bims-mitdis Biomed News
on Mitochondrial disorders
Issue of 2022–10–23
fifty-five papers selected by
Catalina Vasilescu, University of Helsinki
  1. Whole-genome sequencing for mitochondrial disorders identifies unexpected mimics.
  2. Gene therapy for primary mitochondrial diseases: experimental advances and clinical challenges.
  3. The metabolite-controlled ubiquitin conjugase Ubc8 promotes mitochondrial protein import.
  4. Parkin drives pS65-Ub turnover independently of canonical autophagy in Drosophila.
  5. Lipids reroute mitochondria.
  6. Pearson syndrome: a multisystem mitochondrial disease with bone marrow failure.
  7. Parkin coordinates mitochondrial lipid remodeling to execute mitophagy.
  8. Mitochondrial DNA maintenance in Drosophila melanogaster.
  9. TRPV4 acts as a mitochondrial Ca2+-importer and regulates mitochondrial temperature and metabolism.
  10. Silencing of the mitochondrial ribosomal protein L-24 gene activates the oxidative stress response in Caenorhabditis elegans.
  11. Ophthalmic manifestations of MEPAN syndrome.
  12. Decreased anterograde transport coupled with sustained retrograde transport contributes to reduced axonal mitochondrial density in tauopathy neurons.
  13. Prohibitin 1 regulates mtDNA release and downstream inflammatory responses.
  14. Clinical Experience of Neurological Mitochondrial Diseases in Children and Adults: A Single-Center Study.
  15. MTCH2 is a mitochondrial outer membrane protein insertase.
  16. Relief of CoA sequestration and restoration of mitochondrial function in a mouse model of propionic acidemia.
  17. Beyond ATP, new roles of mitochondria.
  18. Tissue-specific mitochondrial HIGD1C promotes oxygen sensitivity in carotid body chemoreceptors.
  19. Mitochondrial stress-induced GFRAL signaling controls diurnal food intake and anxiety-like behavior.
  20. Mammalian oocytes store mRNAs in a mitochondria-associated membraneless compartment.
  21. Structure of a mitochondrial ribosome with fragmented rRNA in complex with membrane-targeting elements.
  22. Splicing variants in NARS2 are associated with milder phenotypes and intra-familial variability.
  23. Single protonation of the reduced quinone in respiratory complex I drives four-proton pumping.
  24. Nitric oxide-driven modifications of lipoic arm inhibit α-ketoacid dehydrogenases.
  25. HIRA loss transforms FH-deficient cells.
  26. Mitochondrial aggregation caused by cytochalasin B compromises the efficiency and safety of three-parent embryo.
  27. Double administration of self-complementary AAV9NDUFS4 prevents Leigh disease in Ndufs4-/- mice.
  28. Biallelic variants in coenzyme Q10 biosynthesis pathway genes cause a retinitis pigmentosa phenotype.
  29. Assessment of Mitochondrial Dysfunctions After Sirtuin Inhibition.
  30. Taking mitochondria to heart.
  31. Mitochondrial DNA population variation is not associated with Alzheimer's in the Japanese population: A consistent finding across global populations.
  32. COX7A2L genetic variants determine cardiorespiratory fitness in mice and human.
  33. Inducible deletion of Raptor and mTOR from adult skeletal muscle impairs muscle contractility and relaxation.
  34. Purine nucleoside phosphorylase controls nicotinamide riboside metabolism in mammalian cells.
  35. A simple screening method for heterozygous female patients with ornithine transcarbamylase deficiency.
  36. A protein that mobilizes the cofactor molecule haem for use in cells.
  37. Deep Learning for Rare Disease: A Scoping Review.
  38. Recent advances in high-throughput single-cell transcriptomics and spatial transcriptomics.
  39. Social isolation induces succinate dehydrogenase dysfunction in anxious mice.
  40. Gut reaction: it's not all about enzymes.
  41. Genetic Basis of Childhood Cardiomyopathy.
  42. The gut drives fat cravings.
  43. Gene-specific machine learning model to predict the pathogenicity of BRCA2 variants.
  44. Didemnin B and ternatin-4 differentially inhibit conformational changes in eEF1A required for aminoacyl-tRNA accommodation into mammalian ribosomes.
  45. Baicalein modulates mitochondrial function by upregulating mitochondrial uncoupling protein-1 (UCP1) expression in brown adipocytes, cytotoxicity, and computational studies.
  46. Skeletal muscle mitochondrial remodeling in heart failure: An update on mechanisms and therapeutic opportunities.
  47. Targeted profiling of human extrachromosomal DNA by CRISPR-CATCH.
  48. Circulating mitochondrial DNA is associated with anemia in newly diagnosed hematologic malignancies.
  49. A performance evaluation study: Variant annotation tools - the enigma of clinical next generation sequencing (NGS) based genetic testing.
  50. Mitochondrial biogenesis, telomere length and cellular senescence in Parkinson's disease and Lewy body dementia.
  51. HiFENS: high-throughput FISH detection of endogenous pre-mRNA splicing isoforms.
  52. Recognition of cyclic dinucleotides and folates by human SLC19A1.
  53. Designing clinical trials for rare diseases: unique challenges and opportunities.
  54. Semi-automated assembly of high-quality diploid human reference genomes.
  55. The beauty of simplicity in membrane biology.
  1. Pract Neurol. 2022 Oct 17. pii: pn-2022-003570. [Epub ahead of print]
    Whole-genome sequencing for mitochondrial disorders identifies unexpected mimics.
    Katherine R Schon, Patrick F Chinnery.
      
    Keywords:  genetics; mitochondrial disorders; neurogenetics
    DOI:  https://doi.org/10.1136/pn-2022-003570
  2. Nat Rev Neurol. 2022 Oct 18.
    Gene therapy for primary mitochondrial diseases: experimental advances and clinical challenges.
    Micol Falabella, Michal Minczuk, Michael G Hanna, Carlo Viscomi, Robert D S Pitceathly.
      The variable clinical and biochemical manifestations of primary mitochondrial diseases (PMDs), and the complexity of mitochondrial genetics, have proven to be a substantial barrier to the development of effective disease-modifying therapies. Encouraging data from gene therapy trials in patients with Leber hereditary optic neuropathy and advances in DNA editing techniques have raised expectations that successful clinical transition of genetic therapies for PMDs is feasible. However, obstacles to the clinical application of genetic therapies in PMDs remain; the development of innovative, safe and effective genome editing technologies and vectors will be crucial to their future success and clinical approval. In this Perspective, we review progress towards the genetic treatment of nuclear and mitochondrial DNA-related PMDs. We discuss advances in mitochondrial DNA editing technologies alongside the unique challenges to targeting mitochondrial genomes. Last, we consider ongoing trials and regulatory requirements.
    DOI:  https://doi.org/10.1038/s41582-022-00715-9
  3. Life Sci Alliance. 2023 Jan;pii: e202201526. [Epub ahead of print]6(1):
    The metabolite-controlled ubiquitin conjugase Ubc8 promotes mitochondrial protein import.
    Saskia Rödl, Fabian den Brave, Markus Räschle, Büsra Kizmaz, Svenja Lenhard, Carina Groh, Hanna Becker, Jannik Zimmermann, Bruce Morgan, Elke Richling, Thomas Becker, Johannes M Herrmann.
      Mitochondria play a key role in cellular energy metabolism. Transitions between glycolytic and respiratory conditions induce considerable adaptations of the cellular proteome. These metabolism-dependent changes are particularly pronounced for the protein composition of mitochondria. Here, we show that the yeast cytosolic ubiquitin conjugase Ubc8 plays a crucial role in the remodeling process when cells transition from respiratory to fermentative conditions. Ubc8 is a conserved and well-studied component of the catabolite control system that is known to regulate the stability of gluconeogenic enzymes. Unexpectedly, we found that Ubc8 also promotes the assembly of the translocase of the outer membrane of mitochondria (TOM) and increases the levels of its cytosol-exposed receptor subunit Tom22. Ubc8 deficiency results in compromised protein import into mitochondria and reduced steady-state levels of mitochondrial proteins. Our observations show that Ubc8, which is controlled by the prevailing metabolic conditions, promotes the switch from glucose synthesis to glucose usage in the cytosol and induces the biogenesis of the mitochondrial TOM machinery to improve mitochondrial protein import during phases of metabolic transition.
    DOI:  https://doi.org/10.26508/lsa.202201526
  4. EMBO Rep. 2022 Oct 17. e202153552
    Parkin drives pS65-Ub turnover independently of canonical autophagy in Drosophila.
    Joanne L Usher, Alvaro Sanchez-Martinez, Ana Terriente-Felix, Po-Lin Chen, Juliette J Lee, Chun-Hong Chen, Alexander J Whitworth.
      Parkinson's disease-related proteins, PINK1 and Parkin, act in a common pathway to maintain mitochondrial quality control. While the PINK1-Parkin pathway can promote autophagic mitochondrial turnover (mitophagy) following mitochondrial toxification in cell culture, alternative quality control pathways are suggested. To analyse the mechanisms by which the PINK1-Parkin pathway operates in vivo, we developed methods to detect Ser65-phosphorylated ubiquitin (pS65-Ub) in Drosophila. Exposure to the oxidant paraquat led to robust, Pink1-dependent pS65-Ub production, while pS65-Ub accumulates in unstimulated parkin-null flies, consistent with blocked degradation. Additionally, we show that pS65-Ub specifically accumulates on disrupted mitochondria in vivo. Depletion of the core autophagy proteins Atg1, Atg5 and Atg8a did not cause pS65-Ub accumulation to the same extent as loss of parkin, and overexpression of parkin promoted turnover of both basal and paraquat-induced pS65-Ub in an Atg5-null background. Thus, we have established that pS65-Ub immunodetection can be used to analyse Pink1-Parkin function in vivo as an alternative to reporter constructs. Moreover, our findings suggest that the Pink1-Parkin pathway can promote mitochondrial turnover independently of canonical autophagy in vivo.
    Keywords:   in vivo ; Parkinson's disease; mitochondria; mitophagy; phospho-ubiquitin
    DOI:  https://doi.org/10.15252/embr.202153552
  5. Nat Metab. 2022 Oct;4(10): 1218
    Lipids reroute mitochondria.
    Michael Attwaters.
      
    DOI:  https://doi.org/10.1038/s42255-022-00662-1
  6. Orphanet J Rare Dis. 2022 10 17. 17(1): 379
    Pearson syndrome: a multisystem mitochondrial disease with bone marrow failure.
    Ayami Yoshimi, Kaori Ishikawa, Charlotte Niemeyer, Sarah C Grünert.
      Pearson syndrome (PS) is a rare fatal mitochondrial disorder caused by single large-scale mitochondrial DNA deletions (SLSMDs). Most patients present with anemia in infancy. Bone marrow cytology with vacuolization in erythroid and myeloid precursors and ring-sideroblasts guides to the correct diagnosis, which is established by detection of SLSMDs. Non hematological symptoms suggesting a mitochondrial disease are often lacking at initial presentation, thus PS is an important differential diagnosis in isolated hypogenerative anemia in infancy. Spontaneous resolution of anemia occurs in two-third of patients at the age of 1-3 years, while multisystem non-hematological complications such as failure to thrive, muscle hypotonia, exocrine pancreas insufficiency, renal tubulopathy and cardiac dysfunction develop during the clinical course. Some patients with PS experience a phenotypical change to Kearns-Sayre syndrome. In the absence of curative therapy, the prognosis of patients with PS is dismal. Most patients die of acute lactic acidosis and multi-organ failure in early childhood. There is a great need for the development of novel therapies to alter the natural history of patients with PS.
