Inefficient thermogenic mitochondrial respiration due to futile proton leak in a mouse model of fragile X syndrome.
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ABSTRACT:
SUBMITTER: Griffiths KK
PROVIDER: S-EPMC7692004 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:By measuring the relationship between protonmotive force and the increment in oxygen consumption by mitochondria treated with submaximal amounts of uncoupler, we have experimentally tested four different models of imperfect coupling of oxidative phosphorylation. The results show that the increased rate of oxygen consumption at high protonmotive force is explained entirely by the dependence on protonmotive force of the passive proton leak conductance of the mitochondrial inner membrane. There is no measurable contribution from redox-slip reactions in the proton pumps caused by high protonmotive force. Neither is there any contribution from increased proton conductance of the membrane or increased redox slip in the respiratory chain caused by high turnover rates of the complexes.
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Project description:An alteration of mitochondrial function can result in disruption of redox homeostasis and is associated with abnormal cancer cell growth. Manganese superoxide dismutase (SOD2) and glutathione peroxidase 4 (Gpx4) are two of the most important antioxidant defense enzymes that protect cells against oxidative stress. We had previously shown that n-3 polyunsaturated fatty acids (PUFA) promote colonocyte apoptosis, a marker of colon cancer risk, in part by enhancing phospholipid oxidation. To elucidate the mechanisms regulating oxidative stress-induced apoptosis in vivo, we fed heterozygous SOD2(Het), Gpx4(Het), and transgenic Gpx4(Tg) mice diets containing either 15% corn oil by weight (CO, enriched in n-6 PUFA) or 3.5% CO + 11.5% fish oil (FO, enriched in n-3 PUFA) for 4 weeks. Our data showed that (i) genetic predeposition to oxidative stress facilitates apoptosis in the mouse colon (Gpx4(Het) > SOD2(Het) > Wt > Gpx4(Tg)), (ii) dietary n-3 PUFA have an additive effect on the induction of apoptosis in Gpx4(Het) and SOD2(Het) mice; and (iii) dietary n-3 PUFA reverse the phenotype in oxidatively protected Gpx4(Tg) mice by elevating apoptosis to a level observed in wild-type (Wt; control) animals. Complimentary experiments examining colonic mitochondrial bioenergetic profiles indicate that FO-fed mice exhibit a significantly (P < 0.05) increased respiration-induced proton leak relative to control CO treatment. This finding was consistent with a loss of membrane potential in response to chronic oxidative stress and supports the contention that n-3 PUFA alter mitochondrial metabolic activity, thereby enhancing apoptosis and reducing colon cancer risk.
| S-EPMC3137683 | biostudies-literature
Project description:In this study, we compared the influence of GDP and GTP on isolated mitochondria respiring under conditions favoring oxidative phosphorylation (OXPHOS) and under conditions excluding this process, i.e., in the presence of carboxyatractyloside, an adenine nucleotide translocase inhibitor, and/or oligomycin, an FOF1-ATP synthase inhibitor. Using mitochondria isolated from rat kidney and human endothelial cells, we found that the action of GDP and GTP can differ diametrically depending on the conditions. Namely, under conditions favoring OXPHOS, both in the absence and presence of linoleic acid, an activator of uncoupling proteins (UCPs), the addition of 1 mM GDP resulted in the state 4 (non-phosphorylating respiration)-state 3 (phosphorylating respiration) transition, which is characteristic of ADP oxidative phosphorylation. In contrast, the addition of 1 mM GTP resulted in a decrease in the respiratory rate and an increase in the membrane potential, which is characteristic of UCP inhibition. The stimulatory effect of GDP, but not GTP, was also observed in inside-out submitochondrial particles prepared from rat kidney mitochondria. However, the effects of GDP and GTP were more similar in the presence of OXPHOS inhibitors. The importance of these observations in connection with the action of UCPs, adenine nucleotide translocase (or other carboxyatractyloside-sensitive carriers), carboxyatractyloside- and purine nucleotide-insensitive carriers, as well as nucleoside-diphosphate kinase (NDPK) are considered. Because the measurements favoring oxidative phosphorylation better reflect in vivo conditions, our study strongly supports the idea that GDP cannot be considered a significant physiological inhibitor of UCP. Moreover, it appears that, under native conditions, GTP functions as a more efficient UCP inhibitor than GDP and ATP.
