bims-mibica 2021-12-12 papers

bims-mibica

Biomed News

on Mitochondrial bioenergetics in cancer

Issue of 2021–12–12
thirty-six papers selected by
Kelsey Fisher-Wellman, East Carolina University

Targeting coenzyme Q10 synthesis overcomes bortezomib resistance in multiple myeloma.
Alterations in sphingolipid composition and mitochondrial bioenergetics represent synergistic therapeutic vulnerabilities linked to multidrug resistance in leukemia.
Mitochondria Metabolomics Reveals a Role of β-Nicotinamide Mononucleotide Metabolism in Mitochondrial DNA Replication.
Controlled power: how biology manages succinate-driven energy release.
2-Deoxy-D-glucose couples mitochondrial DNA replication with mitochondrial fitness and promotes the selection of wild-type over mutant mitochondrial DNA.
Metabolic Rewiring Is Essential for AML Cell Survival to Overcome Autophagy Inhibition by Loss of ATG3.
Differential Effects of Oleic and Palmitic Acids on Lipid Droplet-Mitochondria Interaction in the Hepatic Cell Line HepG2.
Reverting the mode of action of the mitochondrial FOF1-ATPase by Legionella pneumophila preserves its replication niche.
Ameliorating effects of lifelong physical activity on healthy aging and mitochondrial function in human white adipose tissue.
Nm23-H1 activator phenylbutenoid dimer exerts cytotoxic effects on metastatic breast cancer cells by inducing mitochondrial dysfunction only under glucose starvation.
NEAT1 is essential for metabolic changes that promote breast cancer growth and metastasis.
MIROs and DRP1 drive mitochondrial-derived vesicle biogenesis and promote quality control.
Mitochondrial Plasticity Promotes Resistance to Sorafenib and Vulnerability to STAT3 Inhibition in Human Hepatocellular Carcinoma.
Molecular species selectivity of lipid transport creates a mitochondrial sink for di-unsaturated phospholipids.
Inhibition of pyrimidine biosynthesis by strobilurin derivatives induces differentiation of acute myeloid leukemia cells.
Monitoring and Modulating mtDNA G-Quadruplex Dynamics Reveal Its Close Relationship to Cell Glycolysis.
Identification and characterisation of novel MPC1 gene variants causing mitochondrial pyruvate carrier deficiency.
Quantitative assays to measure the transport activity of the mitochondrial calcium uniporter in cell lines or Xenopus oocytes.
Somatic mitochondrial DNA mutations in different grades of glioma.
Acute RyR1 Ca2+ leak enhances NADH-linked mitochondrial respiratory capacity.
A novel PDK1 inhibitor, JX06, inhibits glycolysis and induces apoptosis in multiple myeloma cells.
The mitochondrial protein Opa1 promotes adipocyte browning that is dependent on urea cycle metabolites.
Mitochondrial DNA heteroplasmy is modulated during oocyte development propagating mutation transmission.
Surveying the mitochondrial proteome.
Prolyl endopeptidase-like is a (thio)esterase involved in mitochondrial respiratory chain function.
A predominant enhancer co-amplified with the SOX2 oncogene is necessary and sufficient for its expression in squamous cancer.
Data Processing to Probe the Cellular Hydrogen Peroxide Landscape.
AMP-Activated Protein Kinase Contributes to Apoptosis Induced by the Bcl-2 Inhibitor Venetoclax in Acute Myeloid Leukemia.
Acetoacetate protects macrophages from lactic acidosis-induced mitochondrial dysfunction by metabolic reprograming.
Mechanism of PINK1 activation by autophosphorylation and insights into assembly on the TOM complex.
Glucocorticoid receptor regulates PD-L1 and MHC-I in pancreatic cancer cells to promote immune evasion and immunotherapy resistance.
Coenzyme A fuels T cell anti-tumor immunity.
Proteomic Analysis Identifies NDUFS1 and ATP5O as Novel Markers for Survival Outcome in Prostate Cancer.
Spatial-proteomics reveals phospho-signaling dynamics at subcellular resolution.
Free fatty-acid transport via CD36 drives β-oxidation-mediated hematopoietic stem cell response to infection.
Combined Inhibition of Bcl2 and Bcr-Abl1 Exercises Anti-Leukemia Activity but Does Not Eradicate the Primitive Leukemic Cells.

Mol Omics. 2021 Aug 26.

Targeting coenzyme Q10 synthesis overcomes bortezomib resistance in multiple myeloma.

Esther A Zaal, Harm-Jan de Grooth, Inge Oudaert, Pieter Langerhorst, Sophie Levantovsky, Gijs J J van Slobbe, Jeroen W A Jansen, Eline Menu, Wei Wu, Celia R Berkers.

During the development of drug resistance, multiple myeloma (MM) cells undergo changes to their metabolism. However, how these metabolic changes can be exploited to improve treatment efficacy is not known. Here we demonstrate that targeting coenzyme Q10 (CoQ) biosynthesis through the mevalonate pathway works in synergy with the proteasome inhibitor bortezomib (BTZ) in MM. We show that gene expression signatures relating to the mitochondrial tricarboxylic acid (TCA) cycle and electron transport chain (ETC) predispose to clinical BTZ resistance and poor prognosis in MM patients. Mechanistically, BTZ-resistant cells show increased activity of glutamine-driven TCA cycle and oxidative phosphorylation, together with an increased vulnerability towards ETC inhibition. Moreover, BTZ resistance is accompanied by high levels of the mitochondrial electron carrier CoQ, while the mevalonate pathway inhibitor simvastatin increases cell death and decreases CoQ levels, specifically in BTZ-resistant cells. Both in vitro and in vivo, simvastatin enhances the effect of bortezomib treatment. Our study links CoQ synthesis to drug resistance in MM and provides a novel avenue for improving BTZ responses through statin-induced inhibition of mitochondrial metabolism.

DOI: https://doi.org/10.1039/d1mo00106j
FASEB J. 2022 Jan;36(1): e22094

Alterations in sphingolipid composition and mitochondrial bioenergetics represent synergistic therapeutic vulnerabilities linked to multidrug resistance in leukemia.

Kelsey H Fisher-Wellman, James T Hagen, Miki Kassai, Li-Pin Kao, Margaret A M Nelson, Kelsey L McLaughlin, Hannah S Coalson, Todd E Fox, Su-Fern Tan, David J Feith, Mark Kester, Thomas P Loughran, David F Claxton, Myles C Cabot.

  Modifications in sphingolipid (SL) metabolism and mitochondrial bioenergetics are key factors implicated in cancer cell response to chemotherapy, including chemotherapy resistance. In the present work, we utilized acute myeloid leukemia (AML) cell lines, selected to be refractory to various chemotherapeutics, to explore the interplay between SL metabolism and mitochondrial biology supportive of multidrug resistance (MDR). In agreement with previous findings in cytarabine or daunorubicin resistant AML cells, relative to chemosensitive wildtype controls, HL-60 cells refractory to vincristine (HL60/VCR) presented with alterations in SL enzyme expression and lipidome composition. Such changes were typified by upregulated expression of various ceramide detoxifying enzymes, as well as corresponding shifts in ceramide, glucosylceramide, and sphingomyelin (SM) molecular species. With respect to mitochondria, despite consistent increases in both basal respiration and maximal respiratory capacity, direct interrogation of the oxidative phosphorylation (OXPHOS) system revealed intrinsic deficiencies in HL60/VCR, as well as across multiple MDR model systems. Based on the apparent requirement for augmented SL and mitochondrial flux to support the MDR phenotype, we explored a combinatorial therapeutic paradigm designed to target each pathway. Remarkably, despite minimal cytotoxicity in peripheral blood mononuclear cells (PBMC), co-targeting SL metabolism, and respiratory complex I (CI) induced synergistic cytotoxicity consistently across multiple MDR leukemia models. Together, these data underscore the intimate connection between cellular sphingolipids and mitochondrial metabolism and suggest that pharmacological intervention across both pathways may represent a novel treatment strategy against MDR.

