bims-mitdyn Biomed News
on Mitochondrial dynamics: mechanisms
Issue of 2024–01–28
ten papers selected by
Edmond Chan, Queen’s University, School of Medicine
  1. Motion of VAPB molecules reveals ER-mitochondria contact site subdomains.
  2. Shared structural features of Miro binding control mitochondrial homeostasis.
  3. CLPB disaggregase dysfunction impacts the functional integrity of the proteolytic SPY complex.
  4. Phosphate starvation signaling increases mitochondrial membrane potential through respiration-independent mechanisms.
  5. Drosophila MIC10b can polymerize into cristae-shaping filaments.
  6. Mitochondrial translocation of TFEB regulates complex I and inflammation.
  7. Lithium carbonate revitalizes tumor-reactive CD8+ T cells by shunting lactic acid into mitochondria.
  8. Characterization of a novel variant in the HR1 domain of MFN2 in a patient with ataxia, optic atrophy and sensorineural hearing loss.
  9. Protocol for measuring mitochondrial size in mouse and human liver tissues.
  10. A NOVEL MTORC2-AKT-ROS AXIS TRIGGERS MITOFISSION and MITOPHAGY-ASSOCIATED EXECUTION of COLORECTAL CANCER CELLS UPON DRUG-INDUCED ACTIVATION of MUTANT KRAS.
  1. Nature. 2024 Jan 24.
    Motion of VAPB molecules reveals ER-mitochondria contact site subdomains.
    Christopher J Obara, Jonathon Nixon-Abell, Andrew S Moore, Federica Riccio, David P Hoffman, Gleb Shtengel, C Shan Xu, Kathy Schaefer, H Amalia Pasolli, Jean-Baptiste Masson, Harald F Hess, Christopher P Calderon, Craig Blackstone, Jennifer Lippincott-Schwartz.
      To coordinate cellular physiology, eukaryotic cells rely on the rapid exchange of molecules at specialized organelle-organelle contact sites1,2. Endoplasmic reticulum-mitochondrial contact sites (ERMCSs) are particularly vital communication hubs, playing key roles in the exchange of signalling molecules, lipids and metabolites3,4. ERMCSs are maintained by interactions between complementary tethering molecules on the surface of each organelle5,6. However, due to the extreme sensitivity of these membrane interfaces to experimental perturbation7,8, a clear understanding of their nanoscale organization and regulation is still lacking. Here we combine three-dimensional electron microscopy with high-speed molecular tracking of a model organelle tether, Vesicle-associated membrane protein (VAMP)-associated protein B (VAPB), to map the structure and diffusion landscape of ERMCSs. We uncovered dynamic subdomains within VAPB contact sites that correlate with ER membrane curvature and undergo rapid remodelling. We show that VAPB molecules enter and leave ERMCSs within seconds, despite the contact site itself remaining stable over much longer time scales. This metastability allows ERMCSs to remodel with changes in the physiological environment to accommodate metabolic needs of the cell. An amyotrophic lateral sclerosis-associated mutation in VAPB perturbs these subdomains, likely impairing their remodelling capacity and resulting in impaired interorganelle communication. These results establish high-speed single-molecule imaging as a new tool for mapping the structure of contact site interfaces and reveal that the diffusion landscape of VAPB at contact sites is a crucial component of ERMCS homeostasis.
    DOI:  https://doi.org/10.1038/s41586-023-06956-y
  2. EMBO J. 2024 Jan 24.
    Shared structural features of Miro binding control mitochondrial homeostasis.
    Christian Covill-Cooke, Brian Kwizera, Guillermo López-Doménech, Caleb Od Thompson, Ngaam J Cheung, Ema Cerezo, Martin Peterka, Josef T Kittler, Benoît Kornmann.