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Project description:Fasting hyperinsulinemia precedes the development of type 2 diabetes. However, it is unclear whether fasting insulin hypersecretion is a primary driver of insulin resistance or a consequence of the progressive increase in fasting glycemia induced by insulin resistance in the prediabetic state. Herein, we have discovered a mechanism that specifically regulates non-glucose-stimulated insulin secretion (NGSIS) in pancreatic islets that is activated by nonesterified free fatty acids, the major fuel used by β-cells during fasting. We show that the mitochondrial permeability transition pore regulator cyclophilin D (CypD) promotes NGSIS, but not glucose-stimulated insulin secretion, by increasing mitochondrial proton leak. Islets from prediabetic obese mice show significantly higher CypD-dependent proton leak and NGSIS compared with lean mice. Proton leak-mediated NGSIS is conserved in human islets and is stimulated by exposure to nonesterified free fatty acids at concentrations observed in obese subjects. Mechanistically, proton leak activates islet NGSIS independently of mitochondrial ATP synthesis but ultimately requires closure of the KATP channel. In summary, we have described a novel nonesterified free fatty acid-stimulated pathway that selectively drives pancreatic islet NGSIS, which may be therapeutically exploited as an alternative way to halt fasting hyperinsulinemia and the progression of type 2 diabetes.
| S-EPMC6971491 | biostudies-literature
Project description:Background?-synuclein (SNCA) accumulation in the substantia nigra is one of the characteristic pathologies of Parkinson's disease (PD). A53T missense mutations in the SNCA gene has been proved to enhance the expression of SNCA and accelerate the onset of PD. Mitochondrial dysfunction in SNCA aggregation has been under debate for decades but the causal relationship remains uncertain. At a later stage of PD, the cellular dysfunctions are complicated and multiple factors are tangled. Our aim here is to investigate the mitochondrial functional changes and clarify the main causal mechanism at earlier-stage of PD.MethodsWe used the mutant A53T SNCA-expressed neuro 2a (N2a) cells without detectable cell death to investigate: 1) whether SNCA overexpression impairs the mitochondrial respiration and biogenesis. 2) The role of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) signal in SNCA-induced mitochondria dysfunction.ResultsAccompanying with the increment of SNCA, reactive oxygen species (ROS) accumulation was increased. The maximal respiratory capacity was suppressed. Meanwhile, mitochondrial complex 1 activity and the activity of nicotinamide adenine dinucleotide (NADH) cytochrome C reductase (NCCR) were decreased. Moreover, the mitochondrial DNA (mtDNA) copy number was decreased. On the other hand, the nuclear peroxisome proliferator-activated receptor-gamma coactivator 1? (PGC-1?), Nrf2, and the cytosolic mitochondrial transcription factor A (TFAM) were increased at an early stage and declined thereafter. Above factors triggered by SNCA were reversed by tBHQ, a Nrf2 activator.ConclusionThese results suggested that at an early stage, SNCA-overexpressed increase mtROS accumulation, mitochondrial dysfunction and mtDNA decrement. Nrf2, PGC-1? and TFAM were upregulated to compromise mitochondrial dysfunction. tBHQ effectively reversed the SNCA-induced mitochondrial dysfunction.
| S-EPMC6138761 | biostudies-literature
Project description:The mechanisms of mitochondrial proton (H+) leak under various pathophysiological conditions are poorly understood. In the present study it was hypothesized that different mechanisms underlie H+ leak in cardiac IR (ischaemia/reperfusion) injury and IPC (ischaemic preconditioning). Potential H(+) leak mechanisms examined were UCPs (uncoupling proteins), allosteric activation of the ANT (adenine nucleotide translocase) by AMP, or the PT (permeability transition) pore. Mitochondria isolated from perfused rat hearts that were subjected to IPC exhibited a greater H+ leak than did controls (202+/-27%, P<0.005), and this increased leakage was completely abolished by the UCP inhibitor, GDP, or the ANT inhibitor, CAT (carboxyattractyloside). Mitochondria from hearts subjected to IR injury exhibited a much greater amount of H+ leak than did controls (411+/-28%, P<0.001). The increased leakage after IR was weakly inhibited by GDP, but was inhibited, >50%, by carboxyattractyloside. In addition, it was inhibited by cardioprotective treatment strategies including pre-IR perfusion with the PT pore inhibitors cyclosporin A or sanglifehrin A, the adenylate kinase inhibitor, AP5A (diadenosine pentaphosphate), or IPC. Together these data suggest that the small increase in H+ leak in IPC is mediated by UCPs, while the large increase in H+ leak in IR is mediated by the ANT. Furthermore, under all conditions studied, in situ myocardial O2 efficiency was correlated with isolated mitochondrial H+ leak (r2=0.71). In conclusion, these data suggest that the modulation of H+ leak may have important implications for the outcome of IR injury.
| S-EPMC1462692 | biostudies-other
Project description:Li-Fraumeni syndrome (LFS) is a cancer predisposition disorder caused by germline mutations in TP53 that can lead to increased mitochondrial metabolism in patients. However, the implications of altered mitochondrial function for tumorigenesis in LFS are unclear. Here, we have reported that genetic or pharmacologic disruption of mitochondrial respiration improves cancer-free survival in a mouse model of LFS that expresses mutant p53. Mechanistically, inhibition of mitochondrial function increased autophagy and decreased the aberrant proliferation signaling caused by mutant p53. In a pilot study, LFS patients treated with metformin exhibited decreases in mitochondrial activity concomitant with activation of antiproliferation signaling, thus reproducing the effects of disrupting mitochondrial function observed in LFS mice. These observations indicate that a commonly prescribed diabetic medicine can restrain mitochondrial metabolism and tumorigenesis in an LFS model, supporting its further consideration for cancer prevention in LFS patients.
