PGC-1α is Dispensable for Exercise-Induced Mitochondrial Biogenesis in Skeletal Muscle

Exercise confers numerous health benefits, many of which are thought to stem from exercise-induced mitochondrial biogenesis (EIMB) in skeletal muscle. The transcriptional coactivator PGC-1α, a potent regulator of metabolism in numerous tissues, is widely believed to be required for EIMB. We show here that this is not the case. Mice engineered to lack PGC-1α specifically in skeletal muscle (Myo-PGC-1αKO mice) retained intact EIMB. The exercise capacity of these mice was comparable to littermate controls. Induction of metabolic genes after 2 weeks of in-cage voluntary wheel running was intact. Electron microscopy revealed no gross abnormalities in mitochondria, and the mitochondrial biogenic response to endurance exercise was as robust in Myo-PGC-1αKO mice as in wildtype mice. The induction of enzymatic activity of the electron transport chain by exercise was likewise unperturbed in Myo-PGC-1αKO mice. These data demonstrate that PGC-1α is dispensable for exercise-induced mitochondrial biogenesis in skeletal muscle, in sharp contrast to the prevalent assumption in the field

Translocator Protein-Mediated Stabilization of Mitochondrial Architecture during Inflammation Stress in Colonic Cells.

International audienceChronic inflammation of the gastrointestinal tract increasing the risk of cancer has been described to be linked to the high expression of the mitochondrial translocator protein (18 kDa; TSPO). Accordingly, TSPO drug ligands have been shown to regulate cytokine production and to improve tissue reconstruction. We used HT-29 human colon carcinoma cells to evaluate the role of TSPO and its drug ligands in tumor necrosis factor (TNF)-induced inflammation. TNF-induced interleukin (IL)-8 expression, coupled to reactive oxygen species (ROS) production, was followed by TSPO overexpression. TNF also destabilized mitochondrial ultrastructure, inducing cell death by apoptosis. Treatment with the TSPO drug ligand PK 11195 maintained the mitochondrial ultrastructure, reducing IL-8 and ROS production and cell death. TSPO silencing and overexpression studies demonstrated that the presence of TSPO is essential to control IL-8 and ROS production, so as to maintain mitochondrial ultrastructure and to prevent cell death. Taken together, our data indicate that inflammation results in the disruption of mitochondrial complexes containing TSPO, leading to cell death and epithelia disruption. This work implicates TSPO in the maintenance of mitochondrial membrane integrity and in the control of mitochondrial ROS production, ultimately favoring tissue regeneration

Friedreich's ataxia: the vicious circle hypothesis revisited

Friedreich's ataxia, the most frequent progressive autosomal recessive disorder involving the central and peripheral nervous systems, is mostly associated with unstable expansion of GAA trinucleotide repeats in the first intron of the FXN gene, which encodes the mitochondrial frataxin protein. Since FXN was shown to be involved in Friedreich's ataxia in the late 1990s, the consequence of frataxin loss of function has generated vigorous debate. Very early on we suggested a unifying hypothesis according to which frataxin deficiency leads to a vicious circle of faulty iron handling, impaired iron-sulphur cluster synthesis and increased oxygen radical production. However, data from cell and animal models now indicate that iron accumulation is an inconsistent and late event and that frataxin deficiency does not always impair the activity of iron-sulphur cluster-containing proteins. In contrast, frataxin deficiency appears to be consistently associated with increased sensitivity to reactive oxygen species as opposed to increased oxygen radical production. By compiling the findings of fundamental research and clinical observations we defend here the opinion that the very first consequence of frataxin depletion is indeed an abnormal oxidative status which initiates the pathogenic mechanism underlying Friedreich's ataxia

In Vivo Detection of Succinate by Magnetic Resonance Spectroscopy as a Hallmark of SDHx Mutations in Paraganglioma

International audiencePurpose: Germline mutations in genes encoding mitochon-drial succinate dehydrogenase (SDH) are found in patients with paragangliomas, pheochromocytomas, gastrointestinal stromal tumors, and renal cancers. SDH inactivation leads to a massive accumulation of succinate, acting as an oncometabolite and which levels, assessed on surgically resected tissue are a highly specific biomarker of SDHx-mutated tumors. The aim of this study was to address the feasibility of detecting succinate in vivo by magnetic resonance spectroscopy. Experimental Design: A pulsed proton magnetic resonance spectroscopy (1 H-MRS) sequence was developed, optimized, and applied to image nude mice grafted with Sdhb À/À or wild-type chromaffin cells. The method was then applied to patients with paraganglioma carrying (n ¼ 5) or not (n ¼ 4) an SDHx gene mutation. Following surgery, succinate was measured using gas chromatography/mass spectrometry, and SDH protein expression was assessed by immunohistochemistry in resected tumors. Results: A succinate peak was observed at 2.44 ppm by 1 H-MRS in all Sdhb À/À-derived tumors in mice and in all paragangliomas of patients carrying an SDHx gene mutation, but neither in wild-type mouse tumors nor in patients exempt of SDHx mutation. In one patient, 1 H-MRS results led to the identification of an unsus-pected SDHA gene mutation. In another case, it helped define the pathogenicity of a variant of unknown significance in the SDHB gene. Conclusions: Detection of succinate by 1 H-MRS is a highly specific and sensitive hallmark of SDHx mutations. This non-invasive approach is a simple and robust method allowing in vivo detection of the major biomarker of SDHx-mutated tumors. Clin Cancer Res; 22(5); 1120–9. Ó2015 AACR

Broad AOX expression in a genetically tractable mouse model does not disturb normal physiology

