bims-mitdyn 2023-01-15 papers

bims-mitdyn

Biomed News

on Mitochondrial dynamics: mechanisms

Issue of 2023–01–15
eleven papers selected by
Edmond Chan, Queen’s University, School of Medicine

Small molecule agonist of mitochondrial fusion repairs mitochondrial dysfunction.
OGT controls mammalian cell viability by regulating the proteasome/mTOR/ mitochondrial axis.
Cryo-EM structures of mitochondrial respiratory complex I from Drosophila melanogaster.
Disruption of mitochondrial dynamics triggers muscle inflammation through interorganellar contacts and mitochondrial DNA mislocation.
The mitophagy receptor BNIP3 is critical for the regulation of metabolic homeostasis and mitochondrial function in the nucleus pulposus cells of the intervertebral disc.
Monitoring Mitochondrial Protein Import Using Mitochondrial Targeting Sequence (MTS)-eGFP.
Microtubule-mitochondrial attachment facilitates cell division symmetry and mitochondrial partitioning in fission yeast.
A tale of two pathways: Regulation of proteostasis by UPRmt and MDPs.
Mitochondrial signalling and homeostasis: from cell biology to neurological disease.
Atg1 modulates mitochondrial dynamics to promote germline stem cell maintenance in Drosophila.
Translation initiation of leaderless and polycistronic transcripts in mammalian mitochondria.

Nat Chem Biol. 2023 Jan 12.

Small molecule agonist of mitochondrial fusion repairs mitochondrial dysfunction.

Yingjie Guo, Huan Zhang, Chen Yan, Birong Shen, Yue Zhang, Xiangyang Guo, Sha Sun, Fan Yu, Jiayun Yan, Ronghe Liu, Qianping Zhang, Di Zhang, Haiyang Liu, Yang Liu, Yaoyao Zhang, Wenlei Li, Jiangyu Qin, He Lv, Zhaoxia Wang, Yun Yuan, Jie-Feng Yang, Ya-Ting Zhong, Song Gao, Bing Zhou, Lei Liu, Deling Kong, Xiaojiang Hao, Junjie Hu, Quan Chen.

Membrane dynamics are important to the integrity and function of mitochondria. Defective mitochondrial fusion underlies the pathogenesis of multiple diseases. The ability to target fusion highlights the potential to fight life-threatening conditions. Here we report a small molecule agonist, S89, that specifically promotes mitochondrial fusion by targeting endogenous MFN1. S89 interacts directly with a loop region in the helix bundle 2 domain of MFN1 to stimulate GTP hydrolysis and vesicle fusion. GTP loading or competition by S89 dislodges the loop from the GTPase domain and unlocks the molecule. S89 restores mitochondrial and cellular defects caused by mitochondrial DNA mutations, oxidative stress inducer paraquat, ferroptosis inducer RSL3 or CMT2A-causing mutations by boosting endogenous MFN1. Strikingly, S89 effectively eliminates ischemia/reperfusion (I/R)-induced mitochondrial damage and protects mouse heart from I/R injury. These results reveal the priming mechanism for MFNs and provide a therapeutic strategy for mitochondrial diseases when additional mitochondrial fusion is beneficial.

DOI: https://doi.org/10.1038/s41589-022-01224-y
Proc Natl Acad Sci U S A. 2023 Jan 17. 120(3): e2218332120

OGT controls mammalian cell viability by regulating the proteasome/mTOR/ mitochondrial axis.

Xiang Li, Xiaojing Yue, Hugo Sepulveda, Rajan A Burt, David A Scott, Steven A Carr, Samuel A Myers, Anjana Rao.

