bims-mitdis Biomed News
on Mitochondrial disorders
Issue of 2023–10–08
sixty-six papers selected by
Catalina Vasilescu, Helmholz Munich
  1. Wide diagnostic and genotypic spectrum in patients with suspected mitochondrial disease.
  2. Nuclear genome influences mitochondrial DNA.
  3. Mechanisms and regulation of human mitochondrial transcription.
  4. Metabolomics and mitochondrial dysfunction in cardiometabolic disease.
  5. Improving lysosomal ferroptosis with NMN administration protects against heart failure.
  6. ClpP/ClpX deficiency impairs mitochondrial functions and mTORC1 signaling during spermatogenesis.
  7. Research Advances on Deafness Genes Associated with Mitochondrial tRNA-37 Modifications.
  8. Mitochondrial Targeted Interventions for Aging.
  9. Mitophagy for cardioprotection.
  10. Mitochondrial Membrane Model: Lipids, Elastic Properties and the Changing Curvature of Cardiolipin.
  11. Base editing of organellar DNA with programmable deaminases.
  12. Purification of functional mouse skeletal muscle mitochondria using percoll density gradient centrifugation.
  13. Mitochondrial transfer in hematological malignancies.
  14. Caloric restriction promotes beta cell longevity and delays aging and senescence signatures.
  15. Knockdown of BRAWNIN minimally affect mitochondrial complex III assembly in human cells.
  16. Mitochondrial Cytochrome C deficiency can show the first disease signs in the prenatal stage.
  17. Integrated analysis of transcriptomic and proteomic alterations in mouse models of ALS/FTD identify early metabolic adaptions with similarities to mitochondrial dysfunction disorders.
  18. Implementation of Exome Sequencing to Identify Rare Genetic Diseases.
  19. Inter-organellar nucleic acid communication by a mitochondrial tRNA regulates nuclear metabolic transcription.
  20. The glyoxylate shunt protein ICL-1 protects from mitochondrial superoxide stress through activation of the mitochondrial unfolded protein response.
  21. Lithium isotopes differentially modify mitochondrial amorphous calcium phosphate cluster size distribution and calcium capacity.
  22. Long term survival and abnormal liver fat accumulation in mice with specific thymidine kinase 2 deficiency in liver tissue.
  23. Significant decrease of maternal mitochondria carryover using optimized spindle-chromosomal complex transfer.
  24. Cleavage kinetics of human mitochondrial RNase P and contribution of its non-nuclease subunits.
  25. Deleterious heteroplasmic mitochondrial mutations are associated with an increased risk of overall and cancer-specific mortality.
  26. Complete mitochondrial DNA profile in stroke: a geographical matched case-control study in Spanish population.
  27. Single-Nucleotide Resolution Mapping of N6-Methyladenine in Genomic DNA.
  28. Complementary Approaches to Interrogate Mitophagy Flux in Pancreatic β-Cells.
  29. Gene-specific ACMG/AMP classification criteria for germline APC variants: recommendations from the ClinGen InSiGHT Hereditary Colorectal Cancer / Polyposis Variant Curation Expert Panel.
  30. Conformational analysis of membrane-proximal segments of GDAP1 in a lipidic environment using synchrotron radiation suggests a mode of assembly at the mitochondrial outer membrane.
  31. Light-sensitive phosphorylation regulates enzyme activity and filament assembly of human IMPDH1 retinal splice variants.
  32. Spatial single-cell mass spectrometry defines zonation of the hepatocyte proteome.
  33. Alternate splice variants of the mitochondrial fission protein DNM1L/Drp1 regulate mitochondrial dynamics and cell fate in ovarian cancer.
  34. Second international consensus report on gaps and opportunities for the clinical translation of precision diabetes medicine.
  35. Mitochondrial dysfunction: A fatal blow in depression.
  36. A common benign intronic deletion masking a pathogenic deep intronic PCCB variant - genome sequencing and RNA studies to the rescue.
  37. Cytoplasmic Endonuclease G promotes nonalcoholic fatty liver disease via mTORC2-AKT-ACLY and endoplasmic reticulum stress.
  38. tRNA renovatio: Rebirth through fragmentation.
  39. A Galaxy of informatics resources for MS-based proteomics.
  40. Mitochondrial DNA damage triggers spread of Parkinson's disease-like pathology.
  41. Editorial: Whole Genome Sequencing for rare diseases.
  42. A milder form of molybdenum cofactor deficiency type A presenting as Leigh's syndrome-like phenotype highlighting the secondary mitochondrial dysfunction: a case report.
  43. Expanding what we know about rare genetic diseases.
  44. KDM5-mediated activation of genes required for mitochondrial biology is necessary for viability in Drosophila.
  45. Streamlined and sensitive mono- and di-ribosome profiling in yeast and human cells.
  46. Long-read sequencing resolves a complex structural variant in PRKN Parkinson's disease.
  47. Ketone Bodies Preserve Mitochondria Through Epigenetics.
  48. Mitochondrial DNA heteroplasmy analysis in keratoconus patients from China.
  49. A conserved, non-canonical insert in FIS1 mediates TBC1D15 and DRP1 recruitment for mitochondrial fission.
  50. Machine learning nominates the inositol pathway and novel genes in Parkinson's disease.
  51. Fuelling the fire: de novo lipogenesis primes thermogenesis.
  52. Gene therapies for rare diseases are under threat. Scientists hope to save them.
  53. Metabolic Regulation of Aged Hematopoietic Stem Cells: Key Players and Mechanisms.
  54. Regulation of defective mitochondrial DNA accumulation and transmission in C. elegans by the programmed cell death and aging pathways.
  55. Genomic variant benchmark: if you cannot measure it, you cannot improve it.
  56. Anticodon sequence determines the impact of mistranslating tRNAAla variants.
  57. Knowledge, attitudes and decision regret: a longitudinal survey study of participants offered genome sequencing in the 100,000 Genomes Project.
  58. Association of 2-oxoacid dehydrogenase complexes with respirasomes in mitochondria.
  59. Deciphering RNA splicing logic with interpretable machine learning.
  60. Spatiotemporal regulation of de novo and salvage purine synthesis during brain development.
  61. Analysis of a second-tier test panel in dried blood spot samples using liquid chromatography-tandem mass spectrometry in Catalonia's newborn screening programme.
  62. ApoE4-dependent lysosomal cholesterol accumulation impairs mitochondrial homeostasis and oxidative phosphorylation in human astrocytes.
  63. Refining the phenotypic spectrum of CCDC88A-related PEHO-like syndrome.
  64. Clinical and genetic analyses of a Swedish patient series diagnosed with ataxia.
  65. The mitochondrial amidoxime reducing component (mARC) - from prodrug-activation mechanism to drug-metabolising enzyme and onwards to drug target.
  66. Links between Exercise Capacity, Exercise Training, and Metabolism.
  1. Orphanet J Rare Dis. 2023 Oct 02. 18(1): 307
    Wide diagnostic and genotypic spectrum in patients with suspected mitochondrial disease.
    Kristina Grigalionienė, Birutė Burnytė, Laima Ambrozaitytė, Algirdas Utkus.
       BACKGROUND: Mitochondrial Diseases (MDs) are a diverse group of neurometabolic disorders characterized by impaired mitochondrial oxidative phosphorylation and caused by pathogenic variants in more than 400 genes. The implementation of next-generation sequencing (NGS) technologies helps to increase the understanding of molecular basis and diagnostic yield of these conditions. The purpose of the study was to investigate diagnostic and genotypic spectrum in patients with suspected MD. The comprehensive analysis of mtDNA variants using Sanger sequencing was performed in the group of 83 unrelated individuals with clinically suspected mitochondrial disease. Additionally, targeted next generation sequencing or whole exome sequencing (WES) was performed for 30 patients of the study group.
    RESULTS: The overall diagnostic rate was 21.7% for the patients with suspected MD, increasing to 36.7% in the group of patients where NGS methods were applied. Mitochondrial disease was confirmed in 11 patients (13.3%), including few classical mitochondrial syndromes (MELAS, MERRF, Leigh and Kearns-Sayre syndrome) caused by pathogenic mtDNA variants (8.4%) and MDs caused by pathogenic variants in five nDNA genes. Other neuromuscular diseases caused by pathogenic variants in seven nDNA genes, were confirmed in seven patients (23.3%).
    CONCLUSION: The wide spectrum of identified rare mitochondrial or neurodevelopmental diseases proves that MD suspected patients would mostly benefit from an extensive genetic profiling allowing rapid diagnostics and improving the care of these patients.
    Keywords:  Genetic diagnosis; Mitochondrial disease; Next generation sequencing; Variants; Whole exome sequencing; mtDNA; nDNA
    DOI:  https://doi.org/10.1186/s13023-023-02921-0
  2. Nat Rev Genet. 2023 Oct 06.
    Nuclear genome influences mitochondrial DNA.
    Henry Ertl.
      
    DOI:  https://doi.org/10.1038/s41576-023-00667-w
  3. Nat Rev Mol Cell Biol. 2023 Oct 02.
    Mechanisms and regulation of human mitochondrial transcription.
    Benedict G Tan, Claes M Gustafsson, Maria Falkenberg.
      The expression of mitochondrial genes is regulated in response to the metabolic needs of different cell types, but the basic mechanisms underlying this process are still poorly understood. In this Review, we describe how different layers of regulation cooperate to fine tune initiation of both mitochondrial DNA (mtDNA) transcription and replication in human cells. We discuss our current understanding of the molecular mechanisms that drive and regulate transcription initiation from mtDNA promoters, and how the packaging of mtDNA into nucleoids can control the number of mtDNA molecules available for both transcription and replication. Indeed, a unique aspect of the mitochondrial transcription machinery is that it is coupled to mtDNA replication, such that mitochondrial RNA polymerase is additionally required for primer synthesis at mtDNA origins of replication. We discuss how the choice between replication-primer formation and genome-length RNA synthesis is controlled at the main origin of replication (OriH) and how the recent discovery of an additional mitochondrial promoter (LSP2) in humans may change this long-standing model.
    DOI:  https://doi.org/10.1038/s41580-023-00661-4
  4. Life Sci. 2023 Oct 01. pii: S0024-3205(23)00772-5. [Epub ahead of print] 122137
    Metabolomics and mitochondrial dysfunction in cardiometabolic disease.
    Abhishek Shastry, Kimberly Dunham-Snary.
      Circulating metabolites are indicators of systemic metabolic dysfunction and can be detected through contemporary techniques in metabolomics. These metabolites are involved in numerous mitochondrial metabolic processes including glycolysis, fatty acid β-oxidation, and amino acid catabolism, and changes in abundance of these metabolites is implicated in the pathogenesis of cardiometabolic diseases (CMDs). Epigenetic regulation and direct metabolite-protein interactions modulate the metabolism, both within cells and in the circulation. Dysfunction of multiple mitochondrial components stemming from mitochondrial DNA mutations are implicated in disease pathogenesis. This review will summarize the current state of knowledge regarding: i) the interactions between metabolites found within the mitochondrial environment during CMDs, ii) various metabolites' effects on cellular and systemic function, iii) how harnessing the power of metabolomic analyses represents the next frontier of precision medicine, and iv) how these concepts integrate to expand the clinical potential for translational cardiometabolic medicine.
    Keywords:  Cardiometabolic disease; Circulating metabolites; Metabolic profiling; Metabolomics; Mitochondria; Redox balance
    DOI:  https://doi.org/10.1016/j.lfs.2023.122137
  5. Life Sci Alliance. 2023 Dec;pii: e202302116. [Epub ahead of print]6(12):
    Improving lysosomal ferroptosis with NMN administration protects against heart failure.
    Mikako Yagi, Yura Do, Haruka Hirai, Kenji Miki, Takahiro Toshima, Yukina Fukahori, Daiki Setoyama, Chiaki Abe, Yo-Ichi Nabeshima, Dongchon Kang, Takeshi Uchiumi.
      Myocardial mitochondria are primary sites of myocardial energy metabolism. Mitochondrial disorders are associated with various cardiac diseases. We previously showed that mice with cardiomyocyte-specific knockout of the mitochondrial translation factor p32 developed heart failure from dilated cardiomyopathy. Mitochondrial translation defects cause not only mitochondrial dysfunction but also decreased nicotinamide adenine dinucleotide (NAD+) levels, leading to impaired lysosomal acidification and autophagy. In this study, we investigated whether nicotinamide mononucleotide (NMN) administration, which compensates for decreased NAD+ levels, improves heart failure because of mitochondrial dysfunction. NMN administration reduced damaged lysosomes and improved autophagy, thereby reducing heart failure and extending the lifespan in p32cKO mice. We found that lysosomal damage due to mitochondrial dysfunction induced ferroptosis, involving the accumulation of iron in lysosomes and lipid peroxide. The ameliorative effects of NMN supplementation were found to strongly affect lysosomal function rather than mitochondrial function, particularly lysosome-mediated ferroptosis. NMN supplementation can improve lysosomal, rather than mitochondrial, function and prevent chronic heart failure.
    DOI:  https://doi.org/10.26508/lsa.202302116
  6. Commun Biol. 2023 Oct 05. 6(1): 1012
    ClpP/ClpX deficiency impairs mitochondrial functions and mTORC1 signaling during spermatogenesis.
    Chenxi Guo, Yuan Xiao, Jingkai Gu, Peikun Zhao, Zhe Hu, Jiahuan Zheng, Renwu Hua, Zhuo Hai, Jiaping Su, Jian V Zhang, William S B Yeung, Tianren Wang.
      Caseinolytic protease proteolytic subunit (ClpP) and caseinolytic protease X (ClpX) are mitochondrial matrix peptidases that activate mitochondrial unfolded protein response to maintain protein homeostasis in the mitochondria. However, the role of ClpP and ClpX in spermatogenesis remains largely unknown. In this study, we demonstrated the importance of ClpP/ClpX for meiosis and spermatogenesis with two conditional knockout (cKO) mouse models. We found that ClpP/ClpX deficiency reduced mitochondrial functions and quantity in spermatocytes, affected energy supply during meiosis and attenuated zygotene-pachytene transformation of the male germ cells. The dysregulated spermatocytes finally underwent apoptosis resulting in decreased testicular size and vacuolar structures within the seminiferous tubules. We found mTORC1 pathway was over-activated after deletion of ClpP/ClpX in spermatocytes. Long-term inhibition of the mTORC1 signaling via rapamycin treatment in vivo partially rescue spermatogenesis. The data reveal the critical roles of ClpP and ClpX in regulating meiosis and spermatogenesis.
