bims-celmim Biomed News
on Cellular and mitochondrial metabolism
Issue of 2024–06–02
twenty papers selected by
Marc Segarra Mondejar
  1. Transport activity regulates mitochondrial bioenergetics and biogenesis in renal tubules.
  2. mitoBKCa is functionally expressed in murine and human breast cancer cells and potentially contributes to metabolic reprogramming.
  3. AspSnFR: A genetically encoded biosensor for real-time monitoring of aspartate in live cells.
  4. Reciprocal Regulation of Cardiac β-Oxidation and Pyruvate Dehydrogenase by Insulin.
  5. L-serine deficiency: on the properties of the Asn133Ser variant of human phosphoserine phosphatase.
  6. A mitochondrial-targeted activity-based sensing probe for ratiometric imaging of formaldehyde reveals key regulators of the mitochondrial one-carbon pool.
  7. 13C metabolite tracing reveals glutamine and acetate as critical in vivo fuels for CD8 T cells.
  8. Regulation of TSC2 lysosome translocation and mitochondrial turnover by TSC2 acetylation status.
  9. Breast cancer cell-secreted miR-199b-5p hijacks neurometabolic coupling to promote brain metastasis.
  10. Mitochondrial iron deficiency triggers cytosolic iron overload in PKAN hiPS-derived astrocytes.
  11. Cells dispose of cytoplasmic mitochondrial DNA by nucleoid-phagy.
  12. Hairpin protein partitioning from the ER to lipid droplets involves major structural rearrangements.
  13. Cystine/glutamate antiporter xCT controls skeletal muscle glutathione redox, bioenergetics and differentiation.
  14. Mechanical activation of VE-cadherin stimulates AMPK to increase endothelial cell metabolism and vasodilation.
  15. Oral mitochondrial transplantation using nanomotors to treat ischaemic heart disease.
  16. CPT2 mediated fatty acid oxidation is dispensable for humoral immunity.
  17. High-resolution in situ structures of mammalian respiratory supercomplexes.
  18. Movement of the endoplasmic reticulum is driven by multiple classes of vesicles marked by Rab-GTPases.
  19. Rab30 facilitates lipid homeostasis during fasting.
  20. Targeting the autophagy-NAD axis protects against cell death in Niemann-Pick type C1 disease models.
  1. FASEB J. 2024 May 31. 38(10): e23703
    Transport activity regulates mitochondrial bioenergetics and biogenesis in renal tubules.
    Chih-Jen Cheng, Jonathan M Nizar, Dao-Fu Dai, Chou-Long Huang.
      Renal tubules are featured with copious mitochondria and robust transport activity. Mutations in mitochondrial genes cause congenital renal tubulopathies, and changes in transport activity affect mitochondrial morphology, suggesting mitochondrial function and transport activity are tightly coupled. Current methods of using bulk kidney tissues or cultured cells to study mitochondrial bioenergetics are limited. Here, we optimized an extracellular flux analysis (EFA) to study mitochondrial respiration and energy metabolism using microdissected mouse renal tubule segments. EFA detects mitochondrial respiration and glycolysis by measuring oxygen consumption and extracellular acidification rates, respectively. We show that both measurements positively correlate with sample sizes of a few centimeter-length renal tubules. The thick ascending limbs (TALs) and distal convoluted tubules (DCTs) critically utilize glucose/pyruvate as energy substrates, whereas proximal tubules (PTs) are significantly much less so. Acute inhibition of TALs' transport activity by ouabain treatment reduces basal and ATP-linked mitochondrial respiration. Chronic inhibition of transport activity by 2-week furosemide treatment or deletion of with-no-lysine kinase 4 (Wnk4) decreases maximal mitochondrial capacity. In addition, chronic inhibition downregulates mitochondrial DNA mass and mitochondrial length/density in TALs and DCTs. Conversely, gain-of-function Wnk4 mutation increases maximal mitochondrial capacity and mitochondrial length/density without increasing mitochondrial DNA mass. In conclusion, EFA is a sensitive and reliable method to investigate mitochondrial functions in isolated renal tubules. Transport activity tightly regulates mitochondrial bioenergetics and biogenesis to meet the energy demand in renal tubules. The system allows future investigation into whether and how mitochondria contribute to tubular remodeling adapted to changes in transport activity.
    Keywords:  distal convoluted tubule; extracellular flux analysis; glycolysis; mitochondrial respiration; thick ascending limb; transport activity
    DOI:  https://doi.org/10.1096/fj.202400358RR
  2. Elife. 2024 May 29. pii: RP92511. [Epub ahead of print]12
    mitoBKCa is functionally expressed in murine and human breast cancer cells and potentially contributes to metabolic reprogramming.