  O-GlcNAc transferase (OGT) modifies serine and threonine residues on nuclear and cytosolic proteins with O-linked N-acetylglucosamine (GlcNAc). OGT is essential for mammalian cell viability, but the underlying mechanisms are still enigmatic. We performed a genome-wide CRISPR-Cas9 screen in mouse embryonic stem cells (mESCs) to identify candidates whose depletion rescued the block in cell proliferation induced by OGT deficiency. We show that the block in cell proliferation in OGT-deficient cells stems from mitochondrial dysfunction secondary to mTOR (mechanistic target of rapamycin) hyperactivation. In normal cells, OGT maintains low mTOR activity and mitochondrial fitness through suppression of proteasome activity; in the absence of OGT, increased proteasome activity results in increased steady-state amino acid levels, which in turn promote mTOR lysosomal translocation and activation, and increased oxidative phosphorylation. mTOR activation in OGT-deficient mESCs was confirmed by an independent phospho-proteomic screen. Our study highlights a unique series of events whereby OGT regulates the proteasome/ mTOR/ mitochondrial axis in a manner that maintains homeostasis of intracellular amino acid levels, mitochondrial fitness, and cell viability. A similar mechanism operates in CD8+ T cells, indicating its generality across mammalian cell types. Manipulating OGT activity may have therapeutic potential in diseases in which this signaling pathway is impaired.

Keywords:  OGT; genome-wide CRISPR/Cas9 screen; mTOR; mitochondrion; proteasome

DOI:  https://doi.org/10.1073/pnas.2218332120
Elife. 2023 Jan 09. pii: e84424. [Epub ahead of print]12

Cryo-EM structures of mitochondrial respiratory complex I from Drosophila melanogaster.

Ahmed-Noor A Agip, Injae Chung, Alvaro Sanchez-Martinez, Alexander J Whitworth, Judy Hirst.

  Respiratory complex I powers ATP synthesis by oxidative phosphorylation, exploiting the energy from NADH oxidation by ubiquinone to drive protons across an energy-transducing membrane. Drosophila melanogaster is a candidate model organism for complex I due to its high evolutionary conservation with the mammalian enzyme, well-developed genetic toolkit, and complex physiology for studies in specific cell types and tissues. Here, we isolate complex I from Drosophila and determine its structure, revealing a 43-subunit assembly with high structural homology to its 45-subunit mammalian counterpart, including a hitherto unknown homologue to subunit NDUFA3. The major conformational state of the Drosophila enzyme is the mammalian-type 'ready-to-go' active resting state, with a fully ordered and enclosed ubiquinone-binding site, but a subtly altered global conformation related to changes in subunit ND6. The mammalian-type 'deactive' pronounced resting state is not observed: in two minor states the ubiquinone-binding site is unchanged, but a deactive-type p-bulge is present in ND6-TMH3. Our detailed structural knowledge of Drosophila complex I provides a foundation for new approaches to disentangle mechanisms of complex I catalysis and regulation in bioenergetics and physiology.

Keywords:  D. melanogaster; molecular biophysics; structural biology

DOI:  https://doi.org/10.7554/eLife.84424
Nat Commun. 2023 Jan 06. 14(1): 108

Disruption of mitochondrial dynamics triggers muscle inflammation through interorganellar contacts and mitochondrial DNA mislocation.

Andrea Irazoki, Isabel Gordaliza-Alaguero, Emma Frank, Nikolaos Nikiforos Giakoumakis, Jordi Seco, Manuel Palacín, Anna Gumà, Lykke Sylow, David Sebastián, Antonio Zorzano.

Some forms of mitochondrial dysfunction induce sterile inflammation through mitochondrial DNA recognition by intracellular DNA sensors. However, the involvement of mitochondrial dynamics in mitigating such processes and their impact on muscle fitness remain unaddressed. Here we report that opposite mitochondrial morphologies induce distinct inflammatory signatures, caused by differential activation of DNA sensors TLR9 or cGAS. In the context of mitochondrial fragmentation, we demonstrate that mitochondria-endosome contacts mediated by the endosomal protein Rab5C are required in TLR9 activation in cells. Skeletal muscle mitochondrial fragmentation promotes TLR9-dependent inflammation, muscle atrophy, reduced physical performance and enhanced IL6 response to exercise, which improved upon chronic anti-inflammatory treatment. Taken together, our data demonstrate that mitochondrial dynamics is key in preventing sterile inflammatory responses, which precede the development of muscle atrophy and impaired physical performance. Thus, we propose the targeting of mitochondrial dynamics as an approach to treating disorders characterized by chronic inflammation and mitochondrial dysfunction.

