bims-mimbat Biomed News
on Mitochondrial metabolism in brown adipose tissue
Issue of 2023–02–26
twelve papers selected by
José Carlos de Lima-Júnior, Washington University
  1. Mitochondrial phosphatidylethanolamine modulates UCP1 to promote brown adipose thermogenesis.
  2. Stimulation of the beta-2-adrenergic receptor with salbutamol activates human brown adipose tissue.
  3. The SLC25A47 locus controls gluconeogenesis and energy expenditure.
  4. Differentiation and Imaging of Brown Adipocytes from the Stromal Vascular Fraction of Interscapular Adipose Tissue from Newborn Mice.
  5. Mitochondrial membrane biogenesis: A new pathway for lipid transport mediated by PERK/E-Syt1 complex.
  6. Chemical inhibition of mitochondrial fission via targeting the DRP1-receptor interaction.
  7. The Energetics and Ion Coupling of Cholesterol Transport Through Patched1.
  8. Effects of hepatic mitochondrial pyruvate carrier deficiency on de novo lipogenesis and glycerol-mediated gluconeogenesis in mice.
  9. Mitochondrial Peroxiredoxin 3 Is Rapidly Oxidized and Hyperoxidized by Fatty Acid Hydroperoxides.
  10. PERK recruits E-Syt1 at ER-mitochondria contacts for mitochondrial lipid transport and respiration.
  11. Inhibition of Mitochondrial Uncoupling Proteins Arrests Human Spermatozoa Motility without Compromising Viability.
  12. Isolating Mitochondria, Mitoplasts, and mtDNA from Cultured Mammalian Cells.
  1. Sci Adv. 2023 Feb 24. 9(8): eade7864
    Mitochondrial phosphatidylethanolamine modulates UCP1 to promote brown adipose thermogenesis.
    Jordan M Johnson, Alek D Peterlin, Enrique Balderas, Elahu G Sustarsic, J Alan Maschek, Marisa J Lang, Alejandro Jara-Ramos, Vanja Panic, Jeffrey T Morgan, Claudio J Villanueva, Alejandro Sanchez, Jared Rutter, Irfan J Lodhi, James E Cox, Kelsey H Fisher-Wellman, Dipayan Chaudhuri, Zachary Gerhart-Hines, Katsuhiko Funai.
      Thermogenesis by uncoupling protein 1 (UCP1) is one of the primary mechanisms by which brown adipose tissue (BAT) increases energy expenditure. UCP1 resides in the inner mitochondrial membrane (IMM), where it dissipates membrane potential independent of adenosine triphosphate (ATP) synthase. Here, we provide evidence that phosphatidylethanolamine (PE) modulates UCP1-dependent proton conductance across the IMM to modulate thermogenesis. Mitochondrial lipidomic analyses revealed PE as a signature molecule whose abundance bidirectionally responds to changes in thermogenic burden. Reduction in mitochondrial PE by deletion of phosphatidylserine decarboxylase (PSD) made mice cold intolerant and insensitive to β3 adrenergic receptor agonist-induced increase in whole-body oxygen consumption. High-resolution respirometry and fluorometry of BAT mitochondria showed that loss of mitochondrial PE specifically lowers UCP1-dependent respiration without compromising electron transfer efficiency or ATP synthesis. These findings were confirmed by a reduction in UCP1 proton current in PE-deficient mitoplasts. Thus, PE performs a previously unknown role as a temperature-responsive rheostat that regulates UCP1-dependent thermogenesis.
    DOI:  https://doi.org/10.1126/sciadv.ade7864
  2. Cell Rep Med. 2023 Feb 21. pii: S2666-3791(23)00034-4. [Epub ahead of print]4(2): 100942
    Stimulation of the beta-2-adrenergic receptor with salbutamol activates human brown adipose tissue.
    Maaike E Straat, Carlijn A Hoekx, Floris H P van Velden, Lenka M Pereira Arias-Bouda, Lauralyne Dumont, Denis P Blondin, Mariëtte R Boon, Borja Martinez-Tellez, Patrick C N Rensen.
