bims-celmim Biomed News
on Cellular and mitochondrial metabolism
Issue of 2024–03–24
25 papers selected by
Marc Segarra Mondejar, University of Cologne
  1. The phospholipids cardiolipin and phosphatidylethanolamine differentially regulate MDC biogenesis.
  2. Up-regulation of cholesterol synthesis by lysosomal defects requires a functional mitochondrial respiratory chain.
  3. SIRT4 loss reprograms intestinal nucleotide metabolism to support proliferation following perturbation of homeostasis.
  4. GALDAR: A genetically encoded galactose sensor for visualizing sugar metabolism in vivo.
  5. A Human Brain Map of Mitochondrial Respiratory Capacity and Diversity.
  6. Metabolic regulation of skeletal cell fate and function.
  7. Mitochondria-ER contact sites expand during mitosis.
  8. MCU-independent Ca2+ uptake mediates mitochondrial Ca2+ overload and necrotic cell death in a mouse model of Duchenne muscular dystrophy.
  9. Serine synthesis and catabolism in starved lung cancer and primary bronchial epithelial cells.
  10. Ferredoxin reduction by hydrogen with iron functions as an evolutionary precursor of flavin-based electron bifurcation.
  11. NMR and MS reveal characteristic metabolome atlas and optimize esophageal squamous cell carcinoma early detection.
  12. The roles of autophagy, ferroptosis and pyroptosis in the anti-ovarian cancer mechanism of harmine and their crosstalk.
  13. Knowledge-Based Therapeutics for Tricarboxylic Acid (TCA) Cycle-Deficient Cancers.
  14. Protocol for analyzing energy metabolic pathway dependency in human liver cancer cell lines.
  15. Insulin signalling regulates Pink1 mRNA localization via modulation of AMPK activity to support PINK1 function in neurons.
  16. Synchronized assembly of the oxidative phosphorylation system controls mitochondrial respiration in yeast.
  17. GLUT and HK: two primary and essential key players in Tumor glycolysis.
  18. Luminal transport through intact endoplasmic reticulum limits the magnitude of localized Ca2+ signals.
  19. A novel fluorescent probe for visualizing viscosity changes in lipid droplets during chemotherapy-induced ferroptosis.
  20. Fluorinated Analogues to the Pentuloses of the Pentose Phosphate Pathway.
  21. Activation of polyamine catabolism promotes glutamine metabolism and creates a targetable vulnerability in lung cancer.
  22. The AMPK-related kinase NUAK1 controls cortical axons branching by locally modulating mitochondrial metabolic functions.
  23. Tracing the Diverse Paths of One-Carbon Metabolism in Cancer and Beyond.
  24. A citric acid cycle-deficient Escherichia coli as an efficient chassis for aerobic fermentations.
  25. Metabolic impairments associated with type 2 diabetes mellitus and the potential effects of exercise therapy: An exploratory randomized trial based on untargeted metabolomics.
  1. J Cell Biol. 2024 May 06. pii: e202302069. [Epub ahead of print]223(5):
    The phospholipids cardiolipin and phosphatidylethanolamine differentially regulate MDC biogenesis.
    Tianyao Xiao, Alyssa M English, Zachary N Wilson, J Alan Maschek, James E Cox, Adam L Hughes.
      Cells utilize multiple mechanisms to maintain mitochondrial homeostasis. We recently characterized a pathway that remodels mitochondria in response to metabolic alterations and protein overload stress. This remodeling occurs via the formation of large membranous structures from the mitochondrial outer membrane called mitochondrial-derived compartments (MDCs), which are eventually released from mitochondria and degraded. Here, we conducted a microscopy-based screen in budding yeast to identify factors that regulate MDC formation. We found that two phospholipids, cardiolipin (CL) and phosphatidylethanolamine (PE), differentially regulate MDC biogenesis. CL depletion impairs MDC biogenesis, whereas blocking mitochondrial PE production leads to constitutive MDC formation. Additionally, in response to metabolic MDC activators, cellular and mitochondrial PE declines, and overexpressing mitochondrial PE synthesis enzymes suppress MDC biogenesis. Altogether, our data indicate a requirement for CL in MDC biogenesis and suggest that PE depletion may stimulate MDC formation downstream of MDC-inducing metabolic stress.
    DOI:  https://doi.org/10.1083/jcb.202302069
  2. bioRxiv. 2024 Mar 07. pii: 2024.03.06.583589. [Epub ahead of print]
    Up-regulation of cholesterol synthesis by lysosomal defects requires a functional mitochondrial respiratory chain.
    Francesco Agostini, Leonardo Pereyra, Justin Dale, King Faisal Yambire, Silvia Maglioni, Alfonso Schiavi, Natascia Ventura, Ira Milosevic, Nuno Raimundo.
      Mitochondria and lysosomes are two organelles that carry out both signaling and metabolic roles in the cells. Recent evidence has shown that mitochondria and lysosomes are dependent on one another, as primary defects in one cause secondary defects in the other. Nevertheless, the signaling consequences of primary mitochondrial malfunction and of primary lysosomal defects are not similar, despite in both cases there are impairments of mitochondria and of lysosomes. Here, we used RNA sequencing to obtain transcriptomes from cells with primary mitochondrial or lysosomal defects, to identify what are the global cellular consequences that are associated with malfunction of mitochondria or lysosomes. We used these data to determine what are the pathways that are affected by defects in both organelles, which revealed a prominent role for the cholesterol synthesis pathway. This pathway is transcriptionally up-regulated in cellular and mouse models of lysosomal defects and is transcriptionally down-regulated in cellular and mouse models of mitochondrial defects. We identified a role for post-transcriptional regulation of the transcription factor SREBF1, a master regulator of cholesterol and lipid biosynthesis, in models of mitochondrial respiratory chain deficiency. Furthermore, the retention of Ca 2+ in the lysosomes of cells with mitochondrial respiratory chain defects contributes to the differential regulation of the cholesterol synthesis pathway in the mitochondrial and lysosomal defects tested. Finally, we verified in vivo , using models of mitochondria-associated diseases in C. elegans , that normalization of lysosomal Ca 2+ levels results in partial rescue of the developmental arrest induced by the respiratory chain deficiency.
