bims-mitdyn Biomed News
on Mitochondrial dynamics: mechanisms
Issue of 2022–06–26
nineteen papers selected by
Edmond Chan, Queen’s University, School of Medicine
  1. Tumor necrosis factor induces pathogenic mitochondrial ROS in tuberculosis through reverse electron transport.
  2. Cancer cells depend on environmental lipids for proliferation when electron acceptors are limited.
  3. Adaptive stimulation of macropinocytosis overcomes aspartate limitation in cancer cells under hypoxia.
  4. Structural basis of Tom20 and Tom22 cytosolic domains as the human TOM complex receptors.
  5. Structure of human NADK2 reveals atypical assembly and regulation of NAD kinases from animal mitochondria.
  6. Mitochondrial calcium uniporter promotes phagocytosis-dependent activation of the NLRP3 inflammasome.
  7. A ferroptosis defense mechanism mediated by glycerol-3-phosphate dehydrogenase 2 in mitochondria.
  8. Substrate binding in the mitochondrial ADP/ATP carrier is a step-wise process guiding the structural changes in the transport cycle.
  9. CEND1 deficiency induces mitochondrial dysfunction and cognitive impairment in Alzheimer's disease.
  10. Tollip negatively regulates mitophagy by promoting the mitochondrial processing and cytoplasmic release of PINK1.
  11. Inhibition of mitoNEET induces Pink1-Parkin-mediated mitophagy.
  12. Shaping fuel utilization by mitochondria.
  13. TFAM downregulation promotes autophagy and ESCC survival through mtDNA stress-mediated STING pathway.
  14. Most mitochondrial dGTP is tightly bound to respiratory complex I through the NDUFA10 subunit.
  15. Protocol for engineering and validating a synthetic mitochondrial intermembrane bridge in mammalian cells.
  16. The Use of Seahorse XF Assays to Interrogate Real-Time Energy Metabolism in Cancer Cell Lines.
  17. ncRNA regulation of mitochondrial transcription.
  18. Is Drp1 sufficient to catalyze membrane fission?
  19. Blood flow meets mitophagy.
  1. Science. 2022 Jun 24. 376(6600): eabh2841
    Tumor necrosis factor induces pathogenic mitochondrial ROS in tuberculosis through reverse electron transport.
    Francisco J Roca, Laura J Whitworth, Hiran A Prag, Michael P Murphy, Lalita Ramakrishnan.
      Tumor necrosis factor (TNF) is a critical host resistance factor against tuberculosis. However, excess TNF produces susceptibility by increasing mitochondrial reactive oxygen species (mROS), which initiate a signaling cascade to cause pathogenic necrosis of mycobacterium-infected macrophages. In zebrafish, we identified the mechanism of TNF-induced mROS in tuberculosis. Excess TNF in mycobacterium-infected macrophages elevates mROS production by reverse electron transport (RET) through complex I. TNF-activated cellular glutamine uptake leads to an increased concentration of succinate, a Krebs cycle intermediate. Oxidation of this elevated succinate by complex II drives RET, thereby generating the mROS superoxide at complex I. The complex I inhibitor metformin, a widely used antidiabetic drug, prevents TNF-induced mROS and necrosis of Mycobacterium tuberculosis-infected zebrafish and human macrophages; metformin may therefore be useful in tuberculosis therapy.
    DOI:  https://doi.org/10.1126/science.abh2841
  2. Nat Metab. 2022 Jun 23.
    Cancer cells depend on environmental lipids for proliferation when electron acceptors are limited.
    Zhaoqi Li, Brian W Ji, Purushottam D Dixit, Konstantine Tchourine, Evan C Lien, Aaron M Hosios, Keene L Abbott, Justine C Rutter, Anna M Westermark, Elizabeth F Gorodetsky, Lucas B Sullivan, Matthew G Vander Heiden, Dennis Vitkup.
