bims-mibica 2021-10-10 papers

bims-mibica

Biomed News

on Mitochondrial bioenergetics in cancer

Issue of 2021‒10‒10
38 papers selected by
Kelsey Fisher-Wellman, East Carolina University

The assembly, regulation and function of the mitochondrial respiratory chain.
Respiratory Complex I dysfunction in cancer: from a maze of cellular adaptive responses to potential therapeutic strategies.
TGF-β/HDAC7 axis suppresses TCA cycle metabolism in renal cancer.
Coupling of GABA Metabolism to Mitochondrial Glucose Phosphorylation.
The Gro3p factor: Restoring NAD+/NADH homeostasis to ameliorate mitochondrial disease.
PHDs/CPT1B/VDAC1 axis regulates long-chain fatty acid oxidation in cardiomyocytes.
Increased glucosylceramide production leads to decreased cell energy metabolism and lowered tumor marker expression in non-cancerous liver cells.
IL-10-Mediated Refueling of Exhausted T Cell Mitochondria Boosts Anti-Tumour Immunity.
Intercellular transfer of mitochondrial DNA carrying metastasis-enhancing pathogenic mutations from high- to low-metastatic tumor cells and stromal cells via extracellular vesicles.
Petasin potently inhibits mitochondrial complex I-based metabolism that supports tumor growth and metastasis.
Quantification of mtDNA content in cultured cells by direct droplet digital PCR.
The multifaceted ATPase inhibitory factor 1 (IF1) in energy metabolism reprogramming and mitochondrial dysfunction: a new player in age-associated disorders?
TFAM knockdown-triggered mtDNA-nucleoid aggregation and a decrease in mtDNA copy number induce the reorganization of nucleoid populations and mitochondria-associated ER-membrane contacts.
TXNIP links anticipatory unfolded protein response to estrogen reprogramming glucose metabolism in breast cancer cells.
Structural Basis for Inhibition of ROS-Producing Respiratory Complex I by NADH-OH.
Natural tetramic acids elicit multiple inhibitory actions against mitochondrial machineries presiding over oxidative phosphorylation.
Labeling and Measuring Stressed Mitochondria Using a PINK1-Based Ratiometric Fluorescent Sensor.
Proton export alkalinizes intracellular pH and reprograms carbon metabolism to drive hematopoietic progenitor growth.
Mitochondrial Impairment by MitoBloCK-6 Inhibits Liver Cancer Cell Proliferation.
Beyond Base Excision Repair: An evolving picture of mitochondrial DNA repair.
USP29 coordinates MYC and HIF1α stabilization to promote tumor metabolism and progression.
Mitochondrial-targeting nanoprodrugs to mutually reinforce metabolic inhibition and autophagy for combating resistant cancer.
ATP synthase inhibitory factor subunit 1 regulates islet β-cell function via repression of mitochondrial homeostasis.
Protective mitochondrial fission induced by stress-responsive protein GJA1-20k.
Mild mitochondrial impairment enhances inn3ate immunity and longevity through ATFS-1 and p38 signaling.
Sirt1 promotes tissue regeneration in zebrafish through regulating the mitochondrial unfolded protein response.
A Benzenesulfonamide-based Mitochondrial Uncoupler Induces Endoplasmic Reticulum Stress and Immunogenic Cell Death in Epithelial Ovarian Cancer.
Mitochondrial DNA copy number and heteroplasmy load correlate with skeletal muscle oxidative capacity by P31 MR spectroscopy.
Endogenous S-nitrosocysteine proteomic inventories identify a core of proteins in heart metabolic pathways.
Structure and assembly of the mammalian mitochondrial supercomplex CIII2CIV.
Metformin sensitises hepatocarcinoma cells to methotrexate by targeting dihydrofolate reductase.
Green tea catechins EGCG and ECG enhance the fitness and lifespan of Caenorhabditis elegans by complex I inhibition.
PINK1 drives production of mtDNA-containing extracellular vesicles to promote invasiveness.
Dietary Restriction Extends Lifespan through Metabolic Regulation of Innate Immunity.
ALL blasts drive primary mesenchymal stromal cells to increase asparagine availability during Asparaginase treatment.
Therapeutic targeting of SLC6A8 creatine transporter suppresses colon cancer progression and modulates human creatine levels.
Remodelin, a N-acetyltransferase 10 (NAT10) inhibitor, alters mitochondrial lipid metabolism in cancer cells.
CRISPR screens identify cholesterol biosynthesis as a therapeutic target on stemness and drug resistance of colon cancer.

Nat Rev Mol Cell Biol. 2021 Oct 07.

The assembly, regulation and function of the mitochondrial respiratory chain.

Irene Vercellino, Leonid A Sazanov.

The mitochondrial oxidative phosphorylation system is central to cellular metabolism. It comprises five enzymatic complexes and two mobile electron carriers that work in a mitochondrial respiratory chain. By coupling the oxidation of reducing equivalents coming into mitochondria to the generation and subsequent dissipation of a proton gradient across the inner mitochondrial membrane, this electron transport chain drives the production of ATP, which is then used as a primary energy carrier in virtually all cellular processes. Minimal perturbations of the respiratory chain activity are linked to diseases; therefore, it is necessary to understand how these complexes are assembled and regulated and how they function. In this Review, we outline the latest assembly models for each individual complex, and we also highlight the recent discoveries indicating that the formation of larger assemblies, known as respiratory supercomplexes, originates from the association of the intermediates of individual complexes. We then discuss how recent cryo-electron microscopy structures have been key to answering open questions on the function of the electron transport chain in mitochondrial respiration and how supercomplexes and other factors, including metabolites, can regulate the activity of the single complexes. When relevant, we discuss how these mechanisms contribute to physiology and outline their deregulation in human diseases.

DOI: https://doi.org/10.1038/s41580-021-00415-0
FEBS J. 2021 Oct 04.

Respiratory Complex I dysfunction in cancer: from a maze of cellular adaptive responses to potential therapeutic strategies.

Manuela Sollazzo, Monica De Luise, Silvia Lemma, Licia Bressi, Maria Iorio, Stefano Miglietta, Sara Milioni, Ivana Kurelac, Luisa Iommarini, Giuseppe Gasparre, Anna Maria Porcelli.

  Mitochondria act as key organelles in cellular bioenergetics and biosynthetic processes producing signals that regulate different molecular networks for proliferation and cell death. This ability is also preserved in pathologic contexts such as tumorigenesis, during which bioenergetic changes and metabolic reprogramming confer flexibility favoring cancer cells survival in a hostile microenvironment. Although different studies epitomize mitochondrial dysfunction as a pro-tumorigenic hit, genetic ablation or pharmacological inhibition of respiratory Complex I causing a severe impairment are associated with a low proliferative phenotype. In this scenario, it must be considered that despite the initial delay in growth, cancer cells may become able to resume proliferation exploiting molecular mechanisms to overcome growth arrest. Here we highlight the current knowledge on molecular responses activated by Complex I-defective cancer cells to bypass physiological control systems and to re-adapt their fitness during microenvironment changes. Such adaptive mechanisms could reveal possible novel molecular players in synthetic lethality with Complex I impairment, thus providing new synergistic strategies for mitochondria-based anti-cancer therapy.

Keywords:  Respiratory complex I; adaptive responses; cancer metabolism; mitochondria; tumor microenvironment

DOI:  https://doi.org/10.1111/febs.16218
JCI Insight. 2021 Oct 05. pii: e148438. [Epub ahead of print]

TGF-β/HDAC7 axis suppresses TCA cycle metabolism in renal cancer.

Hyeyoung Nam, Anirban Kundu, Suman Karki, Garrett J Brinkley, Darshan S Chandrashekar, Richard L Kirkman, Juan Liu, Maria V Liberti, Jason W Locasale, Tanecia Mitchell, Sooryanarayana Varambally, Sunil Sudarshan.

  Mounting evidence points to alterations in mitochondrial metabolism in renal cell carcinoma (RCC). However, the mechanisms that regulate the TCA cycle in RCC remain uncharacterized. Here, we demonstrate that loss of TCA cycle enzyme expression is retained in RCC metastatic tissues. Moreover, proteomic analysis demonstrates that reduced TCA cycle enzyme expression is far more pronounced in RCC relative to other tumor types. Loss of TCA cycle enzyme expression is correlated with reduced expression of the transcription factor peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) which is also lost in RCC tissues. PGC-1α re-expression in RCC cells restores the expression of TCA cycle enzymes in vitro and in vivo and leads to enhanced glucose carbon incorporation into TCA cycle intermediates. Mechanistically, TGF-β signaling, in concert with histone deacetylase 7 (HDAC7), suppresses TCA cycle enzyme expression. Our studies show that pharmacologic inhibition of TGF-β restores the expression of TCA cycle enzymes and suppresses tumor growth in an orthotopic model of RCC. Taken together, this investigation reveals a novel role for the TGF-β/HDAC7 axis in global suppression of TCA cycle enzymes in RCC and provides new insight into the molecular basis of altered mitochondrial metabolism in this malignancy.