    DOI:  https://doi.org/10.1038/s42003-023-05372-2
  7. J Int Adv Otol. 2023 Oct;19(5): 414-419
    Research Advances on Deafness Genes Associated with Mitochondrial tRNA-37 Modifications.
    Panpan Bian, Jing Chai, Baicheng Xu.
      As the most common cause of speech disorders, the etiological study of deafness is important for the diagnosis and treatment of deafness. The mitochondrial genome has gradually become a hotspot for deafness genetic research. Mitochondria are the core organelles of energy and material metabolism in eukaryotic cells. Human mitochondria contain 20 amino acids, except for tRNALeu and tRNASer, which have 2 iso-receptors, the other 18 amino acids correspond to unique tRNAs one by one, so mutations in any one tRNA may lead to protein translation defects in mitochondria and thus affect their oxidative phosphorylation process resulting in the corresponding disease phenotype. Mitochondrial tRNAs are extensively modified with base modifications that contribute to the correct folding of tRNAs and maintain their stability. Defective mitochondrial tRNA modifications are closely associated with the development of mitochondrial diseases. The in-depth study found that modification defects of mammalian mitochondrial tRNAs are associated with deafness, especially the nucleotide modification defect of mt-tRNA-37. This article reviews the research on mitochondrial tRNAs, nucleotide modification structure of mitochondrial tRNA-37, and nuclear genes related to modification defects to provide new ideas for the etiological study of deafness.
    DOI:  https://doi.org/10.5152/iao.2023.231107
  8. Cold Spring Harb Perspect Med. 2023 Oct 03. pii: a041199. [Epub ahead of print]
    Mitochondrial Targeted Interventions for Aging.
    Sophia Z Liu, Ying Ann Chiao, Peter S Rabinovitch, David J Marcinek.
      Changes in mitochondrial function play a critical role in the basic biology of aging and age-related disease. Mitochondria are typically thought of in the context of ATP production and oxidant production. However, it is clear that the mitochondria sit at a nexus of cell signaling where they affect metabolite, redox, and energy status, which influence many factors that contribute to the biology of aging, including stress responses, proteostasis, epigenetics, and inflammation. This has led to growing interest in identifying mitochondrial targeted interventions to delay or reverse age-related decline in function and promote healthy aging. In this review, we discuss the diverse roles of mitochondria in the cell. We then highlight some of the most promising strategies and compounds to target aging mitochondria in preclinical testing. Finally, we review the strategies and compounds that have advanced to clinical trials to test their ability to improve health in older adults.
    DOI:  https://doi.org/10.1101/cshperspect.a041199
  9. Basic Res Cardiol. 2023 Oct 05. 118(1): 42
    Mitophagy for cardioprotection.
    Allen Sam Titus, Eun-Ah Sung, Daniela Zablocki, Junichi Sadoshima.
      Mitochondrial function is maintained by several strictly coordinated mechanisms, collectively termed mitochondrial quality control mechanisms, including fusion and fission, degradation, and biogenesis. As the primary source of energy in cardiomyocytes, mitochondria are the central organelle for maintaining cardiac function. Since adult cardiomyocytes in humans rarely divide, the number of dysfunctional mitochondria cannot easily be diluted through cell division. Thus, efficient degradation of dysfunctional mitochondria is crucial to maintaining cellular function. Mitophagy, a mitochondria specific form of autophagy, is a major mechanism by which damaged or unnecessary mitochondria are targeted and eliminated. Mitophagy is active in cardiomyocytes at baseline and in response to stress, and plays an essential role in maintaining the quality of mitochondria in cardiomyocytes. Mitophagy is mediated through multiple mechanisms in the heart, and each of these mechanisms can partially compensate for the loss of another mechanism. However, insufficient levels of mitophagy eventually lead to mitochondrial dysfunction and the development of heart failure. In this review, we discuss the molecular mechanisms of mitophagy in the heart and the role of mitophagy in cardiac pathophysiology, with the focus on recent findings in the field.
    Keywords:  Alternative mitophagy; Drp1; Mitochondrial quality control; Mitophagy
    DOI:  https://doi.org/10.1007/s00395-023-01009-x
  10. Biophys J. 2023 Oct 05. pii: S0006-3495(23)00626-4. [Epub ahead of print]
    Mitochondrial Membrane Model: Lipids, Elastic Properties and the Changing Curvature of Cardiolipin.
    Sukanya Konar, Hina Arif, Christoph Allolio.
      Mammalian and Drosophila Melanogaster model mitochondrial membrane compositions are constructed from experimental data. Simplified compositions for inner and outer mitochondrial membranes are provided, including an asymmetric inner mitochondrial membrane. We performed atomistic molecular dynamics simulations of these membranes and computed their material properties. When comparing these properties to those obtained by extrapolation from their constituting lipids, we find good overall agreement. Finally, we analyzed the curvature effect of cardiolipin, considering ion concentration effects, oxidation and pH. We draw the conclusion that cardiolipin negative curvature is most likely due to counterion effects, such as cation adsorption, in particular of H3O+. This oft-neglected effect might account for the puzzling behavior of this lipid.
    DOI:  https://doi.org/10.1016/j.bpj.2023.10.002
  11. Nat Rev Mol Cell Biol. 2023 Oct 04.
    Base editing of organellar DNA with programmable deaminases.
    Jin-Soo Kim, Jia Chen.
      Mitochondria and chloroplasts are organelles that include their own genomes, which encode key genes for ATP production and carbon dioxide fixation, respectively. Mutations in mitochondrial DNA can cause diverse genetic disorders and are also linked to ageing and age-related diseases, including cancer. Targeted editing of organellar DNA should be useful for studying organellar genes and developing novel therapeutics, but it has been hindered by lack of efficient tools in living cells. Recently, CRISPR-free, protein-only base editors, such as double-stranded DNA deaminase toxin A-derived cytosine base editors (DdCBEs) and adenine base editors (ABEs), have been developed, which enable targeted organellar DNA editing in human cell lines, animals and plants. In this Review, we present programmable deaminases developed for base editing of organellar DNA in vitro and discuss mitochondrial DNA editing in animals, and plastid genome (plastome) editing in plants. We also discuss precision and efficiency limitations of these tools and propose improvements for therapeutic, agricultural and environmental applications.
    DOI:  https://doi.org/10.1038/s41580-023-00663-2
  12. BMC Res Notes. 2023 Sep 30. 16(1): 243
    Purification of functional mouse skeletal muscle mitochondria using percoll density gradient centrifugation.
    Rea Victoria P Anunciado-Koza, Anyonya R Guntur, Calvin P Vary, Carlos A Gartner, Madeleine Nowak, Robert A Koza.
       OBJECTIVE: Our goal was to isolate purified mitochondria from mouse skeletal muscle using a Percoll density gradient and to assess bioenergetic function and purity via Seahorse Extracellular Flux (XF) Analyses and mass spectrometry.
    RESULTS: Mitochondria isolated from murine quadriceps femoris skeletal muscle using a Percoll density gradient method allowed for minimally contaminated preparations with time from tissue harvest to mitochondrial isolation and quantification in about 3-4 h. Percoll purification from 100 to 200 mg fresh tissue yielded ~ 200-400 ug protein. Mitochondrial bioenergetics evaluated using the Seahorse XFe96 analyzer, a high-throughput respirometry platform, showed optimum mitochondrial input at 500 ng with respiratory control ratio ranging from 3.9 to 7.1 using various substrates demonstrating a high degree of functionality. Furthermore, proteomic analysis of Percoll-enriched mitochondria isolated from skeletal muscle using this method showed significant enrichment of mitochondrial proteins indicating high sample purity. This study established a methodology that ensures sufficient high quality mitochondria for downstream analyses such as mitochondrial bioenergetics and proteomics.
    Keywords:  Bioenergetics; Density gradient; Mitochondria; Percoll purification; Skeletal muscle
    DOI:  https://doi.org/10.1186/s13104-023-06519-4
  13. Biomark Res. 2023 Oct 05. 11(1): 89
    Mitochondrial transfer in hematological malignancies.
    Xiaodong Guo, Can Can, Wancheng Liu, Yihong Wei, Xinyu Yang, Jinting Liu, Hexiao Jia, Wenbo Jia, Hanyang Wu, Daoxin Ma.
      Mitochondria are energy-generated organelles and take an important part in biological metabolism. Mitochondria could be transferred between cells, which serves as a new intercellular communication. Mitochondrial transfer improves mitochondrial defects, restores the biological functions of recipient cells, and maintains the high metabolic requirements of tumor cells as well as drug resistance. In recent years, it has been reported mitochondrial transfer between cells of bone marrow microenvironment and hematological malignant cells play a critical role in the disease progression and resistance during chemotherapy. In this review, we discuss the patterns and mechanisms on mitochondrial transfer and their engagement in different pathophysiological contexts and outline the latest knowledge on intercellular transport of mitochondria in hematological malignancies. Besides, we briefly outline the drug resistance mechanisms caused by mitochondrial transfer in cells during chemotherapy. Our review demonstrates a theoretical basis for mitochondrial transfer as a prospective therapeutic target to increase the treatment efficiency in hematological malignancies and improve the prognosis of patients.
    Keywords:  Extracellular mitochondria; Extracellular vesicles; Hematological malignancies; Mitochondrial transfer; Tunneling nanotubes
    DOI:  https://doi.org/10.1186/s40364-023-00529-x
  14. Res Sq. 2023 Sep 20. pii: rs.3.rs-3311459. [Epub ahead of print]
    Caloric restriction promotes beta cell longevity and delays aging and senescence signatures.
    Rafael Arrojo E Drigo, Cristiane Dos Santos, Shristi Shrestha, Matthew Cottam, Guy Perkins, Varda Lev-Ram, Birbickram Roy, Christopher Acree, Keun-Young Kim, Thomas Deerinck, Melanie Cutler, Danielle Dean, Jean-Philippe Cartailler, Patrick MacDonald, Martin Hetzer, Mark Ellisman.
      Caloric restriction (CR) extends organismal lifespan and health span by improving glucose homeostasis mechanisms. How CR affects organellar structure and function of pancreatic beta cells over the lifetime of the animal remains unknown. Here, we used single nucleus transcriptomics to show that CR increases the expression of genes for beta cell identity, protein processing, and organelle homeostasis. Gene regulatory network analysis link this transcriptional phenotype to transcription factors involved in beta cell identity (Mafa) and homeostasis (Atf6). Imaging metabolomics further demonstrates that CR beta cells are more energetically competent. In fact, high-resolution light and electron microscopy indicates that CR reduces beta cell mitophagy and increases mitochondria mass, increasing mitochondrial ATP generation. Finally, we show that long-term CR delays the onset of beta cell aging and senescence to promote longevity by reducing beta cell turnover. Therefore, CR could be a feasible approach to preserve compromised beta cells during aging and diabetes.
    DOI:  https://doi.org/10.21203/rs.3.rs-3311459/v1
  15. Biochim Biophys Acta Mol Cell Res. 2023 Sep 26. pii: S0167-4889(23)00174-X. [Epub ahead of print]1871(1): 119601
    Knockdown of BRAWNIN minimally affect mitochondrial complex III assembly in human cells.
    Ya Wang, Yu Shi, Wen Li, Xinyu Han, Xi Lin, Dandan Liu, Yuyan Lin, Lijun Shen.
      BRAWNIN was found as a mitochondrial respiratory complex III (CIII) assembly factor. Here, we showed that the deletion rather than knockdown of BRAWNIN impaired the assembly of CIII. BRAWNIN levels were affected by nutritional stress and negatively associated with AMPK activation. Although the BRAWNIN knockout via CRISPR/Cas9 led to decreased complex III levels, both biochemical and functional studies of oxidative phosphorylation system (OXPHOS) complexes revealed that knockdown of BRAWNIN neither affected mitochondrial respiration nor impaired the integrity of OXPHOS complexes I-V. Transcriptomic and proteomic profiling further confirmed that the BRAWNIN knockdown had a minimal effect on mitochondrial function. Moreover, only a small proportion of BRAWNIN interacted with the subunits of the OXPHOS complexes, which might be difficult to detect via co-immunoprecipitation and mass spectrometry. Finally, our findings also indicated that although only a minimal amount of BRAWNIN was required for CIII assembly, metabolic analyses revealed that it may fine-tune the pyruvate metabolism route in mitochondria.
    DOI:  https://doi.org/10.1016/j.bbamcr.2023.119601
  16. Eur J Hum Genet. 2023 Oct 04.
    Mitochondrial Cytochrome C deficiency can show the first disease signs in the prenatal stage.
    Caterina Garone.
      
    DOI:  https://doi.org/10.1038/s41431-023-01466-x
  17. Amyotroph Lateral Scler Frontotemporal Degener. 2023 Oct 01. 1-15
    Integrated analysis of transcriptomic and proteomic alterations in mouse models of ALS/FTD identify early metabolic adaptions with similarities to mitochondrial dysfunction disorders.
    Anna Matveeva, Orla Watters, Ani Rukhadze, Niraj Khemka, Debora Gentile, Ivan Fernandez Perez, Irene Llorente-Folch, Cliona Farrell, Elide Lo Cacciato, Joshua Jackson, Antonia Piazzesi, Lena Wischhof, Ina Woods, Luise Halang, Marion Hogg, Amaya Garcia Muñoz, Eugène T Dillon, David Matallanas, Ingrid Arijs, Diether Lambrechts, Daniele Bano, Niamh M C Connolly, Jochen H M Prehn.
       OBJECTIVE: Sporadic and familial amyotrophic lateral sclerosis (ALS) is a fatal progressive neurodegenerative disease that results in loss of motor neurons and, in some patients, associates with frontotemporal dementia (FTD). Apart from the accumulation of proteinaceous deposits, emerging literature indicates that aberrant mitochondrial bioenergetics may contribute to the onset and progression of ALS/FTD. Here we sought to investigate the pathophysiological signatures of mitochondrial dysfunction associated with ALS/FTD.
    METHODS: By means of label-free mass spectrometry (MS) and mRNA sequencing (mRNA-seq), we report pre-symptomatic changes in the cortices of TDP-43 and FUS mutant mouse models. Using tissues from transgenic mouse models of mitochondrial diseases as a reference, we performed comparative analyses and extracted unique and common mitochondrial signatures that revealed neuroprotective compensatory mechanisms in response to early damage.
