bims-mitdis Biomed News
on Mitochondrial disorders
Issue of 2025–03–30
sixty-two papers selected by
Catalina Vasilescu, Helmholz Munich
  1. A cellular assay to determine the fusion capacity of MFN2 variants linked to Charcot-Marie-Tooth disease of type 2 A.
  2. Dependence of mitochondrial calcium signalling and dynamics on the disaggregase, CLPB.
  3. The Balance of MFN2 and OPA1 in Mitochondrial Dynamics, Cellular Homeostasis, and Disease.
  4. Unclogging of the TOM complex under import stress.
  5. Mitochondrial DNA Pathogenic Variants in Ophthalmic Diseases: A Review.
  6. The Pathogenesis of Very Long-Chain Acyl-CoA Dehydrogenase Deficiency.
  7. A map of mitochondrial biology reveals the energy landscape of the human brain.
  8. H2S remodels mitochondrial ultrastructure and destabilizes respiratory supercomplexes.
  9. A New Perspective on the Role of Alterations in Mitochondrial Proteins Involved in ATP Synthesis and Mobilization in Cardiomyopathies.
  10. Mitochondrial DNA variants revealed by whole exome sequencing: from screening to diagnosis and follow-up.
  11. Current Understanding of Pathogenic Mechanisms and Disease Models of Citrin Deficiency.
  12. Diverse Phenotypes of Mitochondrial Disease With Varying Levels of Heteroplasmy.
  13. Mechanistic insights into impaired β-oxidation and its role in mitochondrial dysfunction: A comprehensive review.
  14. OmicsCam Enables Trimodal Profiling of Mitochondrial Genome Editing.
  15. Organoids as 3D Models for Studying Exogenous Mitochondrial Transplantation.
  16. Mitochondrial Proteome Reveals Metabolic Tuning by Restricted Insulin Signaling to Promote Longevity in Caenorhabditis elegans.
  17. Cell biology: A new dynamin superfamily protein remodels mitochondrial dynamics.
  18. The fate of mitochondrial respiratory complexes in aging.
  19. In situ architecture of the human prohibitin complex.
  20. Adult-onset Leigh syndrome with recurrent seizures and peripheral neuropathy due to the 9176T > C mutation: a case report and literature review.
  21. Mitochondria targeted nanoparticles for the treatment of mitochondrial dysfunction-associated brain disorders.
  22. Multiple Inhibitory Mechanisms of DS16570511 Targeting Mitochondrial Calcium Uptake: Insights from Biochemical Analysis of Rat Liver Mitochondria.
  23. Mitochondrial respiration defects in lymphoblast cell lines from patients with Dravet syndrome.
  24. Ameliorating TIMM50 Loss Slows Senescence by Improving Mitochondrial Structure and Function.
  25. Dual-Modality Imaging Unveil Inner Mitochondrial Membrane Viscosity and Respiratory Dynamics in Mitophagy.
  26. Plasticity of the mammalian integrated stress response.
  27. A human brain map of mitochondrial respiratory capacity and diversity.
  28. Mitochondria transplantation transiently rescues cerebellar neurodegeneration improving mitochondrial function and reducing mitophagy in mice.
  29. Clonal expansion dictates the efficacy of mitochondrial lineage tracing in single cells.
  30. A leigh syndrome mutation perturbs long-range energy coupling in respiratory complex I.
  31. Therapeutic Efficacy of Small Extracellular Vesicles Loaded with ROCK Inhibitor in Parkinson's Disease.
  32. First map of human brain mitochondria is 'groundbreaking' achievement.
  33. Application of rapid clinical exome sequencing technology in the diagnosis of critically ill pediatric patients with suspected genetic diseases.
  34. Identification of IDH3G, encoding the gamma subunit of mitochondrial isocitrate dehydrogenase, as a novel candidate gene for X-linked retinitis pigmentosa.
  35. Glycyl-tRNA sequestration is a unifying mechanism underlying GARS1-associated peripheral neuropathy.
  36. mitoXplorer 3.0, A Web Tool for Exploring Mitochondrial Dynamics in Single-cell RNA-seq Data.
  37. Patient-specific mutation of a contact site protein Tomm70 causes neurodegeneration in a zebrafish model.
  38. AlphaFold is running out of data - so drug firms are building their own version.
  39. Pharma companies collaborate on secure data sharing for AI.
  40. Unique Signatures of Highly Constrained Genes Across Publicly Available Genomic Databases.
  41. Vitamin A-Retinoic Acid Contributes to Muscle Stem Cell andMitochondrial Function Loss in Old Age.
  42. Haem biosynthesis regulates BCAA catabolism and thermogenesis in brown adipose tissue.
  43. Leveraging base excision repair for efficient adenine base editing of mitochondrial DNA.
  44. Serine mistranslation induces the integrated stress response through the P stalk.
  45. Transgenic iPSC Lines with Genetically Encoded MitoTimer to Study Mitochondrial Biogenesis in Dopaminergic Neurons with Tauopathy.
  46. Clioquinol induces mitochondrial toxicity in SH-SY5Y neuroblastoma cells by affecting the respiratory chain complex IV and OPA1 dynamin-like GTPase.
  47. Dnmt3a overexpression disrupts skeletal muscle homeostasis, promotes an aging-like phenotype, and reduces metabolic elasticity.
  48. The Role of Mitophagy in Cardiac Metabolic Remodeling of Heart Failure: Insights of Molecular Mechanisms and Therapeutic Prospects.
  49. TGFB signaling induces mitophagy via PLSCR3-mediated cardiolipin externalization in conjunction with a BNIP3L/NIX-, BNIP3-, and FUNDC1-dependent mechanism.
  50. Protocol for spatial metabolomics and isotope tracing in the mouse brain.
  51. Oxidation of retromer complex controls mitochondrial translation.
  52. Non-canonical hemoglobin: An updated review on its ubiquitous expression.
  53. Unraveling undiagnosed rare disease cases by HiFi long-read genome sequencing.
  54. Using implementation science to navigate the complexity of integrating genomics into healthcare.
  55. MAM kinases: physiological roles, related diseases, and therapeutic perspectives-a systematic review.
  56. High-throughput screening of human genetic variants by pooled prime editing.
  57. Methylmap: visualization of modified nucleotides for large cohort sizes.
  58. A BCAA-haem axis regulates brown fat function.
  59. Systematic high-throughput evaluation reveals FrCas9's superior specificity and efficiency for therapeutic genome editing.
  60. The Genetic Epidemiology of Parkinson's Disease view - Authors' reply.
  61. Protocol for generating transgene-free naive human induced pluripotent stem cells from somatic cells using modified Sendai viral system.
  62. Reductive Stress and Mitochondrial Dysfunction: The Hidden Link in Chronic Disease.
  1. Sci Rep. 2025 Mar 22. 15(1): 9971
    A cellular assay to determine the fusion capacity of MFN2 variants linked to Charcot-Marie-Tooth disease of type 2 A.
    Chloe Barsa, Julian Perrin, Claudine David, Arnaud Mourier, Manuel Rojo.
      Charcot-Marie-Tooth Disease (CMT) is an inherited peripheral neuropathy with two main forms: demyelinating CMT1 and axonal CMT2. The most frequent subtype of CMT2 (CMT2A) is linked to mutations of MFN2, encoding a ubiquitously expressed GTP-binding protein anchored to the mitochondrial outer membrane and essential for mitochondrial fusion. The use of Next-Generation Sequencing has led to the identification of increasing numbers of MFN2 variants, yet many of them remain of unknown significance, depriving patients of a clear diagnosis. In this work, we establish a cellular assay allowing to assess the impact of 12 known MFN2 variants linked to CMT2A on mitochondrial fusion. The functional analysis revealed that out of the 12 selected MFN2 mutations, only six exhibited reduced fusion activity. The classification of MFN2 variants according to the results of the functional assay revealed a correlation between the fusion capacity, the age at onset of CMT2A and computational variant effect predictions relying on the analysis of the protein sequence. The functional assay and the results obtained will assist and improve the classification of novel MFN2 variants identified in patients.
    Keywords:  CMT2A; Charcot–Marie-Tooth disease; MFN2; Mitochondrial dynamics; Mitochondrial fusion; Single nucleotide variants; Variant effect predictor; Variants of unknown significance
    DOI:  https://doi.org/10.1038/s41598-025-93702-1
  2. Nat Commun. 2025 Mar 21. 16(1): 2810
    Dependence of mitochondrial calcium signalling and dynamics on the disaggregase, CLPB.
    Donato D'Angelo, Víctor H Sánchez-Vázquez, Benjamín Cartes-Saavedra, Denis Vecellio Reane, Ryan R Cupo, Hilda Delgado de la Herran, Giorgia Ghirardo, James Shorter, Ron A Wevers, Saskia B Wortmann, Fabiana Perocchi, Rosario Rizzuto, Anna Raffaello, György Hajnóczky.
      Cells utilize protein disaggregases to avoid abnormal protein aggregation that causes many diseases. Among these, caseinolytic peptidase B protein homolog (CLPB) is localized in the mitochondrial intermembrane space and linked to human disease. Upon CLPB loss, MICU1 and MICU2, regulators of the mitochondrial calcium uniporter complex (mtCU), and OPA1, a main mediator of mitochondrial fusion, become insoluble but the functional outcome remains unclear. In this work we demonstrate that CLPB is required to maintain mitochondrial calcium signalling and fusion dynamics. CLPB loss results in altered mtCU composition, interfering with mitochondrial calcium uptake independently of cytosolic calcium and mitochondrial membrane potential. Additionally, OPA1 decreases, and aggregation occurs, accompanied by mitochondrial fragmentation. Disease-associated mutations in the CLPB gene present in skin fibroblasts from patients also display mitochondrial calcium and structural changes. Thus, mtCU and fusion activity are dependent on CLPB, and their impairments might contribute to the disease caused by CLPB variants.
    DOI:  https://doi.org/10.1038/s41467-025-57641-9
  3. Biomolecules. 2025 Mar 18. pii: 433. [Epub ahead of print]15(3):
    The Balance of MFN2 and OPA1 in Mitochondrial Dynamics, Cellular Homeostasis, and Disease.
    Paola Zanfardino, Alessandro Amati, Mirko Perrone, Vittoria Petruzzella.
      Mitochondrial dynamics, governed by fusion and fission, are crucial for maintaining cellular homeostasis, energy production, and stress adaptation. MFN2 and OPA1, key regulators of mitochondrial fusion, play essential roles beyond their structural functions, influencing bioenergetics, intracellular signaling, and quality control mechanisms such as mitophagy. Disruptions in these processes, often caused by MFN2 or OPA1 mutations, are linked to neurodegenerative diseases like Charcot-Marie-Tooth disease type 2A (CMT2A) and autosomal dominant optic atrophy (ADOA). This review explores the molecular mechanisms underlying mitochondrial fusion, the impact of MFN2 and OPA1 dysfunction on oxidative phosphorylation and autophagy, and their role in disease progression. Additionally, we discuss the divergent cellular responses to MFN2 and OPA1 mutations, particularly in terms of proliferation, senescence, and metabolic signaling. Finally, we highlight emerging therapeutic strategies to restore mitochondrial integrity, including mTOR modulation and autophagy-targeted approaches, with potential implications for neurodegenerative disorders.
    Keywords:  autophagy; mTOR signaling; mitochondria; mitochondrial dynamics; mitophagy; neurodegenerative diseases; oxidative phosphorylation; proliferation; senescence
    DOI:  https://doi.org/10.3390/biom15030433
  4. Biol Chem. 2025 Mar 28.
    Unclogging of the TOM complex under import stress.
    Joshua Jackson, Thomas Becker.
