The function of the respiratory supercomplexes: the plasticity model

Mitochondria are important organelles not only as efficient ATP generators but also in controlling and regulating many cellular processes. Mitochondria are dynamic compartments that rearrange under stress response and changes in food availability or oxygen concentrations. The mitochondrial electron transport chain parallels these rearrangements to achieve an optimum performance and therefore requires a plastic organization within the inner mitochondrial membrane. This consists in a balanced distribution between free respiratory complexes and supercomplexes. The mechanisms by which the distribution and organization of supercomplexes can be adjusted to the needs of the cells are still poorly understood. The aim of this review is to focus on the functional role of the respiratory supercomplexes and its relevance in physiology. This article is part of a Special Issue entitled: Dynamic and ultrastructure of bioenergetic membranes and their components.This study was supported by grants from the Ministerio de Ciencia e Innovación (SAF2012-32776 & CSD200700020); the Comunidad de Madrid (CAM/API1009); and the Marie Curie Career Integration Grant (UEO/MCA1108). RA-P is an investigator of the Ramon y Cajal research program from the Ministerio de Economía y Competitividad. The CNIC is supported by the Ministerio de Economía y Competitividad and the Pro-CNIC Foundation.S

Protein Phosphorylation and Prevention of Cytochrome Oxidase Inhibition by ATP: Coupled Mechanisms of Energy Metabolism Regulation

SummaryRapid regulation of oxidative phosphorylation is crucial for mitochondrial adaptation to swift changes in fuels availability and energy demands. An intramitochondrial signaling pathway regulates cytochrome oxidase (COX), the terminal enzyme of the respiratory chain, through reversible phosphorylation. We find that PKA-mediated phosphorylation of a COX subunit dictates mammalian mitochondrial energy fluxes and identify the specific residue (S58) of COX subunit IV-1 (COXIV-1) that is involved in this mechanism of metabolic regulation. Using protein mutagenesis, molecular dynamics simulations, and induced fit docking, we show that mitochondrial energy metabolism regulation by phosphorylation of COXIV-1 is coupled with prevention of COX allosteric inhibition by ATP. This regulatory mechanism is essential for efficient oxidative metabolism and cell survival. We propose that S58 COXIV-1 phosphorylation has evolved as a metabolic switch that allows mammalian mitochondria to rapidly toggle between energy utilization and energy storage

MicroRNA-661 modulates redox and metabolic homeostasis in colon cancer

Cancer cell survival and metastasis are dependent on metabolic reprogramming that is capable of increasing resistance to oxidative and energetic stress. Targeting these two processes can be crucial for cancer progression. Herein, we describe the role of microRNA-661 (miR661) as epigenetic regulator of colon cancer (CC) cell metabolism. MicroR661 induces a global increase in reactive oxygen species, specifically in mitochondrial superoxide anions, which appears to be mediated by decreased carbohydrate metabolism and pentose phosphate pathway, and by a higher dependency on mitochondrial respiration. MicroR661 overexpression in non-metastatic human CC cells induces an epithelial-to-mesenchymal transition phenotype, and a reduced tolerance to metabolic stress. This seems to be a general effect of miR661 in CC, since metastatic CC cell metabolism is also compromised upon miR661 overexpression. We propose hexose-6-phosphate dehydrogenase and pyruvate kinase M2 as two key players related to the observed metabolic reprogramming. Finally, the clinical relevance of miR661 expression levels in stage-II and III CC patients is discussed. In conclusion, we propose miR661 as a potential modulator of redox and metabolic homeostasis in CC.This work was supported by Ministerio de Econom ıa y Competitividad del Gobierno de España (MINECO/FEDER Plan Nacional I+D+i AGL201348943-C2 and AGL2016-76736-C3-3-R), Gobierno regional de la Comunidad de Madrid (P2013/ABI2728, ALIBIRD-CM) and EU Structural Funds.S

Are Zinc-Finger Domains of Protein Kinase C Dynamic Structures That Unfold by Lipid or Redox Activation?