    RESULTS: In this regard, upregulation of both Acyl-CoA Synthetase Long-Chain Family Member 3 (ACSL3) and mitochondrial tyrosyl-tRNA synthetase 2 (YARS2) were the most representative change in pre-symptomatic ALS/FTD tissues, suggesting that fatty acid beta-oxidation and mitochondrial protein translation are mechanisms of adaptation in response to ALS/FTD pathology.
    CONCLUSIONS: Together, our unbiased integrative analyses unveil novel molecular components that may influence mitochondrial homeostasis in the earliest phase of ALS.
    Keywords:  Amyotrophic lateral sclerosis (ALS); bioinformatics; frontotemporal dementia (FTD); lipid metabolism; master regulator analysis; mitochondrial dysfunction
    DOI:  https://doi.org/10.1080/21678421.2023.2261979
  18. Methods Mol Biol. 2024 ;2719 79-98
    Implementation of Exome Sequencing to Identify Rare Genetic Diseases.
    Prajna Udupa, Debasish Kumar Ghosh.
      Modern high-throughput genomic testing using next-generation sequencing (NGS) has led to a significant increase in the successful diagnosis of rare genetic disorders. Recent advances in NGS tools and techniques have led to accurate and timely diagnosis of a large proportion of genetic diseases by finding sequence variations in clinical samples. One of the NGS techniques, exome sequencing (ES), is considered as a powerful and easily approachable method for genetic disorders in terms of rapid and cost-effective diagnostic yields. In this chapter, we describe an overview of whole exome sequencing (ES) in the context of experimental and analytical methodologies. Approaches to ES include sequencing capture technique, quality control processes at various stages of sequencing analysis, exome data filtering strategy that incorporates both primary and secondary filtering, and prioritization of candidate variants in diagnosing genetic diseases.
    Keywords:  Capture kit; Exome sequencing; In-silico analysis; Library preparation; Variant annotation
    DOI:  https://doi.org/10.1007/978-1-0716-3461-5_5
  19. bioRxiv. 2023 Sep 22. pii: 2023.09.21.558912. [Epub ahead of print]
    Inter-organellar nucleic acid communication by a mitochondrial tRNA regulates nuclear metabolic transcription.
    Christopher Rouya, King Faisal Yambire, Mark L Derbyshire, Hanan Alwaseem, Sohail F Tavazoie.
      Efficient communication between mitochondria and the nucleus underlies homoeostatic metabolic control, though the involved mitochondrial factors and their mechanisms are poorly defined. Here, we report the surprising detection of multiple mitochondrial-derived transfer RNAs (mito-tRNAs) within the nuclei of human cells. Focused studies of nuclear-transported mito-tRNA-asparagine (mtAsn) revealed that its cognate charging enzyme (NARS2) is also present in the nucleus. MtAsn promoted interaction of NARS2 with histone deacetylase 2 (HDAC2), and repressed HDAC2 association with specific chromatin loci. Perturbation of this axis using antisense oligonucleotides promoted nucleotide biogenesis and enhanced breast cancer growth, and RNA and nascent transcript sequencing demonstrated specific alterations in the transcription of nuclear genes. These findings uncover nucleic-acid mediated communication between two organelles and the existence of a machinery for nuclear gene regulation by a mito-tRNA that restricts tumor growth through metabolic control.
    Highlights: Multiple mitochondrial-derived tRNAs are detected in human cell nucleiMtAsn promotes binding between NARS2 and HDAC2Metabolic alterations driven by mtAsn impact cell proliferationMtAsn inhibition releases HDAC2 to bind and transcriptionally regulate multiple nuclear genes.
    DOI:  https://doi.org/10.1101/2023.09.21.558912
  20. Free Radic Biol Med. 2023 Sep 25. pii: S0891-5849(23)00654-8. [Epub ahead of print]208 771-779
    The glyoxylate shunt protein ICL-1 protects from mitochondrial superoxide stress through activation of the mitochondrial unfolded protein response.
    Guoqiang Wang, Ricardo Laranjeiro, Stephanie LeValley, Jeremy M Van Raamsdonk, Monica Driscoll.
      Disrupting mitochondrial superoxide dismutase (SOD) causes neonatal lethality in mice and death of flies within 24 h after eclosion. Deletion of mitochondrial sod genes in C. elegans impairs fertility as well, but surprisingly is not detrimental to survival of progeny generated. The comparison of metabolic pathways among mouse, flies and nematodes reveals that mice and flies lack the glyoxylate shunt, a shortcut that bypasses part of the tricarboxylic acid (TCA) cycle. Here we show that ICL-1, the sole protein that catalyzes the glyoxylate shunt, is critical for protection against embryonic lethality resulting from elevated levels of mitochondrial superoxide. In exploring the mechanism by which ICL-1 protects against ROS-mediated embryonic lethality, we find that ICL-1 is required for the efficient activation of mitochondrial unfolded protein response (UPRmt) and that ATFS-1, a key UPRmt transcription factor and an activator of icl-1 gene expression, is essential to limit embryonic/neonatal lethality in animals lacking mitochondrial SOD. In sum, we identify a biochemical pathway that highlights a molecular strategy for combating toxic mitochondrial superoxide consequences in cells.
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2023.09.029
  21. Front Physiol. 2023 ;14 1200119
    Lithium isotopes differentially modify mitochondrial amorphous calcium phosphate cluster size distribution and calcium capacity.
    Marshall L Deline, Joshua Straub, Manisha Patel, Pratigya Subba, Martin Grashei, Frits H A van Heijster, Philip Pirkwieser, Veronika Somoza, James D Livingstone, Michael Beazely, Brian Kendall, Michel J P Gingras, Zoya Leonenko, Carmen Höschen, Gertraud Harrington, Katharina Kuellmer, Wangqing Bian, Franz Schilling, Matthew P A Fisher, Matthew E Helgeson, Tobias Fromme.
      Lithium is commonly prescribed as a mood stabilizer in a variety of mental health conditions, yet its molecular mode of action is incompletely understood. Many cellular events associated with lithium appear tied to mitochondrial function. Further, recent evidence suggests that lithium bioactivities are isotope specific. Here we focus on lithium effects related to mitochondrial calcium handling. Lithium protected against calcium-induced permeability transition and decreased the calcium capacity of liver mitochondria at a clinically relevant concentration. In contrast, brain mitochondrial calcium capacity was increased by lithium. Surprisingly, 7Li acted more potently than 6Li on calcium capacity, yet 6Li was more effective at delaying permeability transition. The size distribution of amorphous calcium phosphate colloids formed in vitro was differentially affected by lithium isotopes, providing a mechanistic basis for the observed isotope specific effects on mitochondrial calcium handling. This work highlights a need to better understand how mitochondrial calcium stores are structurally regulated and provides key considerations for future formulations of lithium-based therapeutics.
    Keywords:  amorphous calcium phosphate; calcium; isotope distribution; lithium; lithium bioactivity; mitochondria; mitochondrial calcium
    DOI:  https://doi.org/10.3389/fphys.2023.1200119
  22. PLoS One. 2023 ;18(10): e0285242
    Long term survival and abnormal liver fat accumulation in mice with specific thymidine kinase 2 deficiency in liver tissue.
    Qian Zhao, Xiaoshan Zhou, Jingyi Yan, Raoul Kuiper, Sophie Curbo, Anna Karlsson.
      Deficiency in thymidine kinase 2 (TK2) causes mitochondrial DNA depletion. Liver mitochondria are severely affected in Tk2 complete knockout models and have been suggested to play a role in the pathogenesis of the Tk2 knockout phenotype, characterized by loss of hypodermal fat tissue, growth retardation and reduced life span. Here we report a liver specific Tk2 knockout (KO) model to further study mechanisms contributing to the phenotypic changes associated with Tk2 deficiency. Interestingly, the liver specific Tk2 KO mice had a normal life span despite a much lower mtDNA level in liver tissue. Mitochondrial DNA encoded peptide COXI did not differ between the Tk2 KO and control mice. However, the relative liver weight was significantly increased in the male Tk2 KO mouse model. Histology analysis indicated an increased lipid accumulation. We conclude that other enzyme activities can partly compensate Tk2 deficiency to maintain mtDNA at a low but stable level throughout the life span of the liver specific Tk2 KO mice. The lower level of mtDNA was sufficient for survival but led to an abnormal lipid accumulation in liver tissue.
    DOI:  https://doi.org/10.1371/journal.pone.0285242
  23. PLoS Biol. 2023 Oct;21(10): e3002313
    Significant decrease of maternal mitochondria carryover using optimized spindle-chromosomal complex transfer.
    Xiaoyu Liao, Wenzhi Li, Kaibo Lin, Wei Jin, Shaozhen Zhang, Yao Wang, Meng Ma, Yating Xie, Weina Yu, Zhiguang Yan, Hongyuan Gao, Leiwen Zhao, Jiqiang Si, Yun Wang, Jiaying Lin, Chen Chen, Li Chen, Yanping Kuang, Qifeng Lyu.
      Mutations in mitochondrial DNA (mtDNA) contribute to a variety of serious multi-organ human diseases, which are strictly inherited from the maternal germline. However, there is currently no curative treatment. Attention has been focused on preventing the transmission of mitochondrial diseases through mitochondrial replacement (MR) therapy, but levels of mutant mtDNA can often unexpectedly undergo significant changes known as mitochondrial genetic drift. Here, we proposed a novel strategy to perform spindle-chromosomal complex transfer (SCCT) with maximal residue removal (MRR) in metaphase II (MII) oocytes, thus hopefully eliminated the transmission of mtDNA diseases. With the MRR procedure, we initially investigated the proportions of mtDNA copy numbers in isolated karyoplasts to those of individual oocytes. Spindle-chromosomal morphology and copy number variation (CNV) analysis also confirmed the safety of this method. Then, we reconstructed oocytes by MRR-SCCT, which well developed to blastocysts with minimal mtDNA residue and normal chromosomal copy numbers. Meanwhile, we optimized the manipulation order between intracytoplasmic sperm injection (ICSI) and SCC transfer and concluded that ICSI-then-transfer was conducive to avoid premature activation of reconstructed oocytes in favor of normal fertilization. Offspring of mice generated by embryos transplantation in vivo and embryonic stem cells derivation further presented evidences for competitive development competence and stable mtDNA carryover without genetic drift. Importantly, we also successfully accomplished SCCT in human MII oocytes resulting in tiny mtDNA residue and excellent embryo development through MRR manipulation. Taken together, our preclinical mouse and human models of the MRR-SCCT strategy not only demonstrated efficient residue removal but also high compatibility with normal embryo development, thus could potentially be served as a feasible clinical treatment to prevent the transmission of inherited mtDNA diseases.
    DOI:  https://doi.org/10.1371/journal.pbio.3002313
  24. Nucleic Acids Res. 2023 Oct 02. pii: gkad713. [Epub ahead of print]
    Cleavage kinetics of human mitochondrial RNase P and contribution of its non-nuclease subunits.
    Elisa Vilardo, Ursula Toth, Enxhi Hazisllari, Roland K Hartmann, Walter Rossmanith.
      RNase P is the endonuclease responsible for the 5' processing of precursor tRNAs (pre-tRNAs). Unlike the single-subunit protein-only RNase P (PRORP) found in plants or protists, human mitochondrial RNase P is a multi-enzyme assembly that in addition to the homologous PRORP subunit comprises a methyltransferase (TRMT10C) and a dehydrogenase (SDR5C1) subunit; these proteins, but not their enzymatic activities, are required for efficient pre-tRNA cleavage. Here we report a kinetic analysis of the cleavage reaction by human PRORP and its interplay with TRMT10C-SDR5C1 including 12 different mitochondrial pre-tRNAs. Surprisingly, we found that PRORP alone binds pre-tRNAs with nanomolar affinity and can even cleave some of them at reduced efficiency without the other subunits. Thus, the ancient binding mode, involving the tRNA elbow and PRORP's PPR domain, appears basically retained by human PRORP, and its metallonuclease domain is in principle correctly folded and functional. Our findings support a model according to which the main function of TRMT10C-SDR5C1 is to direct PRORP's nuclease domain to the cleavage site, thereby increasing the rate and accuracy of cleavage. This functional dependence of human PRORP on an extra tRNA-binding protein complex likely reflects an evolutionary adaptation to the erosion of canonical structural features in mitochondrial tRNAs.
    DOI:  https://doi.org/10.1093/nar/gkad713
  25. Nat Commun. 2023 09 30. 14(1): 6113
    Deleterious heteroplasmic mitochondrial mutations are associated with an increased risk of overall and cancer-specific mortality.
    Yun Soo Hong, Stephanie L Battle, Wen Shi, Daniela Puiu, Vamsee Pillalamarri, Jiaqi Xie, Nathan Pankratz, Nicole J Lake, Monkol Lek, Jerome I Rotter, Stephen S Rich, Charles Kooperberg, Alex P Reiner, Paul L Auer, Nancy Heard-Costa, Chunyu Liu, Meng Lai, Joanne M Murabito, Daniel Levy, Megan L Grove, Alvaro Alonso, Richard Gibbs, Shannon Dugan-Perez, Lukasz P Gondek, Eliseo Guallar, Dan E Arking.
      Mitochondria carry their own circular genome and disruption of the mitochondrial genome is associated with various aging-related diseases. Unlike the nuclear genome, mitochondrial DNA (mtDNA) can be present at 1000 s to 10,000 s copies in somatic cells and variants may exist in a state of heteroplasmy, where only a fraction of the DNA molecules harbors a particular variant. We quantify mtDNA heteroplasmy in 194,871 participants in the UK Biobank and find that heteroplasmy is associated with a 1.5-fold increased risk of all-cause mortality. Additionally, we functionally characterize mtDNA single nucleotide variants (SNVs) using a constraint-based score, mitochondrial local constraint score sum (MSS) and find it associated with all-cause mortality, and with the prevalence and incidence of cancer and cancer-related mortality, particularly leukemia. These results indicate that mitochondria may have a functional role in certain cancers, and mitochondrial heteroplasmic SNVs may serve as a prognostic marker for cancer, especially for leukemia.
    DOI:  https://doi.org/10.1038/s41467-023-41785-7
  26. Mitochondrion. 2023 Oct 02. pii: S1567-7249(23)00084-3. [Epub ahead of print]
    Complete mitochondrial DNA profile in stroke: a geographical matched case-control study in Spanish population.