DOI: https://doi.org/10.1038/s41467-022-35732-1
Autophagy. 2023 Jan 10. 1-23

The mitophagy receptor BNIP3 is critical for the regulation of metabolic homeostasis and mitochondrial function in the nucleus pulposus cells of the intervertebral disc.

Vedavathi Madhu, Miriam Hernandez-Meadows, Paige K Boneski, Yunping Qiu, Anyonya R Guntur, Irwin J Kurland, Ruteja A Barve, Makarand V Risbud.

  The contribution of mitochondria to the metabolic function of hypoxic NP cells has been overlooked. We have shown that NP cells contain networked mitochondria and that mitochondrial translocation of BNIP3 mediates hypoxia-induced mitophagy. However, whether BNIP3 also plays a role in governing mitochondrial function and metabolism in hypoxic NP cells is not known. BNIP3 knockdown altered mitochondrial morphology, and number, and increased mitophagy. Interestingly, BNIP3 deficiency in NP cells reduced glycolytic capacity reflected by lower production of lactate/H+ and lower ATP production rate. Widely targeted metabolic profiling and flux analysis using 1-2-13C-glucose showed that the BNIP3 loss resulted in redirection of glycolytic flux into pentose phosphate and hexosamine biosynthesis as well as pyruvate resulting in increased TCA flux. An overall reduction in one-carbon metabolism was noted suggesting reduced biosynthesis. U13C-glutamine flux analysis showed preservation of glutamine utilization to maintain TCA intermediates. The transcriptomic analysis of the BNIP3-deficient cells showed dysregulation of cellular functions including membrane and cytoskeletal integrity, ECM-growth factor signaling, and protein quality control with an overall increase in themes related to angiogenesis and innate immune response. Importantly, we observed strong thematic similarities with the transcriptome of a subset of human degenerative samples. Last, we noted increased autophagic flux, decreased disc height index and aberrant COL10A1/collagen X expression, signs of early disc degeneration in young adult bnip3 knockout mice. These results suggested that in addition to mitophagy regulation, BNIP3 plays a role in maintaining mitochondrial function and metabolism, and dysregulation of mitochondrial homeostasis could promote disc degeneration.Abbreviations: ECAR extracellular acidification rate; HIF hypoxia inducible factor; MFA metabolic flux analysis; NP nucleus pulposus; OCR oxygen consumption rate; ShBnip3 short-hairpin Bnip3.

Keywords:  BNIP3; disc degeneration; hypoxia; intervertebral disc; metabolism; mitochondria; mitophagy; nucleus pulposus

DOI:  https://doi.org/10.1080/15548627.2022.2162245
Bio Protoc. 2022 Dec 20. pii: e4578. [Epub ahead of print]12(24):

Monitoring Mitochondrial Protein Import Using Mitochondrial Targeting Sequence (MTS)-eGFP.

Jonas B Michaelis, Süleyman Bozkurt, Jasmin A Schäfer, Christian Münch.

  Mitochondria are cellular organelles essential for the function and survival of eukaryotic cells. Nearly all mitochondrial proteins are nuclear-encoded and require mitochondrial import upon their synthesis in the cytosol. Various approaches have been described to study mitochondrial protein import, such as monitoring the entry of radiolabeled proteins into purified mitochondria or quantifying newly synthesized proteins within mitochondria by proteomics. Here, we provide a detailed protocol for a commonly used and straightforward assay that quantitatively examines mitochondrial protein import by monitoring the co-localization of mitochondrially targeted enhanced green fluorescent protein (eGFP) with the mitochondrial fluorescence dye MitoTracker TM Deep Red FM by live cell imaging. We describe the preparation and use of a stable mammalian cell line inducibly expressing a mitochondrial targeting sequence (MTS)-eGFP, followed by quantitative image analysis using an open-source ImageJ-based plugin. This inducible expression system avoids the need for transient transfection while enabling titration of MTS-eGFP expression and thereby avoiding protein folding stress. Overall, the assay provides a simple and robust approach to assess mitochondrial import capacity of cells in various disease-related settings. This protocol was validated in: Mol Cell (2021), DOI: 10.1016/j.molcel.2021.11.004 Graphical abstract.