Keywords:  Cancer; Cell Biology; Mitochondria; Molecular biology

DOI:  https://doi.org/10.1172/jci.insight.148438
Neurochem Res. 2021 Oct 08.

Coupling of GABA Metabolism to Mitochondrial Glucose Phosphorylation.

Joao Paulo Cavalcanti-de-Albuquerque, Eduardo de-Souza-Ferreira, Denise Pires de Carvalho, Antonio Galina.

  Glucose and oxygen (O2) are vital to the brain. Glucose metabolism and mitochondria play a pivotal role in this process, culminating in the increase of reactive O2 species. Hexokinase (HK) is a key enzyme on glucose metabolism and is coupled to the brain mitochondrial redox modulation by recycling ADP for oxidative phosphorylation (OXPHOS). GABA shunt is an alternative pathway to GABA metabolism that increases succinate levels, a Krebs cycle intermediate. Although glucose and GABA metabolisms are intrinsically connected, their interplay coordinating mitochondrial function is poorly understood. Here, we hypothesize that the HK and the GABA shunt interact to control mitochondrial metabolism differently in the cortex and the hypothalamus. The GABA shunt stimulated mitochondrial O2 consumption and H2O2 production higher in hypothalamic synaptosomes (HSy) than cortical synaptosomes (CSy). The GABA shunt increased the HK coupled to OXPHOS activity in both population of synaptosomes, but the rate of activation was higher in HSy than CSy. Significantly, malonate and vigabatrin blocked the effects of the GABA shunt in the HK activity coupled to OXPHOS. It indicates that the glucose phosphorylation is linked to GABA and Krebs cycle reactions. Together, these data shed light on the HK and SDH role on the metabolism of each region fed by GABA turnover, which depends on the neurons' metabolic route.

Keywords:  Bioenergetics; Brain; GABA shunt; Hexokinase; Mitochondria; Synaptosome

DOI:  https://doi.org/10.1007/s11064-021-03463-2
Cell Metab. 2021 Oct 05. pii: S1550-4131(21)00425-3. [Epub ahead of print]33(10): 1905-1907

The Gro3p factor: Restoring NAD+/NADH homeostasis to ameliorate mitochondrial disease.

Divakar S Mithal, Navdeep S Chandel.

Leigh syndrome, a mitochondrial disease, can be modeled in mice with a deficiency in mitochondrial complex I that results in a decreased NAD+/NADH ratio. In this issue of Cell Metabolism, Liu et al. (2021) identify glycerol-3-phosphate (Gro3P) biosynthesis as a method for regenerating cytosolic NAD+ to ameliorate pathology in this mitochondrial disease model.

DOI: https://doi.org/10.1016/j.cmet.2021.09.006
Cell Rep. 2021 Oct 05. pii: S2211-1247(21)01221-3. [Epub ahead of print]37(1): 109767

PHDs/CPT1B/VDAC1 axis regulates long-chain fatty acid oxidation in cardiomyocytes.

Aude Angelini, Pradip K Saha, Antrix Jain, Sung Yun Jung, Randall L Mynatt, Xinchun Pi, Liang Xie.

  Cardiac metabolism is a high-oxygen-consuming process, showing a preference for long-chain fatty acid (LCFA) as the fuel source under physiological conditions. However, a metabolic switch (favoring glucose instead of LCFA) is commonly reported in ischemic or late-stage failing hearts. The mechanism regulating this metabolic switch remains poorly understood. Here, we report that loss of PHD2/3, the cellular oxygen sensors, blocks LCFA mitochondria uptake and β-oxidation in cardiomyocytes. In high-fat-fed mice, PHD2/3 deficiency improves glucose metabolism but exacerbates the cardiac defects. Mechanistically, we find that PHD2/3 bind to CPT1B, a key enzyme of mitochondrial LCFA uptake, promoting CPT1B-P295 hydroxylation. Further, we show that CPT1B-P295 hydroxylation is indispensable for its interaction with VDAC1 and LCFA β-oxidation. Finally, we demonstrate that a CPT1B-P295A mutant constitutively binds to VDAC1 and rescues LCFA metabolism in PHD2/3-deficient cardiomyocytes. Together, our data identify an oxygen-sensitive regulatory axis involved in cardiac metabolism.

Keywords:  cardiac metabolism switch; cardiomyocyte; carnitine O-palmitoyltransferase 1b; heart failure; hypoxia; long-chain fatty acid; myocardial infarction; prolyl hydroxylase domain protein; voltage-dependent anion channel

DOI:  https://doi.org/10.1016/j.celrep.2021.109767
Cell Mol Life Sci. 2021 Oct 09.

Increased glucosylceramide production leads to decreased cell energy metabolism and lowered tumor marker expression in non-cancerous liver cells.

Marthe-Susanna Wegner, Nina Schömel, Ellen M Olzomer, Sandra Trautmann, Catherine Olesch, Frances L Byrne, Bernhard Brüne, Robert Gurke, Nerea Ferreirós, Andreas Weigert, Gerd Geisslinger, Kyle L Hoehn.

  Hepatocellular carcinoma (HCC) is one of the most difficult cancer types to treat. Liver cancer is often diagnosed at late stages and therapeutic treatment is frequently accompanied by development of multidrug resistance. This leads to poor outcomes for cancer patients. Understanding the fundamental molecular mechanisms leading to liver cancer development is crucial for developing new therapeutic approaches, which are more efficient in treating cancer. Mice with a liver specific UDP-glucose ceramide glucosyltransferase (UGCG) knockout (KO) show delayed diethylnitrosamine (DEN)-induced liver tumor growth. Accordingly, the rationale for our study was to determine whether UGCG overexpression is sufficient to drive cancer phenotypes in liver cells. We investigated the effect of UGCG overexpression (OE) on normal murine liver (NMuLi) cells. Increased UGCG expression results in decreased mitochondrial respiration and glycolysis, which is reversible by treatment with EtDO-P4, an UGCG inhibitor. Furthermore, tumor markers such as FGF21 and EPCAM are lowered following UGCG OE, which could be related to glucosylceramide (GlcCer) and lactosylceramide (LacCer) accumulation in glycosphingolipid-enriched microdomains (GEMs) and subsequently altered signaling protein phosphorylation. These cellular processes lead to decreased proliferation in NMuLi/UGCG OE cells. Our data show that increased UGCG expression itself does not induce pro-cancerous processes in normal liver cells, which indicates that increased GlcCer expression leads to different outcomes in different cancer types.

Keywords:  GEMs; Glycolysis; HCC marker; Mitochondrial ROS; Oxidative phosphorylation

DOI:  https://doi.org/10.1007/s00018-021-03958-9
Immunometabolism. 2021 Sep 24. 3(4): e210030

IL-10-Mediated Refueling of Exhausted T Cell Mitochondria Boosts Anti-Tumour Immunity.

Dylan Ryan, Christian Frezza.

  Immunotherapy has underscored a revolution in cancer treatment. Yet, many patients fail to respond due to T cell exhaustion. Here, an intervention that restores mitochondrial function reversed the exhausted T cell phenotype to promote cytotoxicity and durable anti-tumour responses in vivo.

Keywords:  IL-10; T cell; exhaustion; immunotherapy; metabolism; mitochondria

DOI:  https://doi.org/10.20900/immunometab20210030
BMC Mol Cell Biol. 2021 Oct 07. 22(1): 52

Intercellular transfer of mitochondrial DNA carrying metastasis-enhancing pathogenic mutations from high- to low-metastatic tumor cells and stromal cells via extracellular vesicles.

Keizo Takenaga, Nobuko Koshikawa, Hiroki Nagase.