    Ana Onieva, Joan Martin, Daniel R Cuesta-Aguirre, Violeta Planells, Marta Coronado-Zamora, Katrin Beyer, Tomas Vega, Jose Eugenio Lozano, Cristina Santos, M Pilar Aluja.
       INTRODUCTION: Stroke, the second leading cause of death worldwide, is a complex disease influenced by many risk factors among which we can find reactive oxygen species (ROS). Since mitochondria are the main producers of cellular ROS, nowadays studies are trying to elucidate the role of these organelles and its DNA (mtDNA) variation in stroke risk. The aim of the present study was to perform a comprehensive evaluation of the association between mtDNA mutations and mtDNA content and stroke risk.
    MATERIAL AND METHODS: Homoplasmic and heteroplasmic mutations of the mtDNA were analysed in a case-controls study using 110 stroke cases and their corresponding control individuals. Mitochondrial DNA copy number (mtDNA-CN) was analysed in 73 of those case-control pairs.
    RESULTS: Our results suggest that haplogroup V, specifically variants m.72C>T, m.4580G>A, m.15904C>T and m.16298T>C have a protective role in relation to stroke risk. On the contrary, variants m.73A>G, m.11719G>A and m.14766C>T appear to be genetic risk factors for stroke. In this study, we found no statistically significant association between stroke risk and mitochondrial DNA copy number.
    CONCLUSIONS: These results demonstrate the possible role of mtDNA genetics on the pathogenesis of stroke, probably through alterations in mitochondrial ROS production.
    Keywords:  Mitochondrial DNA; Mitochondrial DNA copy number; Mitochondrial haplogroups; Stroke
    DOI:  https://doi.org/10.1016/j.mito.2023.10.001
  27. ACS Cent Sci. 2023 Sep 27. 9(9): 1799-1809
    Single-Nucleotide Resolution Mapping of N6-Methyladenine in Genomic DNA.
    Cheng-Jie Ma, Gaojie Li, Wen-Xuan Shao, Yi-Hao Min, Ping Wang, Jiang-Hui Ding, Neng-Bin Xie, Min Wang, Feng Tang, Yu-Qi Feng, Weimin Ci, Yinsheng Wang, Bi-Feng Yuan.
      N6-Methyladenine (6mA) is a naturally occurring DNA modification in both prokaryotes and eukaryotes. Herein, we developed a deaminase-mediated sequencing (DM-seq) method for genome-wide mapping of 6mA at single-nucleotide resolution. The method capitalizes on the selective deamination of adenine, but not 6mA, in DNA mediated by an evolved adenine deaminase, ABE8e. By employing this method, we achieved genome-wide mapping of 6mA in Escherichia coli and in mammalian mitochondrial DNA (mtDNA) at single-nucleotide resolution. We found that the 6mA sites are mainly located in the GATC motif in the E. coli genome. We also identified 17 6mA sites in mtDNA of HepG2 cells, where all of the 6mA sites are distributed in the heavy strand of mtDNA. We envision that DM-seq will be a valuable tool for uncovering new functions of 6mA in DNA and for exploring its potential roles in mitochondria-related human diseases.
    DOI:  https://doi.org/10.1021/acscentsci.3c00481
  28. J Vis Exp. 2023 09 15.
    Complementary Approaches to Interrogate Mitophagy Flux in Pancreatic β-Cells.
    Elena Levi-D'Ancona, Vaibhav Sidarala, Scott A Soleimanpour.
      Mitophagy is a quality control mechanism necessary to maintain optimal mitochondrial function. Dysfunctional β-cell mitophagy results in insufficient insulin release. Advanced quantitative assessments of mitophagy often require the use of genetic reporters. The mt-Keima mouse model, which expresses a mitochondria-targeted pH-sensitive dual-excitation ratiometric probe for quantifying mitophagy via flow cytometry, has been optimized in β-cells. The ratio of acidic-to-neutral mt-Keima wavelength emissions can be used to robustly quantify mitophagy. However, using genetic mitophagy reporters can be challenging when working with complex genetic mouse models or difficult-to-transfect cells, such as primary human islets. This protocol describes a novel complementary dye-based method to quantify β-cell mitophagy in primary islets using MtPhagy. MtPhagy is a pH-sensitive, cell-permeable dye that accumulates in the mitochondria and increases its fluorescence intensity when mitochondria are in low pH environments, such as lysosomes during mitophagy. By combining the MtPhagy dye with Fluozin-3-AM, a Zn2+ indicator that selects for β-cells, and Tetramethylrhodamine, ethyl ester (TMRE) to assess mitochondrial membrane potential, mitophagy flux can be quantified specifically in β-cells via flow cytometry. These two approaches are highly complementary, allowing for flexibility and precision in assessing mitochondrial quality control in numerous β-cell models.
    DOI:  https://doi.org/10.3791/65789
  29. Genet Med. 2023 Oct 03. pii: S1098-3600(23)01008-0. [Epub ahead of print] 100992
    Gene-specific ACMG/AMP classification criteria for germline APC variants: recommendations from the ClinGen InSiGHT Hereditary Colorectal Cancer / Polyposis Variant Curation Expert Panel.
    InSiGHT - ClinGen Hereditary Colon Cancer / Polyposis Variant Curation Expert Panel
       PURPOSE: The Hereditary Colorectal Cancer/Polyposis Variant Curation Expert Panel (VCEP) was established by the International Society for Gastrointestinal Hereditary Tumours (InSiGHT) and the Clinical Genome Resource (ClinGen), who set out to develop recommendations for the interpretation of germline APC variants underlying Familial Adenomatous Polyposis (FAP), the most frequent hereditary polyposis syndrome.
    METHODS: Through a rigorous process of database analysis, literature review, and expert elicitation, the APC VCEP derived gene-specific modifications to the ACMG/AMP (American College of Medical Genetics and Genomics and Association for Molecular Pathology) variant classification guidelines and validated such criteria through the pilot classification of 58 variants.
    RESULTS: The APC-specific criteria represented gene- and disease-informed specifications, including a quantitative approach to allele frequency thresholds, a stepwise decision tool for truncating variants, and semiquantitative evaluations of experimental and clinical data. Using the APC-specific criteria, 47% (27/58) of pilot variants were reclassified including 14 previous variants of uncertain significance (VUS).
    CONCLUSIONS: The APC-specific ACMG/AMP criteria preserved the classification of well-characterised variants on ClinVar while substantially reducing the number of VUS by 56% (14/25). Moving forward, the APC VCEP will continue to interpret prioritised lists of VUS, the results of which will represent the most authoritative variant classification for widespread clinical use.
    Keywords:  ACMG/AMP variant classification guidelines; APC, ClinGen; Familial adenomatous polyposis; InSiGHT; variant interpretation
    DOI:  https://doi.org/10.1016/j.gim.2023.100992
  30. Biophys Chem. 2023 Sep 25. pii: S0301-4622(23)00164-3. [Epub ahead of print]303 107113
    Conformational analysis of membrane-proximal segments of GDAP1 in a lipidic environment using synchrotron radiation suggests a mode of assembly at the mitochondrial outer membrane.
    Aleksi Sutinen, Nykola C Jones, Søren Vrønning Hoffmann, Salla Ruskamo, Petri Kursula.
      The mitochondrial outer membrane creates a diffusion barrier between the cytosol and the mitochondrial intermembrane space, allowing the exchange of metabolic products, important for efficient mitochondrial function in neurons. The ganglioside-induced differentiation-associated protein 1 (GDAP1) is a mitochondrial outer membrane protein with a critical role in mitochondrial dynamics and metabolic balance in neurons. Missense mutations in the GDAP1 gene are linked to the most common human peripheral neuropathy, Charcot-Marie-Tooth disease (CMT). GDAP1 is a distant member of the glutathione-S-transferase (GST) superfamily, with unknown enzymatic properties or functions at the molecular level. The structure of the cytosol-facing GST-like domain has been described, but there is no consensus on how the protein interacts with the mitochondrial outer membrane. Here, we describe a model for GDAP1 assembly on the membrane using peptides vicinal to the GDAP1 transmembrane domain. We used oriented circular dichroism spectroscopy (OCD) with synchrotron radiation to study the secondary structure and orientation of GDAP1 segments at the outer and inner surfaces of the outer mitochondrial membrane. These experiments were complemented by small-angle X-ray scattering, providing the first experimental structural models for full-length human GDAP1. The results indicate that GDAP1 is bound into the membrane via a single transmembrane helix, flanked by two peripheral helices interacting with the outer and inner leaflets of the mitochondrial outer membrane in different orientations. Impairment of these interactions could be a mechanism for CMT in the case of missense mutations affecting these segments instead of the GST-like domain.
    Keywords:  Charcot-Marie-Tooth disease; GDAP1; Mitochondrial outer membrane; Protein biophysics
    DOI:  https://doi.org/10.1016/j.bpc.2023.107113
  31. bioRxiv. 2023 Sep 21. pii: 2023.09.21.558867. [Epub ahead of print]
    Light-sensitive phosphorylation regulates enzyme activity and filament assembly of human IMPDH1 retinal splice variants.
    S John Calise, Audrey G O'Neill, Anika L Burrell, Miles S Dickinson, Josephine Molfino, Charlie Clarke, Joel Quispe, David Sokolov, Rubén M Buey, Justin M Kollman.
      Inosine monophosphate dehydrogenase (IMPDH) is the rate-limiting enzyme in de novo guanosine triphosphate (GTP) synthesis and is controlled by feedback inhibition and allosteric regulation. IMPDH assembles into micron-scale filaments in cells, which desensitizes the enzyme to feedback inhibition by GTP and boosts nucleotide production. The vertebrate retina expresses two tissue-specific splice variants IMPDH1(546) and IMPDH1(595). IMPDH1(546) filaments adopt high and low activity conformations, while IMPDH1(595) filaments maintain high activity. In bovine retinas, residue S477 is preferentially phosphorylated in the dark, but the effects on IMPDH1 activity and regulation are unclear. Here, we generated phosphomimetic mutants to investigate structural and functional consequences of phosphorylation in IMPDH1 variants. The S477D mutation re-sensitized both variants to GTP inhibition, but only blocked assembly of IMPDH1(595) filaments and not IMPDH1(546) filaments. Cryo-EM structures of both variants showed that S477D specifically blocks assembly of the high activity assembly interface, still allowing assembly of low activity IMPDH1(546) filaments. Finally, we discovered that S477D exerts a dominant-negative effect in cells, preventing endogenous IMPDH filament assembly. By modulating the structure and higher-order assembly of IMPDH, phosphorylation at S477 acts as a mechanism for downregulating retinal GTP synthesis in the dark, when nucleotide turnover is decreased. Like IMPDH1, many other metabolic enzymes dynamically assemble filamentous polymers that allosterically regulate activity. Our work suggests that posttranslational modifications may be yet another layer of regulatory control to finely tune activity by modulating filament assembly in response to changing metabolic demands.
    SIGNIFICANCE STATEMENT: Over 20 different metabolic enzymes form micron-scale filaments in cells, suggesting that filament assembly is a conserved mechanism for regulating diverse metabolic pathways. Filament assembly regulates catalytic activity of many of these enzymes, including inosine monophosphate dehydrogenase (IMPDH), the rate-limiting enzyme in de novo GTP biosynthesis. The vertebrate retina expresses two IMPDH1 splice variants that are critical for maintaining nucleotide levels required for phototransduction. Here, we show that filament assembly by these variants is itself controlled by phosphorylation at a single residue, adding further complexity to the tight regulation of nucleotide metabolism in the retina. Phosphorylation and other posttranslational modifications are likely to be a general regulatory mechanism controlling filament assembly by enzymes in many different metabolic pathways.
    DOI:  https://doi.org/10.1101/2023.09.21.558867
  32. Nat Methods. 2023 Oct;20(10): 1530-1536
    Spatial single-cell mass spectrometry defines zonation of the hepatocyte proteome.
    Florian A Rosenberger, Marvin Thielert, Maximilian T Strauss, Lisa Schweizer, Constantin Ammar, Sophia C Mädler, Andreas Metousis, Patricia Skowronek, Maria Wahle, Katherine Madden, Janine Gote-Schniering, Anna Semenova, Herbert B Schiller, Edwin Rodriguez, Thierry M Nordmann, Andreas Mund, Matthias Mann.
      Single-cell proteomics by mass spectrometry is emerging as a powerful and unbiased method for the characterization of biological heterogeneity. So far, it has been limited to cultured cells, whereas an expansion of the method to complex tissues would greatly enhance biological insights. Here we describe single-cell Deep Visual Proteomics (scDVP), a technology that integrates high-content imaging, laser microdissection and multiplexed mass spectrometry. scDVP resolves the context-dependent, spatial proteome of murine hepatocytes at a current depth of 1,700 proteins from a cell slice. Half of the proteome was differentially regulated in a spatial manner, with protein levels changing dramatically in proximity to the central vein. We applied machine learning to proteome classes and images, which subsequently inferred the spatial proteome from imaging data alone. scDVP is applicable to healthy and diseased tissues and complements other spatial proteomics and spatial omics technologies.
    DOI:  https://doi.org/10.1038/s41592-023-02007-6
  33. bioRxiv. 2023 Sep 20. pii: 2023.09.20.558501. [Epub ahead of print]
    Alternate splice variants of the mitochondrial fission protein DNM1L/Drp1 regulate mitochondrial dynamics and cell fate in ovarian cancer.
    Zaineb Javed, Dong Hui Shin, Weihua Pan, Sierra R White, Yeon Soo Kim, Amal Taher Elhaw, Shriya Kamlapurkar, Ya-Yun Cheng, J Cory Benson, Ahmed Emam Abdelnaby, Rebecca Phaeton, Hong-Gang Wang, Shengyu Yang, Mara Lisa Grove Sullivan, Simon C Watkins, Steven J Mullett, Stacy Lynn Gelhaus, Nam Y Lee, Lan Coffman, Katherine Marie Aird, Mohamed Trebak, Mythreye Karthikeyan, Vonn Walter, Nadine Hempel.