      Production of oxidized biomass, which requires regeneration of the cofactor NAD+, can be a proliferation bottleneck that is influenced by environmental conditions. However, a comprehensive quantitative understanding of metabolic processes that may be affected by NAD+ deficiency is currently missing. Here, we show that de novo lipid biosynthesis can impose a substantial NAD+ consumption cost in proliferating cancer cells. When electron acceptors are limited, environmental lipids become crucial for proliferation because NAD+ is required to generate precursors for fatty acid biosynthesis. We find that both oxidative and even net reductive pathways for lipogenic citrate synthesis are gated by reactions that depend on NAD+ availability. We also show that access to acetate can relieve lipid auxotrophy by bypassing the NAD+ consuming reactions. Gene expression analysis demonstrates that lipid biosynthesis strongly anti-correlates with expression of hypoxia markers across tumor types. Overall, our results define a requirement for oxidative metabolism to support biosynthetic reactions and provide a mechanistic explanation for cancer cell dependence on lipid uptake in electron acceptor-limited conditions, such as hypoxia.
    DOI:  https://doi.org/10.1038/s42255-022-00588-8
  3. Nat Metab. 2022 Jun 20.
    Adaptive stimulation of macropinocytosis overcomes aspartate limitation in cancer cells under hypoxia.
    Javier Garcia-Bermudez, Michael A Badgley, Sheela Prasad, Lou Baudrier, Yuyang Liu, Konnor La, Mariluz Soula, Robert T Williams, Norihiro Yamaguchi, Rosa F Hwang, Laura J Taylor, Elisa de Stanchina, Bety Rostandy, Hanan Alwaseem, Henrik Molina, Dafna Bar-Sagi, Kıvanç Birsoy.
      Stress-adaptive mechanisms enable tumour cells to overcome metabolic constraints under nutrient and oxygen shortage. Aspartate is an endogenous metabolic limitation under hypoxic conditions, but the nature of the adaptive mechanisms that contribute to aspartate availability and hypoxic tumour growth are poorly understood. Here we identify GOT2-catalysed mitochondrial aspartate synthesis as an essential metabolic dependency for the proliferation of pancreatic tumour cells under hypoxic culture conditions. In contrast, GOT2-catalysed aspartate synthesis is dispensable for pancreatic tumour formation in vivo. The dependence of pancreatic tumour cells on aspartate synthesis is bypassed in part by a hypoxia-induced potentiation of extracellular protein scavenging via macropinocytosis. This effect is mutant KRAS dependent, and is mediated by hypoxia-inducible factor 1 (HIF1A) and its canonical target carbonic anhydrase-9 (CA9). Our findings reveal high plasticity of aspartate metabolism and define an adaptive regulatory role for macropinocytosis by which mutant KRAS tumours can overcome nutrient deprivation under hypoxic conditions.
    DOI:  https://doi.org/10.1038/s42255-022-00583-z
  4. Proc Natl Acad Sci U S A. 2022 Jun 28. 119(26): e2200158119
    Structural basis of Tom20 and Tom22 cytosolic domains as the human TOM complex receptors.
    Jiayue Su, Desheng Liu, Fan Yang, Mei-Qing Zuo, Chang Li, Meng-Qiu Dong, Shan Sun, Sen-Fang Sui.
      Mitochondrial preproteins synthesized in cytosol are imported into mitochondria by a multisubunit translocase of the outer membrane (TOM) complex. Functioned as the receptor, the TOM complex components, Tom 20, Tom22, and Tom70, recognize the presequence and further guide the protein translocation. Their deficiency has been linked with neurodegenerative diseases and cardiac pathology. Although several structures of the TOM complex have been reported by cryoelectron microscopy (cryo-EM), how Tom22 and Tom20 function as TOM receptors remains elusive. Here we determined the structure of TOM core complex at 2.53 Å and captured the structure of the TOM complex containing Tom22 and Tom20 cytosolic domains at 3.74 Å. Structural analysis indicates that Tom20 and Tom22 share a similar three-helix bundle structural feature in the cytosolic domain. Further structure-guided biochemical analysis reveals that the Tom22 cytosolic domain is responsible for binding to the presequence, and the helix H1 is critical for this binding. Altogether, our results provide insights into the functional mechanism of the TOM complex recognizing and transferring preproteins across the mitochondrial membrane.