      Mitochondrial functions and biogenesis depend on the import of more than 1,000 proteins which are synthesized as precursor proteins on cytosolic ribosomes. Mitochondrial protein translocases sort the precursor proteins into the mitochondrial sub-compartments: outer and inner membrane, the intermembrane space and the matrix. The translocase of the outer mitochondrial membrane (TOM complex) constitutes the major import site for most of these precursor proteins. Defective protein translocases, premature folding of the precursor, or depletion of the membrane potential can cause clogging of the TOM channel by a precursor protein. This clogging impairs further protein import and leads to accumulation of precursor proteins in the cell that perturbates protein homeostasis, leading to proteotoxic stress in the cell. Therefore, unclogging of the translocon is critical for maintaining mitochondrial and cellular function. Ubiquitylation and AAA-ATPases play a central role in the extraction of the precursor proteins to deliver them to the proteasome for degradation. Here we summarize our understanding of the molecular mechanisms that remove such translocation-stalled precursor proteins from the translocation channel to regenerate the TOM complex for protein import.
    Keywords:  AAA ATPases; TOM complex; mitochondria; protein import; quality control; ubiquitylation
    DOI:  https://doi.org/10.1515/hsz-2025-0110
  5. Genes (Basel). 2025 Mar 17. pii: 347. [Epub ahead of print]16(3):
    Mitochondrial DNA Pathogenic Variants in Ophthalmic Diseases: A Review.
    Khaled K Abu-Amero, Bashaer Almadani, Shereen Abualkhair, Syed Hameed, Altaf A Kondkar, Andrea Sollazzo, Angeli Christy Yu, Massimo Busin, Giorgio Zauli.
      Mitochondria are vital organelles responsible for ATP production and metabolic regulation, essential for energy-intensive cells such as retinal ganglion cells. Dysfunction in mitochondrial oxidative phosphorylation or mitochondrial DNA (mtDNA) pathogenic variants can disrupt ATP synthesis, cause oxidative stress, and lead to cell death. This has profound implications for tissues such as the retina, optic nerve, and retinal pigment epithelium, which are dependent on robust mitochondrial function. In this review, we provide a comprehensive compilation of pathogenic variants in the mtDNA associated with various ophthalmic diseases, including Leber's hereditary optic neuropathy, chronic progressive external ophthalmoplegia, Leigh syndrome, mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes, among others. We highlight the genetic variants implicated in these conditions, their pathogenic roles, and the phenotypic consequences of mitochondrial dysfunction in ocular tissues. In addition to well-established mutations, we also discuss the emerging evidence of the role of mtDNA's variants in complex multifactorial diseases, such as non-arteritic anterior ischemic optic neuropathy, primary open-angle glaucoma, and age-related macular degeneration. The review aims to serve as a valuable resource for clinicians and researchers, providing a detailed overview of mtDNA pathogenic variants and their clinical significance in the context of mitochondrial-related eye diseases.
    Keywords:  DNA pathogenic variant; mitochondria; ophthalmology
    DOI:  https://doi.org/10.3390/genes16030347
  6. Biomolecules. 2025 Mar 14. pii: 416. [Epub ahead of print]15(3):
    The Pathogenesis of Very Long-Chain Acyl-CoA Dehydrogenase Deficiency.
    Shashwat Sharma, Matthew McKenzie.
      Living systems require energy to maintain their existence and perform tasks such as cell division. This energy is stored in several molecular forms in nature, specifically lipids, carbohydrates, and amino acids. At a cellular level, energy is extracted from these complex molecules and transferred to adenosine triphosphate (ATP) in the cytoplasm and mitochondria. Within the mitochondria, fatty acid β-oxidation (FAO) and oxidative phosphorylation (OXPHOS) are crucial metabolic processes involved in generating ATP, with defects in these pathways causing mitochondrial disease. Very long-chain acyl-CoA dehydrogenase deficiency (VLCADD) is a fatty acid β-oxidation disorder (FAOD) affecting 1 to 2 individuals per 100,000. Similar to other mitochondrial disorders, there is no cure for VLCADD, with symptomatic treatment comprising dietary management and supplementation with medium-chain fatty acids to bypass the enzyme deficiency. While this addresses the primary defect in VLCADD, there is growing evidence that other aspects of mitochondrial function are also affected in VLCADD, including secondary defects in OXPHOS function. Here, we review our current understanding of VLCADD with a focus on the associated biochemical and molecular defects that can disrupt multiple aspects of mitochondrial function. We describe the interactions between FAO proteins and the OXPHOS complexes and how these interactions are critical for maintaining the activity of both metabolic pathways. In particular, we describe what is now known about the protein-protein interactions between VLCAD and the OXPHOS supercomplex and how their disruption contributes to overall VLCADD pathogenesis.
    Keywords:  FAO; FAOD; OXPHOS; VLCAD; VLCADD; fatty acid oxidation disorder; fatty acid β-oxidation; oxidative phosphorylation; very long-chain acyl-CoA dehydrogenase; very long-chain acyl-CoA dehydrogenase deficiency
    DOI:  https://doi.org/10.3390/biom15030416
  7. Nature. 2025 Mar 26.
    A map of mitochondrial biology reveals the energy landscape of the human brain.
      
    Keywords:  Brain; Metabolism; Neuroscience
    DOI:  https://doi.org/10.1038/d41586-025-00872-z
  8. J Biol Chem. 2025 Mar 20. pii: S0021-9258(25)00282-0. [Epub ahead of print] 108433
    H2S remodels mitochondrial ultrastructure and destabilizes respiratory supercomplexes.
    David A Hanna, Brandon Chen, Yatrik M Shah, Oleh Khalimonchuk, Brian Cunniff, Ruma Banerjee.
      Mitochondrial form and function are intimately interconnected, responding to cellular stresses and changes in energy demand. Hydrogen sulfide, a product of amino acid metabolism, has dual roles as an electron transport chain substrate and complex IV (CIV) inhibitor, leading to a reductive shift, which has pleiotropic metabolic consequences. Luminal sulfide concentration in colon is high due to microbial activity, and in this study, we demonstrate that chronic sulfide exposure of colonocyte-derived cells leads to lower Mic60 and Mic19 expression that is correlated with a profound loss of cristae and lower mitochondrial networking. Sulfide-induced depolarization of the inner mitochondrial membrane activates Oma1-dependent cleavage of Opa1 and is associated with a profound loss of CI and CIV activities associated with respirasomes. Our study reveals a potential role for sulfide as an endogenous modulator of mitochondrial dynamics and suggests that this regulation is corrupted in hereditary or acquired diseases associated with elevated sulfide.
    Keywords:  Cristae; cristae; electron transport chain; hydrogen sulfide; mitochondrial dynamics; respirasome
    DOI:  https://doi.org/10.1016/j.jbc.2025.108433
  9. Int J Mol Sci. 2025 Mar 19. pii: 2768. [Epub ahead of print]26(6):
    A New Perspective on the Role of Alterations in Mitochondrial Proteins Involved in ATP Synthesis and Mobilization in Cardiomyopathies.
    Melissa Vázquez-Carrada, María Magdalena Vilchis-Landeros, Héctor Vázquez-Meza, Daniel Uribe-Ramírez, Deyamira Matuz-Mares.
      The heart requires a continuous energy supply to sustain its unceasing contraction-relaxation cycle. Mitochondria, a double-membrane organelle, generate approximately 90% of cellular energy as adenosine triphosphate (ATP) through oxidative phosphorylation, utilizing the electrochemical gradient established by the respiratory chain. Mitochondrial function is compromised by damage to mitochondrial DNA, including point mutations, deletions, duplications, or inversions. Additionally, disruptions to proteins associated with mitochondrial membranes regulating metabolic homeostasis can impair the respiratory chain's efficiency. This results in diminished ATP production and increased generation of reactive oxygen species. This review provides an overview of mutations affecting mitochondrial transporters and proteins involved in mitochondrial energy synthesis, particularly those involved in ATP synthesis and mobilization, and it examines their role in the pathogenesis of specific cardiomyopathies.
    Keywords:  ATP synthesis; cardiomyopathies; mitochondria; mitochondrial DNA; point mutations
    DOI:  https://doi.org/10.3390/ijms26062768
  10. Neurogenetics. 2025 Mar 26. 26(1): 38
    Mitochondrial DNA variants revealed by whole exome sequencing: from screening to diagnosis and follow-up.
    Sebastian Skoczylas, Tomasz Płoszaj, Karolina Gadzalska, Monika Gorządek, Paulina Jakiel, Ewa Juścińska, Maria Malarska, Magdalena Traczyk-Borszyńska, Hanna Biezynska, Magdalena Rychlik, Agata Pastorczak, Agnieszka Zmysłowska.
      Mutations in mitochondrial DNA play a crucial role in several diseases, but interpreting the clinical significance of mitochondrial DNA variants is challenging due to heteroplasmy, age-related loss of variants and evolving phenotypes. The aim of study was to identify mitochondrial pathogenic variants and explore their potential future association with specific phenotypes in patients during their lifetime, for both known and novel variants. We used a Python pipeline to analyse exome sequencing data from 418 patients (median age: 15 years; 52.9% males and 47.1% females), mostly diagnosed with neurological disorders, developmental and intellectual disabilities, behavioural and sensory disorders, cardiovascular and metabolic abnormalities, renal diseases and others. Screening identified 1,000 unique variants with heteroplasmy levels greater than 10% and 192 unique variants with 1-10% heteroplasmy, excluding hypervariable regions. Among these variants, four confirmed pathogenic variants were detected according to MITOMAP (m.1555 A > G, m.3243 A > G, m.9035T > C, and m.11778G > A), each identified in one patient. The application of pathogenicity and frequency criteria led to the identification of three unique variants and one in monozygotic twin sister with low levels of heteroplasmy, which were confirmed by next-generation sequencing. Finally, one of them, the variant m.15897G > A, was recognised as likely pathogenic (PP3, PS2). Our study highlights the complexity of diagnosing mitochondrial diseases associated with mtDNA mutations and emphasises the need for a comprehensive genotype-phenotype approach to correctly identify causal variants.
    Keywords:  Mitochondrial diseases; Whole exome sequencing; mtdna; mtdna variants
    DOI:  https://doi.org/10.1007/s10048-025-00820-z
  11. J Inherit Metab Dis. 2025 Mar;48(2): e70021
    Current Understanding of Pathogenic Mechanisms and Disease Models of Citrin Deficiency.
    Denis Lacabanne, Alice P Sowton, Bosco Jose, Edmund R S Kunji, Sotiria Tavoulari.
      Citrin deficiency (CD) is a complex mitochondrial disease with three different age-related stages: neonatal intrahepatic cholestasis caused by CD (NICCD), failure to thrive and dyslipidemia caused by CD (FTTDCD), and type II citrullinemia (CTLN2), recently renamed adolescent and adult CD (AACD). While highly prevalent in the Asian population, CD is pan-ethnic and remains severely underdiagnosed. The disease is caused by the dysfunction or absence of the mitochondrial aspartate/glutamate carrier 2 (AGC2/SLC25A13), also known as citrin. Citrin deficiency results in a direct impairment of the malate-aspartate shuttle and the urea cycle, with expected knock-on effects on a multitude of other metabolic pathways, leading to a complicated pathophysiology. Here, we discuss our current knowledge of the molecular mechanism of substrate transport by citrin, including recent advances  suggesting against its calcium regulation. We also discuss the different types of pathogenic variants found in CD patients and new insights into their pathogenic mechanisms. Additionally, we provide a summary and assessment of the efforts to develop preclinical models as well as treatments for the disease.
    Keywords:  citrin deficiency; disease models; mitochondrial transport; urea cycle disorders
    DOI:  https://doi.org/10.1002/jimd.70021
  12. JCEM Case Rep. 2025 Apr;3(4): luaf020
    Diverse Phenotypes of Mitochondrial Disease With Varying Levels of Heteroplasmy.
    Rebecca John, Aaron Chapla, Geeta Chacko, Sangeetha Yoganathan, Maya Mary Thomas, Nihal Thomas.