    Helmut Bischof, Selina Maier, Piotr Koprowski, Bogusz Kulawiak, Sandra Burgstaller, Joanna Jasińska, Kristian Serafimov, Monika Zochowska, Dominic Gross, Werner Schroth, Lucas Matt, David Arturo Juarez Lopez, Ying Zhang, Irina Bonzheim, Florian A Büttner, Falko Fend, Matthias Schwab, Andreas L Birkenfeld, Roland Malli, Michael Lämmerhofer, Piotr Bednarczyk, Adam Szewczyk, Robert Lukowski.
      Alterations in the function of K+ channels such as the voltage- and Ca2+-activated K+ channel of large conductance (BKCa) reportedly promote breast cancer (BC) development and progression. Underlying molecular mechanisms remain, however, elusive. Here, we provide electrophysiological evidence for a BKCa splice variant localized to the inner mitochondrial membrane of murine and human BC cells (mitoBKCa). Through a combination of genetic knockdown and knockout along with a cell permeable BKCa channel blocker, we show that mitoBKCa modulates overall cellular and mitochondrial energy production, and mediates the metabolic rewiring referred to as the 'Warburg effect', thereby promoting BC cell proliferation in the presence and absence of oxygen. Additionally, we detect mitoBKCa and BKCa transcripts in low or high abundance, respectively, in clinical BC specimens. Together, our results emphasize, that targeting mitoBKCa could represent a treatment strategy for selected BC patients in future.
    Keywords:  K+ channels; Kcnma1; Slo1; Warburg effect; biosensors; breast cancer; cancer biology; cell biology; human; metabolic reprogramming; mitoBKCa; mouse
    DOI:  https://doi.org/10.7554/eLife.92511
  3. Cell Chem Biol. 2024 May 20. pii: S2451-9456(24)00179-X. [Epub ahead of print]
    AspSnFR: A genetically encoded biosensor for real-time monitoring of aspartate in live cells.
    Lars Hellweg, Martin Pfeifer, Miroslaw Tarnawski, Shao Thing-Teoh, Lena Chang, Andrea Bergner, Jana Kress, Julien Hiblot, Tabea Wiedmer, Giulio Superti-Furga, Jürgen Reinhardt, Kai Johnsson, Philipp Leippe.
      Aspartate is crucial for nucleotide synthesis, ammonia detoxification, and maintaining redox balance via the malate-aspartate-shuttle (MAS). To disentangle these multiple roles of aspartate metabolism, tools are required that measure aspartate concentrations in real time and in live cells. We introduce AspSnFR, a genetically encoded green fluorescent biosensor for intracellular aspartate, engineered through displaying and screening biosensor libraries on mammalian cells. In live cells, AspSnFR is able to precisely and quantitatively measure cytosolic aspartate concentrations and dissect its production from glutamine. Combining high-content imaging of AspSnFR with pharmacological perturbations exposes differences in metabolic vulnerabilities of aspartate levels based on nutrient availability. Further, AspSnFR facilitates tracking of aspartate export from mitochondria through SLC25A12, the MAS' key transporter. We show that SLC25A12 is a rapidly responding and direct route to couple Ca2+ signaling with mitochondrial aspartate export. This establishes SLC25A12 as a crucial link between cellular signaling, mitochondrial respiration, and metabolism.
    DOI:  https://doi.org/10.1016/j.chembiol.2024.05.002
  4. J Biol Chem. 2024 May 23. pii: S0021-9258(24)01913-6. [Epub ahead of print] 107412
    Reciprocal Regulation of Cardiac β-Oxidation and Pyruvate Dehydrogenase by Insulin.
    Abdallah Elnwasany, Heba A Ewida, Ivan Menendez-Montes, Monika Mizerska, Xiaorong Fu, Chai-Wan Kim, Jay D Horton, Shawn C Burgess, Beverly A Rothermel, Pamela A Szweda, Luke I Szweda.