    Keywords:  TOM complex; Tom20; Tom22; cryo-EM; mitochondria
    DOI:  https://doi.org/10.1073/pnas.2200158119
  5. Proc Natl Acad Sci U S A. 2022 Jun 28. 119(26): e2200923119
    Structure of human NADK2 reveals atypical assembly and regulation of NAD kinases from animal mitochondria.
    Jin Du, Michael Estrella, Kristina Solorio-Kirpichyan, Philip D Jeffrey, Alexei Korennykh.
      All kingdoms of life produce essential nicotinamide dinucleotide NADP(H) using NAD kinases (NADKs). A panel of published NADK structures from bacteria, eukaryotic cytosol, and yeast mitochondria revealed similar tetrameric enzymes. Here, we present the 2.8-Å structure of the human mitochondrial kinase NADK2 with a bound substrate, which is an exception to this uniformity, diverging both structurally and biochemically from NADKs. We show that NADK2 harbors a unique tetramer disruptor/dimerization element, which is conserved in mitochondrial kinases of animals (EMKA) and absent from other NADKs. EMKA stabilizes the NADK2 dimer but prevents further NADK2 oligomerization by blocking the tetramerization interface. This structural change bears functional consequences and alters the activation mechanism of the enzyme. Whereas tetrameric NADKs undergo cooperative activation via oligomerization, NADK2 is a constitutively active noncooperative dimer. Thus, our data point to a unique regulation of NADP(H) synthesis in animal mitochondria achieved via structural adaptation of the NADK2 kinase.
    Keywords:  NADK; NADK2; cooperative; dimer; structure
    DOI:  https://doi.org/10.1073/pnas.2200923119
  6. Proc Natl Acad Sci U S A. 2022 Jun 28. 119(26): e2123247119
    Mitochondrial calcium uniporter promotes phagocytosis-dependent activation of the NLRP3 inflammasome.
    Hong Dong, Bao Zhao, Jianwen Chen, Zihao Liu, Xinghui Li, Lupeng Li, Haitao Wen.
      Mitochondria, a highly metabolically active organelle, have been shown to play an essential role in regulating innate immune function. Mitochondrial Ca2+ uptake via the mitochondrial Ca2+ uniporter (MCU) is an essential process regulating mitochondrial metabolism by targeting key enzymes involved in the tricarboxylic acid cycle (TCA). Accumulative evidence suggests MCU-dependent mitochondrial Ca2+ signaling may bridge the metabolic reprogramming and regulation of immune cell function. However, the mechanism by which MCU regulates inflammation and its related disease remains elusive. Here we report a critical role of MCU in promoting phagocytosis-dependent activation of NLRP3 (nucleotide-binding domain, leucine-rich repeat containing family, pyrin domain-containing 3) inflammasome by inhibiting phagolysosomal membrane repair. Myeloid deletion of MCU (McuΔmye) resulted in an attenuated phagolysosomal rupture, leading to decreased caspase-1 cleavage and interleukin (IL)-1β release, in response to silica or alum challenge. In contrast, other inflammasome agonists such as adenosine triphosphate (ATP), nigericin, poly(dA:dT), and flagellin induced normal IL-1β release in McuΔmye macrophages. Mechanistically, we demonstrated that decreased NLRP3 inflammasome activation in McuΔmye macrophages was caused by improved phagolysosomal membrane repair mediated by ESCRT (endosomal sorting complex required for transport)-III complex. Furthermore, McuΔmye mice showed a pronounced decrease in immune cell recruitment and IL-1β production in alum-induced peritonitis, a typical IL-1-dependent inflammation model. In sum, our results identify a function of MCU in promoting phagocytosis-dependent NLRP3 inflammatory response via an ESCRT-mediated phagolysosomal membrane repair mechanism.
    Keywords:  ESCRT; MCU; inflammasome; phagosome
    DOI:  https://doi.org/10.1073/pnas.2123247119
  7. Proc Natl Acad Sci U S A. 2022 Jun 28. 119(26): e2121987119
    A ferroptosis defense mechanism mediated by glycerol-3-phosphate dehydrogenase 2 in mitochondria.
