bims-mitdyn Biomed News
on Mitochondrial dynamics: mechanisms
Issue of 2021‒08‒22
fifteen papers selected by
Edmond Chan
Queen’s University, School of Medicine

  1. iScience. 2021 Aug 20. 24(8): 102895
      The mitochondrial uniporter is a Ca2+-selective ion-conducting channel in the inner mitochondrial membrane that is involved in various cellular processes. The components of this uniporter, including the pore-forming membrane subunit MCU and the modulatory subunits MCUb, EMRE, MICU1, and MICU2, have been identified in recent years. Previously, extensive studies revealed various aspects of uniporter activities and proposed multiple regulatory models of mitochondrial Ca2+ uptake. Recently, the individual auxiliary components of the uniporter and its holocomplex have been structurally characterized, providing the first insight into the component structures and their spatial relationship within the context of the uniporter. Here, we review recent uniporter structural studies in an attempt to establish an architectural framework, elucidating the mechanism that governs mitochondrial Ca2+ uptake and regulation, and to address some apparent controversies. This information could facilitate further characterization of mitochondrial Ca2+ permeation and a better understanding of uniporter-related disease conditions.
    Keywords:  Ion; Membranes; Molecular biology
  2. Proc Natl Acad Sci U S A. 2021 Aug 24. pii: e2101674118. [Epub ahead of print]118(34):
      The inability of adult mammalian cardiomyocytes to proliferate underpins the development of heart failure following myocardial injury. Although the newborn mammalian heart can spontaneously regenerate for a short period of time after birth, this ability is lost within the first week after birth in mice, partly due to increased mitochondrial reactive oxygen species (ROS) production which results in oxidative DNA damage and activation of DNA damage response. This increase in ROS levels coincides with a postnatal switch from anaerobic glycolysis to fatty acid (FA) oxidation by cardiac mitochondria. However, to date, a direct link between mitochondrial substrate utilization and oxidative DNA damage is lacking. Here, we generated ROS-sensitive fluorescent sensors targeted to different subnuclear compartments (chromatin, heterochromatin, telomeres, and nuclear lamin) in neonatal rat ventricular cardiomyocytes, which allowed us to determine the spatial localization of ROS in cardiomyocyte nuclei upon manipulation of mitochondrial respiration. Our results demonstrate that FA utilization by the mitochondria induces a significant increase in ROS detection at the chromatin level compared to other nuclear compartments. These results indicate that mitochondrial metabolic perturbations directly alter the nuclear redox status and that the chromatin appears to be particularly sensitive to the prooxidant effect of FA utilization by the mitochondria.
    Keywords:  metabolism; mitochondria; reactive oxygen species
  3. Elife. 2021 Aug 20. pii: e69142. [Epub ahead of print]10
      The trafficking of specific protein cohorts to correct subcellular locations at correct times is essential for every signaling and regulatory process in biology. Gene perturbation screens could provide a powerful approach to probe the molecular mechanisms of protein trafficking, but only if protein localization or mislocalization can be tied to a simple and robust phenotype for cell selection, such as cell proliferation or fluorescence-activated cell sorting (FACS). To empower the study of protein trafficking processes with gene perturbation, we developed a genetically-encoded molecular tool named HiLITR. HiLITR converts protein colocalization into proteolytic release of a membrane-anchored transcription factor, which drives the expression of a chosen reporter gene. Using HiLITR in combination with FACS-based CRISPRi screening in human cell lines, we identified genes that influence the trafficking of mitochondrial and ER tail-anchored proteins. We show that loss of the SUMO E1 component SAE1 results in mislocalization and destabilization of many mitochondrial tail-anchored proteins. We also demonstrate a distinct regulatory role for EMC10 in the ER membrane complex, opposing the transmembrane-domain insertion activity of the complex. Through transcriptional integration of complex cellular functions, HiLITR expands the scope of biological processes that can be studied by genetic perturbation screening technologies.
