bims-mitdyn Biomed News
on Mitochondrial dynamics: mechanisms
Issue of 2023–01–08
thirteen papers selected by
Edmond Chan, Queen’s University, School of Medicine
  1. Potent ClpP agonists with anticancer properties bind with improved structural complementarity and alter the mitochondrial N-terminome.
  2. A multipoint guidance mechanism for β-barrel folding on the SAM complex.
  3. Exercise preserves physical fitness during aging through AMPK and mitochondrial dynamics.
  4. Functional crosstalk between the TIM22 complex and YME1 machinery maintains mitochondrial proteostasis and integrity.
  5. The endoplasmic reticulum kinase PERK interacts with the oxidoreductase ERO1 to metabolically adapt mitochondria.
  6. The mitochondrial Hsp70 controls the assembly of the F1FO-ATP synthase.
  7. Human mitochondria require mtRF1 for translation termination at non-canonical stop codons.
  8. Endoplasmic reticulum protein BIK binds to and inhibits mitochondria-localized anti-apoptotic proteins.
  9. Lactate metabolism is essential in early-onset mitochondrial myopathy.
  10. The nucleoprotein of influenza A virus inhibits the innate immune response by inducing mitophagy.
  11. Inner mitochondrial membrane protein Prohibitin 1 mediates Nix-induced, Parkin-independent mitophagy.
  12. Fluorescent pulse-chase labeling to monitor long-term mitochondrial degradation in primary hippocampal neurons.
  13. Monitoring mitochondrial translation by pulse SILAC.
  1. Structure. 2022 Dec 15. pii: S0969-2126(22)00488-9. [Epub ahead of print]
    Potent ClpP agonists with anticancer properties bind with improved structural complementarity and alter the mitochondrial N-terminome.
    Mark F Mabanglo, Keith S Wong, Marim M Barghash, Elisa Leung, Stephanie H W Chuang, Afshan Ardalan, Emily M Majaesic, Cassandra J Wong, Shen Zhang, Henk Lang, Donald S Karanewsky, Andrew A Iwanowicz, Lee M Graves, Edwin J Iwanowicz, Anne-Claude Gingras, Walid A Houry.
      The mitochondrial ClpP protease is responsible for mitochondrial protein quality control through specific degradation of proteins involved in several metabolic processes. ClpP overexpression is also required in many cancer cells to eliminate reactive oxygen species (ROS)-damaged proteins and to sustain oncogenesis. Targeting ClpP to dysregulate its function using small-molecule agonists is a recent strategy in cancer therapy. Here, we synthesized imipridone-derived compounds and related chemicals, which we characterized using biochemical, biophysical, and cellular studies. Using X-ray crystallography, we found that these compounds have enhanced binding affinities due to their greater shape and charge complementarity with the surface hydrophobic pockets of ClpP. N-terminome profiling of cancer cells upon treatment with one of these compounds revealed the global proteomic changes that arise and identified the structural motifs preferred for protein cleavage by compound-activated ClpP. Together, our studies provide the structural and molecular basis by which dysregulated ClpP affects cancer cell viability and proliferation.
    Keywords:  ClpP agonist; ClpP protease; HYTANE mass spectrometry; N-terminome; TR compounds; X-ray crystallography; cancer; drug design; imipridones; mitochondria
    DOI:  https://doi.org/10.1016/j.str.2022.12.002
  2. Nat Struct Mol Biol. 2023 Jan 05.
    A multipoint guidance mechanism for β-barrel folding on the SAM complex.
    Hironori Takeda, Jon V Busto, Caroline Lindau, Akihisa Tsutsumi, Kentaro Tomii, Kenichiro Imai, Yu Yamamori, Takatsugu Hirokawa, Chie Motono, Iniyan Ganesan, Lena-Sophie Wenz, Thomas Becker, Masahide Kikkawa, Nikolaus Pfanner, Nils Wiedemann, Toshiya Endo.