  BACKGROUND: Mitochondrial DNA (mtDNA) carrying certain pathogenic mutations or single nucleotide variants (SNVs) enhances the invasion and metastasis of tumor cells, and some of these mutations are homoplasmic in tumor cells and even in tumor tissues. On the other hand, intercellular transfer of mitochondria and cellular components via extracellular vesicles (EVs) and tunneling nanotubes (TNTs) has recently attracted intense attention in terms of cell-to-cell communication in the tumor microenvironment. It remains unclear whether metastasis-enhancing pathogenic mutant mtDNA in tumor cells is intercellularly transferred between tumor cells and stromal cells. In this study, we investigated whether mtDNA with the NADH dehydrogenase subunit 6 (ND6) G13997A pathogenic mutation in highly metastatic cells can be horizontally transferred to low-metastatic cells and stromal cells in the tumor microenvironment.RESULTS: When MitoTracker Deep Red-labeled high-metastatic Lewis lung carcinoma A11 cells carrying the ND6 G13997A mtDNA mutation were cocultured with CellLight mitochondria-GFP-labeled low-metastatic P29 cells harboring wild-type mtDNA, bidirectional transfer of red- and green-colored vesicles, probably mitochondria-related EVs, was observed in a time-dependent manner. Similarly, intercellular transfer of mitochondria-related EVs occurred between A11 cells and α-smooth muscle actin (α-SMA)-positive cancer-associated fibroblasts (CAFs, WA-mFib), macrophages (RAW264.7) and cytotoxic T cells (CTLL-2). Intercellular transfer was suppressed by inhibitors of EV release. The large and small EV fractions (L-EV and S-EV, respectively) prepared from the conditioned medium by differential ultracentrifugation both were found to contain mtDNA, although only S-EVs were efficiently incorporated into the cells. Several subpopulations had evidence of LC3-II and contained degenerated mitochondrial components in the S-EV fraction, signaling to the existence of autophagy-related S-EVs. Interestingly, the S-EV fraction contained a MitoTracker-positive subpopulation, which was inhibited by the respiration inhibitor antimycin A, indicating the presence of mitochondria with membrane potential. It was also demonstrated that mtDNA was transferred into mtDNA-less ρ0 cells after coculture with the S-EV fraction. In syngeneic mouse subcutaneous tumors formed by a mixture of A11 and P29 cells, the mitochondria-related EVs released from A11 cells reached distantly positioned P29 cells and CAFs.
CONCLUSIONS: These results suggest that metastasis-enhancing pathogenic mtDNA derived from metastatic tumor cells is transferred to low-metastatic tumor cells and stromal cells via S-EVs in vitro and in the tumor microenvironment, inferring a novel mechanism of enhancement of metastatic potential during tumor progression.

Keywords:  Extracellular vesicles; Intercellular transfer; Lung cancer; Metastasis; Mitochondria; Mitochondrial DNA; Mutation; Tumor microenvironment

DOI:  https://doi.org/10.1186/s12860-021-00391-5
J Clin Invest. 2021 Sep 01. pii: e139933. [Epub ahead of print]131(17):

Petasin potently inhibits mitochondrial complex I-based metabolism that supports tumor growth and metastasis.

Kazuki Heishima, Nobuhiko Sugito, Tomoyoshi Soga, Masashi Nishikawa, Yuko Ito, Ryo Honda, Yuki Kuranaga, Hiroki Sakai, Ryo Ito, Takayuki Nakagawa, Hiroshi Ueda, Yukihiro Akao.

  Mitochondrial electron transport chain complex I (ETCC1) is the essential core of cancer metabolism, yet potent ETCC1 inhibitors capable of safely suppressing tumor growth and metastasis in vivo are limited. From a plant extract screening, we identified petasin (PT) as a highly potent ETCC1 inhibitor with a chemical structure distinct from conventional inhibitors. PT had at least 1700 times higher activity than that of metformin or phenformin and induced cytotoxicity against a broad spectrum of tumor types. PT administration also induced prominent growth inhibition in multiple syngeneic and xenograft mouse models in vivo. Despite its higher potency, it showed no apparent toxicity toward nontumor cells and normal organs. Also, treatment with PT attenuated cellular motility and focal adhesion in vitro as well as lung metastasis in vivo. Metabolome and proteome analyses revealed that PT severely depleted the level of aspartate, disrupted tumor-associated metabolism of nucleotide synthesis and glycosylation, and downregulated major oncoproteins associated with proliferation and metastasis. These findings indicate the promising potential of PT as a potent ETCC1 inhibitor to target the metabolic vulnerability of tumor cells.

Keywords:  Amino acid metabolism; Cancer; Metabolism; Mitochondria; Oncology

DOI:  https://doi.org/10.1172/JCI139933
Mitochondrion. 2021 Oct 01. pii: S1567-7249(21)00140-9. [Epub ahead of print]

Quantification of mtDNA content in cultured cells by direct droplet digital PCR.

Natalya Kozhukhar, Anthony Fant, Mikhail F Alexeyev.

  Although alterations in cellular mitochondrial DNA (mtDNA) content have been linked to various pathological conditions, the mechanisms that govern mtDNA copy number (mtCN) control remain poorly understood. Moreover, techniques for mtDNA quantification do not allow for direct comparisons of absolute mtCNs between labs. Here we report the development of a direct droplet digital PCR technique for the determination of mtCNs in whole-cell lysates. Using this technique, we demonstrate that cellular mtDNA content can fluctuate in culture by as much as 50% and provide evidence for both cell proliferation-coupled and uncoupled mtDNA replication.

Keywords:  Mitochondrial DNA; X-ray; cell cycle; cell proliferation; mtDNA copy number; mtDNA replication

DOI:  https://doi.org/10.1016/j.mito.2021.09.014
Antioxid Redox Signal. 2021 Oct 04.

The multifaceted ATPase inhibitory factor 1 (IF1) in energy metabolism reprogramming and mitochondrial dysfunction: a new player in age-associated disorders?

Emilia Gore, Thibaut Duparc, Annelise Genoux, Bertrand Perret, Souad Najib, Laurent O Martinez.

Significance The mitochondrial oxidative phosphorylation (OXPHOS) system, comprising the electron transport chain and ATP synthase, generates membrane potential, drives ATP synthesis, governs energy metabolism, and maintains redox balance. OXPHOS dysfunction is associated with a plethora of diseases ranging from rare inherited disorders to common conditions including diabetes, cancer, neurodegenerative diseases, as well as aging. There has been great interest in studying regulators of OXPHOS. Among these, ATPase inhibitory factor 1 (IF1) is an endogenous inhibitor of ATP synthase that has long been thought to avoid the consumption of cellular ATP when ATP synthase acts as an ATP hydrolysis enzyme. Recent Advances Recent data indicates that IF1 inhibits ATP synthesis and is involved in a multitude of mitochondrial-related functions, such as mitochondrial quality control, energy metabolism, redox balance, and cell fate. IF1 also inhibits the ATPase activity of cell-surface ATP synthase, and it is used as a cardiovascular disease biomarker. Critical Issues Although recent data have led to a paradigm shift regarding IF1 functions, these have been poorly studied in entire organisms and in different organs. The understanding of the cellular biology of IF1 is, therefore, still limited. The aim of this review was to provide an overview of the current understanding of the role of IF1 in mitochondrial functions, health, and diseases. Future Directions Further investigations of IF1 functions at the cell, organ, and whole-organism levels and in different pathophysiological conditions will help decipher the controversies surrounding its involvement in mitochondrial function and could unveil therapeutic strategies in human pathology.

DOI: https://doi.org/10.1089/ars.2021.0137
Biochem Biophys Rep. 2021 Dec;28 101142

TFAM knockdown-triggered mtDNA-nucleoid aggregation and a decrease in mtDNA copy number induce the reorganization of nucleoid populations and mitochondria-associated ER-membrane contacts.

Koit Aasumets, Yuliya Basikhina, Jaakko L Pohjoismäki, Steffi Goffart, Joachim Gerhold.

  The correct organization of mitochondrial DNA (mtDNA) in nucleoids and the contacts of mitochondria with the ER play an important role in maintaining the mitochondrial genome distribution within the cell. Mitochondria-associated ER membranes (MAMs) consist of interacting proteins and lipids located in the outer mitochondrial membrane and ER membrane, forming a platform for the mitochondrial inner membrane-associated genome replication factory as well as connecting the nucleoids with the mitochondrial division machinery. We show here that knockdown of a core component of mitochondrial nucleoids, TFAM, causes changes in the mitochondrial nucleoid populations, which subsequently impact ER-mitochondria membrane contacts. Knockdown of TFAM causes a significant decrease in the copy number of mtDNA as well as aggregation of mtDNA nucleoids. At the same time, it causes significant upregulation of the replicative TWNK helicase in the membrane-associated nucleoid fraction. This is accompanied by a transient elevation of MAM proteins, indicating a rearrangement of the linkage between ER and mitochondria triggered by changes in mitochondrial nucleoids. Reciprocal knockdown of the mitochondrial replicative helicase TWNK causes a decrease in mtDNA copy number and modifies mtDNA membrane association, however, it does not cause nucleoid aggregation and considerable alterations of MAM proteins in the membrane-associated fraction. Our explanation is that the aggregation of mitochondrial nucleoids resulting from TFAM knockdown triggers a compensatory mechanism involving the reorganization of both mitochondrial nucleoids and MAM. These results could provide an important insight into pathological conditions associated with impaired nucleoid organization or defects of mtDNA distribution.