      The heart alters the rate and relative oxidation of fatty acids and glucose based on availability and energetic demand. Insulin plays a crucial role in this process diminishing fatty acid and increasing glucose oxidation when glucose availability increases. Loss of insulin sensitivity and metabolic flexibility can result in cardiovascular disease. It is therefore important to identify mechanisms by which insulin regulates substrate utilization in the heart. Mitochondrial pyruvate dehydrogenase (PDH) is the key regulatory site for the oxidation of glucose for ATP production. Nevertheless, the impact of insulin on PDH activity has not been fully delineated, particularly in the heart. We sought in vivo evidence that insulin stimulates cardiac PDH and that this process is driven by inhibition of fatty acid oxidation. Mice injected with insulin exhibited dephosphorylation and activation of cardiac PDH. This was accompanied by an increase in the content of malonyl-CoA, an inhibitor of carnitine palmitoyltransferase 1 (CPT1) and, thus, mitochondrial import of fatty acids. Administration of the CPT1 inhibitor oxfenicine was sufficient to activate PDH. Malonyl-CoA is produced by acetyl-CoA carboxylase (ACC). Pharmacologic inhibition or knockout of cardiac ACC diminished insulin-dependent production of malonyl-CoA and activation of PDH. Finally, circulating insulin and cardiac glucose utilization exhibit daily rhythms reflective of nutritional status. We demonstrate that time of day-dependent changes in PDH activity are mediated, in part, by ACC-dependent production of malonyl-CoA. Thus, by inhibiting fatty acid oxidation, insulin reciprocally activates PDH. These studies identify potential molecular targets to promote cardiac glucose oxidation and treat heart disease.
    Keywords:  Acetyl-CoA Carboxylase; Heart; Insulin; Malonyl-CoA; Mitochondria; Pyruvate Dehydrogenase; β-Oxidation
    DOI:  https://doi.org/10.1016/j.jbc.2024.107412
  5. Sci Rep. 2024 05 30. 14(1): 12463
    L-serine deficiency: on the properties of the Asn133Ser variant of human phosphoserine phosphatase.
    Loredano Pollegioni, Barbara Campanini, Jean-Marc Good, Zoraide Motta, Giulia Murtas, Valeria Buoli Comani, Despina-Christina Pavlidou, Noëlle Mercier, Laureane Mittaz-Crettol, Silvia Sacchi, Francesco Marchesani.
      The non-essential amino acid L-serine is involved in a number of metabolic pathways and in the brain its level is largely due to the biosynthesis from the glycolytic intermediate D-3-phosphoglycerate by the phosphorylated pathway (PP). This cytosolic pathway is made by three enzymes proposed to generate a reversible metabolon named the "serinosome". Phosphoserine phosphatase (PSP) catalyses the last and irreversible step, representing the driving force pushing L-serine synthesis. Genetic defects of the PP enzymes result in strong neurological phenotypes. Recently, we identified the homozygous missense variant [NM_004577.4: c.398A > G p.(Asn133Ser)] in the PSPH, the PSP encoding gene, in two siblings with a neurodevelopmental syndrome and a myelopathy. The recombinant Asn133Ser enzyme does not show significant alterations in protein conformation and dimeric oligomerization state, as well as in enzymatic activity and functionality of the reconstructed PP. However, the Asn133Ser variant is less stable than wild-type PSP, a feature also apparent at cellular level. Studies on patients' fibroblasts also highlight a strong decrease in the level of the enzymes of the PP, a partial nuclear and perinuclear localization of variant PSP and a stronger perinuclear aggregates formation. We propose that these alterations contribute to the formation of a dysfunctional serinosome and thus to the observed reduction of L-serine, glycine and D-serine levels (the latter playing a crucial role in modulating NMDA receptors). The characterization of patients harbouring the Asn133Ser PSP substitution allows to go deep into the molecular mechanisms related to L-serine deficit and to suggest treatments to cope with the observed amino acids alterations.
    Keywords:  Genetic disease; Phosphorylated pathway; Serine deficiency; Structure–function relationships
    DOI:  https://doi.org/10.1038/s41598-024-63164-y
  6. Chem Sci. 2024 May 29. 15(21): 8080-8088
    A mitochondrial-targeted activity-based sensing probe for ratiometric imaging of formaldehyde reveals key regulators of the mitochondrial one-carbon pool.
    Logan Tenney, Vanha N Pham, Thomas F Brewer, Christopher J Chang.
      Formaldehyde (FA) is both a highly reactive environmental genotoxin and an endogenously produced metabolite that functions as a signaling molecule and one-carbon (1C) store to regulate 1C metabolism and epigenetics in the cell. Owing to its signal-stress duality, cells have evolved multiple clearance mechanisms to maintain FA homeostasis, acting to avoid the established genotoxicity of FA while also redirecting FA-derived carbon units into the biosynthesis of essential nucleobases and amino acids. The highly compartmentalized nature of FA exposure, production, and regulation motivates the development of chemical tools that enable monitoring of transient FA fluxes with subcellular resolution. Here we report a mitochondrial-targeted, activity-based sensing probe for ratiometric FA detection, MitoRFAP-2, and apply this reagent to monitor endogenous mitochondrial sources and sinks of this 1C unit. We establish the utility of subcellular localization by showing that MitoRFAP-2 is sensitive enough to detect changes in mitochondrial FA pools with genetic and pharmacological modulation of enzymes involved in 1C and amino acid metabolism, including the pervasive, less active genetic mutant aldehyde dehydrogenase 2*2 (ALDH2*2), where previous, non-targeted versions of FA sensors are not. Finally, we used MitoRFAP-2 to comparatively profile basal levels of FA across a panel of breast cancer cell lines, finding that FA-dependent fluorescence correlates with expression levels of enzymes involved in 1C metabolism. By showcasing the ability of MitoRFAP-2 to identify new information on mitochondrial FA homeostasis, this work provides a starting point for the design of a broader range of chemical probes for detecting physiologically important aldehydes with subcellular resolution and a useful reagent for further studies of 1C biology.
