bims-mitdyn Biomed News
on Mitochondrial dynamics: mechanisms
Issue of 2021‒06‒20
fourteen papers selected by
Edmond Chan
Queen’s University, School of Medicine

  1. JCI Insight. 2021 Jun 17. pii: 138835. [Epub ahead of print]
      Cancer cells re-program cellular metabolism to maintain adequate nutrient pools to sustain proliferation. Moreover, autophagy is a regulated mechanism to breakdown dysfunctional cellular components and recycle cellular nutrients. However, the requirement for autophagy and the integration in cancer cell metabolism is not clear in colon cancer. Here we show a cell-autonomous dependency of autophagy for cell growth in colorectal cancer. Loss of epithelial autophagy inhibits tumor growth in both sporadic and colitis associated cancer models. Genetic and pharmacological inhibition of autophagy inhibits cell growth in colon cancer-derived cell lines and patient-derived enteroid models. Importantly, normal colon epithelium and patient-derived normal enteroid growth was not decreased following autophagy inhibition. To couple the role of autophagy to cellular metabolism, a cell culture screen in conjunction with metabolomic analysis was performed. We identified a critical role of autophagy to maintain mitochondrial metabolites for growth. Loss of mitochondrial recycling through inhibition of mitophagy hinders colon cancer cell growth. These findings have revealed a cell-autonomous role of autophagy that plays a critical role in regulating nutrient pools in vivo and in cell models and provides therapeutic targets for colon cancer.
    Keywords:  Colorectal cancer; Gastroenterology; Oncology
  2. Cell Rep. 2021 Jun 15. pii: S2211-1247(21)00594-5. [Epub ahead of print]35(11): 109235
      T regulatory (Treg) cells are crucial to maintain immune tolerance and repress antitumor immunity, but the mechanisms governing their cellular redox homeostasis remain elusive. We report that glutathione peroxidase 4 (Gpx4) prevents Treg cells from lipid peroxidation and ferroptosis in regulating immune homeostasis and antitumor immunity. Treg-specific deletion of Gpx4 impairs immune homeostasis without substantially affecting survival of Treg cells at steady state. Loss of Gpx4 results in excessive accumulation of lipid peroxides and ferroptosis of Treg cells upon T cell receptor (TCR)/CD28 co-stimulation. Neutralization of lipid peroxides and blockade of iron availability rescue ferroptosis of Gpx4-deficient Treg cells. Moreover, Gpx4-deficient Treg cells elevate generation of mitochondrial superoxide and production of interleukin-1β (IL-1β) that facilitates T helper 17 (TH17) responses. Furthermore, Treg-specific ablation of Gpx4 represses tumor growth and concomitantly potentiates antitumor immunity. Our studies establish a crucial role for Gpx4 in protecting activated Treg cells from lipid peroxidation and ferroptosis and offer a potential therapeutic strategy to improve cancer treatment.
    Keywords:  B16 melanoma; Gpx4; T(H)17 responses; TCR stimulation; Treg cells; ferroptosis; lipid peroxidation; mitochondria; tumor immune evasion
  3. Cell Rep. 2021 Jun 15. pii: S2211-1247(21)00596-9. [Epub ahead of print]35(11): 109237
      The formation of stress granules (SGs) is an essential aspect of the cellular response to many kinds of stress, but its adaptive role is far from clear. SG dysfunction is implicated in aging-onset neurodegenerative diseases, prompting interest in their physiological function. Here, we report that during starvation stress, SGs interact with mitochondria and regulate metabolic remodeling. We show that SG formation leads to a downregulation of fatty acid β-oxidation (FAO) through the modulation of mitochondrial voltage-dependent anion channels (VDACs), which import fatty acids (FAs) into mitochondria. The subsequent decrease in FAO during long-term starvation reduces oxidative damage and rations FAs for longer use. Failure to form SGs, whether caused by the genetic deletion of SG components or an amyotrophic lateral sclerosis (ALS)-associated mutation, translates into an inability to downregulate FAO. Because metabolic dysfunction is a common pathological element of neurodegenerative diseases, including ALS, our findings provide a direction for studying the clinical relevance of SGs.