      Miro proteins are universally conserved mitochondrial calcium-binding GTPases that regulate a multitude of mitochondrial processes, including transport, clearance, and lipid trafficking. The exact role of Miro in these functions is unclear but involves binding to a variety of client proteins. How this binding is operated at the molecular level and whether and how it is important for mitochondrial health, however, remains unknown. Here, we show that known Miro interactors-namely, CENPF, Trak, and MYO19-all use a similar short motif to bind the same structural element: a highly conserved hydrophobic pocket in the first calcium-binding domain of Miro. Using these Miro-binding motifs, we identified direct interactors de novo, including MTFR1/2/1L, the lipid transporters Mdm34 and VPS13D, and the ubiquitin E3-ligase Parkin. Given the shared binding mechanism of these functionally diverse clients and its conservation across eukaryotes, we propose that Miro is a universal mitochondrial adaptor coordinating mitochondrial health.
    Keywords:  AlphaFold; ERMES; Lipid Transport; Mitophagy; Organelle Transport
    DOI:  https://doi.org/10.1038/s44318-024-00028-1
  3. J Cell Biol. 2024 Mar 04. pii: e202305087. [Epub ahead of print]223(3):
    CLPB disaggregase dysfunction impacts the functional integrity of the proteolytic SPY complex.
    Megan J Baker, Kai Uwe Blau, Alexander J Anderson, Catherine S Palmer, Laura F Fielden, Jordan J Crameri, Dusanka Milenkovic, David R Thorburn, Ann E Frazier, Thomas Langer, Diana Stojanovski.
      CLPB is a mitochondrial intermembrane space AAA+ domain-containing disaggregase. CLPB mutations are associated with 3-methylglutaconic aciduria and neutropenia; however, the molecular mechanism underscoring disease and the contribution of CLPB substrates to disease pathology remains unknown. Interactions between CLPB and mitochondrial quality control (QC) factors, including PARL and OPA1, have been reported, hinting at dysregulation of organelle QC in disease. Utilizing proteomic and biochemical approaches, we show a stress-specific aggregation phenotype in a CLPB-null environment and define the CLPB substrate profile. We illustrate an interplay between intermembrane space proteins including CLPB, HAX1, HTRA2, and the inner membrane quality control proteins (STOML2, PARL, YME1L1; SPY complex), with CLPB deficiency impeding SPY complex function by virtue of protein aggregation in the intermembrane space. We conclude that there is an interdependency of mitochondrial QC components at the intermembrane space/inner membrane interface, and perturbations to this network may underscore CLPB disease pathology.
    DOI:  https://doi.org/10.1083/jcb.202305087
  4. Elife. 2024 Jan 22. pii: e84282. [Epub ahead of print]13
    Phosphate starvation signaling increases mitochondrial membrane potential through respiration-independent mechanisms.
    Yeyun Ouyang, Mi-Young Jeong, Corey N Cunningham, Jordan A Berg, Ashish G Toshniwal, Casey E Hughes, Kristina Seiler, Jonathan G Van Vranken, Ahmad A Cluntun, Geanette Lam, Jacob M Winter, Emel Akdoǧan, Katja K Dove, Sara M Nowinski, Matthew West, Greg Odorizzi, Steven P Gygi, Cory D Dunn, Dennis R Winge, Jared Rutter.
      Mitochondrial membrane potential directly powers many critical functions of mitochondria, including ATP production, mitochondrial protein import, and metabolite transport. Its loss is a cardinal feature of aging and mitochondrial diseases, and cells closely monitor membrane potential as an indicator of mitochondrial health. Given its central importance, it is logical that cells would modulate mitochondrial membrane potential in response to demand and environmental cues, but there has been little exploration of this question. We report that loss of the Sit4 protein phosphatase in yeast increases mitochondrial membrane potential, both through inducing the electron transport chain and the phosphate starvation response. Indeed, a similarly elevated mitochondrial membrane potential is also elicited simply by phosphate starvation or by abrogation of the Pho85-dependent phosphate sensing pathway. This enhanced membrane potential is primarily driven by an unexpected activity of the ADP/ATP carrier. We also demonstrate that this connection between phosphate limitation and enhancement of mitochondrial membrane potential is observed in primary and immortalized mammalian cells as well as in Drosophila. These data suggest that mitochondrial membrane potential is subject to environmental stimuli and intracellular signaling regulation and raise the possibility for therapeutic enhancement of mitochondrial function even in defective mitochondria.