    DOI:  https://doi.org/10.1101/2024.03.06.583589
  3. Cell Rep. 2024 Mar 19. pii: S2211-1247(24)00303-6. [Epub ahead of print]43(4): 113975
    SIRT4 loss reprograms intestinal nucleotide metabolism to support proliferation following perturbation of homeostasis.
    Sarah A Tucker, Song-Hua Hu, Sejal Vyas, Albert Park, Shakchhi Joshi, Aslihan Inal, Tiffany Lam, Emily Tan, Kevin M Haigis, Marcia C Haigis.
      The intestine is a highly metabolic tissue, but the metabolic programs that influence intestinal crypt proliferation, differentiation, and regeneration are still emerging. Here, we investigate how mitochondrial sirtuin 4 (SIRT4) affects intestinal homeostasis. Intestinal SIRT4 loss promotes cell proliferation in the intestine following ionizing radiation (IR). SIRT4 functions as a tumor suppressor in a mouse model of intestinal cancer, and SIRT4 loss drives dysregulated glutamine and nucleotide metabolism in intestinal adenomas. Intestinal organoids lacking SIRT4 display increased proliferation after IR stress, along with increased glutamine uptake and a shift toward de novo nucleotide biosynthesis over salvage pathways. Inhibition of de novo nucleotide biosynthesis diminishes the growth advantage of SIRT4-deficient organoids after IR stress. This work establishes SIRT4 as a modulator of intestinal metabolism and homeostasis in the setting of DNA-damaging stress.
    Keywords:  CP: Cancer; SIRT4; glutamine; intestinal organoids; irradiation; nucleotide biosynthesis; nucleotide metabolism; sirtuin
    DOI:  https://doi.org/10.1016/j.celrep.2024.113975
  4. PLoS Biol. 2024 Mar;22(3): e3002549
    GALDAR: A genetically encoded galactose sensor for visualizing sugar metabolism in vivo.
    Uğurcan Sakizli, Tomomi Takano, Sa Kan Yoo.
      Sugar metabolism plays a pivotal role in sustaining life. Its dynamics within organisms is less understood compared to its intracellular metabolism. Galactose, a hexose stereoisomer of glucose, is a monosaccharide transported via the same transporters with glucose. Galactose feeds into glycolysis and regulates protein glycosylation. Defects in galactose metabolism are lethal for animals. Here, by transgenically implementing the yeast galactose sensing system into Drosophila, we developed a genetically encoded sensor, GALDAR, which detects galactose in vivo. Using this heterologous system, we revealed dynamics of galactose metabolism in various tissues. Notably, we discovered that intestinal stem cells do not uptake detectable levels of galactose or glucose. GALDAR elucidates the role for galactokinase in metabolism of galactose and a transition of galactose metabolism during the larval period. This work provides a new system that enables analyses of in vivo sugar metabolism.
    DOI:  https://doi.org/10.1371/journal.pbio.3002549
  5. bioRxiv. 2024 Mar 07. pii: 2024.03.05.583623. [Epub ahead of print]
    A Human Brain Map of Mitochondrial Respiratory Capacity and Diversity.
    Eugene V Mosharov, Ayelet M Rosenberg, Anna S Monzel, Corey A Osto, Linsey Stiles, Gorazd B Rosoklija, Andrew J Dwork, Snehal Bindra, Ya Zhang, Masashi Fujita, Madeline B Mariani, Mihran Bakalian, David Sulzer, Philip L De Jager, Vilas Menon, Orian S Shirihai, J John Mann, Mark Underwood, Maura Boldrini, Michel Thiebaut de Schotten, Martin Picard.
      Mitochondrial oxidative phosphorylation (OxPhos) powers brain activity 1,2 , and mitochondrial defects are linked to neurodegenerative and neuropsychiatric disorders 3,4 , underscoring the need to define the brain's molecular energetic landscape 5-10 . To bridge the cognitive neuroscience and cell biology scale gap, we developed a physical voxelization approach to partition a frozen human coronal hemisphere section into 703 voxels comparable to neuroimaging resolution (3×3×3 mm). In each cortical and subcortical brain voxel, we profiled mitochondrial phenotypes including OxPhos enzyme activities, mitochondrial DNA and volume density, and mitochondria-specific respiratory capacity. We show that the human brain contains a diversity of mitochondrial phenotypes driven by both topology and cell types. Compared to white matter, grey matter contains >50% more mitochondria. We show that the more abundant grey matter mitochondria also are biochemically optimized for energy transformation, particularly among recently evolved cortical brain regions. Scaling these data to the whole brain, we created a backward linear regression model integrating several neuroimaging modalities 11 , thereby generating a brain-wide map of mitochondrial distribution and specialization that predicts mitochondrial characteristics in an independent brain region of the same donor brain. This new approach and the resulting MitoBrainMap of mitochondrial phenotypes provide a foundation for exploring the molecular energetic landscape that enables normal brain functions, relating it to neuroimaging data, and defining the subcellular basis for regionalized brain processes relevant to neuropsychiatric and neurodegenerative disorders.
    DOI:  https://doi.org/10.1101/2024.03.05.583623
  6. Nat Rev Endocrinol. 2024 Mar 18.
    Metabolic regulation of skeletal cell fate and function.