      Mitochondrial diseases have a wide spectrum of clinical presentations. Heteroplasmy, the presence of wild type and mutated mitochondrial deoxyribonucleic acid (DNA) in a single cell, is typical of mitochondrial disorders. It can show varying levels between cells of the same tissue, between organs in a single individual as well as between members of the same family. We describe below a woman who presented to us for management of pancreatic diabetes. Her daughter had a history of recurrent bouts of myopathy; evaluation was suggestive of having a mitochondrial etiology. Subsequently, mitochondrial genetic testing revealed positivity for m.3243A>G variant with a heteroplasmy of 45% in the blood in the daughter and 15% in the proband. We highlight how differences in the heteroplasmy and threshold levels among members of the same family resulted in a variable spectrum of clinical disease. Family screening of members identified with mitochondrial disease is of utmost significance to ensure early diagnosis and therapy.
    Keywords:  diabetes mellitus; heteroplasmy; mitochondrial diseases; muscular diseases
    DOI:  https://doi.org/10.1210/jcemcr/luaf020
  13. Diabetes Res Clin Pract. 2025 Mar 23. pii: S0168-8227(25)00143-3. [Epub ahead of print] 112129
    Mechanistic insights into impaired β-oxidation and its role in mitochondrial dysfunction: A comprehensive review.
    Amna Aqeel, Areeba Akram, Minahil Ali, Maryam Iqbal, Mehral Aslam, Rukhma Sattar, Fatima Iftikhar Shah.
      Mitochondria, also known as the powerhouse of cells, have an important role in cellular metabolism and energy production. However, during Mitochondrial Dysfunction (MD), it is known to generate reactive oxidative species and induce cellular apoptosis. A number of research findings have linked MD to various diseases, highlighting its critical role in maintaining health and contributing to disease development. In this regard, recent research has revealed that disruptions in lipid metabolism, especially in fatty acid oxidation, are significant contributors to MD. However, the precise mechanisms by which these defects lead to disease remain poorly understood. This review explores how disruptions in lipid metabolism are responsible for triggering oxidative stress, inflammation, and cellular damage, leading to impaired mitochondrial function. By examining specific fatty acid oxidation disorders, such as carnitine palmitoyltransferase deficiency, medium-chain acyl-CoA dehydrogenase deficiency, and very long-chain acyl-CoA dehydrogenase deficiency, this review aims to uncover the underlying molecular pathways connecting lipid metabolism to mitochondrial dysfunction. Furthermore, MD is a common underlying mechanism in a wide array of diseases, including neurodegenerative disorders, and metabolic syndromes. Understanding the mechanisms behind mitochondrial malfunction may aid in the development of tailored therapies to restore mitochondrial health and treat intricate health conditions.
    Keywords:  Fatty acid oxidation disorders (FAODs); Lipid metabolism; Mitochondrial dysfunction; Neurodegenerative diseases
    DOI:  https://doi.org/10.1016/j.diabres.2025.112129
  14. Anal Chem. 2025 Mar 25.
    OmicsCam Enables Trimodal Profiling of Mitochondrial Genome Editing.
    Guoqiang Zhou, Ting Li, Jingjing Du, Chengpeng He, Yu Yang, Guanju Chen, Jie Li, Bin Shen, Weilin Pu, Jingwei Zhang, Zhenglong Gu.
      Mitochondrial DNA (mtDNA) editing can generate cellular and animal models of mitochondrial genetic disorders and holds promise for future ex vivo and in vivo therapeutic applications. However, due to the quantitative nature of mitochondrion genetics, as more base-editing tools evolve, it is crucial to evaluate not only their efficiency and specificity on the sequence level but also the resulting molecular phenotypes. Here, we devised a novel Omics Carrier microcapsule, abbreviated as OmicsCam, that achieves homogeneous reactions within a heterogeneous carrier membrane, enabling highly efficient multistep biochemistry workflows. Incorporating magnetic beads into the carrier enables high-throughput automation. We demonstrated simultaneous trimodal assessment of mtDNA editing efficiency, postediting cellular transcriptome, and chromatin accessibility in minute cell samples containing as few as 25,000 cells. Applying OmicsCam to two TALE-DdCBE-edited human cell lines revealed that ND4 gene knockdown led to the downregulation of the mitochondrial oxidative phosphorylation pathway and changes in NF-Y transcription factor-associated histone modification pathways in the cell nucleus. Our study provides the most comprehensive analysis of mitochondrial gene editing efficiency and molecular phenotypes to date, which not only facilitates the establishment of mitochondrial genotype-molecular phenotype relationships but also helps assess the global safety of mitochondrial genome nucleases prior to clinical use.
    DOI:  https://doi.org/10.1021/acs.analchem.4c05251
  15. Adv Exp Med Biol. 2025 Mar 26.
    Organoids as 3D Models for Studying Exogenous Mitochondrial Transplantation.
    Ismail Eş, Oner Ulger.
      Mitochondria play a critical role in cellular communication, cell proliferation, and apoptosis, which make them essential to maintaining cellular health. Recently, mitochondrial transplantation has emerged as a promising therapeutic approach to treat conditions such as ischemia, neurodegenerative diseases, and cardiovascular disorders by restoring mitochondrial function in damaged cells. Despite its potential, understanding mitochondrial behavior in vivo remains challenging; however, organoid models, which are three-dimensional structures derived from stem cells that mimic human tissues, offer a solution to study mitochondrial function and transplantation strategies under controlled conditions. These models are particularly necessary in studies, as they can replicate disease conditions and consequently enable researchers to investigate mitochondrial dynamics and therapeutic integration. Developing organoid systems optimized for mitochondrial transplantation requires exploring factors that influence mitochondrial uptake, refining transplantation strategies, and understanding their role in cellular regeneration in order to advance in the field of mitochondrial research.
    Keywords:  3D cell culture models; Delivery methods in organoids; Mitochondria; Mitochondrial transplantation; Organoids
    DOI:  https://doi.org/10.1007/5584_2025_857
  16. Biology (Basel). 2025 Mar 09. pii: 279. [Epub ahead of print]14(3):
    Mitochondrial Proteome Reveals Metabolic Tuning by Restricted Insulin Signaling to Promote Longevity in Caenorhabditis elegans.
    Xuanxuan Guo, Jiuwei Lu, Long Miao, Enzhi Shen.
      Aging is a time-dependent process of functional decline influenced by genetic and environmental factors. Age-related mitochondrial changes remain incompletely understood. Here, we found that compared to the wild type, the mitochondria of long-lived daf-2 C. elegans maintain youthful morphology and function. Through quantitative proteomic analysis on isolated mitochondria, we identified 257 differentially expressed candidates. Analysis of these changed mitochondrial proteins reveals a significant upregulation of five key mitochondrial metabolic pathways in daf-2 mutants, including branched-chain amino acids (BCAA), reactive oxygen species (ROS), propionate, β-alanine, and fatty acids (FA), all of which are related to daf-2-mediated longevity. In addition, mitochondrial ribosome protein abundance slightly decreased in daf-2 mutants. A mild reduction in mitochondrial elongation factor G (gfm-1) by RNAi extends the lifespan of wild type while decreasing lipid metabolic process and cytoplasmic fatty acid metabolism, suggesting that proper inhibition of mitochondrial translation activity might be important for lifespan extension. Overall, our findings indicate that mitochondrial metabolic modulation contributes to the longevity of daf-2 mutants and further highlights the crucial role of mitochondria in aging.
    Keywords:  DAF-2; aging; metabolism; mitochondria; quantitative proteomics
    DOI:  https://doi.org/10.3390/biology14030279
  17. Curr Biol. 2025 Mar 24. pii: S0960-9822(25)00132-0. [Epub ahead of print]35(6): R218-R221
    Cell biology: A new dynamin superfamily protein remodels mitochondrial dynamics.
    Hassan Hashimi.
      Dynamin superfamily proteins mediate mitochondrial fusion in fungi and animals. A new study expands the taxonomic reach of this superfamily and provides insights into the roles these proteins play by investigating MfnL, a family member involved in trypanosomal mitochondrial dynamics. Importantly, MfnL occurs widely in eukaryotes and prokaryotes.
    DOI:  https://doi.org/10.1016/j.cub.2025.01.069
  18. Trends Cell Biol. 2025 Mar 26. pii: S0962-8924(25)00042-X. [Epub ahead of print]
    The fate of mitochondrial respiratory complexes in aging.
    Hanna Salmonowicz, Karolina Szczepanowska.
      While mitochondrial dysfunction is one of the canonical hallmarks of aging, it remains only vaguely defined. Its core feature embraces defects in energy-producing molecular machinery, the mitochondrial respiratory complexes (MRCs). The causes and consequences of these defects hold research attention. In this review, we assess the lifecycle of respiratory complexes, from biogenesis to degradation, and look closely at the mechanisms that could underpin their dysfunction in aged cells. We discuss how these processes could be altered by aging and expand on the fate of MRCs in age-associated pathologies. Given the complexity behind MRC maintenance and functionality, several traits could contribute to the phenomenon known as age-associated mitochondrial dysfunction. New advances will help us better understand the fate of this machinery in aging and age-related diseases.
    Keywords:  OXPHOS; age-associated diseases; dysfunction; mitochondria; protein complexes, aging hallmarks
    DOI:  https://doi.org/10.1016/j.tcb.2025.02.008
  19. Nat Cell Biol. 2025 Mar 21.
    In situ architecture of the human prohibitin complex.
    Felix Lange, Michael Ratz, Jan-Niklas Dohrke, Maxence Le Vasseur, Dirk Wenzel, Peter Ilgen, Dietmar Riedel, Stefan Jakobs.
      Prohibitins are a highly conserved family of proteins that have been implicated in a variety of functions including mitochondrial stress signalling and housekeeping, cell cycle progression, apoptosis, lifespan regulation and many others. The human prohibitins prohibitin 1 and prohibitin 2 have been proposed to act as scaffolds within the mitochondrial inner membrane, but their molecular organization has remained elusive. Here we determined the molecular organization of the human prohibitin complex within the mitochondrial inner membrane using an integrative structural biology approach combining quantitative western blotting, cryo-electron tomography, subtomogram averaging and molecular modelling. The proposed bell-shaped structure consists of 11 alternating prohibitin 1 and prohibitin 2 molecules. This study reveals an average of about 43 prohibitin complexes per crista, covering 1-3% of the crista membrane area. These findings provide a structural basis for understanding the functional contributions of prohibitins to the integrity and spatial organization of the mitochondrial inner membrane.
    DOI:  https://doi.org/10.1038/s41556-025-01620-1
  20. BMC Neurol. 2025 Mar 26. 25(1): 128
    Adult-onset Leigh syndrome with recurrent seizures and peripheral neuropathy due to the 9176T > C mutation: a case report and literature review.
    Yashi Liao, Yaxin Lai, Xinxin Chen, Shanshan Zhao.
       BACKGROUND: Leigh syndrome (LS) is an inherited form of mitochondrial encephalopathy associated with various gene mutations of the oxidative phosphorylation system, typically occurring in infancy or early childhood and resulting in disability or even death. However, few late-onset cases have been reported.
    OBJECTIVE: The objective of this case report was to investigate the radiological and clinical characteristics of an adult patient diagnosed with Leigh syndrome.
    CASE PRESENTATION: This article describes a patient who presented with recurrent generalized seizures, peripheral neuropathy and hypertension and was ultimately diagnosed with Leigh syndrome with a mitochondrial gene variant, c.9176T > C (p.Leu217Pro), in 20,315 of the MT-ATP6 gene. Here, we discuss the possible pathogenesis of its clinical manifestations according to the related literature and review the current therapeutic approaches and prognosis of LS.
    CONCLUSION: A possible diagnosis of LS should be taken into consideration when patients with characteristic neuroimaging findings of LS demonstrate recurrent seizures, peripheral neuropathy, or hypertension, and genetic analysis should be carried out for differential diagnosis.
    Keywords:  Basal ganglia; Hypertension; Leigh syndrome; MT-ATP6 gene; Mitochondrial diseases; Peripheral neuropathy; Recurrent seizure
    DOI:  https://doi.org/10.1186/s12883-025-04135-2
  21. Front Bioeng Biotechnol. 2025 ;13 1563701
    Mitochondria targeted nanoparticles for the treatment of mitochondrial dysfunction-associated brain disorders.