Keywords:  Mitochondrial DNA; Mitochondrial transcription factor A (TFAM); Nucleoids; Organellar membranes; TWNK helicase

DOI:  https://doi.org/10.1016/j.bbrep.2021.101142
Endocrinology. 2021 Oct 06. pii: bqab212. [Epub ahead of print]

TXNIP links anticipatory unfolded protein response to estrogen reprogramming glucose metabolism in breast cancer cells.

Yuanzhong Wang, Shiuan Chen.

  Estrogen and estrogen receptor (ER) play a fundamental role in breast cancer. To adapt the rapid proliferation of ER+ breast cancer cells, estrogen increases glucose uptake and reprograms glucose metabolism. Meanwhile, estrogen/ER activates the anticipatory unfolded protein response (UPR) preparing cancer cells for the increased protein production required for subsequent cell proliferation. Here, we report that thioredoxin-interacting protein (TXNIP) is an important regulator of glucose metabolism in ER+ breast cancer cells, and estrogen/ER increases glucose uptake and reprograms glucose metabolism via activating anticipatory unfolded protein response (UPR) and subsequently repressing TXNIP expression. By using two widely used ER+ breast cancer cell lines MCF7 and T47D, we showed that MCF7 cells express high TXNIP levels and exhibit mitochondrial oxidative phosphorylation (OXPHOS) phenotype, while T47D cells express low TXNIP levels and display aerobic glycolysis (Warburg effect) phenotype. Knockdown of TXNIP promoted glucose uptake and Warburg effect, while forced overexpression of TXNIP inhibited glucose uptake and Warburg effect. We further showed that estrogen represses TXNIP expression and activates UPR sensor inositol-requiring enzyme 1 (IRE1) via ER in the breast cancer cells, and IRE1 activity is required for estrogen suppression of TXNIP expression and estrogen-induced cell proliferation. Together, our study suggests that TXNIP is involved in estrogen-induced glucose uptake and metabolic reprogramming in ER+ breast cancer cells, and links anticipatory UPR to estrogen reprogramming glucose metabolism.

Keywords:  TXNIP; breast cancer; estrogen; glucose metabolism; unfolded protein response

DOI:  https://doi.org/10.1210/endocr/bqab212
Angew Chem Int Ed Engl. 2021 Oct 06.

Structural Basis for Inhibition of ROS-Producing Respiratory Complex I by NADH-OH.

Thorsten Friedrich, Marta Vranas, Daniel Wohlwend, Danye Qiu, Stefan Gerhardt, Christian Trncik, Mehrosh Pervaiz, Kevin Ritter, Stefan Steimle, Antonio Randazzo, Oliver Einsle, Stefan Günther, Henning J Jessen, Alexander Kotlyar.

  NADH:ubiquinone oxidoreductase, respiratory complex I, plays a central role in cellular energy metabolism. As a major source of reactive oxygen species (ROS) it affects ageing and mitochondrial dysfunction. The novel inhibitor NADH-OH specifically blocks NADH oxidation and ROS production by complex I in nM concentrations. Attempts to elucidate its structure by NMR failed. Here, we report by X-ray crystallographic analysis the structure of NADH-OH bound in the active site of a soluble fragment of complex I at 2.0 Å resolution. We have identified key amino acid residues specific and essential for binding NADH-OH. Furthermore, the structure sheds light on the specificity of NADH-OH towards the unique Rossmann-fold of complex I and indicates a regulatory role for mitochondrial ROS generation. In addition, NADH-OH displays a lead-structure for synthesis of a novel class of ROS suppressors.

Keywords:  Electron transport; NADH:ubiquinone oxidoreductase; Structural Biology; inhibitors; reactive oxygen species

DOI:  https://doi.org/10.1002/anie.202112165
Biosci Biotechnol Biochem. 2021 Oct 08. pii: zbab176. [Epub ahead of print]

Natural tetramic acids elicit multiple inhibitory actions against mitochondrial machineries presiding over oxidative phosphorylation.

Yufu Unten, Masatoshi Murai, Katsuyuki Sakai, Yukihiro Asami, Takenori Yamamoto, Takahiro Masuya, Hideto Miyoshi.

  The mitochondrial machineries presiding over ATP synthesis via oxidative phosphorylation are promising druggable targets. Fusaramin, a 3-acyl tetramic acid isolated from Fusarium concentricum FKI-7550, is an inhibitor of oxidative phosphorylation in Saccharomyces cerevisiae mitochondria, although its target has yet to be identified. Fusaramin significantly interfered with [3H]ADP uptake by yeast mitochondria at the concentration range inhibiting oxidative phosphorylation. A photoreactive fusaramin derivative (pFS-5) specifically labeled voltage-dependent anion channel 1 (VDAC1), which facilitates trafficking of ADP/ATP across the outer mitochondrial membrane. These results strongly suggest that the inhibition of oxidative phosphorylation by fusaramin is predominantly attributable to the impairment of VDAC1 functions. Fusaramin also inhibited FoF1-ATP synthase and ubiquinol-cytochrome c oxidoreductase (complex III) at concentrations higher than those required for the VDAC inhibition. Considering that other tetramic acid derivatives are reported to inhibit FoF1-ATP synthase and complex III, natural tetramic acids were found to elicit multiple inhibitory actions against mitochondrial machineries.

Keywords:  VDAC1; fusaramin; mitochondria; oxidative phosphorylation; tetramic acid

DOI:  https://doi.org/10.1093/bbb/zbab176
J Biol Chem. 2021 Oct 05. pii: S0021-9258(21)01082-6. [Epub ahead of print] 101279

Labeling and Measuring Stressed Mitochondria Using a PINK1-Based Ratiometric Fluorescent Sensor.

Rie Uesugi, Shunsuke Ishii, Akira Matsuura, Eisuke Itakura.

  Mitochondria are essential organelles that carry out a number of pivotal metabolic processes and maintain cellular homeostasis. Mitochondrial dysfunction caused by various stresses is associated with many diseases such as type 2 diabetes, obesity, cancer, heart failure, neurodegenerative disorders, and aging. Therefore, it is important to understand the stimuli that induce mitochondrial stress. However, broad analysis of mitochondrial stress has not been carried out to date. Here, we present a set of fluorescent tools, called mito-Pain (mitochondrial PINK1 accumulation index), which enables the labeling of stressed mitochondria. Mito-Pain utilizes PINK1 stabilization on mitochondria and quantifies mitochondrial stress levels by comparison with PINK1-GFP, which is stabilized under mitochondrial stress, and RFP-Omp25, which is constitutively localized on mitochondria. To identify compounds that induce mitochondrial stress, we screened a library of 3374 compounds using mito-Pain and identified 57 compounds as mitochondrial stress inducers. Furthermore, we classified each compound into several categories based on mitochondrial response: depolarization, mitochondrial morphology, or Parkin recruitment. Parkin recruitment to mitochondria was often associated with mitochondrial depolarization and aggregation, suggesting that Parkin is recruited to heavily damaged mitochondria. In addition, many of the compounds led to various mitochondrial morphological changes, including fragmentation, aggregation, elongation, and swelling, with or without Parkin recruitment or mitochondrial depolarization. We also found that several compounds induced an ectopic response of Parkin, leading to the formation of cytosolic puncta dependent on PINK1. Thus, mito-Pain enables the detection of stressed mitochondria under a wide variety of conditions and provide insights into mitochondrial quality control systems.

Keywords:  PTEN‐induced putative kinase 1 (PINK1); Parkin; mitochondria; mitochondrial membrane potential; mitochondrial sensor; mitochondrial stress

DOI:  https://doi.org/10.1016/j.jbc.2021.101279
Blood. 2021 Oct 05. pii: blood.2021011563. [Epub ahead of print]

Proton export alkalinizes intracellular pH and reprograms carbon metabolism to drive hematopoietic progenitor growth.

Cheuk Him Man, Francois Emile Mercier, Nian Liu, Wentao Dong, Gregory Stephanopoulos, Li Jiang, Yookyung Jung, Charles Lin, Anskar Yh Leung, David T Scadden.