      While brown adipose tissue (BAT) is activated by the beta-3-adrenergic receptor (ADRB3) in rodents, in human brown adipocytes, the ADRB2 is dominantly present and responsible for noradrenergic activation. Therefore, we performed a randomized double-blinded crossover trial in young lean men to compare the effects of single intravenous bolus of the ADRB2 agonist salbutamol without and with the ADRB1/2 antagonist propranolol on glucose uptake by BAT, assessed by dynamic 2-[18F]fluoro-2-deoxy-D-glucose positron emission tomography-computed tomography scan (i.e., primary outcome). Salbutamol, compared with salbutamol with propranolol, increases glucose uptake by BAT, without affecting the glucose uptake by skeletal muscle and white adipose tissue. The salbutamol-induced glucose uptake by BAT positively associates with the increase in energy expenditure. Notably, participants with high salbutamol-induced glucose uptake by BAT have lower body fat mass, waist-hip ratio, and serum LDL-cholesterol concentration. In conclusion, specific ADRB2 agonism activates human BAT, which warrants investigation of ADRB2 activation in long-term studies (EudraCT: 2020-004059-34).
    Keywords:  brown fat; cardiometabolic diseases; energy expenditure; glucose metabolism; indirect calorimetry; lipoprotein metabolism; positron emission tomography; propranolol; responders
    DOI:  https://doi.org/10.1016/j.xcrm.2023.100942
  3. Proc Natl Acad Sci U S A. 2023 Feb 28. 120(9): e2216810120
    The SLC25A47 locus controls gluconeogenesis and energy expenditure.
    Jin-Seon Yook, Zachary H Taxin, Bo Yuan, Satoshi Oikawa, Christopher Auger, Beste Mutlu, Pere Puigserver, Sheng Hui, Shingo Kajimura.
      Mitochondria provide essential metabolites and adenosine triphosphate (ATP) for the regulation of energy homeostasis. For instance, liver mitochondria are a vital source of gluconeogenic precursors under a fasted state. However, the regulatory mechanisms at the level of mitochondrial membrane transport are not fully understood. Here, we report that a liver-specific mitochondrial inner-membrane carrier SLC25A47 is required for hepatic gluconeogenesis and energy homeostasis. Genome-wide association studies found significant associations between SLC25A47 and fasting glucose, HbA1c, and cholesterol levels in humans. In mice, we demonstrated that liver-specific depletion of SLC25A47 impaired hepatic gluconeogenesis selectively from lactate, while significantly enhancing whole-body energy expenditure and the hepatic expression of FGF21. These metabolic changes were not a consequence of general liver dysfunction because acute SLC25A47 depletion in adult mice was sufficient to enhance hepatic FGF21 production, pyruvate tolerance, and insulin tolerance independent of liver damage and mitochondrial dysfunction. Mechanistically, SLC25A47 depletion leads to impaired hepatic pyruvate flux and malate accumulation in the mitochondria, thereby restricting hepatic gluconeogenesis. Together, the present study identified a crucial node in the liver mitochondria that regulates fasting-induced gluconeogenesis and energy homeostasis.
    Keywords:  bioenergetics; metabolism; mitochondria; obesity; type 2 diabetes
    DOI:  https://doi.org/10.1073/pnas.2216810120
  4. J Vis Exp. 2023 Feb 03.
    Differentiation and Imaging of Brown Adipocytes from the Stromal Vascular Fraction of Interscapular Adipose Tissue from Newborn Mice.
    Ana-María Figueroa, Francisca Stolzenbach, Pablo Tapia, Víctor Cortés.
      Brown adipose tissue (BAT) is only present in mammals and has a thermogenic function. Brown adipocytes are characterized by a multilocular cytoplasm with multiple lipid droplets, a central nucleus, a high mitochondrial content, and the expression of uncoupling protein 1 (UCP1). BAT has been proposed as a potential therapeutic target for obesity and its associated metabolic disorders due to its ability to dissipate metabolic energy as heat. To investigate BAT function and regulation, brown adipocyte culturing is indispensable. The present protocol optimizes tissue processing and cell differentiation for culturing brown adipocytes from newborn mice. Additionally, procedures for the imaging of differentiated adipocytes with both confocal immunofluorescence and transmission electron microscopy are shown. In the brown adipocytes differentiated with the techniques described herein, the major defining features of classical BAT are preserved, including high UCP1 levels, increased mitochondrial mass, and very close physical contact between the lipid droplets and mitochondria, making this method a valuable tool for BAT studies.
    DOI:  https://doi.org/10.3791/64604
  5. J Cell Biol. 2023 Mar 06. pii: e202301132. [Epub ahead of print]222(3):
    Mitochondrial membrane biogenesis: A new pathway for lipid transport mediated by PERK/E-Syt1 complex.
    Sébastien Leterme, Morgane Michaud.