Keywords:  HL-60 cells; acute myeloid leukemia; chemotherapy resistance; mitochondrial bioenergetics; sphingolipids; vincristine

DOI:  https://doi.org/10.1096/fj.202101194RRR
J Biochem. 2021 Dec 04. pii: mvab136. [Epub ahead of print]

Mitochondria Metabolomics Reveals a Role of β-Nicotinamide Mononucleotide Metabolism in Mitochondrial DNA Replication.

Tomoko Nomiyama, Daiki Setoyama, Takehiro Yasukawa, Dongchon Kang.

  Mitochondrial DNA (mtDNA) replication is tightly regulated and necessary for cellular homeostasis; however, its relationship with mitochondrial metabolism remains unclear. Advances in metabolomics integrated with the rapid isolation of mitochondria will allow for remarkable progress in analyzing mitochondrial metabolism. Here, we propose a novel methodology for mitochondria-targeted metabolomics, which employs a quick isolation procedure using a hemolytic toxin from Streptococcus pyogenes streptolysin O (SLO). SLO-isolation of mitochondria from cultured HEK293 cells is time- and labor-saving for simultaneous multi-sample processing and has been applied to various other cell lines in this study. Furthermore, our method can detect the time-dependent reduction in mitochondrial ATP in response to a glycolytic inhibitor 2-deoxyglucose, indicating the suitability to prepare metabolite analysis-competent mitochondria. Using this methodology, we searched for specific mitochondrial metabolites associated with mtDNA replication activation, and nucleotides and NAD+ were identified to be prominently altered. Most notably, treatment of β-Nicotinamide Mononucleotide (β-NMN), a precursor of NAD+, to HEK293 cells activated and improved the rate of mtDNA replication by increasing nucleotides in mitochondria and decreasing their degradation products: nucleosides. Our results suggest that β-NMN metabolism play a role in supporting mtDNA replication by maintaining the nucleotide pool balance in the mitochondria.

Keywords:  beta-nicotinamide mononucleotide (β-NMN); metabolomics; mitochondrial DNA; nucleotide metabolism; streptolysin O

DOI:  https://doi.org/10.1093/jb/mvab136
Biochem Soc Trans. 2021 Dec 09. pii: BST20211032. [Epub ahead of print]

Controlled power: how biology manages succinate-driven energy release.

Shona A Mookerjee, Akos A Gerencser, Mark A Watson, Martin D Brand.

  Oxidation of succinate by mitochondria can generate a higher protonmotive force (pmf) than can oxidation of NADH-linked substrates. Fundamentally, this is because of differences in redox potentials and gearing. Biology adds kinetic constraints that tune the oxidation of NADH and succinate to ensure that the resulting mitochondrial pmf is suitable for meeting cellular needs without triggering pathology. Tuning within an optimal range is used, for example, to shift ATP consumption between different consumers. Conditions that overcome these constraints and allow succinate oxidation to drive pmf too high can cause pathological generation of reactive oxygen species. We discuss the thermodynamic properties that allow succinate oxidation to drive pmf higher than NADH oxidation, and discuss the evidence for kinetic tuning of ATP production and for pathologies resulting from substantial succinate oxidation in vivo.

Keywords:  bioenergetics; ischaemia-reperfusion injury; membrane potential; mitochondria; reactive oxygen species; thermodynamics

DOI:  https://doi.org/10.1042/BST20211032
Nat Commun. 2021 Dec 06. 12(1): 6997

2-Deoxy-D-glucose couples mitochondrial DNA replication with mitochondrial fitness and promotes the selection of wild-type over mutant mitochondrial DNA.

Boris Pantic, Daniel Ives, Mara Mennuni, Diego Perez-Rodriguez, Uxoa Fernandez-Pelayo, Amaia Lopez de Arbina, Mikel Muñoz-Oreja, Marina Villar-Fernandez, Thanh-Mai Julie Dang, Lodovica Vergani, Iain G Johnston, Robert D S Pitceathly, Robert McFarland, Michael G Hanna, Robert W Taylor, Ian J Holt, Antonella Spinazzola.

Pathological variants of human mitochondrial DNA (mtDNA) typically co-exist with wild-type molecules, but the factors driving the selection of each are not understood. Because mitochondrial fitness does not favour the propagation of functional mtDNAs in disease states, we sought to create conditions where it would be advantageous. Glucose and glutamine consumption are increased in mtDNA dysfunction, and so we targeted the use of both in cells carrying the pathogenic m.3243A>G variant with 2-Deoxy-D-glucose (2DG), or the related 5-thioglucose. Here, we show that both compounds selected wild-type over mutant mtDNA, restoring mtDNA expression and respiration. Mechanistically, 2DG selectively inhibits the replication of mutant mtDNA; and glutamine is the key target metabolite, as its withdrawal, too, suppresses mtDNA synthesis in mutant cells. Additionally, by restricting glucose utilization, 2DG supports functional mtDNAs, as glucose-fuelled respiration is critical for mtDNA replication in control cells, when glucose and glutamine are scarce. Hence, we demonstrate that mitochondrial fitness dictates metabolite preference for mtDNA replication; consequently, interventions that restrict metabolite availability can suppress pathological mtDNAs, by coupling mitochondrial fitness and replication.

DOI: https://doi.org/10.1038/s41467-021-26829-0
Cancers (Basel). 2021 Dec 06. pii: 6142. [Epub ahead of print]13(23):

Metabolic Rewiring Is Essential for AML Cell Survival to Overcome Autophagy Inhibition by Loss of ATG3.

Fatima Baker, Ibrahim H Polat, Khalil Abou-El-Ardat, Islam Alshamleh, Marlyn Thoelken, Daniel Hymon, Andrea Gubas, Sebastian E Koschade, Jonas B Vischedyk, Manuel Kaulich, Harald Schwalbe, Shabnam Shaid, Christian H Brandts.

  Autophagy is an important survival mechanism that allows recycling of nutrients and removal of damaged organelles and has been shown to contribute to the proliferation of acute myeloid leukemia (AML) cells. However, little is known about the mechanism by which autophagy- dependent AML cells can overcome dysfunctional autophagy. In our study we identified autophagy related protein 3 (ATG3) as a crucial autophagy gene for AML cell proliferation by conducting a CRISPR/Cas9 dropout screen with a library targeting around 200 autophagy-related genes. shRNA-mediated loss of ATG3 impaired autophagy function in AML cells and increased their mitochondrial activity and energy metabolism, as shown by elevated mitochondrial ROS generation and mitochondrial respiration. Using tracer-based NMR metabolomics analysis we further demonstrate that the loss of ATG3 resulted in an upregulation of glycolysis, lactate production, and oxidative phosphorylation. Additionally, loss of ATG3 strongly sensitized AML cells to the inhibition of mitochondrial metabolism. These findings highlight the metabolic vulnerabilities that AML cells acquire from autophagy inhibition and support further exploration of combination therapies targeting autophagy and mitochondrial metabolism in AML.

Keywords:  ATG3; acute myeloid leukemia; autophagy; autophagy inhibition; metabolic rewiring

DOI:  https://doi.org/10.3390/cancers13236142
Front Nutr. 2021 ;8 775382

Differential Effects of Oleic and Palmitic Acids on Lipid Droplet-Mitochondria Interaction in the Hepatic Cell Line HepG2.

Andrea Eynaudi, Francisco Díaz-Castro, Juan Carlos Bórquez, Roberto Bravo-Sagua, Valentina Parra, Rodrigo Troncoso.