Proton export is often considered a detoxifying process in animal cells with monocarboxylate symporters co-exporting excessive lactate and protons during glycolysis or the Warburg effect. Here we report a novel mechanism by which lactate/H+ export is sufficient to induce cell growth. Increased lactate/proton export induces intracellular alkalization that selectively activates catalysis by key metabolic gatekeeper enzymes, HK1/PKM2/G6PDH, thereby enhancing glycolytic and pentose phosphate pathway carbon flux. The result is increased nucleotide levels, NADPH/NADP+ ratio and cell proliferation. Simply increasing the lactate/proton symporter, MCT4, or sodium-proton antiporter, NHE1 was sufficient to increase intracellular-pH (pHi) and give normal hematopoietic cells a significant competitive growth advantage in vivo. This process does not require additional cytokine triggers and is exploited in malignancy where leukemogenic mutations epigenetically increase MCT4. Inhibiting MCT4 decreased intracellular pH, carbon flux and eliminated acute myeloid leukemia-initiating-cells without cytotoxic chemotherapy. Intracellular alkalization is a primitive mechanism by which proton partitioning can directly reprogram carbon metabolism for cell growth.

DOI: https://doi.org/10.1182/blood.2021011563
Front Cell Dev Biol. 2021 ;9 725474

Mitochondrial Impairment by MitoBloCK-6 Inhibits Liver Cancer Cell Proliferation.

Yaschar Kabiri, Anna Fuhrmann, Anna Becker, Luisa Jedermann, Carola Eberhagen, Ann-Christine König, Tiago Barros Silva, Fernanda Borges, Stefanie M Hauck, Bernhard Michalke, Percy Knolle, Hans Zischka.

  Augmenter of liver regeneration (ALR) is a critical multi-isoform protein with its longer isoform, located in the mitochondrial intermembrane space, being part of the mitochondrial disulfide relay system (DRS). Upregulation of ALR was observed in multiple forms of cancer, among them hepatocellular carcinoma (HCC). To shed light into ALR function in HCC, we used MitoBloCK-6 to pharmacologically inhibit ALR, resulting in profound mitochondrial impairment and cancer cell proliferation deficits. These effects were mostly reversed by supplementation with bioavailable hemin b, linking ALR function to mitochondrial iron homeostasis. Since many tumor cells are known for their increased iron demand and since increased iron levels in cancer are associated with poor clinical outcome, these results help to further advance the intricate relation between iron and mitochondrial homeostasis in liver cancer.

Keywords:  HCC; disulfide relay system; heme; iron; mitochondria

DOI:  https://doi.org/10.3389/fcell.2021.725474
Biosci Rep. 2021 Oct 05. pii: BSR20211320. [Epub ahead of print]

Beyond Base Excision Repair: An evolving picture of mitochondrial DNA repair.

Kathrin Allkanjari, Robert A Baldock.

  Mitochondria are highly specialised organelles required for cellular processes including ATP-production through cellular respiration and controlling apoptosis. Mitochondria contain their own DNA genome which encodes both protein and RNA required for cellular respiration. Each cell may contain hundreds to thousands of copies of the mitochondrial genome, which is essential for normal cellular function - deviation of mitochondrial DNA (mtDNA) copy number is associated with cellular aging and disease. Furthermore, mtDNA lesions can arise from both endogenous or exogenous sources and must either be tolerated or corrected to preserve mitochondrial function. Importantly, replication of damaged mtDNA can lead to stalling and introduction of mutations or genetic loss, mitochondria have adapted mechanisms to repair damaged DNA. These mechanisms rely on nuclear encoded DNA repair proteins that are translocated into the mitochondria. Despite the presence of many known nuclear DNA repair proteins being found in the mitochondrial proteome, it remains to be established which DNA repair mechanisms are functional in mammalian mitochondria. Here, we summarise the existing and emerging research, alongside examining proteomic evidence, demonstrating that mtDNA damage can be repaired using Base Excision Repair (BER), Homologous Recombination (HR) and Microhomology-mediated End Joining (MMEJ). Critically, these repair mechanisms do not operate in isolation and evidence for interplay between pathways and repair associated with replication is discussed. Importantly, characterising non-canonical functions of key proteins and understanding the bespoke pathways used to tolerate, repair or bypass DNA damage will be fundamental in fully understanding the causes of mitochondrial genome mutations and mitochondrial dysfunction.

Keywords:  DNA replication and recombination; DNA synthesis and repair; genome integrity; mtDNA

DOI:  https://doi.org/10.1042/BSR20211320
Oncogene. 2021 Oct 02.

USP29 coordinates MYC and HIF1α stabilization to promote tumor metabolism and progression.

Rongfu Tu, Wenqian Kang, Mengjie Yang, Liyuan Wang, Qing Bao, Zhi Chen, Yang Dong, Jingchao Wang, Jue Jiang, Hudan Liu, Guoliang Qing.

Tumor cells must rewire cellular metabolism to satisfy the demands of unbridled growth and proliferation. How these metabolic processes are integrated to fuel cancer cell growth remains largely unknown. Deciphering the regulatory mechanisms is vital to develop targeted strategies for tumor-selective therapies. We herein performed an unbiased and functional siRNA screen against 96 deubiquitinases, which play indispensable roles in cancer and are emerging as therapeutic targets, and identified USP29 as a top candidate essential for metabolic reprogramming that support biosynthesis and survival in tumor cells. Integrated metabolic flux analysis and molecular investigation reveal that USP29 directly deubiquitinates and stabilizes MYC and HIF1α, two master regulators of metabolic reprogramming, enabling adaptive response of tumor cells in both normoxia and hypoxia. Systemic knockout of Usp29 depleted MYC and HIF1α in MYC-driven neuroblastoma and B cell lymphoma, inhibited critical metabolic targets and significantly prolonged survival of tumor-bearing mice. Strikingly, mice homozygous null for the Usp29 gene are viable, fertile, and display no gross phenotypic abnormalities. Altogether, these results demonstrate that USP29 selectively coordinates MYC and HIF1α to integrate metabolic processes critical for cancer cell growth, and therapeutic targeting of USP29, a potentially targetable enzyme, could create a unique vulnerability given deregulation of MYC and HIF1α frequently occurs in human cancers.

DOI: https://doi.org/10.1038/s41388-021-02031-w
Biomaterials. 2021 Sep 28. pii: S0142-9612(21)00525-1. [Epub ahead of print]278 121168

Mitochondrial-targeting nanoprodrugs to mutually reinforce metabolic inhibition and autophagy for combating resistant cancer.

Min Sun, Congyu Wang, Mingchen Lv, Zhen Fan, Jianzhong Du.

  Abnormal energy metabolism is one of the hallmarks of cancer and closely linked to therapy resistance. However, existing metabolic inhibitors suffer from inefficient cell enrichment and therapeutic effects. In this work, we developed an effective strategy to mutually reinforce the metabolic inhibition and autophagy for enhanced tumor killing efficacy and combating resistant cancer. First, mitochondrial homing moiety triphenylphosphonium and metabolic inhibitor lonidamine were grafted onto polylysine. After self-assembly of this functionalized polylysine, ferrocene and glucose oxidase were immobilized to afford additional chemotherapy functions, and the final product was named as FG/T-Nanoprodrug. Effective mitochondrial targeting and metabolic inhibition were observed in resistant cancer cells. In addition, owing to the inhibited metabolism, less glucose is consumed to allow FG/T-Nanoprodrug to produce excess reactive oxygen species (ROS) by glucose oxidase and ferrocene. The enhanced chemodynamic therapy increases the mitochondrial permeability to promote the release of cytochrome c from mitochondria, ultimately induces high levels of autophagy. The FG/T-Nanoprodrug demonstrated superior mutually reinforcing of metabolic inhibition (up to 3.7-fold compared to free lonidamine) and autophagy (up to 125.3-fold compared to free lonidamine) to effectively kill resistant cancer cell both in vitro and in vivo. Overall, this strategy could pave a new way to efficient treatment of resistant cancer and other metabolically abnormal diseases.

Keywords:  Autophagy; Chemodynamic therapy; Lonidamine; Nanoprodrug; Polylysine

DOI:  https://doi.org/10.1016/j.biomaterials.2021.121168
Lab Invest. 2021 Oct 04.

ATP synthase inhibitory factor subunit 1 regulates islet β-cell function via repression of mitochondrial homeostasis.

Kailiang Zhang, Rong Bao, Fengyuan Huang, Kevin Yang, Yishu Ding, Lothar Lauterboeck, Masasuke Yoshida, Qinqiang Long, Qinglin Yang.