      Mitochondrial β-barrel proteins are essential for the transport of metabolites, ions and proteins. The sorting and assembly machinery (SAM) mediates their folding and membrane insertion. We report the cryo-electron microscopy structure of the yeast SAM complex carrying an early eukaryotic β-barrel folding intermediate. The lateral gate of Sam50 is wide open and pairs with the last β-strand (β-signal) of the substrate-the 19-β-stranded Tom40 precursor-to form a hybrid barrel in the membrane plane. The Tom40 barrel grows and curves, guided by an extended bridge with Sam50. Tom40's first β-segment (β1) penetrates into the nascent barrel, interacting with its inner wall. The Tom40 amino-terminal segment then displaces β1 to promote its pairing with Tom40's last β-strand to complete barrel formation with the assistance of Sam37's dynamic α-protrusion. Our study thus reveals a multipoint guidance mechanism for mitochondrial β-barrel folding.
    DOI:  https://doi.org/10.1038/s41594-022-00897-2
  3. Proc Natl Acad Sci U S A. 2023 Jan 10. 120(2): e2204750120
    Exercise preserves physical fitness during aging through AMPK and mitochondrial dynamics.
    Juliane Cruz Campos, Luiz Henrique Marchesi Bozi, Barbara Krum, Luiz Roberto Grassmann Bechara, Nikolas Dresch Ferreira, Gabriel Santos Arini, Rudá Prestes Albuquerque, Annika Traa, Takafumi Ogawa, Alexander M van der Bliek, Afshin Beheshti, Edward T Chouchani, Jeremy M Van Raamsdonk, T Keith Blackwell, Julio Cesar Batista Ferreira.
      Exercise is a nonpharmacological intervention that improves health during aging and a valuable tool in the diagnostics of aging-related diseases. In muscle, exercise transiently alters mitochondrial functionality and metabolism. Mitochondrial fission and fusion are critical effectors of mitochondrial plasticity, which allows a fine-tuned regulation of organelle connectiveness, size, and function. Here we have investigated the role of mitochondrial dynamics during exercise in the model organism Caenorhabditis elegans. We show that in body-wall muscle, a single exercise session induces a cycle of mitochondrial fragmentation followed by fusion after a recovery period, and that daily exercise sessions delay the mitochondrial fragmentation and physical fitness decline that occur with aging. Maintenance of proper mitochondrial dynamics is essential for physical fitness, its enhancement by exercise training, and exercise-induced remodeling of the proteome. Surprisingly, among the long-lived genotypes we analyzed (isp-1,nuo-6, daf-2, eat-2, and CA-AAK-2), constitutive activation of AMP-activated protein kinase (AMPK) uniquely preserves physical fitness during aging, a benefit that is abolished by impairment of mitochondrial fission or fusion. AMPK is also required for physical fitness to be enhanced by exercise, with our findings together suggesting that exercise may enhance muscle function through AMPK regulation of mitochondrial dynamics. Our results indicate that mitochondrial connectivity and the mitochondrial dynamics cycle are essential for maintaining physical fitness and exercise responsiveness during aging and suggest that AMPK activation may recapitulate some exercise benefits. Targeting mechanisms to optimize mitochondrial fission and fusion, as well as AMPK activation, may represent promising strategies for promoting muscle function during aging.
    Keywords:  C. elegans; aging; exercise; mitochondrial fission; mitochondrial fusion
    DOI:  https://doi.org/10.1073/pnas.2204750120
  4. J Cell Sci. 2023 Jan 05. pii: jcs.260060. [Epub ahead of print]
    Functional crosstalk between the TIM22 complex and YME1 machinery maintains mitochondrial proteostasis and integrity.
    Abhishek Kumar, Tejashree Pradip Waingankar, Patrick D'Silva.
      The TIM22 pathway cargos are essential for sustaining mitochondrial homeostasis as an excess of these proteins leads to proteostatic stress and cell death. Yme1 is an inner membrane metalloprotease that regulates protein quality control with chaperone-like and proteolytic activities. Although the mitochondrial translocase and protease machinery are critical for organelle health, their functional association remains unexplored. The present study unravels a novel genetic connection between the TIM22 complex and YME1 machinery in Saccharomyces cerevisiae required for maintaining mitochondrial health. Our genetic analyses indicate that impairment in the TIM22 complex rescues the respiratory growth defects of cells without Yme1. Further, Yme1 is essential for the stability of the TIM22 complex and regulates the proteostasis of the TIM22 pathway substrates. Moreover, impairment in the TIM22 complex suppressed the mitochondrial structural and functional defects of Yme1 devoid cells. In summary, excessive levels of the TIM22 pathway substrates could be one of the reasons for respiratory growth defects of cells lacking Yme1, and compromising the TIM22 complex can compensate for the imbalance in mitochondrial proteostasis caused by the loss of Yme1.