    DOI:  https://doi.org/10.1039/d4sc01183j
  7. Sci Adv. 2024 May 31. 10(22): eadj1431
    13C metabolite tracing reveals glutamine and acetate as critical in vivo fuels for CD8 T cells.
    Eric H Ma, Michael S Dahabieh, Lisa M DeCamp, Irem Kaymak, Susan M Kitchen-Goosen, Brandon M Oswald, Joseph Longo, Dominic G Roy, Mark J Verway, Radia M Johnson, Bozena Samborska, Lauren R Duimstra, Catherine A Scullion, Mya Steadman, Matthew Vos, Thomas P Roddy, Connie M Krawczyk, Kelsey S Williams, Ryan D Sheldon, Russell G Jones.
      Infusion of 13C-labeled metabolites provides a gold standard for understanding the metabolic processes used by T cells during immune responses in vivo. Through infusion of 13C-labeled metabolites (glucose, glutamine, and acetate) in Listeria monocytogenes-infected mice, we demonstrate that CD8 T effector (Teff) cells use metabolites for specific pathways during specific phases of activation. Highly proliferative early Teff cells in vivo shunt glucose primarily toward nucleotide synthesis and leverage glutamine anaplerosis in the tricarboxylic acid (TCA) cycle to support adenosine triphosphate and de novo pyrimidine synthesis. In addition, early Teff cells rely on glutamic-oxaloacetic transaminase 1 (Got1)-which regulates de novo aspartate synthesis-for effector cell expansion in vivo. CD8 Teff cells change fuel preference over the course of infection, switching from glutamine- to acetate-dependent TCA cycle metabolism late in infection. This study provides insights into the dynamics of Teff metabolism, illuminating distinct pathways of fuel consumption associated with CD8 Teff cell function in vivo.
    DOI:  https://doi.org/10.1126/sciadv.adj1431
  8. Sci Rep. 2024 May 31. 14(1): 12521
    Regulation of TSC2 lysosome translocation and mitochondrial turnover by TSC2 acetylation status.
    Patricia Marqués, Jesús Burillo, Carlos González-Blanco, Beatriz Jiménez, Gema García, Ana García-Aguilar, Sarai Iglesias-Fortes, Ángela Lockwood, Carlos Guillén.
      Sirtuin1 (SIRT1) activity decreases the tuberous sclerosis complex 2 (TSC2) lysine acetylation status, inhibiting the mechanistic target of rapamycin complex 1 (mTORC1) signalling and concomitantly, activating autophagy. This study analyzes the role of TSC2 acetylation levels in its translocation to the lysosome and the mitochondrial turnover in both mouse embryonic fibroblast (MEF) and in mouse insulinoma cells (MIN6) as a model of pancreatic β cells. Resveratrol (RESV), an activator of SIRT1 activity, promotes TSC2 deacetylation and its translocation to the lysosome, inhibiting mTORC1 activity. An improvement in mitochondrial turnover was also observed in cells treated with RESV, associated with an increase in the fissioned mitochondria, positive autophagic and mitophagic fluxes and an enhancement of mitochondrial biogenesis. This study proves that TSC2 in its deacetylated form is essential for regulating mTORC1 signalling and the maintenance of the mitochondrial quality control, which is involved in the homeostasis of pancreatic beta cells and prevents from several metabolic disorders such as Type 2 Diabetes Mellitus.
    Keywords:  Acetylation; Lysosome; Mitophagy; Pancreatic β cells; TSC2; mTORC1
    DOI:  https://doi.org/10.1038/s41598-024-63525-7
  9. Nat Commun. 2024 May 29. 15(1): 4549
    Breast cancer cell-secreted miR-199b-5p hijacks neurometabolic coupling to promote brain metastasis.
    Xianhui Ruan, Wei Yan, Minghui Cao, Ray Anthony M Daza, Miranda Y Fong, Kaifu Yang, Jun Wu, Xuxiang Liu, Melanie Palomares, Xiwei Wu, Arthur Li, Yuan Chen, Rahul Jandial, Nicholas C Spitzer, Robert F Hevner, Shizhen Emily Wang.