Mitochondrial homeostasis is crucial for the function of pancreatic β-cells. ATP synthase inhibitory factor subunit 1 (IF1) is a mitochondrial protein interacting with ATP synthase to inhibit its enzyme activity. IF1 may also play a role in maintaining ATP synthase oligomerization and mitochondrial inner membrane formation. A recent study confirmed IF1 expresses in β-cells. IF1 knockdown in cultured INS-1E β-cells enhances glucose-induced insulin release. However, the role of IF1 in islet β-cells remains little known. The present study investigates islets freshly isolated from mouse lines with global IF1 knockout (IF1-/-) and overexpression (OE). The glucose-stimulated insulin secretion was increased in islets from IF1-/- mice but decreased in islets from IF1 OE mice. Transmitted Electronic Microscopic assessment of isolated islets revealed that the number of matured insulin granules (with dense core) was relatively higher in IF1-/-, but fewer in IF1 OE islets than those of controlled islets. The mitochondrial ultrastructure within β-cells of IF1 overexpressed islets was comparable with those of wild-type mice, whereas those in IF1-/- β-cells showed increased mitochondrial mass. Mitochondrial network analysis in cultured INS-1 β-cells showed a similar pattern with an increased mitochondrial network in IF1 knockdown cells. IF1 overexpressed INS-1 β-cells showed a compromised rate of mitochondrial oxidative phosphorylation with attenuated cellular ATP content. In contrast, INS-1 cells with IF1 knockdown showed markedly increased cellular respiration with improved ATP production. These results support that IF1 is a negative regulator of insulin production and secretion via inhibiting mitochondrial mass and respiration in β-cells. Therefore, inhibiting IF1 to improve β-cell function in patients can be a novel therapeutic strategy to treat diabetes.

DOI: https://doi.org/10.1038/s41374-021-00670-x
Elife. 2021 10 05. pii: e69207. [Epub ahead of print]10

Protective mitochondrial fission induced by stress-responsive protein GJA1-20k.

Daisuke Shimura, Esther Nuebel, Rachel Baum, Steven E Valdez, Shaohua Xiao, Junco S Warren, Joseph A Palatinus, TingTing Hong, Jared Rutter, Robin M Shaw.

  The Connexin43 gap junction gene GJA1 has one coding exon, but its mRNA undergoes internal translation to generate N-terminal truncated isoforms of Connexin43 with the predominant isoform being only 20 kDa in size (GJA1-20k). Endogenous GJA1-20k protein is not membrane bound and has been found to increase in response to ischemic stress, localize to mitochondria, and mimic ischemic preconditioning protection in the heart. However, it is not known how GJA1-20k benefits mitochondria to provide this protection. Here, using human cells and mice, we identify that GJA1-20k polymerizes actin around mitochondria which induces focal constriction sites. Mitochondrial fission events occur within about 45 s of GJA1-20k recruitment of actin. Interestingly, GJA1-20k mediated fission is independent of canonical Dynamin-Related Protein 1 (DRP1). We find that GJA1-20k-induced smaller mitochondria have decreased reactive oxygen species (ROS) generation and, in hearts, provide potent protection against ischemia-reperfusion injury. The results indicate that stress responsive internally translated GJA1-20k stabilizes polymerized actin filaments to stimulate non-canonical mitochondrial fission which limits ischemic-reperfusion induced myocardial infarction.

Keywords:  GJA1-20k; actin dynamics; cell biology; human; ischemia/reperfusion; mitochondria; mitochondria dynamics; mouse; organ protection

DOI:  https://doi.org/10.7554/eLife.69207
EMBO Rep. 2021 Oct 07. e52964

Mild mitochondrial impairment enhances inn3ate immunity and longevity through ATFS-1 and p38 signaling.

Juliane C Campos, Ziyun Wu, Paige D Rudich, Sonja K Soo, Meeta Mistry, Julio Cb Ferreira, T Keith Blackwell, Jeremy M Van Raamsdonk.

  While mitochondrial function is essential for life in all multicellular organisms, a mild impairment of mitochondrial function can extend longevity in model organisms. By understanding the molecular mechanisms involved, these pathways might be targeted to promote healthy aging. In studying two long-lived mitochondrial mutants in C. elegans, we found that disrupting subunits of the mitochondrial electron transport chain results in upregulation of genes involved in innate immunity, which is driven by the mitochondrial unfolded protein response (mitoUPR) but also dependent on the canonical p38-mediated innate immune signaling pathway. Both of these pathways are required for the increased resistance to bacterial pathogens and extended longevity of the long-lived mitochondrial mutants, as is the FOXO transcription factor DAF-16. This work demonstrates that both the p38-mediated innate immune signaling pathway and the mitoUPR act in concert on the same innate immunity genes to promote pathogen resistance and longevity and that input from the mitochondria can extend longevity by signaling through these pathways. This indicates that multiple evolutionarily conserved genetic pathways controlling innate immunity also function to modulate lifespan.

Keywords:   C. elegans ; aging; innate immunity; mitochondria; mitochondrial unfolded protein response

DOI:  https://doi.org/10.15252/embr.202152964
iScience. 2021 Oct 22. 24(10): 103118

Sirt1 promotes tissue regeneration in zebrafish through regulating the mitochondrial unfolded protein response.

Yi-Fan Lin, Jessica Sam, Todd Evans.

  The mitochondrial unfolded protein response (UPRmt) is an organellar stress signaling pathway that functions to detect and restore disruption of mitochondrial proteostasis. The UPRmt is involved in a wide range of physiological and disease conditions, including aging, stem cell maintenance, innate immunity, neurodegeneration, and cancer. Here we report that the UPRmt is integral to zebrafish fin regeneration. Taking advantage of a novel zebrafish UPRmt reporter, we observed that UPRmt activation occurs in regenerating fin tissue shortly after injury. Through chemical and genetic approaches, we discovered that the Sirt1-UPRmt pathway, best known for its role in promoting lifespan extension, is crucial for fin regeneration. The metabolism of NAD+ is an important contributor to Sirt1 activity in this context. We propose that Sirt1 activation induces mitochondrial biogenesis in injured fin tissue, which leads to UPRmt activation and promotes tissue regeneration.

Keywords:  Cell biology; Developmental biology; Molecular biology

DOI:  https://doi.org/10.1016/j.isci.2021.103118
Mol Cancer Ther. 2021 Oct 08. pii: molcanther.MCT-21-0396-A.2021. [Epub ahead of print]

A Benzenesulfonamide-based Mitochondrial Uncoupler Induces Endoplasmic Reticulum Stress and Immunogenic Cell Death in Epithelial Ovarian Cancer.

Fangfang Bi, Ziyan Jiang, Wonmin Park, Tobias Mp Hartwich, Zhiping Ge, Kay Yi Chong, Kevin Yang, Madeline J Morrisson, Dongin Kim, Jaeyeon Kim, Wen Zhang, Liliia M Kril, David S Watt, Chunming Liu, Yang Yang-Hartwich.

Epithelial ovarian cancer (EOC) is a leading cause of death from gynecologic malignancies and requires new therapeutic strategies to improve clinical outcomes. EOCs metastasize in the abdominal cavity through dissemination in the peritoneal fluid and ascites, efficiently adapt to the nutrient-deprived microenvironment, and resist current chemotherapeutic agents. Accumulating evidence suggests that mitochondrial oxidative phosphorylation is critical for the adaptation of EOC cells to this otherwise hostile microenvironment. Although chemical mitochondrial uncouplers can impair mitochondrial functions and thereby target multiple, essential pathways for cancer cell proliferation, traditional mitochondria uncouplers often cause toxicity that precludes their clinical application. In this study, we demonstrated that a mitochondrial uncoupler, specifically 2,5-dichloro-N-(4-nitronaphthalen-1-yl)benzenesulfonamide, hereinafter named Y3, was an antineoplastic agent in ovarian cancer models. Y3 treatment activated AMP-activated protein kinase and resulted in the activation of endoplasmic reticulum stress sensors as well as growth inhibition and apoptosis in ovarian cancer cells in vitro. Y3 was well tolerated in vivo and effectively suppressed tumor progression in three mouse models of EOC, and Y3 also induced immunogenic cell death of cancer cells that involved the release of damage-associated molecular patterns and the activation of antitumor adaptive immune responses. These findings suggest that mitochondrial uncouplers hold promise in developing new anticancer therapies that delay tumor progression and protect ovarian cancer patients against relapse.

DOI: https://doi.org/10.1158/1535-7163.MCT-21-0396
Aging Cell. 2021 Oct 06. e13487

Mitochondrial DNA copy number and heteroplasmy load correlate with skeletal muscle oxidative capacity by P31 MR spectroscopy.

Qu Tian, Ann Zenobia Moore, Richard Oppong, Jun Ding, Marta Zampino, Kenneth W Fishbein, Richard G Spencer, Luigi Ferrucci.