  Fatty acid overload, either of the saturated palmitic acid (PA) or the unsaturated oleic acid (OA), causes triglyceride accumulation into specialized organelles termed lipid droplets (LD). However, only PA overload leads to liver damage mediated by mitochondrial dysfunction. Whether these divergent outcomes stem from differential effects of PA and OA on LD and mitochondria joint dynamics remains to be uncovered. Here, we contrast how both fatty acids impact the morphology and interaction between both organelles and mitochondrial bioenergetics in HepG2 cells. Using confocal microscopy, we showed that short-term (2-24 h) OA overload promotes more and bigger LD accumulation than PA. Oxygen polarography indicated that both treatments stimulated mitochondrial respiration; however, OA favored an overall build-up of the mitochondrial potential, and PA evoked mitochondrial fragmentation, concomitant with an ATP-oriented metabolism. Even though PA-induced a lesser increase in LD-mitochondria proximity than OA, those LD associated with highly active mitochondria suggest that they interact mainly to fuel fatty acid oxidation and ATP synthesis (that is, metabolically "active" LD). On the contrary, OA overload seemingly stimulated LD-mitochondria interaction mainly for LD growth (thus metabolically "passive" LDs). In sum, these differences point out that OA readily accumulates in LD, likely reducing their toxicity, while PA preferably stimulates mitochondrial oxidative metabolism, which may contribute to liver damage progression.

Keywords:  fatty acids; hepatocytes; lipid droplets; mitochondria; oxygen consumption

DOI:  https://doi.org/10.3389/fnut.2021.775382
Elife. 2021 Dec 09. pii: e71978. [Epub ahead of print]10

Reverting the mode of action of the mitochondrial FOF1-ATPase by Legionella pneumophila preserves its replication niche.

Pedro Escoll, Lucien Platon, Mariatou Dramé, Tobias Sahr, Silke Schmidt, Christophe Rusniok, Carmen Buchrieser.

  Legionella pneumophila, the causative agent of Legionnaires'; disease, a severe pneumonia, injects via a type-IV-secretion-system (T4SS) more than 300 proteins into macrophages, its main host cell in humans. Certain of these proteins are implicated in reprogramming the metabolism of infected cells by reducing mitochondrial oxidative phosphorylation (OXPHOS) early after infection. Here we show that despite reduced OXPHOS, the mitochondrial membrane potential (Δψm) is maintained during infection of primary human monocyte-derived macrophages (hMDMs). We reveal that L. pneumophila reverses the ATP-synthase activity of the mitochondrial FOF1-ATPase to ATP-hydrolase activity in a T4SS-dependent manner, which leads to a conservation of the Δψm, preserves mitochondrial polarization and prevents macrophage cell death. Analyses of T4SS effectors known to target mitochondrial functions revealed that LpSpl is partially involved in conserving the Δψm, but not LncP and MitF. The inhibition of the L. pneumophila-induced 'reverse mode' of the FOF1-ATPase collapsed the Δψm and caused cell death in infected cells. Single-cell analyses suggested that bacterial replication occurs preferentially in hMDMs that conserved the Δψm and showed delayed cell death. This direct manipulation of the mode of activity of the FOF1-ATPase is a newly identified feature of L. pneumophila allowing to delay host cell death and thereby to preserve the bacterial replication niche during infection.

Keywords:  infectious disease; microbiology

DOI:  https://doi.org/10.7554/eLife.71978
J Gerontol A Biol Sci Med Sci. 2021 Dec 07. pii: glab356. [Epub ahead of print]

Ameliorating effects of lifelong physical activity on healthy aging and mitochondrial function in human white adipose tissue.

Anders Gudiksen, Albina Qoqaj, Stine Ringholm, Jørgen Wojtaszewski, Peter Plomgaard, Henriette Pilegaard.

  Growing old is patently among the most prominent risk factors for lifestyle related diseases and deterioration in physical performance. Aging in particular affects mitochondrial homeostasis and maintaining a well-functioning mitochondrial pool is imperative in order to avoid age-associated metabolic decline. White adipose tissue (WAT) is a key organ in energy balance and impaired mitochondrial function in adipocytes has been associated with increased low-grade inflammation, altered metabolism, excessive ROS production and an accelerated aging phenotype. Exercise training improves mitochondrial health but whether lifelong exercise training can sufficiently maintain WAT mitochondrial function is currently unknown. Therefore, to dissect the role and dose-dependence of lifelong exercise training on aging WAT metabolic parameters and mitochondrial function, young and older untrained, as well as moderately and highly exercise trained older male subjects were recruited and abdominal subcutaneous (s)WAT biopsies and venous blood samples were obtained to measure mitochondrial function and key metabolic factors in WAT and plasma. Mitochondrial intrinsic respiratory capacity was lower in sWAT from older than in young subjects. In spite of this, maximal mitochondrial respiration per wet weight, markers of oxidative capacity, and mitophagic capacity were increased in sWAT from lifelong highly exercise trained than all other groups. Furthermore, ROS emission was generally lower in sWAT from lifelong highly exercise trained than older untrained subjects. Taken together, aging reduces intrinsic mitochondrial respiration in human sWAT, but lifelong high volume exercise training increases oxidative capacity by increasing mitochondrial volume likely contributing to healthy aging.

Keywords:  ADIPOSE TISSUE; EXERCISE TRAINING; METABOLISM; MITOCHONDRIA; ROS

DOI:  https://doi.org/10.1093/gerona/glab356
Sci Rep. 2021 Dec 07. 11(1): 23549

Nm23-H1 activator phenylbutenoid dimer exerts cytotoxic effects on metastatic breast cancer cells by inducing mitochondrial dysfunction only under glucose starvation.

Bokyung Kim, Jae-Jin Lee, Ji Soo Shin, Ji-Wan Suh, Sunhee Jung, Geum-Sook Hwang, Hee-Yoon Lee, Kong-Joo Lee.

Mitochondrial oxidative phosphorylation (OXPHOS) has become an attractive target in anti-cancer studies in recent years. In this study, we found that a small molecule phenylbutenoid dimer NMac1 (Nm23-H1 activator 1), (±)-trans-3-(3,4-dimethoxyphenyl)-4-[(E)-3,4-dimethoxystyryl]cyclohex-1-ene, a previously identified anti-metastatic agent, has novel anti-proliferative effect only under glucose starvation in metastatic breast cancer cells. NMac1 causes significant activation of AMPK by decreasing ATP synthesis, lowers mitochondrial membrane potential (MMP, ΔΨm), and inhibits oxygen consumption rate (OCR) under glucose starvation. These effects of NMac1 are provoked by a consequence of OXPHOS complex I inhibition. Through the structure-activity relationship (SAR) study of NMac1 derivatives, NMac24 was identified as the most effective compound in anti-proliferation. NMac1 and NMac24 effectively suppress cancer cell proliferation in 3D-spheroid in vivo-like models only under glucose starvation. These results suggest that NMac1 and NMac24 have the potential as anti-cancer agents having cytotoxic effects selectively in glucose restricted cells.

DOI: https://doi.org/10.1038/s41598-021-02729-7
Cell Metab. 2021 Dec 07. pii: S1550-4131(21)00539-8. [Epub ahead of print]33(12): 2380-2397.e9

NEAT1 is essential for metabolic changes that promote breast cancer growth and metastasis.

Mi Kyung Park, Li Zhang, Kyung-Won Min, Jung-Hyun Cho, Chih-Chen Yeh, Hyesu Moon, Daniel Hormaechea-Agulla, Hyejin Mun, Seungbeom Ko, Ji Won Lee, Sonali Jathar, Aubrey S Smith, Yixin Yao, Nguyen Thu Giang, Hong Ha Vu, Victoria C Yan, Mary C Bridges, Antonis Kourtidis, Florian Muller, Jeong Ho Chang, Su Jung Song, Shinichi Nakagawa, Tetsuro Hirose, Je-Hyun Yoon, Min Sup Song.