    Keywords:  D. melanogaster; S. cerevisiae; cell biology; human
    DOI:  https://doi.org/10.7554/eLife.84282
  5. Life Sci Alliance. 2024 Apr;pii: e202302177. [Epub ahead of print]7(4):
    Drosophila MIC10b can polymerize into cristae-shaping filaments.
    Till Stephan, Stefan Stoldt, Mariam Barbot, Travis D Carney, Felix Lange, Mark Bates, Peter Bou Dib, Kaushik Inamdar, Halyna R Shcherbata, Michael Meinecke, Dietmar Riedel, Sven Dennerlein, Peter Rehling, Stefan Jakobs.
      Cristae are invaginations of the mitochondrial inner membrane that are crucial for cellular energy metabolism. The formation of cristae requires the presence of a protein complex known as MICOS, which is conserved across eukaryotic species. One of the subunits of this complex, MIC10, is a transmembrane protein that supports cristae formation by oligomerization. In Drosophila melanogaster, three MIC10-like proteins with different tissue-specific expression patterns exist. We demonstrate that CG41128/MINOS1b/DmMIC10b is the major MIC10 orthologue in flies. Its loss destabilizes MICOS, disturbs cristae architecture, and reduces the life span and fertility of flies. We show that DmMIC10b has a unique ability to polymerize into bundles of filaments, which can remodel mitochondrial crista membranes. The formation of these filaments relies on conserved glycine and cysteine residues, and can be suppressed by the co-expression of other Drosophila MICOS proteins. These findings provide new insights into the regulation of MICOS in flies, and suggest potential mechanisms for the maintenance of mitochondrial ultrastructure.
    DOI:  https://doi.org/10.26508/lsa.202302177
  6. EMBO Rep. 2024 Jan 23.
    Mitochondrial translocation of TFEB regulates complex I and inflammation.
    Chiara Calabrese, Hendrik Nolte, Melissa R Pitman, Raja Ganesan, Philipp Lampe, Raymond Laboy, Roberto Ripa, Julia Fischer, Ruhi Polara, Sameer Kumar Panda, Sandhya Chipurupalli, Saray Gutierrez, Daniel Thomas, Stuart M Pitson, Adam Antebi, Nirmal Robinson.
      TFEB is a master regulator of autophagy, lysosome biogenesis, mitochondrial metabolism, and immunity that works primarily through transcription controlled by cytosol-to-nuclear translocation. Emerging data indicate additional regulatory interactions at the surface of organelles such as lysosomes. Here we show that TFEB has a non-transcriptional role in mitochondria, regulating the electron transport chain complex I to down-modulate inflammation. Proteomics analysis reveals extensive TFEB co-immunoprecipitation with several mitochondrial proteins, whose interactions are disrupted upon infection with S. Typhimurium. High resolution confocal microscopy and biochemistry confirms TFEB localization in the mitochondrial matrix. TFEB translocation depends on a conserved N-terminal TOMM20-binding motif and is enhanced by mTOR inhibition. Within the mitochondria, TFEB and protease LONP1 antagonistically co-regulate complex I, reactive oxygen species and the inflammatory response. Consequently, during infection, lack of TFEB specifically in the mitochondria exacerbates the expression of pro-inflammatory cytokines, contributing to innate immune pathogenesis.
    Keywords:  LONP1; Metabolism; Mitochondria; Salmonella; TFEB
    DOI:  https://doi.org/10.1038/s44319-024-00058-0
  7. Nat Immunol. 2024 Jan 23.
    Lithium carbonate revitalizes tumor-reactive CD8+ T cells by shunting lactic acid into mitochondria.