Keywords:   Live cell imaging ; Microscopy ; Mitochondria ; Mitochondrial protein import ; Protein translocation

DOI:  https://doi.org/10.21769/BioProtoc.4578
J Cell Sci. 2023 Jan 01. pii: jcs260705. [Epub ahead of print]136(1):

Microtubule-mitochondrial attachment facilitates cell division symmetry and mitochondrial partitioning in fission yeast.

Leeba Ann Chacko, Felix Mikus, Nicholas Ariotti, Gautam Dey, Vaishnavi Ananthanarayanan.

  Association with microtubules inhibits the fission of mitochondria in Schizosaccharomyces pombe. Here, we show that this attachment of mitochondria to microtubules is an important cell-intrinsic factor in determining cell division symmetry. By comparing mutant cells that exhibited enhanced attachment and no attachment of mitochondria to microtubules (Dnm1Δ and Mmb1Δ, respectively), we show that microtubules in these mutants displayed aberrant dynamics compared to wild-type cells, which resulted in errors in nuclear positioning. This translated to cell division asymmetry in a significant proportion of both Dnm1Δ and Mmb1Δ cells. Asymmetric division in Dnm1Δ and Mmb1Δ cells resulted in unequal distribution of mitochondria, with the daughter cell that received more mitochondria growing faster than the other daughter cell. Taken together, we show the existence of homeostatic feedback controls between mitochondria and microtubules in fission yeast, which directly influence mitochondrial partitioning and, thereby, cell growth. This article has an associated First Person interview with the first author of the paper.

Keywords:  Cell division; Microtubules; Mitochondria; Mitochondrial partitioning

DOI:  https://doi.org/10.1242/jcs.260705
Curr Opin Neurobiol. 2023 Jan 06. pii: S0959-4388(22)00167-2. [Epub ahead of print]78 102673

A tale of two pathways: Regulation of proteostasis by UPRmt and MDPs.

Angela Johns, Ryo Higuchi-Sanabria, Max A Thorwald, David Vilchez.

Mitochondrial fitness is critical to organismal health and its impairment is associated with aging and age-related diseases. As such, numerous quality control mechanisms exist to preserve mitochondrial stability, including the unfolded protein response of the mitochondria (UPRmt). The UPRmt is a conserved mechanism that drives the transcriptional activation of mitochondrial chaperones, proteases, autophagy (mitophagy), and metabolism to promote restoration of mitochondrial function under stress conditions. UPRmt has direct ramifications in aging, and its activation is often ascribed to improve health whereas its dysfunction tends to correlate with disease. This review pairs a description of the most recent findings within the field of UPRmt with a more poorly understood field: mitochondria-derived peptides (MDPs). Similar to UPRmt, MDPs are microproteins derived from the mitochondria that can impact organismal health and longevity. We then highlight a tantalizing interconnection between UPRmt and MDPs wherein both mechanisms may be efficiently coordinated to maintain organismal health.

DOI: https://doi.org/10.1016/j.conb.2022.102673
Trends Neurosci. 2023 Jan 10. pii: S0166-2236(22)00239-9. [Epub ahead of print]

Mitochondrial signalling and homeostasis: from cell biology to neurological disease.

Jack J Collier, Monika Oláhová, Thomas G McWilliams, Robert W Taylor.

  Efforts to understand how mitochondrial dysfunction contributes to neurodegeneration have primarily focussed on the role of mitochondria in neuronal energy metabolism. However, progress in understanding the etiological nature of emerging mitochondrial functions has yielded new ideas about the mitochondrial basis of neurological disease. Studies aimed at deciphering how mitochondria signal through interorganellar contacts, vesicular trafficking, and metabolic transmission have revealed that mitochondrial regulation of immunometabolism, cell death, organelle dynamics, and neuroimmune interplay are critical determinants of neural health. Moreover, the homeostatic mechanisms that exist to protect mitochondrial health through turnover via nanoscale proteostasis and lysosomal degradation have become integrated within mitochondrial signalling pathways to support metabolic plasticity and stress responses in the nervous system. This review highlights how these distinct mitochondrial pathways converge to influence neurological health and contribute to disease pathology.