    Shiqi Wu, Chao Mao, Lavanya Kondiparthi, Masha V Poyurovsky, Kellen Olszewski, Boyi Gan.
      Mechanisms of defense against ferroptosis (an iron-dependent form of cell death induced by lipid peroxidation) in cellular organelles remain poorly understood, hindering our ability to target ferroptosis in disease treatment. In this study, metabolomic analyses revealed that treatment of cancer cells with glutathione peroxidase 4 (GPX4) inhibitors results in intracellular glycerol-3-phosphate (G3P) depletion. We further showed that supplementation of cancer cells with G3P attenuates ferroptosis induced by GPX4 inhibitors in a G3P dehydrogenase 2 (GPD2)-dependent manner; GPD2 deletion sensitizes cancer cells to GPX4 inhibition-induced mitochondrial lipid peroxidation and ferroptosis, and combined deletion of GPX4 and GPD2 synergistically suppresses tumor growth by inducing ferroptosis in vivo. Mechanistically, inner mitochondrial membrane-localized GPD2 couples G3P oxidation with ubiquinone reduction to ubiquinol, which acts as a radical-trapping antioxidant to suppress ferroptosis in mitochondria. Taken together, these results reveal that GPD2 participates in ferroptosis defense in mitochondria by generating ubiquinol.
    Keywords:  GPD2; cell death; ferroptosis; lipid peroxidation; mitochondria
    DOI:  https://doi.org/10.1073/pnas.2121987119
  8. Nat Commun. 2022 Jun 23. 13(1): 3585
    Substrate binding in the mitochondrial ADP/ATP carrier is a step-wise process guiding the structural changes in the transport cycle.
    Vasiliki Mavridou, Martin S King, Sotiria Tavoulari, Jonathan J Ruprecht, Shane M Palmer, Edmund R S Kunji.
      Mitochondrial ADP/ATP carriers import ADP into the mitochondrial matrix and export ATP to the cytosol to fuel cellular processes. Structures of the inhibited cytoplasmic- and matrix-open states have confirmed an alternating access transport mechanism, but the molecular details of substrate binding remain unresolved. Here, we evaluate the role of the solvent-exposed residues of the translocation pathway in the process of substrate binding. We identify the main binding site, comprising three positively charged and a set of aliphatic and aromatic residues, which bind ADP and ATP in both states. Additionally, there are two pairs of asparagine/arginine residues on opposite sides of this site that are involved in substrate binding in a state-dependent manner. Thus, the substrates are directed through a series of binding poses, inducing the conformational changes of the carrier that lead to their translocation. The properties of this site explain the electrogenic and reversible nature of adenine nucleotide transport.
    DOI:  https://doi.org/10.1038/s41467-022-31366-5
  9. Cell Death Differ. 2022 Jun 22.
    CEND1 deficiency induces mitochondrial dysfunction and cognitive impairment in Alzheimer's disease.
    Wenting Xie, Dong Guo, Jieyin Li, Lei Yue, Qi Kang, Guimiao Chen, Tingwen Zhou, Han Wang, Kai Zhuang, Lige Leng, Huifang Li, Zhenyi Chen, Weiwei Gao, Jie Zhang.
      Alzheimer's disease (AD) is the most common form of neurodegenerative disease featured with memory loss and cognitive function impairments. Chronic mitochondrial stress is a vital pathogenic factor for AD and finally leads to massive neuronal death. However, the underlying mechanism is unclear. By proteomic analysis, we identified a new mitochondrial protein, cell-cycle exit and neuronal differentiation 1 (CEND1), which was decreased significantly in the brain of 5xFAD mice. CEND1 is a neuronal specific protein and locates in the presynaptic mitochondria. Depletion of CEND1 leads to increased mitochondrial fission mediated by upregulation of dynamin related protein 1 (Drp1), resulting in abnormal mitochondrial functions. CEND1 deficiency leads to cognitive impairments in mice. Overexpression of CEND1 in the hippocampus of 5xFAD mice rescued cognitive deficits. Moreover, we identified that CDK5/p25 interacted with and phosphorylated CEND1 which promoted its degradation. Our study provides new mechanistic insights in mitochondrial function regulations by CEND1 in Alzheimer's disease.