    Jingwei Ma, Liang Tang, Yaoyao Tan, Jingxuan Xiao, Keke Wei, Xin Zhang, Yuan Ma, Shuai Tong, Jie Chen, Nannan Zhou, Li Yang, Zhang Lei, Yonggang Li, Jiadi Lv, Junwei Liu, Huafeng Zhang, Ke Tang, Yi Zhang, Bo Huang.
      The steady flow of lactic acid (LA) from tumor cells to the extracellular space via the monocarboxylate transporter symport system suppresses antitumor T cell immunity. However, LA is a natural energy metabolite that can be oxidized in the mitochondria and could potentially stimulate T cells. Here we show that the lactate-lowering mood stabilizer lithium carbonate (LC) can inhibit LA-mediated CD8+ T cell immunosuppression. Cytoplasmic LA increased the pumping of protons into lysosomes. LC interfered with vacuolar ATPase to block lysosomal acidification and rescue lysosomal diacylglycerol-PKCθ signaling to facilitate monocarboxylate transporter 1 localization to mitochondrial membranes, thus transporting LA into the mitochondria as an energy source for CD8+ T cells. These findings indicate that targeting LA metabolism using LC could support cancer immunotherapy.
    DOI:  https://doi.org/10.1038/s41590-023-01738-0
  8. F1000Res. 2021 ;10 606
    Characterization of a novel variant in the HR1 domain of MFN2 in a patient with ataxia, optic atrophy and sensorineural hearing loss.
    Govinda Sharma, Mashiat Zaman, Rasha Sabouny, Matthew Joel, Kristina Martens, Davide Martino, A P Jason de Koning, Gerald Pfeffer, Timothy E Shutt.
      Background: Pathogenic variants in MFN2 cause Charcot-Marie-Tooth disease (CMT) type 2A (CMT2A) and are the leading cause of the axonal subtypes of CMT. CMT2A is characterized by predominantly distal motor weakness and muscle atrophy, with highly variable severity and onset age. Notably, some MFN2 variants can also lead to other phenotypes such as optic atrophy, hearing loss and lipodystrophy. Despite the clear link between MFN2 and CMT2A, our mechanistic understanding of how dysfunction of the MFN2 protein causes human disease pathologies remains incomplete. This lack of understanding is due in part to the multiple cellular roles of MFN2. Though initially characterized for its role in mediating mitochondrial fusion, MFN2 also plays important roles in mediating interactions between mitochondria and other organelles, such as the endoplasmic reticulum and lipid droplets. Additionally, MFN2 is also important for mitochondrial transport, mitochondrial autophagy, and has even been implicated in lipid transfer. Though over 100 pathogenic MFN2 variants have been described to date, only a few have been characterized functionally, and even then, often only for one or two functions. Method: Several MFN2-mediated functions were characterized in fibroblast cells from a patient presenting with cerebellar ataxia, deafness, blindness, and diffuse cerebral and cerebellar atrophy, who harbours a novel homozygous MFN2 variant, D414V, which is found in a region of the HR1 domain of MFN2 where few pathogenic variants occur. Results: We found evidence for impairment of several MFN2-mediated functions. Consistent with reduced mitochondrial fusion, patient fibroblasts exhibited more fragmented mitochondrial networks and had reduced mtDNA copy number. Additionally, patient fibroblasts had reduced oxygen consumption, fewer mitochondrial-ER contacts, and altered lipid droplets that displayed an unusual perinuclear distribution. Conclusion: Overall, this work characterizes D414V as a novel variant in MFN2 and expands the phenotypic presentation of MFN2 variants to include cerebellar ataxia.