      Aberrant mitochondrial fission/fusion dynamics have previously been reported in cancer cells. While post translational modifications are known regulators of GTPases of the mitochondrial fission/fusion machinery, we show for the first time that alternate splice variants of the fission protein Drp1 (DNM1L) have specific and unique roles in ovarian cancer, adding to the complexity of mitochondrial fission/fusion regulation in tumor cells. We find that ovarian cancer specimens express a Drp1 alternate splice transcript variant lacking exon 16 of the variable domain. High expression of Drp1 lacking exon 16 relative to other transcripts is associated with poor patient outcome. Unlike the unspliced variant, expression of Drp1 lacking exon 16 leads to decreased association of Drp1 to mitochondrial fission sites, more fused mitochondrial networks, enhanced respiration and TCA cycle metabolites, and is associated with a more tumorigenic phenotype. These effects can also be reversed by specific siRNA-mediated inhibition of the endogenously expressed transcript lacking exon 16. Moreover, lack of exon 16 abrogates mitochondrial fission in response to pro-apoptotic stimuli and leads to decreased sensitivity to chemotherapeutics. These data emphasize the significance of the pathophysiological consequences of Drp1 alternate splicing and divergent functions of Drp1 splice variants, and strongly warrant consideration of Drp1 splicing in future studies.
    DOI:  https://doi.org/10.1101/2023.09.20.558501
  34. Nat Med. 2023 Oct 05.
    Second international consensus report on gaps and opportunities for the clinical translation of precision diabetes medicine.
    Deirdre K Tobias, Jordi Merino, Abrar Ahmad, Catherine Aiken, Jamie L Benham, Dhanasekaran Bodhini, Amy L Clark, Kevin Colclough, Rosa Corcoy, Sara J Cromer, Daisy Duan, Jamie L Felton, Ellen C Francis, Pieter Gillard, Véronique Gingras, Romy Gaillard, Eram Haider, Alice Hughes, Jennifer M Ikle, Laura M Jacobsen, Anna R Kahkoska, Jarno L T Kettunen, Raymond J Kreienkamp, Lee-Ling Lim, Jonna M E Männistö, Robert Massey, Niamh-Maire Mclennan, Rachel G Miller, Mario Luca Morieri, Jasper Most, Rochelle N Naylor, Bige Ozkan, Kashyap Amratlal Patel, Scott J Pilla, Katsiaryna Prystupa, Sridharan Raghavan, Mary R Rooney, Martin Schön, Zhila Semnani-Azad, Magdalena Sevilla-Gonzalez, Pernille Svalastoga, Wubet Worku Takele, Claudia Ha-Ting Tam, Anne Cathrine B Thuesen, Mustafa Tosur, Amelia S Wallace, Caroline C Wang, Jessie J Wong, Jennifer M Yamamoto, Katherine Young, Chloé Amouyal, Mette K Andersen, Maxine P Bonham, Mingling Chen, Feifei Cheng, Tinashe Chikowore, Sian C Chivers, Christoffer Clemmensen, Dana Dabelea, Adem Y Dawed, Aaron J Deutsch, Laura T Dickens, Linda A DiMeglio, Monika Dudenhöffer-Pfeifer, Carmella Evans-Molina, María Mercè Fernández-Balsells, Hugo Fitipaldi, Stephanie L Fitzpatrick, Stephen E Gitelman, Mark O Goodarzi, Jessica A Grieger, Marta Guasch-Ferré, Nahal Habibi, Torben Hansen, Chuiguo Huang, Arianna Harris-Kawano, Heba M Ismail, Benjamin Hoag, Randi K Johnson, Angus G Jones, Robert W Koivula, Aaron Leong, Gloria K W Leung, Ingrid M Libman, Kai Liu, S Alice Long, William L Lowe, Robert W Morton, Ayesha A Motala, Suna Onengut-Gumuscu, James S Pankow, Maleesa Pathirana, Sofia Pazmino, Dianna Perez, John R Petrie, Camille E Powe, Alejandra Quinteros, Rashmi Jain, Debashree Ray, Mathias Ried-Larsen, Zeb Saeed, Vanessa Santhakumar, Sarah Kanbour, Sudipa Sarkar, Gabriela S F Monaco, Denise M Scholtens, Elizabeth Selvin, Wayne Huey-Herng Sheu, Cate Speake, Maggie A Stanislawski, Nele Steenackers, Andrea K Steck, Norbert Stefan, Julie Støy, Rachael Taylor, Sok Cin Tye, Gebresilasea Gendisha Ukke, Marzhan Urazbayeva, Bart Van der Schueren, Camille Vatier, John M Wentworth, Wesley Hannah, Sara L White, Gechang Yu, Yingchai Zhang, Shao J Zhou, Jacques Beltrand, Michel Polak, Ingvild Aukrust, Elisa de Franco, Sarah E Flanagan, Kristin A Maloney, Andrew McGovern, Janne Molnes, Mariam Nakabuye, Pål Rasmus Njølstad, Hugo Pomares-Millan, Michele Provenzano, Cécile Saint-Martin, Cuilin Zhang, Yeyi Zhu, Sungyoung Auh, Russell de Souza, Andrea J Fawcett, Chandra Gruber, Eskedar Getie Mekonnen, Emily Mixter, Diana Sherifali, Robert H Eckel, John J Nolan, Louis H Philipson, Rebecca J Brown, Liana K Billings, Kristen Boyle, Tina Costacou, John M Dennis, Jose C Florez, Anna L Gloyn, Maria F Gomez, Peter A Gottlieb, Siri Atma W Greeley, Kurt Griffin, Andrew T Hattersley, Irl B Hirsch, Marie-France Hivert, Korey K Hood, Jami L Josefson, Soo Heon Kwak, Lori M Laffel, Siew S Lim, Ruth J F Loos, Ronald C W Ma, Chantal Mathieu, Nestoras Mathioudakis, James B Meigs, Shivani Misra, Viswanathan Mohan, Rinki Murphy, Richard Oram, Katharine R Owen, Susan E Ozanne, Ewan R Pearson, Wei Perng, Toni I Pollin, Rodica Pop-Busui, Richard E Pratley, Leanne M Redman, Maria J Redondo, Rebecca M Reynolds, Robert K Semple, Jennifer L Sherr, Emily K Sims, Arianne Sweeting, Tiinamaija Tuomi, Miriam S Udler, Kimberly K Vesco, Tina Vilsbøll, Robert Wagner, Stephen S Rich, Paul W Franks.
      Precision medicine is part of the logical evolution of contemporary evidence-based medicine that seeks to reduce errors and optimize outcomes when making medical decisions and health recommendations. Diabetes affects hundreds of millions of people worldwide, many of whom will develop life-threatening complications and die prematurely. Precision medicine can potentially address this enormous problem by accounting for heterogeneity in the etiology, clinical presentation and pathogenesis of common forms of diabetes and risks of complications. This second international consensus report on precision diabetes medicine summarizes the findings from a systematic evidence review across the key pillars of precision medicine (prevention, diagnosis, treatment, prognosis) in four recognized forms of diabetes (monogenic, gestational, type 1, type 2). These reviews address key questions about the translation of precision medicine research into practice. Although not complete, owing to the vast literature on this topic, they revealed opportunities for the immediate or near-term clinical implementation of precision diabetes medicine; furthermore, we expose important gaps in knowledge, focusing on the need to obtain new clinically relevant evidence. Gaps include the need for common standards for clinical readiness, including consideration of cost-effectiveness, health equity, predictive accuracy, liability and accessibility. Key milestones are outlined for the broad clinical implementation of precision diabetes medicine.
    DOI:  https://doi.org/10.1038/s41591-023-02502-5
  35. Biomed Pharmacother. 2023 Oct 04. pii: S0753-3322(23)01450-6. [Epub ahead of print]167 115652
    Mitochondrial dysfunction: A fatal blow in depression.
    Yu Song, Huan Cao, Chengchao Zuo, Zhongya Gu, Yaqi Huang, Jinfeng Miao, Yufeng Fu, Yu Guo, Yongsheng Jiang, Furong Wang.
      Mitochondria maintain the normal physiological function of nerve cells by producing sufficient cellular energy and performing crucial roles in maintaining the metabolic balance through intracellular Ca2+ homeostasis, oxidative stress, and axonal development. Depression is a prevalent psychiatric disorder with an unclear pathophysiology. Damage to the hippocampal neurons is a key component of the plasticity regulation of synapses and plays a critical role in the mechanism of depression. There is evidence suggesting that mitochondrial dysfunction is associated with synaptic impairment. The maintenance of mitochondrial homeostasis includes quantitative maintenance and quality control of mitochondria. Mitochondrial biogenesis produces new and healthy mitochondria, and mitochondrial dynamics cooperates with mitophagy to remove damaged mitochondria. These processes maintain mitochondrial population stability and exert neuroprotective effects against early depression. In contrast, mitochondrial dysfunction is observed in various brain regions of patients with major depressive disorders. The accumulation of defective mitochondria accelerates cellular nerve dysfunction. In addition, impaired mitochondria aggravate alterations in the brain microenvironment, promoting neuroinflammation and energy depletion, thereby exacerbating the development of depression. This review summarizes the influence of mitochondrial dysfunction and the underlying molecular pathways on the pathogenesis of depression. Additionally, we discuss the maintenance of mitochondrial homeostasis as a potential therapeutic strategy for depression.
    Keywords:  Major depressive disorder; Mitochondrial dysfunction; Mitochondrial quality control; Neuroinflammation
    DOI:  https://doi.org/10.1016/j.biopha.2023.115652
  36. Mol Genet Metab. 2023 Sep 26. pii: S1096-7192(23)00332-3. [Epub ahead of print]140(3): 107702
    A common benign intronic deletion masking a pathogenic deep intronic PCCB variant - genome sequencing and RNA studies to the rescue.
    Alina Kurolap, Dalit Barel, Nava Shaul Lotan, Isaiah Wexler, Chofit Chai Gadot, Adi Mory, Ortal Barel, Shlomo Almashanu, Hagit Baris Feldman.
      Propionic acidemia (PA) is an autosomal recessive metabolic disorder caused by variants in PCCA or PCCB, both sub-units of the propionyl-CoA carboxylase (PCC) enzyme. PCC is required for the catabolism of certain amino acids and odd-chain fatty acids. In its absence, the accumulated toxic metabolites cause metabolic acidosis, neurologic symptoms, multi-organ dysfunction and possible death. The clinical presentation of PA is highly variable, with typical onset in the neonatal or early infantile period. We encountered two families, whose children were diagnosed with PA. Exome sequencing (ES) failed to identify a pathogenic variant, and we proceeded with genome sequencing (GS), demonstrating homozygosity to a deep intronic PCCB variant. RNA analysis established that this variant creates a pseudoexon with a premature stop codon. The parents are variant carriers, though three of them display pseudo-homozygosity due to a common large benign intronic deletion on the second allele. The parental presumed homozygosity merits special attention, as it masked the causative variant at first, which was resolved only by RNA studies. Arriving at a rapid diagnosis, whether biochemical or genetic, can be crucial in directing lifesaving care, concluding the diagnostic odyssey, and allowing the family prenatal testing in subsequent pregnancies. This study demonstrates the power of integrative genetic studies in reaching a diagnosis, utilizing GS and RNA analysis to overcome ES limitations and define pathogenicity. Importantly, it highlights that intronic deletions should be taken into consideration when analyzing genomic data, so that pseudo-homozygosity would not be misinterpreted as true homozygosity, and pathogenic variants will not be mislabeled as benign.
    Keywords:  Deep intronic variant; Genome sequencing; PCCB; Propionic acidemia; Propionic aciduria
    DOI:  https://doi.org/10.1016/j.ymgme.2023.107702
  37. Nat Commun. 2023 Oct 04. 14(1): 6201
    Cytoplasmic Endonuclease G promotes nonalcoholic fatty liver disease via mTORC2-AKT-ACLY and endoplasmic reticulum stress.
    Wenjun Wang, Junyang Tan, Xiaomin Liu, Wenqi Guo, Mengmeng Li, Xinjie Liu, Yanyan Liu, Wenyu Dai, Liubing Hu, Yimin Wang, Qiuxia Lu, Wen Xing Lee, Hong-Wen Tang, Qinghua Zhou.
      Endonuclease G (ENDOG), a nuclear-encoded mitochondrial intermembrane space protein, is well known to be translocated into the nucleus during apoptosis. Recent studies have shown that ENDOG might enter the mitochondrial matrix to regulate mitochondrial genome cleavage and replication. However, little is known about the role of ENDOG in the cytosol. Our previous work showed that cytoplasmic ENDOG competitively binds with 14-3-3γ, which released TSC2 to repress mTORC1 signaling and induce autophagy. Here, we demonstrate that cytoplasmic ENDOG could also release Rictor from 14-3-3γ to activate the mTORC2-AKT-ACLY axis, resulting in acetyl-CoA production. Importantly, we observe that ENDOG could translocate to the ER, bind with Bip, and release IRE1a/PERK to activate the endoplasmic reticulum stress response, promoting lipid synthesis. Taken together, we demonstrate that loss of ENDOG suppresses acetyl-CoA production and lipid synthesis, along with reducing endoplasmic reticulum stress, which eventually alleviates high-fat diet-induced nonalcoholic fatty liver disease in female mice.
    DOI:  https://doi.org/10.1038/s41467-023-41757-x
  38. Mol Cell. 2023 Sep 29. pii: S1097-2765(23)00739-6. [Epub ahead of print]
    tRNA renovatio: Rebirth through fragmentation.
    Bernhard Kuhle, Qi Chen, Paul Schimmel.
      tRNA function is based on unique structures that enable mRNA decoding using anticodon trinucleotides. These structures interact with specific aminoacyl-tRNA synthetases and ribosomes using 3D shape and sequence signatures. Beyond translation, tRNAs are increasingly recognized as versatile signaling molecules, and they interplay with other RNAs and cellular proteins. These roles arose through a long process of evolution. tRNA fragmentation in particular is not merely random degradation but rather an act of re-creation, generating specific shorter molecules, called tRNA-derived small RNAs (tsRNAs), which exploit their linear sequences and newly arranged 3D structures for unexpected biological functions. Emerging methods to uncover full tRNA/tsRNA sequences and modifications, combined with techniques to study RNA structures and to integrate AI-powered predictions, will enable comprehensive investigations of tRNA fragmentation products and new interaction potentials in relation to their biological functions. We anticipate that these directions will herald a new era for understanding biological complexity and advancing pharmaceutical engineering.