    Amy Claire Buck, Gerald J Maarman, Admire Dube, Soraya Bardien.
      Mitochondria play a significant role in several cellular activities and their function in health and disease has become an important area of research. Since the brain is a high-energy-demanding organ, it is particularly vulnerable to mitochondrial dysfunction. This has been implicated in several brain disorders including neurodegenerative, psychiatric and neurological disorders, e.g., Parkinson's disease and schizophrenia. Significant efforts are underway to develop mitochondria-targeting pharmaceutical interventions. However, the complex mitochondrial membrane network restricts the entry of therapeutic compounds into the mitochondrial matrix. Nanoparticles (NPs) present a novel solution to this limitation, while also increasing the stability of the therapeutic moieties and improving their bioavailability. This article provides a detailed overview of studies that have investigated the treatment of mitochondrial dysfunction in brain disorders using either targeted or non-targeted NPs as drug delivery systems. All the NPs showed improved mitochondrial functioning including a reduction in reactive oxygen species (ROS) production, an improvement in overall mitochondrial respiration and a reversal of toxin-induced mitochondrial damage. However, the mitochondrial-targeted NPs showed an advantage over the non-targeted NPs as they were able to improve or rescue mitochondrial dynamics and biogenesis, and they required a lower concentration of the in vivo therapeutic dosage of the drug load to show an effect. Consequently, mitochondria-targeted NPs are a promising therapeutic approach. Future studies should exploit advances in nanotechnology, neuroscience and chemistry to design NPs that can cross the blood-brain barrier and selectively target dysfunctional mitochondria, to improve treatment outcomes.
    Keywords:  brain disorders; mitochondria-targeted nanoparticles; mitochondrial dysfunction; nanomedicine; therapy
    DOI:  https://doi.org/10.3389/fbioe.2025.1563701
  22. Int J Mol Sci. 2025 Mar 16. pii: 2670. [Epub ahead of print]26(6):
    Multiple Inhibitory Mechanisms of DS16570511 Targeting Mitochondrial Calcium Uptake: Insights from Biochemical Analysis of Rat Liver Mitochondria.
    Akiko Yamada, Akira Watanabe, Atsushi Nara, Tsubasa Inokuma, Masatake Asano, Yasuo Shinohara, Takenori Yamamoto.
      Mitochondrial calcium (Ca2+) uptake plays a key role in mitochondrial physiology and disease development. This process is regulated by the mitochondrial calcium uniporter (MCU) complex. DS16570511 is a membrane-permeable drug that inhibits mitochondrial Ca2+ uptake, although its inhibitory mechanisms remain unclear. In this study, we evaluated the effects of DS16570511 on various mitochondrial functions through biochemical analyses. We found that DS16570511 affects multiple mitochondrial functions and exhibits variable potency in inhibiting individual processes. Specifically, DS16570511 not only inhibits MCU, its initially reported target, but also respiratory chain complexes and FoF1-adenosine triphosphatase/adenine nucleotide translocator, particularly respiratory chain complex II. Furthermore, the carboxyl group at the molecular terminus of DS16570511 plays a critical role in its inhibitory effects on mitochondrial Ca2+ uptake through respiratory chain complex II inhibition. These findings enhance our understanding of the mechanisms by which DS16570511 inhibits mitochondrial Ca2+ uptake and provide valuable insights for the clinical application of mitochondrial Ca2+ uptake inhibitors.
    Keywords:  DS16570511; inhibitor; mitochondrial calcium uniporter; rat liver
    DOI:  https://doi.org/10.3390/ijms26062670
  23. Epilepsia. 2025 Mar 22.
    Mitochondrial respiration defects in lymphoblast cell lines from patients with Dravet syndrome.
    Anna G Figueroa, Ruth E Fulton, Rajeswari Banerji, Kelly G Knupp, Manisha N Patel.
       OBJECTIVE: Dravet syndrome (DS) is a developmental and epileptic encephalopathy with early life intractable seizures and lifelong comorbidities. There is growing evidence linking energy metabolism to DS, from mitochondrial respiration deficits in skeletal muscle and fibroblasts from children with DS to responsiveness to ketogenic diets. Lymphoblast cell lines (LCLs) have revealed metabolic alterations in neurological disorders, suggesting their utility for studying systemic bioenergetics. In this pilot study, we used LCLs from patients with DS to evaluate energy metabolism.
    METHODS: LCLs were established from eight children with DS (DS-LCLs) and sex-/age-matched controls (control-LCLs). Extracellular flux analysis measured glycolytic function, mitochondrial respiration, and fatty acid oxidation (FAO). High-resolution respirometry was used to determine sites of mitochondrial respiration defects. Mitochondrial content and membrane potential were analyzed using high-content screening methods.
    RESULTS: DS-LCLs exhibit impaired bioenergetics, characterized by deficiencies in mitochondrial respiration with 25% lower baseline and adenosine triphosphate-linked respiration. Similarly, maximal mitochondrial capacity was 26% lower, leading to a 40% decrease in respiratory reserves. They exhibit a metabolic shift toward FAO, indicated by increased endogenous fatty acid utilization to counter cellular stress. Mitochondrial oxygen flux was impaired, with greatest deficiency in complex I, and reduced complex II activity. Leak respiration, mitochondrial content, membrane potential, and glycolytic function were unaffected.
    SIGNIFICANCE: LCLs from patients with DS reveal reduced mitochondrial respiratory capacity. These preliminary findings may enhance our understanding of energy metabolism in DS pathogenesis. Beyond helping identify new therapies, this model may noninvasively serve as a surrogate for evaluating metabolic function throughout a patient's life.
    Keywords:  developmental epileptic encephalopathy; energy metabolism; fatty acid oxidation; lymphoblast; mitochondria; patient‐derived cell line
    DOI:  https://doi.org/10.1111/epi.18382
  24. Adv Biol (Weinh). 2025 Mar 24. e2400597
    Ameliorating TIMM50 Loss Slows Senescence by Improving Mitochondrial Structure and Function.
    Amrita Nepalia, Deepak Kumar Saini.
      Mitochondrial dysfunction is an irrefutable hallmark of cellular senescence and aging. The dysfunction is marked by increased mitochondrial volume and reduced function, typified by low Adenosine Triphosphate (ATP) production and higher Reactive Oxygen Species (ROS) generation. Over the years, this dysfunction has been linked to Electron Transport Chain (ETC) malfunction and low NAD levels, augmented by poor mitophagy. However, the genetic regulation of mitochondrial dysfunction is still not clear. Here, using several senescence models, the first report on the role of the downregulation of a mitochondrial protein, Translocase of Inner Mitochondrial Membrane 50 (TIMM50), in senescence is presented. The downregulation of TIMM50 is also sufficient for triggering senescence through impaired mitochondrial function, characterized using a variety of mitochondrial function assessment assays. Reduced levels of TIMM50 initiated all the hallmarks of senescence, and overexpression significantly slowed senescence onset in response to an external trigger. The pathway analysis revealed that TIMM50 loss is mediated by the sirtuin1-dependent downregulation of CCAAT enhancer binding protein alpha (CEBPα), a transcription activator for TIMM50 expression. To establish the translational value of the observation, screening several potential anti-aging compounds revealed TIMM50 stabilizing and senescence-delaying effects only for verapamil and mitochondrial ROS quencher, Mito (2-(2,2,6,6-Tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl)triphenylphosphonium chloride (MitoTEMPO), both known anti-aging entities. Overall, TIMM50 is identified as the key mitochondrial protein whose downregulation is a critical step in initiating cellular senescence.
    Keywords:  TIMM50; aging; cellular senescence; mitochondria; sirtuin
    DOI:  https://doi.org/10.1002/adbi.202400597
  25. Anal Chem. 2025 Mar 27.
    Dual-Modality Imaging Unveil Inner Mitochondrial Membrane Viscosity and Respiratory Dynamics in Mitophagy.
    Fei Peng, Xiangnan Ai, Bin Hao, Xiaoyu Bu, Zixuan Zhao, Linshuai Yang, Baoxiang Gao.
      Mitophagy is a vital lysosome-dependent process that maintains mitochondrial integrity and cellular homeostasis, where respiration and inner mitochondrial membrane (IMM) viscosity play key roles. Despite its critical importance, achieving a high-resolution and dynamic visualization of respiration and IMM viscosity during mitophagy remains a significant challenge. In this study, we designed two innovative fluorescent probes: SiR-C8, a viscosity-sensitive rotor-type probe based on silicon-rhodamine, specifically targeting the IMM, and OR-ATP, a rhodamine-derived probe utilizing an intramolecular spirolactam structure to respond to mitochondrial ATP levels. Leveraging fluorescence intensity and lifetime dual-modality imaging, we successfully enabled the high-resolution, real-time monitoring of lysosome-dependent mitophagy. Remarkably, our results unveiled a progressive increase in IMM viscosity alongside a significant attenuation in mitochondrial respiration during mitophagy induced by starvation, carbonyl cyanide, m-chlorophenyl hydrazone (CCCP), and Oligomycin. Significantly, utilizing structured illumination microscopy super-resolution imaging, we have uncovered a novel mitochondrial quality control mechanism by which lysosomes selectively engulf locally damaged mitochondrial regions. This discovery provides novel insights into the intricate processes governing mitophagy and introduces an innovative platform for studying mitochondrial dynamics, dysfunction, and their implications for cellular homeostasis and pathology.
    DOI:  https://doi.org/10.1021/acs.analchem.5c00464
  26. Nature. 2025 Mar 26.
    Plasticity of the mammalian integrated stress response.
    Chien-Wen Chen, David Papadopoli, Krzysztof J Szkop, Bo-Jhih Guan, Mohammed Alzahrani, Jing Wu, Raul Jobava, Mais M Asraf, Dawid Krokowski, Anastasios Vourekas, William C Merrick, Anton A Komar, Antonis E Koromilas, Myriam Gorospe, Matthew J Payea, Fangfang Wang, Benjamin L L Clayton, Paul J Tesar, Ashleigh Schaffer, Alexander Miron, Ilya Bederman, Eckhard Jankowsky, Christine Vogel, Leoš Shivaya Valášek, Jonathan D Dinman, Youwei Zhang, Boaz Tirosh, Ola Larsson, Ivan Topisirovic, Maria Hatzoglou.
      An increased level of phosphorylation of eukaryotic translation initiation factor 2 subunit-α (eIF2α, encoded by EIF2S1; eIF2α-p) coupled with decreased guanine nucleotide exchange activity of eIF2B is a hallmark of the 'canonical' integrated stress response (c-ISR)1. It is unclear whether impaired eIF2B activity in human diseases including leukodystrophies2, which occurs in the absence of eIF2α-p induction, is synonymous with the c-ISR. Here we describe a mechanism triggered by decreased eIF2B activity, distinct from the c-ISR, which we term the split ISR (s-ISR). The s-ISR is characterized by translational and transcriptional programs that are different from those observed in the c-ISR. Opposite to the c-ISR, the s-ISR requires eIF4E-dependent translation of the upstream open reading frame 1 and subsequent stabilization of ATF4 mRNA. This is followed by altered expression of a subset of metabolic genes (for example, PCK2), resulting in metabolic rewiring required to maintain cellular bioenergetics when eIF2B activity is attenuated. Overall, these data demonstrate a plasticity of the mammalian ISR, whereby the loss of eIF2B activity in the absence of eIF2α-p induction activates the eIF4E-ATF4-PCK2 axis to maintain energy homeostasis.
    DOI:  https://doi.org/10.1038/s41586-025-08794-6
  27. Nature. 2025 Mar 26.
    A human brain map of mitochondrial respiratory capacity and diversity.
    Eugene V Mosharov, Ayelet M Rosenberg, Anna S Monzel, Corey A Osto, Linsey Stiles, Gorazd B Rosoklija, Andrew J Dwork, Snehal Bindra, Alex Junker, Ya Zhang, Masashi Fujita, Madeline B Mariani, Mihran Bakalian, David Sulzer, Philip L De Jager, Vilas Menon, Orian S Shirihai, J John Mann, Mark D Underwood, Maura Boldrini, Michel Thiebaut de Schotten, Martin Picard.