    Keywords:  ALS; VDAC; fatty acid oxidation; lipid droplet; lipid metabolism; metabolic adaptation; mitochondria; porin; starvation; stress granule
  4. Nat Commun. 2021 06 16. 12(1): 3673
      Mitochondrial ribosomes (mitoribosomes) synthesize a critical set of proteins essential for oxidative phosphorylation. Therefore, mitoribosomal function is vital to the cellular energy supply. Mitoribosome biogenesis follows distinct molecular pathways that remain poorly understood. Here, we determine the cryo-EM structures of mitoribosomes isolated from human cell lines with either depleted or overexpressed mitoribosome assembly factor GTPBP5, allowing us to capture consecutive steps during mitoribosomal large subunit (mt-LSU) biogenesis. Our structures provide essential insights into the last steps of 16S rRNA folding, methylation and peptidyl transferase centre (PTC) completion, which require the coordinated action of nine assembly factors. We show that mammalian-specific MTERF4 contributes to the folding of 16S rRNA, allowing 16 S rRNA methylation by MRM2, while GTPBP5 and NSUN4 promote fine-tuning rRNA rearrangements leading to PTC formation. Moreover, our data reveal an unexpected involvement of the elongation factor mtEF-Tu in mt-LSU assembly, where mtEF-Tu interacts with GTPBP5, similar to its interaction with tRNA during translational elongation.
  5. Nat Commun. 2021 06 16. 12(1): 3671
      Mitochondrial ribosomes are specialized for the synthesis of membrane proteins responsible for oxidative phosphorylation. Mammalian mitoribosomes have diverged considerably from the ancestral bacterial ribosomes and feature dramatically reduced ribosomal RNAs. The structural basis of the mammalian mitochondrial ribosome assembly is currently not well understood. Here we present eight distinct assembly intermediates of the human large mitoribosomal subunit involving seven assembly factors. We discover that the NSUN4-MTERF4 dimer plays a critical role in the process by stabilizing the 16S rRNA in a conformation that exposes the functionally important regions of rRNA for modification by the MRM2 methyltransferase and quality control interactions with the conserved mitochondrial GTPase MTG2 that contacts the sarcin-ricin loop and the immature active site. The successive action of these factors leads to the formation of the peptidyl transferase active site of the mitoribosome and the folding of the surrounding rRNA regions responsible for interactions with tRNAs and the small ribosomal subunit.
  6. Nat Commun. 2021 06 16. 12(1): 3672
      Ribosome biogenesis requires auxiliary factors to promote folding and assembly of ribosomal proteins and RNA. Particularly, maturation of the peptidyl transferase center (PTC) is mediated by conserved GTPases, but the molecular basis is poorly understood. Here, we define the mechanism of GTPase-driven maturation of the human mitochondrial large ribosomal subunit (mtLSU) using endogenous complex purification, in vitro reconstitution and cryo-EM. Structures of transient native mtLSU assembly intermediates that accumulate in GTPBP6-deficient cells reveal how the biogenesis factors GTPBP5, MTERF4 and NSUN4 facilitate PTC folding. Addition of recombinant GTPBP6 reconstitutes late mtLSU biogenesis in vitro and shows that GTPBP6 triggers a molecular switch and progression to a near-mature PTC state. Additionally, cryo-EM analysis of GTPBP6-treated mature mitochondrial ribosomes reveals the structural basis for the dual-role of GTPBP6 in ribosome biogenesis and recycling. Together, these results provide a framework for understanding step-wise PTC folding as a critical conserved quality control checkpoint.
  7. J Biol Chem. 2021 Jun 15. pii: S0021-9258(21)00680-3. [Epub ahead of print] 100880
      More than half a century ago, reversible protein phosphorylation was first linked to mitochondrial metabolism through the regulation of pyruvate dehydrogenase. Since this discovery, the number of identified mitochondrial protein phosphorylation sites has increased by orders of magnitude, driven largely by technological advances in mass spectrometry-based phosphoproteomics. However, the majority of these modifications remain uncharacterized, rendering their function and relevance unclear. Nonetheless, recent studies have shown that disruption of resident mitochondrial protein phosphatases causes substantial metabolic dysfunction across organisms, suggesting that proper management of mitochondrial phosphorylation is vital for organellar and organismal homeostasis. While these data suggest that phosphorylation within mitochondria is of critical importance, significant gaps remain in our knowledge of how these modifications influence organellar function. Here, we curate publicly available datasets to map the extent of protein phosphorylation within mammalian mitochondria and to highlight the known functions of mitochondrial-resident phosphatases. We further propose models by which phosphorylation may affect mitochondrial enzyme activities, protein import and processing, and overall organellar homeostasis.