    Steve Stegen, Geert Carmeliet.
      Bone development and bone remodelling during adult life are highly anabolic processes requiring an adequate supply of oxygen and nutrients. Bone-forming osteoblasts and bone-resorbing osteoclasts interact closely to preserve bone mass and architecture and are often located close to blood vessels. Chondrocytes within the developing growth plate ensure that bone lengthening occurs before puberty, but these cells function in an avascular environment. With ageing, numerous bone marrow adipocytes appear, often with negative effects on bone properties. Many studies have now indicated that skeletal cells have specific metabolic profiles that correspond to the nutritional microenvironment and their stage-specific functions. These metabolic networks provide not only skeletal cells with sufficient energy, but also biosynthetic intermediates that are necessary for proliferation and extracellular matrix synthesis. Moreover, these metabolic pathways control redox homeostasis to avoid oxidative stress and safeguard cell survival. Finally, several intracellular metabolites regulate the activity of epigenetic enzymes and thus control the fate and function of skeletal cells. The metabolic profile of skeletal cells therefore not only reflects their cellular state, but can also drive cellular activity. Insight into skeletal cell metabolism will thus not only advance our understanding of skeletal development and homeostasis, but also of skeletal disorders, such as osteoarthritis, diabetic bone disease and bone malignancies.
    DOI:  https://doi.org/10.1038/s41574-024-00969-x
  7. iScience. 2024 Apr 19. 27(4): 109379
    Mitochondria-ER contact sites expand during mitosis.
    Fang Yu, Raphael Courjaret, Lama Assaf, Asha Elmi, Ayat Hammad, Melanie Fisher, Mark Terasaki, Khaled Machaca.
      Mitochondria-ER contact sites (MERCS) are involved in energy homeostasis, redox and Ca2+ signaling, and inflammation. MERCS are heavily studied; however, little is known about their regulation during mitosis. Here, we show that MERCS expand during mitosis in three cell types using various approaches, including transmission electron microscopy, serial EM coupled to 3D reconstruction, and a split GFP MERCS marker. We further show enhanced Ca2+ transfer between the ER and mitochondria using either direct Ca2+ measurements or by quantifying the activity of Ca2+-dependent mitochondrial dehydrogenases. Collectively, our results support a lengthening of MERCS in mitosis that is associated with improved Ca2+ coupling between the two organelles. This augmented Ca2+ coupling could be important to support the increased energy needs of the cell during mitosis.
    Keywords:  Biological sciences; Cell biology; Natural sciences
    DOI:  https://doi.org/10.1016/j.isci.2024.109379
  8. Sci Rep. 2024 Mar 21. 14(1): 6751
    MCU-independent Ca2+ uptake mediates mitochondrial Ca2+ overload and necrotic cell death in a mouse model of Duchenne muscular dystrophy.
    Michael J Bround, Eaman Abay, Jiuzhou Huo, Julian R Havens, Allen J York, Donald M Bers, Jeffery D Molkentin.
      Mitochondrial Ca2+ overload can mediate mitochondria-dependent cell death, a major contributor to several human diseases. Indeed, Duchenne muscular dystrophy (MD) is driven by dysfunctional Ca2+ influx across the sarcolemma that causes mitochondrial Ca2+ overload, organelle rupture, and muscle necrosis. The mitochondrial Ca2+ uniporter (MCU) complex is the primary characterized mechanism for acute mitochondrial Ca2+ uptake. One strategy for preventing mitochondrial Ca2+ overload is deletion of the Mcu gene, the pore forming subunit of the MCU-complex. Conversely, enhanced MCU-complex Ca2+ uptake is achieved by deleting the inhibitory Mcub gene. Here we show that myofiber-specific Mcu deletion was not protective in a mouse model of Duchenne MD. Specifically, Mcu gene deletion did not reduce muscle histopathology, did not improve muscle function, and did not prevent mitochondrial Ca2+ overload. Moreover, myofiber specific Mcub gene deletion did not augment Duchenne MD muscle pathology. Interestingly, we observed MCU-independent Ca2+ uptake in dystrophic mitochondria that was sufficient to drive mitochondrial permeability transition pore (MPTP) activation and skeletal muscle necrosis, and this same type of activity was observed in heart, liver, and brain mitochondria. These results demonstrate that mitochondria possess an uncharacterized MCU-independent Ca2+ uptake mechanism that is sufficient to drive MPTP-dependent necrosis in MD in vivo.
    DOI:  https://doi.org/10.1038/s41598-024-57340-3
  9. Cancer Metab. 2024 Mar 21. 12(1): 9
    Serine synthesis and catabolism in starved lung cancer and primary bronchial epithelial cells.
    Theresa Haitzmann, Katharina Schindlmaier, Tobias Frech, Ayusi Mondal, Visnja Bubalo, Barbara Konrad, Gabriele Bluemel, Philipp Stiegler, Stefanie Lackner, Andelko Hrzenjak, Thomas Eichmann, Harald C Köfeler, Katharina Leithner.