  Accelerated glycolysis is the main metabolic change observed in cancer, but the underlying molecular mechanisms and their role in cancer progression remain poorly understood. Here, we show that the deletion of the long noncoding RNA (lncRNA) Neat1 in MMTV-PyVT mice profoundly impairs tumor initiation, growth, and metastasis, specifically switching off the penultimate step of glycolysis. Mechanistically, NEAT1 directly binds and forms a scaffold bridge for the assembly of PGK1/PGAM1/ENO1 complexes and thereby promotes substrate channeling for high and efficient glycolysis. Notably, NEAT1 is upregulated in cancer patients and correlates with high levels of these complexes, and genetic and pharmacological blockade of penultimate glycolysis ablates NEAT1-dependent tumorigenesis. Finally, we demonstrate that Pinin mediates glucose-stimulated nuclear export of NEAT1, through which it exerts isoform-specific and paraspeckle-independent functions. These findings establish a direct role for NEAT1 in regulating tumor metabolism, provide new insights into the Warburg effect, and identify potential targets for therapy.

Keywords:  ENO1; NEAT1; PGAM1; PGK1; Pinin; Warburg effect; aerobic glycolysis; breast cancer; long noncoding RNA; tumor metabolism

DOI:  https://doi.org/10.1016/j.cmet.2021.11.011
Nat Cell Biol. 2021 Dec 06.

MIROs and DRP1 drive mitochondrial-derived vesicle biogenesis and promote quality control.

Tim König, Hendrik Nolte, Mari J Aaltonen, Takashi Tatsuta, Michiel Krols, Thomas Stroh, Thomas Langer, Heidi M McBride.

Mitochondrial-derived vesicles (MDVs) are implicated in diverse physiological processes-for example, mitochondrial quality control-and are linked to various neurodegenerative diseases. However, their specific cargo composition and complex molecular biogenesis are still unknown. Here we report the proteome and lipidome of steady-state TOMM20+ MDVs. We identified 107 high-confidence MDV cargoes, which include all β-barrel proteins and the TOM import complex. MDV cargoes are delivered as fully assembled complexes to lysosomes, thus representing a selective mitochondrial quality control mechanism for multi-subunit complexes, including the TOM machinery. Moreover, we define key biogenesis steps of phosphatidic acid-enriched MDVs starting with the MIRO1/2-dependent formation of thin membrane protrusions pulled along microtubule filaments, followed by MID49/MID51/MFF-dependent recruitment of the dynamin family GTPase DRP1 and finally DRP1-dependent scission. In summary, we define the function of MDVs in mitochondrial quality control and present a mechanistic model for global GTPase-driven MDV biogenesis.

DOI: https://doi.org/10.1038/s41556-021-00798-4
Cancers (Basel). 2021 Nov 30. pii: 6029. [Epub ahead of print]13(23):

Mitochondrial Plasticity Promotes Resistance to Sorafenib and Vulnerability to STAT3 Inhibition in Human Hepatocellular Carcinoma.

Shusil K Pandit, Giada Sandrini, Jessica Merulla, Valentina Nobili, Xin Wang, Alessia Zangari, Andrea Rinaldi, Dheeraj Shinde, Giuseppina M Carbone, Carlo V Catapano.

  The multi-kinase inhibitor sorafenib is a primary treatment modality for advanced-stage hepatocellular carcinoma (HCC). However, the therapeutic benefits are short-lived due to innate and acquired resistance. Here, we examined how HCC cells respond to sorafenib and adapt to continuous and prolonged exposure to the drug. Sorafenib-adapted HCC cells show a profound reprogramming of mitochondria function and marked activation of genes required for mitochondrial protein translation and biogenesis. Mitochondrial ribosomal proteins and components of translation and import machinery are increased in sorafenib-resistant cells and sorafenib-refractory HCC patients show similar alterations. Sorafenib-adapted cells also exhibited increased serine 727 phosphorylated (pSer727) STAT3, the prevalent form in mitochondria, suggesting that STAT3 might be an actionable target to counteract resistance. Consistently, a small-molecule STAT3 inhibitor reduces pSer727, reverts mitochondrial alterations, and enhances the response to sorafenib in resistant cells. These results sustain the importance of mitochondria plasticity in response to sorafenib and identify a clinically actionable strategy for improving the treatment efficacy in HCC patients.

Keywords:  OPB-111077; STAT3; drug resistance; hepatocellular carcinoma; liver cancer; mitochondria; mitochondrial protein translation; mitochondrial ribosomal proteins; sorafenib; tyrosine kinase inhibitors

DOI:  https://doi.org/10.3390/cancers13236029
EMBO J. 2021 Dec 07. e106837

Molecular species selectivity of lipid transport creates a mitochondrial sink for di-unsaturated phospholipids.

Mike F Renne, Xue Bao, Margriet Wj Hokken, Adolf S Bierhuizen, Martin Hermansson, Richard R Sprenger, Tom A Ewing, Xiao Ma, Ruud C Cox, Jos F Brouwers, Cedric H De Smet, Christer S Ejsing, Anton Ipm de Kroon.

  Mitochondria depend on the import of phospholipid precursors for the biosynthesis of phosphatidylethanolamine (PE) and cardiolipin, yet the mechanism of their transport remains elusive. A dynamic lipidomics approach revealed that mitochondria preferentially import di-unsaturated phosphatidylserine (PS) for subsequent conversion to PE by the mitochondrial PS decarboxylase Psd1p. Several protein complexes tethering mitochondria to the endomembrane system have been implicated in lipid transport in yeast, including the endoplasmic reticulum (ER)-mitochondrial encounter structure (ERMES), ER-membrane complex (EMC), and the vacuole and mitochondria patch (vCLAMP). By limiting the availability of unsaturated phospholipids, we created conditions to investigate the mechanism of lipid transfer and the contributions of the tethering complexes in vivo. Under these conditions, inactivation of ERMES components or of the vCLAMP component Vps39p exacerbated accumulation of saturated lipid acyl chains, indicating that ERMES and Vps39p contribute to the mitochondrial sink for unsaturated acyl chains by mediating transfer of di-unsaturated phospholipids. These results support the concept that intermembrane lipid flow is rate-limited by molecular species-dependent lipid efflux from the donor membrane and driven by the lipid species' concentration gradient between donor and acceptor membrane.

Keywords:  lipid transport; membrane contact sites; membrane lipid homeostasis; membrane lipid unsaturation; mitochondria

DOI:  https://doi.org/10.15252/embj.2020106837
Leuk Lymphoma. 2021 Dec 08. 1-9

Inhibition of pyrimidine biosynthesis by strobilurin derivatives induces differentiation of acute myeloid leukemia cells.

Xing Shen, Lu Zhang, Shuang Xing, Xue-Wen Zhang, Guo-Lin Xiong, Yu-Wen Cong, He Xiao, Xin-Ru Wang, Zu-Yin Yu.

  All-trans retinoic acid-based differentiation therapies have succeeded in the treatment of acute promyelocytic leukemia, which is a rare subtype of acute myeloid leukemia (AML). Their clinical efficacy is negligible, however, for other subtypes of AML. Here, we showed that strobilurin derivatives, a well-established class of inhibitors of mitochondrial electron transport chain (ETC) complex III, possessed differentiation-inducing activity in AML cells. Impairment of mitochondrial ETC activity was involved in the differentiation effects of strobilurin derivatives, where reactive oxygen species generation appeared unnecessary. Conversely, strobilurin derivative-mediated differentiation was triggered by pyrimidine deficiency, which resulted from the inhibition of the mitochondrial-coupled dihydroorotate dehydrogenase enzyme. Moreover, strobilurin derivative-mediated pyrimidine depletion led to the activation of the Akt/mTOR cascade, which was required for the differentiation. Our study provided evidence that strobilurin derivatives may represent a novel class of differentiation-inducing agents for the treatment of AML.

Keywords:  Akt; Strobilurin derivatives; acute myeloid leukemia; differentiation; mitochondria; pyrimidines biosynthesis

DOI:  https://doi.org/10.1080/10428194.2021.2008382
J Am Chem Soc. 2021 Dec 06.

Monitoring and Modulating mtDNA G-Quadruplex Dynamics Reveal Its Close Relationship to Cell Glycolysis.