      Despite decades of extensive research, mitochondrial lipid transport is a process far from fully understood. In this issue, Sassano et al. (2023. J. Cell Biol.https://doi.org/10.1083/jcb.202206008) identified a new complex, composed of E-Syt1 and PERK, which mediates lipid transport at ER-mitochondria contact sites and regulates mitochondrial functions in human cells.
    DOI:  https://doi.org/10.1083/jcb.202301132
  6. Cell Chem Biol. 2023 Feb 17. pii: S2451-9456(23)00033-8. [Epub ahead of print]
    Chemical inhibition of mitochondrial fission via targeting the DRP1-receptor interaction.
    Jun Yang, Peihao Chen, Yu Cao, Shanshan Liu, Wei Wang, Lin Li, Jiaojiao Li, Zhaodi Jiang, Yan Ma, She Chen, Sanduo Zheng, Xiangbing Qi, Hui Jiang.
      Mitochondrial fission is critical for mitochondrial dynamics and homeostasis. The dynamin superfamily GTPase DRP1 is recruited by three functionally redundant receptors, MFF, MiD49, and MiD51, to mitochondria to drive fission. Here, we exploit high-content live-cell imaging to screen for mitochondrial fission inhibitors and have developed a covalent compound, mitochondrial division inhibitor (MIDI). MIDI treatment potently blocks mitochondrial fragmentation induced by mitochondrial toxins and restores mitochondrial morphology in fusion-defective cells carrying pathogenic mitofusin and OPA1 mutations. Mechanistically, MIDI does not affect DRP1 tetramerization nor DRP1 GTPase activity but does block DRP1 recruitment to mitochondria. Subsequent biochemical and cellular characterizations reveal an unexpected mechanism that MIDI targets DRP1 interaction with multiple receptors via covalent interaction with DRP1-C367. Taken together, beyond developing a potent mitochondrial fission inhibitor that profoundly impacts mitochondrial morphogenesis, our study establishes proof of concept for developing protein-protein interaction inhibitors targeting DRP1.
    Keywords:  DRP1 inhibitor; MFF; MIDI; MiD49/51; OPA1; mitochondrial dynamics; mitochondrial fission; mitofusin
    DOI:  https://doi.org/10.1016/j.chembiol.2023.02.002
  7. bioRxiv. 2023 Feb 15. pii: 2023.02.14.528445. [Epub ahead of print]
    The Energetics and Ion Coupling of Cholesterol Transport Through Patched1.
    T Bertie Ansell, Robin A Corey, Lucrezia Vittoria Viti, Maia Kinnebrew, Rajat Rohatgi, Christian Siebold, Mark S P Sansom.
      Patched1 (PTCH1) is the principal tumour suppressor protein of the mammalian Hedgehog (HH) signalling pathway, implicated in embryogenesis and tissue homeostasis. PTCH1 inhibits the Class F G protein-coupled receptor Smoothened (SMO) via a debated mechanism involving modulating accessible cholesterol levels within ciliary membranes. Using extensive molecular dynamics (MD) simulations and free energy calculations to evaluate cholesterol transport through PTCH1, we find an energetic barrier of ~15-20 kJ mol -1 for cholesterol export. In simulations we identify cation binding sites within the PTCH1 transmembrane domain (TMD) which may provide the energetic impetus for cholesterol transport. In silico data are coupled to in vivo biochemical assays of PTCH1 mutants to probe coupling between transmembrane motions and PTCH1 activity. Using complementary simulations of Dispatched1 (DISP1) we find that transition between 'inward-open' and solvent 'occluded' states is accompanied by Na + induced pinching of intracellular helical segments. Thus, our findings illuminate the energetics and ion-coupling stoichiometries of PTCH1 transport mechanisms, whereby 1-3 Na + or 2-3 K + couple to cholesterol export, and provide the first molecular description of transitions between distinct transport states.
    DOI:  https://doi.org/10.1101/2023.02.14.528445
  8. bioRxiv. 2023 Feb 18. pii: 2023.02.17.528992. [Epub ahead of print]
    Effects of hepatic mitochondrial pyruvate carrier deficiency on de novo lipogenesis and glycerol-mediated gluconeogenesis in mice.
    Nicole K H Yiew, Daniel Ferguson, Kevin Cho, Stanislaw Deja, Chaowapong Jarasvaraparn, Andrew J Lutkewitte, Sandip Mukherjee, Xiaorong Fu, Jason M Singer, Gary J Patti, Shawn C Burgess, Brian N Finck.