      Serine and glycine give rise to important building blocks in proliferating cells. Both amino acids are either synthesized de novo or taken up from the extracellular space. In lung cancer, serine synthesis gene expression is variable, yet, expression of the initial enzyme, phosphoglycerate dehydrogenase (PHGDH), was found to be associated with poor prognosis. While the contribution of de novo synthesis to serine pools has been shown to be enhanced by serine starvation, the impact of glucose deprivation, a commonly found condition in solid cancers is poorly understood. Here, we utilized a stable isotopic tracing approach to assess serine and glycine de novo synthesis and uptake in different lung cancer cell lines and normal bronchial epithelial cells in variable serine, glycine, and glucose conditions. Under low glucose supplementation (0.2 mM, 3-5% of normal plasma levels), serine de novo synthesis was maintained or even activated. As previously reported, also gluconeogenesis supplied carbons from glutamine to serine and glycine under these conditions. Unexpectedly, low glucose treatment consistently enhanced serine to glycine conversion, along with an up-regulation of the mitochondrial one-carbon metabolism enzymes, serine hydroxymethyltransferase (SHMT2) and methylenetetrahydrofolate dehydrogenase (MTHFD2). The relative contribution of de novo synthesis greatly increased in low serine/glycine conditions. In bronchial epithelial cells, adaptations occurred in a similar fashion as in cancer cells, but serine synthesis and serine to glycine conversion, as assessed by label enrichments and gene expression levels, were generally lower than in (PHGDH positive) cancer cells. In summary, we found a variable contribution of glucose or non-glucose carbon sources to serine and glycine and a high adaptability of the downstream one-carbon metabolism pathway to variable glucose supply.
    Keywords:  Glycine; Lung cancer; Metabolism; Serine; Starvation
    DOI:  https://doi.org/10.1186/s40170-024-00337-3
  10. Proc Natl Acad Sci U S A. 2024 Mar 26. 121(13): e2318969121
    Ferredoxin reduction by hydrogen with iron functions as an evolutionary precursor of flavin-based electron bifurcation.
    Max Brabender, Delfina P Henriques Pereira, Natalia Mrnjavac, Manon Laura Schlikker, Zen-Ichiro Kimura, Jeerus Sucharitakul, Karl Kleinermanns, Harun Tüysüz, Wolfgang Buckel, Martina Preiner, William F Martin.
      Autotrophic theories for the origin of metabolism posit that the first cells satisfied their carbon needs from CO2 and were chemolithoautotrophs that obtained their energy and electrons from H2. The acetyl-CoA pathway of CO2 fixation is central to that view because of its antiquity: Among known CO2 fixing pathways it is the only one that is i) exergonic, ii) occurs in both bacteria and archaea, and iii) can be functionally replaced in full by single transition metal catalysts in vitro. In order to operate in cells at a pH close to 7, however, the acetyl-CoA pathway requires complex multi-enzyme systems capable of flavin-based electron bifurcation that reduce low potential ferredoxin-the physiological donor of electrons in the acetyl-CoA pathway-with electrons from H2. How can the acetyl-CoA pathway be primordial if it requires flavin-based electron bifurcation? Here, we show that native iron (Fe0), but not Ni0, Co0, Mo0, NiFe, Ni2Fe, Ni3Fe, or Fe3O4, promotes the H2-dependent reduction of aqueous Clostridium pasteurianum ferredoxin at pH 8.5 or higher within a few hours at 40 °C, providing the physiological function of flavin-based electron bifurcation, but without the help of enzymes or organic redox cofactors. H2-dependent ferredoxin reduction by iron ties primordial ferredoxin reduction and early metabolic evolution to a chemical process in the Earth's crust promoted by solid-state iron, a metal that is still deposited in serpentinizing hydrothermal vents today.
    Keywords:  acetyl CoA pathway; origin of life; origin of metabolism; serpentinization; transition metals
    DOI:  https://doi.org/10.1073/pnas.2318969121
  11. Nat Commun. 2024 Mar 19. 15(1): 2463
    NMR and MS reveal characteristic metabolome atlas and optimize esophageal squamous cell carcinoma early detection.
    Yan Zhao, Changchun Ma, Rongzhi Cai, Lijing Xin, Yongsheng Li, Lixin Ke, Wei Ye, Ting Ouyang, Jiahao Liang, Renhua Wu, Yan Lin.
      Metabolic changes precede malignant histology. However, it remains unclear whether detectable characteristic metabolome exists in esophageal squamous cell carcinoma (ESCC) tissues and biofluids for early diagnosis. Here, we conduct NMR- and MS-based metabolomics on 1,153 matched ESCC tissues, normal mucosae, pre- and one-week post-operative sera and urines from 560 participants across three hospitals, with machine learning and WGCNA. Aberrations in 'alanine, aspartate and glutamate metabolism' proved to be prevalent throughout the ESCC evolution, consistently identified by NMR and MS, and reflected in 16 serum and 10 urine metabolic signatures in both discovery and validation sets. NMR-based simplified panels of any five serum or urine metabolites outperform clinical serological tumor markers (AUC = 0.984 and 0.930, respectively), and are effective in distinguishing early-stage ESCC in test set (serum accuracy = 0.994, urine accuracy = 0.879). Collectively, NMR-based biofluid screening can reveal characteristic metabolic events of ESCC and be feasible for early detection (ChiCTR2300073613).
    DOI:  https://doi.org/10.1038/s41467-024-46837-0
  12. Sci Rep. 2024 03 18. 14(1): 6504
    The roles of autophagy, ferroptosis and pyroptosis in the anti-ovarian cancer mechanism of harmine and their crosstalk.
    Jun Zhu, Hong Zhu, Qing Zhu, Shi Lei Xu, Lu Xiao, Ming Yue Zhang, Jun Gao.