      Breast cancer metastasis to the brain is a clinical challenge rising in prevalence. However, the underlying mechanisms, especially how cancer cells adapt a distant brain niche to facilitate colonization, remain poorly understood. A unique metabolic feature of the brain is the coupling between neurons and astrocytes through glutamate, glutamine, and lactate. Here we show that extracellular vesicles from breast cancer cells with a high potential to develop brain metastases carry high levels of miR-199b-5p, which shows higher levels in the blood of breast cancer patients with brain metastases comparing to those with metastatic cancer in other organs. miR-199b-5p targets solute carrier transporters (SLC1A2/EAAT2 in astrocytes and SLC38A2/SNAT2 and SLC16A7/MCT2 in neurons) to hijack the neuron-astrocyte metabolic coupling, leading to extracellular retention of these metabolites and promoting cancer cell growth. Our findings reveal a mechanism through which cancer cells of a non-brain origin reprogram neural metabolism to fuel brain metastases.
    DOI:  https://doi.org/10.1038/s41467-024-48740-0
  10. Cell Death Dis. 2024 May 25. 15(5): 361
    Mitochondrial iron deficiency triggers cytosolic iron overload in PKAN hiPS-derived astrocytes.
    Paolo Santambrogio, Anna Cozzi, Chiara Balestrucci, Maddalena Ripamonti, Valeria Berno, Eugenia Cammarota, Andrea Stefano Moro, Sonia Levi.
      Disease models of neurodegeneration with brain iron accumulation (NBIA) offer the possibility to explore the relationship between iron dyshomeostasis and neurodegeneration. We analyzed hiPS-derived astrocytes from PANK2-associated neurodegeneration (PKAN), an NBIA disease characterized by progressive neurodegeneration and high iron accumulation in the globus pallidus. Previous data indicated that PKAN astrocytes exhibit alterations in iron metabolism, general impairment of constitutive endosomal trafficking, mitochondrial dysfunction and acquired neurotoxic features. Here, we performed a more in-depth analysis of the interactions between endocytic vesicles and mitochondria via superresolution microscopy experiments. A significantly lower number of transferrin-enriched vesicles were in contact with mitochondria in PKAN cells than in control cells, confirming the impaired intracellular fate of cargo endosomes. The investigation of cytosolic and mitochondrial iron parameters indicated that mitochondrial iron availability was substantially lower in PKAN cells compared to that in the controls. In addition, PKAN astrocytes exhibited defects in tubulin acetylation/phosphorylation, which might be responsible for unregulated vesicular dynamics and inappropriate iron delivery to mitochondria. Thus, the impairment of iron incorporation into these organelles seems to be the cause of cell iron delocalization, resulting in cytosolic iron overload and mitochondrial iron deficiency, triggering mitochondrial dysfunction. Overall, the data elucidate the mechanism of iron accumulation in CoA deficiency, highlighting the importance of mitochondrial iron deficiency in the pathogenesis of disease.
    DOI:  https://doi.org/10.1038/s41419-024-06757-9
  11. Nat Cell Biol. 2024 May 29.
    Cells dispose of cytoplasmic mitochondrial DNA by nucleoid-phagy.
      
    DOI:  https://doi.org/10.1038/s41556-024-01421-y
  12. Nat Commun. 2024 May 27. 15(1): 4504
    Hairpin protein partitioning from the ER to lipid droplets involves major structural rearrangements.
    Ravi Dhiman, Rehani S Perera, Chetan S Poojari, Haakon T A Wiedemann, Reinhard Kappl, Christopher W M Kay, Jochen S Hub, Bianca Schrul.
      Lipid droplet (LD) function relies on proteins partitioning between the endoplasmic reticulum (ER) phospholipid bilayer and the LD monolayer membrane to control cellular adaptation to metabolic changes. It has been proposed that these hairpin proteins integrate into both membranes in a similar monotopic topology, enabling their passive lateral diffusion during LD emergence at the ER. Here, we combine biochemical solvent-accessibility assays, electron paramagnetic resonance spectroscopy and intra-molecular crosslinking experiments with molecular dynamics simulations, and determine distinct intramembrane positionings of the ER/LD protein UBXD8 in ER bilayer and LD monolayer membranes. UBXD8 is deeply inserted into the ER bilayer with a V-shaped topology and adopts an open-shallow conformation in the LD monolayer. Major structural rearrangements are required to enable ER-to-LD partitioning. Free energy calculations suggest that such structural transition is unlikely spontaneous, indicating that ER-to-LD protein partitioning relies on more complex mechanisms than anticipated and providing regulatory means for this trans-organelle protein trafficking.