    Keywords:  AI-based RNA structure prediction; RNA structure; biological complexity; deep learning; evolution; non-canonical tRNA functions; tRNA; tRNA fragments; tRNA-derived small RNA
    DOI:  https://doi.org/10.1016/j.molcel.2023.09.016
  39. Expert Rev Proteomics. 2023 Oct 03.
    A Galaxy of informatics resources for MS-based proteomics.
    Subina Mehta, Matthias Bernt, Matthew Chambers, Matthias Fahrner, Melanie Christine Föll, Bjoern Gruening, Carlos Horro, James E Johnson, Valentin Loux, Andrew T Rajczewski, Oliver Schilling, Yves Vandenbrouck, Ove Johan Ragnar Gustafsson, W C Mike Thang, Cameron Hyde, Gareth Price, Pratik D Jagtap, Timothy J Griffin.
       INTRODUCTION: Continuous advances in mass spectrometry (MS) technologies have enabled deeper and more reproducible proteome characterization, and a better understanding of biological systems when integrated with other 'omics data. Bioinformatic resources meeting analysis requirements of increasingly complex MS-based proteomic data, and associated multi-omic data, are critically needed. These requirements included availability of software spanning diverse types of analyses, along with scalability for large-scale, compute-intensive applications and mechanisms to ease adoption of the software.
    AREAS COVERED: The Galaxy ecosystem meets these requirements by offering a multitude of open-source tools for MS-based proteomics analyses and applications, all in an adaptable, scalable, and accessible computing environment. A thriving global community maintains these software and associated training resources to empower researcher-driven analyses.
    EXPERT OPINION: The community-supported Galaxy ecosystem remains a crucial contributor to basic biological and clinical studies using MS-based proteomics. In addition to the current status of Galaxy-based resources, we describe ongoing developments for meeting emerging challenges in MS-based proteomic informatics. We hope this review will catalyze increased use of Galaxy by researchers employing MS-based proteomics and inspire software developers to join the community and implement new tools, workflows, and associated training content that will add further value to this already rich ecosystem.
    Keywords:  Bioinformatics; Computational workflows; Galaxy platform; Mass-spectrometry; Multi-omics; Reproducibility; proteomics
    DOI:  https://doi.org/10.1080/14789450.2023.2265062
  40. Mol Psychiatry. 2023 Oct 02.
    Mitochondrial DNA damage triggers spread of Parkinson's disease-like pathology.
    Emilie Tresse, Joana Marturia-Navarro, Wei Qi Guinevere Sew, Marina Cisquella-Serra, Elham Jaberi, Lluis Riera-Ponsati, Natasha Fauerby, Erling Hu, Oliver Kretz, Susana Aznar, Shohreh Issazadeh-Navikas.
      In the field of neurodegenerative diseases, especially sporadic Parkinson's disease (sPD) with dementia (sPDD), the question of how the disease starts and spreads in the brain remains central. While prion-like proteins have been designated as a culprit, recent studies suggest the involvement of additional factors. We found that oxidative stress, damaged DNA binding, cytosolic DNA sensing, and Toll-Like Receptor (TLR)4/9 activation pathways are strongly associated with the sPDD transcriptome, which has dysregulated type I Interferon (IFN) signaling. In sPD patients, we confirmed deletions of mitochondrial (mt)DNA in the medial frontal gyrus, suggesting a potential role of damaged mtDNA in the disease pathophysiology. To explore its contribution to pathology, we used spontaneous models of sPDD caused by deletion of type I IFN signaling (Ifnb-/-/Ifnar-/- mice). We found that the lack of neuronal IFNβ/IFNAR leads to oxidization, mutation, and deletion in mtDNA, which is subsequently released outside the neurons. Injecting damaged mtDNA into mouse brain induced PDD-like behavioral symptoms, including neuropsychiatric, motor, and cognitive impairments. Furthermore, it caused neurodegeneration in brain regions distant from the injection site, suggesting that damaged mtDNA triggers spread of PDD characteristics in an "infectious-like" manner. We also discovered that the mechanism through which damaged mtDNA causes pathology in healthy neurons is independent of Cyclic GMP-AMP synthase and IFNβ/IFNAR, but rather involves the dual activation of TLR9/4 pathways, resulting in increased oxidative stress and neuronal cell death, respectively. Our proteomic analysis of extracellular vesicles containing damaged mtDNA identified the TLR4 activator, Ribosomal Protein S3 as a key protein involved in recognizing and extruding damaged mtDNA. These findings might shed light on new molecular pathways through which damaged mtDNA initiates and spreads PD-like disease, potentially opening new avenues for therapeutic interventions or disease monitoring.
    DOI:  https://doi.org/10.1038/s41380-023-02251-4
  41. Front Med (Lausanne). 2023 ;10 1267930
    Editorial: Whole Genome Sequencing for rare diseases.
    Chiara Di Resta, Valeria D'Argenio.
      
    Keywords:  Whole Genome Sequencing; diagnosis; pathogenesis; rare diseases; treatment
    DOI:  https://doi.org/10.3389/fmed.2023.1267930
  42. Front Neurol. 2023 ;14 1214137
    A milder form of molybdenum cofactor deficiency type A presenting as Leigh's syndrome-like phenotype highlighting the secondary mitochondrial dysfunction: a case report.
    Montaha Almudhry, Asuri N Prasad, C Anthony Rupar, Keng Yeow Tay, Suzanne Ratko, Mary E Jenkins, Chitra Prasad.
       Background: Molybdenum cofactor deficiency (MoCD) (OMIM# 252150) is an autosomal-recessive disorder caused by mutations in four genes involved in the molybdenum cofactor (MOCO) biosynthesis pathway.
    Objectives: We report a milder phenotype in a patient with MOCS1 gene mutation who presented with a Leigh-like presentation.
    Case report: We present the case of a 10-year-old boy who was symptomatic at the age of 5 months with sudden onset of dyskinesia, nystagmus, and extrapyramidal signs following a febrile illness. Initial biochemical, radiological, and histopathological findings a Leigh syndrome-like phenotype; however, whole-exome sequencing detected compound heterozygous mutations in MOCS1 gene, c.1133 G>C and c.217C>T, confirming an underlying MoCD. This was biochemically supported by low uric acid level of 80 (110-282 mmol/L) and low cystine level of 0 (3-49), and a urine S-sulfocysteine at 116 (0-15) mmol/mol creatinine. The patient was administered methionine- and cystine-free formulas. The patient has remained stable, with residual intellectual, speech, and motor sequelae.
    Conclusion: This presentation expands the phenotypic variability of late-onset MoCD A and highlights the role of secondary mitochondrial dysfunction in its pathogenesis.
    Keywords:  Leigh-like phenotype; MOCS1; dystonia; molybdenum cofactor deficiency; stroke
    DOI:  https://doi.org/10.3389/fneur.2023.1214137
  43. Eur J Hum Genet. 2023 10;31(10): 1091-1092
    Expanding what we know about rare genetic diseases.
    Alisdair McNeill.
      
    DOI:  https://doi.org/10.1038/s41431-023-01453-2
  44. Development. 2023 Oct 06. pii: dev.202024. [Epub ahead of print]
    KDM5-mediated activation of genes required for mitochondrial biology is necessary for viability in Drosophila.
    Michael F Rogers, Owen J Marshall, Julie Secombe.
      Histone modifying proteins play critical roles in the precise regulation of the transcriptional programs that coordinate development. KDM5 family proteins interact with chromatin through demethylating H3K4me3 as well as demethylase-independent mechanisms that remain less understood. To gain fundamental insights into the transcriptional activities of KDM5 proteins, we examined the essential roles of the single kdm5 ortholog of Drosophila during development. KDM5 performs critical functions in the larval neuroendocrine prothoracic gland, providing a model to define its role in regulating key gene expression programs. Integrating genome binding and transcriptomic data, we identify that KDM5 regulates the expression of genes critical for the function and maintenance of mitochondria, and we find that loss of KDM5 causes morphological changes to mitochondria. This is key to the developmental functions of KDM5, as expression of the mitochondrial biogenesis transcription factor Ets97D, homolog of GABPα, is able to suppress the altered mitochondrial morphology as well as the lethality of kdm5 null animals. Together, these data establish KDM5-mediated cellular functions that are important for normal development and could contribute to KDM5-linked disorders when dysregulated.
    Keywords:  Demethylase; KDM5; Mitochondria; Prothoracic gland; Transcription
    DOI:  https://doi.org/10.1242/dev.202024
  45. Nat Methods. 2023 Oct 02.
    Streamlined and sensitive mono- and di-ribosome profiling in yeast and human cells.
    Lucas Ferguson, Heather E Upton, Sydney C Pimentel, Amanda Mok, Liana F Lareau, Kathleen Collins, Nicholas T Ingolia.
      Ribosome profiling has unveiled diverse regulation and perturbations of translation through a transcriptome-wide survey of ribosome occupancy, read out by sequencing of ribosome-protected messenger RNA fragments. Generation of ribosome footprints and their conversion into sequencing libraries is technically demanding and sensitive to biases that distort the representation of physiological ribosome occupancy. We address these challenges by producing ribosome footprints with P1 nuclease rather than RNase I and replacing RNA ligation with ordered two-template relay, a single-tube protocol for sequencing library preparation that incorporates adaptors by reverse transcription. Our streamlined approach reduced sequence bias and enhanced enrichment of ribosome footprints relative to ribosomal RNA. Furthermore, P1 nuclease preserved distinct juxtaposed ribosome complexes informative about yeast and human ribosome fates during translation initiation, stalling and termination. Our optimized methods for mRNA footprint generation and capture provide a richer translatome profile with low input and fewer technical challenges.
    DOI:  https://doi.org/10.1038/s41592-023-02028-1
  46. medRxiv. 2023 Aug 21. pii: 2023.08.14.23293948. [Epub ahead of print]
    Long-read sequencing resolves a complex structural variant in PRKN Parkinson's disease.
    Kensuke Daida, Manabu Funayama, Kimberley J Billingsley, Laksh Malik, Abigail Miano-Burkhardt, Hampton L Leonard, Mary B Makarious, Hirotaka Iwaki, Jinhui Ding, J Raphael Gibbs, Mayu Ishiguro, Hiroyo Yoshino, Kotaro Ogaki, Genko Oyama, Kenya Nishioka, Risa Nonaka, Wado Akamatsu, Cornelis Blauwendraat, Nobutaka Hattori.
       Background: PRKN mutations are the most common cause of young onset and autosomal recessive Parkinson's disease (PD). PRKN is located in FRA6E which is one of the common fragile sites in the human genome, making this region prone to structural variants. However, complex structural variants such as inversions of PRKN are seldom reported, suggesting that there are potentially unrevealed complex pathogenic PRKN structural variants.
    Objectives: To identify complex structural variants in PRKN using long-read sequencing.
    Methods: We investigated the genetic cause of monozygotic twins presenting with a young onset dystonia-parkinsonism using targeted sequencing, whole exome sequencing, multiple ligation probe amplification, and long-read. We assessed the presence and frequency of complex inversions overlapping PRKN using whole-genome sequencing data of AMP-PD and UK-Biobank datasets.
    Results: Multiple ligation probe amplification identified a heterozygous exon 3 deletion in PRKN and long-read sequencing identified a large novel inversion spanning over 7Mb, including a large part of the coding DNA sequence of PRKN . We could diagnose the affected subjects as compound heterozygous carriers of PRKN . We analyzed whole genome sequencing data of 43,538 participants of the UK-Biobank and 4,941 participants of the AMP-PD datasets. Nine inversions in the UK-Biobank and two in AMP PD were identified and were considered potentially damaging and likely to affect PRKN isoforms.
    Conclusions: This is the first report describing a large 7Mb inversion involving breakpoints outside of PRKN . This study highlights the importance of using long-read whole genome sequencing for structural variant analysis in unresolved young-onset PD cases.
    DOI:  https://doi.org/10.1101/2023.08.14.23293948
  47. JACC Basic Transl Sci. 2023 Sep;8(9): 1138-1140
    Ketone Bodies Preserve Mitochondria Through Epigenetics.
    Risa Mukai, Junichi Sadoshima.
      
    Keywords:  PGC-1α; epigenetics; heart failure; histone methylation; ketone bodies
    DOI:  https://doi.org/10.1016/j.jacbts.2023.05.013
  48. Front Genet. 2023 ;14 1251951
    Mitochondrial DNA heteroplasmy analysis in keratoconus patients from China.
    Liyan Xu, Kaili Yang, Qi Fan, Yuwei Gu, Shengwei Ren.
      Background: Mitochondrial DNA (mtDNA) variants have been implicated in keratoconus (KC). The present study aimed to characterize the mtDNA heteroplasmy profile in KC and explore the association of mitochondrial heteroplasmic levels with KC. Methods: Mitochondrial sequencing of peripheral blood samples and corneal tomography were conducted in 300 KC cases and 300 matched controls. The number of heteroplasmic and homoplasmic variants was calculated across the mitochondrial genome. Spearman's correlation was used to analyze the correlation between the number of heteroplasmic variants and age. The association of mtDNA heteroplasmic level with KC was analyzed by logistic regression analysis. Moreover, the relationship between mitochondrial heteroplasmic levels and clinical parameters was determined by linear regression analysis. Results: The distribution of mtDNA heteroplasmic variants showed the highest number of heteroplasmic variants in the non-coding region, while the COX3 gene exhibited the highest number in protein-coding genes. Comparisons of the number of heteroplasmic and homoplasmic non-synonymous variants in protein-coding genes revealed no significant differences between KC cases and controls (all p > 0.05). In addition, the number of heteroplasmic variants was positively associated with age in all subjects (r = 0.085, p = 0.037). The logistic regression analyses indicated that the heteroplasmic levels of m.16180_16181delAA was associated with KC (p < 0.005). Linear regression analyses demonstrated that the heteroplasmic levels of m.16180_16181delAA and m.302A>C were not correlated with thinnest corneal thickness (TCT), steep keratometry (Ks), and flat keratometry (Kf) (all p > 0.05) in KC cases and controls separately. Conclusion: The current study characterized the mtDNA heteroplasmy profile in KC, and revealed that the heteroplasmic levels of m.16180_16181delAA were associated with KC.
    Keywords:  heteroplasmy; keratoconus; m.16180_16181delAA; mitochondrial DNA; variant
    DOI:  https://doi.org/10.3389/fgene.2023.1251951
  49. J Biol Chem. 2023 Sep 28. pii: S0021-9258(23)02331-1. [Epub ahead of print] 105303
    A conserved, non-canonical insert in FIS1 mediates TBC1D15 and DRP1 recruitment for mitochondrial fission.