      The liver coordinates the systemic response to nutrient deprivation and availability by producing glucose from gluconeogenesis during fasting and synthesizing lipids via de novo lipogenesis (DNL) when carbohydrates are abundant. Mitochondrial pyruvate metabolism is believed to play important roles in both gluconeogenesis and DNL. We examined the effects of hepatocyte-specific mitochondrial pyruvate carrier (MPC) deletion on the fasting-refeeding response. Rates of DNL during refeeding were impaired by liver MPC deletion, but this did not reduce intrahepatic lipid content. During fasting, glycerol is converted to glucose by two pathways; a cytosolic pathway essentially reversing glycolysis and a mitochondrial pathway requiring the MPC. MPC deletion reduced the incorporation of 13 C-glycerol into TCA cycle metabolites but not into newly synthesized glucose. However, suppression of glycerol metabolism did not affect glucose concentrations in fasted hepatocyte-specific MPC-deficient mice. Thus, glycerol-mediated gluconeogenesis and glucose production by kidney and intestine may compensate for MPC deficiency in hepatocytes.
    DOI:  https://doi.org/10.1101/2023.02.17.528992
  9. Antioxidants (Basel). 2023 Feb 07. pii: 408. [Epub ahead of print]12(2):
    Mitochondrial Peroxiredoxin 3 Is Rapidly Oxidized and Hyperoxidized by Fatty Acid Hydroperoxides.
    Giuliana Cardozo, Mauricio Mastrogiovanni, Ari Zeida, Nicolás Viera, Rafael Radi, Aníbal M Reyes, Madia Trujillo.
      Human peroxiredoxin 3 (HsPrx3) is a thiol-based peroxidase responsible for the reduction of most hydrogen peroxide and peroxynitrite formed in mitochondria. Mitochondrial disfunction can lead to membrane lipoperoxidation, resulting in the formation of lipid-bound fatty acid hydroperoxides (LFA-OOHs) which can be released to become free fatty acid hydroperoxides (fFA-OOHs). Herein, we report that HsPrx3 is oxidized and hyperoxidized by fFA-OOHs including those derived from arachidonic acid and eicosapentaenoic acid peroxidation at position 15 with remarkably high rate constants of oxidation (>3.5 × 107 M-1s-1) and hyperoxidation (~2 × 107 M-1s-1). The endoperoxide-hydroperoxide PGG2, an intermediate in prostanoid synthesis, oxidized HsPrx3 with a similar rate constant, but was less effective in causing hyperoxidation. Biophysical methodologies suggest that HsPrx3 can bind hydrophobic structures. Indeed, molecular dynamic simulations allowed the identification of a hydrophobic patch near the enzyme active site that can allocate the hydroperoxide group of fFA-OOHs in close proximity to the thiolate in the peroxidatic cysteine. Simulations performed using available and herein reported kinetic data indicate that HsPrx3 should be considered a main target for mitochondrial fFA-OOHs. Finally, kinetic simulation analysis support that mitochondrial fFA-OOHs formation fluxes in the range of nM/s are expected to contribute to HsPrx3 hyperoxidation, a modification that has been detected in vivo under physiological and pathological conditions.
    Keywords:  antioxidant systems; fatty acid hydroperoxide; kinetics; lipid peroxidation; mitochondria; peroxiredoxin
    DOI:  https://doi.org/10.3390/antiox12020408
  10. J Cell Biol. 2023 Mar 06. pii: e202206008. [Epub ahead of print]222(3):
    PERK recruits E-Syt1 at ER-mitochondria contacts for mitochondrial lipid transport and respiration.
    Maria Livia Sassano, Alexander R van Vliet, Ellen Vervoort, Sofie Van Eygen, Chris Van den Haute, Benjamin Pavie, Joris Roels, Johannes V Swinnen, Marco Spinazzi, Leen Moens, Kristina Casteels, Isabelle Meyts, Paolo Pinton, Saverio Marchi, Leila Rochin, Francesca Giordano, Blanca Felipe-Abrio, Patrizia Agostinis.