    Keywords:  Mitochondrial DNA deletion; Natural history; Pearson syndrome
    DOI:  https://doi.org/10.1186/s13023-022-02538-9
  7. EMBO Rep. 2022 Oct 18. e55191
    Parkin coordinates mitochondrial lipid remodeling to execute mitophagy.
    Chao-Chieh Lin, Jin Yan, Meghan D Kapur, Kristi L Norris, Cheng-Wei Hsieh, De Huang, Nicolas Vitale, Kah-Leong Lim, Ziqiang Guan, Xiao-Fan Wang, Jen-Tsan Chi, Wei-Yuan Yang, Tso-Pang Yao.
      Autophagy has emerged as the prime machinery for implementing organelle quality control. In the context of mitophagy, the ubiquitin E3 ligase Parkin tags impaired mitochondria with ubiquitin to activate autophagic degradation. Although ubiquitination is essential for mitophagy, it is unclear how ubiquitinated mitochondria activate autophagosome assembly locally to ensure efficient destruction. Here, we report that Parkin activates lipid remodeling on mitochondria targeted for autophagic destruction. Mitochondrial Parkin induces the production of phosphatidic acid (PA) and its subsequent conversion to diacylglycerol (DAG) by recruiting phospholipase D2 and activating the PA phosphatase, Lipin-1. The production of DAG requires mitochondrial ubiquitination and ubiquitin-binding autophagy receptors, NDP52 and optineurin (OPTN). Autophagic receptors, via Golgi-derived vesicles, deliver an autophagic activator, EndoB1, to ubiquitinated mitochondria. Inhibition of Lipin-1, NDP52/OPTN, or EndoB1 results in a failure to produce mitochondrial DAG, autophagosomes, and mitochondrial clearance, while exogenous cell-permeable DAG can induce autophagosome production. Thus, mitochondrial DAG production acts downstream of Parkin to enable the local assembly of autophagosomes for the efficient disposal of ubiquitinated mitochondria.
    Keywords:  Lipin-1; PLD2; Parkin; diacylglycerol; mitophagy
    DOI:  https://doi.org/10.15252/embr.202255191
  8. Biosci Rep. 2022 Oct 18. pii: BSR20211693. [Epub ahead of print]
    Mitochondrial DNA maintenance in Drosophila melanogaster.
    Ana P C Rodrigues, Audrey C Novaes, Grzegorz L Ciesielski, Marcos T Oliveira.
      All 37 mitochondrial DNA (mtDNA)-encoded genes involved with oxidative phosphorylation and intramitochondrial protein synthesis, and several nuclear-encoded genes involved with mtDNA replication, transcription, repair and recombination are conserved between the fruit fly Drosophilamelanogaster and mammals. This, in addition to its easy genetic tractability, has made Drosophila a useful model for our understanding of animal mtDNA maintenance and human mtDNA diseases. However, there are key differences between the Drosophila and mammalian systems that feature the diversity of mtDNA maintenance processes inside animal cells. Here, we review what is known about mtDNA maintenance in Drosophila, highlighting areas for which more research is warranted and providing a perspective preliminary in silico and in vivo analyses of the tissue specificity of mtDNA maintenance processes in this model organism. Our results suggest new roles (or the lack thereof) for well-known maintenance proteins, such as the helicase Twinkle and the accessory subunit of DNA polymerase γ, and for other Drosophila gene products that may even aid in shedding light on mtDNA maintenance in other animals. We hope to provide the reader some interesting paths that can be taken to help our community show how Drosophila may impact future mtDNA maintenance research.
    Keywords:  DNA synthesis and repair; Drosophila melanogaster; mitochondria; nucleic acids
    DOI:  https://doi.org/10.1042/BSR20211693
  9. Mitochondrion. 2022 Oct 16. pii: S1567-7249(22)00085-X. [Epub ahead of print]
    TRPV4 acts as a mitochondrial Ca2+-importer and regulates mitochondrial temperature and metabolism.
    Tusar Kanta Acharya, Ashutosh Kumar, Rakesh Kumar Majhi, Shamit Kumar, Ranabir Chakraborty, Ankit Tiwari, Karl-Heinz Smalla, Xiao Liu, Young-Tae Chang, Eckart D Gundelfinger, Chandan Goswami.
      TRPV4 is associated with the development of neuropathic pain, sensory defects, muscular dystrophies, neurodegenerative disorders, Charcot Marie Tooth and skeletal dysplasia. In all these cases, mitochondrial abnormalities are prominent. Here, we demonstrate that TRPV4, localizes to a subpopulation of mitochondria in various cell lines. Improper expression and/or function of TRPV4 induces several mitochondrial abnormalities. TRPV4 is also involved in the regulation of mitochondrial numbers, Ca2+-levels and mitochondrial temperature. Accordingly, several naturally occurring TRPV4 mutations affect mitochondrial morphology and distribution. These findings may help in understanding the significance of mitochondria in TRPV4-mediated channelopathies possibly classifying them as mitochondrial diseases.
    Keywords:  Mitochondrial Ca(2+); Neuro-Muscular Dystrophies Mitochondrial diseases; TRP channels; mitochondrial dynamics; mitochondrial temperature
    DOI:  https://doi.org/10.1016/j.mito.2022.10.001
  10. Biochim Biophys Acta Gen Subj. 2022 Oct 17. pii: S0304-4165(22)00173-8. [Epub ahead of print] 130255
    Silencing of the mitochondrial ribosomal protein L-24 gene activates the oxidative stress response in Caenorhabditis elegans.
    Graziella Ficociello, Emily Schifano, Michela Di Nottia, Alessandra Torraco, Rosalba Carrozzo, Daniela Uccelletti, Arianna Montanari.
      The mitochondrial translation machinery allows the synthesis of the mitochondrial-encoded subunits of the electron transport chain. Defects in this process lead to mitochondrial physiology failure; in humans, they are associated with early-onset, extremely variable and often fatal disorder. The use of a simple model to study the mitoribosomal defects is mandatory to overcome the difficulty to analyze the impact of pathological mutations in humans. In this paper we study in nematode Caenorhabditis elegans the silencing effect of the mrpl-24 gene, coding for the mitochondrial ribosomal protein L-24 (MRPL-24). This is a structural protein of the large subunit 39S of the mitoribosome and its effective physiological function is not completely elucidated. We have evaluated the nematode's fitness fault and investigated the mitochondrial defects associated with MRPL-24 depletion. The oxidative stress response activation due to the mitochondrial alteration has been also investigated as a compensatory physiological mechanism. For the first time, we demonstrated that MRPL-24 reduction increases the expression of detoxifying enzymes such as SOD-3 and GST-4 through the involvement of transcription factor SKN-1.
    BACKGROUND: In humans, mutations in genes encoding mitochondrial ribosomal proteins (MRPs) often cause early-onset, severe, fatal and extremely variable clinical defects. Mitochondrial ribosomal protein L-24 (MRPL24) is a structural protein of the large subunit 39S of the mitoribosome. It is highly conserved in different species and its effective physiological function is not completely elucidated.
    METHODS: We characterized the MRPL24 functionality using the animal model Caenorhabditis elegans. We performed the RNA mediated interference (RNAi) by exposing the nematodes' embryos to double-stranded RNA (dsRNA) specific for the MRPL-24 coding sequence. We investigated for the first time in C. elegans, the involvement of the MRPL-24 on the nematode's fitness and its mitochondrial physiology.
    RESULTS: Mrpl-24 silencing in C. elegans negatively affected the larval development, progeny production and body bending. The analysis of mitochondrial functionality revealed loss of mitochondrial network and impairment of mitochondrial functionality, as the decrease of oxygen consumption rate and the ROS production, as well as reduction of mitochondrial protein synthesis. Finally, the MRPL-24 depletion activated the oxidative stress response, increasing the expression levels of two detoxifying enzymes, SOD-3 and GST-4.
    CONCLUSIONS: In C. elegans the MRPL-24 depletion activated the oxidative stress response. This appears as a compensatory mechanism to the alteration of the mitochondrial functionality and requires the involvement of transcription factor SKN-1.
    GENERAL SIGNIFICANCE: C. elegans resulted in a good model for the study of mitochondrial disorders and its use as a simple and pluricellular organism could open interesting perspectives to better investigate the pathologic mechanisms underlying these devastating diseases.
    Keywords:  Caenorhabditis elegans; MRPL-24 mitoribosomal protein; Mitochondrial translation; Oxidative stress response; SKN-1 transcription factor
    DOI:  https://doi.org/10.1016/j.bbagen.2022.130255
  11. Ophthalmic Genet. 2022 Oct 19. 1-6
    Ophthalmic manifestations of MEPAN syndrome.
    Priya R Gupta, Sidney M Gospe.
       BACKGROUND: Mitochondrial enoyl CoA reductase protein-associated neurodegeneration (MEPAN) syndrome is an ultra-rare autosomal recessive disorder caused by loss-of-function mutations in the MECR gene. The syndrome is characterized by dystonia in early childhood, basal ganglia signal abnormalities on MRI, and subsequent optic atrophy, with relative sparing of cognition. We characterize the ophthalmic manifestations observed in a patient with MEPAN syndrome, as a detailed account of ocular findings has not been published to date.
    METHODS: Case study of a patient with genetically confirmed MEPAN syndrome, with full ophthalmic evaluation including slit-lamp exam, sensorimotor exam, fundus photography, retinal ocular coherence tomography (OCT), electroretinography, visual evoked potentials, and visual field testing.
    RESULTS: The patient exhibited decreased visual acuity of 20/150 in both eyes with moderate dyschromatopsia on pseudoisochromatic plate testing, while peripheral vision was largely intact on Goldmann visual field testing. Fundus exam revealed bilateral optic atrophy with pallor most pronounced temporally, corresponding to OCT findings of diffuse retinal nerve fiber layer thinning most prominent in the papillomacular bundle region and severe ganglion cell layer thinning in the maculae. She also displayed a high frequency horizontal end-gaze nystagmus and symmetric bilateral external ophthalmoplegia.
    CONCLUSIONS: The pattern of bilateral optic atrophy in our patient with MEPAN syndrome shows predilection for the papillomacular bundle, similar to that seen in other mitochondrial disorders with optic neuropathy, such as Leber Hereditary Optic Neuropathy and Dominant Optic Atrophy. Our patient's external ophthalmoplegia is another neuro-ophthalmic finding that may be seen in patients with heritable mitochondrial disease, either as an isolated ocular phenotype or within a constellation of systemic manifestations.
    Keywords:  MECR; MEPAN syndrome; mitochondrial fatty acid synthesis disorder; ophthalmoplegia; optic atrophy
    DOI:  https://doi.org/10.1080/13816810.2022.2135112
  12. Front Mol Neurosci. 2022 ;15 927195
    Decreased anterograde transport coupled with sustained retrograde transport contributes to reduced axonal mitochondrial density in tauopathy neurons.
    Anusruti Sabui, Mitali Biswas, Pramod Rajaram Somvanshi, Preethi Kandagiri, Madhavi Gorla, Fareed Mohammed, Prasad Tammineni.