    DOI:  https://doi.org/10.1038/s41418-022-01027-7
  10. BMB Rep. 2022 Jun 21. pii: 5642. [Epub ahead of print]
    Tollip negatively regulates mitophagy by promoting the mitochondrial processing and cytoplasmic release of PINK1.
    Woo Hyun Shin, Kwang Chul Chung.
      PTEN-induced putative kinase 1 (PINK1) is a serine/threonine kinase that phosphorylates several substrates and exerts neuroprotective effects against stress-induced apoptotic cell death. Mutations in PINK1 have been linked to autosomal recessive forms of Parkinson's disease (PD). Mitophagy is a type of autophagy that selectively promotes mitochondrial turnover and prevents the accumulation of dysfunctional mitochondria to maintain cellular homeostasis. Toll-interacting protein (Tollip) was initially identified as a negative regulator of IL-1β receptor signaling, suppressing inflammatory TLR signaling cascades. Recently, Tollip has been reported to play a role in autophagy and is implicated in neurodegeneration. In this study, we determined whether Tollip was functionally linked to PINK1-mediated mitophagy. Our results demonstrated that Tollip promoted the mitochondrial processing of PINK1 and altered the localization of PINK1, predominantly to the cytosol. This action was attributed to increased binding of PINK1 to mitochondrial processing peptidase β (MPPβ) and the subsequent increase in MPPβ-mediated mitochondrial PINK1 cleavage. Furthermore, Tollip suppressed mitophagy following carbonyl cyanide m-chlorophenylhydrazone-induced mitochondrial dysfunction. These findings suggest that Tollip inhibits mitophagy via the PINK1/parkin pathway upon mitochondrial damage, leading to the blockade of PINK1-mediated neuroprotection.
  11. BMB Rep. 2022 Jun 21. pii: 5590. [Epub ahead of print]
    Inhibition of mitoNEET induces Pink1-Parkin-mediated mitophagy.
    Seunghee Lee, Sangguk Lee, Seon-Jin Lee, Su Wol Chung.
      MitoNEET, a mitochondrial outer membrane protein containing the Asn-Glu-Glu-Thr (NEET) sequence, controls the formation of intermitochondrial junctions and confers autophagy resistance. Moreover, mitoNEET as a mitochondrial substrate undergoes ubiquitination by activated Parkin during the initiation of mitophagy. Therefore, mitoNEET is linked to the regulation of autophagy and mitophagy. Mitophagy is the selective removal of the damaged or unnecessary mitochondria, which is crucial to sustaining mitochondrial quality control. In numerous human diseases, the accumulation of damaged mitochondria by impaired mitophagy has been observed. However, the therapeutic strategy targeting of mitoNEET as a mitophagy-enhancing mediator requires further research. Herein, we confirmed that mitophagy is indeed activated by mitoNEET inhibition. CCCP (carbonyl cyanide m-chlorophenyl hydrazone), which leads to mitochondrial depolarization, induces mitochondrial dysfunction and superoxide production. This, in turn, contributes to the induction of mitophagy; mitoNEET protein levels were initially increased before an increase in LC3-Ⅱ protein following CCCP treatment. Pharmacological inhibition of mitoNEET using mitoNEET Ligand-1 (NL-1) promoted accumulation of Pink1 and Parkin, which are mitophagy-associated proteins, and activation of mitochondria-lysosome crosstalk, in comparison to CCCP alone. Inhibition of mitoNEET using NL-1, or mitoNEET shRNA transfected into RAW264.7 cells, abrogated CCCP-induced ROS and mitochondrial cell death; additionally, it activated the expression of PGC-1α and SOD2, regulators of oxidative metabolism. In particular, the increase in PGC-1α, which is a major regulator of mitochondrial biogenesis, promotes mitochondrial quality control. These results indicated that mitoNEET is a potential therapeutic target in numerous human diseases to enhance mitophagy and protect cells by maintaining a network of healthy mitochondria.
  12. Curr Biol. 2022 Jun 20. pii: S0960-9822(22)00765-5. [Epub ahead of print]32(12): R618-R623
    Shaping fuel utilization by mitochondria.