    Keywords:  biochemistry; cell biology; chemical biology; human
  4. EMBO Rep. 2021 Aug 17. e52445
      In eukaryotic cells, proteins are targeted to their final subcellular locations with precise timing. A key underlying mechanism is the active transport of cognate mRNAs, which in many systems can be linked intimately to membrane trafficking. A prominent example is the long-distance endosomal transport of mRNAs and their local translation. Here, we describe current highlights of fundamental mechanisms of the underlying transport process as well as of biological functions ranging from endosperm development in plants to fungal pathogenicity and neuronal processes. Translation of endosome-associated mRNAs often occurs at the cytoplasmic surface of endosomes, a process that is needed for membrane-assisted formation of heteromeric protein complexes and for accurate subcellular targeting of proteins. Importantly, endosome-coupled translation of mRNAs encoding mitochondrial proteins, for example, seems to be particularly important for efficient organelle import and for regulating subcellular mitochondrial activity. In essence, these findings reveal a new mechanism of loading newly synthesised proteins onto endocytic membranes enabling intimate crosstalk between organelles. The novel link between endosomes and mitochondria adds an inspiring new level of complexity to trafficking and organelle biology.
    Keywords:  RNA transport; endosomes; local translation; microtubules; mitochondria; organelle
  5. Commun Biol. 2021 Aug 16. 4(1): 974
      Human ATP-dependent Lon protease (LONP1) forms homohexameric, ring-shaped complexes. Depletion of LONP1 causes aggregation of a broad range of proteins in the mitochondrial matrix and decreases the levels of their soluble forms. The ATP hydrolysis activity, but not protease activity, of LONP1 is critical for its chaperone-like anti-aggregation activity. LONP1 forms a complex with the import machinery and an incoming protein, and protein aggregation is linked with matrix protein import. LONP1 also contributes to the degradation of imported, aberrant, unprocessed proteins using its protease activity. Taken together, our results show that LONP1 functions as a gatekeeper for specific proteins imported into the mitochondrial matrix.
  6. Cell Death Dis. 2021 Aug 17. 12(9): 796
      Mitochondria are highly dynamic organelles with strict quality control processes that maintain cellular homeostasis. Within axons, coordinated cycles of fission-fusion mediated by dynamin related GTPase protein (DRP1) and mitofusins (MFN), together with regulated motility of healthy mitochondria anterogradely and damaged/oxidized mitochondria retrogradely, control mitochondrial shape, distribution and size. Disruption of this tight regulation has been linked to aberrant oxidative stress and mitochondrial dysfunction causing mitochondrial disease and neurodegeneration. Although pharmacological induction of Parkinson's disease (PD) in humans/animals with toxins or in mice overexpressing α-synuclein (α-syn) exhibited mitochondrial dysfunction and oxidative stress, mice lacking α-syn showed resistance to mitochondrial toxins; yet, how α-syn influences mitochondrial dynamics and turnover is unclear. Here, we isolate the mechanistic role of α-syn in mitochondrial homeostasis in vivo in a humanized Drosophila model of Parkinson's disease (PD). We show that excess α-syn causes fragmented mitochondria, which persists with either truncation of the C-terminus (α-syn1-120) or deletion of the NAC region (α-synΔNAC). Using in vivo oxidation reporters Mito-roGFP2-ORP1/GRX1 and MitoTimer, we found that α-syn-mediated fragments were oxidized/damaged, but α-syn1-120-induced fragments were healthy, suggesting that the C-terminus is required for oxidation. α-syn-mediated oxidized fragments showed biased retrograde motility, but α-syn1-120-mediated healthy fragments did not, demonstrating that the C-terminus likely mediates the retrograde motility of oxidized mitochondria. Depletion/inhibition or excess DRP1-rescued α-syn-mediated fragmentation, oxidation, and the biased retrograde motility, indicating that DRP1-mediated fragmentation is likely upstream of oxidation and motility changes. Further, excess PINK/Parkin, two PD-associated proteins that function to coordinate mitochondrial turnover via induction of selective mitophagy, rescued α-syn-mediated membrane depolarization, oxidation and cell death in a C-terminus-dependent manner, suggesting a functional interaction between α-syn and PINK/Parkin. Taken together, our findings identify distinct roles for α-syn in mitochondrial homeostasis, highlighting a previously unknown pathogenic pathway for the initiation of PD.
  7. STAR Protoc. 2021 Sep 17. 2(3): 100721
      Disruption of mitochondrial morphology occurs during various diseases, but the biological significance is not entirely clear. Here, we describe a detailed step-by-step protocol for a chemically inducible dimerization system-based synthetic protein device, termed inducible counter mitochondrial morphology. This system allows artificial manipulation of mitochondrial morphology on a timescale of minutes in living mammalian cells. We also describe an AI-assisted imaging processing approach. For complete details on the use and execution of this protocol, please refer to Miyamoto et al., 2021.