  The association between blood-based estimates of mitochondrial DNA parameters, mitochondrial DNA copy number (mtDNA-CN) and heteroplasmy load, with skeletal muscle bioenergetic capacity was evaluated in 230 participants of the Baltimore Longitudinal Study of Aging (mean age:74.7 years, 53% women). Participants in the study sample had concurrent data on muscle oxidative capacity (τPCr ) assessed by 31 P magnetic resonance spectroscopy, and mitochondrial DNA parameters estimated from whole-genome sequencing data. In multivariable linear regression models, adjusted for age, sex, extent of phosphocreatine (PCr) depletion, autosomal sequencing coverage, white blood cell total, and differential count, as well as platelet count, mtDNA-CN and heteroplasmy load were not significantly associated with τPCr (both p > 0.05). However, in models evaluating whether the association between mtDNA-CN and τPCr varied by heteroplasmy load, there was a significant interaction between mtDNA-CN and heteroplasmy load (p = 0.037). In stratified analysis, higher mtDNA-CN was significantly associated with lower τPCr among participants with high heteroplasmy load (n = 84, β (SE) = -0.236 (0.115), p-value = 0.044), but not in those with low heteroplasmy load (n = 146, β (SE) = 0.046 (0.119), p-value = 0.702). Taken together, mtDNA-CN and heteroplasmy load provide information on muscle bioenergetics. Thus, mitochondrial DNA parameters may be considered proxy measures of mitochondrial function that can be used in large epidemiological studies, especially when comparing subgroups.

Keywords:  aging; mitochondrial DNA; skeletal muscle

DOI:  https://doi.org/10.1111/acel.13487
Redox Biol. 2021 Oct 01. pii: S2213-2317(21)00313-X. [Epub ahead of print]47 102153

Endogenous S-nitrosocysteine proteomic inventories identify a core of proteins in heart metabolic pathways.

Benjamin Lau, Hossein Fazelinia, Ipsita Mohanty, Serena Raimo, Margarita Tenopoulou, Paschalis-Thomas Doulias, Harry Ischiropoulos.

  Protein cysteine residues are essential for protein folding, participate in enzymatic catalysis, and coordinate the binding of metal ions to proteins. Enzymatically catalyzed and redox-dependent post-translational modifications of cysteine residues are also critical for signal transduction and regulation of protein function and localization. S-nitrosylation, the addition of a nitric oxide equivalent to a cysteine residue, is a redox-dependent modification. In this study, we curated and analyzed four different studies that employed various chemoselective platforms coupled to mass spectrometry to precisely identify S-nitrosocysteine residues in mouse heart proteins. Collectively 1974 S-nitrosocysteine residues in 761 proteins were identified and 33.4% were identified in two or more studies. A core of 75 S-nitrosocysteine residues in 44 proteins were identified in all four studies. Bioinformatic analysis of each study indicated a significant enrichment of mitochondrial proteins participating in metabolism. Regulatory proteins in glycolysis, TCA cycle, oxidative phosphorylation and ATP production, long chain fatty acid β-oxidation, and ketone and amino acid metabolism constitute the major functional pathways impacted by protein S-nitrosylation. In the cardiovascular system, nitric oxide signaling regulates vasodilation and cardiac muscle contractility. The meta-analysis of the proteomic data supports the hypothesis that nitric oxide signaling via protein S-nitrosylation is also a regulator of cardiomyocyte metabolism that coordinates fuel utilization to maximize ATP production. As such, protein cysteine S-nitrosylation represents a third functional dimension of nitric oxide signaling in the cardiovascular system to ensure optimal cardiac function.

Keywords:  Cardiovascular system; Nitric oxide; Proteomics; S-nitrosylation

DOI:  https://doi.org/10.1016/j.redox.2021.102153
Nature. 2021 Oct 06.

Structure and assembly of the mammalian mitochondrial supercomplex CIII2CIV.

Irene Vercellino, Leonid A Sazanov.

The enzymes of the mitochondrial electron transport chain are key players of cell metabolism. Despite being active when isolated, in vivo they associate into supercomplexes1, whose precise role is debated. Supercomplexes CIII2CIV1-2 (refs. 2,3), CICIII2 (ref. 4) and CICIII2CIV (respirasome)5-10 exist in mammals, but in contrast to CICIII2 and the respirasome, to date the only known eukaryotic structures of CIII2CIV1-2 come from Saccharomyces cerevisiae11,12 and plants13, which have different organization. Here we present the first, to our knowledge, structures of mammalian (mouse and ovine) CIII2CIV and its assembly intermediates, in different conformations. We describe the assembly of CIII2CIV from the CIII2 precursor to the final CIII2CIV conformation, driven by the insertion of the N terminus of the assembly factor SCAF1 (ref. 14) deep into CIII2, while its C terminus is integrated into CIV. Our structures (which include CICIII2 and the respirasome) also confirm that SCAF1 is exclusively required for the assembly of CIII2CIV and has no role in the assembly of the respirasome. We show that CIII2 is asymmetric due to the presence of only one copy of subunit 9, which straddles both monomers and prevents the attachment of a second copy of SCAF1 to CIII2, explaining the presence of one copy of CIV in CIII2CIV in mammals. Finally, we show that CIII2 and CIV gain catalytic advantage when assembled into the supercomplex and propose a role for CIII2CIV in fine tuning the efficiency of electron transfer in the electron transport chain.

DOI: https://doi.org/10.1038/s41586-021-03927-z
Cell Death Dis. 2021 Oct 02. 12(10): 902

Metformin sensitises hepatocarcinoma cells to methotrexate by targeting dihydrofolate reductase.

Yinghui Wang, Hui Lu, Linchong Sun, Xin Chen, Haoran Wei, Caixia Suo, Junru Feng, Mengqiu Yuan, Shengqi Shen, Weidong Jia, Ying Wang, Huafeng Zhang, Zijun Li, Xiuying Zhong, Ping Gao.

Metformin, the first-line drug for type II diabetes, has recently been considered an anticancer agent. However, the molecular target and underlying mechanism of metformin's anti-cancer effects remain largely unclear. Herein, we report that metformin treatment increases the sensitivity of hepatocarcinoma cells to methotrexate (MTX) by suppressing the expression of the one-carbon metabolism enzyme DHFR. We show that the combination of metformin and MTX blocks nucleotide metabolism and thus effectively inhibits cell cycle progression and tumorigenesis. Mechanistically, metformin not only transcriptionally represses DHFR via E2F4 but also promotes lysosomal degradation of the DHFR protein. Notably, metformin dramatically increases the response of patient-derived hepatocarcinoma organoids to MTX without obvious toxicity to organoids derived from normal liver tissue. Taken together, our findings identify an important role for DHFR in the suppressive effects of metformin on therapeutic resistance, thus revealing a therapeutically targetable potential vulnerability in hepatocarcinoma.

DOI: https://doi.org/10.1038/s41419-021-04199-1
Aging (Albany NY). 2021 Oct 04. 13(undefined):

Green tea catechins EGCG and ECG enhance the fitness and lifespan of Caenorhabditis elegans by complex I inhibition.

Jing Tian, Caroline Geiss, Kim Zarse, Corina T Madreiter-Sokolowski, Michael Ristow.

  Green tea catechins are associated with a delay in aging. We have designed the current study to investigate the impact and to unveil the target of the most abundant green tea catechins, epigallocatechin gallate (EGCG) and epicatechin gallate (ECG). Experiments were performed in Caenorhabditis elegans to analyze cellular metabolism, ROS homeostasis, stress resistance, physical exercise capacity, health- and lifespan, and the underlying signaling pathways. Besides, we examined the impact of EGCG and ECG in isolated murine mitochondria. A concentration of 2.5 μM EGCG and ECG enhanced health- and lifespan as well as stress resistance in C. elegans. Catechins hampered mitochondrial respiration in C. elegans after 6-12 h and the activity of complex I in isolated rodent mitochondria. The impaired mitochondrial respiration was accompanied by a transient drop in ATP production and a temporary increase in ROS levels in C. elegans. After 24 h, mitochondrial respiration and ATP levels got restored, and ROS levels even dropped below control conditions. The lifespan increases induced by EGCG and ECG were dependent on AAK-2/AMPK and SIR-2.1/SIRT1, as well as on PMK-1/p38 MAPK, SKN-1/NRF2, and DAF-16/FOXO. Long-term effects included significantly diminished fat content and enhanced SOD and CAT activities, required for the positive impact of catechins on lifespan. In summary, complex I inhibition by EGCG and ECG induced a transient drop in cellular ATP levels and a temporary ROS burst, resulting in SKN-1 and DAF-16 activation. Through adaptative responses, catechins reduced fat content, enhanced ROS defense, and improved healthspan in the long term.

Keywords:  C. elegans; aging; mitochondria; polyphenols; reactive oxygen species

DOI:  https://doi.org/10.18632/aging.203597
J Cell Biol. 2021 Dec 06. pii: e202006049. [Epub ahead of print]220(12):

PINK1 drives production of mtDNA-containing extracellular vesicles to promote invasiveness.

Nicolas Rabas, Sarah Palmer, Louise Mitchell, Shehab Ismail, Andrea Gohlke, Joel S Riley, Stephen W G Tait, Payam Gammage, Leandro Lemgruber Soares, Iain R Macpherson, Jim C Norman.