      Mitochondrial oxidative phosphorylation (OXPHOS) powers brain activity1,2, and mitochondrial defects are linked to neurodegenerative and neuropsychiatric disorders3,4. To understand the basis of brain activity and behaviour, there is a need to define the molecular energetic landscape of the brain5-10. Here, to bridge the scale gap between cognitive neuroscience and cell biology, we developed a physical voxelization approach to partition a frozen human coronal hemisphere section into 703 voxels comparable to neuroimaging resolution (3 × 3 × 3 mm). In each cortical and subcortical brain voxel, we profiled mitochondrial phenotypes, including OXPHOS enzyme activities, mitochondrial DNA and volume density, and mitochondria-specific respiratory capacity. We show that the human brain contains diverse mitochondrial phenotypes driven by both topology and cell types. Compared with white matter, grey matter contains >50% more mitochondria. Moreover, the mitochondria in grey matter are biochemically optimized for energy transformation, particularly among recently evolved cortical brain regions. Scaling these data to the whole brain, we created a backwards linear regression model that integrates several neuroimaging modalities11 to generate a brain-wide map of mitochondrial distribution and specialization. This model predicted mitochondrial characteristics in an independent brain region of the same donor brain. This approach and the resulting MitoBrainMap of mitochondrial phenotypes provide a foundation for exploring the molecular energetic landscape that enables normal brain function. This resource also relates to neuroimaging data and defines the subcellular basis for regionalized brain processes relevant to neuropsychiatric and neurodegenerative disorders. All data are available at http://humanmitobrainmap.bcblab.com .
    DOI:  https://doi.org/10.1038/s41586-025-08740-6
  28. Nat Commun. 2025 Mar 22. 16(1): 2839
    Mitochondria transplantation transiently rescues cerebellar neurodegeneration improving mitochondrial function and reducing mitophagy in mice.
    Shu-Jiao Li, Qian-Wen Zheng, Jie Zheng, Jin-Bao Zhang, Hui Liu, Jing-Jing Tie, Kun-Long Zhang, Fei-Fei Wu, Xiao-Dong Li, Shuai Zhang, Xin Sun, Yan-Ling Yang, Ya-Yun Wang.
      Cerebellar ataxia is the primary manifestation of cerebellar degenerative diseases, and mitochondrial dysfunction in Purkinje cells (PCs) plays a critical role in disease progression. In this study, we investigated the feasibility of mitochondria transplantation as a potential therapeutic approach to rescue cerebellar neurodegeneration and elucidate the associated mechanisms. We constructed a conditional Drp1 knockout model in PCs (PCKO mice), characterized by progressive ataxia. Drp1 knockout resulted in pervasive and progressive apoptosis of PCs and significant activation of surrounding glial cells. Mitochondrial dysfunction, which triggers mitophagy, is a key pathogenic factor contributing to morphological and functional damage in PCs. Transplanting liver-derived mitochondria into the cerebellum of 1-month-old PCKO mice improved mitochondrial function, reduced mitophagy, delayed apoptosis of PCs, and alleviated cerebellar ataxia for up to 3 weeks. These findings demonstrate that mitochondria transplantation holds promise as a therapeutic approach for cerebellar degenerative diseases.
    DOI:  https://doi.org/10.1038/s41467-025-58189-4
  29. Genome Biol. 2025 Mar 26. 26(1): 70
    Clonal expansion dictates the efficacy of mitochondrial lineage tracing in single cells.
    Xin Wang, Kun Wang, Weixing Zhang, Zhongjie Tang, Hao Zhang, Yuying Cheng, Da Zhou, Chao Zhang, Wen-Zhao Zhong, Qing Ma, Jin Xu, Zheng Hu.
       BACKGROUND: Mitochondrial DNA (mtDNA) variants hold promise as endogenous barcodes for tracking human cell lineages, but their efficacy as reliable lineage markers are hindered by the complex dynamics of mtDNA in somatic tissues.
    RESULTS: Here, we use computational modeling and single-cell genomics to thoroughly interrogate the origin and clonal dynamics of mtDNA variants across various biological settings. Our findings reveal that the majority of mtDNA variants which are specifically present in a cell subpopulation, termed subpopulation-specific variants, are pre-existing heteroplasmies in the first cell instead of de novo somatic mutations during divisions. Moreover, subpopulation-specific variants demonstrate limited discriminatory power among different genuine lineages under weak clonal expansion; however, certain subpopulation-specific variants with consistently high frequencies among a subpopulation are capable of faithfully labeling cell lineages in scenarios of stringent clonal expansion, such as strongly expanded T cell populations in diseased conditions and clonal hematopoiesis in aged individuals. Inspired by our simulations, we introduce a lineage informative score, facilitating the identification of reliable mitochondrial lineage tracing markers across different modalities of single-cell genomic data.
    CONCLUSIONS: Combining computational modeling and single-cell sequencing, our study reveals that the performance of mitochondrial lineage tracing is highly dependent on the extent of clonal expansion, which thus should be considered when applying mitochondrial lineage tracing.
    Keywords:  Clonal dynamics; Lineage tracing; Single-cell genomics; mtDNA variants
    DOI:  https://doi.org/10.1186/s13059-025-03540-7
  30. Chem Sci. 2025 Mar 21.
    A leigh syndrome mutation perturbs long-range energy coupling in respiratory complex I.
    Franziska Hoeser, Patricia Saura, Caroline Harter, Ville R I Kaila, Thorsten Friedrich.
      Respiratory complex I is a central enzyme of cellular energy metabolism that couples electron transfer with proton translocation across a biological membrane. In doing so, it powers oxidative phosphorylation that drives energy consuming processes. Mutations in complex I lead to severe neurodegenerative diseases in humans. However, the biochemical consequences of these mutations remain largely unknown. Here, we use the Escherichia coli complex I as a model to biochemically characterize the F124LMT-ND5 mutation found in patients suffering from Leigh syndrome. We show that the mutation drastically perturbs proton translocation and electron transfer activities to the same extent, despite the remarkable 140 Å distance between the mutated position and the electron transfer domain. Our molecular dynamics simulations suggest that the disease-causing mutation induces conformational changes that hamper the propagation of an electric wave through an ion-paired network essential for proton translocation. Our findings imply that malfunction of the proton translocation domain is entirely transmitted to the electron transfer domain underlining the action-at-a-distance coupling in the proton-coupled electron transfer of respiratory complex I.
    DOI:  https://doi.org/10.1039/d4sc04036h
  31. Pharmaceutics. 2025 Mar 13. pii: 365. [Epub ahead of print]17(3):
    Therapeutic Efficacy of Small Extracellular Vesicles Loaded with ROCK Inhibitor in Parkinson's Disease.
    Candy Carbajal, Myosotys Rodriguez, Florida Owens, Nicole Stone, Dileepkumar Veeragoni, Rebecca Z Fan, Kim Tieu, Nazira El-Hage.
      Background/Objectives: Parkinson's disease (PD) is a rapidly growing neurological disorder in the developed world, affecting millions over the age of 60. The decline in motor functions occurs due to a progressive loss of midbrain dopaminergic neurons, resulting in lowered dopamine levels and impaired muscle function. Studies show defective mitochondrial autophagy (or "mitophagy") links to PD. Rho-associated coiled-coil containing protein kinases (ROCK) 1 and ROCK2 are serine/threonine kinases, and their inhibition can enhance neuroprotection in PD by promoting mitophagy. Methods: We examine the effects of ROCK inhibitor SR3677, delivered via macrophage-derived small extracellular vesicles (sEVs) to Parkin Q311X(A) PD mouse models. sEVs with SR3677, administered intranasally, increased mitophagy gene expression, reduced inflammatory factors, and elevated dopamine levels in brain tissues. Results: ROCK2 expression decreased, showing the drug's inhibitory effect. sEV-SR3677 treatment was more effective than treatment with the drug alone, although sham EVs showed lower effects. This suggests that EV-SR3677 not only activates mitochondrial processes but also promotes the degradation of damaged mitochondria through autophagy. Mitochondrial functional assays and oxygen consumption in ex vivo glial cultures revealed that sEV-SR3677 significantly improved mitochondrial respiration compared to that in untreated or SR3677-only treated cells. Conclusion: We demonstrated the efficacy of ROCK2 inhibition on mitochondrial function via sEV-SR3677 in the PD mouse model, necessitating further studies to explore design challenges and mechanisms of sEV-SR3677 as mitochondria-targeted therapy for PD.
    Keywords:  Parkinson’s disease; ROCK; drug delivery system; extracellular vesicles; intranasal delivery
    DOI:  https://doi.org/10.3390/pharmaceutics17030365
  32. Nature. 2025 Mar 26.
    First map of human brain mitochondria is 'groundbreaking' achievement.
    Nora Bradford.
      
    Keywords:  Brain; Metabolism; Neuroscience
    DOI:  https://doi.org/10.1038/d41586-025-00848-z
  33. Front Genet. 2025 ;16 1526077
    Application of rapid clinical exome sequencing technology in the diagnosis of critically ill pediatric patients with suspected genetic diseases.
    Xuejun Ouyang, Yu Zhang, Tian Yu, Qian Zhang, Lei Xu, Victor Wei Zhang, Bin Wang.
       Purpose: This study evaluates the efficacy of rapid clinical exome sequencing (CES) and mitochondrial DNA (mtDNA) sequencing for diagnosing genetic disorders in critically ill pediatric patients.
    Methods: A multi-centre investigation was conducted, enrolling critically ill pediatric patients suspected of having genetic disorders from March 2019 to December 2020. Peripheral blood samples from patients and their parents were analyzed using CES (proband-parent) and mtDNA sequencing (proband-mother) based on Next-Generation Sequencing (NGS) technology.
    Results: The study included 44 pediatric patients (24 males, 20 females) with a median age of 27 days. The median turnaround time for genetic tests was 9.5 days. Genetic disorders were diagnosed in 25 patients (56.8%): 5 with chromosome microduplication/deletion syndromes (11.3%), 1 with UPD-related disease (2.3%), and 19 with monogenic diseases (43.2%). De novo variants were identified in nine patients (36.0%). A neonate was diagnosed with two genetic disorders due to a homozygous SLC25A20 variant and an MT-TL1 gene variation.
    Conclusion: Rapid genetic diagnosis is crucial for critically ill pediatric patients with suspected genetic disorders. CES and mtDNA sequencing offer precise and timely results, guiding treatment and reducing mortality and disability, making them suitable primary diagnostic tools.
    Keywords:  clinical exome sequencing; critical illness; mtDNA sequencing; pediatric; rapid genetic diagnosis
    DOI:  https://doi.org/10.3389/fgene.2025.1526077
  34. Genet Med. 2025 Mar 19. pii: S1098-3600(25)00065-6. [Epub ahead of print] 101418
    Identification of IDH3G, encoding the gamma subunit of mitochondrial isocitrate dehydrogenase, as a novel candidate gene for X-linked retinitis pigmentosa.
    Lorenzo Bianco, Julien Navarro, Christelle Michiels, Riccardo Sangermano, Christel Condroyer, Aline Antonio, Alessio Antropoli, Camille Andrieu, Emily M Place, Eric A Pierce, Said El Shamieh, Vasily Smirnov, Vasiliki Kalatzis, Luke Mansard, Anne-Françoise Roux, Béatrice Bocquet, José-Alain Sahel, Isabelle Meunier, Kinga M Bujakowska, Isabelle Audo, Christina Zeitz.
       PURPOSE: Retinitis pigmentosa (RP) is a genetically heterogeneous group of retinal degenerative disorders characterized by the loss rod and cone photoreceptors, leading to visual impairment and blindness. To date, X-linked RP has been associated with variants in three genes (RPGR, RP2, OFD1), while genetic defects at three loci (RP6, RP24, RP34) are yet unidentified. The aim of this study was to identify a novel candidate gene underlying X-linked RP.