    Keywords:  Biotechnology and bioengineering; Cell Biology; Molecular/Chemical Probes; Protein Biochemistry
  8. Nat Methods. 2021 Aug 19.
      Mitochondria display complex morphology and movements, which complicates their segmentation and tracking in time-lapse images. Here, we introduce Mitometer, an algorithm for fast, unbiased, and automated segmentation and tracking of mitochondria in live-cell two-dimensional and three-dimensional time-lapse images. Mitometer requires only the pixel size and the time between frames to identify mitochondrial motion and morphology, including fusion and fission events. The segmentation algorithm isolates individual mitochondria via a shape- and size-preserving background removal process. The tracking algorithm links mitochondria via differences in morphological features and displacement, followed by a gap-closing scheme. Using Mitometer, we show that mitochondria of triple-negative breast cancer cells are faster, more directional, and more elongated than those in their receptor-positive counterparts. Furthermore, we show that mitochondrial motility and morphology in breast cancer, but not in normal breast epithelia, correlate with metabolic activity. Mitometer is an unbiased and user-friendly tool that will help resolve fundamental questions regarding mitochondrial form and function.
  9. Commun Biol. 2021 Aug 19. 4(1): 989
      Mouse models of genetic mitochondrial disorders are generally used to understand specific molecular defects and their biochemical consequences, but rarely to map compensatory changes allowing survival. Here we took advantage of the extraordinary mitochondrial resilience of hepatic Lrpprc knockout mice to explore this question using native proteomics profiling and lipidomics. In these mice, low levels of the mtRNA binding protein LRPPRC induce a global mitochondrial translation defect and a severe reduction (>80%) in the assembly and activity of the electron transport chain (ETC) complex IV (CIV). Yet, animals show no signs of overt liver failure and capacity of the ETC is preserved. Beyond stimulation of mitochondrial biogenesis, results show that the abundance of mitoribosomes per unit of mitochondria is increased and proteostatic mechanisms are induced in presence of low LRPPRC levels to preserve a balance in the availability of mitochondrial- vs nuclear-encoded ETC subunits. At the level of individual organelles, a stabilization of residual CIV in supercomplexes (SCs) is observed, pointing to a role of these supramolecular arrangements in preserving ETC function. While the SC assembly factor COX7A2L could not contribute to the stabilization of CIV, important changes in membrane glycerophospholipid (GPL), most notably an increase in SC-stabilizing cardiolipins species (CLs), were observed along with an increased abundance of other supramolecular assemblies known to be stabilized by, and/or participate in CL metabolism. Together these data reveal a complex in vivo network of molecular adjustments involved in preserving mitochondrial integrity in energy consuming organs facing OXPHOS defects, which could be therapeutically exploited.
  10. Bio Protoc. 2021 Jul 20. 11(14): e4087
      The crucial role of hexokinase 2 (HK2) in the metabolic rewiring of tumors is now well established, which makes it a suitable target for the design of novel therapies. However, hexokinase activity is central to glucose utilization in all tissues; thus, enzymatic inhibition of HK2 can induce severe adverse effects. In an effort to find a selective anti-neoplastic strategy, we exploited an alternative approach based on HK2 detachment from its location on the outer mitochondrial membrane. We designed a HK2-targeting peptide named HK2pep, corresponding to the N-terminal hydrophobic domain of HK2 and armed with a metalloprotease cleavage sequence and a polycation stretch shielded by a polyanion sequence. In the tumor microenvironment, metalloproteases unleash polycations to allow selective plasma membrane permeation in neoplastic cells. HK2pep delivery induces the detachment of HK2 from mitochondria-associated membranes (MAMs) and mitochondrial Ca2+ overload caused by the opening of inositol-3-phosphate receptors on the endoplasmic reticulum (ER) and Ca2+ entry through the plasma membrane leading to Ca2+-mediated calpain activation and mitochondrial depolarization. As a result, HK2pep rapidly elicits death of diverse tumor cell types and dramatically reduces in vivo tumor mass. HK2pep does not affect hexokinase enzymatic activity, avoiding any noxious effect on non-transformed cells. Here, we make available a detailed protocol for the use of HK2pep and to investigate its biological effects, providing a comprehensive panel of assays to quantitate both HK2 enzymatic activity and changes in mitochondrial functions, Ca2+ flux, and cell viability elicited by HK2pep treatment of tumor cells. Graphical abstract: Flowchart for the analysis of the effects of HK2 detachment from MAMs.