    DOI:  https://doi.org/10.1038/s41467-024-48843-8
  13. Redox Biol. 2024 May 25. pii: S2213-2317(24)00191-5. [Epub ahead of print]73 103213
    Cystine/glutamate antiporter xCT controls skeletal muscle glutathione redox, bioenergetics and differentiation.
    Michel N Kanaan, Chantal A Pileggi, Charbel Y Karam, Luke S Kennedy, Claire Fong-McMaster, Miroslava Cuperlovic-Culf, Mary-Ellen Harper.
      Cysteine, the rate-controlling amino acid in cellular glutathione synthesis is imported as cystine, by the cystine/glutamate antiporter, xCT, and subsequently reduced to cysteine. As glutathione redox is important in muscle regeneration in aging, we hypothesized that xCT exerts upstream control over skeletal muscle glutathione redox, metabolism and regeneration. Bioinformatic analyses of publicly available datasets revealed that expression levels of xCT and GSH-related genes are inversely correlated with myogenic differentiation genes. Muscle satellite cells (MuSCs) isolated from Slc7a11sut/sut mice, which harbour a mutation in the Slc7a11 gene encoding xCT, required media supplementation with 2-mercaptoethanol to support cell proliferation but not myotube differentiation, despite persistently lower GSH. Slc7a11sut/sut primary myotubes were larger compared to WT myotubes, and also exhibited higher glucose uptake and cellular oxidative capacities. Immunostaining of myogenic markers (Pax7, MyoD, and myogenin) in cardiotoxin-damaged tibialis anterior muscle fibres revealed greater MuSC activation and commitment to differentiation in Slc7a11sut/sut muscle compared to WT mice, culminating in larger myofiber cross-sectional areas at 21 days post-injury. Slc7a11sut/sut mice subjected to a 5-week exercise training protocol demonstrated enhanced insulin tolerance compared to WT mice, but blunted muscle mitochondrial biogenesis and respiration in response to exercise training. Our results demonstrate that the absence of xCT inhibits cell proliferation but promotes myotube differentiation by regulating cellular metabolism and glutathione redox. Altogether, these results support the notion that myogenesis is a redox-regulated process and may help inform novel therapeutic approaches for muscle wasting and dysfunction in aging and disease.
    Keywords:  Glutathione; Glycolysis; Mitochondria; Myogenesis; Oxidative phosphorylation; Redox
    DOI:  https://doi.org/10.1016/j.redox.2024.103213
  14. bioRxiv. 2024 May 13. pii: 2024.05.09.593171. [Epub ahead of print]
    Mechanical activation of VE-cadherin stimulates AMPK to increase endothelial cell metabolism and vasodilation.
    Nicholas M Cronin, Logan W Dawson, Kris A DeMali.
      Endothelia cells respond to mechanical force by stimulating cellular signaling, but how these pathways are linked to elevations in cell metabolism and whether metabolism supports the mechanical response remains poorly understood. Here, we show that application of force to VE-cadherin stimulates liver kinase B1 (LKB1) to activate AMP-activated protein kinase (AMPK), a master regulator of energy homeostasis. VE-cadherin stimulated AMPK increases eNOS activity and localization to the plasma membrane as well as reinforcement of the actin cytoskeleton and cadherin adhesion complex, and glucose uptake. We present evidence for the increase in metabolism being necessary to fortify the adhesion complex, actin cytoskeleton, and cellular alignment. Together these data extend the paradigm for how mechanotransduction and metabolism are linked to include a connection to vasodilation, thereby providing new insight into how diseases involving contractile, metabolic, and vasodilatory disturbances arise.
    DOI:  https://doi.org/10.1101/2024.05.09.593171
  15. Nat Nanotechnol. 2024 May 27.
    Oral mitochondrial transplantation using nanomotors to treat ischaemic heart disease.
    Ziyu Wu, Lin Chen, Wenyan Guo, Jun Wang, Haiya Ni, Jianing Liu, Wentao Jiang, Jian Shen, Chun Mao, Min Zhou, Mimi Wan.
      Mitochondrial transplantation is an important therapeutic strategy for restoring energy supply in patients with ischaemic heart disease (IHD); however, it is limited by the invasiveness of the transplantation method and loss of mitochondrial activity. Here we report successful mitochondrial transplantation by oral administration for IHD therapy. A nitric-oxide-releasing nanomotor is modified on the mitochondria surface to obtain nanomotorized mitochondria with chemotactic targeting ability towards damaged heart tissue due to nanomotor action. The nanomotorized mitochondria are packaged in enteric capsules to protect them from gastric acid erosion. After oral delivery the mitochondria are released in the intestine, where they are quickly absorbed by intestinal cells and secreted into the bloodstream, allowing delivery to the damaged heart tissue. The regulation of disease microenvironment by the nanomotorized mitochondria can not only achieve rapid uptake and high retention of mitochondria by damaged cardiomyocytes but also maintains high activity of the transplanted mitochondria. Furthermore, results from animal models of IHD indicate that the accumulated nanomotorized mitochondria in the damaged heart tissue can regulate cardiac metabolism at the transcriptional level, thus preventing IHD progression. This strategy has the potential to change the therapeutic strategy used to treat IHD.