Keywords:  immunity; inflammation; metabolism; mitochondrial-derived vesicles; mitochondria–lysosome axis; quality control

DOI:  https://doi.org/10.1016/j.tins.2022.12.001
Biochem Biophys Res Commun. 2022 Nov 30. pii: S0006-291X(22)01633-3. [Epub ahead of print]643 192-202

Atg1 modulates mitochondrial dynamics to promote germline stem cell maintenance in Drosophila.

Minal S Ayachit, Bhupendra V Shravage.

  Mitochondrial dynamics (fusion and fission) are necessary for stem cell maintenance and differentiation. However, the relationship between mitophagy, mitochondrial dynamics and stem cell exhaustion needs to be clearly understood. Here we report the multifaceted role of Atg1 in mitophagy, mitochondrial dynamics and stem cell maintenance in female germline stem cells (GSCs) in Drosophila. We found that depletion of Atg1 in GSCs leads to impaired autophagy and mitophagy as measured by reduced formation of autophagosomes, increased accumulation of p62/Ref (2)P and accumulation of damaged mitochondria. Disrupting Atg1 function led to mitochondrial fusion in developing cysts. The fusion resulted from an increase in Marf levels in both GSCs and cysts, and the fusion phenotype could be rescued by overexpression of Drp1 or by depleting Marf via RNAi in Atg1-depleted cyst cells. Interestingly, double knockdown of both Atg1:Drp1 led to the significant loss of germ cells (GCs) as compared to Atg1KD and Drp1KD. Strikingly, Atg1:Marf double knockdown leads to a dramatic loss of GSCs, GCs and a total loss of vitellogenic stages, suggesting a block in oogenesis. Overall, our results demonstrate that Drp1, Marf and Atg1 function together to influence female GSC maintenance, their differentiation into cysts and oogenesis in Drosophila.

Keywords:  Atg1; Drp1; Homeostasis; Marf; Mitochondria; Mitophagy; Oogenesis

DOI:  https://doi.org/10.1016/j.bbrc.2022.11.076
Nucleic Acids Res. 2023 Jan 11. pii: gkac1233. [Epub ahead of print]

Translation initiation of leaderless and polycistronic transcripts in mammalian mitochondria.

Cristina Remes, Anas Khawaja, Sarah F Pearce, Adam M Dinan, Shreekara Gopalakrishna, Miriam Cipullo, Vasileios Kyriakidis, Jingdian Zhang, Xaquin Castro Dopico, Olessya Yukhnovets, Ilian Atanassov, Andrew E Firth, Barry Cooperman, Joanna Rorbach.

The synthesis of mitochondrial OXPHOS complexes is central to cellular metabolism, yet many molecular details of mitochondrial translation remain elusive. It has been commonly held view that translation initiation in human mitochondria proceeded in a manner similar to bacterial systems, with the mitoribosomal small subunit bound to the initiation factors, mtIF2 and mtIF3, along with initiator tRNA and an mRNA. However, unlike in bacteria, most human mitochondrial mRNAs lack 5' leader sequences that can mediate small subunit binding, raising the question of how leaderless mRNAs are recognized by mitoribosomes. By using novel in vitro mitochondrial translation initiation assays, alongside biochemical and genetic characterization of cellular knockouts of mitochondrial translation factors, we describe unique features of translation initiation in human mitochondria. We show that in vitro, leaderless mRNA transcripts can be loaded directly onto assembled 55S mitoribosomes, but not onto the mitoribosomal small subunit (28S), in a manner that requires initiator fMet-tRNAMet binding. In addition, we demonstrate that in human cells and in vitro, mtIF3 activity is not required for translation of leaderless mitochondrial transcripts but is essential for translation of ATP6 in the case of the bicistronic ATP8/ATP6 transcript. Furthermore, we show that mtIF2 is indispensable for mitochondrial protein synthesis. Our results demonstrate an important evolutionary divergence of the mitochondrial translation system and further our fundamental understanding of a process central to eukaryotic metabolism.

DOI: https://doi.org/10.1093/nar/gkac1233