      This study aimed to investigate the role of autophagy, ferroptosis, and pyroptosis in the antitumour mechanism of harmine (Har) and its crosstalk in ovarian cancer. By transmission electron microscopy, we found that compared with those in the control group, the cytoplasm of human ovarian cancer cells (SKOV3) treated with Har showed increased numbers of autophagic vesicles, decreased intracellular mitochondrial volume, increased bilayer membrane density, and decreased cristae. Western blot, immunofluorescence, and monodasylcadaverine (MDC) staining all suggested that Har promoted autophagy in SKOV3 cells. LY294002 and siFOXO3 rescued the inhibition of the PI3K/AKT/mTOR/FOXO3 signalling pathway and the promotion of autophagy by Har. Additionally, the levels of ferroptosis- and pyroptosis-related proteins and the levels of Fe2+ , glutathione (GSH), malondialdehyde (MDA), and superoxide dismutase (SOD) suggested that Har promoted ferroptosis and pyroptosis in SKOV3 cells. Interestingly, pretreatment with chloroquine (CQ), erastin, rapamycin (Rap), or ferrostatin-1 (Fer-1) increased or reversed the ferroptosis and pyroptosis promoted by Har, respectively. In vivo, the volume of tumours in the Har group was decreased, and immunohistochemistry revealed decreased levels of Ki-67 and GPX4 and increased levels of ATG5 and NARL3. In conclusion, Har exerts its anti-ovarian cancer effect not only by promoting autophagy by regulating the PI3K/AKT/mTOR/FOXO3 signalling pathway but also by promoting ferroptosis and pyroptosis. Additionally, there is complex crosstalk between autophagy, ferroptosis, and pyroptosis in ovarian cancer.
    DOI:  https://doi.org/10.1038/s41598-024-57196-7
  13. Cold Spring Harb Perspect Med. 2024 Mar 19. pii: a041536. [Epub ahead of print]
    Knowledge-Based Therapeutics for Tricarboxylic Acid (TCA) Cycle-Deficient Cancers.
    Daniel Peled, Ruth Casey, Eyal Gottlieb.
      With the foundation pre-laid, research in the new millennium has readily excavated and expanded upon the architectural framework laid out by Otto Warburg's seminal work in a new wave of "westward expansion," ever widening our understanding of cancer metabolism beyond the telescopic vision seen over a century ago. On this path, the unique circuitry of the cancer metabolic program has been elucidated, illuminating mutations of conserved cellular pathways implicated in tumorigenesis. Paramount among these are mutations in tricarboxylic acid cycle enzymes, succinate dehydrogenase, and fumarate hydratase, leading to deleterious accumulations in metabolic intermediates, "oncometabolites," the pilots of the disease process. In this work, we seek to reflect on the advancements in the field in recent years, updating knowledge on the exact biochemical mechanisms at the helm of the tumor, providing rationale for clinical trials currently underway, and anticipating directions for the future on this expansive frontier.
    DOI:  https://doi.org/10.1101/cshperspect.a041536
  14. STAR Protoc. 2024 Mar 18. pii: S2666-1667(24)00129-1. [Epub ahead of print]5(2): 102964
    Protocol for analyzing energy metabolic pathway dependency in human liver cancer cell lines.
    Sk Ramiz Islam, Sebabrata Maity, Oishee Chakrabarti, Soumen Kanti Manna.
      Cellular energy metabolism analysis is complex, expensive, and indirect. We present a protocol to analyze relative contribution of metabolic pathways to ATP production by directly measuring ATP levels. We describe steps for cell counting and seeding in 96-well plate, treating with metformin, and systematic inhibition with metabolic inhibitors. We then detail procedures for a viability and ATP assay and calculating energy metabolism dependency. This high-throughput and accessible protocol works with any cell line and allows for flexible perturbation studies.
    Keywords:  Cancer; Metabolism; Molecular Biology
    DOI:  https://doi.org/10.1016/j.xpro.2024.102964
  15. Nat Metab. 2024 Mar 19.
    Insulin signalling regulates Pink1 mRNA localization via modulation of AMPK activity to support PINK1 function in neurons.
    J Tabitha Hees, Simone Wanderoy, Jana Lindner, Marlena Helms, Hariharan Murali Mahadevan, Angelika B Harbauer.
      Mitochondrial quality control failure is frequently observed in neurodegenerative diseases. The detection of damaged mitochondria by stabilization of PTEN-induced kinase 1 (PINK1) requires transport of Pink1 messenger RNA (mRNA) by tethering it to the mitochondrial surface. Here, we report that inhibition of AMP-activated protein kinase (AMPK) by activation of the insulin signalling cascade prevents Pink1 mRNA binding to mitochondria. Mechanistically, AMPK phosphorylates the RNA anchor complex subunit SYNJ2BP within its PDZ domain, a phosphorylation site that is necessary for its interaction with the RNA-binding protein SYNJ2. Notably, loss of mitochondrial Pink1 mRNA association upon insulin addition is required for PINK1 protein activation and its function as a ubiquitin kinase in the mitophagy pathway, thus placing PINK1 function under metabolic control. Induction of insulin resistance in vitro by the key genetic Alzheimer risk factor apolipoprotein E4 retains Pink1 mRNA at the mitochondria and prevents proper PINK1 activity, especially in neurites. Our results thus identify a metabolic switch controlling Pink1 mRNA localization and PINK1 activity via insulin and AMPK signalling in neurons and propose a mechanistic connection between insulin resistance and mitochondrial dysfunction.
    DOI:  https://doi.org/10.1038/s42255-024-01007-w
  16. Dev Cell. 2024 Mar 18. pii: S1534-5807(24)00110-2. [Epub ahead of print]
    Synchronized assembly of the oxidative phosphorylation system controls mitochondrial respiration in yeast.
    Daiana N Moretti-Horten, Carlotta Peselj, Asli Aras Taskin, Lisa Myketin, Uwe Schulte, Oliver Einsle, Friedel Drepper, Marcin Luzarowski, F-Nora Vögtle.