    Keywords:  Mitochondrial DNA maintenance; Mitochondrial protein translocation; Mitochondrial proteostasis; Mitochondrial quality control; TIM22 complex; YME1 machinery
    DOI:  https://doi.org/10.1242/jcs.260060
  5. Cell Rep. 2022 Dec 23. pii: S2211-1247(22)01798-3. [Epub ahead of print] 111899
    The endoplasmic reticulum kinase PERK interacts with the oxidoreductase ERO1 to metabolically adapt mitochondria.
    Arthur Bassot, Junsheng Chen, Kei Takahashi-Yamashiro, Megan C Yap, Christine Silvia Gibhardt, Giang N T Le, Saaya Hario, Yusuke Nasu, Jack Moore, Tomas Gutiérrez, Lucas Mina, Heather Mast, Audric Moses, Rakesh Bhat, Klaus Ballanyi, Hélène Lemieux, Roberto Sitia, Ester Zito, Ivan Bogeski, Robert E Campbell, Thomas Simmen.
      Endoplasmic reticulum (ER) homeostasis requires molecular regulators that tailor mitochondrial bioenergetics to the needs of protein folding. For instance, calnexin maintains mitochondria metabolism and mitochondria-ER contacts (MERCs) through reactive oxygen species (ROS) from NADPH oxidase 4 (NOX4). However, induction of ER stress requires a quick molecular rewiring of mitochondria to adapt to new energy needs. This machinery is not characterized. We now show that the oxidoreductase ERO1⍺ covalently interacts with protein kinase RNA-like ER kinase (PERK) upon treatment with tunicamycin. The PERK-ERO1⍺ interaction requires the C-terminal active site of ERO1⍺ and cysteine 216 of PERK. Moreover, we show that the PERK-ERO1⍺ complex promotes oxidization of MERC proteins and controls mitochondrial dynamics. Using proteinaceous probes, we determined that these functions improve ER-mitochondria Ca2+ flux to maintain bioenergetics in both organelles, while limiting oxidative stress. Therefore, the PERK-ERO1⍺ complex is a key molecular machinery that allows quick metabolic adaptation to ER stress.
    Keywords:  CP: Metabolism; CP: Molecular biology; ER; ER stress; ERO1; MAMs; MERCs; PERK; bioenergetics; endoplasmic reticulum; mitochondria; mitochondria-associated membranes; mitochondria-endoplasmic reticulum contacts; oxidoreductase
    DOI:  https://doi.org/10.1016/j.celrep.2022.111899
  6. Nat Commun. 2023 Jan 03. 14(1): 39
    The mitochondrial Hsp70 controls the assembly of the F1FO-ATP synthase.
    Jiyao Song, Liesa Steidle, Isabelle Steymans, Jasjot Singh, Anne Sanner, Lena Böttinger, Dominic Winter, Thomas Becker.
      The mitochondrial F1FO-ATP synthase produces the bulk of cellular ATP. The soluble F1 domain contains the catalytic head that is linked via the central stalk and the peripheral stalk to the membrane embedded rotor of the FO domain. The assembly of the F1 domain and its linkage to the peripheral stalk is poorly understood. Here we show a dual function of the mitochondrial Hsp70 (mtHsp70) in the formation of the ATP synthase. First, it cooperates with the assembly factors Atp11 and Atp12 to form the F1 domain of the ATP synthase. Second, the chaperone transfers Atp5 into the assembly line to link the catalytic head with the peripheral stalk. Inactivation of mtHsp70 leads to integration of assembly-defective Atp5 variants into the mature complex, reflecting a quality control function of the chaperone. Thus, mtHsp70 acts as an assembly and quality control factor in the biogenesis of the F1FO-ATP synthase.
