular survival by interfering with mitochondrial metabolism. The protonophoreinduced recovering of NAC-provoked decrease in Dw indicates that an ATP synthasefavored route of H 1 re-entry to the matrix is essentially switched off by NAC while other respiratory chain complexes remain intact. These data may be important for therapeutic timing of treatments with NAC. ' 2010 International Society for Advancement of Cytometry Key terms luteal cells; antioxidant; redox imbalance; dihydrorhodamine 123; flow cytometry; mitochondrial dysfunction; viability THE corpus luteum is a transient ovarian steroidogenic gland, which develops within few days from the ovulated follicle to establish embryonic development. Intense vascularization is essential for maturation and maintenance of a healthy gland (1,2). The mature mid-luteal phase corpus luteum secretes large quantities of progesterone, a derivative from its mitochondrially formed precursor, pregnenolone.Steroidogenesis-dependent oxy-radicals and oxidants are generated during intramitochondrial conversion of cholesterol through the activity of NADPH-dependent cholesterol monooxygenase (side chain cleaving). It catalyzes in a phospholipid-assisted manner the formation of pregnenolone. This process is accompanied by an electron leakage into the mitochondrial matrix (3) and the synthesis of 4-methylpental as byproduct (4,5). In addition, regulatory constituents driving enhanced steroidogenesis are generated by the cyclooxygenase and lipoxygenase pathways which require reactive oxygen species. For instance, cyclooxygenase has an absolute requirement for hydroperoxide to catalyze oxygenation of lipids with a lipoperoxidation as the first step in forming prostanoids (6). In turn, high concentration of antioxidants and oxidant-detoxifying enzym...

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Section: Discussionsupporting

confidence: 77%

“…This approach allowed us to examine the response to both NAC and rotenone, a plasma membrane-impermeable inhibitor of the complex I of the respiratory chain. These tests were performed because the complex I has been shown to be one of the main cellular sources of ROS and to largely determine the mitochondrial activity (21,22).…”

Section: Discussionmentioning

confidence: 99%

N‐acetylcysteine impairs survival of luteal cells through mitochondrial dysfunction

Löhrke

Weitzel

et al. 2010

Cytometry Pt A

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“…These results strongly suggest that effects of ∆lac-acetogenins on the redox status of the free radical intermediate in complex I, probably the region of the ubisemiquinone binding site, are fairly different from those of ordinary complex I inhibitors. The rates of superoxide production induced by ordinary inhibitors were not identical, whereas the difference in the rates was less significant than that determined as hydrogen peroxide generation with intact mitochondria (22).…”

Section: Crucial Structural Factors In the Thf Ring Moietymentioning

confidence: 64%

Synthesis and Inhibition Mechanism of Δlac-Acetogenins, a Novel Type of Inhibitor of Bovine Heart Mitochondrial Complex I

et al. 2004

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We have synthesized ∆lac-acetogenins that are new acetogenin mimics possessing two n-alkyl tails without an R, -unsaturated γ-lactone ring and suggested that their inhibition mechanism may be different from that of common acetogenins [Hamada et al. (2004) Biochemistry 43, 3651-3658]. To elucidate the inhibition mechanism of ∆lac-acetogenins in more detail, we carried out wide structural modifications of original ∆lac-acetogenins and characterized the inhibitory action with bovine heart mitochondrial complex I. In contrast to common acetogenins, both the presence of adjacent bis-THF rings and the stereochemistry around the hydroxylated bis-THF rings are important structural factors required for potent inhibition. The inhibitory potency of a derivative possessing an n-butylphenyl ether structure (compound 7) appeared to be superior to that of the original ∆lac-acetogenins and equivalent to that of bullatacin, one of the most potent natural acetogenins. Double-inhibitor titration of steady-state complex I activity showed that the extent of inhibition of compound 7 and bullatacin is not additive, suggesting that the binding sites of the two inhibitors are not identical. Competition tests using a fluorescent ligand indicated that the binding site of compound 7 does not overlap with that of other complex I inhibitors. The effects of compound 7 on superoxide production from complex I are also different from those of other complex I inhibitors. Our results clearly demonstrate that ∆lac-acetogenins are a novel type of inhibitor acting at the terminal electron-transfer step of bovine complex I.