      Mitochondria are essential organelle required for neuronal homeostasis. Mitochondria supply ATP and buffer calcium at synaptic terminals. However, the complex structural geometry of neurons poses a unique challenge in transporting mitochondria to synaptic terminals. Kinesin motors supply mitochondria to the axonal compartments, while cytoplasmic dynein is required for retrograde transport. Despite the importance of presynaptic mitochondria, how and whether axonal mitochondrial transport and distribution are altered in tauopathy neurons remain poorly studied. In the current study, we have shown that anterograde transport of mitochondria is reduced in P301L neurons, while there is no change in the retrograde transport. Consistently, axonal mitochondrial abundance is reduced in P301L neurons. We further studied the possible role of two opposing motor proteins on mitochondrial transport and found that mitochondrial association of kinesin is decreased significantly in P301L cells. Interestingly, fitting our experimental data into mathematical equations suggested a possible rise in dynein activity to maintain retrograde flux in P301L cells. Our data indicate that decreased kinesin-mediated transport coupled with sustained retrograde transport might reduce axonal mitochondria in tauopathy neurons, thus contributing to the synaptic deficits in Alzheimer's disease (AD) and other tauopathies.
    Keywords:  Alzheimer’s disease (AD); P301L; axonal transport; dynein; kinesin; mitochondria; tau
    DOI:  https://doi.org/10.3389/fnmol.2022.927195
  13. EMBO J. 2022 Oct 17. e111173
    Prohibitin 1 regulates mtDNA release and downstream inflammatory responses.
    Hao Liu, Hualin Fan, Pengcheng He, Haixia Zhuang, Xiao Liu, Meiting Chen, Wenwei Zhong, Yi Zhang, Cien Zhen, Yanling Li, Huilin Jiang, Tian Meng, Yiming Xu, Guojun Zhao, Du Feng.
      Exposure of mitochondrial DNA (mtDNA) to the cytosol activates innate immune responses. But the mechanisms by which mtDNA crosses the inner mitochondrial membrane are unknown. Here, we found that the inner mitochondrial membrane protein prohibitin 1 (PHB1) plays a critical role in mtDNA release by regulating permeability across the mitochondrial inner membrane. Loss of PHB1 results in alterations in mitochondrial integrity and function. PHB1-deficient macrophages, serum from myeloid-specific PHB1 KO (Phb1MyeKO) mice, and peripheral blood mononuclear cells from neonatal sepsis patients show increased interleukin-1β (IL-1β) levels. PHB1 KO mice are also intolerant of lipopolysaccharide shock. Phb1-depleted macrophages show increased cytoplasmic release of mtDNA and inflammatory responses. This process is suppressed by cyclosporine A and VBIT-4, which inhibit the mitochondrial permeability transition pore (mPTP) and VDAC oligomerization. Inflammatory stresses downregulate PHB1 expression levels in macrophages. Under normal physiological conditions, the inner mitochondrial membrane proteins, AFG3L2 and SPG7, are tethered to PHB1 to inhibit mPTP opening. Downregulation of PHB1 results in enhanced interaction between AFG3L2 and SPG7, mPTP opening, mtDNA release, and downstream inflammatory responses.
    Keywords:  AFG3L2; MIMP; PHB; SPG7; mtDNA
    DOI:  https://doi.org/10.15252/embj.2022111173
  14. Balkan J Med Genet. 2021 Nov;24(2): 5-14
    Clinical Experience of Neurological Mitochondrial Diseases in Children and Adults: A Single-Center Study.
    M Rogac, D Neubauer, L Leonardis, N Pecaric, M Meznaric, A Maver, W Sperl, B M Garavaglia, E Lamantea, B Peterlin.
      The goal of the study was to retrospectively evaluate a cohort of children and adults with mitochondrial diseases (MDs) in a single-center experience. Neurological clinical examination, brain magnetic resonance imaging (MRI) and spectroscopy, muscle biopsy, metabolic and molecular-genetic analysis were evaluated in 26 children and 36 adult patients with MD in Slovenia from 2004 to 2018. Nijmegen MD criteria (MDC) were applied to all patients and the need for a muscle biopsy was estimated. Exome-sequencing was used in half of the patients. Twenty children (77.0%) and 12 adults (35.0%) scored a total of ≥8 on MDC, a result that is compatible with the diagnosis of definite MD. Yield of exome-sequencing was 7/22 (31.0%), but the method was not applied systematically in all patients from the beginning of diagnostics. Brain MRI morphological changes, which can be an imaging clue for the diagnosis of MD, were found in 17/24 children (71.0%). In 7/26 (29.0%) children, and in 20/30 (67.0%) adults, abnormal mitochondria were found on electron microscopy (EM) and ragged-red fibers were found in 16/30 (53.0%) adults. Respiratory chain enzymes (RCEs) and/or pyruvate dehydrogenase complex (PDHc) activities were abnormal in all the children and six adult cases. First, our data revealed that MDC was useful in the clinical diagnosis of MD, and second, until the use of NGS methods, extensive, laborious and invasive diagnostic procedures were performed to reach a final diagnosis. In patients with suspected MD, there is a need to prioritize molecular diagnosis with the more modern next-generation sequencing (NGS) method.
    Keywords:  Exome-sequencing; Magnetic resonance imaging (MRI); Mitochondrial disease (MD); Muscle biopsy; Nijmegen mitochondrial disease criteria (MDC)
    DOI:  https://doi.org/10.2478/bjmg-2021-0019
  15. Science. 2022 Oct 21. 378(6617): 317-322
    MTCH2 is a mitochondrial outer membrane protein insertase.
    Alina Guna, Taylor A Stevens, Alison J Inglis, Joseph M Replogle, Theodore K Esantsi, Gayathri Muthukumar, Kelly C L Shaffer, Maxine L Wang, Angela N Pogson, Jeff J Jones, Brett Lomenick, Tsui-Fen Chou, Jonathan S Weissman, Rebecca M Voorhees.
      In the mitochondrial outer membrane, α-helical transmembrane proteins play critical roles in cytoplasmic-mitochondrial communication. Using genome-wide CRISPR screens, we identified mitochondrial carrier homolog 2 (MTCH2), and its paralog MTCH1, and showed that it is required for insertion of biophysically diverse tail-anchored (TA), signal-anchored, and multipass proteins, but not outer membrane β-barrel proteins. Purified MTCH2 was sufficient to mediate insertion into reconstituted proteoliposomes. Functional and mutational studies suggested that MTCH2 has evolved from a solute carrier transporter. MTCH2 uses membrane-embedded hydrophilic residues to function as a gatekeeper for the outer membrane, controlling mislocalization of TAs into the endoplasmic reticulum and modulating the sensitivity of leukemia cells to apoptosis. Our identification of MTCH2 as an insertase provides a mechanistic explanation for the diverse phenotypes and disease states associated with MTCH2 dysfunction.
    DOI:  https://doi.org/10.1126/science.add1856
  16. J Inherit Metab Dis. 2022 Oct 17.
    Relief of CoA sequestration and restoration of mitochondrial function in a mouse model of propionic acidemia.
    Chitra Subramanian, Matthew W Frank, Rajendra Tangallapally, Mi-Kyung Yun, Stephen W White, Richard E Lee, Charles O Rock, Suzanne Jackowski.
      Propionic acidemia (PA, OMIM 606054) is a devastating inborn error of metabolism arising from mutations that reduce the activity of the mitochondrial enzyme propionyl-CoA carboxylase (PCC). The defects in PCC reduce the concentrations of nonesterified coenzyme A (CoASH), thus compromising mitochondrial function and disrupting intermediary metabolism. Here, we use a hypomorphic PA mouse model to test the effectiveness of BBP-671 in correcting the metabolic imbalances in PA. BBP-671 is a high-affinity allosteric pantothenate kinase activator that counteracts feedback inhibition of the enzyme to increase the intracellular concentration of CoA. Liver CoASH and acetyl-CoA are depressed in PA mice and BBP-671 treatment normalizes the cellular concentrations of these two key cofactors. Hepatic propionyl-CoA is also reduced by BBP-671 leading to an improved intracellular C3:C2-CoA ratio. Elevated plasma C3:C2-carnitine ratio and methylcitrate, hallmark biomarkers of PA, are significantly reduced by BBP-671. The large elevations of malate and α-ketoglutarate in the urine of PA mice are biomarkers for compromised tricarboxylic acid cycle activity and BBP-671 therapy reduces the amounts of both metabolites. Furthermore, the low survival of PA mice is restored to normal by BBP-671. These data show that BBP-671 relieves CoA sequestration, improves mitochondrial function, reduces plasma PA biomarkers and extends the lifespan of PA mice, providing the preclinical foundation for the therapeutic potential of BBP-671. This article is protected by copyright. All rights reserved.
    DOI:  https://doi.org/10.1002/jimd.12570
  17. Biochem (Lond). 2022 Aug;44(4): 2-8
    Beyond ATP, new roles of mitochondria.
    Ram Prosad Chakrabarty, Navdeep S Chandel.
      Mitochondria, special double-membraned intracellular compartments or 'organelles', are popularly known as the 'powerhouses of the cell', as they generate the bulk of ATP used to fuel cellular biochemical reactions. Mitochondria are also well known for generating metabolites for the synthesis of macromolecules (e.g., carbohydrates, proteins, lipids and nucleic acids). In the mid-1990s, new evidence suggesting that mitochondria, beyond their canonical roles in bioenergetics and biosynthesis, can act as signalling organelles began to emerge, bringing a dramatic shift in our view of mitochondria's roles in controlling cell function. Over the next two and half decades, works from multiple groups have demonstrated how mitochondrial signalling can dictate diverse physiological and pathophysiological outcomes. In this article, we will briefly discuss different mechanisms by which mitochondria can communicate with cytosol and other organelles to regulate cell fate and function and exert paracrine effects. Our molecular understanding of mitochondrial communication with the rest of the cell, i.e. mitochondrial signalling, could reveal new therapeutic strategies to improve health and ameliorate diseases.
    DOI:  https://doi.org/10.1042/bio_2022_119
  18. Elife. 2022 Oct 18. pii: e78915. [Epub ahead of print]11
    Tissue-specific mitochondrial HIGD1C promotes oxygen sensitivity in carotid body chemoreceptors.
    Alba Timón-Gómez, Alexandra L Scharr, Nicholas Y Wong, Erwin Ni, Arijit Roy, Min Liu, Julisia Chau, Jack L Lampert, Homza Hireed, Noah S Kim, Masood Jan, Alexander R Gupta, Ryan W Day, James M Gardner, Richard J A Wilson, Antoni Barrientos, Andy J Chang.
      Mammalian carotid body arterial chemoreceptors function as an early warning system for hypoxia, triggering acute life-saving arousal and cardiorespiratory reflexes. To serve this role, carotid body glomus cells are highly sensitive to decreases in oxygen availability. While the mitochondria and plasma membrane signaling proteins have been implicated in oxygen sensing by glomus cells, the mechanism underlying their mitochondrial sensitivity to hypoxia compared to other cells is unknown. Here, we identify HIGD1C, a novel hypoxia-inducible gene domain factor isoform, as an electron transport chain Complex IV-interacting protein that is almost exclusively expressed in the carotid body and is therefore not generally necessary for mitochondrial function. Importantly, HIGD1C is required for carotid body oxygen sensing and enhances Complex IV sensitivity to hypoxia. Thus, we propose that HIGD1C promotes exquisite oxygen sensing by the carotid body, illustrating how specialized mitochondria can be used as sentinels of metabolic stress to elicit essential adaptive behaviors.