    DOI:  https://doi.org/10.1038/s41467-022-35720-5
  7. Nat Commun. 2023 Jan 03. 14(1): 30
    Human mitochondria require mtRF1 for translation termination at non-canonical stop codons.
    Annika Krüger, Cristina Remes, Dmitrii Igorevich Shiriaev, Yong Liu, Henrik Spåhr, Rolf Wibom, Ilian Atanassov, Minh Duc Nguyen, Barry S Cooperman, Joanna Rorbach.
      The mitochondrial translation machinery highly diverged from its bacterial counterpart. This includes deviation from the universal genetic code, with AGA and AGG codons lacking cognate tRNAs in human mitochondria. The locations of these codons at the end of COX1 and ND6 open reading frames, respectively, suggest they might function as stop codons. However, while the canonical stop codons UAA and UAG are known to be recognized by mtRF1a, the release mechanism at AGA and AGG codons remains a debated issue. Here, we show that upon the loss of another member of the mitochondrial release factor family, mtRF1, mitoribosomes accumulate specifically at AGA and AGG codons. Stalling of mitoribosomes alters COX1 transcript and protein levels, but not ND6 synthesis. In addition, using an in vitro reconstituted mitochondrial translation system, we demonstrate the specific peptide release activity of mtRF1 at the AGA and AGG codons. Together, our results reveal the role of mtRF1 in translation termination at non-canonical stop codons in mitochondria.
    DOI:  https://doi.org/10.1038/s41467-022-35684-6
  8. J Biol Chem. 2023 Jan 02. pii: S0021-9258(22)01306-0. [Epub ahead of print] 102863
    Endoplasmic reticulum protein BIK binds to and inhibits mitochondria-localized anti-apoptotic proteins.
    Elizabeth J Osterlund, Nehad Hirmiz, Dang Nguyen, James M Pemberton, Qiyin Fang, David W Andrews.
      The pro-apoptotic BH3-only endoplasmic reticulum (ER) resident protein BIK, positively regulates mitochondrial outer membrane permeabilization (MOMP), the point-of-no-return in apoptosis. It is generally accepted that BIK functions at a distance from mitochondria by binding and sequestering anti-apoptotic proteins at the ER thereby promoting ER calcium release. Although BIK is predominantly localized to the ER, we detect by FLIM-FRET microscopy, BH3 region-dependent direct binding between BIK and mitochondria-localized chimeric mutants of the anti-apoptotic proteins BCL-XL and BCL-2 in both BMK and MCF-7 cells. Direct binding was accompanied by cell-type specific differential relocalization in response to co-expression of either BIK or one of its target binding partners, BCL-XL, when co-expressed in cells. In BMK cells with genetic deletion of both BAX and BAK (BMK-DKO) our data suggest a fraction of BIK protein moves towards mitochondria in response to the expression of a mitochondria-localized BCL-XL mutant. In contrast, in MCF-7 cells our data suggest BIK is localized at both ER and mitochondria-associated endoplasmic reticulum membranes (MAMs) and binds to the mitochondria-localized BCL-XL mutant via relocalization of BCL-XL to ER and MAMs. Rather than functioning at a distance, our data suggest BIK initiates MOMP via direct interactions with ER and mitochondria-localized anti-apoptotic proteins, that occur via ER-mitochondria contact sites, and/or by relocalization of either BIK or anti-apoptotic proteins in cells.
    Keywords:  BCL-2 family; BCL-2 interacting killer; BIK; FLIM-FRET; apoptosis; subcellular localization fluorescence lifetime imaging microscopy
    DOI:  https://doi.org/10.1016/j.jbc.2022.102863
  9. Sci Adv. 2023 Jan 04. 9(1): eadd3216
    Lactate metabolism is essential in early-onset mitochondrial myopathy.
    Zhenkang Chen, Bogdan Bordieanu, Rushendhiran Kesavan, Nicholas P Lesner, Siva Sai Krishna Venigalla, Spencer D Shelton, Ralph J DeBerardinis, Prashant Mishra.