The cystine-glutamate antiporter, xCT, supports a glutathione synthesis program enabling cancer cells to cope with metabolically stressful microenvironments. Up-regulated xCT, in combination with glutaminolysis, leads to increased extracellular glutamate, which promotes invasive behavior by activating metabotropic glutamate receptor 3 (mGluR3). Here we show that activation of mGluR3 in breast cancer cells activates Rab27-dependent release of extracellular vesicles (EVs), which can transfer invasive characteristics to "recipient" tumor cells. These EVs contain mitochondrial DNA (mtDNA), which is packaged via a PINK1-dependent mechanism. We highlight mtDNA as a key EV cargo necessary and sufficient for intercellular transfer of invasive behavior by activating Toll-like receptor 9 in recipient cells, and this involves increased endosomal trafficking of pro-invasive receptors. We propose that an EV-mediated mechanism, through which altered cellular metabolism in one cell influences endosomal trafficking in other cells, is key to generation and dissemination of pro-invasive microenvironments during mammary carcinoma progression.

DOI: https://doi.org/10.1083/jcb.202006049
Cell Metab. 2021 Oct 05. pii: S1550-4131(21)00379-X. [Epub ahead of print]33(10): 2090

Dietary Restriction Extends Lifespan through Metabolic Regulation of Innate Immunity.

Ziyun Wu, Meltem Isik, Natalie Moroz, Michael J Steinbaugh, Peng Zhang, T Keith Blackwell.

DOI: https://doi.org/10.1016/j.cmet.2021.08.016
Blood Adv. 2021 Oct 06. pii: bloodadvances.2020004041. [Epub ahead of print]

ALL blasts drive primary mesenchymal stromal cells to increase asparagine availability during Asparaginase treatment.

Martina Chiu, Giuseppe Taurino, Erica Dander, Donatella Bardelli, Alessandra Fallati, Roberta Andreoli, Massimiliano G Bianchi, Cecilia Carubbi, Giulia Pozzi, Laura Galuppo, Prisco Mirandola, Carmelo Rizzari, Saverio Tardito, Andrea Biondi, Giovanna D'Amico, Ovidio Bussolati.

Mechanisms underlying the resistance of Acute Lymphoblastic Leukemia (ALL) blasts to L-asparaginase are still incompletely known. Here we demonstrate that human primary bone marrow mesenchymal stromal cells (MSCs) successfully adapt to L-asparaginase and markedly protect leukemic blasts from the enzyme-dependent cytotoxicity through an amino acid trade-off. ALL blasts synthesize and secrete glutamine, thus increasing extracellular glutamine availability for stromal cells. In turn, MSCs use glutamine, either synthesized through Glutamine Synthetase (GS) or imported, to produce asparagine, which is then extruded to sustain asparagine-auxotroph leukemic cells. GS inhibition prevents mesenchymal cells adaptation to L-asparaginase, lowers glutamine secretion by ALL blasts, and markedly hinders the protection exerted by MSCs on leukemic cells. The pro-survival amino acid exchange is hindered by the inhibition or silencing of the asparagine efflux transporter SNAT5, which is induced in mesenchymal cells by ALL blasts. Consistently, primary MSCs from ALL patients express higher levels of SNAT5 (p < 0.05), secrete more asparagine (p < 0.05), and protect leukemic blasts (p < 0.05) better than MSCs isolated from healthy donors. In conclusion, ALL blasts arrange a pro-leukemic amino acid trade-off with bone marrow mesenchymal cells, which depends on GS and SNAT5 and promotes leukemic cell survival during L-asparaginase treatment.

DOI: https://doi.org/10.1182/bloodadvances.2020004041
Sci Adv. 2021 Oct 08. 7(41): eabi7511

Therapeutic targeting of SLC6A8 creatine transporter suppresses colon cancer progression and modulates human creatine levels.

Isabel Kurth, Norihiro Yamaguchi, Celia Andreu-Agullo, Helen S Tian, Subhasree Sridhar, Shugaku Takeda, Foster C Gonsalves, Jia Min Loo, Afsar Barlas, Katia Manova-Todorova, Robert Busby, Johanna C Bendell, James Strauss, Marwan Fakih, Autumn J McRee, Andrew E Hendifar, Lee S Rosen, Andrea Cercek, Robert Wasserman, Michael Szarek, Scott L Spector, Syed Raza, Masoud F Tavazoie, Sohail F Tavazoie.

[Figure: see text].

DOI: https://doi.org/10.1126/sciadv.abi7511
J Cell Biochem. 2021 Oct 04.

Remodelin, a N-acetyltransferase 10 (NAT10) inhibitor, alters mitochondrial lipid metabolism in cancer cells.

Mahmood Hassan Dalhat, Mohammed Razeeth Shait Mohammed, Abrar Ahmad, Mohammad Imran Khan, Hani Choudhry.

  Remodelin is a small molecule inhibitor of N-acetyltransferase 10 (NAT10), reported to reverse the effect of cancer conditions such as epithelial to mesenchymal transition, hypoxia, and drug resistance. We analysed RNA seq data of siNAT10 and found many metabolic pathways were altered, this made us perform unbiased metabolic analysis. Here we performed untargeted metabolomics in Remodelin treated cancer cells using high-performance liquid chromatography-tandem mass spectrometry. Statistical analysis revealed a total number of 138 of which 52 metabolites were significantly modified in Remodelin treated cells. Among the most significantly altered metabolites, we identified metabolites related with mitochondrial fatty acid elongation (MFAE) and mitochondrial beta-oxidation such as lauroyl-CoA, cholesterol, triglycerides, (S)-3-hydroxyhexadecanoyl-CoA, and NAD+ . Furthermore, assessment showed alteration in expression of Enoyl-CoA hydratase, short chain 1, mitochondrial (ECHS1), and Mitochondrial trans-2-enoyl-CoA reductase (MECR) genes, associated with MFAE pathway. We also found statistically significant decrease in total cholesterol and triglycerides in Remodelin treated cancer cells. Overall, our results showed that Remodelin alters mitochondrial fatty acid metabolism and lipid accumulation in cancer cells. Finally, we validated these results in NAT10 knockdown cancer cells and found that NAT10 reduction results in alteration in gene expression associated with mitochondrial fatty acid metabolism, clearly suggesting the possible role of NAT10 in maintaining mitochondrial fatty acid metabolism.

Keywords:  NAT10; cancer; metabolomics; remodelin

DOI:  https://doi.org/10.1002/jcb.30155
Oncogene. 2021 Oct 07.

CRISPR screens identify cholesterol biosynthesis as a therapeutic target on stemness and drug resistance of colon cancer.

Shanshan Gao, Fraser Soares, Shiyan Wang, Chi Chun Wong, Huarong Chen, Zhenjie Yang, Weixin Liu, Minnie Y Y Go, Musaddeque Ahmed, Yong Zeng, Catherine Adell O'Brien, Joseph J Y Sung, Housheng Hansen He, Jun Yu.

Cancer stem cells (CSCs) are responsible for tumor progression, recurrence, and drug resistance. To identify genetic vulnerabilities of colon cancer, we performed targeted CRISPR dropout screens comprising 657 Drugbank targets and 317 epigenetic regulators on two patient-derived colon CSC-enriched spheroids. Next-generation sequencing of pooled genomic DNAs isolated from surviving cells yielded therapeutic candidates. We unraveled 44 essential genes for colon CSC-enriched spheroids propagation, including key cholesterol biosynthetic genes (HMGCR, FDPS, and GGPS1). Cholesterol biosynthesis was induced in colon cancer tissues, especially CSC-enriched spheroids. The genetic and pharmacological inhibition of HMGCR/FDPS impaired self-renewal capacity and tumorigenic potential of the spheroid models in vitro and in vivo. Mechanistically, HMGCR or FDPS depletion impaired cancer stemness characteristics by activating TGF-β signaling, which in turn downregulated expression of inhibitors of differentiation (ID) proteins, key regulators of cancer stemness. Cholesterol and geranylgeranyl diphosphate (GGPP) rescued the growth inhibitory and signaling effect of HMGCR/FDPS blockade, implying a direct role of these metabolites in modulating stemness. Finally, cholesterol biosynthesis inhibitors and 5-FU demonstrated antitumor synergy in colon CSC-enriched spheroids, tumor organoids, and xenografts. Taken together, our study unravels novel genetic vulnerabilities of colon CSC-enriched spheroids and suggests cholesterol biosynthesis as a potential target in conjunction with traditional chemotherapy for colon cancer treatment.

DOI: https://doi.org/10.1038/s41388-021-01882-7