      Control of protein stoichiometry is essential for cell function. Mitochondrial oxidative phosphorylation (OXPHOS) presents a complex stoichiometric challenge as the ratio of the electron transport chain (ETC) and ATP synthase must be tightly controlled, and assembly requires coordinated integration of proteins encoded in the nuclear and mitochondrial genome. How correct OXPHOS stoichiometry is achieved is unknown. We identify the MitochondrialRegulatory hub for respiratoryAssembly (MiRA) platform, which synchronizes ETC and ATP synthase biogenesis in yeast. Molecularly, this is achieved by a stop-and-go mechanism: the uncharacterized protein Mra1 stalls complex IV assembly. Two "Go" signals are required for assembly progression: binding of the complex IV assembly factor Rcf2 and Mra1 interaction with an Atp9-translating mitoribosome induce Mra1 degradation, allowing synchronized maturation of complex IV and the ATP synthase. Failure of the stop-and-go mechanism results in cell death. MiRA controls OXPHOS assembly, ensuring correct stoichiometry of protein machineries encoded by two different genomes.
    Keywords:  complex stoichiometry; mitochondria; mitoribosome; protein complex assembly; protein import; protein quality control; respiratory chain
    DOI:  https://doi.org/10.1016/j.devcel.2024.02.011
  17. Semin Cancer Biol. 2024 Mar 15. pii: S1044-579X(24)00020-8. [Epub ahead of print]
    GLUT and HK: two primary and essential key players in Tumor glycolysis.
    Dhiraj Yadav, Anubha Yadav, Sujata Bhattacharya, Akansha Dagar, Vinit Kumar, Reshma Rani.
      Cancer cells reprogram their metabolism to become "glycolysis-dominant," which enables them to meet their energy and macromolecule needs and enhancing their rate of survival. This glycolytic-dominancy is known as the "Warburg effect", a significant factor in the growth and invasion of malignant tumors. Many studies confirmed that members of the GLUT family, specifically HK-II from the HK family play a pivotal role in the Warburg effect, and are closely associated with glucose transportation followed by glucose metabolism in cancer cells. Overexpression of GLUTs and HK-II correlates with aggressive tumor behaviour and tumor microenvironment making them attractive therapeutic targets. Several studies have proven that the regulation of GLUTs and HK-II expression improves the treatment outcome for various tumors. Therefore, small molecule inhibitors targeting GLUT and HK-II show promise in sensitizing cancer cells to treatment, either alone or in combination with existing therapies including chemotherapy, radiotherapy, immunotherapy, and photodynamic therapy. Despite existing therapies, viable methods to target the glycolysis of cancer cells are currently lacking to increase the effectiveness of cancer treatment. This review explores the current understanding of GLUT and HK-II in cancer metabolism, recent inhibitor developments, and strategies for future drug development, offering insights into improving cancer treatment efficacy.
    Keywords:  Cancer cells reprograming; GLUT; Glucose transporter; Warburg effect; hexokinase
    DOI:  https://doi.org/10.1016/j.semcancer.2024.03.001
  18. Proc Natl Acad Sci U S A. 2024 Mar 26. 121(13): e2312172121
    Luminal transport through intact endoplasmic reticulum limits the magnitude of localized Ca2+ signals.
    Cécile C Crapart, Zubenelgenubi C Scott, Tasuku Konno, Aman Sharma, Pierre Parutto, David M D Bailey, Laura M Westrate, Edward Avezov, Elena F Koslover.
      The endoplasmic reticulum (ER) forms an interconnected network of tubules stretching throughout the cell. Understanding how ER functionality relies on its structural organization is crucial for elucidating cellular vulnerability to ER perturbations, which have been implicated in several neuronal pathologies. One of the key functions of the ER is enabling Ca[Formula: see text] signaling by storing large quantities of this ion and releasing it into the cytoplasm in a spatiotemporally controlled manner. Through a combination of physical modeling and live-cell imaging, we demonstrate that alterations in ER shape significantly impact its ability to support efficient local Ca[Formula: see text] releases, due to hindered transport of luminal content within the ER. Our model reveals that rapid Ca[Formula: see text] release necessitates mobile luminal buffer proteins with moderate binding strength, moving through a well-connected network of ER tubules. These findings provide insight into the functional advantages of normal ER architecture, emphasizing its importance as a kinetically efficient intracellular Ca[Formula: see text] delivery system.
    Keywords:  calcium signaling; endoplasmic reticulum; intracellular transport; mathematical modeling; organelles
    DOI:  https://doi.org/10.1073/pnas.2312172121
  19. Anal Chim Acta. 2024 Apr 22. pii: S0003-2670(24)00223-X. [Epub ahead of print]1299 342422
    A novel fluorescent probe for visualizing viscosity changes in lipid droplets during chemotherapy-induced ferroptosis.
    Di Wei, Yingshu Dai, Jing Cao, Nanyan Fu.
       BACKGROUND: Ferroptosis, as a novel form of cell death, is becoming one of the hot topics in cancer treatment research. It differs from necrosis and autophagy in that it involves the accumulation of lipid peroxides and is triggered by iron dependency. Recent studies have suggested that this mechanism may alter the viscosity or structure of lipid droplets (LDs). The relationship between LDs viscosity and ferroptosis remains an active area of research with limited reports at present. Additionally, there is a lack of effective anticancer drugs targeting the ferroptosis pathway to promote ferroptosis in tumour cells. Therefore, the development of tools to detect viscosity changes during ferroptosis and targeted therapeutic strategies is of great significance.
    RESULTS: By coupling 1,3-indandione with naphthalimide, including decamethylamine as a LDs recognition group, we designed and synthesized an environmental fluorescent probe that induces intramolecular charge transfer (TICT) effects. Notably, the diffusion and transport of intracellular substances may be affected in highly viscous environments. Under such conditions, intracellular iron ions may accumulate, leading to peroxide production and cellular damage, which can trigger ferroptosis. Therefore, WD-1 achieved excellent in situ bioimaging of LDs targeting and its viscosity during ferroptosis in HeLa cells and zebrafish. Furthermore, it was observed that WD-1 effectively differentiated between malignant and normal cells during this process, highlighting its potential significance in distinguishing cellular states. In addition, we used the antitumour drug paclitaxel to study ferroptosis in cancer cells. These findings not only provide an excellent tool for the development of the ferroptosis response, but also are crucial for understanding the biological properties of LDs during the ferroptosis response.