      Myopathies secondary to mitochondrial electron transport chain (ETC) dysfunction can result in devastating disease. While the consequences of ETC defects have been extensively studied in culture, little in vivo data are available. Using a mouse model of severe, early-onset mitochondrial myopathy, we characterized the proteomic, transcriptomic, and metabolic characteristics of disease progression. Unexpectedly, ETC dysfunction in muscle results in reduced expression of glycolytic enzymes in our animal model and patient muscle biopsies. The decrease in glycolysis was mediated by loss of constitutive Hif1α signaling, down-regulation of the purine nucleotide cycle enzyme AMPD1, and activation of AMPK. In vivo isotope tracing experiments indicated that myopathic muscle relies on lactate import to supply central carbon metabolites. Inhibition of lactate import reduced steady-state levels of tricarboxylic acid cycle intermediates and compromised the life span of myopathic mice. These data indicate an unexpected mode of metabolic reprogramming in severe mitochondrial myopathy that regulates disease progression.
    DOI:  https://doi.org/10.1126/sciadv.add3216
  10. Autophagy. 2023 Jan 01. 1-18
    The nucleoprotein of influenza A virus inhibits the innate immune response by inducing mitophagy.
    Bo Zhang, Shuai Xu, Minxuan Liu, Yanli Wei, Qian Wang, Wentao Shen, Cao-Qi Lei, Qiyun Zhu.
      Mitophagy is a form of autophagy that plays a key role in maintaining the homeostasis of functional mitochondria in the cell. Viruses have evolved various strategies to manipulate mitophagy to escape host immune responses and promote virus replication. In this study, the nucleoprotein (NP) of H1N1 virus (PR8 strain) was identified as a regulator of mitophagy. We revealed that NP-mediated mitophagy leads to the degradation of the mitochondria-anchored protein MAVS, thereby blocking MAVS-mediated antiviral signaling and promoting virus replication. The NP-mediated mitophagy is dependent on the interaction of NP with MAVS and the cargo receptor TOLLIP. Moreover, Y313 of NP is a key residue for the MAVS-NP interaction and NP-mediated mitophagy. The NPY313F mutation significantly attenuates the virus-induced mitophagy and the virus replication in vitro and in vivo. Taken together, our findings uncover a novel mechanism by which the NP of influenza virus induces mitophagy to attenuate innate immunity.Abbreviations: ACTB: actin beta; ATG7: autophagy related 7; ATG12: autophagy related 12; CCCP: carbonyl cyanide 3-chlorophenyl hydrazone; co-IP: co-immunoprecipitation; COX4/COXIV: cytochrome c oxidase subunit 4; DAPI: 4',6-diamidino-2-phenylindole, dihydrochloride; EID50: 50% egg infective dose; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFP: green fluorescent protein; HEK: human embryonic kidney; hpi: hours post-infection; IAV: influenza A virus; IFN: interferon; IP: immunoprecipitation; LAMP1: lysosomal associated membrane protein 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAVS: mitochondrial antiviral signaling protein; Mdivi-1: mitochondrial division inhibitor 1; MLD50: 50% mouse lethal dose; MOI: multiplicity of infection; NBR1: NBR1 autophagy cargo receptor; NP: nucleoprotein; PB1: basic polymerase 1; RFP: red fluorescent protein; RIGI: RNA sensor RIG-I; RIGI-N: RIGI-CARD; SeV: Sendai virus; SQSTM1/p62: sequestosome 1; TIMM23: translocase of inner mitochondrial membrane 23; TOLLIP: toll interacting protein; TOMM20: translocase of outer mitochondrial membrane 20; TUBA: tubulin alpha; Vec: empty vector; vRNP: viral ribonucleoprotein.
    Keywords:  Influenza A virus; MAVS; TOLLIP; mitophagy; nucleoprotein
    DOI:  https://doi.org/10.1080/15548627.2022.2162798
  11. Sci Rep. 2023 Jan 02. 13(1): 18
    Inner mitochondrial membrane protein Prohibitin 1 mediates Nix-induced, Parkin-independent mitophagy.