    Keywords:  biochemistry; chemical biology; human; mouse; neuroscience; rat
    DOI:  https://doi.org/10.7554/eLife.78915
  19. Life Sci Alliance. 2022 Nov;pii: e202201495. [Epub ahead of print]5(11):
    Mitochondrial stress-induced GFRAL signaling controls diurnal food intake and anxiety-like behavior.
    Carla Igual Gil, Bethany M Coull, Wenke Jonas, Rachel N Lippert, Susanne Klaus, Mario Ost.
      Growth differentiation factor 15 (GDF15) is a mitochondrial stress-induced cytokine that modulates energy balance in an endocrine manner. However, the importance of its brainstem-restricted receptor GDNF family receptor alpha-like (GFRAL) to mediate endocrine GDF15 signaling to the brain upon mitochondrial dysfunction is still unknown. Using a mouse model with muscle-specific mitochondrial dysfunction, we here show that GFRAL is required for activation of systemic energy metabolism via daytime-restricted anorexia but not responsible for muscle wasting. We further find that muscle mitochondrial stress response involves a GFRAL-dependent induction of hypothalamic corticotropin-releasing hormone, without elevated corticosterone levels. Finally, we identify that GFRAL signaling governs an anxiety-like behavior in male mice with muscle mitochondrial dysfunction, with females showing a less robust GFRAL-dependent anxiety-like phenotype. Together, we here provide novel evidence of a mitochondrial stress-induced muscle-brain crosstalk via the GDF15-GFRAL axis to modulate food intake and anxiogenic behavior.
    DOI:  https://doi.org/10.26508/lsa.202201495
  20. Science. 2022 Oct 21. 378(6617): eabq4835
    Mammalian oocytes store mRNAs in a mitochondria-associated membraneless compartment.
    Shiya Cheng, Gerrit Altmeppen, Chun So, Luisa M Welp, Sarah Penir, Torben Ruhwedel, Katerina Menelaou, Katarina Harasimov, Alexandra Stützer, Martyn Blayney, Kay Elder, Wiebke Möbius, Henning Urlaub, Melina Schuh.
      Full-grown oocytes are transcriptionally silent and must stably maintain the messenger RNAs (mRNAs) needed for oocyte meiotic maturation and early embryonic development. However, where and how mammalian oocytes store maternal mRNAs is unclear. Here, we report that mammalian oocytes accumulate mRNAs in a mitochondria-associated ribonucleoprotein domain (MARDO). MARDO assembly around mitochondria was promoted by the RNA-binding protein ZAR1 and directed by an increase in mitochondrial membrane potential during oocyte growth. MARDO foci coalesced into hydrogel-like matrices that clustered mitochondria. Maternal mRNAs stored in the MARDO were translationally repressed. Loss of ZAR1 disrupted the MARDO, dispersed mitochondria, and caused a premature loss of MARDO-localized mRNAs. Thus, a mitochondria-associated membraneless compartment controls mitochondrial distribution and regulates maternal mRNA storage, translation, and decay to ensure fertility in mammals.
    DOI:  https://doi.org/10.1126/science.abq4835
  21. Nat Commun. 2022 Oct 17. 13(1): 6132
    Structure of a mitochondrial ribosome with fragmented rRNA in complex with membrane-targeting elements.
    Victor Tobiasson, Ieva Berzina, Alexey Amunts.
      Mitoribosomes of green algae display a great structural divergence from their tracheophyte relatives, with fragmentation of both rRNA and proteins as a defining feature. Here, we report a 2.9 Å resolution structure of the mitoribosome from the alga Polytomella magna harbouring a reduced rRNA split into 13 fragments. We found that the rRNA contains a non-canonical reduced form of the 5S, as well as a permutation of the LSU domain I. The mt-5S rRNA is stabilised by mL40 that is also found in mitoribosomes lacking the 5S, which suggests an evolutionary pathway. Through comparison to other ribosomes with fragmented rRNAs, we observe that the pattern is shared across large evolutionary distances, and between cellular compartments, indicating an evolutionary convergence and supporting the concept of a primordial fragmented ribosome. On the protein level, eleven peripherally associated HEAT-repeat proteins are involved in the binding of 3' rRNA termini, and the structure features a prominent pseudo-trimer of one of them (mL116). Finally, in the exit tunnel, mL128 constricts the tunnel width of the vestibular area, and mL105, a homolog of a membrane targeting component mediates contacts with an inner membrane bound insertase. Together, the structural analysis provides insight into the evolution of the ribosomal machinery in mitochondria.
    DOI:  https://doi.org/10.1038/s41467-022-33582-5
  22. Eur J Med Genet. 2022 Oct 14. pii: S1769-7212(22)00224-5. [Epub ahead of print] 104643
    Splicing variants in NARS2 are associated with milder phenotypes and intra-familial variability.
    Samira Ait-El-Mkadem Saadi, Elsa Kaphan, Amaya Morales Jaurrieta, Konstantina Fragaki, Annabelle Chaussenot, Sylvie Bannwarth, André Maues De Paula, Véronique Paquis-Flucklinger, Cécile Rouzier.
      Biallelic rare variants in NARS2 that encode the mitochondrial asparaginyl-tRNA synthetase are associated with a wide spectrum of clinical phenotypes ranging from severe neurodegenerative disorders to isolated mitochondrial myopathy or deafness. To date, only a small number of patients with NARS2 variants have been reported, and possible genotype-phenotype correlations are still lacking. Here, we present three siblings who had an early-onset hearing loss, while one developed severe symptoms in adulthood associated with early intellectual impairment, refractory seizures, moderate axonal sensorimotor neuropathy, and atypical psychiatric symptoms. Biochemical analysis revealed impairment of the activity and assembly of the respiratory chain complexes in this patient's muscle and fibroblasts. Whole Exome Sequencing allowed identification of a heterozygous variant NM_024678.5(NARS2):c.822G > C (p.Gln274His) that is known to be pathogenic and to affect splicing of the NARS2 gene, but was unable to detect a second variant in this gene. Coverage analysis and Sanger sequencing led to identification of a novel intronic deletion NM_024678.5(NARS2):c.922-21_922-19del in the three siblings in trans with the c.822G > C. Functional analysis by RT-PCR showed that this deletion was causing aberrant splicing and led to exon 9 skipping in NARS2 mRNA in patient fibroblasts. Our work expands the phenotype and genotype spectrum of NARS2-related disorders. We provide evidence of the pathogenic effect of a novel intronic deletion in the NARS2 gene and report on additional adult patients with a large intrafamilial variability associated with splice variants in this gene. More specifically, we detail the phenotype of the oldest living patient to date with NARS2 variants and, for the first time, we report the psychiatric symptoms associated with this gene. Our work confirms the complexity of genotype-phenotype correlation in patients with pathogenic NARS2 variants.
    Keywords:  Aberrant splicing; Aminoacyl-tRNA synthetase; Genotype-phenotype; NARS2; Splice variant
    DOI:  https://doi.org/10.1016/j.ejmg.2022.104643
  23. FEBS Lett. 2022 Oct 17.
    Single protonation of the reduced quinone in respiratory complex I drives four-proton pumping.
    Alexei A Stuchebrukhov, Tomoyuki Hayashi.
      Complex I is a key proton-pumping enzyme in bacterial and mitochondrial respiratory electron transport chains. Using quantum chemistry and electrostatic calculations, we have examined the pKa of the reduced quinone QH-/QH2 in the catalytic cavity of complex I. We find that pKa(QH-/QH2) is very high, above 20. This means that the energy of a single protonation reaction of the doubly reduced quinone (i.e. the reduced semiquinone QH-) is sufficient to drive four protons across the membrane with a potential of 180mV. Based on these calculations, we propose a possible scheme of redox-linked proton pumping by complex I. The model explains how the energy of the protonation reaction can be divided equally between four pumping units of the pump, and how a single proton can drive translocation of four additional protons in multiple pumping blocks.
    Keywords:  Mitochondria; NADH dehydrogenase; coupled electron-proton transfer; proton pumping; proton translocation; respiratory complex I
    DOI:  https://doi.org/10.1002/1873-3468.14518
  24. Nat Chem Biol. 2022 Oct 20.
    Nitric oxide-driven modifications of lipoic arm inhibit α-ketoacid dehydrogenases.
    Gretchen L Seim, Steven V John, Nicholas L Arp, Zixiang Fang, David J Pagliarini, Jing Fan.
      Pyruvate dehydrogenase complex (PDHC) and oxoglutarate dehydrogenase complex (OGDC), which belong to the mitochondrial α-ketoacid dehydrogenase family, play crucial roles in cellular metabolism. These multi-subunit enzyme complexes use lipoic arms covalently attached to their E2 subunits to transfer an acyl group to coenzyme A (CoA). Here, we report a novel mechanism capable of substantially inhibiting PDHC and OGDC: reactive nitrogen species (RNS) can covalently modify the thiols on their lipoic arms, generating a series of adducts that block catalytic activity. S-Nitroso-CoA, a product between RNS and the E2 subunit's natural substrate, CoA, can efficiently deliver these modifications onto the lipoic arm. We found RNS-mediated inhibition of PDHC and OGDC occurs during classical macrophage activation, driving significant rewiring of cellular metabolism over time. This work provides a new mechanistic link between RNS and mitochondrial metabolism with potential relevance for numerous physiological and pathological conditions in which RNS accumulate.
    DOI:  https://doi.org/10.1038/s41589-022-01153-w
  25. Sci Adv. 2022 Oct 21. 8(42): eabq8297
    HIRA loss transforms FH-deficient cells.
    Lorea Valcarcel-Jimenez, Connor Rogerson, Cissy Yong, Christina Schmidt, Ming Yang, Monica Cremades-Rodelgo, Victoria Harle, Victoria Offord, Kim Wong, Ariane Mora, Alyson Speed, Veronica Caraffini, Maxine Gia Binh Tran, Eamonn R Maher, Grant D Stewart, Sakari Vanharanta, David J Adams, Christian Frezza.
      Fumarate hydratase (FH) is a mitochondrial enzyme that catalyzes the reversible hydration of fumarate to malate in the tricarboxylic acid (TCA) cycle. Germline mutations of FH lead to hereditary leiomyomatosis and renal cell carcinoma (HLRCC), a cancer syndrome characterized by a highly aggressive form of renal cancer. Although HLRCC tumors metastasize rapidly, FH-deficient mice develop premalignant cysts in the kidneys, rather than carcinomas. How Fh1-deficient cells overcome these tumor-suppressive events during transformation is unknown. Here, we perform a genome-wide CRISPR-Cas9 screen to identify genes that, when ablated, enhance the proliferation of Fh1-deficient cells. We found that the depletion of the histone cell cycle regulator (HIRA) enhances proliferation and invasion of Fh1-deficient cells in vitro and in vivo. Mechanistically, Hira loss activates MYC and its target genes, increasing nucleotide metabolism specifically in Fh1-deficient cells, independent of its histone chaperone activity. These results are instrumental for understanding mechanisms of tumorigenesis in HLRCC and the development of targeted treatments for patients.