Xiu-Cai Chen, Gui-Xue Tang, Wen-Hua Luo, Wen Shao, Jing Dai, Shu-Tang Zeng, Zhi-Shu Huang, Shuo-Bin Chen, Jia-Heng Tan.

The mitochondrial DNA G-quadruplex (mtDNA G4) is a potential regulatory element for the regulation of mitochondrial functions; however, its relevance and specific roles in diseases remain largely unknown. Here, we engineered a set of chemical probes, including MitoISCH, an mtDNA G4-specific fluorescent probe, together with MitoPDS, a mitochondria-targeted G4-stabilizing agent, to thoroughly investigate mtDNA G4s. Using MitoISCH to monitor previously intractable dynamics of mtDNA G4s, we surprisingly found that their formation was prevalent only in endothelial and cancer cells that rely on glycolysis for energy production. Consistent with this, promotion of mtDNA G4 folding by MitoPDS in turn caused glycolysis-related gene activation and glycolysis enhancement. Remarkably, this close relationship among mtDNA G4s, glycolysis, and cancer cells further allowed MitoISCH to accumulate in tumors and label them in vivo. Our work reveals an unprecedented link between mtDNA G4s and cell glycolysis, suggesting that mtDNA G4s may be a novel cancer biomarker and therapeutic target deserving further exploration.

DOI: https://doi.org/10.1021/jacs.1c08860
J Inherit Metab Dis. 2021 Dec 06.

Identification and characterisation of novel MPC1 gene variants causing mitochondrial pyruvate carrier deficiency.

Huafang Jiang, Ahmad Alahmad, Song Fu, Xiaoling Fu, Zhimei Liu, Xiaodi Han, Lanlan Li, Tianyu Song, Manting Xu, Shanshan Liu, Junling Wang, Buthaina Albash, Ahmad Alaqeel, Vasilescu Catalina, Holger Prokisch, Robert W Taylor, Robert McFarland, Fang Fang.



Keywords:  Leigh-like syndrome; MPC1; glutamine; mitochondria; mitochondrial pyruvate carrier deficiency; treatment

DOI:  https://doi.org/10.1002/jimd.12462
STAR Protoc. 2021 Dec 17. 2(4): 100979

Quantitative assays to measure the transport activity of the mitochondrial calcium uniporter in cell lines or Xenopus oocytes.

Madison X Rodriguez, Anna M Van Keuren, Ming-Feng Tsai.

  The mitochondrial calcium uniporter, which mediates mitochondrial Ca2+ uptake, regulates key cellular functions, including intracellular Ca2+ signaling, cell-fate determination, and mitochondrial bioenergetics. Here, we describe two complementary strategies to quantify the uniporter's transport activity. First, we detail a mitochondrial Ca2+ radionuclide uptake assay in cultured cell lines. Second, we describe electrophysiological recordings of the uniporter expressed in Xenopus oocytes. These approaches enable a detailed kinetic analysis of the uniporter to link its molecular properties to physiological functions. For complete details on the use and execution of this protocol, please refer to Tsai and Tsai (2018) and Phillips et al. (2019).

Keywords:  Biophysics; Cell Biology; Cell-based Assays; Molecular Biology

DOI:  https://doi.org/10.1016/j.xpro.2021.100979
Per Med. 2021 Dec 07.

Somatic mitochondrial DNA mutations in different grades of glioma.

Bee Hong Soon, Nadiah Abu, Nor Azian Abdul Murad, Sue-Mian Then, Azizi Abu Bakar, Farizal Fadzil, Jegan Thanabalan, Mohd Saffari Mohd Haspani, Charng Jeng Toh, Ramesh Kumar, Ainul Syahrilfazli Jaafar, Anis Nabillah Mohd Azli, Mohd Syakir Mohd Azahar, Sanmugarajah Paramasvaran, Kamalanathan Palaniandy, Azmi Mohd Tamil, Rahman Jamal.

  Aim: Mitochondrial DNA (mtDNA) alterations play an important role in the multistep processes of cancer development. Gliomas are among the most diagnosed brain cancer. The relationship between mtDNA alterations and different grades of gliomas are still elusive. This study aimed to elucidate the profile of somatic mtDNA mutations in different grades of gliomas and correlate it with clinical phenotype. Materials & methods: Forty histopathologically confirmed glioma tissue samples and their matched blood were collected and subjected for mtDNA sequencing. Results & conclusion: About 75% of the gliomas harbored at least one somatic mutation in the mtDNA gene, and 45% of these mutations were pathogenic. Mutations were scattered across the mtDNA genome, and the commonest nonsynonymous mutations were located at complex I and IV of the mitochondrial respiratory chain. These findings may have implication for future research to determine the mitochondrial energetics and its downstream metabolomics on gliomas.

Keywords:  Malaysia; cancer; genome; glioblastoma; gliomas; microarray; mitochondria; mtDNA; mutation; somatic mutation

DOI:  https://doi.org/10.2217/pme-2021-0033
Nat Commun. 2021 Dec 10. 12(1): 7219

Acute RyR1 Ca2+ leak enhances NADH-linked mitochondrial respiratory capacity.

Nadège Zanou, Haikel Dridi, Steven Reiken, Tanes Imamura de Lima, Chris Donnelly, Umberto De Marchi, Manuele Ferrini, Jeremy Vidal, Leah Sittenfeld, Jerome N Feige, Pablo M Garcia-Roves, Isabel C Lopez-Mejia, Andrew R Marks, Johan Auwerx, Bengt Kayser, Nicolas Place.

Sustained ryanodine receptor (RyR) Ca2+ leak is associated with pathological conditions such as heart failure or skeletal muscle weakness. We report that a single session of sprint interval training (SIT), but not of moderate intensity continuous training (MICT), triggers RyR1 protein oxidation and nitrosylation leading to calstabin1 dissociation in healthy human muscle and in in vitro SIT models (simulated SIT or S-SIT). This is accompanied by decreased sarcoplasmic reticulum Ca2+ content, increased levels of mitochondrial oxidative phosphorylation proteins, supercomplex formation and enhanced NADH-linked mitochondrial respiratory capacity. Mechanistically, (S-)SIT increases mitochondrial Ca2+ uptake in mouse myotubes and muscle fibres, and decreases pyruvate dehydrogenase phosphorylation in human muscle and mouse myotubes. Countering Ca2+ leak or preventing mitochondrial Ca2+ uptake blunts S-SIT-induced adaptations, a result supported by proteomic analyses. Here we show that triggering acute transient Ca2+ leak through RyR1 in healthy muscle may contribute to the multiple health promoting benefits of exercise.

DOI: https://doi.org/10.1038/s41467-021-27422-1
Biochem Biophys Res Commun. 2021 Dec 01. pii: S0006-291X(21)01627-2. [Epub ahead of print]587 153-159

A novel PDK1 inhibitor, JX06, inhibits glycolysis and induces apoptosis in multiple myeloma cells.

Yawara Kawano, Takayuki Sasano, Yuichiro Arima, Saki Kushima, Kenichi Tsujita, Masao Matsuoka, Hiroyuki Hata.

  Pyruvate dehydrogenase kinase 1 (PDK1) is a Ser/Thr kinase that inactivates mitochondrial pyruvate dehydrogenase (PDH), leading to switch of glucose metabolism from mitochondrial oxidation to aerobic glycolysis. We previously reported that PDK1 inhibition is a potent therapeutic strategy in multiple myeloma (MM). However, availability of PDK1 inhibitors, which are effective at low concentrations, are limited at present, making PDK1 inhibition difficult to apply in the clinic. In the present study, we examined the efficacy and mechanism of action of JX06, a novel PDK1 inhibitor, against MM cells. We confirmed that PDK1 is highly expressed in normal plasma cells and MM cells using publicly available gene expression datasets. JX06 suppressed cell growth and induced apoptosis against MM cells from approximately 0.5 μM JX06 treatment reduced PDH phosphorylation, suggesting that JX06 is indeed inhibiting PDK1. Intracellular metabolite analysis revealed that JX06 treatment reduced metabolites associated with glucose metabolism of MM cells. Additionally, JX06 in combination with a well-known proteasome inhibitor, bortezomib, significantly increased MM cell death, which raises the possibility of combination use of JX06 with proteasome inhibitors in the clinic. These findings demonstrate that PDK1 can be potentially targeted by JX06 in MM through glycolysis inhibition, leading to a novel therapeutic strategy in MM.