      The integrity of ER-mitochondria appositions ensures transfer of ions and phospholipids (PLs) between these organelles and exerts crucial effects on mitochondrial bioenergetics. Malfunctions within the ER-mitochondria contacts altering lipid trafficking homeostasis manifest in diverse pathologies, but the molecular effectors governing this process remain ill-defined. Here, we report that PERK promotes lipid trafficking at the ER-mitochondria contact sites (EMCS) through a non-conventional, unfolded protein response-independent, mechanism. PERK operates as an adaptor for the recruitment of the ER-plasma membrane tether and lipid transfer protein (LTP) Extended-Synaptotagmin 1 (E-Syt1), within the EMCS. In resting cells, the heterotypic E-Syt1-PERK interaction endorses transfer of PLs between the ER and mitochondria. Weakening the E-Syt1-PERK interaction or removing the lipid transfer SMP-domain of E-Syt1, compromises mitochondrial respiration. Our findings unravel E-Syt1 as a PERK interacting LTP and molecular component of the lipid trafficking machinery of the EMCS, which critically maintains mitochondrial homeostasis and fitness.
    DOI:  https://doi.org/10.1083/jcb.202206008
  11. Antioxidants (Basel). 2023 Feb 08. pii: 409. [Epub ahead of print]12(2):
    Inhibition of Mitochondrial Uncoupling Proteins Arrests Human Spermatozoa Motility without Compromising Viability.
    David F Carrageta, Laís Freire-Brito, Bárbara Guerra-Carvalho, João C Ribeiro, Bruno S Monteiro, Alberto Barros, Pedro F Oliveira, Mariana P Monteiro, Marco G Alves.
      Mitochondrial uncoupling proteins (UCPs) are central in the regulation of mitochondrial activity and reactive oxygen species (ROS) production. High oxidative stress is a major cause of male infertility; however, UCPs expression and function in human spermatozoa are still unknown. Herein, we aimed to assess the expression and function of the different homologs (UCP1-6) in human spermatozoa. For this purpose, we screened for the mRNA expression of all UCP homologs. Protein expression and immunolocalization of UCP1, UCP2, and UCP3 were also assessed. Highly motile spermatozoa were isolated from human normozoospermic seminal samples (n = 16) and incubated with genipin, an inhibitor of UCPs (0, 0.5, 5, and 50 µM) for 3 h at 37 °C. Viability and total motility were assessed. Mitochondrial membrane potential and ROS production were evaluated. Media were collected and the metabolic profile and antioxidant potential were analyzed by 1H-NMR and FRAP, respectively. The expression of all UCP homologs (UCP1-6) mRNA by human spermatozoa is herein reported for the first time. UCP1-3 are predominant at the head equatorial segment, whereas UCP1 and UCP2 are also expressed at the spermatozoa midpiece, where mitochondria are located. The inhibition of UCPs by 50 µM genipin, resulting in the UCP3 inhibition, did not compromise sperm cell viability but resulted in irreversible total motility loss that persisted despite washing or incubation with theophylline, a cAMP activator. These effects were associated with decreased mitochondrial membrane potential and lactate production. No differences concerning UCP3 expression, however, were observed in spermatozoa from normozoospermic versus asthenozoospermic men (n = 6). The inhibition of UCPs did not increase ROS production, possibly due to the decreased mitochondrial activity and genipin antioxidant properties. In sum, UCPs are major regulators of human spermatozoa motility and metabolism. The discovery and characterization of UCPs' role in human spermatozoa can shed new light on spermatozoa ROS-related pathways and bioenergetics physiology.
    Keywords:  UCPs; genipin; mitochondria; mitochondrial uncoupling proteins; motility; oxidative stress; spermatozoa
    DOI:  https://doi.org/10.3390/antiox12020409
  12. Methods Mol Biol. 2023 ;2615 17-30
    Isolating Mitochondria, Mitoplasts, and mtDNA from Cultured Mammalian Cells.
    Katja E Menger, Thomas J Nicholls.
      Mitochondria are double membrane-bound eukaryotic organelles with roles in a range of cellular activities including energy conversion, apoptosis, cell signalling, and the biosynthesis of enzyme cofactors. Mitochondria contain their own genome, called mtDNA, which encodes subunits of the oxidative phosphorylation machinery as well as the rRNA and tRNA molecules required for their translation within mitochondria. The ability to isolate highly purified mitochondria from cells has been instrumental in a number of studies of mitochondrial function. Differential centrifugation is a long-established method for the isolation of mitochondria. Cells are subjected to osmotic swelling and disruption, followed by centrifugation in isotonic sucrose solutions to separate mitochondria from other cellular components. We present a method using this principle for the isolation of mitochondria from cultured mammalian cell lines. Mitochondria purified by this method can be further fractionated to investigate protein localization, or act as a starting point to purify mtDNA.
    Keywords:  DNA extraction; Mitochondria; Mitochondrial DNA; Mitochondrial isolation; Protein localization
    DOI:  https://doi.org/10.1007/978-1-0716-2922-2_2
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