    METHODS: Participants were identified from cohorts of genetically unsolved male individuals affected by RP, who underwent genome sequencing, exome sequencing, or candidate gene screening via direct Sanger sequencing at three referral centers. Specifically, two probands were identified at the National Reference Centre for Rare Retinal Diseases REFERET (Paris, France), two at Massachusetts Eye and Ear Hospital (Boston, MA, USA), and one at the National Reference Centre for Inherited Sensory Diseases MAOLYA (Montpellier, France). The pathogenicity of the identified variants was assessed using bioinformatic predictions, protein expression analyses, and mitochondrial function assays.
    RESULTS: We identified four rare single nucleotide variants in IDH3G (HGNC:5386), located at the RP34 locus on the X chromosome, as well as a complete gene deletion, in five unrelated male individuals affected with nonsyndromic RP. The variants segregated with the phenotype in all available family members. In all cases, the disease severity was intermediate. None had high myopia. IDH3G encodes the γ subunit of mitochondrial isocitrate dehydrogenase (IDH3), an enzyme involved in the citric acid cycle, which is expressed in the inner segments of photoreceptors. Variants in IDH3A and IDH3B, encoding the other subunits of IDH3, have already been associated with nonsyndromic autosomal recessive RP. Bioinformatic predictions and functional assays support a pathogenic role for the variants identified in this study, possibly though partial loss of enzymatic activity and mitochondrial function.
    CONCLUSIONS: Our findings suggest that variants in IDH3G are a novel cause of X-linked RP, possibly by impairing mitochondrial function and ultimately resulting in photoreceptor degeneration.
    Keywords:  IDH3G; X-linked; inherited retinal diseases; mitochondrial; retinitis pigmentosa
    DOI:  https://doi.org/10.1016/j.gim.2025.101418
  35. Nucleic Acids Res. 2025 Mar 20. pii: gkaf201. [Epub ahead of print]53(6):
    Glycyl-tRNA sequestration is a unifying mechanism underlying GARS1-associated peripheral neuropathy.
    Natalia Mora, Erik F J Slot, Vanessa Lewandowski, Maria P Menafra, Moushami Mallik, Pascal van Lith, Céline Sijlmans, Nick van Bakel, Zoya Ignatova, Erik Storkebaum.
      Dominantly inherited mutations in eight cytosolic aminoacyl-tRNA synthetase genes cause hereditary motor and sensory neuropathy, characterized by degeneration of peripheral motor and sensory axons. We previously identified a pathogenic gain-of-toxic function mechanism underlying peripheral neuropathy (PN) caused by heterozygous mutations in the GARS1 gene, encoding glycyl-tRNA synthetase (GlyRS). Specifically, PN-mutant GlyRS variants sequester tRNAGly, which depletes the cellular tRNAGly pool, leading to insufficient glycyl-tRNAGly available to the ribosome and consequently ribosome stalling at glycine codons. Given that GlyRS functions as a homodimer, a subset of PN-GlyRS mutations might alternatively cause peripheral neuropathy through a dominant negative loss-of-function mechanism. To explore this possibility, we here generated three novel PN-GlyRS Drosophila models expressing human PN-GlyRS (hGlyRS) variants that do not alter the overall GlyRS protein charge (S211F and H418R) or the single reported PN-GlyRS variant that renders the GlyRS protein charge more negative (K456Q). High-level expression of hGlyRS-K456Q did not induce peripheral neuropathy and the K456Q variant does not affect aminoacylation activity, suggesting that K456Q is not a pathogenic mutation. Expression of hGlyRS-S211F or hGlyRS-H418R in Drosophila did induce peripheral neuropathy and de novo protein synthesis defects. Genetic and biochemical evidence indicates that these phenotypes were attributable to tRNAGly sequestration rather than a dominant negative mechanism. Our data identify tRNAGly sequestration as a unifying pathogenic mechanism underlying PN-GlyRS. Thus, elevating tRNAGly levels may constitute a therapeutic approach for all PN-GlyRS patients, irrespective of their disease-causing mutation.
    DOI:  https://doi.org/10.1093/nar/gkaf201
  36. J Mol Biol. 2025 Feb 12. pii: S0022-2836(25)00070-1. [Epub ahead of print] 169004
    mitoXplorer 3.0, A Web Tool for Exploring Mitochondrial Dynamics in Single-cell RNA-seq Data.
    Margaux Haering, Andrea Del Bondio, Helene Puccio, Bianca H Habermann.
      Mitochondria are essential eukaryotic organelles, primarily recognized for their roles in ATP production, cellular metabolism and signalling. It is widely accepted that their structure, composition and function differ across cell types. However, little is known about mitochondrial variability within the same cell type. A comprehensive understanding of mitochondrial function and dynamics requires investigation at both, the individual cell type and single-cell resolution. Based on our mitoXplorer 2.0 web tool, we introduce mitoXplorer 3.0 with new features adapted for analysing single-cell sequencing data, focusing only on mitochondria. We developed a formatting script, scXplorer, which generates mitoXplorer 3.0 compatible files for data upload. The script generates pseudo-bulk transcriptomes of cell types from scRNA-seq data, enabling differential expression analysis and subsequent mitochondria-centric analysis with mitoXplorer classical interfaces. It also creates a single-cell expression matrix only containing mitochondria-associated genes (mito-genes), which can be analysed for cell-to-cell variability with novel, interactive interfaces created for mitoXplorer 3.0: these new interfaces help to identify sub-clusters of cell types based only on mito-genes and offer in-depth mitochondria-centric analysis of subpopulations. We demonstrate the usability and predictive power of mitoXplorer 3.0 through analysis of single-cell transcriptome data from a Spinocerebellar Ataxia Type 1 study. Our analysis identified several mitochondrial processes and genes significantly affected in SCA1 Purkinje cells, potentially contributing to mitochondrial dysfunction and subsequent Purkinje cell degeneration in this disease. MitoXplorer 3.0 is freely available at https://mitoxplorer3.ibdm.univ-amu.fr.
    Keywords:  data integration; mitoXplorer; mitochondria; single-nuclei sequencing; visual data mining
    DOI:  https://doi.org/10.1016/j.jmb.2025.169004
  37. Dis Model Mech. 2025 Mar 28. pii: dmm.052029. [Epub ahead of print]
    Patient-specific mutation of a contact site protein Tomm70 causes neurodegeneration in a zebrafish model.
    Vranda Garg, Ralf Heinrich, Roshan Priyarangana Perera, Till Ischebeck, Wiebke Möbius, Torben Ruhwedel, Patricia Scholz, Gabriela Salinas, Christian Dullin, Martin C Göpfert, Jacob Engelmann, Roland Dosch, Bart R H Geurten.
      Tomm70 is a receptor at the contact site between mitochondria and the endoplasmic reticulum, and has been identified as a risk gene for hereditary spastic paraplegia. Furthermore, de novo missense mutations in TOMM70 have been identified to cause neurological impairments in two unrelated patients. Here, we show that mutant zebrafish ruehreip25ca also harbor a missense mutation in tomm70, affecting the same conserved isoleucine residue as in one of the human patients. Using this model, we demonstrate how loss of Tomm70 function leads to impairment. At the molecular level, the mutation affects the interaction of Tomm70 with the endoplasmic reticulum protein Lam6, a known sterol transporter. At the neuronal level, the mutation impairs mitochondrial transport to the axons and dendrites, leading to demyelination of large calibre axons in the spinal cord. These neurodegenerative defects in zebrafish are associated with reduced endurance, swimming efficiency, and alterations in the C-start escape response, which correlate with decreased spiking in giant Mauthner neurons. Thus, in zebrafish, a mutation in the endoplasmic reticulum-mitochondria contact site protein Tomm70 recreates some of the neurodegenerative phenotypes characteristic of hereditary spastic paraplegia.
    Keywords:  Endoplasmic reticulum-mitochondria contact site (ER-MCS); Neurodegeneration; Tomm70; Zebrafish
    DOI:  https://doi.org/10.1242/dmm.052029
  38. Nature. 2025 Mar 27.
    AlphaFold is running out of data - so drug firms are building their own version.
    Ewen Callaway.
      
    Keywords:  Databases; Drug discovery; Machine learning; Structural biology
    DOI:  https://doi.org/10.1038/d41586-025-00868-9
  39. Nat Med. 2025 Mar 26.
    Pharma companies collaborate on secure data sharing for AI.
    Karen O'Leary.
      
    Keywords:  Drug discovery; Machine Learning; Technology
    DOI:  https://doi.org/10.1038/d41591-025-00021-z
  40. Genet Med. 2025 Mar 20. pii: S1098-3600(25)00060-7. [Epub ahead of print] 101413
    Unique Signatures of Highly Constrained Genes Across Publicly Available Genomic Databases.
    Klaus Schmitz-Abe, Qifei Li, Sunny Greene, Michela Borrelli, Shiyu Luo, Madesh C Ramesh, Pankaj B Agrawal.
       PURPOSE: Publicly available genomic databases are critical in understanding human genetic variation. They also provide unique insights into patterns of genetic constraints and their relationship with human disease.
    METHODS: We utilized one of the largest publicly available databases, gnomAD, to determine genes that are highly constrained for only loss of function (LoF), only missense, and both LoF/missense variants. We identified their unique signatures and explored their causal relationship with human diseases. Those genes were also evaluated for chromosomal location, tissue level expression, gene ontology analysis, and gene family categorization using multiple publicly available databases.
    RESULTS: We identified unique patterns of inheritance, protein size, and enrichment in distinct molecular pathways for those constrained genes associated with human disease. In addition, we identified genes that are currently not known to cause human disease, which may be excellent gene discovery candidates.
    CONCLUSIONS: We elucidate biological pathways of highly constrained genes that expand our understanding of critical cellular proteins. The findings can also advance research in rare diseases.
    Keywords:  ClinVar; HGMD; Z score; constrained genes; gnomAD
    DOI:  https://doi.org/10.1016/j.gim.2025.101413
  41. JCI Insight. 2025 Mar 25. pii: e183706. [Epub ahead of print]
    Vitamin A-Retinoic Acid Contributes to Muscle Stem Cell andMitochondrial Function Loss in Old Age.
    Paula M Fraczek, Pamela Duran, Benjamin A Yang, Valeria Ferre, Leanne Alawieh, Jesus A Castor-Macias, Vivian T Wong, Steve D Guzman, Celeste Piotto, Klimentini Itsani, Jacqueline A Larouche, Carlos A Aguilar.
      Adult stem cells decline in number and function in old age and identifying factors that can delay or revert age-associated adult stem cell dysfunction are vital for maintaining healthy lifespan. Here we show that Vitamin A, a micronutrient that is derived from diet and metabolized into retinoic acid, acts as an antioxidant and transcriptional regulator in muscle stem cells. We first show that obstruction of dietary Vitamin A in young animals drives mitochondrial and cell cycle dysfunction in muscle stem cells that mimics old age. Next, we pharmacologically targeted retinoic acid signaling in myoblasts and aged muscle stem cells ex vivo and in vivo and observed reductions in oxidative damage, enhanced mitochondrial function, and improved maintenance of quiescence through fatty acid oxidation. We next detected the receptor for vitamin A derived retinol, stimulated by retinoic acid 6 or Stra6, was diminished with muscle stem cell activation and in old age. To understand the relevance of Stra6 loss, we knocked down Stra6 and observed an accumulation of mitochondrial reactive oxygen species, as well as changes in mitochondrial morphology and respiration. These results demonstrate that Vitamin A regulates mitochondria and metabolism in muscle stem cells and highlight a unique mechanism connecting stem cell function with vitamin intake.
    Keywords:  Adult stem cells; Aging; Muscle; Muscle biology; Stem cells
    DOI:  https://doi.org/10.1172/jci.insight.183706
  42. Nat Metab. 2025 Mar 25.
    Haem biosynthesis regulates BCAA catabolism and thermogenesis in brown adipose tissue.