    Kibrom M Alula, Yaritza Delgado-Deida, Rosemary Callahan, Andreas Till, Lucia Underwood, Winston E Thompson, Rhonda F Souza, Themistocles Dassopoulos, Joseph Onyiah, K Venuprasad, Arianne L Theiss.
      Autophagy of damaged mitochondria, called mitophagy, is an important organelle quality control process involved in the pathogenesis of inflammation, cancer, aging, and age-associated diseases. Many of these disorders are associated with altered expression of the inner mitochondrial membrane (IMM) protein Prohibitin 1. The mechanisms whereby dysfunction occurring internally at the IMM and matrix activate events at the outer mitochondrial membrane (OMM) to induce mitophagy are not fully elucidated. Using the gastrointestinal epithelium as a model system highly susceptible to autophagy inhibition, we reveal a specific role of Prohibitin-induced mitophagy in maintaining intestinal homeostasis. We demonstrate that Prohibitin 1 induces mitophagy in response to increased mitochondrial reactive oxygen species (ROS) through binding to mitophagy receptor Nix/Bnip3L and independently of Parkin. Prohibitin 1 is required for ROS-induced Nix localization to mitochondria and maintaining homeostasis of epithelial cells highly susceptible to mitochondrial dysfunction.
    DOI:  https://doi.org/10.1038/s41598-022-26775-x
  12. STAR Protoc. 2022 Dec 16. pii: S2666-1667(22)00702-X. [Epub ahead of print]3(4): 101822
    Fluorescent pulse-chase labeling to monitor long-term mitochondrial degradation in primary hippocampal neurons.
    Jordan R Schneider, Chantell S Evans.
      The accumulation of dysfunctional mitochondria is a hallmark of neurodegenerative diseases, yet the dynamics of mitochondrial turnover in neurons are unclear. Here, we describe a protocol to monitor the degradation of spectrally distinct, "aged" mitochondrial populations. We describe the preparation and transfection of primary rat hippocampal neuron cultures. We detail a mitochondrial-damaging assay, a SNAP pulse-chase labeling paradigm, and live imaging to visualize the mitochondrial network. Finally, we provide steps to quantify mitochondrial turnover via lysosomal fusion. For complete details on the use and execution of this protocol, please refer to Evans and Holzbaur (2020a).
    Keywords:  Cell Biology; Cell culture; Cell-based Assays; Metabolism; Microscopy; Molecular Biology; Molecular/Chemical Probes; Neuroscience
    DOI:  https://doi.org/10.1016/j.xpro.2022.101822
  13. J Biol Chem. 2023 Jan 02. pii: S0021-9258(22)01308-4. [Epub ahead of print] 102865
    Monitoring mitochondrial translation by pulse SILAC.
    Koshi Imami, Matthias Selbach, Yasushi Ishihama.
      Mitochondrial ribosomes are specialized to translate the 13 membrane proteins encoded in the mitochondrial genome, which shapes the oxidative phosphorylation (OXPHOS) complexes essential for cellular energy metabolism. Despite the importance of mitochondrial translation control, it is challenging to identify and quantify the mitochondrial-encoded proteins due to their hydrophobic nature and low abundance. Here, we introduce a mass spectrometry-based proteomic method that combines biochemical isolation of mitochondria with pulse stable isotope labeling by amino acids in cell culture (pSILAC). Our method provides the highest protein identification rate with the shortest measurement time among currently available methods, enabling us to quantify 12 out of the 13 mitochondrial-encoded proteins. We applied this method to uncover the global picture of (post-)translational regulation of both mitochondrial- and nuclear-encoded subunits of OXPHOS complexes. We found that inhibition of mitochondrial translation led to degradation of orphan nuclear-encoded subunits that are considered to form subcomplexes with the mitochondrial-encoded subunits. This method should be readily applicable to study mitochondrial translation programs in many contexts, including oxidative stress and mitochondrial disease.
    Keywords:  Mitochondria; OXPHOS; Protein complex; Proteomics; Translation; pulse SILAC
    DOI:  https://doi.org/10.1016/j.jbc.2022.102865
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