    Keywords:   Ca2+; Anti-neoplastic strategy; Cancer; Cell-penetrating peptide; Hexokinase 2; Mitochondria; Mitochondria-associated membranes
  11. Biochim Biophys Acta Mol Cell Res. 2021 Aug 13. pii: S0167-4889(21)00170-1. [Epub ahead of print] 119116
      The neuropathological hallmarks of Alzheimer's Disease are plaques and neurofibrillary tangles. Yet, Alzheimer's is a complex disease with many contributing factors, such as energy-metabolic changes, which have been documented in autopsy brains from individuals with Alzheimer's and animal disease models alike. One conceivable explanation is that the interplay of age-related extracellular and intracellular alterations pertaining to Alzheimer's, such as cerebrovascular changes, protein aggregates and inflammation, evoke a mitochondrial response. However, it is not clear if and how mitochondria can contribute to Alzheimer's pathophysiology. This study focuses on one particular aspect of this question by investigating the functional interaction between the microtubule-associated protein tau and the mitochondrial inner membrane fusion machinery, which shows alterations in Alzheimer's brains. OPA1 is an essential inner membrane-fusion protein regulated by the two membrane proteases OMA1 and YME1L1. Assessment of OPA1 proteolysis-usually found in dividing mitochondria-and posttranslational tau modifications in mouse and human neuroblastoma cells under different experimental conditions clarified the relationship between these two pathways: OPA1 hydrolysis and phosphorylation or dephosphorylation of tau may coincide, but are not causally related. OPA1 cleavage did not alter tau's phosphorylation pattern. Conversely, tau's phosphorylation state did not induce nor correlate with OPA1 proteolysis. These results irrefutably demonstrate that there is no direct functional interaction between posttranslational tau modifications and the regulation of the OMA1-OPA1 pathway, which implies a common root cause modulating both pathways in Alzheimer's.
    Keywords:  Alzheimer’s; Mitochondria; OMA1; fusion; tau hyperphosphorylation
  12. Curr Res Physiol. 2021 ;4 163-176
      Folding of the mitochondrial inner membrane (IM) into cristae greatly increases the ATP-generating surface area, S IM, per unit volume but also creates diffusional bottlenecks that could limit reaction rates inside mitochondria. This study explores possible effects of inner membrane folding on mitochondrial ATP output, using a mathematical model for energy metabolism developed by the Jafri group and two- and three-dimensional spatial models for mitochondria, implemented on the Virtual Cell platform. Simulations demonstrate that cristae are micro-compartments functionally distinct from the cytosol. At physiological steady states, standing gradients of ADP form inside cristae that depend on the size and shape of the compartments, and reduce local flux (rate per unit area) of the adenine nucleotide translocase. This causes matrix ADP levels to drop, which in turn reduces the flux of ATP synthase. The adverse effects of membrane folding on reaction fluxes increase with crista length and are greater for lamellar than tubular crista. However, total ATP output per mitochondrion is the product of flux of ATP synthase and S IM which can be two-fold greater for mitochondria with lamellar than tubular cristae, resulting in greater ATP output for the former. The simulations also demonstrate the crucial role played by intracristal kinases (adenylate kinase, creatine kinase) in maintaining the energy advantage of IM folding.
    Keywords:  ATP synthesis; Computational modeling; Cristae; Energy metabolism; Kinases; Mitochondria
  13. EMBO Rep. 2021 Aug 19. e53790
      Zellweger spectrum disorder (ZSD) is the most severe peroxisomal biogenesis disorder (PBD). Why ZSD patients not only loose functional peroxisomes but also present with severe mitochondrial dysfunction was a long-standing mystery. In this issue, Nuebel et al (2021) identified that loss of peroxisomes leads to re-routing of peroxisomal proteins to mitochondria, thereby impairing mitochondrial structure and function. The findings provide the first molecular understanding of the mitochondrial-peroxisomal link in ZSD.
  14. Curr Opin Microbiol. 2021 Aug 16. pii: S1369-5274(21)00101-6. [Epub ahead of print]63 189-194
      Invading microbes occupy the host cytosol and take up nutrients on which host organelles are also dependent. Thus, host organelles are poised to interact with intracellular microbes. Despite the essential role of host mitochondria in cellular metabolic homeostasis and in mediating cellular responses to microbial infection, we know little of how these organelles interact with intracellular pathogens, and how such interactions affect disease pathogenesis. Here, we give an overview of the different classes of physical and metabolic interactions reported to occur between mitochondria and eukaryotic pathogens. Investigating the underlying molecular mechanisms and functions of such interactions will reveal novel aspects of infection biology.