Plants and many lower organisms, but not mammals, express alternative oxidases (AOXs) that branch the mitochondrial respiratory chain, transferring electrons directly from ubiquinol to oxygen without proton pumping. Thus, they maintain electron flow under conditions when the classical respiratory chain is impaired, limiting excess production of oxygen radicals and supporting redox and metabolic homeostasis. AOX from Ciona intestinalis has been used to study and mitigate mitochondrial impairments in mammalian cell lines, Drosophila disease models and, most recently, in the mouse, where multiple lentivector-AOX transgenes conferred substantial expression in specific tissues. Here, we describe a genetically tractable mouse model in which Ciona AOX has been targeted to the Rosa26 locus for ubiquitous expression. The AOX(Rosa26) mouse exhibited only subtle phenotypic effects on respiratory complex formation, oxygen consumption or the global metabolome, and showed an essentially normal physiology. AOX conferred robust resistance to inhibitors of the respiratory chain in organello; moreover, animals exposed to a systemically applied LD50 dose of cyanide did not succumb. The AOX(Rosa26) mouse is a useful tool to investigate respiratory control mechanisms and to decipher mitochondrial disease aetiology in vivo.Peer reviewe

Cyanide resistant respiration and the alternative oxidase pathway : A from to mammals

In a large number of organisms covering all phyla, the mitochondrial respiratory chain harbors, in addition to the conventional elements, auxiliary proteins that confer adaptive metabolic plasticity. The alternative oxidase (AOX) represents one of the most studied auxiliary proteins, initially identified in plants. In contrast to the standard respiratory chain, the AOX mediates a thermogenic cyanide-resistant respiration; a phenomenon that has been of great interest for over 2 centuries in that energy is not conserved when electrons flow through it. Here we summarize centuries of studies starting from the early observations of thermogenicity in plants and the identification of cyanide resistant respiration, to the fascinating discovery of the AOX and its current applications in animals under normal and pathological conditions.Peer reviewe

DAPIT Over-Expression Modulates Glucose Metabolism and Cell Behaviour in HEK293T Cells

Introduction Diabetes Associated Protein in Insulin-sensitive Tissues (DAPIT) is a subunit of mitochondrial ATP synthase and has also been found to associate with the vacuolar H+-ATPase. Its expression is particularly high in cells with elevated aerobic metabolism and in epithelial cells that actively transport nutrients and ions. Deletion of DAPIT is known to induce loss of mitochondrial ATP synthase but the effects of its over-expression are obscure. Results In order to study the consequences of high expression of DAPIT, we constructed a transgenic cell line that constitutively expressed DAPIT in human embryonal kidney cells, HEK293T. Enhanced DAPIT expression decreased mtDNA content and mitochondrial mass, and saturated respiratory chain by decreasing H+-ATP synthase activity. DAPIT over-expression also increased mitochondrial membrane potential and superoxide level, and translocated the transcription factors hypoxia inducible factor 1α (Hif1α) and β-catenin to the nucleus. Accordingly, cells over-expressing DAPIT used more glucose and generated a larger amount of lactate compared to control cells. Interestingly, these changes were associated with an epithelial to mesenchymal (EMT)-like transition by changing E-cadherin to N-cadherin and up-regulating several key junction/adhesion proteins. At physiological level, DAPIT over-expression slowed down cell growth by G1 arrest and migration, and enhanced cell detachment. Several cancers also showed an increase in genomic copy number of Usmg5 (gene encoding DAPIT), thereby providing strong correlative evidence for DAPIT possibly having oncogenic function in cancers. Conclusions DAPIT over-expression thus appears to modulate mitochondrial functions and alter cellular regulations, promote anaerobic metabolism and induce EMT-like transition. We propose that DAPIT over-expression couples the changes in mitochondrial metabolism to physiological and pathophysiological regulations, and suggest it could play a critical role in H+-ATP synthase dysfunctions.Public Library of Science open acces

Mitochondrial activities in human cultured skin fibroblasts contaminated by Mycoplasma hyorhinis

BACKGROUND: Mycoplasma contaminations are a recurrent problem in the use of cultured cells, including human cells, especially as it has been shown to impede cell cycle, triggering cell death under various conditions. More specific consequences on cell metabolism are poorly known. RESULTS: Here we report the lack of significant consequence of a heavy contamination by the frequently encountered mycoplasma strain, M. hyorhinis, on the determination of respiratory chain activities, but the potential interference when assaying citrate synthase. Contamination by M. hyorhinis was detected by fluorescent imaging and further quantified by the determination of the mycoplasma-specific phosphate acetyltransferase activity. Noticeably, this latter activity was not found equally distributed in various mycoplasma types, being exceptionally high in M. hyorhinis. CONCLUSION: While we observed a trend for respiration reduction in heavily contaminated cells, no significant and specific targeting of any respiratory chain components could be identified. This suggested a potential interference with cell metabolism rather than direct interaction with respiratory chain components

Glutathione precursors replenish decreased glutathione pool in cystinotic cell

Abstract Cystinosis is an inherited disorder due to mutations in the CTNS gene which encodes cystinosin, a lysosomal transmembrane protein involved in cystine export to the cytosol. Both accumulation of cystine in the lysosome and decreased cystine in the cytosol may participate in the pathogenic mechanism underlying the disease. We observed that cystinotic cell lines have moderate decrease of glutathione content during exponential growth phase. This resulted in increased solicitation of oxidative defences of the cell denoted by concurrent superoxide dismutase induction, although without major oxidative insult under our experimental conditions. Finally, decreased glutathione content in cystinotic cell lines could be counterbalanced by a series of exogenous precursors of cysteine, denoting that lysosomal cystine export is a natural source of cellular cysteine in the studied cell lines