| S-EPMC5199691 | biostudies-literature
Project description:Loss of the gene (Fmr1) encoding Fragile X mental retardation protein (FMRP) causes increased mRNA translation and aberrant synaptic development. We find neurons of the Fmr1-/y mouse have a mitochondrial inner membrane leak contributing to a "leak metabolism." In human Fragile X syndrome (FXS) fibroblasts and in Fmr1-/y mouse neurons, closure of the ATP synthase leak channel by mild depletion of its c-subunit or pharmacological inhibition normalizes stimulus-induced and constitutive mRNA translation rate, decreases lactate and key glycolytic and tricarboxylic acid (TCA) cycle enzyme levels, and triggers synapse maturation. FMRP regulates leak closure in wild-type (WT), but not FX synapses, by stimulus-dependent ATP synthase β subunit translation; this increases the ratio of ATP synthase enzyme to its c-subunit, enhancing ATP production efficiency and synaptic growth. In contrast, in FXS, inability to close developmental c-subunit leak prevents stimulus-dependent synaptic maturation. Therefore, ATP synthase c-subunit leak closure encourages development and attenuates autistic behaviors.
| S-EPMC7484101 | biostudies-literature
Project description:The kinetics of the formation and relaxation of transmembrane electric potential (Deltapsi) during the complete single turnover of CcO was studied in the bovine heart mitochondrial and the aa(3)-type Paracoccus denitrificans enzymes incorporated into proteoliposome membrane. The real-time Deltapsi kinetics was followed by the direct electrometry technique. The prompt oxidation of CcO and formation of the activated, oxidized (O(H)) state of the enzyme leaves the enzyme trapped in the open state that provides an internal leak for protons and thus facilitates dissipation of Deltapsi (tau(app) < or = 0.5-0.8 s). By contrast, when the enzyme in the O(H) state is rapidly re-reduced by sequential electron delivery, Deltapsi dissipates much slower (tau(app) > 3 s). In P. denitrificans CcO proteoliposomes the accelerated Deltapsi dissipation is slowed down by a mutational block of the proton conductance through the D-, but not K-channel. We concluded that in contrast to the other intermediates the O(H) state of CcO is vulnerable to the elevated internal proton leak that proceeds via the D-channel.
| S-EPMC2711495 | biostudies-literature
Project description:BackgroundType 2 diabetes mellitus and muscle insulin resistance have been associated with reduced capacity of skeletal muscle mitochondria, possibly as a result of increased intake of dietary fat. Here, we examined the hypothesis that a prolonged high-fat diet consumption (HFD) increases the saturation of muscle mitochondrial membrane phospholipids causing impaired mitochondrial oxidative capacity and possibly insulin resistance.MethodologyC57BL/6J mice were fed an 8-week or 20-week low fat diet (10 kcal%; LFD) or HFD (45 kcal%). Skeletal muscle mitochondria were isolated and fatty acid (FA) composition of skeletal muscle mitochondrial phospholipids was analyzed by thin-layer chromatography followed by GC. High-resolution respirometry was used to assess oxidation of pyruvate and fatty acids by mitochondria. Insulin sensitivity was estimated by HOMA-IR.Principal findingsAt 8 weeks, mono-unsaturated FA (16?1n7, 18?1n7 and 18?1n9) were decreased (-4.0%, p<0.001), whereas saturated FA (16?0) were increased (+3.2%, p<0.001) in phospholipids of HFD vs. LFD mitochondria. Interestingly, 20 weeks of HFD descreased mono-unsaturated FA while n-6 poly-unsaturated FA (18?2n6, 20?4n6, 22?5n6) showed a pronounced increase (+4.0%, p<0.001). Despite increased saturation of muscle mitochondrial phospholipids after the 8-week HFD, mitochondrial oxidation of both pyruvate and fatty acids were similar between LFD and HFD mice. After 20 weeks of HFD, the increase in n-6 poly-unsaturated FA was accompanied by enhanced maximal capacity of the electron transport chain (+49%, p?=?0.002) and a tendency for increased ADP-stimulated respiration, but only when fuelled by a lipid-derived substrate. Insulin sensitivity in HFD mice was reduced at both 8 and 20 weeks.Conclusions/interpretationOur findings do not support the concept that prolonged HF feeding leads to increased saturation of skeletal muscle mitochondrial phospholipids resulting in a decrease in mitochondrial fat oxidative capacity and (muscle) insulin resistance.
| S-EPMC3225362 | biostudies-literature