    Keywords:  mitochondria; phosphoproteomics; protein kinase; protein phosphatase; protein phosphorylation
  8. Mol Biol Cell. 2021 Jun 16. mbcE21030097
      Membrane contact sites (MCSs) between the endoplasmic reticulum (ER) and mitochondria are emerging as critical hubs for diverse cellular events, and alterations in the extent of these contacts are linked to neurodegenerative diseases. However, the mechanisms that control ER-mitochondrial interactions are so far elusive. Here, we demonstrate a key role of vacuolar protein sorting-associated protein 13D (VPS13D) in the negative regulation of ER-mitochondrial MCSs. VPS13D suppression results in extensive ER-mitochondrial tethering, a phenotype that can be substantially rescued by suppression of the tethering proteins VAPB and PTPIP51. VPS13D interacts with valosin-containing protein (VCP/p97) to control the level of ER-resident VAPB at contacts. VPS13D is required for the stability of p97. Functionally, VPS13D suppression leads to severe defects in the mitochondrial morphology, mitochondrial cellular distribution and mitochondrial DNA synthesis. Together our results suggest that VPS13D negatively regulates the ER-mitochondrial MCSs partially through its interactions with VCP/p97. [Media: see text].
  9. Oncogene. 2021 Jun 12.
      SOS1 ablation causes specific defective phenotypes in MEFs including increased levels of intracellular ROS. We showed that the mitochondria-targeted antioxidant MitoTEMPO restores normal endogenous ROS levels, suggesting predominant involvement of mitochondria in generation of this defective SOS1-dependent phenotype. The absence of SOS1 caused specific alterations of mitochondrial shape, mass, and dynamics accompanied by higher percentage of dysfunctional mitochondria and lower rates of electron transport in comparison to WT or SOS2-KO counterparts. SOS1-deficient MEFs also exhibited specific alterations of respiratory complexes and their assembly into mitochondrial supercomplexes and consistently reduced rates of respiration, glycolysis, and ATP production, together with distinctive patterns of substrate preference for oxidative energy metabolism and dependence on glucose for survival. RASless cells showed defective respiratory/metabolic phenotypes reminiscent of those of SOS1-deficient MEFs, suggesting that the mitochondrial defects of these cells are mechanistically linked to the absence of SOS1-GEF activity on cellular RAS targets. Our observations provide a direct mechanistic link between SOS1 and control of cellular oxidative stress and suggest that SOS1-mediated RAS activation is required for correct mitochondrial dynamics and function.