    DOI:  https://doi.org/10.1126/sciadv.abq8297
  26. Mol Hum Reprod. 2022 Oct 20. pii: gaac036. [Epub ahead of print]
    Mitochondrial aggregation caused by cytochalasin B compromises the efficiency and safety of three-parent embryo.
    Ying Li, Sanbao Shi, Jin Yuan, Xi Xiao, Dongmei Ji, Jianxin Pan, Zhunyuan Min, Hao Wang, Hongying Sha, Yazhong Ji.
      It is widely accepted that cytochalasin B (CB) is required in enucleation of the oocyte in order to stabilize the cytoplasm. However, CB treatment results in the uneven distribution of mitochondria, with aggregation towards the nucleus, which might compromise the efficiency and safety of a three-parent embryo. Here, we demonstrated that CB treatment affected mitochondrial dynamics, spindle morphology and mitochondrial DNA carryover in a concentration-dependent manner. Our results showed that mouse oocytes treated with over 1 μg/ml CB exhibited more aggregated pattern of mitochondria and diminished filamentous actin expression. Abnormal fission of mitochondria together with changes in spindle morphology increased as CB concentration escalated. Based on the results of mouse experiments, we further revealed the practical value of these findings in human oocytes. Chip-based digital PCR and pyrosequencing revealed that the mitochondrial carryover in reconstituted human embryos was significantly reduced by modifying the concentration of CB from the standard 5 μg/ml to 1 μg/ml before spindle transfer and pronuclear transfer. In conclusion, our findings provide an optimal manipulation for improving the efficiency and safety of mitochondrial replacement therapy.
    Keywords:  cytochalasin B; germ cells; mitochondrial replacement; mtDNA heteroplasmy; oocyte
    DOI:  https://doi.org/10.1093/molehr/gaac036
  27. Brain. 2022 Oct 21. 145(10): 3405-3414
    Double administration of self-complementary AAV9NDUFS4 prevents Leigh disease in Ndufs4-/- mice.
    Samantha Corrà, Raffaele Cerutti, Valeria Balmaceda, Carlo Viscomi, Massimo Zeviani.
      Leigh disease, or subacute necrotizing encephalomyelopathy, a genetically heterogeneous condition consistently characterized by defective mitochondrial bioenergetics, is the most common oxidative-phosphorylation related disease in infancy. Both neurological signs and pathological lesions of Leigh disease are mimicked by the ablation of the mouse mitochondrial respiratory chain subunit Ndufs4-/-, which is part of, and crucial for, normal Complex I activity and assembly, particularly in the brains of both children and mice. We previously conveyed the human NDUFS4 gene to the mouse brain using either single-stranded adeno-associated viral 9 recombinant vectors or the PHP.B adeno-associated viral vector. Both these approaches significantly prolonged the lifespan of the Ndufs4-/- mouse model but the extension of the survival was limited to a few weeks by the former approach, whereas the latter was applicable to a limited number of mouse strains, but not to primates. Here, we exploited the recent development of new, self-complementary adeno-associated viral 9 vectors, in which the transcription rate of the recombinant gene is markedly increased compared with the single-stranded adeno-associated viral 9 and can be applied to all mammals, including humans. Either single intra-vascular or double intra-vascular and intra-cerebro-ventricular injections were performed at post-natal Day 1. The first strategy ubiquitously conveyed the human NDUFS4 gene product in Ndufs4-/- mice, doubling the lifespan from 45 to ≈100 days after birth, when the mice developed rapidly progressive neurological failure. However, the double, contemporary intra-vascular and intra-cerebroventricular administration of self-complementary-adeno-associated viral NDUFS4 prolonged healthy lifespan up to 9 months of age. These mice were well and active at euthanization, at 6, 7, 8 and 9 months of age, to investigate the brain and other organs post-mortem. Robust expression of hNDUFS4 was detected in different cerebral areas preserving normal morphology and restoring Complex I activity and assembly. Our results warrant further investigation on the translatability of self-complementary-adeno-associated viral 9 NDUFS4-based therapy in the prodromal phase of the disease in mice and eventually humans.
    Keywords:   Ndufs4 ; Complex I; Leigh disease; gene therapy; mitochondrial disease
    DOI:  https://doi.org/10.1093/brain/awac182
  28. NPJ Genom Med. 2022 Oct 20. 7(1): 60
    Biallelic variants in coenzyme Q10 biosynthesis pathway genes cause a retinitis pigmentosa phenotype.
    Genomics England Research Consortium
      The aim of this study was to investigate coenzyme Q10 (CoQ10) biosynthesis pathway defects in inherited retinal dystrophy. Individuals affected by inherited retinal dystrophy (IRD) underwent exome or genome sequencing for molecular diagnosis of their condition. Following negative IRD gene panel analysis, patients carrying biallelic variants in CoQ10 biosynthesis pathway genes were identified. Clinical data were collected from the medical records. Haplotypes harbouring the same missense variant were characterised from family genome sequencing (GS) data and direct Sanger sequencing. Candidate splice variants were characterised using Oxford Nanopore Technologies single molecule sequencing. The CoQ10 status of the human plasma was determined in some of the study patients. 13 individuals from 12 unrelated families harboured candidate pathogenic genotypes in the genes: PDSS1, COQ2, COQ4 and COQ5. The PDSS1 variant c.589 A > G was identified in three affected individuals from three unrelated families on a possible ancestral haplotype. Three variants (PDSS1 c.468-25 A > G, PDSS1 c.722-2 A > G, COQ5 c.682-7 T > G) were shown to lead to cryptic splicing. 6 affected individuals were diagnosed with non-syndromic retinitis pigmentosa and 7 had additional clinical findings. This study provides evidence of CoQ10 biosynthesis pathway gene defects leading to non-syndromic retinitis pigmentosa in some cases. Intronic variants outside of the canonical splice-sites represent an important cause of disease. RT-PCR nanopore sequencing is effective in characterising these splice defects.
    DOI:  https://doi.org/10.1038/s41525-022-00330-z
  29. Methods Mol Biol. 2023 ;2589 269-291
    Assessment of Mitochondrial Dysfunctions After Sirtuin Inhibition.
    Christian Marx, Lisa Marx-Blümel, Jürgen Sonnemann, Zhao-Qi Wang.
      Posttranslational modifications are important for protein functions and cellular signaling pathways. The acetylation of lysine residues is catalyzed by histone acetyltransferases (HATs) and removed by histone deacetylases (HDACs), with the latter being grouped into four phylogenetic classes. The class III of the HDAC family, the sirtuins (SIRTs), contributes to gene expression, genomic stability, cell metabolism, and tumorigenesis. Thus, several specific SIRT inhibitors (SIRTi) have been developed to target cancer cell proliferation. Here we provide an overview of methods to study SIRT-dependent cell metabolism and mitochondrial functionality. The chapter describes metabolic flux analysis using Seahorse analyzers, methods for normalization of Seahorse data, flow cytometry and fluorescence microscopy to determine the mitochondrial membrane potential, mitochondrial content per cell and mitochondrial network structures, and Western blot analysis to measure mitochondrial proteins.
    Keywords:  Flow cytometry; Metabolism; Mitochondria; SIRT; Seahorse analysis; Sirtuin inhibition; Western blot
    DOI:  https://doi.org/10.1007/978-1-0716-2788-4_18
  30. Sci Signal. 2022 Oct 18. 15(756): eadf2995
    Taking mitochondria to heart.
    Wei Wong.
      Long-chain fatty acids redirect the uptake of mitochondria released from adipocytes from macrophages to the heart.
    DOI:  https://doi.org/10.1126/scisignal.adf2995
  31. PLoS One. 2022 ;17(10): e0276169
    Mitochondrial DNA population variation is not associated with Alzheimer's in the Japanese population: A consistent finding across global populations.
    Johanna Wong, Jannetta S Steyn, Ilse S Pienaar, Joanna L Elson.
      Several mitochondrial DNA (mtDNA) haplogroup association studies have suggested that common mtDNA variants are associated with multifactorial diseases, including Alzheimer's disease (AD). However, such studies have also produced conflicting results. A new mtDNA association model, the 'variant load model' (VLM), has been applied to multiple disease phenotypes. Application of the VLM in a 2017 study failed to find different variant loads in AD patients compared to controls, in two cohorts of European origin. The study also suggested a lower variant load in healthy elderly individuals, but could offer no replicate cohort to support this observation. Here, the VLM is applied to Japanese mtDNA sequences; in doing so, we explored the role of mtDNA variation in AD and ageing in a different global population. Consistent with the previous findings using the VLM in two populations of European origin, we found no evidence for an association between rarer, non-haplogroup associated variation and the development of AD. However, the result in the context of ageing that suggested those with fewer mildly deleterious mutations might undergo healthier ageing, was not replicated. In contrast to our previous study, our present results suggest that those living to advanced old age may harbour more mildly deleterious mtDNA variations. Importantly our analysis showed this finding is not primarily being driven by many rare population variants dispersed across the mtDNA, but by a few more frequent variants with high MutPred scores. It is suggested the variants in question do not exert a mildly deleterious effect in their most frequent haplogroup context.
    DOI:  https://doi.org/10.1371/journal.pone.0276169
  32. Nat Metab. 2022 Oct;4(10): 1336-1351
    COX7A2L genetic variants determine cardiorespiratory fitness in mice and human.
    Giorgia Benegiamo, Maroun Bou Sleiman, Martin Wohlwend, Sandra Rodríguez-López, Ludger J E Goeminne, Pirkka-Pekka Laurila, Marie Klevjer, Minna K Salonen, Jari Lahti, Pooja Jha, Sara Cogliati, José Antonio Enriquez, Ben M Brumpton, Anja Bye, Johan G Eriksson, Johan Auwerx.
      Mitochondrial respiratory complexes form superassembled structures called supercomplexes. COX7A2L is a supercomplex-specific assembly factor in mammals, although its implication for supercomplex formation and cellular metabolism remains controversial. Here we identify a role for COX7A2L for mitochondrial supercomplex formation in humans. By using human cis-expression quantitative trait loci data, we highlight genetic variants in the COX7A2L gene that affect its skeletal muscle expression specifically. The most significant cis-expression quantitative trait locus is a 10-bp insertion in the COX7A2L 3' untranslated region that increases messenger RNA stability and expression. Human myotubes harboring this insertion have more supercomplexes and increased respiration. Notably, increased COX7A2L expression in the muscle is associated with lower body fat and improved cardiorespiratory fitness in humans. Accordingly, specific reconstitution of Cox7a2l expression in C57BL/6J mice leads to higher maximal oxygen consumption, increased lean mass and increased energy expenditure. Furthermore, Cox7a2l expression in mice is induced specifically in the muscle upon exercise. These findings elucidate the genetic basis of mitochondrial supercomplex formation and function in humans and show that COX7A2L plays an important role in cardiorespiratory fitness, which could have broad therapeutic implications in reducing cardiovascular mortality.
    DOI:  https://doi.org/10.1038/s42255-022-00655-0
  33. J Physiol. 2022 Oct 18.
    Inducible deletion of Raptor and mTOR from adult skeletal muscle impairs muscle contractility and relaxation.
    Martina Baraldo, Sabrina Zorzato, Achille Homère Tchampda Dondjang, Alessia Geremia, Leonardo Nogara, Ana Georgia Dumitras, Marta Canato, Lorenzo Marcucci, Hendrik Nolte, Bert Blaauw.