    Ugochukwu K Ihenacho, Rafael Toro, Rana H Mansour, R Blake Hill.
      Mitochondrial fission protein 1 (FIS1) is conserved in all eukaryotes, yet its function in metazoans is thought divergent from lower eukaryotes like fungi. To address this discrepancy, structure-based sequence alignments revealed a conserved but non-canonical three-residue insert (Ser-X-X) in a turn of FIS1, suggesting a conserved function. In vertebrate FIS1, this insert is serine (S45), lysine (K46), and tyrosine (Y47). To determine the biological role of the "SKY insert" in vertebrates, three variants were evaluated for their fold and tested in HCT116 cells for altered mitochondrial morphology and recruitment of effectors, DRP1 and TBC1D15. Substitution of the SKY insert with three alanine residues (AAA) or deletion of the insert (ΔSKY) did not substantially alter the fold or thermal stability of the protein. Replacing SKY with a canonical turn (ΔSKYD49G) introduced significant conformational heterogeneity by NMR that was removed upon deletion of a known regulatory region, the FIS1 arm. Expression of AAA fragmented mitochondria into perinuclear clumps associated with increased mitochondrial DRP1 similar to the wild-type protein. In contrast, the expression of ΔSKY variants led to elongated mitochondrial networks and reduced mitochondrial DRP1 by colocalization analysis, although DRP1 coimmunoprecipitates were highly enriched with ΔSKY variants. Co-expression of YFP-TBC1D15 with ΔSKY variants rescued mitochondrial morphology, despite a reduced ability to drive YFP-TBC1D15 into punctate structures that is found upon co-expression with wildtype FIS1 or the AAA variant. In support YFP-TBC1D15 coimmunoprecipitates were poorly enriched with ΔSKY variants. Co-expression of YFP-TBC1D15 also revealed a gain of function phenotype with the AAA variant compared to wildtype. Collectively these results show that FIS1 can be modulated by conserved residues, thus supporting a unifying model whereby FIS1 activity is effectively governed by intramolecular interactions between the regulatory FIS1 arm and an S-X-X insert that is conserved across eukaryotes.
    Keywords:  Mitochondria; dynamin; fission; mitophagy; nuclear magnetic resonance (NMR); organelle dynamic; peroxisome; protein motif; repeat proteins; tetratricopeptide repeat
    DOI:  https://doi.org/10.1016/j.jbc.2023.105303
  50. Brain. 2023 Oct 06. pii: awad345. [Epub ahead of print]
    Machine learning nominates the inositol pathway and novel genes in Parkinson's disease.
    Eric Yu, Roxanne Larivière, Rhalena A Thomas, Lang Liu, Konstantin Senkevich, Shady Rahayel, Jean-François Trempe, Edward A Fon, Ziv Gan-Or.
      There are 78 loci associated with Parkinson's disease (PD) in the most recent genome-wide association study (GWAS), yet the specific genes driving these associations are mostly unknown. Herein, we aimed to nominate the top candidate gene from each PD locus, and identify variants and pathways potentially involved in PD. We trained a machine learning model to predict PD-associated genes from GWAS loci using genomic, transcriptomic, and epigenomic data from brain tissues and dopaminergic neurons. We nominated candidate genes in each locus, identified novel pathways potentially involved in PD, such as the inositol phosphate biosynthetic pathway (INPP5F, IP6K2, ITPKB, PPIP5K2). Specific common coding variants in SPNS1 and MLX may be involved in PD, and burden tests of rare variants further support that CNIP3, LSM7, NUCKS1 and the polyol/inositol phosphate biosynthetic pathway are associated with PD. Functional studies are needed to further analyze the involvements of these genes and pathways in PD.
    Keywords:  GWAS; Parkinson’s disease; gene prioritization; machine learning
    DOI:  https://doi.org/10.1093/brain/awad345
  51. Nat Metab. 2023 Oct 02.
    Fuelling the fire: de novo lipogenesis primes thermogenesis.
    S Rodríguez-Fdez, A Vidal-Puig.
      
    DOI:  https://doi.org/10.1038/s42255-023-00865-0
  52. Nature. 2023 Oct 06.
    Gene therapies for rare diseases are under threat. Scientists hope to save them.
    Heidi Ledford.
      
    Keywords:  Business; Drug discovery; Gene therapy; Medical research; Society
    DOI:  https://doi.org/10.1038/d41586-023-03109-z
  53. Exp Hematol. 2023 Sep 29. pii: S0301-472X(23)01733-2. [Epub ahead of print]
    Metabolic Regulation of Aged Hematopoietic Stem Cells: Key Players and Mechanisms.
    Nazanin Karimnia, James Harris, Shen Y Heazlewood, Benjamin Cao, Susan K Nilsson.
      Aging is accompanied by a gradual decline in the function and regenerative capacity of hematopoietic stem cells (HSCs), which leads to increased susceptibility to blood disorders. Recent studies have highlighted the critical role of metabolic regulation in governing the fate and function of HSCs, and alterations in metabolism play a critical role in the age-related changes observed in HSCs. Metabolic processes including glycolysis, mitochondrial function, nutrient sensing and inflammation, profoundly influence the maintenance, self-renewal and differentiation potential of the HSC pool. This review focuses on the metabolic alterations that occur in HSCs during aging and the systemic factors which contribute to HSC metabolic dysregulation, leading to impaired cellular function and reduced regenerative capacity. We highlight the impact of age-associated changes in oxidative stress, mitochondrial dysfunction, nutrient availability and inflammation on HSC metabolism and function. Targeting metabolic pathways and modulating key regulators of metabolism hold promise for reducing age-related HSC dysregulation, thus maintaining functional potential as a path towards healthy aging. Exploiting these metabolic interventions has the potential to improve hemopoietic recovery, enhance immune function and pave the way for novel therapeutic interventions to combat age-related blood disorders.
    Keywords:  Aging; autophagy; hematopoietic stem cell; metabolism; mitochondria; niche; nutrients
    DOI:  https://doi.org/10.1016/j.exphem.2023.09.006
  54. Elife. 2023 10 02. pii: e79725. [Epub ahead of print]12
    Regulation of defective mitochondrial DNA accumulation and transmission in C. elegans by the programmed cell death and aging pathways.
    Sagen Flowers, Rushali Kothari, Yamila N Torres Cleuren, Melissa R Alcorn, Chee Kiang Ewe, Geneva Alok, Samantha L Fiallo, Pradeep M Joshi, Joel H Rothman.
      The heteroplasmic state of eukaryotic cells allows for cryptic accumulation of defective mitochondrial genomes (mtDNA). 'Purifying selection' mechanisms operate to remove such dysfunctional mtDNAs. We found that activators of programmed cell death (PCD), including the CED-3 and CSP-1 caspases, the BH3-only protein CED-13, and PCD corpse engulfment factors, are required in C. elegans to attenuate germline abundance of a 3.1-kb mtDNA deletion mutation, uaDf5, which is normally stably maintained in heteroplasmy with wildtype mtDNA. In contrast, removal of CED-4/Apaf1 or a mutation in the CED-4-interacting prodomain of CED-3, do not increase accumulation of the defective mtDNA, suggesting induction of a non-canonical germline PCD mechanism or non-apoptotic action of the CED-13/caspase axis. We also found that the abundance of germline mtDNAuaDf5 reproducibly increases with age of the mothers. This effect is transmitted to the offspring of mothers, with only partial intergenerational removal of the defective mtDNA. In mutants with elevated mtDNAuaDf5 levels, this removal is enhanced in older mothers, suggesting an age-dependent mechanism of mtDNA quality control. Indeed, we found that both steady-state and age-dependent accumulation rates of uaDf5 are markedly decreased in long-lived, and increased in short-lived, mutants. These findings reveal that regulators of both PCD and the aging program are required for germline mtDNA quality control and its intergenerational transmission.
    Keywords:  C. elegans; aging; cell biology; genetics; genomics; heteroplasmy; insulin signaling; programmed cell death; purifying selection; uaDf5
    DOI:  https://doi.org/10.7554/eLife.79725
  55. Genome Biol. 2023 Oct 05. 24(1): 221
    Genomic variant benchmark: if you cannot measure it, you cannot improve it.
    Sina Majidian, Daniel Paiva Agustinho, Chen-Shan Chin, Fritz J Sedlazeck, Medhat Mahmoud.
      Genomic benchmark datasets are essential to driving the field of genomics and bioinformatics. They provide a snapshot of the performances of sequencing technologies and analytical methods and highlight future challenges. However, they depend on sequencing technology, reference genome, and available benchmarking methods. Thus, creating a genomic benchmark dataset is laborious and highly challenging, often involving multiple sequencing technologies, different variant calling tools, and laborious manual curation. In this review, we discuss the available benchmark datasets and their utility. Additionally, we focus on the most recent benchmark of genes with medical relevance and challenging genomic complexity.
    Keywords:  Benchmark datasets; Genetic variation; Indels; Medical genes; SNPs; Sequencing technology; Structural variant
    DOI:  https://doi.org/10.1186/s13059-023-03061-1
  56. RNA Biol. 2023 Jan;20(1): 791-804
    Anticodon sequence determines the impact of mistranslating tRNAAla variants.
    Ecaterina Cozma, Megha Rao, Madison Dusick, Julie Genereaux, Ricard A Rodriguez-Mias, Judit Villén, Christopher J Brandl, Matthew D Berg.
      Transfer RNAs (tRNAs) maintain translation fidelity through accurate charging by their cognate aminoacyl-tRNA synthetase and codon:anticodon base pairing with the mRNA at the ribosome. Mistranslation occurs when an amino acid not specified by the genetic message is incorporated into proteins and has applications in biotechnology, therapeutics and is relevant to disease. Since the alanyl-tRNA synthetase uniquely recognizes a G3:U70 base pair in tRNAAla and the anticodon plays no role in charging, tRNAAla variants with anticodon mutations have the potential to mis-incorporate alanine. Here, we characterize the impact of the 60 non-alanine tRNAAla anticodon variants on the growth of Saccharomyces cerevisiae. Overall, 36 tRNAAla anticodon variants decreased growth in single- or multi-copy. Mass spectrometry analysis of the cellular proteome revealed that 52 of 57 anticodon variants, not decoding alanine or stop codons, induced mistranslation when on single-copy plasmids. Variants with G/C-rich anticodons resulted in larger growth deficits than A/U-rich variants. In most instances, synonymous anticodon variants impact growth differently, with anticodons containing U at base 34 being the least impactful. For anticodons generating the same amino acid substitution, reduced growth generally correlated with the abundance of detected mistranslation events. Differences in decoding specificity, even between synonymous anticodons, resulted in each tRNAAla variant mistranslating unique sets of peptides and proteins. We suggest that these differences in decoding specificity are also important in determining the impact of tRNAAla anticodon variants.
    Keywords:  Mistranslation; expanded decoding; genetic code; tRNA biology; tRNAAla
    DOI:  https://doi.org/10.1080/15476286.2023.2257471
  57. Eur J Hum Genet. 2023 Oct 03.
    Knowledge, attitudes and decision regret: a longitudinal survey study of participants offered genome sequencing in the 100,000 Genomes Project.
    Michelle Peter, Jennifer Hammond, Saskia C Sanderson, Jana Gurasashvili, Amy Hunter, Beverly Searle, Christine Patch, Lyn S Chitty, Melissa Hill, Celine Lewis.
      We used cross-sectional surveys to compare the knowledge, attitudes, and decision regret of participants who had consented for genome sequencing (GS) for rare disease diagnosis in the 100,000 Genomes Project (100kGP) across two timepoints (at the time of consenting for GS (T1) and 12-18 months later (T2)). At T1, participants (n = 504) completed a survey that included measures of general knowledge of GS ("Knowledge of Genome Sequencing" (KOGS)), specific knowledge of GS and attitudes towards GS ("General attitudes" and "Specific attitudes"). At T2, participants (n = 296) completed these same assessments (apart from the specific knowledge scale) together with an assessment of decision regret towards GS ("Decisional Regret Scale"). At 12-18 months after consenting for GS, participants' basic knowledge of GS had remained stable. General knowledge of GS varied across topics; concepts underlying more general information about genetics were better understood than the technical details of genomic testing. Attitudes towards GS at T2 were generally positive, and feelings towards GS (both positive and negative) remained unchanged. However, those who were more positive about the test at the outset had greater specific knowledge (as opposed to general knowledge) of GS. Finally, although the majority of participants indicated feeling little regret towards undergoing GS, those with low positive attitude and high negative attitude about GS at T1 reported greater decision regret at T2. Careful assessment of patient knowledge about and attitudes towards GS at the time of offering testing is crucial for supporting informed decision making and mitigating later regret.
    DOI:  https://doi.org/10.1038/s41431-023-01470-1
  58. FEBS J. 2023 Oct 03.
    Association of 2-oxoacid dehydrogenase complexes with respirasomes in mitochondria.
    Konstantin S Plokhikh, Semen V Nesterov, Yuriy M Chesnokov, Anton G Rogov, Roman A Kamyshinsky, Aleksandr L Vasiliev, Lev S Yaguzhinsky, Raif G Vasilov.
      In this work, cryo-electron tomography (cryo-ET) was used to study the localization of 2-oxoacid dehydrogenase complexes (OADCs) in cardiac mitochondria and mitochondrial inner membrane samples. Two classes of ordered OADC inner cores with different symmetries were distinguished and their quaternary structures modeled. One class corresponds to pyruvate dehydrogenase complexes and the other to dehydrogenase complexes of α-ketoglutarate and branched-chain α-ketoacids. OADCs were shown to be localized in close proximity to membrane-embedded respirasomes, as observed both in densely packed lamellar cristae of cardiac mitochondria and in ruptured mitochondrial samples where the dense packing is absent. This suggests the specificity of the OADC -respirasome interaction, which allows localized NADH/NAD+ exchange between OADCs and complex I of the respiratory chain. The importance of this local coupling lies in the fact that OADCs are the link between respiration, glycolysis and amino acid metabolism. The coupling of these basic metabolic processes can vary in different tissues and conditions and may be involved in the development of various pathologies. This article shows that this important and previously missing parameter of mitochondrial complex coupling can be successfully assessed using cryo-ET.