    DOI:  https://doi.org/10.1038/s41565-024-01681-7
  16. bioRxiv. 2024 May 18. pii: 2024.05.15.594133. [Epub ahead of print]
    CPT2 mediated fatty acid oxidation is dispensable for humoral immunity.
    Meilu Li, Xian Zhou, Xingxing Zhu, Yanfeng Li, Taro Hitosugi, Yuzhen Li, Hu Zeng.
      B cell activation is accompanied by dynamic metabolic reprogramming, supported by a multitude of nutrients that include glucose, amino acids and fatty acids. While several studies have indicated that fatty acid mitochondrial oxidation is critical for immune cell functions, contradictory findings have been reported. Carnitine palmitoyltransferase II (CPT2) is a critical enzyme for long-chain fatty acid oxidation in mitochondria. Here, we test the requirement of CPT2 for humoral immunity using a mouse model with a lymphocyte specific deletion of CPT2. Stable 13 C isotope tracing reveals highly reduced fatty acid-derived citrate production in CPT2 deficient B cells. Yet, CPT2 deficiency has no significant impact on B cell development, B cell activation, germinal center formation, and antibody production upon either thymus-dependent or -independent antigen challenges. Together, our findings indicate that CPT2 mediated fatty acid oxidation is dispensable for humoral immunity, highlighting the metabolic flexibility of lymphocytes.
    DOI:  https://doi.org/10.1101/2024.05.15.594133
  17. Nature. 2024 May 29.
    High-resolution in situ structures of mammalian respiratory supercomplexes.
    Wan Zheng, Pengxin Chai, Jiapeng Zhu, Kai Zhang.
      Mitochondria play a pivotal part in ATP energy production through oxidative phosphorylation, which occurs within the inner membrane through a series of respiratory complexes1-4. Despite extensive in vitro structural studies, determining the atomic details of their molecular mechanisms in physiological states remains a major challenge, primarily because of loss of the native environment during purification. Here we directly image porcine mitochondria using an in situ cryo-electron microscopy approach. This enables us to determine the structures of various high-order assemblies of respiratory supercomplexes in their native states. We identify four main supercomplex organizations: I1III2IV1, I1III2IV2, I2III2IV2 and I2III4IV2, which potentially expand into higher-order arrays on the inner membranes. These diverse supercomplexes are largely formed by 'protein-lipids-protein' interactions, which in turn have a substantial impact on the local geometry of the surrounding membranes. Our in situ structures also capture numerous reactive intermediates within these respiratory supercomplexes, shedding light on the dynamic processes of the ubiquinone/ubiquinol exchange mechanism in complex I and the Q-cycle in complex III. Structural comparison of supercomplexes from mitochondria treated under different conditions indicates a possible correlation between conformational states of complexes I and III, probably in response to environmental changes. By preserving the native membrane environment, our approach enables structural studies of mitochondrial respiratory supercomplexes in reaction at high resolution across multiple scales, from atomic-level details to the broader subcellular context.
    DOI:  https://doi.org/10.1038/s41586-024-07488-9
  18. bioRxiv. 2024 May 15. pii: 2024.05.14.592021. [Epub ahead of print]
    Movement of the endoplasmic reticulum is driven by multiple classes of vesicles marked by Rab-GTPases.
    Allison Langley, Sarah Abeling-Wang, Erinn Wagner, John Salogiannis.
      Peripheral endoplasmic reticulum (ER) tubules move along microtubules to interact with various organelles through membrane contact sites (MCS). Traditionally, ER moves by either sliding along stable microtubules via molecular motors or attaching to the plus ends of dynamic microtubules through tip attachment complexes (TAC). A recently discovered third process, hitchhiking, involves motile vesicles pulling ER tubules along microtubules. Previous research showed that ER hitchhikes on Rab5- and Rab7-marked endosomes, but it is uncertain if other Rab-vesicles can do the same. In U2OS cells, we screened Rabs for their ability to cotransport with ER tubules and found that ER hitchhikes on post-Golgi vesicles marked by Rab6 (isoforms a and b). Rab6-ER hitchhiking occurs independently of ER-endolysosome contacts and TAC-mediated ER movement. Disrupting either Rab6 or the motility of Rab6-vesicles reduces overall ER movement. Conversely, relocating these vesicles to the cell periphery causes peripheral ER accumulation, indicating that Rab6-vesicle motility is crucial for a subset of ER movements. Proximal post-Golgi vesicles marked by TGN46 are involved in Rab6-ER hitchhiking, while other post-Golgi vesicles (Rabs 8/10/11/13/14) are not essential for ER movement. Our further analysis finds that ER to Golgi vesicles marked by Rab1 are also capable of driving a subset of ER movements. Taken together, our findings suggest that ER hitchhiking on Rab-vesicles is a significant mode of ER movement.