       KEY POINTS: Skeletal muscle wasting and weakness have been associated with different pathological conditions, including sarcopenia and muscular dystrophy, and is accompanied by altered mTOR signaling Mammalian Target of Rapamycin (mTOR) plays a crucial role in the maintenance of muscle mass and functionality We found that the loss of both mTOR and Raptor results in contractile abnormalities, with severe muscle weakness and delayed relaxation following tetanic stimulation These results are associated with alterations in the expression of genes involved in sarcomere organization and calcium handling, and with an impairment in calcium reuptake after contraction Taken together, these results reveal a mechanistic insight into the role of mTOR in muscle contractility ABSTRACT: Skeletal muscle weakness has been associated with different pathological conditions, including sarcopenia and muscular dystrophy, and is accompanied by altered mTOR signaling. Here we wanted to better elucidate the functional role of mTOR on muscle contractility. Most loss of function studies for mTOR signaling have used the drug rapamycin to inhibit some of the signaling downstream of mTOR. However, as rapamycin does not completely inhibit all mTOR signaling, we generated a double k.o. for mTOR and for the scaffold protein of mTORC1, Raptor, in skeletal muscle. We found that dk.o. mice results in a more severe phenotype compared to Raptor or mTOR deletion alone. Indeed, they display muscle weakness, increased fiber denervation, and a slower muscle relaxation following tetanic stimulation. This is accompanied by a shift towards slow-twitch fibers and changes in the expression levels of calcium-related genes, like Serca1 and Casq1. Indeed, dk.o. mice show a decrease in calcium decay kinetics after tetanus in vivo, suggestive of a reduced calcium reuptake. In addition, RNA sequencing analysis revealed that many downregulated genes are linked to sarcomere organization, like Tcap and Fhod3. These results suggest a key role for mTOR signaling in maintaining a proper fiber relaxation in skeletal muscle. Abstract figure legend This article is protected by copyright. All rights reserved.
    Keywords:  Raptor; calcium; mTOR; muscle force; relaxation; skeletal muscle
    DOI:  https://doi.org/10.1113/JP283686
  34. J Biol Chem. 2022 Oct 17. pii: S0021-9258(22)01058-4. [Epub ahead of print] 102615
    Purine nucleoside phosphorylase controls nicotinamide riboside metabolism in mammalian cells.
    Andrey Kropotov, Veronika Kulikova, Ljudmila Solovjeva, Alexander Yakimov, Kirill Nerinovski, Maria Svetlova, Julia Sudnitsyna, Alena Plusnina, Maria Antipova, Mikhail Khodorkovskiy, Marie E Migaud, Stepan Gambaryan, Mathias Ziegler, Andrey Nikiforov.
      Nicotinamide riboside (NR) is an effective precursor of nicotinamide adenine dinucleotide (NAD) in human and animal cells. NR supplementation can increase the level of NAD in various tissues and thereby improve physiological functions that are weakened or lost in experimental models of aging or various human pathologies. However, there are also reports questioning the efficacy of NR supplementation. Indeed, the mechanisms of its utilization by cells are not fully understood. Herein, we investigated the role of purine nucleoside phosphorylase (PNP) in NR metabolism in mammalian cells. Using both PNP overexpression and genetic knockout, we show that after being imported into cells by members of the equilibrative nucleoside transporter family, NR is predominantly metabolized by PNP, resulting in nicotinamide (Nam) accumulation. Intracellular cleavage of NR to Nam is prevented by the potent PNP inhibitor Immucillin H in various types of mammalian cells. In turn, suppression of PNP activity potentiates NAD synthesis from NR. Combining pharmacological inhibition of PNP with NR supplementation in mice, we demonstrate that the cleavage of the riboside to Nam is strongly diminished, maintaining high levels of NR in blood, kidney and liver. Moreover, we show that PNP inhibition stimulates Nam mononucleotide and NAD+ synthesis from NR in vivo, in particular, in the kidney. Thus, we establish PNP as a major regulator of NR metabolism in mammals and provide evidence that the health benefits of NR supplementation could be greatly enhanced by concomitant downregulation of PNP activity.
    Keywords:  NAD biosynthesis; human; metabolism; mouse; nicotinamide adenine dinucleotide (NAD); nicotinamide riboside; purine nucleoside phosphorylase
    DOI:  https://doi.org/10.1016/j.jbc.2022.102615
  35. Mol Genet Metab. 2022 Oct 12. pii: S1096-7192(22)00418-8. [Epub ahead of print]137(3): 301-307
    A simple screening method for heterozygous female patients with ornithine transcarbamylase deficiency.
    Hiroyuki Iijima, Mitsuru Kubota.
      Ornithine transcarbamylase deficiency (OTCD), caused by X-linked OTC mutations, is characterized by life-threatening hyperammonemia. Heterozygous female patients are often asymptomatic and usually have milder disease than affected male patients, but can have higher morbidity and mortality rates if the disease progresses prior to diagnosis. Our purpose was to establish a screening method for female heterozygotes with OTCD. We retrospectively identified female patients who underwent plasma amino acid analysis at the National Center for Child Health and Development, using data from electronic medical records from March 2002 to September 2021. We extracted patient age, medical history, and biochemical data, including plasma amino acid levels. Patients were categorized into several groups according to their underlying diseases; those with underlying diseases that could potentially affect plasma amino acid levels, such as mitochondrial disease or short bowel syndrome, were excluded, except for untreated OTCD. Biochemical values were compared between OTCD patients and others using the Mann-Whitney U test. The receiver operator characteristic analysis was performed to assess the diagnostic capability for detecting OTCD in each subject. For patients with multiple test data, the most recent of the measurement dates was used in the analysis. The data sets of 976 patients were included. There were significant differences in values of glutamine, citrulline, arginine, and ammonia, but the diagnostic capacity of each alone was inadequate. By contrast, the (glutamine + glycine)/(citrulline + arginine) ratio was appropriate for discriminating heterozygous female patients with OTCD, with a sensitivity of 100% and specificity of 98.6% when the cutoff level was 15.8; the AUC for this discrimination was 0.996 (95% confidence interval, 0.992 to 1.000). These findings could help identify heterozygous female patients with OTCD before the onset of clinical disease.
    Keywords:  Amino acids; Female heterozygotes; Hyperammonemia; Ornithine transcarbamylase deficiency; Screening; Urea cycle
    DOI:  https://doi.org/10.1016/j.ymgme.2022.10.003
  36. Nature. 2022 Oct 19.
    A protein that mobilizes the cofactor molecule haem for use in cells.
      
    Keywords:  Biochemistry; Cell biology; Metabolism
    DOI:  https://doi.org/10.1038/d41586-022-03149-x
  37. J Biomed Inform. 2022 Oct 15. pii: S1532-0464(22)00232-5. [Epub ahead of print] 104227
    Deep Learning for Rare Disease: A Scoping Review.
    Junghwan Lee, Cong Liu, Junyoung Kim, Zhehuan Chen, Yingcheng Sun, James R Rogers, Wendy K Chung, Chunhua Weng.
      Although individually rare, collectively more than 7,000 rare diseases affect about 10% of patients. Each of the rare diseases impacts the quality of life for patients and their families, and incurs significant societal costs. The low prevalence of each rare disease causes formidable challenges in accurately diagnosing and caring for these patients and engaging participants in research to advance treatments. Deep learning has advanced many scientific fields and has been applied to many healthcare tasks. This study reviewed the current uses of deep learning to advance rare disease research. Among the 332 reviewed articles, we found that deep learning has been actively used for rare neoplastic diseases (250/332), followed by rare genetic diseases (170/332) and rare neurological diseases (127/332). Convolutional neural networks (307/332) were the most frequently used deep learning architecture, presumably because image data were the most commonly available data type in rare disease research. Diagnosis is the main focus of rare disease research using deep learning (263/332). We summarized the challenges and future research directions for leveraging deep learning to advance rare disease research.
    Keywords:  Deep Learning; Machine Learning; Rare Disease
    DOI:  https://doi.org/10.1016/j.jbi.2022.104227
  38. Lab Chip. 2022 Oct 18.
    Recent advances in high-throughput single-cell transcriptomics and spatial transcriptomics.
    Xiaohan Shen, Yichun Zhao, Zhuo Wang, Qihui Shi.
      Single-cell RNA sequencing (scRNA-seq) has been developed for characterizing the transcriptome of cells that are rare but of biological significance. With cell barcoding and microchip technologies, a suite of high-throughput scRNA-seq protocols enable transcriptome profiling in thousands of individual cells at single-cell resolution for classifying cell types, discovering novel cell populations, investigating cellular heterogeneity and elucidating lineage trajectories. Microchip technologies including microfluidics- and microwell-based platforms play a major role in high-throughput scRNA-seq. As the emerging technology, spatial transcriptomics integrates cellular transcriptomics with their spatial coordinates within tissues for spatially deciphering cellular composition, heterogeneity and cell-cell communications. Spatial transcriptomics has been increasingly recognized as one of the most powerful tools for discovering new biology and advancing precision medicine. Microfluidics as an enabling technology plays an increasingly important role in spatial transcriptomics. We review the technological spectrum and advances in high-throughput scRNA-seq and spatial transcriptomics, discuss their advantages and limitations, and pitch into new biology learned from these new tools.
    DOI:  https://doi.org/10.1039/d2lc00633b
  39. Neurochem Int. 2022 Oct 14. pii: S0197-0186(22)00159-0. [Epub ahead of print] 105434
    Social isolation induces succinate dehydrogenase dysfunction in anxious mice.
    Saki Watanabe, Alzahra J Al Omran, Amy S Shao, Zeyu Zhang, Chen Xue, Jifeng Zhang, Junji Watanabe, Jing Liang.
      We have previously reported social isolation induces anxiety-like behavior, cognition decline, and reduction in brain ATP levels in mice. These changes were ameliorated by treatment with dihydromyricetin (DHM), a compound that positively modulates γ-aminobutyric A (GABAA) receptor. To gain further insight into the subcellular mechanisms underlying these changes, we utilized a social isolation-induced anxiety mouse model and investigated changes in mitochondrial oxidative capacity via the electron transport chain. We found that 4 weeks of social isolation decreased ATP levels by 43% and succinate dehydrogenase capacity by 52% of the control, while daily DHM (2 mg/kg oral) administration restored succinate dehydrogenase capacity. These results suggest that social isolation decreased mitochondrial capacity to generate ATP. DHM can be developed to be a therapeutic against anxiety and mitochondrial stress.
    Keywords:  Anxiety; Complex II; Dihydromyricetin; Social isolation; Stress; Succinate dehydrogenase
    DOI:  https://doi.org/10.1016/j.neuint.2022.105434
  40. Nat Metab. 2022 Oct;4(10): 1219-1220
    Gut reaction: it's not all about enzymes.
    Stephan Kamrad, Anna E Lindell, Kiran R Patil.
      
    DOI:  https://doi.org/10.1038/s42255-022-00637-2
  41. Circ Genom Precis Med. 2022 Oct 11. e003686
    Genetic Basis of Childhood Cardiomyopathy.
    Richard D Bagnall, Emma S Singer, Julie Wacker, Natalie Nowak, Jodie Ingles, Ingrid King, Ivan Macciocca, Joshua Crowe, Anne Ronan, Robert G Weintraub, Christopher Semsarian.