    Keywords:  Ataxia; MFN2; Mitochondria; Mitochondrial Fusion
    DOI:  https://doi.org/10.12688/f1000research.53230.2
  9. STAR Protoc. 2024 Jan 18. pii: S2666-1667(24)00007-8. [Epub ahead of print]5(1): 102842
    Protocol for measuring mitochondrial size in mouse and human liver tissues.
    Balamurugan Ramatchandirin, Miho Iijima, Arisa Ikeda, Alexia Pearah, Sally Radovick, Fredric E Wondisford, Hiromi Sesaki, Ling He.
      Mitochondrial dynamic process is important for cell viability, metabolic activity, and mitochondria health. Here, we present a protocol for measuring mitochondrial size through immunofluorescence staining, confocal imaging, and analysis in ImageJ. We describe the steps for tissue processing, antigen retrieval, mitochondrial staining using an integrating immunofluorescence assay, and computerized image analysis to measure each mitochondrial size in mouse and human liver tissues. This protocol reduces tissue sample volume and processing time for the preparation of primary cells. For complete details on the use and execution of this protocol, please refer to Pearah et al.1.
    Keywords:  Cell Biology; Health Sciences; Metabolism
    DOI:  https://doi.org/10.1016/j.xpro.2024.102842
  10. Autophagy. 2024 Jan 23.
    A NOVEL MTORC2-AKT-ROS AXIS TRIGGERS MITOFISSION and MITOPHAGY-ASSOCIATED EXECUTION of COLORECTAL CANCER CELLS UPON DRUG-INDUCED ACTIVATION of MUTANT KRAS.
    Kartini Iskandar, Jonathan Foo, Angeline Qiu Xia Liew, Haiyuxin Zhu, Deepika Raman, Jayshree L Hirpara, Yan Yi Leong, Maria V Babak, Anna A Kirsanova, Anne-Sophie Armand, Franck Oury, Gregory Bellot, Shazib Pervaiz.
      RAS is one of the most commonly mutated oncogenes associated with multiple cancer hallmarks. Notably, RAS activation induces intracellular reactive oxygen species (ROS) generation, which we previously demonstrated as a trigger for autophagy-associated execution of mutant KRAS-expressing cancer cells. Here we report that drug (merodantoin; C1)-induced activation of mutant KRAS promotes phospho-AKT S473-dependent ROS-mediated S616 phosphorylation and mitochondrial localization of DNM1L/DRP1 (dynamin 1 like) and cleavage of the fusion-associated protein OPA1 (OPA1 mitochondrial dynamin like GTPase). Interestingly, accumulation of the outer mitochondrial membrane protein VDAC1 (voltage dependent anion channel 1) is observed in mutant KRAS-expressing cells upon exposure to C1. Conversely, silencing VDAC1 abolishes C1-induced mitophagy, and gene knockdown of either KRAS, AKT or DNM1L rescues ROS-dependent VDAC1 accumulation and stability, thus suggesting an axis of mutant active KRAS-phospho-AKT S473-ROS-DNM1L-VDAC1 in mitochondrial morphology change and cancer cell execution. Importantly, we identified MTOR (mechanistic target of rapamycin kinsase) complex 2 (MTORC2) as the upstream mediator of AKT phosphorylation at S473 in our model. Pharmacological or genetic inhibition of MTORC2 abrogated C1-induced phosphorylation of AKT S473, ROS generation and mitophagy induction, as well as rescued tumor colony forming ability and migratory capacity. Finally, increase in thermal stability of KRAS, AKT and DNM1L were observed upon exposure to C1 only in mutant KRAS-expressing cells. Taken together, our work has unraveled a novel mechanism of selective targeting of mutant KRAS-expressing cancers via MTORC2-mediated AKT activation and ROS-dependent mitofission, which could have potential therapeutic implications given the relative lack of direct RAS-targeting strategies in cancer.
    Keywords:  AKT; DNM1L; KRAS; MTORC2; ROS; mitofission
    DOI:  https://doi.org/10.1080/15548627.2024.2307224
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