    SIGNIFICANCE STATEMENT: Peripheral endoplasmic reticulum tubules move on microtubules by either attaching to motors (cargo adaptor-mediated), dynamic microtubule-plus ends (tip attachment complexes) or motile vesicles (hitchhiking) but the prevalence of each mode is not clearPost-Golgi vesicles marked by Rab6/TGN46 and ER to Golgi vesicles marked by Rab1 drive ER movementsER hitchhiking on multiple classes of vesicles (endolysosomal, post-Golgi and ER to Golgi) marked by Rabs plays a prominent role in ER movement.
    DOI:  https://doi.org/10.1101/2024.05.14.592021
  19. Nat Commun. 2024 May 25. 15(1): 4469
    Rab30 facilitates lipid homeostasis during fasting.
    Danielle M Smith, Brian Y Liu, Michael J Wolfgang.
      To facilitate inter-tissue communication and the exchange of proteins, lipoproteins, and metabolites with the circulation, hepatocytes have an intricate and efficient intracellular trafficking system regulated by small Rab GTPases. Here, we show that Rab30 is induced in the mouse liver by fasting, which is amplified in liver-specific carnitine palmitoyltransferase 2 knockout mice (Cpt2L-/-) lacking the ability to oxidize fatty acids, in a Pparα-dependent manner. Live-cell super-resolution imaging and in vivo proximity labeling demonstrates that Rab30-marked vesicles are highly dynamic and interact with proteins throughout the secretory pathway. Rab30 whole-body, liver-specific, and Rab30; Cpt2 liver-specific double knockout (DKO) mice are viable with intact Golgi ultrastructure, although Rab30 deficiency in DKO mice suppresses the serum dyslipidemia observed in Cpt2L-/- mice. Corresponding with decreased serum triglyceride and cholesterol levels, DKO mice exhibit decreased circulating but not hepatic ApoA4 protein, indicative of a trafficking defect. Together, these data suggest a role for Rab30 in the selective sorting of lipoproteins to influence hepatocyte and circulating triglyceride levels, particularly during times of excessive lipid burden.
    DOI:  https://doi.org/10.1038/s41467-024-48959-x
  20. Cell Death Dis. 2024 May 31. 15(5): 382
    Targeting the autophagy-NAD axis protects against cell death in Niemann-Pick type C1 disease models.
    Tetsushi Kataura, Lucia Sedlackova, Congxin Sun, Gamze Kocak, Niall Wilson, Peter Banks, Faisal Hayat, Sergey Trushin, Eugenia Trushina, Oliver D K Maddocks, John E Oblong, Satomi Miwa, Masaya Imoto, Shinji Saiki, Daniel Erskine, Marie E Migaud, Sovan Sarkar, Viktor I Korolchuk.
      Impairment of autophagy leads to an accumulation of misfolded proteins and damaged organelles and has been implicated in plethora of human diseases. Loss of autophagy in actively respiring cells has also been shown to trigger metabolic collapse mediated by the depletion of nicotinamide adenine dinucleotide (NAD) pools, resulting in cell death. Here we found that the deficit in the autophagy-NAD axis underpins the loss of viability in cell models of a neurodegenerative lysosomal storage disorder, Niemann-Pick type C1 (NPC1) disease. Defective autophagic flux in NPC1 cells resulted in mitochondrial dysfunction due to impairment of mitophagy, leading to the depletion of both the reduced and oxidised forms of NAD as identified via metabolic profiling. Consequently, exhaustion of the NAD pools triggered mitochondrial depolarisation and apoptotic cell death. Our chemical screening identified two FDA-approved drugs, celecoxib and memantine, as autophagy activators which effectively restored autophagic flux, NAD levels, and cell viability of NPC1 cells. Of biomedical relevance, either pharmacological rescue of the autophagy deficiency or NAD precursor supplementation restored NAD levels and improved the viability of NPC1 patient fibroblasts and induced pluripotent stem cell (iPSC)-derived cortical neurons. Together, our findings identify the autophagy-NAD axis as a mechanism of cell death and a target for therapeutic interventions in NPC1 disease, with a potential relevance to other neurodegenerative disorders.
    DOI:  https://doi.org/10.1038/s41419-024-06770-y
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