       BACKGROUND: The causes of cardiomyopathy in children are less well described than in adults. We evaluated the clinical diagnoses and genetic causes of childhood cardiomyopathy and outcomes of cascade genetic testing in family members.
    METHODS: We recruited children from a pediatric cardiology service or genetic heart diseases clinic. We performed Sanger, gene panel, exome or genome sequencing and classified variants for pathogenicity using American College of Molecular Genetics and Genomics guidelines.
    RESULTS: Cardiomyopathy was diagnosed in 221 unrelated children aged ≤18 years. Children mostly had hypertrophic cardiomyopathy (n=98, 44%) or dilated cardiomyopathy (n=89, 40%). The highest genetic testing diagnostic yields were in restrictive cardiomyopathy (n=16, 80%) and hypertrophic cardiomyopathy (n=65, 66%), and lowest in dilated cardiomyopathy (n=26, 29%) and left ventricular noncompaction (n=3, 25%). Pathogenic variants were primarily found in genes encoding sarcomere proteins, with TNNT2 and TNNI3 variants associated with more severe clinical outcomes. Ten children (4.5%) had multiple pathogenic variants. Genetic test results prompted review of clinical diagnosis in 14 families with syndromic, mitochondrial or metabolic gene variants. Cascade genetic testing in 127 families confirmed 24 de novo variants, recessive inheritance in 8 families, and supported reclassification of 12 variants.
    CONCLUSIONS: Genetic testing of children with cardiomyopathy supports a precise clinical diagnosis, which may inform prognosis.
    Keywords:  cardiomyopathy; child; family; genetics; sarcomere
    DOI:  https://doi.org/10.1161/CIRCGEN.121.003686
  42. Nat Metab. 2022 Oct;4(10): 1217
    The gut drives fat cravings.
    Joseph Willson.
      
    DOI:  https://doi.org/10.1038/s42255-022-00661-2
  43. Front Genet. 2022 ;13 982930
    Gene-specific machine learning model to predict the pathogenicity of BRCA2 variants.
    Mohannad N Khandakji, Borbala Mifsud.
      Background: Existing BRCA2-specific variant pathogenicity prediction algorithms focus on the prediction of the functional impact of a subtype of variants alone. General variant effect predictors are applicable to all subtypes, but are trained on putative benign and pathogenic variants and do not account for gene-specific information, such as hotspots of pathogenic variants. Local, gene-specific information have been shown to aid variant pathogenicity prediction; therefore, our aim was to develop a BRCA2-specific machine learning model to predict pathogenicity of all types of BRCA2 variants. Methods: We developed an XGBoost-based machine learning model to predict pathogenicity of BRCA2 variants. The model utilizes general variant information such as position, frequency, and consequence for the canonical BRCA2 transcript, as well as deleteriousness prediction scores from several tools. We trained the model on 80% of the expert reviewed variants by the Evidence-Based Network for the Interpretation of Germline Mutant Alleles (ENIGMA) consortium and tested its performance on the remaining 20%, as well as on an independent set of variants of uncertain significance with experimentally determined functional scores. Results: The novel gene-specific model predicted the pathogenicity of ENIGMA BRCA2 variants with an accuracy of 99.9%. The model also performed excellently on predicting the functional consequence of the independent set of variants (accuracy was up to 91.3%). Conclusion: This new, gene-specific model is an accurate method for interpreting the pathogenicity of variants in the BRCA2 gene. It is a valuable addition for variant classification and can prioritize unreviewed variants for functional analysis or expert review.
    Keywords:  VUS; breast cancer; in-silico predictions; variant pathogenicity; variant prioritization
    DOI:  https://doi.org/10.3389/fgene.2022.982930
  44. Elife. 2022 Oct 20. pii: e81608. [Epub ahead of print]11
    Didemnin B and ternatin-4 differentially inhibit conformational changes in eEF1A required for aminoacyl-tRNA accommodation into mammalian ribosomes.
    Manuel F Juette, Jordan D Carelli, Emily J Rundlet, Alan Brown, Sichen Shao, Angelica Ferguson, Michael R Wasserman, Mikael Holm, Jack Taunton, Scott C Blanchard.
      Rapid and accurate mRNA translation requires efficient codon-dependent delivery of the correct aminoacyl-tRNA (aa-tRNA) to the ribosomal A site. In mammals, this fidelity-determining reaction is facilitated by the GTPase elongation factor-1 alpha (eEF1A), which escorts aa-tRNA as an eEF1A(GTP)-aa-tRNA ternary complex into the ribosome. The structurally unrelated cyclic peptides didemnin B and ternatin-4 bind to the eEF1A(GTP)-aa-tRNA ternary complex and inhibit translation but have different effects on protein synthesis in vitro and in vivo. Here, we employ single-molecule fluorescence imaging and cryogenic electron microscopy to determine how these natural products inhibit translational elongation on mammalian ribosomes. By binding to a common site on eEF1A, didemnin B and ternatin-4 trap eEF1A in an intermediate state of aa-tRNA selection, preventing eEF1A release and aa-tRNA accommodation on the ribosome. We also show that didemnin B and ternatin-4 exhibit distinct effects on the dynamics of aa-tRNA selection that inform on observed disparities in their inhibition efficacies and physiological impacts. These integrated findings underscore the value of dynamics measurements in assessing the mechanism of small-molecule inhibition and highlight potential of single-molecule methods to reveal how distinct natural products differentially impact the human translation mechanism.
    Keywords:  Didemnin; Ternatin; biochemistry; chemical biology; eEF1A; human; o. cuniculus; ribosome; smFRET
    DOI:  https://doi.org/10.7554/eLife.81608
  45. Int J Biol Macromol. 2022 Oct 14. pii: S0141-8130(22)02224-3. [Epub ahead of print]
    Baicalein modulates mitochondrial function by upregulating mitochondrial uncoupling protein-1 (UCP1) expression in brown adipocytes, cytotoxicity, and computational studies.
    Md Reyad-Ul-Ferdous, Yongfeng Song.
       BACKGROUND: Obesity, fatty liver, type 2 diabetes, and Non-alcoholic fatty liver disease (NAFLD) are all metabolic diseases caused by excess food consumption. Existing drug molecules had negative side effects and caused other diseases to develop (Orlistat causes angioedema, and menstrual irregularities; megestrol acetate causes hypertension, and insomnia). By enhancing lipid consumption and increasing nonshivering thermogenesis, targeting mitochondrial uncoupling protein-1 (UCP1) expression in adipocytes could be an auspicious treatment strategy against obesity or metabolic disorders associated with obesity.
    METHODS: We used previously produced UCP1-A-GFP reporter cell lines in this investigation to find new pharmacological compounds against obesity or metabolic syndrome, which we then tested in cellular analysis, cytotoxicity, mitochondrial function, mitochondrial DNA quantification, mitochondrial ATP production, and in-silico models.
    RESULTS: Baicalein was discovered to play a critical role in obesity prevention via altering mitochondrial function. Baicalein lowers ATP generation while increasing considerable UCP1 gene expression in brown adipocytes. As a result, cellular thermogenesis is boosted. The HEK293T cell line is harmless by baicalein. The investigation by the in-silico study revealed drug-protein interaction and UCP1 binding. Thus, our research clarifies baicalein's therapeutic role in metabolic and obesity-related illnesses via modulating mitochondrial activity (Supplementary Fig. 2).
    CONCLUSIONS: Further studies are required in both murine and human models to understand the full mechanism of action by mitochondrial modulation. Drug development investigation also requires to development of a precise formulation.
    Keywords:  Drug screening; In-silico study; Mitochondrial function; Mitochondrial uncoupling protein-1 (UCP1); Obesity
    DOI:  https://doi.org/10.1016/j.ijbiomac.2022.09.285
  46. Biomed Pharmacother. 2022 Nov;pii: S0753-3322(22)01222-7. [Epub ahead of print]155 113833
    Skeletal muscle mitochondrial remodeling in heart failure: An update on mechanisms and therapeutic opportunities.
    Jiayu Lv, Yumeng Li, Shuqing Shi, Xia Xu, Huaqin Wu, Bingxuan Zhang, Qingqiao Song.
      Patients with heart failure (HF) usually present with skeletal muscle diseases of varying severity, ranging from early fatigue on exercise to sarcopenia, sarcopenic obesity or cachexia, and frailty, which are significant predictors of HF prognosis. Abnormal mitochondrial metabolism has been identified as one of the earliest signs of skeletal muscle injury in HF and is associated with pathological alterations in muscle, manifested as muscle wasting, myocyte atrophy and apoptosis, fiber type shift, impaired contractile coupling, and muscle fat infiltration. In this review, we update the evidence for skeletal muscle mitochondrial remodeling in HF patients or animal models, including the impairments in mitochondrial ultrastructure, oxidative metabolism, electron transport chain (ETC), phosphorylation apparatus, phosphotransfer system, and quality control. We also focus on molecular regulatory mechanisms upstream of mitochondria, including circulating factors (e.g., RAAS, TNF-α IL-6, IGF-1, GH, ghrelin, adiponectin, NO) and molecular signals within myocytes (e.g., PGC-1α, PPARs, AMPK, SIRT1/3, ROS, and MuRF1). Besides the therapies targeting the signaling pathways mentioned above, such as AdipoRon and elamipretide, we further summarize other potential pharmacological approaches like inhibitors of sodium-glucose cotransporter 2 (SGLT2) and dipeptidyl peptidase-4 (DPP-4), as well as some natural products, which may have the beneficial effects on improving the skeletal muscle mitochondrial function of HF. Targeting myocyte mitochondrial biogenesis, oxidative metabolism, oxidative phosphorylation, and reduction of oxidative stress injury are promising future opportunities for the prevention and management of skeletal muscle myopathy in HF.
    Keywords:  Heart failure; Mitochondria; Molecular mechanisms; Pharmacological targets; Skeletal muscle
    DOI:  https://doi.org/10.1016/j.biopha.2022.113833
  47. Nat Genet. 2022 Oct 17.
    Targeted profiling of human extrachromosomal DNA by CRISPR-CATCH.
    King L Hung, Jens Luebeck, Siavash R Dehkordi, Caterina I Colón, Rui Li, Ivy Tsz-Lo Wong, Ceyda Coruh, Prashanthi Dharanipragada, Shirley H Lomeli, Natasha E Weiser, Gatien Moriceau, Xiao Zhang, Chris Bailey, Kathleen E Houlahan, Wenting Yang, Rocío Chamorro González, Charles Swanton, Christina Curtis, Mariam Jamal-Hanjani, Anton G Henssen, Julie A Law, William J Greenleaf, Roger S Lo, Paul S Mischel, Vineet Bafna, Howard Y Chang.