    Keywords:  2-oxoacid; cryo-electron tomography; dehydrogenase complex; mitochondria; pyruvate
    DOI:  https://doi.org/10.1111/febs.16965
  59. Proc Natl Acad Sci U S A. 2023 Oct 10. 120(41): e2221165120
    Deciphering RNA splicing logic with interpretable machine learning.
    Susan E Liao, Mukund Sudarshan, Oded Regev.
      Machine learning methods, particularly neural networks trained on large datasets, are transforming how scientists approach scientific discovery and experimental design. However, current state-of-the-art neural networks are limited by their uninterpretability: Despite their excellent accuracy, they cannot describe how they arrived at their predictions. Here, using an "interpretable-by-design" approach, we present a neural network model that provides insights into RNA splicing, a fundamental process in the transfer of genomic information into functional biochemical products. Although we designed our model to emphasize interpretability, its predictive accuracy is on par with state-of-the-art models. To demonstrate the model's interpretability, we introduce a visualization that, for any given exon, allows us to trace and quantify the entire decision process from input sequence to output splicing prediction. Importantly, the model revealed uncharacterized components of the splicing logic, which we experimentally validated. This study highlights how interpretable machine learning can advance scientific discovery.
    Keywords:  RNA splicing; artificial intelligence; interpretable machine learning
    DOI:  https://doi.org/10.1073/pnas.2221165120
  60. eNeuro. 2023 Sep 28. pii: ENEURO.0159-23.2023. [Epub ahead of print]
    Spatiotemporal regulation of de novo and salvage purine synthesis during brain development.
    Tomoya Mizukoshi, Seiya Yamada, Shin-Ichi Sakakibara.
      The levels of purines, essential molecules to sustain eukaryotic cell homeostasis, are regulated by the coordination of the de novo and salvage synthesis pathways. In the embryonic central nervous system (CNS), the de novo pathway is considered crucial to meet the requirements for the active proliferation of neural stem/progenitor cells (NSPCs). However, how these two pathways are balanced or separately utilized during CNS development remains poorly understood. In this study, we showed a dynamic shift in pathway utilization, with greater reliance on the de novo pathway during embryonic stages and on the salvage pathway in postnatal-adult mouse brain. The pharmacological effects of various purine synthesis inhibitors in vitro and the expression profile of purine synthesis enzymes indicated that NSPCs in the embryonic cerebrum mainly utilize the de novo pathway. Simultaneously, NSPCs in the cerebellum require both the de novo and the salvage pathways. In vivo administration of de novo inhibitors resulted in severe hypoplasia of the forebrain cortical region, indicating a gradient of purine demand along the anteroposterior axis of the embryonic brain, with cortical areas of the dorsal forebrain having higher purine requirements than ventral or posterior areas such as the striatum and thalamus. This histological defect of the neocortex was accompanied by strong downregulation of the mechanistic target of rapamycin complex 1 (mTORC1)/ribosomal protein S6 kinase (S6K)/S6 signaling cascade, a crucial pathway for cell metabolism, growth, and survival. These findings indicate the importance of the spatiotemporal regulation of both purine pathways for mTORC1 signaling and proper brain development.Significance StatementBrain development requires a balance of de novo and salvage purine synthetic pathways. However, the utilization of these pathways during brain development remains poorly understood. This study provides evidence that the spatiotemporal regulation of these two purine synthesis pathways is essential for normal brain development. We revealed that inhibition of de novo purine synthesis results in the downregulation of mammalian/mechanistic target of rapamycin (mTOR) signaling, leading to malformations in specific embryonic brain regions such as the cerebral neocortex. These results suggest a temporal and spatial gradient of purine demand during embryonic brain development. These findings could improve our understanding of neurological diseases caused by defects in purine metabolism.
    Keywords:  NSPCs; cortical development; de novo pathway; mTOR; purine metabolism; salvage pathway
    DOI:  https://doi.org/10.1523/ENEURO.0159-23.2023
  61. Clin Chem Lab Med. 2023 Oct 06.
    Analysis of a second-tier test panel in dried blood spot samples using liquid chromatography-tandem mass spectrometry in Catalonia's newborn screening programme.
    Sonia Pajares-García, José Manuel González de Aledo-Castillo, José Eduardo Flores-Jiménez, Tatiana Collado, Judit Pérez, Abraham José Paredes-Fuentes, Ana Argudo-Ramírez, Rosa María López-Galera, Blanca Prats, Judit García-Villoria.
       OBJECTIVES: Acylcarnitine and amino acid analyses of dried blood spot (DBS) samples using tandem mass spectrometry in newborn screening (NBS) programmes can generate false positive (FP) results. Therefore, implementation of second-tier tests (2TTs) using DBS samples has become increasingly important to avoid FPs. The most widely used 2TT metabolites include methylmalonic acid, 3-hydroxypropionic acid, methylcitric acid, and homocysteine.
    METHODS: We simultaneously measured 46 underivatised metabolites, including organic acids, acylglycine and acylcarnitine isomers, homocysteine, and orotic acid, in DBS samples using tandem mass spectrometry. To validate this method, we analysed samples from 147 healthy newborns, 160 patients with genetic disorders diagnosed via NBS, 20 patients with acquired vitamin B12 deficiency, 10 newborns receiving antibiotic treatment, and nine external quality control samples.
    RESULTS: The validation study revealed that 31 metabolites showed good analytical performance. Furthermore, this method detected key metabolites for all diseases associated with increased levels of the following acylcarnitines: C3, C4, C5, C4DC\C5OH, and C5DC. The sensitivity of this method to detect all diseases was 100 %, and the specificity was 74-99 %, except for glutaric aciduria type 1. This method can also be used to diagnose mitochondrial fatty acid β-oxidation disorders (FAODs) and urea cycle defects (UCDs).
    CONCLUSIONS: We have described a 2TT panel of 31 metabolites in DBS samples based on an easy and rapid method without derivatisation. Its implementation allowed us to distinguish between different organic acidurias, some FAODs, and UCDs. This new strategy has increased the efficiency of our NBS programme by reducing FP and false negative results, second sample requests, and the time required for diagnosis.
    Keywords:  acylcarnitines; acylglycines; mass spectrometry; newborn screening; organic acids; second-tier test
    DOI:  https://doi.org/10.1515/cclm-2023-0216
  62. Cell Rep. 2023 Sep 29. pii: S2211-1247(23)01195-6. [Epub ahead of print]42(10): 113183
    ApoE4-dependent lysosomal cholesterol accumulation impairs mitochondrial homeostasis and oxidative phosphorylation in human astrocytes.
    Hyein Lee, Sukhee Cho, Mi-Jin Kim, Yeo Jin Park, Eunji Cho, Yeon Suk Jo, Yong-Seok Kim, Jung Yi Lee, Themis Thoudam, Seung-Hwa Woo, Se-In Lee, Juyeong Jeon, Young-Sam Lee, Byung-Chang Suh, Jong Hyuk Yoon, Younghoon Go, In-Kyu Lee, Jinsoo Seo.
      Recent developments in genome sequencing have expanded the knowledge of genetic factors associated with late-onset Alzheimer's disease (AD). Among them, genetic variant ε4 of the APOE gene (APOE4) confers the greatest disease risk. Dysregulated glucose metabolism is an early pathological feature of AD. Using isogenic ApoE3 and ApoE4 astrocytes derived from human induced pluripotent stem cells, we find that ApoE4 increases glycolytic activity but impairs mitochondrial respiration in astrocytes. Ultrastructural and autophagy flux analyses show that ApoE4-induced cholesterol accumulation impairs lysosome-dependent removal of damaged mitochondria. Acute treatment with cholesterol-depleting agents restores autophagic activity, mitochondrial dynamics, and associated proteomes, and extended treatment rescues mitochondrial respiration in ApoE4 astrocytes. Taken together, our study provides a direct link between ApoE4-induced lysosomal cholesterol accumulation and abnormal oxidative phosphorylation.
    Keywords:  ApoE4; CP: Cell biology; CP: Metabolism; glucose metabolism; human astrocytes; lysosomal cholesterol accumulation; mitophagy
    DOI:  https://doi.org/10.1016/j.celrep.2023.113183
  63. Am J Med Genet A. 2023 Oct 05.
    Refining the phenotypic spectrum of CCDC88A-related PEHO-like syndrome.
    Mahmoud Y Issa, Mona A Hafez, Samir M Mounir, Sherif F Abdel Ghafar, Maha S Zaki, Mohamed S Abdel-Hamid.
      Progressive encephalopathy with edema, hypsarrhythmia, and optic atrophy (PEHO) and PEHO-like syndromes are very rare infantile disorders characterized by profound intellectual disability, hypotonia, convulsions, optic, and progressive brain atrophy. Many causative genes for PEHO and PEHO-like syndromes have been identified including CCDC88A. So far, only five patients from two unrelated families with biallelic CCDC88A variants have been reported in the literature. Herein, we describe a new family from Egypt with a lethal epileptic encephalopathy. Our patient was the youngest child born to a highly consanguineous couple and had a family history of five deceased sibs with the same condition. She presented with postnatal microcephaly, poor visual responsiveness, and epilepsy. Her brain MRI showed abnormal cortical gyration with failure of opercularization of the insula, hypogenesis of corpus callosum, colpocephaly, reduced white matter, hypoplastic vermis, and brain stem. Whole exome sequencing identified a new homozygous frameshift variant in CCDC88A gene (c.1795_1798delACAA, p.Thr599ValfsTer4). Our study presents the third reported family with this extremely rare disorder. We also reviewed all described cases to better refine the phenotypic spectrum associated with biallelic loss of function variants in the CCDC88A gene.
    Keywords:  CCDC88A gene; PEHO like syndrome; brain dysplasia; microcephaly; whole exome sequencing
    DOI:  https://doi.org/10.1002/ajmg.a.63425
  64. J Neurol. 2023 Oct 03.
    Clinical and genetic analyses of a Swedish patient series diagnosed with ataxia.
    Sorina Gorcenco, Efthymia Kafantari, Joel Wallenius, Christin Karremo, Erik Alinder, Sigurd Dobloug, Maria Landqvist Waldö, Elisabet Englund, Hans Ehrencrona, Klas Wictorin, Kristina Karrman, Andreas Puschmann.
      Hereditary ataxia is a heterogeneous group of complex neurological disorders. Next-generation sequencing methods have become a great help in clinical diagnostics, but it may remain challenging to determine if a genetic variant is the cause of the patient's disease. We compiled a consecutive single-center series of 87 patients from 76 families with progressive ataxia of known or unknown etiology. We investigated them clinically and genetically using whole exome or whole genome sequencing. Test methods were selected depending on family history, clinical phenotype, and availability. Genetic results were interpreted based on the American College of Medical Genetics criteria. For high-suspicion variants of uncertain significance, renewed bioinformatical and clinical evaluation was performed to assess the level of pathogenicity. Thirty (39.5%) of the 76 families had received a genetic diagnosis at the end of our study. We present the predominant etiologies of hereditary ataxia in a Swedish patient series. In two families, we established a clinical diagnosis, although the genetic variant was classified as "of uncertain significance" only, and in an additional three families, results are pending. We found a pathogenic variant in one family, but we suspect that it does not explain the complete clinical picture. We conclude that correctly interpreting genetic variants in complex neurogenetic diseases requires genetics and clinical expertise. The neurologist's careful phenotyping remains essential to confirm or reject a diagnosis, also by reassessing clinical findings after a candidate genetic variant is suggested. Collaboration between neurology and clinical genetics and combining clinical and research approaches optimizes diagnostic yield.
    Keywords:  Hereditary ataxia; Next-generation sequencing; Post-NGS phenotyping
    DOI:  https://doi.org/10.1007/s00415-023-11990-x
  65. J Biol Chem. 2023 Sep 29. pii: S0021-9258(23)02334-7. [Epub ahead of print] 105306
    The mitochondrial amidoxime reducing component (mARC) - from prodrug-activation mechanism to drug-metabolising enzyme and onwards to drug target.
    Michel A Struwe, Axel J Scheidig, Bernd Clement.
      The mitochondrial amidoxime reducing component (mARC) is one of five known molybdenum enzymes in eukaryotes. mARC belongs to the MOSC domain superfamily, a large group of so far poorly studied molybdoenzymes. mARC was initially discovered as the enzyme activating N-hydroxylated prodrugs of basic amidines but has since been shown to also reduce a variety of other N-oxygenated compounds, e.g., toxic nucleobase analogues. Under certain circumstances, mARC might also be involved in reductive nitric oxide synthesis through reduction of nitrite. Recently, mARC enzymes have received a lot of attention due to their apparent involvement in lipid metabolism and, in particular, because many genome-wide association studies have shown a common variant of human mARC1 to have a protective effect against liver disease. The mechanism linking mARC enzymes with lipid metabolism remains unknown. Here, we give a comprehensive overview of what is currently known about mARC enzymes, their substrates, structure and apparent involvement in human disease.
    Keywords:  Molybdenum; drug metabolism; enzyme; liver; mARC; metalloprotein; nitric oxide
    DOI:  https://doi.org/10.1016/j.jbc.2023.105306
  66. Compr Physiol. 2023 Sep 28. 13(4): 5115-5155
    Links between Exercise Capacity, Exercise Training, and Metabolism.
    Alena Spagnolo, Sebastian Klug, Christina Schenkl, Michael Schwarzer.
      Exercise capacity of an individual describes the ability to perform physical activity. This exercise capacity is influenced by intrinsic factors such as genetic constitution and extrinsic factors such as exercise training. On the metabolic level exercise and metabolism are linked. As an important site of metabolism and the main source for ATP needed for muscle contraction, mitochondrial function can determine exercise capacity, and exercise inversely influences mitochondrial function. It has been suggested that exercise mediates many of its effects due to such metabolic changes. Although extrinsic factors affect exercise capacity, a major part of an individual's exercise capacity is genetically determined, and extrinsic factors can only improve on this baseline. Looking at the effect of exercise capacity on and with disease, the two go hand in hand. On one hand, disease is negatively affecting an individual's exercise capacity; on the other hand, exercise offers an effective treatment option. Combining these factors, exercise capacity is an often-ignored prognostic variable for life expectancy as well as morbidity and mortality. In this review, we aim to provide the current knowledge on the links between inherited and acquired exercise capacity, as well as the mechanisms in which metabolism interacts with exercise capacity. © 2023 American Physiological Society. Compr Physiol 13:5115-5155, 2023.
    DOI:  https://doi.org/10.1002/cphy.c230004
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