    SIGNIFICANCE AND NOVELTY: Based on a powerful tool of fluorescent probe with in vivo bioimaging, we developed WD-1 to track the impact of paclitaxel on the process of ferroptosis in living cells. Therefore, we preliminarily believe that paclitaxel may affect the occurrence of ferroptosis and control apoptosis in cancer cells. These findings not only serve as an exceptional tool for advancing our understanding of the ferroptosis response, but furthermore play a vital role in comprehending the biological characteristics of LDs in relation to ferroptosis.
    Keywords:  Bioimaging in cells and zebrafish; Ferroptosis; Fluorescent probe; Lipid droplets; Paclitaxel
    DOI:  https://doi.org/10.1016/j.aca.2024.342422
  20. European J Org Chem. 2023 Aug 14. 26(31): e202300339
    Fluorinated Analogues to the Pentuloses of the Pentose Phosphate Pathway.
    Lukas Scheibelberger, Toda Stankovic, Kaja Liepert, Andreas Kienzle, Eva-Maria Patronas, Theresa Balber, Markus Mitterhauser, Arvand Haschemi, Katharina Pallitsch.
      Fluorinated carbohydrates are valuable tools for enzymological studies due to their increased metabolic stability compared to their non-fluorinated analogues. Replacing different hydroxyl groups within the same monosaccharide by fluorine allows to influence a wide range of sugar-receptor interactions and enzymatic transformations. In the past, this principle was frequently used to study the metabolism of highly abundant carbohydrates, while the metabolic fate of rare sugars is still poorly studied. Rare sugars, however, are key intermediates of many metabolic routes, such as the pentose phosphate pathway (PPP). Here we present the design and purely chemical synthesis of a set of three deoxyfluorinated analogues of the rare sugars d-xylulose and d-ribulose: 1-deoxy-1-fluoro-d-ribulose (1DFRu), 3-deoxy-3-fluoro-d-ribulose (3DFRu) and 3-deoxy-3-fluoro-d-xylulose (3DFXu). Together with a designed set of potential late-stage radio-fluorination precursors, they have the potential to become useful tools for studies on the complex equilibria of the non-oxidative PPP.
    Keywords:  d-Ribulose; d-Xylulose; deoxyfluorinated carbohydrates; pentose-phosphate-pathway; rare sugars
    DOI:  https://doi.org/10.1002/ejoc.202300339
  21. Proc Natl Acad Sci U S A. 2024 Mar 26. 121(13): e2319429121
    Activation of polyamine catabolism promotes glutamine metabolism and creates a targetable vulnerability in lung cancer.
    Xinlu Han, Deyu Wang, Liao Yang, Ning Wang, Jianliang Shen, Jinghan Wang, Lei Zhang, Li Chen, Shenglan Gao, Wei-Xing Zong, Yongbo Wang.
      Polyamines are a class of small polycationic alkylamines that play essential roles in both normal and cancer cell growth. Polyamine metabolism is frequently dysregulated and considered a therapeutic target in cancer. However, targeting polyamine metabolism as monotherapy often exhibits limited efficacy, and the underlying mechanisms are incompletely understood. Here we report that activation of polyamine catabolism promotes glutamine metabolism, leading to a targetable vulnerability in lung cancer. Genetic and pharmacological activation of spermidine/spermine N1-acetyltransferase 1 (SAT1), the rate-limiting enzyme of polyamine catabolism, enhances the conversion of glutamine to glutamate and subsequent glutathione (GSH) synthesis. This metabolic rewiring ameliorates oxidative stress to support lung cancer cell proliferation and survival. Simultaneous glutamine limitation and SAT1 activation result in ROS accumulation, growth inhibition, and cell death. Importantly, pharmacological inhibition of either one of glutamine transport, glutaminase, or GSH biosynthesis in combination with activation of polyamine catabolism synergistically suppresses lung cancer cell growth and xenograft tumor formation. Together, this study unveils a previously unappreciated functional interconnection between polyamine catabolism and glutamine metabolism and establishes cotargeting strategies as potential therapeutics in lung cancer.
    Keywords:  SAT1; glutamine metabolism; lung cancer; polyamine catabolism
    DOI:  https://doi.org/10.1073/pnas.2319429121
  22. Nat Commun. 2024 Mar 21. 15(1): 2487
    The AMPK-related kinase NUAK1 controls cortical axons branching by locally modulating mitochondrial metabolic functions.
    Marine Lanfranchi, Sozerko Yandiev, Géraldine Meyer-Dilhet, Salma Ellouze, Martijn Kerkhofs, Raphael Dos Reis, Audrey Garcia, Camille Blondet, Alizée Amar, Anita Kneppers, Hélène Polvèche, Damien Plassard, Marc Foretz, Benoit Viollet, Kei Sakamoto, Rémi Mounier, Cyril F Bourgeois, Olivier Raineteau, Evelyne Goillot, Julien Courchet.