Keywords:  JX06; Multiple myeloma; PDK1

DOI:  https://doi.org/10.1016/j.bbrc.2021.11.102
Nat Metab. 2021 Dec 06.

The mitochondrial protein Opa1 promotes adipocyte browning that is dependent on urea cycle metabolites.

Camilla Bean, Matteo Audano, Tatiana Varanita, Francesca Favaretto, Marta Medaglia, Marco Gerdol, Lena Pernas, Fabio Stasi, Marta Giacomello, Stèphanie Herkenne, Maheswany Muniandy, Sini Heinonen, Emma Cazaly, Miina Ollikainen, Gabriella Milan, Alberto Pallavicini, Kirsi H Pietiläinen, Roberto Vettor, Nico Mitro, Luca Scorrano.

White to brown/beige adipocytes conversion is a possible therapeutic strategy to tackle the current obesity epidemics. While mitochondria are key for energy dissipation in brown fat, it is unknown if they can drive adipocyte browning. Here, we show that the mitochondrial cristae biogenesis protein optic atrophy 1 (Opa1) facilitates cell-autonomous adipocyte browning. In two cohorts of patients with obesity, including weight discordant monozygotic twin pairs, adipose tissue OPA1 levels are reduced. In the mouse, Opa1 overexpression favours white adipose tissue expandability as well as browning, ultimately improving glucose tolerance and insulin sensitivity. Transcriptomics and metabolomics analyses identify the Jumanji family chromatin remodelling protein Kdm3a and urea cycle metabolites, including fumarate, as effectors of Opa1-dependent browning. Mechanistically, the higher cyclic adenosine monophosphate (cAMP) levels in Opa1 pre-adipocytes activate cAMP-responsive element binding protein (CREB), which transcribes urea cycle enzymes. Flux analyses in pre-adipocytes indicate that Opa1-dependent fumarate accumulation depends on the urea cycle. Conversely, adipocyte-specific Opa1 deletion curtails urea cycle and beige differentiation of pre-adipocytes, and is rescued by fumarate supplementation. Thus, the urea cycle links the mitochondrial dynamics protein Opa1 to white adipocyte browning.

DOI: https://doi.org/10.1038/s42255-021-00497-2
Sci Adv. 2021 Dec 10. 7(50): eabi5657

Mitochondrial DNA heteroplasmy is modulated during oocyte development propagating mutation transmission.

Haixin Zhang, Marco Esposito, Mikael G Pezet, Juvid Aryaman, Wei Wei, Florian Klimm, Claudia Calabrese, Stephen P Burr, Carolina H Macabelli, Carlo Viscomi, Mitinori Saitou, Marcos R Chiaratti, James B Stewart, Nick Jones, Patrick F Chinnery.

[Figure: see text].

DOI: https://doi.org/10.1126/sciadv.abi5657
Nat Cell Biol. 2021 Dec 06.

Surveying the mitochondrial proteome.

Dominic Winter, Thomas Becker.

DOI: https://doi.org/10.1038/s41556-021-00801-y
iScience. 2021 Dec 17. 24(12): 103460

Prolyl endopeptidase-like is a (thio)esterase involved in mitochondrial respiratory chain function.

Karen Rosier, Molly T McDevitt, Joél Smet, Brendan J Floyd, Maxime Verschoore, Maria J Marcaida, Craig A Bingman, Irma Lemmens, Matteo Dal Peraro, Jan Tavernier, Benjamin F Cravatt, Natalia V Gounko, Katlijn Vints, Yenthe Monnens, Kritika Bhalla, Laetitia Aerts, Edrees H Rashan, Arnaud V Vanlander, Rudy Van Coster, Luc Régal, David J Pagliarini, John W M Creemers.

  Deficiency of the serine hydrolase prolyl endopeptidase-like (PREPL) causes a recessive metabolic disorder characterized by neonatal hypotonia, feeding difficulties, and growth hormone deficiency. The pathophysiology of PREPL deficiency and the physiological substrates of PREPL remain largely unknown. In this study, we connect PREPL with mitochondrial gene expression and oxidative phosphorylation by analyzing its protein interactors. We demonstrate that the long PREPLL isoform localizes to mitochondria, whereas PREPLS remains cytosolic. Prepl KO mice showed reduced mitochondrial complex activities and disrupted mitochondrial gene expression. Furthermore, mitochondrial ultrastructure was abnormal in a PREPL-deficient patient and Prepl KO mice. In addition, we reveal that PREPL has (thio)esterase activity and inhibition of PREPL by Palmostatin M suggests a depalmitoylating function. We subsequently determined the crystal structure of PREPL, thereby providing insight into the mechanism of action. Taken together, PREPL is a (thio)esterase rather than a peptidase and PREPLL is involved in mitochondrial homeostasis.

Keywords:  Molecular biology; Molecular medicine; Structural biology

DOI:  https://doi.org/10.1016/j.isci.2021.103460
Nat Commun. 2021 Dec 08. 12(1): 7139

A predominant enhancer co-amplified with the SOX2 oncogene is necessary and sufficient for its expression in squamous cancer.

Yanli Liu, Zhong Wu, Jin Zhou, Dinesh K A Ramadurai, Katelyn L Mortenson, Estrella Aguilera-Jimenez, Yifei Yan, Xiaojun Yang, Alison M Taylor, Katherine E Varley, Jason Gertz, Peter S Choi, Andrew D Cherniack, Xingdong Chen, Adam J Bass, Swneke D Bailey, Xiaoyang Zhang.

Amplification and overexpression of the SOX2 oncogene represent a hallmark of squamous cancers originating from diverse tissue types. Here, we find that squamous cancers selectively amplify a 3' noncoding region together with SOX2, which harbors squamous cancer-specific chromatin accessible regions. We identify a single enhancer e1 that predominantly drives SOX2 expression. Repression of e1 in SOX2-high cells causes collapse of the surrounding enhancers, remarkable reduction in SOX2 expression, and a global transcriptional change reminiscent of SOX2 knockout. The e1 enhancer is driven by a combination of transcription factors including SOX2 itself and the AP-1 complex, which facilitates recruitment of the co-activator BRD4. CRISPR-mediated activation of e1 in SOX2-low cells is sufficient to rebuild the e1-SOX2 loop and activate SOX2 expression. Our study shows that squamous cancers selectively amplify a predominant enhancer to drive SOX2 overexpression, uncovering functional links among enhancer activation, chromatin looping, and lineage-specific copy number amplifications of oncogenes.

DOI: https://doi.org/10.1038/s41467-021-27055-4
Methods Mol Biol. 2022 ;2385 153-160

Data Processing to Probe the Cellular Hydrogen Peroxide Landscape.

Fernando Antunes, Paula Brito.

  Hydrogen peroxide (H2O2) regulates signaling pathways by modulating the activity of redox-sensitive proteins denominated redox switches. The magnitude of the transient variations in localized H2O2 pools during signaling events and how these variations impact redox switches present in the cell remain elusive. A canonical model with two chemical reactions comprising the oxidation/reduction cycle of a redox switch is described. The model is dimensionless with respect to the redox switch concentration. Thus, the time-series data required to apply the equations deduced is the percentage of oxidation of a redox switch, avoiding the application of absolute concentrations that are often difficult to measure experimentally. Here, we describe detailed protocols for the processing of experimental data with the canonical model to probe the absolute concentrations of H2O2 found in the vicinity of redox switches and probes, as well as the kinetic parameters that describe the reduction and oxidation of redox switches. The protocols are an analytical tool that helps to depict the cellular hydrogen peroxide signaling landscape, giving new insights on H2O2 signaling mechanisms, and hold the potential to be a framework for a future redox kinetomics analytical platform.