    Dylan J Duerre, Julia K Hansen, Steven V John, Annie Jen, Noah D Carrillo, Hoang Bui, Yutong Bao, Matias Fabregat, J Leon Catrow, Li-Yu Chen, Katherine A Overmyer, Evgenia Shishkova, Quentinn Pearce, Mark P Keller, Richard A Anderson, Vincent L Cryns, Alan D Attie, James E Cox, Joshua J Coon, Jing Fan, Andrea Galmozzi.
      The distinctive colour of brown adipose tissue (BAT) is attributed to its high content of haem-rich mitochondria. However, the mechanisms by which BAT regulates intracellular haem levels remain largely unexplored. Here we demonstrate that haem biosynthesis is the primary source of haem in brown adipocytes. Inhibiting haem biosynthesis results in an accumulation of the branched-chain amino acids (BCAAs) valine and isoleucine, owing to a haem-associated metabolon that channels BCAA-derived carbons into haem biosynthesis. Haem synthesis-deficient brown adipocytes display reduced mitochondrial respiration and lower UCP1 levels than wild-type cells. Although exogenous haem supplementation can restore intracellular haem levels and mitochondrial function, UCP1 downregulation persists. This sustained UCP1 suppression is linked to epigenetic regulation induced by the accumulation of propionyl-CoA, a byproduct of disrupted haem synthesis. Finally, disruption of haem biosynthesis in BAT impairs thermogenic response and, in female but not male mice, hinders the cold-induced clearance of circulating BCAAs in a sex-hormone-dependent manner. These findings establish adipose haem biosynthesis as a key regulator of thermogenesis and sex-dependent BCAA homeostasis.
    DOI:  https://doi.org/10.1038/s42255-025-01253-6
  43. Nat Biotechnol. 2025 Mar 25.
    Leveraging base excision repair for efficient adenine base editing of mitochondrial DNA.
    Yuhang Fan, Wenchao Xu, Bao-Qing Gao, Huichao Qin, Xiaoyi Wu, Jia Wei, Qingyang Ni, Lina Zhou, Jiangchao Xiang, Jing Wu, Bei Yang, Li Yang, Jia Chen.
      Transcription activator-like effector-linked deaminases (TALEDs) use their single-stranded DNA (ssDNA)-specific adenosine deaminase TadA8e to mediate A-to-G editing in mitochondrial DNA (mtDNA). The working mechanism of this process is unknown, hindering the development of more effective TALEDs. Here we reveal that TALED-mediated A-to-G editing relies on the formation of an ssDNA region through base excision repair (BER), which is triggered by double-stranded DNA-specific cytidine deaminase (DddA)-induced C-to-U deamination. We develop a series of enhanced TALEDs (eTALED6s) with increased editing efficiency by replacing DddA with the high-activity variant DddA6 and fusing human uracil DNA glycosylase to TadA8e. By further engineering TadA8e, the resulting eTALED6Rs induces efficient on-target editing with reduced bystander editing and off-target editing at the DNA and RNA levels. Lastly, we use eTALED6 and eTALED6R to install a pathogenic mutation in mtDNA. Revealing the mechanism of TALED-mediated A-to-G editing demonstrates that enhancing BER increases editing efficiency.
    DOI:  https://doi.org/10.1038/s41587-025-02608-w
  44. J Biol Chem. 2025 Mar 25. pii: S0021-9258(25)00296-0. [Epub ahead of print] 108447
    Serine mistranslation induces the integrated stress response through the P stalk.
    Hong Zhang, Jiqiang Ling.
      Aminoacyl-tRNA synthetases (aaRSs) are essential enzymes that support robust and accurate protein synthesis. A rapidly expanding number of studies show that mutations in aaRSs lead to multiple human diseases, including neurological disorders and cancer. How aaRS mutations impact human health is not fully understood. In particular, our knowledge of how aminoacylation errors affect stress responses and fitness in eukaryotic cells remains limited. The integrated stress response (ISR) is an adaptive mechanism in response to multiple stresses. However, chronic activation of the ISR contributes to the development of multiple diseases such as neuropathies. In this study, we show that Ser misincorporation into Ala and Thr codons, resulting from either aaRS editing defects or mutations in tRNAs, actives the ISR. We further demonstrate that activation of the ISR by Ser mistranslation does not depend on the accumulation of uncharged tRNAs, but rather requires the P stalk associated with the ribosome, implying that ribosome stalling and collision are involved. Our work highlights that certain types of aminoacylation errors can lead to chronic activation of the ISR, potentially affecting fitness and disease progression.
    Keywords:  AlaRS; ThrRS; Translational fidelity; stress response; tRNA misacylation
    DOI:  https://doi.org/10.1016/j.jbc.2025.108447
  45. Biomedicines. 2025 Feb 21. pii: 550. [Epub ahead of print]13(3):
    Transgenic iPSC Lines with Genetically Encoded MitoTimer to Study Mitochondrial Biogenesis in Dopaminergic Neurons with Tauopathy.
    Julia A Nadtochy, Sergey P Medvedev, Elena V Grigor'eva, Sophia V Pavlova, Julia M Minina, Anton V Chechushkov, Anastasia A Malakhova, Liudmila V Kovalenko, Suren M Zakian.
      Background: Tauopathy has been identified as a prevalent causative agent of neurodegenerative diseases, including frontotemporal dementia with parkinsonism-17 (FTDP-17). This rare hereditary neurodegenerative condition is characterised by the manifestation of parkinsonism and behavioural changes. The majority of cases of FTDP-17 are associated with mutations in the MAPT gene, which encodes the tau protein. MAPT mutations lead to disruption of the balance between 3R and 4R tau forms, which causes destabilisation of microtubules and impairment of cellular organelle functions, particularly mitochondrial dysfunction. The development of model systems and tools for studying the molecular, genetic, and biochemical mechanisms underlying FTDP-17 and testing therapies at the cellular level is an urgent necessity. Methods: In this study, we generated transgenic lines of induced pluripotent stem cells (iPSCs) from a patient carrying the pathogenic mutation c.2013T > G (rs63750756, p.N279K) of MAPT and a healthy donor. A doxycycline-controlled transgene of the genetically encoded biosensor MitoTimer was integrated into the AAVS1 locus of these cells. The MitoTimer biosensor allows for lifetime monitoring of the turnover of mitochondria in neuronal cells derived from directed iPSC differentiation. The fact that transcription of the transgene can be induced by doxycycline provides additional possibilities for pulse labelling of newly formed mitochondria. Results: Transgenic iPSC lines provide a unique tool to study the molecular and genetic mechanisms of FTDP-17 caused by the presence of the c.2013T > G (p.N279K) mutation, as well as to test potential drugs in vitro.
    Keywords:  CRISPR/Cas9; MAPT:c.2013T > G; MitoTimer biosensor; frontotemporal dementia with parkinsonism-17; induced pluripotent stem cells; mitochondrial dysfunction
    DOI:  https://doi.org/10.3390/biomedicines13030550
  46. FEBS Lett. 2025 Mar 24.
    Clioquinol induces mitochondrial toxicity in SH-SY5Y neuroblastoma cells by affecting the respiratory chain complex IV and OPA1 dynamin-like GTPase.
    Masato Katsuyama, Noriaki Arakawa, Takeshi Yaoi, En Kimura, Misaki Matsumoto, Kazumi Iwata, Atsushi Umemura, Chihiro Yabe-Nishimura.
      Clioquinol has been thought of as the causative drug of subacute myelo-optic neuropathy (SMON). The underlying mechanisms of clioquinol toxicity, however, have not been elucidated in detail. Here, we revealed that clioquinol (20 μm) suppressed the expression of SCO1 and SCO2 copper chaperones for mitochondrial respiratory chain Complex IV (cytochrome c oxidase) in SH-SY5Y neuroblastoma cells. The assembly of Complex IV components and Complex IV activity were suppressed in clioquinol-treated cells. Clioquinol (10-50 μm) decreased cellular ATP levels in glucose-free media. Clioquinol (10-50 μm) induced OMA1 mitochondrial protease-dependent degradation of the dynamin-related GTPase OPA1 and suppressed the expression of CHCHD10 and CHCHD2 involved in the maintenance of cristae structure. These results suggest that mitochondrial toxicity is one of the mechanisms of clioquinol-induced neuronal cell death.
    Keywords:  CHCHD10; OMA1; SCO2; SMON; clioquinol; mitochondria
    DOI:  https://doi.org/10.1002/1873-3468.70033
  47. iScience. 2025 Apr 18. 28(4): 112144
    Dnmt3a overexpression disrupts skeletal muscle homeostasis, promotes an aging-like phenotype, and reduces metabolic elasticity.
    Mamoru Oyabu, Yuto Ohira, Mariko Fujita, Kiyoshi Yoshioka, Runa Kawaguchi, Atsushi Kubo, Yukino Hatazawa, Hinako Yukitoshi, Huascar Pedro Ortuste Quiroga, Naoki Horii, Fumihito Miura, Hiromitsu Araki, Masaki Okano, Izuho Hatada, Hitoshi Gotoh, Tatsuya Yoshizawa, So-Ichiro Fukada, Yoshihiro Ogawa, Takashi Ito, Kengo Ishihara, Yusuke Ono, Yasutomi Kamei.
      Mammalian aging is reportedly driven by the loss of epigenetic information; however, its impact on skeletal muscle aging remains unclear. This study shows that aging mouse skeletal muscle exhibits increased DNA methylation, and overexpression of DNA methyltransferase 3a (Dnmt3a) induces an aging-like phenotype. Muscle-specific Dnmt3a overexpression leads to an increase in central nucleus-positive myofibers, predominantly in fast-twitch fibers, a shift toward slow-twitch fibers, elevated inflammatory and senescence markers, mitochondrial OXPHOS complex I reduction, and decreased basal autophagy. Dnmt3a overexpression resulted in reduced muscle mass and strength and impaired endurance exercise capacity with age, accompanied by an enhanced inflammatory signature. In addition, Dnmt3a overexpression reduced not only sensitivity to starvation-induced muscle atrophy but also the restorability from muscle atrophy. These findings suggest that increased DNA methylation disrupts skeletal muscle homeostasis, promotes an aging-like phenotype, and reduces muscle metabolic elasticity.
    Keywords:  Age; Epigenetics; Integrative aspects of cell biology; Model organism; Transcriptomics
    DOI:  https://doi.org/10.1016/j.isci.2025.112144
  48. J Cardiovasc Transl Res. 2025 Mar 26.
    The Role of Mitophagy in Cardiac Metabolic Remodeling of Heart Failure: Insights of Molecular Mechanisms and Therapeutic Prospects.
    Fangying Yan, Liwen Bao.
      Heart failure (HF) treatment remains one of the major challenges in cardiovascular disease management, and its pathogenesis requires further exploration. Cardiac metabolic remodeling is of great significance as a key pathological process in the progression of HF. The complex alterations of metabolic substrates and associated enzymes in mitochondria create a vicious cycle in HF. These changes lead to increased reactive oxygen species, altered mitochondrial Ca2+ handling, and the accumulation of fatty acids, contributing to impaired mitochondrial function. In this context, mitophagy plays a significant role in clearing damaged mitochondria, thereby maintaining mitochondrial function and preserving cardiac function by modulating metabolic remodeling in HF. This article aims to explore the role of mitophagy in cardiac metabolic remodeling in HF, especially in obesity cardiomyopathy, diabetic cardiomyopathy, and excessive afterload-induced heart failure, thoroughly analyze its molecular mechanisms, and review the therapeutic strategies and prospects based on the regulation of mitophagy.
    Keywords:  Cardiac metabolic remodeling; Heart failure; Mitophagy; Molecular mechanisms; Therapeutic prospects
    DOI:  https://doi.org/10.1007/s12265-025-10606-1
  49. Autophagy. 2025 Mar 22.
    TGFB signaling induces mitophagy via PLSCR3-mediated cardiolipin externalization in conjunction with a BNIP3L/NIX-, BNIP3-, and FUNDC1-dependent mechanism.
    Jiong Yan, Xin Chen, Swati Choksi, Zheng-Gang Liu.