      Extrachromosomal DNA (ecDNA) is a common mode of oncogene amplification but is challenging to analyze. Here, we adapt CRISPR-CATCH, in vitro CRISPR-Cas9 treatment and pulsed field gel electrophoresis of agarose-entrapped genomic DNA, previously developed for bacterial chromosome segments, to isolate megabase-sized human ecDNAs. We demonstrate strong enrichment of ecDNA molecules containing EGFR, FGFR2 and MYC from human cancer cells and NRAS ecDNA from human metastatic melanoma with acquired therapeutic resistance. Targeted enrichment of ecDNA versus chromosomal DNA enabled phasing of genetic variants, identified the presence of an EGFRvIII mutation exclusively on ecDNAs and supported an excision model of ecDNA genesis in a glioblastoma model. CRISPR-CATCH followed by nanopore sequencing enabled single-molecule ecDNA methylation profiling and revealed hypomethylation of the EGFR promoter on ecDNAs. We distinguished heterogeneous ecDNA species within the same sample by size and sequence with base-pair resolution and discovered functionally specialized ecDNAs that amplify select enhancers or oncogene-coding sequences.
    DOI:  https://doi.org/10.1038/s41588-022-01190-0
  48. Leuk Lymphoma. 2022 Oct 19. 1-10
    Circulating mitochondrial DNA is associated with anemia in newly diagnosed hematologic malignancies.
    Liang Yue, Yu-Hang Li, Rui-Lin Ma, Jing-Wen Niu, Hong-Tu Cui, Yao Sun, Zhi-Min Yun, Hai-Long Zhuo, Lu-Ming Wan, Su-Bo Li, Xue Zhang, Cheng-Jun Wu, Liang-Ding Hu, Ying-Xia Tan.
      Recent reports discovered that red blood cells (RBCs) could scavenge cell-free mitochondrial DNA (mtDNA), which drives the accelerated erythrophagocytosis and innate immune activation characterized by anemia and inflammatory cytokine production. However, the clinical value of the circulating mtDNA copy number alterations in hematologic malignancies is poorly understood. Our data showed that in comparison to healthy group, the patients group had significantly higher mtDNA and histone H4 levels. Moreover, we observed that RBC-bound mtDNA and histone H4 were negatively correlated with hemoglobin in patients. In addition, cytokines and chemokines levels in patients differed significantly from normal controls (21 higher, 7 lower). Our study suggested that both circulating mtDNA and histone H4 were associated with anemia in hematologic malignancies, which helps to further understand the potential mechanism of anemia development in patients with hematologic malignancies. This information may play a vital role in the specific therapeutic interventions for leukemia in the future.
    Keywords:  Mitochondrial DNA; RBC; anemia; hematologic malignancies
    DOI:  https://doi.org/10.1080/10428194.2022.2133537
  49. J Pathol Inform. 2022 ;13 100130
    A performance evaluation study: Variant annotation tools - the enigma of clinical next generation sequencing (NGS) based genetic testing.
    Sachleen Tuteja, Sabah Kadri, Kai Lee Yap.
      Dramatically expanding our ability for clinical genetic testing for inherited conditions and complex diseases such as cancer, next generation sequencing (NGS) technologies are allowing for rapid interrogation of thousands of genes and identification of millions of variants. Variant annotation, the process of assigning functional information to DNA variants based on the standardized Human Genome Variation Society (HGVS) nomenclature, is a fundamental challenge in the analysis of NGS data that has led to the development of many bioinformatic algorithms. In this study, we evaluated the performance of 3 variant annotation tools: Alamut® Batch, Ensembl Variant Effect Predictor (VEP), and ANNOVAR, benchmarked by a manually curated ground-truth set of 298 variants from the medical exome database at the Molecular Diagnostics Laboratory at Lurie Children's Hospital. Of the 3 tools, VEP produces the most accurate variant annotations (HGVS nomenclature for 297 of the 298 variants) due to usage of updated gene transcript versions within the algorithm. Alamut® Batch called 296 of the 298 variants correctly; strikingly, ANNOVAR exhibited the greatest number of discrepancies (20 of the 298 variants, 93.3% concordance with ground-truth set). Adoption of validated methods of variant annotation is critical in post-analytical phases of clinical testing.
    Keywords:  ANNOVAR; Alamut®; Gene panel; Genetic testing; VEP; Variant annotation
    DOI:  https://doi.org/10.1016/j.jpi.2022.100130
  50. Sci Rep. 2022 Oct 20. 12(1): 17578
    Mitochondrial biogenesis, telomere length and cellular senescence in Parkinson's disease and Lewy body dementia.
    Muhammad Asghar, Amani Odeh, Ahmad Jouni Fattahi, Alexandra Edwards Henriksson, Aurelie Miglar, Shervin Khosousi, Per Svenningsson.
      Progressive age is the single major risk factor for neurodegenerative diseases. Cellular aging markers during Parkinson's disease (PD) have been implicated in previous studies, however the majority of studies have investigated the association of individual cellular aging hallmarks with PD but not jointly. Here, we have studied the association of PD with three aging hallmarks (telomere attrition, mitochondrial dysfunction, and cellular senescence) in blood and the brain tissue. Our results show that PD patients had 20% lower mitochondrial DNA copies but 26% longer telomeres in blood compared to controls. Moreover, telomere length in blood was positively correlated with medication (Levodopa Equivalent Daily Dose, LEDD) and disease duration. Similar results were found in brain tissue, where patients with Parkinson's disease (PD), Parkinson's disease dementia (PDD) and Dementia with Lewy Bodies (DLB) showed (46-95%) depleted mtDNA copies, but (7-9%) longer telomeres compared to controls. In addition, patients had lower mitochondrial biogenesis (PGC-1α and PGC-1β) and higher load of a cellular senescence marker in postmortem prefrontal cortex tissue, with DLB showing the highest effect among the patient groups. Our results suggest that mitochondrial dysfunction (copy number and biogenesis) in blood might be a valuable marker to assess the risk of PD. However, further studies with larger sample size are needed to evaluate these findings.
    DOI:  https://doi.org/10.1038/s41598-022-22400-z
  51. Nucleic Acids Res. 2022 Oct 16. pii: gkac869. [Epub ahead of print]
    HiFENS: high-throughput FISH detection of endogenous pre-mRNA splicing isoforms.
    Asaf Shilo, Gianluca Pegoraro, Tom Misteli.
      Splicing factors play an essential role in regulation of alternative pre-mRNA splicing. While much progress has been made in delineating the mechanisms of the splicing machinery, the identity of signal transduction pathways and upstream factors that regulate splicing factor activity is largely unknown. A major challenge in the discovery of upstream regulatory factors of pre-mRNA splicing is the scarcity of functional genomics screening methods to monitor splicing outcomes of endogenous genes. Here, we have developed HiFENS (high throughput FISH detection of endogenous splicing isoforms), a high-throughput imaging assay based on hybridization chain reaction (HCR) and used HiFENS to screen for cellular factors that regulate alternative splicing of endogenous genes. We demonstrate optimized detection with high specificity of endogenous splicing isoforms and multiplexing of probes for accurate detection of splicing outcomes with single cell resolution. As proof-of-principle, we perform an RNAi screen of 702 human kinases and identify potential candidate upstream splicing regulators of the FGFR2 gene. HiFENS should be a useful tool for the unbiased delineation of cellular pathways involved in alternative splicing regulation.
    DOI:  https://doi.org/10.1093/nar/gkac869
  52. Nature. 2022 Oct 20.
    Recognition of cyclic dinucleotides and folates by human SLC19A1.
    Qixiang Zhang, Xuyuan Zhang, Yalan Zhu, Panpan Sun, Liwei Zhang, Junxiao Ma, Yong Zhang, Lingan Zeng, Xiaohua Nie, Yina Gao, Zhaolong Li, Songqing Liu, Jizhong Lou, Ang Gao, Liguo Zhang, Pu Gao.
      Cyclic dinucleotides (CDNs) are ubiquitous signaling molecules in all domains of life1,2. Mammalian cells produce one CDN, 2'3'-cGAMP, by cyclic GMP-AMP synthase upon detecting cytosolic DNA signals3-7. 2'3'-cGAMP, as well as bacterial and synthetic CDN analogs, can act as second messengers to activate stimulator of interferon genes (STING) and elicit broad downstream responses8-21. Extracellular CDNs must traverse the cell membrane to activate STING, a process that is critically dependent on the solute carrier SLC19A122,23. In addition, SLC19A1 represents the major transporter for folate nutrients and antifolate therapeutics24,25, thereby placing SLC19A1 as a key factor in multiple physiological and pathological processes. How SLC19A1 recognizes and transports CDNs and folate/antifolate is unknown. Here we report cryo-electron microscopy structures of human SLC19A1 (hSLC19A1) in a substrate-free state and in complexes with multiple CDNs from different sources, a predominant natural folate, and a new-generation antifolate drug. Structural and mutagenesis results demonstrate that hSLC19A1 utilizes unique yet divergent mechanisms to recognize CDN- and folate-type substrates. Two CDN molecules bind within the hSLC19A1 cavity as a compact dual-molecule unit, while folate or antifolate binds as a monomer and occupies a distinct pocket of the cavity. Moreover, the structures allow accurate mapping and potential mechanistic interpretation of loss-of-activity and disease-related mutations of hSLC19A1. Our work provides a framework for understanding the mechanism of SLC19 family transporters and serves as a foundation for the development of potential therapeutics.
    DOI:  https://doi.org/10.1038/s41586-022-05452-z
  53. Nat Rev Methods Primers. 2022 Dec;pii: s43586-022-00100-2. [Epub ahead of print]2(1):
    Designing clinical trials for rare diseases: unique challenges and opportunities.
    Chiara Pizzamiglio, Hilary J Vernon, Michael G Hanna, Robert D S Pitceathly.
      Orphan drug development is a rapidly expanding field. Nevertheless, clinical trials for rare diseases can present inherent challenges. Optimal study design and partnerships between academia and industry are therefore required for the successful development, delivery and clinical approval of effective therapies in this group of disorders.
    DOI:  https://doi.org/10.1038/s43586-022-00100-2
  54. Nature. 2022 Oct 19.
    Semi-automated assembly of high-quality diploid human reference genomes.
    Human Pangenome Reference Consortium
      The current human reference genome, GRCh38, represents over 20 years of effort to generate a high-quality assembly, which has benefitted society1,2. However, it still has many gaps and errors, and does not represent a biological genome as it is a blend of multiple individuals3,4. Recently, a high-quality telomere-to-telomere reference, CHM13, was generated with the latest long-read technologies, but it was derived from a hydatidiform mole cell line with a nearly homozygous genome5. To address these limitations, the Human Pangenome Reference Consortium formed with the goal of creating high-quality, cost-effective, diploid genome assemblies for a pangenome reference that represents human genetic diversity6. Here, in our first scientific report, we determined which combination of current genome sequencing and assembly approaches yield the most complete and accurate diploid genome assembly with minimal manual curation. Approaches that used highly accurate long reads and parent-child data with graph-based haplotype phasing during assembly outperformed those that did not. Developing a combination of the top-performing methods, we generated our first high-quality diploid reference assembly, containing only approximately four gaps per chromosome on average, with most chromosomes within ±1% of the length of CHM13. Nearly 48% of protein-coding genes have non-synonymous amino acid changes between haplotypes, and centromeric regions showed the highest diversity. Our findings serve as a foundation for assembling near-complete diploid human genomes at scale for a pangenome reference to capture global genetic variation from single nucleotides to structural rearrangements.
    DOI:  https://doi.org/10.1038/s41586-022-05325-5
  55. Nat Cell Biol. 2022 Oct 20.
    The beauty of simplicity in membrane biology.
    Jeanne C Stachowiak, Tomas Kirchhausen.
      
    DOI:  https://doi.org/10.1038/s41556-022-01015-6
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