      The cellular mechanisms underlying axonal morphogenesis are essential to the formation of functional neuronal networks. We previously identified the autism-linked kinase NUAK1 as a central regulator of axon branching through the control of mitochondria trafficking. However, (1) the relationship between mitochondrial position, function and axon branching and (2) the downstream effectors whereby NUAK1 regulates axon branching remain unknown. Here, we report that mitochondria recruitment to synaptic boutons supports collateral branches stabilization rather than formation in mouse cortical neurons. NUAK1 deficiency significantly impairs mitochondrial metabolism and axonal ATP concentration, and upregulation of mitochondrial function is sufficient to rescue axonal branching in NUAK1 null neurons in vitro and in vivo. Finally, we found that NUAK1 regulates axon branching through the mitochondria-targeted microprotein BRAWNIN. Our results demonstrate that NUAK1 exerts a dual function during axon branching through its ability to control mitochondrial distribution and metabolic activity.
    DOI:  https://doi.org/10.1038/s41467-024-46146-6
  23. Cold Spring Harb Perspect Med. 2024 Mar 19. pii: a041533. [Epub ahead of print]
    Tracing the Diverse Paths of One-Carbon Metabolism in Cancer and Beyond.
    Esther W Lim, Christian M Metallo.
      One-carbon (1C) metabolism is a network of biochemical reactions distributed across organelles that delivers folate-activated 1C units to support macromolecule synthesis, methylation, and reductive homeostasis. Fluxes through these pathways are up-regulated in highly proliferative cancer cells, and anti-folates, which target enzymes within the 1C pathway, have long been used in the treatment of cancer. In this work, we review fundamental aspects of 1C metabolism and place it in context with other biosynthetic and redox pathways, such that 1C metabolism acts to bridge pathways across compartments. We further discuss the importance of stable-isotope-tracing techniques combined with mass spectrometry analysis to study 1C metabolism and conclude by highlighting therapeutic approaches that could exploit cancer cells' dependency on 1C metabolism.
    DOI:  https://doi.org/10.1101/cshperspect.a041533
  24. Nat Commun. 2024 Mar 15. 15(1): 2372
    A citric acid cycle-deficient Escherichia coli as an efficient chassis for aerobic fermentations.
    Hang Zhou, Yiwen Zhang, Christopher P Long, Xuesen Xia, Yanfen Xue, Yanhe Ma, Maciek R Antoniewicz, Yong Tao, Baixue Lin.
      Tricarboxylic acid cycle (TCA cycle) plays an important role for aerobic growth of heterotrophic bacteria. Theoretically, eliminating TCA cycle would decrease carbon dissipation and facilitate chemicals biosynthesis. Here, we construct an E. coli strain without a functional TCA cycle that can serve as a versatile chassis for chemicals biosynthesis. We first use adaptive laboratory evolution to recover aerobic growth in minimal medium of TCA cycle-deficient E. coli. Inactivation of succinate dehydrogenase is a key event in the evolutionary trajectory. Supply of succinyl-CoA is identified as the growth limiting factor. By replacing endogenous succinyl-CoA dependent enzymes, we obtain an optimized TCA cycle-deficient E. coli strain. As a proof of concept, the strain is engineered for high-yield production of four separate products. This work enhances our understanding of the role of the TCA cycle in E. coli metabolism and demonstrates the advantages of using TCA cycle-deficient E. coli strain for biotechnological applications.
    DOI:  https://doi.org/10.1038/s41467-024-46655-4
  25. PLoS One. 2024 ;19(3): e0300593
    Metabolic impairments associated with type 2 diabetes mellitus and the potential effects of exercise therapy: An exploratory randomized trial based on untargeted metabolomics.
    Furong Zhang, Xixi Chen, Mingxiao Yang, Xiaoyu Shen, Yiliang Wang, Dongling Zhong, Fang Zeng, Rongjiang Jin.
       BACKGROUND: Type 2 diabetes mellitus (T2DM) is a common condition that is characterized by metabolic impairments. Exercise therapy has proven effective in improving the physiological and psychological states of patients with T2DM; however, the influence of different exercise modalities on metabolic profiles is not fully understood. This study first aimed to investigate the metabolic changes associated with T2DM among patients and then to evaluate the potential physiological effects of different exercise modalities (Tai Chi and brisk walking) on their metabolic profiles.
    METHODS: This study included 20 T2DM patients and 11 healthy subjects. Patients were randomly allocated to either the Tai Chi or walking group to perform Dijia simplified 24-form Tai Chi or brisk walking (80-100 m/min), with 90 minutes each time, three times per week for 12 weeks, for a total of 36 sessions. The healthy group maintained daily living habits without intervention. Glycemic tests were conducted at the baseline and after 12 weeks. Serum and urine samples were collected for untargeted metabolomic analyses at baseline and 12 weeks to examine the differential metabolic profiles between T2DM and healthy subjects, and the metabolic alterations of T2DM patients before and after exercise therapy.
    RESULTS: Compared to the healthy group, T2DM patients exhibited metabolic disturbances in carbohydrates (fructose, mannose, galactose, glycolysis/gluconeogenesis), lipids (inositol phosphate), and amino acids (arginine, proline, cysteine, methionine, valine, leucine, and isoleucine) metabolism, including 20 differential metabolites in the serum and six in the urine. After exercise, the glycemic results showed insignificant changes. However, patients who practiced Tai Chi showed significant improvements in their post-treatment metabolic profiles compared to baseline, with nine serum and six urine metabolites, including branch-chained amino acids (BCAAs); while those in the walking group had significantly altered nine serum and four urine metabolites concerning steroid hormone biosynthesis and arachidonic acid metabolism compared to baseline.
    CONCLUSION: T2DM patients displayed impaired carbohydrate, lipid, and amino acid metabolism, and exercise therapy improved their metabolic health. Different modalities may act through different pathways. Tai Chi may improve disrupted BCAAs metabolism, whereas brisk walking mainly regulates steroid hormone biosynthesis and arachidonic acid metabolism.
    DOI:  https://doi.org/10.1371/journal.pone.0300593
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