Keywords:  Hydrogen peroxide; Kinetic parameters; Oxidation–reduction cycle; Rate constants; Redox biology

DOI:  https://doi.org/10.1007/978-1-0716-1767-0_8
Cancers (Basel). 2021 Nov 27. pii: 5966. [Epub ahead of print]13(23):

AMP-Activated Protein Kinase Contributes to Apoptosis Induced by the Bcl-2 Inhibitor Venetoclax in Acute Myeloid Leukemia.

Noémie Legrand, Amandine Pradier, Laury Poulain, Sarah Mouche, Rudy Birsen, Clément Larrue, Federico Simonetta, Jerome Tamburini.

  The treatment of acute myeloid leukemia (AML) remains a challenge especially among the elderly. The Bcl-2 inhibitor venetoclax recently showed significant survival benefits in AML patients when combined to low-dose cytarabine or azacitidine. Bcl-2 inhibition initiate mitochondrial apoptosis, but also respiration and cellular ATP production in AML. AMP-Activated Protein Kinase (AMPK) is a central energy sensor activated by increased AMP:ATP ratio to restore the cellular energy balance. Unexpectedly, we observed that venetoclax inhibited AMPK activity through caspase-dependent degradation of AMPK subunits in AML cells. On the other hand, genetic models of AMPK invalidation and re-expression suggested that AMPK participated to the early stages of apoptotic response through a negative regulation of multi-domain anti-apoptotic effectors such as Mcl-1 or Bcl-xL. Together our results suggested a new link between AMPK and Bcl-2-dependent mitochondrial apoptosis that participated to the anti-leukemic activity of venetoclax in AML.

Keywords:  AML; AMPK; venetoclax

DOI:  https://doi.org/10.3390/cancers13235966
Nat Commun. 2021 Dec 08. 12(1): 7115

Acetoacetate protects macrophages from lactic acidosis-induced mitochondrial dysfunction by metabolic reprograming.

Clément Adam, Léa Paolini, Naïg Gueguen, Guillaume Mabilleau, Laurence Preisser, Simon Blanchard, Pascale Pignon, Florence Manero, Morgane Le Mao, Alain Morel, Pascal Reynier, Céline Beauvillain, Yves Delneste, Vincent Procaccio, Pascale Jeannin.

Lactic acidosis, the extracellular accumulation of lactate and protons, is a consequence of increased glycolysis triggered by insufficient oxygen supply to tissues. Macrophages are able to differentiate from monocytes under such acidotic conditions, and remain active in order to resolve the underlying injury. Here we show that, in lactic acidosis, human monocytes differentiating into macrophages are characterized by depolarized mitochondria, transient reduction of mitochondrial mass due to mitophagy, and a significant decrease in nutrient absorption. These metabolic changes, resembling pseudostarvation, result from the low extracellular pH rather than from the lactosis component, and render these cells dependent on autophagy for survival. Meanwhile, acetoacetate, a natural metabolite produced by the liver, is utilized by monocytes/macrophages as an alternative fuel to mitigate lactic acidosis-induced pseudostarvation, as evidenced by retained mitochondrial integrity and function, retained nutrient uptake, and survival without the need of autophagy. Our results thus show that acetoacetate may increase tissue tolerance to sustained lactic acidosis.

DOI: https://doi.org/10.1038/s41467-021-27426-x
Mol Cell. 2021 Nov 26. pii: S1097-2765(21)00991-6. [Epub ahead of print]

Mechanism of PINK1 activation by autophosphorylation and insights into assembly on the TOM complex.

Shafqat Rasool, Simon Veyron, Naoto Soya, Mohamed A Eldeeb, Gergely L Lukacs, Edward A Fon, Jean-François Trempe.

  Mutations in PINK1 cause autosomal-recessive Parkinson's disease. Mitochondrial damage results in PINK1 import arrest on the translocase of the outer mitochondrial membrane (TOM) complex, resulting in the activation of its ubiquitin kinase activity by autophosphorylation and initiation of Parkin-dependent mitochondrial clearance. Herein, we report crystal structures of the entire cytosolic domain of insect PINK1. Our structures reveal a dimeric autophosphorylation complex targeting phosphorylation at the invariant Ser205 (human Ser228). The dimer interface requires insert 2, which is unique to PINK1. The structures also reveal how an N-terminal helix binds to the C-terminal extension and provide insights into stabilization of PINK1 on the core TOM complex.

Keywords:  PINK1; Parkin; Parkinson; TOM; X-ray crystallography; kinase; mass spectrometry; mitochondria; phosphorylation; ubiquitin

DOI:  https://doi.org/10.1016/j.molcel.2021.11.012
Nat Commun. 2021 Dec 06. 12(1): 7041

Glucocorticoid receptor regulates PD-L1 and MHC-I in pancreatic cancer cells to promote immune evasion and immunotherapy resistance.

Yalan Deng, Xianghou Xia, Yang Zhao, Zilong Zhao, Consuelo Martinez, Wenjuan Yin, Jun Yao, Qinglei Hang, Weiche Wu, Jie Zhang, Yang Yu, Weiya Xia, Fan Yao, Di Zhao, Yutong Sun, Haoqiang Ying, Mien-Chie Hung, Li Ma.

Despite unprecedented responses of some cancers to immune checkpoint blockade (ICB) therapies, the application of checkpoint inhibitors in pancreatic cancer has been unsuccessful. Glucocorticoids and glucocorticoid receptor (GR) signaling are long thought to suppress immunity by acting on immune cells. Here we demonstrate a previously undescribed tumor cell-intrinsic role for GR in activating PD-L1 expression and repressing the major histocompatibility complex class I (MHC-I) expression in pancreatic ductal adenocarcinoma (PDAC) cells through transcriptional regulation. In mouse models of PDAC, either tumor cell-specific depletion or pharmacologic inhibition of GR leads to PD-L1 downregulation and MHC-I upregulation in tumor cells, which in turn promotes the infiltration and activity of cytotoxic T cells, enhances anti-tumor immunity, and overcomes resistance to ICB therapy. In patients with PDAC, GR expression correlates with high PD-L1 expression, low MHC-I expression, and poor survival. Our results reveal GR signaling in cancer cells as a tumor-intrinsic mechanism of immunosuppression and suggest that therapeutic targeting of GR is a promising way to sensitize pancreatic cancer to immunotherapy.

DOI: https://doi.org/10.1038/s41467-021-27349-7
Cell Metab. 2021 Dec 07. pii: S1550-4131(21)00538-6. [Epub ahead of print]33(12): 2415-2427.e6

Coenzyme A fuels T cell anti-tumor immunity.

Michael St Paul, Samuel D Saibil, SeongJun Han, Kavita Israni-Winger, Scott C Lien, Rob C Laister, Azin Sayad, Susanne Penny, Rodabe N Amaria, Lauren E Haydu, Carlos R Garcia-Batres, Meghan Kates, David T Mulder, Céline Robert-Tissot, Matthew J Gold, Charles W Tran, Alisha R Elford, Linh T Nguyen, Trevor J Pugh, Devanand M Pinto, Jennifer A Wargo, Pamela S Ohashi.

  Metabolic programming is intricately linked to the anti-tumor properties of T cells. To study the metabolic pathways associated with increased anti-tumor T cell function, we utilized a metabolomics approach to characterize three different CD8+ T cell subsets with varying degrees of anti-tumor activity in murine models, of which IL-22-producing Tc22 cells displayed the most robust anti-tumor activity. Tc22s demonstrated upregulation of the pantothenate/coenzyme A (CoA) pathway and a requirement for oxidative phosphorylation (OXPHOS) for differentiation. Exogenous administration of CoA reprogrammed T cells to increase OXPHOS and adopt the CD8+ Tc22 phenotype independent of polarizing conditions via the transcription factors HIF-1α and the aryl hydrocarbon receptor (AhR). In murine tumor models, treatment of mice with the CoA precursor pantothenate enhanced the efficacy of anti-PDL1 antibody therapy. In patients with melanoma, pre-treatment plasma pantothenic acid levels were positively correlated with the response to anti-PD1 therapy. Collectively, our data demonstrate that pantothenate and its metabolite CoA drive T cell polarization, bioenergetics, and anti-tumor immunity.