    Lukas Alan, Luca Scorrano.
      Mitochondria are central to cellular metabolism. They provide intermediate metabolites that are used in biosynthetic pathways and they process diet-derived nutrients into the energy-rich compound ATP. Mitochondrial ATP biosynthesis is a marvel of thermodynamic efficiency. Via the tricarboxylic acid cycle (TCA) and fatty acid β-oxidation, mitochondria extract electrons from dietary carbon compounds and pass them to nucleotides that ultimately deliver them to the respiratory chain complexes located in invaginations in the inner mitochondrial membrane (IMM) known as cristae. The respiratory chain complexes donate electrons in stepwise redox reactions to molecular oxygen and, with the exception of complex II, use the liberated energy to pump protons across the proton-impermeable IMM, generating a proton electrochemical gradient. This gradient is then utilized by the ATP synthase, which, in a rotary mechanism, catalyzes the formation of the high-energy γ-phosphate chemical bond between ADP and inorganic phosphate. The conversion of the chemical energy of carbon compounds into a physical, vectorial form of energy (the electrochemical gradient) maximizes the yield of the ATP biosynthetic process and is perhaps one of the foundations of life as we know it.
    DOI:  https://doi.org/10.1016/j.cub.2022.05.006
  13. Oncogene. 2022 Jun 24.
    TFAM downregulation promotes autophagy and ESCC survival through mtDNA stress-mediated STING pathway.
    Yujia Li, Qi Yang, Hui Chen, Xiaotian Yang, Jingru Han, Xiaojuan Yao, Xiajie Wei, Jiaoyang Si, Huanling Yao, Hongliang Liu, Lixin Wan, Hushan Yang, Yanming Wang, Dengke Bao.
      The dynamics of mitochondrial biogenesis regulation is critical in maintaining cellular homeostasis for immune regulation and tumor prevention. Here, we report that mitochondrial biogenesis disruption through TFAM reduction significantly impairs mitochondrial function, induces autophagy, and promotes esophageal squamous cell carcinoma (ESCC) growth. We found that TFAM protein reduction promotes mitochondrial DNA (mtDNA) release into the cytosol, induces cytosolic mtDNA stress, subsequently activates the cGAS-STING signaling pathway, thereby stimulating autophagy and ESCC growth. STING depletion or mtDNA degradation by DNase I abrogates mtDNA stress response, attenuates autophagy, and decreases the growth of TFAM depleted cells. In addition, autophagy inhibitor also ameliorates mitochondrial dysfunction-induced activation of the cGAS-STING signaling pathway and ESCC growth. In conclusion, our results indicate that mtDNA stress induced by mitochondria biogenesis perturbation activates the cGAS-STING pathway and autophagy to promote ESCC growth, revealing an underappreciated therapeutic strategy for ESCC.
    DOI:  https://doi.org/10.1038/s41388-022-02365-z
  14. Commun Biol. 2022 Jun 23. 5(1): 620
    Most mitochondrial dGTP is tightly bound to respiratory complex I through the NDUFA10 subunit.
    David Molina-Granada, Emiliano González-Vioque, Marris G Dibley, Raquel Cabrera-Pérez, Antoni Vallbona-Garcia, Javier Torres-Torronteras, Leonid A Sazanov, Michael T Ryan, Yolanda Cámara, Ramon Martí.
      Imbalanced mitochondrial dNTP pools are known players in the pathogenesis of multiple human diseases. Here we show that, even under physiological conditions, dGTP is largely overrepresented among other dNTPs in mitochondria of mouse tissues and human cultured cells. In addition, a vast majority of mitochondrial dGTP is tightly bound to NDUFA10, an accessory subunit of complex I of the mitochondrial respiratory chain. NDUFA10 shares a deoxyribonucleoside kinase (dNK) domain with deoxyribonucleoside kinases in the nucleotide salvage pathway, though no specific function beyond stabilizing the complex I holoenzyme has been described for this subunit. We mutated the dNK domain of NDUFA10 in human HEK-293T cells while preserving complex I assembly and activity. The NDUFA10E160A/R161A shows reduced dGTP binding capacity in vitro and leads to a 50% reduction in mitochondrial dGTP content, proving that most dGTP is directly bound to the dNK domain of NDUFA10. This interaction may represent a hitherto unknown mechanism regulating mitochondrial dNTP availability and linking oxidative metabolism to DNA maintenance.