  10. mSphere. 2021 Jun 16. e0032721
      Mitochondrial cristae are polymorphic invaginations of the inner membrane that are the fabric of cellular respiration. Both the mitochondrial contact site and cristae organization system (MICOS) and the F1FO-ATP synthase are vital for sculpting cristae by opposing membrane-bending forces. While MICOS promotes negative curvature at crista junctions, dimeric F1FO-ATP synthase is crucial for positive curvature at crista rims. Crosstalk between these two complexes has been observed in baker's yeast, the model organism of the Opisthokonta supergroup. Here, we report that this property is conserved in Trypanosoma brucei, a member of the Discoba clade that separated from the Opisthokonta ∼2 billion years ago. Specifically, one of the paralogs of the core MICOS subunit Mic10 interacts with dimeric F1FO-ATP synthase, whereas the other core Mic60 subunit has a counteractive effect on F1FO-ATP synthase oligomerization. This is evocative of the nature of MICOS-F1FO-ATP synthase crosstalk in yeast, which is remarkable given the diversification that these two complexes have undergone during almost 2 eons of independent evolution. Furthermore, we identified a highly diverged, putative homolog of subunit e, which is essential for the stability of F1FO-ATP synthase dimers in yeast. Just like subunit e, it is preferentially associated with dimers and interacts with Mic10, and its silencing results in severe defects to cristae and the disintegration of F1FO-ATP synthase dimers. Our findings indicate that crosstalk between MICOS and dimeric F1FO-ATP synthase is a fundamental property impacting crista shape throughout eukaryotes. IMPORTANCE Mitochondria have undergone profound diversification in separate lineages that have radiated since the last common ancestor of eukaryotes some eons ago. Most eukaryotes are unicellular protists, including etiological agents of infectious diseases, like Trypanosoma brucei. Thus, the study of a broad range of protists can reveal fundamental features shared by all eukaryotes and lineage-specific innovations. Here, we report that two different protein complexes, MICOS and F1FO-ATP synthase, known to affect mitochondrial architecture, undergo crosstalk in T. brucei, just as in baker's yeast. This is remarkable considering that these complexes have otherwise undergone many changes during their almost 2 billion years of independent evolution. Thus, this crosstalk is a fundamental property needed to maintain proper mitochondrial structure even if the constituent players considerably diverged.
    Keywords:  ATP synthase; MICOS; Trypanosoma; evolution; mitochondria
  11. Elife. 2021 Jun 16. pii: e63104. [Epub ahead of print]10
      Typified by oxidative phosphorylation (OXPHOS), mitochondria catalyze a wide variety of cellular processes seemingly critical for malignant growth. As such, there is considerable interest in targeting mitochondrial metabolism in cancer. However, notwithstanding the few drugs targeting mutant dehydrogenase activity, nearly all hopeful 'mito-therapeutics' cannot discriminate cancerous from non-cancerous OXPHOS and thus suffer from a limited therapeutic index. The present project was based on the premise that the development of efficacious mitochondrial-targeted anti-cancer compounds requires answering two fundamental questions: 1) is mitochondrial bioenergetics in fact different between cancer and non-cancer cells? and 2) If so, what are the underlying mechanisms? Such information is particularly critical for the subset of human cancers, including acute myeloid leukemia (AML), in which alterations in mitochondrial metabolism are implicated in various aspects of cancer biology (e.g., clonal expansion and chemoresistance). Herein, we leveraged an in-house diagnostic biochemical workflow to comprehensively evaluate mitochondrial bioenergetic efficiency and capacity in various hematological cell types, with a specific focus on OXPHOS dynamics in AML. Consistent with prior reports, clonal cell expansion, characteristic of leukemia, was universally associated with a hyper-metabolic phenotype which included increases in basal and maximal glycolytic and respiratory flux. However, despite having nearly 2-fold more mitochondria per cell, clonally expanding hematopoietic stem cells, leukemic blasts, as well as chemoresistant AML were all consistently hallmarked by intrinsic limitations in oxidative ATP synthesis (i.e., OXPHOS). Remarkably, by performing experiments across a physiological span of ATP free energy (i.e, ΔGATP), we provide direct evidence that, rather than contributing to cellular ΔGATP, leukemic mitochondria are particularly poised to consume ATP. Relevant to AML biology, acute restoration of OXPHOS kinetics proved highly cytotoxic to leukemic blasts, suggesting that active OXPHOS repression supports aggressive disease dissemination in AML. Taken together, these findings argue against ATP being the primary output of mitochondria in leukemia and provide proof-of-principle that restoring, rather than disrupting, OXPHOS and/or cellular ΔGATP in cancer may represent an untapped therapeutic avenue for combatting hematological malignancy and chemoresistance.