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“…− production by complex I under basal conditions in a process that involves reduced ubiquinone. 47,48 NDUFS2 establishes a groove with NDUFS7 to form the ubiquinone binding site. [23][24][25] GB cleavage of NDUFS2 may alter the ubiquinone binding site to favor ROS production in the reverse electron transfer mode.…”

Section: Discussionmentioning

confidence: 99%

Granzyme B-induced mitochondrial ROS are required for apoptosis

et al. 2014

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Caspases and the cytotoxic lymphocyte protease granzyme B (GB) induce reactive oxygen species (ROS) formation, loss of transmembrane potential and mitochondrial outer membrane permeabilization (MOMP). Whether ROS are required for GBmediated apoptosis and how GB induces ROS is unclear. Here, we found that GB induces cell death in an ROS-dependent manner, independently of caspases and MOMP. GB triggers ROS increase in target cell by directly attacking the mitochondria to cleave NDUFV1, NDUFS1 and NDUFS2 subunits of the NADH: ubiquinone oxidoreductase complex I inside mitochondria. This leads to mitocentric ROS production, loss of complex I and III activity, disorganization of the respiratory chain, impaired mitochondrial respiration and loss of the mitochondrial cristae junctions. Furthermore, we have also found that GB-induced mitocentric ROS are necessary for optimal apoptogenic factor release, rapid DNA fragmentation and lysosomal rupture. Interestingly, scavenging the ROS delays and reduces many of the features of GB-induced death. Consequently, GB-induced ROS significantly promote apoptosis. Cell Death and Differentiation (2015) 22, 862-874; doi:10.1038/cdd.2014.180; published online 31 October 2014To induce cell death, human granzyme B (GB) activates effector caspase-3 or acts directly on key caspase substrates, such as the proapoptotic BH3 only Bcl-2 family member Bid, inhibitor of caspase-activated DNase (ICAD), poly-(ADPribose) polymerase-1 (PARP-1), lamin B, nuclear mitotic apparatus protein 1 (NUMA1), catalytic subunit of the DNAdependent protein kinase (DNA-PKcs) and tubulin. [1][2][3] Consequently, caspase inhibitors have little effect on human GB-mediated cell death and DNA fragmentation. 2 GB causes reactive oxygen species (ROS) production, dissipation of the mitochondrial transmembrane potential (ΔΨm) and MOMP, which leads to the release of apoptogenic factors such as cytochrome c (Cyt c), HtrA2/Omi, endonuclease G (Endo G), Smac/Diablo and apoptosis-inducing factor, from the mitochondrial intermembrane space to the cytosol. [4][5][6][7][8][9][10][11] Interestingly, cells deficient for Bid, Bax and Bak are still sensitive to human GB-induced cell death, 5,11-13 suggesting that human GB targets the mitochondria in another way that needs to be characterized. Altogether, much attention has been focused on the importance of MOMP in the execution of GB-mediated cell death, leaving unclear whether ROS production is a bystander effect or essential to the execution of GBinduced apoptosis. The mitochondrial NADH: ubiquinone oxidoreductase complex I is a key determinant in steadystate ROS production. This 1 MDa complex, composed of 44 subunits, 14 couples the transfer of two electrons from NADH to ubiquinone with the translocation of four protons to generate the ΔΨm. The importance of ROS has been previously demonstrated for caspase-3 and granzyme A (GA) pathways through the cleavage of NDUFS1 and NDUFS3, respectively. [15][16][17][18] GA induces cell death in a Bcl-2-insensitive and caspase-and MOMP-indepen...

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Inhibitors of the Quinone-binding Site Allow Rapid Superoxide Production from Mitochondrial NADH:Ubiquinone Oxidoreductase (Complex I)

Cited by 430 publications

References 33 publications

N‐acetylcysteine impairs survival of luteal cells through mitochondrial dysfunction

N‐acetylcysteine impairs survival of luteal cells through mitochondrial dysfunction

Synthesis and Inhibition Mechanism of Δlac-Acetogenins, a Novel Type of Inhibitor of Bovine Heart Mitochondrial Complex I

Granzyme B-induced mitochondrial ROS are required for apoptosis

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