    DOI:  https://doi.org/10.1038/s42003-022-03568-6
  15. STAR Protoc. 2022 Sep 16. 3(3): 101454
    Protocol for engineering and validating a synthetic mitochondrial intermembrane bridge in mammalian cells.
    Gunjan Purohit, Martonio Ponte Viana, Oleh Khalimonchuk.
      Membrane contact sites are recognized as critical means of intercompartmental communication. Here, we describe a protocol for engineering and validating a synthetic bridge between the inner and outer mitochondrial membranes to support functioning of the endogenous mitochondrial contact site and cristae organizing system (MICOS). A chimeric protein, MitoT, is stably expressed in cultured mammalian cells to bridge the mitochondrial membranes. This approach can be a valuable tool to study the function of the MICOS complex and associated proteins. For complete details on the use and execution of this protocol, please refer to Viana et al. (2021).
    Keywords:  Biotechnology and bioengineering; Cell Biology; Cell Membrane; Cell culture; Cell isolation; Flow Cytometry/Mass Cytometry; Metabolism; Microscopy; Molecular Biology
    DOI:  https://doi.org/10.1016/j.xpro.2022.101454
  16. Methods Mol Biol. 2022 ;2508 225-234
    The Use of Seahorse XF Assays to Interrogate Real-Time Energy Metabolism in Cancer Cell Lines.
    Jenna K Caines, David A Barnes, Mark D Berry.
      The propensity of cancer cells to preferentially undergo anaerobic metabolism despite oxygen being abundant is referred to as the Warburg effect. Measuring cellular metabolism is therefore central to understanding the cellular physiology of cancer cells. The Seahorse XFe Analyzer series allows real-time measurement of cellular metabolism. In the basic assay, two parameters, the oxygen consumption rate (OCR) and the extracellular acidification rate (ECAR), are used to determine real-time changes in the energy needs of live cells: OCR provides a measure of aerobic mitochondrial respiration; ECAR gives a measure of anaerobic glycolysis. Through the use of various respiration inhibitors, the Seahorse assay allows baseline respiration rate and total aerobic and anaerobic ATP production to be determined under a variety of experimental conditions. Here we describe the protocol for completing the Seahorse Real-Time ATP Rate Assay for adherent and suspension cancer cell lines. Depending on individual experimental results, more refined subsequent assays can then be performed to specifically determine, for example, the ability to utilize different substrates by the cell lines in the presence and absence of pharmacological and/or genetic interventions.
    Keywords:  ATP; Cancer cell metabolism; Energy metabolism; Glycolysis; Mitochondria; Seahorse
    DOI:  https://doi.org/10.1007/978-1-0716-2376-3_17
  17. Nat Rev Mol Cell Biol. 2022 Jun 21.
    ncRNA regulation of mitochondrial transcription.
    Eytan Zlotorynski.
      
    DOI:  https://doi.org/10.1038/s41580-022-00509-3
  18. Proc Natl Acad Sci U S A. 2022 Jul 05. 119(27): e2201709119
    Is Drp1 sufficient to catalyze membrane fission?
    Krishnendu Roy, Thomas J Pucadyil.
      
    DOI:  https://doi.org/10.1073/pnas.2201709119
  19. J Cell Biol. 2022 Jul 04. pii: e202206033. [Epub ahead of print]221(7):
    Blood flow meets mitophagy.
    Emir Bora Akmeriç, Holger Gerhardt.
      Since its discovery as a mechanosensitive transcription factor in endothelial networks, Klf2's varying expression levels under different blood flow patterns remained a mystery. In this study, Coon et al. (2022. J. Cell Biol.https://doi.org/10.1083/jcb.202109144) discover a connection between sustained laminar shear stress and mitochondrial flux that contributes to Klf2's transcriptional dynamics.
    DOI:  https://doi.org/10.1083/jcb.202206033
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