    Keywords:  biochemistry; cancer biology; chemical biology; human
  12. FASEB J. 2021 Jul;35(7): e21678
      Hypertension is associated with excessive reactive oxygen species (ROS) production in vascular cells. Mitochondria undergo fusion and fission, a process playing a role in mitochondrial function. OPA1 is essential for mitochondrial fusion. Loss of OPA1 is associated with ROS production and cell dysfunction. We hypothesized that mitochondria fusion could reduce oxidative stress that defect in fusion would exacerbate hypertension. Using (a) Opa1 haploinsufficiency in isolated resistance arteries from Opa1+/- mice, (b) primary vascular cells from Opa1+/- mice, and (c) RNA interference experiments with siRNA against Opa1 in vascular cells, we investigated the role of mitochondria fusion in hypertension. In hypertension, Opa1 haploinsufficiency induced altered mitochondrial cristae structure both in vascular smooth muscle and endothelial cells but did not modify protein level of long and short forms of OPA1. In addition, we demonstrated an increase of mitochondrial ROS production, associated with a decrease of superoxide dismutase 1 protein expression. We also observed an increase of apoptosis in vascular cells and a decreased VSMCs proliferation. Blood pressure, vascular contractility, as well as endothelium-dependent and -independent relaxation were similar in Opa1+/- , WT, L-NAME-treated Opa1+/- and WT mice. Nevertheless, chronic NO-synthase inhibition with L-NAME induced a greater hypertension in Opa1+/- than in WT mice without compensatory arterial wall hypertrophy. This was associated with a stronger reduction in endothelium-dependent relaxation due to excessive ROS production. Our results highlight the protective role of mitochondria fusion in the vasculature during hypertension by limiting mitochondria ROS production.
    Keywords:  Opa1; hypertension; mitochondria; oxidative stress; vascular function
  13. FASEB J. 2021 Jul;35(7): e21688
      The mitochondria-associated membrane (MAM) is a functional subdomain of the endoplasmic reticulum membrane that tethers to the mitochondrial outer membrane and is essential for cellular homeostasis. A defect in MAM is involved in various neurological diseases, including amyotrophic lateral sclerosis (ALS). Recently, we and others reported that MAM was disrupted in the models expressing several ALS-linked genes, including SOD1, SIGMAR1, VAPB, TARDBP, and FUS, suggesting that MAM disruption is deeply involved in the pathomechanism of ALS. However, it is still uncertain whether MAM disruption is a common pathology in ALS, mainly due to the absence of a simple, quantitative tool for monitoring the status of MAM. In this study, to examine the effects of various ALS-causative genes on MAM, we created the following two novel MAM reporters: MAMtracker-Luc and MAMtracker-Green. The MAMtrackers could detect MAM disruption caused by suppression of SIGMAR1 or the overexpression of ALS-linked mutant SOD1 in living cells. Moreover, the MAMtrackers have an advantage in their ability to monitor reversible changes in the MAM status induced by nutritional conditions. We used the MAMtrackers with an expression plasmid library of ALS-causative genes and noted that 76% (16/21) of the genes altered MAM integrity. Our results suggest that MAM disruption is a common pathological feature in ALS. Furthermore, we anticipate our MAMtrackers, which are suitable for high-throughput assays, to be valuable tools to understand MAM dynamics.
    Keywords:  ER-mitochondria contact; amyotrophic lateral sclerosis (ALS); mitochondria-associated membrane (MAM); organelle; sigma-1 receptor (Sig1R)
  14. iScience. 2021 Jun 25. 24(6): 102498
      Mitochondria regulate the immune response after dengue virus (DENV) infection. Microarray analysis of genes identified the upregulation of mitochondrial cytidine/uridine monophosphate kinase 2 (CMPK2) by DENV infection. We used small interfering RNA-mediated knockdown (KD) and CRISPR-Cas9 knockout (KO) approaches, to investigate the role of CMPK2 in mouse and human cells. The results showed that CMPK2 was critical in DENV-induced antiviral cytokine release and mitochondrial oxidative stress and mitochondrial DNA release to the cytosol. The DENV-induced activation of Toll-like receptor (TLR)-9, inflammasome pathway, and cell migration was suppressed by CMPK2 depletion; however, viral production increased under CMPK2 deficiency. Examining mouse bone marrow-derived dendritic cells from interferon-alpha (IFN-α) receptor-KO mice and signal transducer and activator of transcription 1 (STAT1)-KO mice, we confirmed that CMPK2-mediated antiviral activity occurred in IFN-dependent and IFN-independent manners. In sum, CMPK2 is a critical factor in DENV-induced immune responses to determine innate immunity.
    Keywords:  Immunology; Molecular biology; Virology