Protein kinase C (PKC) is activated by lipid second messengers or redox action, raising the question whether these activation modes involve the same or alternate mechanisms. Here we show that both lipid activators and oxidation target the zinc-finger domains of PKC, suggesting a unifying activation mechanism. We found that lipid agonist-binding or redox action leads to zinc release and disassembly of zinc fingers, thus triggering large-scale unfolding that underlies conversion to the active enzyme. These results suggest that PKC zinc fingers, originally considered purely structural devices, are in fact redox-sensitive flexible hinges, whose conformation is controlled both by redox conditions and lipid agonists. Antioxid. Redox Signal. 14, 757-766.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90473/1/ars-2E2010-2E3773.pd

Activation of Serine One-Carbon Metabolism by Calcineurin A beta 1 Reduces Myocardial Hypertrophy and Improves Ventricular Function

BACKGROUND In response to pressure overload, the heart develops ventricular hypertrophy that progressively decompensates and leads to heart failure. This pathological hypertrophy is mediated, among others, by the phosphatase calcineurin and is characterized by metabolic changes that impair energy production by mitochondria. OBJECTIVES The authors aimed to determine the role of the calcineurin splicing variant CnA beta 1 in the context of cardiac hypertrophy and its mechanism of action. METHODS Transgenic mice overexpressing CnAb1 specifically in cardiomyocytes and mice lacking the unique C-terminal domain in CnA beta 1 (CnA beta 1(Delta i12) mice) were used. Pressure overload hypertrophy was induced by transaortic constriction. Cardiac function was measured by echocardiography. Mice were characterized using various molecular analyses. RESULTS In contrast to other calcineurin isoforms, the authors show here that cardiac-specific overexpression of CnA beta 1 in transgenic mice reduces cardiac hypertrophy and improves cardiac function. This effect is mediated by activation of serine and one-carbon metabolism, and the production of antioxidant mediators that prevent mitochondrial protein oxidation and preserve ATP production. The induction of enzymes involved in this metabolic pathway by CnAb1 is dependent on mTOR activity. Inhibition of serine and one-carbon metabolism blocks the beneficial effects of CnA beta 1. CnA beta 1(Delta i12) mice show increased cardiac hypertrophy and declined contractility. CONCLUSIONS The metabolic reprogramming induced by CnAb1 redefines the role of calcineurin in the heart and shows for the first time that activation of the serine and one-carbon pathway has beneficial effects on cardiac hypertrophy and function, paving the way for new therapeutic approaches. (J Am Coll Cardiol 2018; 71: 654-67) (C) 2018 The Authors. Published by Elsevier on behalf of the American College of Cardiology Foundation. This is an open access article under the CC BY-NC-ND license (http://creativecommons. org/licenses/by-nc-nd/4.0/).This work was supported by grants from the European Union (CardioNeT-ITN-289600 and CardioNext-608027 to Dr. Lara-Pezzi; Meet-ITN-317433 to Dr. Enriquez; UE0/MCA1108 to Dr. Acin-Perez), from the Spanish Ministry of Economy and Competitiveness (SAF2015-65722-R and SAF2012-31451 to Dr. Lara-Pezzi; SAF2015-71521-REDC, BFU2013-50448, and SAF2012-32776 to Dr. Enriquez; RyC-2011-07826 to Dr. Acin-Perez; BIO2012-37926 and BIO2015-67580-P to Dr. Vazquez), from the Spanish Carlos III Institute of Health (CPII14/00027 to Dr. Lara-Pezzi; RD12/0042/066 to Drs. Garcia-Pavia and Lara-Pezzi), from the Regional Government of Madrid (2010-BMD-2321 ``Fibroteam´´ to Dr. Lara-Pezzi; 2011-BMD-2402 ``Mitolab´´ to Dr. Enriquez) and the FIS-ISCIII (PRB2-IPT13/0001 and RD12/0042/0056-RIC-RETICS to Dr. Vazquez). This work was also supported by the Plan Estatal de IthornDthornI 2013-2016-European Regional Development Fund (FEDER) ``A way of making Europe,´´ Spain. The CNIC is supported by the Spanish Ministry of Economy and Competitiveness and by the Pro-CNIC Foundation and is a Severo Ochoa Center of Excellence (MINECO award SEV-2015-0505). Drs. Vazquez and Garcia-Pavia have served as consultants for VL39. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose. Drs. Padron-Barthe, Villalba-Orero, and Gomez-Salinero contributed equally to this work and are joint first authors. Robyn Shaw, MD, PhD, served as Guest Editor for this paper.S

Increased localization of APP-C99 in mitochondria-associated ER membranes causes mitochondrial dysfunction in Alzheimer disease