Keywords:  CD8(+) T cells; IL-22; Tc1; Tc17; Tc22; anti-PD1; coenzyme A; immunometabolism; immunotherapy; pantothenate

DOI:  https://doi.org/10.1016/j.cmet.2021.11.010
Cancers (Basel). 2021 Nov 30. pii: 6036. [Epub ahead of print]13(23):

Proteomic Analysis Identifies NDUFS1 and ATP5O as Novel Markers for Survival Outcome in Prostate Cancer.

Robert Wiebringhaus, Matteo Pecoraro, Heidi A Neubauer, Karolína Trachtová, Bettina Trimmel, Maritta Wieselberg, Jan Pencik, Gerda Egger, Christoph Krall, Richard Moriggl, Matthias Mann, Brigitte Hantusch, Lukas Kenner.

  We aimed to identify novel markers for aggressive prostate cancer in a STAT3-low proteomics-derived dataset of mitochondrial proteins by immunohistochemical analysis and correlation with transcriptomic data and biochemical recurrence in a STAT3 independent PCa cohort. Formalin-fixed paraffin-embedded tissue (FFPE) sample selection for proteomic analysis and tissue-microarray (TMA) generation was conducted from a cohort of PCa patients. Retrospective data analysis was performed with the same cohort. 153 proteins differentially expressed between STAT3-low and STAT3-high samples were identified. Out of these, 46 proteins were associated with mitochondrial processes including oxidative phosphorylation (OXPHOS), and 45 proteins were upregulated, including NDUFS1/ATP5O. In a STAT3 independent PCa cohort, high expression of NDUFS1/ATP5O was confirmed by immunocytochemistry (IHC) and was significantly associated with earlier biochemical recurrence (BCR). mRNA expression levels for these two genes were significantly higher in intra-epithelial neoplasia and in PCa compared to benign prostate glands. NDUFS1/ATP5O levels are increased both at the mRNA and protein level in aggressive PCa. Our results provide evidence that NDUFS1/ATP5O could be used to identify high-risk PCa patients.

Keywords:  ATP5O; FFPE-proteomics; NDUFS1; OXPHOS; STAT3; prostate cancer; transcriptomics

DOI:  https://doi.org/10.3390/cancers13236036
Nat Commun. 2021 Dec 07. 12(1): 7113

Spatial-proteomics reveals phospho-signaling dynamics at subcellular resolution.

Ana Martinez-Val, Dorte B Bekker-Jensen, Sophia Steigerwald, Claire Koenig, Ole Østergaard, Adi Mehta, Trung Tran, Krzysztof Sikorski, Estefanía Torres-Vega, Ewa Kwasniewicz, Sólveig Hlín Brynjólfsdóttir, Lisa B Frankel, Rasmus Kjøbsted, Nicolai Krogh, Alicia Lundby, Simon Bekker-Jensen, Fridtjof Lund-Johansen, Jesper V Olsen.

Dynamic change in subcellular localization of signaling proteins is a general concept that eukaryotic cells evolved for eliciting a coordinated response to stimuli. Mass spectrometry-based proteomics in combination with subcellular fractionation can provide comprehensive maps of spatio-temporal regulation of protein networks in cells, but involves laborious workflows that does not cover the phospho-proteome level. Here we present a high-throughput workflow based on sequential cell fractionation to profile the global proteome and phospho-proteome dynamics across six distinct subcellular fractions. We benchmark the workflow by studying spatio-temporal EGFR phospho-signaling dynamics in vitro in HeLa cells and in vivo in mouse tissues. Finally, we investigate the spatio-temporal stress signaling, revealing cellular relocation of ribosomal proteins in response to hypertonicity and muscle contraction. Proteomics data generated in this study can be explored through https://SpatialProteoDynamics.github.io .

DOI: https://doi.org/10.1038/s41467-021-27398-y
Nat Commun. 2021 Dec 08. 12(1): 7130

Free fatty-acid transport via CD36 drives β-oxidation-mediated hematopoietic stem cell response to infection.

Jayna J Mistry, Charlotte Hellmich, Jamie A Moore, Aisha Jibril, Iain Macaulay, Mar Moreno-Gonzalez, Federica Di Palma, Naiara Beraza, Kristian M Bowles, Stuart A Rushworth.

Acute infection is known to induce rapid expansion of hematopoietic stem cells (HSCs), but the mechanisms supporting this expansion remain incomplete. Using mouse models, we show that inducible CD36 is required for free fatty acid uptake by HSCs during acute infection, allowing the metabolic transition from glycolysis towards β-oxidation. Mechanistically, high CD36 levels promote FFA uptake, which enables CPT1A to transport fatty acyl chains from the cytosol into the mitochondria. Without CD36-mediated FFA uptake, the HSCs are unable to enter the cell cycle, subsequently enhancing mortality in response to bacterial infection. These findings enhance our understanding of HSC metabolism in the bone marrow microenvironment, which supports the expansion of HSCs during pathogenic challenge.

DOI: https://doi.org/10.1038/s41467-021-27460-9
J Clin Med. 2021 Nov 29. pii: 5606. [Epub ahead of print]10(23):

Combined Inhibition of Bcl2 and Bcr-Abl1 Exercises Anti-Leukemia Activity but Does Not Eradicate the Primitive Leukemic Cells.

Michele Massimino, Paolo Vigneri, Stefania Stella, Elena Tirrò, Maria Stella Pennisi, Laura Nunziatina Parrinello, Calogero Vetro, Livia Manzella, Fabio Stagno, Francesco Di Raimondo.

   BACKGROUND: The management of Philadelphia Chromosome-positive (Ph+) hematological malignancies is strictly correlated to the use of BCR-ABL1 tyrosine kinase inhibitors (TKIs). However, these drugs do not induce leukemic stem cells death and their persistence may generate a disease relapse. Published reports indicated that Venetoclax, a selective BCL2 inhibitor, could be effective in Ph+ diseases, as BCL2 anti-apoptotic activity is modulated by BCR-ABL1 kinase. We, therefore, investigated if BCL2 inhibition, alone or combined with Nilotinib, a BCR-ABL1 inhibitor, affects the primitive and committed Ph+ cells survival.
METHODS: We used Ph+ cells isolated from leukemic patients at diagnosis. To estimate the therapeutic efficacy of BCL2 and BCR-ABL1 inhibition we employed long-term culture, proliferation and apoptosis assay. Immunoblot was used to evaluate the ability of treatment to interfere with the down-stream targets of BCR-ABL1.
RESULTS: Blocking BCL2, we observed reduced proliferation and clonogenic potential of CML CD34-positive cells and this cytotoxicity was improved by combination with BCR-ABL1 inhibitor. However, BCL2 inhibition, alone or in combination regiment with BCR-ABL1 inhibitor, did not reduce the self-renewal of primitive leukemic cells, while strongly induced cell death on primary Ph+ Acute Lymphoblastic Leukemia (ALL).
CONCLUSION: Our results suggest that primitive CML leukemic cells are not dependent on BCL2 for their persistence and support that committed CML and Ph + ALL cells are dependent by BCL2 and BCR-ABL1 cooperation for their survival. The antileukemic activity of BCL2 and BCR-ABL1 dual targeting may be a useful therapeutic strategy for Ph+ ALL patients.

Keywords:  ALL; BCL2; BCR-ABL1; CML; LTC-IC; Nilotinib; Venetoclax; stem cell

DOI:  https://doi.org/10.3390/jcm10235606