In the amyloidogenic pathway associated with Alzheimer disease (AD), the amyloid precursor protein (APP) is cleaved by beta-secretase to generate a 99-aa C-terminal fragment (C99) that is then cleaved by c-secretase to generate the beta-amyloid (Ab) found in senile plaques. In previous reports, we and others have shown that c-secretase activity is enriched in mitochondria-associated endoplasmic reticulum (ER) membranes (MAM) and that ER-mitochondrial connectivity and MAM function are upregulated in AD. We now show that C99, in addition to its localization in endosomes, can also be found in MAM, where it is normally processed rapidly by c-secretase. In cell models of AD, however, the concentration of unprocessed C99 increases in MAM regions, resulting in elevated sphingolipid turnover and an altered lipid composition of both MAM and mitochondrial membranes. In turn, this change in mitochondrial membrane composition interferes with the proper assembly and activity of mitochondrial respiratory supercomplexes, thereby likely contributing to the bioenergetic defects characteristic of AD.We thank Drs. Orian Shirihai and Marc Liesa (UCLA) for assistance with the Seahorse measurements, Dr. Huaxi Xu (Sanford Burnham Institute) for the APP-DKO MEFs and Dr. Mark Mattson (NIH) for the PS1 knock-in mice, Drs. Arancio and Teich for the APP-KO mice tissues used in these studies, Dr. Hua Yang (Columbia University) for mouse husbandry, and Drs. Marc Tambini, Ira Tabas, and Serge Przedborski for helpful comments. This work was supported by the Fundacion Alfonso Martin Escudero (to M.P.); the Alzheimer's Drug Discovery Foundation, the Ellison Medical Foundation, the Muscular Dystrophy Association, the U.S. Department of Defense W911NF-12-1-9159 and W911F-15-1-0169), and the J. Willard and Alice S. Marriott Foundation (to E.A.S.); the U.S. National Institutes of Health (P01-HD080642 and P01-HD032062 to E.A.S.; NS071571 and HD071593 to M.F.M.; R01-NS056049 and P50-AG008702 to G.D.P.; 1S10OD016214-01A1 to G.S.P. and F.P.M, and K01-AG045335 to E.A.-G.), the Lucien Cote Early Investigator Award in Clinical Genetics from the Parkinson's Disease Foundation (PDF-CEI-1364 and PDF-CEI-1240) to C.G.-L., and National Defense Science and Engineering Graduate Fellowship (FA9550-11-C-0028) to R.R.A.S

Allotopic expression of mitochondrial-encoded genes in mammals: achieved goal, undemonstrated mechanism or impossible task?

Mitochondrial-DNA diseases have no effective treatments. Allotopic expression—synthesis of a wild-type version of the mutated protein in the nuclear-cytosolic compartment and its importation into mitochondria—has been proposed as a gene-therapy approach. Allotopic expression has been successfully demonstrated in yeast, but in mammalian mitochondria results are contradictory. The evidence available is based on partial phenotype rescue, not on the incorporation of a functional protein into mitochondria. Here, we show that reliance on partial rescue alone can lead to a false conclusion of successful allotopic expression. We recoded mitochondrial mt-Nd6 to the universal genetic code, and added the N-terminal mitochondrial-targeting sequence of cytochrome c oxidase VIII (C8) and the HA epitope (C8Nd6HA). The protein apparently co-localized with mitochondria, but a significant part of it seemed to be located outside mitochondria. Complex I activity and assembly was restored, suggesting successful allotopic expression. However, careful examination of transfected cells showed that the allotopically-expressed protein was not internalized in mitochondria and that the selected clones were in fact revertants for the mt-Nd6 mutation. These findings demonstrate the need for extreme caution in the interpretation of functional rescue experiments and for clear-cut controls to demonstrate true rescue of mitochondrial function by allotopic expression

Mitochondrial DNA mutations affect calcium handling in differentiated neurons

Mutations in the mitochondrial genome are associated with a wide range of neurological symptoms, but many aspects of the basic neuronal pathology are not understood. One candidate mechanism, given the well-established role of mitochondria in calcium buffering, is a deficit in neuronal calcium homoeostasis. We therefore examined calcium responses in the neurons derived from various ‘cybrid’ embryonic stem cell lines carrying different mitochondrial DNA mutations. Brief (∼50 ms), focal glutamatergic stimuli induced a transient rise in intracellular calcium concentration, which was visualized by bulk loading the cells with the calcium dye, Oregon Green BAPTA-1. Calcium entered the neurons through N-methyl-d-aspartic acid and voltage-gated calcium channels, as has been described in many other neuronal classes. Intriguingly, while mitochondrial mutations did not affect the calcium transient in response to single glutamatergic stimuli, they did alter the responses to repeated stimuli, with each successive calcium transient decaying ever more slowly in mitochondrial mutant cell lines. A train of stimuli thus caused intracellular calcium in these cells to be significantly elevated for many tens of seconds. These results suggest that calcium-handling deficits are likely to contribute to the pathological phenotype seen in patients with mitochondrial DNA mutations