      Selective clearance of damaged mitochondria through mitophagy is crucial for the maintenance of mitochondrial homeostasis. While mitophagy can be activated by various mitochondrial toxins, the physiologically relevant signal that triggers mitophagy is less studied. TGFB/TGFβ signaling has been linked to autophagic induction, but its specific role in mitophagy is not well understood. Here, we discovered a novel mitophagy induction paradigm stimulated by TGFB1. The mitophagic response is exclusively mediated by SMAD2, SMAD3, and SMAD4 underlying the TGFB receptor signaling. The transcriptional regulation activates genes involved in the canonical autophagic pathway which is required for the TGFB1-induced mitophagy. Moreover, TGFB1 signaling promotes mitophagic flux by upregulating PLSCR3 that externalizes cardiolipin in conjunction with the MAP1LC3/LC3/GABARAPs-interacting receptor proteins (BNIP3L/NIX, BNIP3, and FUNDC1)-dependent mechanism. Overall, our study characterized the essential components engaged in the TGFB1-induced mitophagy and demonstrated that TGFB is an important signal that induces mitophagy.
    Keywords:  ATG8; BNIP3; BNIP3L/NIX; PLSCR3; TGFB/TGFβ; mitophagy
    DOI:  https://doi.org/10.1080/15548627.2025.2483441
  50. STAR Protoc. 2025 Mar 27. pii: S2666-1667(25)00122-4. [Epub ahead of print]6(2): 103716
    Protocol for spatial metabolomics and isotope tracing in the mouse brain.
    Watit Sontising, Francine Yanchik-Slade, Carlos Rodriguez-Navas, Md Amir Hossen, Manuel Gonzalez-Rodriguez, Jake Vaynshteyn, Shinichi Yamaguchi, Mohamed Nazim Boutaghou, Isaac Marin-Valencia.
      Mass spectrometry imaging enables high-resolution spatial chemical mapping, yet its application for dynamic analysis with tracers poses challenges. Here, we present a protocol for spatial metabolomics and isotope tracing in the mouse brain. We describe steps for tracer administration, tissue collection, and cryosectioning. We then detail procedures for matrix application, ion identification, and data analysis. This protocol delivers high-quality spatial metabolomics data and is well suited for region-specific tracing analysis in the brain.
    Keywords:  mass spectrometry; metabolism; neuroscience
    DOI:  https://doi.org/10.1016/j.xpro.2025.103716
  51. Nature. 2025 Mar 26.
    Oxidation of retromer complex controls mitochondrial translation.
    Junbing Zhang, Md Yousuf Ali, Harrison Byron Chong, Pei-Chieh Tien, James Woods, Carolina Noble, Tristan Vornbäumen, Zehra Ordulu, Anthony P Possemato, Stefan Harry, Jay Miguel Fonticella, Lina Fellah, Drew Harrison, Maolin Ge, Neha Khandelwal, Yingfei Huang, Maëva Chauvin, Anica Tamara Bischof, Grace Marie Hambelton, Magdy Farag Gohar, Siwen Zhang, MinGyu Choi, Sara Bouberhan, Esther Oliva, Mari Mino-Kenudson, Natalya N Pavlova, Michael Lawrence, Justin F Gainor, Sean A Beausoleil, Nabeel Bardeesy, Raul Mostoslavsky, David Pépin, Christopher J Ott, Brian Liau, Liron Bar-Peled.
      Reactive oxygen species (ROS) underlie human pathologies including cancer and neurodegeneration1,2. However, the proteins that sense ROS levels and regulate their production through their cysteine residues remain ill defined. Here, using systematic base-editing and computational screens, we identify cysteines in VPS35, a member of the retromer trafficking complex3, that phenocopy inhibition of mitochondrial translation when mutated. We find that VPS35 underlies a reactive metabolite-sensing pathway that lowers mitochondrial translation to decrease ROS levels. Intracellular hydrogen peroxide oxidizes cysteine residues in VPS35, resulting in retromer dissociation from endosomal membranes and subsequent plasma membrane remodelling. We demonstrate that plasma membrane localization of the retromer substrate SLC7A1 is required to sustain mitochondrial translation. Furthermore, decreasing VPS35 levels or oxidation of its ROS-sensing cysteines confers resistance to ROS-generating chemotherapies, including cisplatin, in ovarian cancer models. Thus, we identify that intracellular ROS levels are communicated to the plasma membrane through VPS35 to regulate mitochondrial translation, connecting cytosolic ROS sensing to mitochondrial ROS production.
    DOI:  https://doi.org/10.1038/s41586-025-08756-y
  52. Redox Biol. 2025 Mar 18. pii: S2213-2317(25)00115-6. [Epub ahead of print]82 103602
    Non-canonical hemoglobin: An updated review on its ubiquitous expression.
    Emily C Reed, Jacob D Kim, Adam J Case.
      Hemoglobin, once thought to be exclusive to erythrocytes, has been identified to be expressed in various cell types over the past several decades. While hemoglobin's function within erythrocytes is primarily characterized as a gaseous transport molecule, its function within non-erythrocyte cells varies among different cell types, and in many cases, remains to be fully elucidated. Despite this variability, hemoglobin expression seems to broadly function as a redox modulator, whether it is involved in the hypoxic response, mitochondrial function, antioxidant balance or, like in erythrocytes, gas transport. This review provides an updated summary of the most recent discoveries of hemoglobin in non-erythrocyte cells. While discussing the function and regulation of this ubiquitous protein, we additionally compare these cell-specific details to identify commonalities throughout the diverse group of hemoglobin-expressing cells. Lastly, we discuss potential implications of non-canonical hemoglobin in various disease states such neurodegeneration, autoimmune disorders, psychological trauma, and hemoglobinopathies, while providing future directions for hemoglobin research.
    Keywords:  Antioxidant; Hbα; Hbβ; Hemoglobin; Mitochondria; Reactive oxygen species; Redox
    DOI:  https://doi.org/10.1016/j.redox.2025.103602
  53. Genome Res. 2025 Mar 26.
    Unraveling undiagnosed rare disease cases by HiFi long-read genome sequencing.
    Solve-RD DITF-ITHACA
      Solve-RD is a pan-European rare disease (RD) research program that aims to identify disease-causing genetic variants in previously undiagnosed RD families. We utilized 10-fold coverage HiFi long-read sequencing (LRS) for detecting causative structural variants (SVs), single-nucleotide variants (SNVs), insertion-deletions (indels), and short tandem repeat (STR) expansions in previously studied RD families without a clear molecular diagnosis. Our cohort includes 293 individuals from 114 genetically undiagnosed RD families selected by European Reference Network (ERN) experts. Of these, 21 families were affected by so-called "unsolvable" syndromes for which genetic causes remain unknown and for which prior testing was not a prerequisite. The remaining 93 families had at least one individual affected by a rare neurological, neuromuscular, or epilepsy disorder without a genetic diagnosis despite extensive prior testing. Clinical interpretation and orthogonal validation of variants in known disease genes yielded 12 novel genetic diagnoses due to de novo and rare inherited SNVs, indels, SVs, and STR expansions. In an additional five families, we identified a candidate disease-causing variant, including an MCF2/FGF13 fusion and a PSMA3 deletion. However, no common genetic cause was identified in any of the "unsolvable" syndromes. Taken together, we found (likely) disease-causing genetic variants in 11.8% of previously unsolved families and additional candidate disease-causing SVs in another 5.4% of these families. In conclusion, our results demonstrate the potential added value of HiFi long-read genome sequencing in undiagnosed rare diseases.
    DOI:  https://doi.org/10.1101/gr.279414.124
  54. Nat Med. 2025 Mar 21.
    Using implementation science to navigate the complexity of integrating genomics into healthcare.
    Stephanie Best, Jeffrey Braithwaite, Ilias Goranitis, Danya F Vears, Monica Ferrie, Clara L Gaff, Andrew J Mallett, Tiffany Boughtwood, Kathryn N North, Zornitza Stark.
      
    DOI:  https://doi.org/10.1038/s41591-025-03588-9
  55. Cell Mol Biol Lett. 2025 Mar 28. 30(1): 35
    MAM kinases: physiological roles, related diseases, and therapeutic perspectives-a systematic review.
    A Anjana Mohan, Priti Talwar.
      Mitochondria-associated membranes (MAMs) are tethering regions amid the membranes of the endoplasmic reticulum (ER) and mitochondria. They are a lipid raft-like structure occupied by various proteins that facilitates signal transduction between the two organelles. The MAM proteome participates in cellular functions such as calcium (Ca2+) homeostasis, lipid synthesis, ER stress, inflammation, autophagy, mitophagy, and apoptosis. The human kinome is a superfamily of homologous proteins consisting of 538 kinases. MAM-associated kinases participate in the aforementioned cellular functions and act as cell fate executors. Studies have proved the dysregulated kinase interactions in MAM as an etiology for various diseases including cancer, diabetes mellitus, neurodegenerative diseases, cardiovascular diseases (CVDs), and obesity. Several small kinase inhibitory molecules have been well explored as promising drug candidates in clinical trials with an accelerating impact in the field of precision medicine. This review narrates the physiological actions, pathophysiology, and therapeutic potential of MAM-associated kinases with recent updates in the field.
    Keywords:  Cancer; Diabetes; ER stress; Kinases; MAM; Mitophagy; Neurodegenerative disease; Therapeutics
    DOI:  https://doi.org/10.1186/s11658-025-00714-w
  56. Cell Genom. 2025 Mar 18. pii: S2666-979X(25)00070-9. [Epub ahead of print] 100814
    High-throughput screening of human genetic variants by pooled prime editing.
    Michael Herger, Christina M Kajba, Megan Buckley, Ana Cunha, Molly Strom, Gregory M Findlay.
      Multiplexed assays of variant effect (MAVEs) enable scalable functional assessment of human genetic variants. However, established MAVEs are limited by exogenous expression of variants or constraints of genome editing. Here, we introduce a pooled prime editing (PE) platform to scalably assay variants in their endogenous context. We first improve efficiency of PE in HAP1 cells, defining optimal prime editing guide RNA (pegRNA) designs and establishing enrichment of edited cells via co-selection. We next demonstrate negative selection screening by testing over 7,500 pegRNAs targeting SMARCB1 and observing depletion of efficiently installed loss-of-function (LoF) variants. We then screen for LoF variants in MLH1 via 6-thioguanine selection, testing 65.3% of all possible SNVs in a 200-bp region including exon 10 and 362 non-coding variants from ClinVar spanning a 60-kb region. The platform's overall accuracy for discriminating pathogenic variants indicates that it will be highly valuable for identifying new variants underlying diverse human phenotypes across large genomic regions.
    Keywords:  MAVE; MLH1; PE; SMARCB1; functional genomics; genome-editing technology; multiplexed assay of variant effect; precision medicine; prime editing; saturation mutagenesis
    DOI:  https://doi.org/10.1016/j.xgen.2025.100814
  57. BMC Bioinformatics. 2025 Mar 26. 26(1): 91
    Methylmap: visualization of modified nucleotides for large cohort sizes.
    Elise Coopman, Svenn D'Hert, Rosa Rademakers, Wouter De Coster.
       BACKGROUND: Over the years, there has been growing interest in epigenetics, where nucleotide modifications are increasingly recognized for their roles in health and disease. Understanding methylation patterns at the nucleotide level has become pivotal for advancing this field. However, visualizing these modifications, particularly in cohorts of more than a few individuals, remains a challenge.
    RESULTS: Here, we present methylmap, a tool developed to visualize modified nucleotide frequencies for regions of interest, specifically optimized for cohort sizes with more than a few individuals. Furthermore, methylmap features the visualization of the haplotype-specific methylation status of 226 individuals of the 1000 Genomes Project ONT Sequencing Consortium, sequenced using the Oxford Nanopore Technologies PromethION. This resource provides the research community with a comprehensive and complete overview of genome-wide methylation patterns.
    CONCLUSIONS: Methylmap offers an easy-to-use platform to facilitate epigenetic research. It is available both as a web application at https://methylmap.bioinf.be and as a command-line tool through Bioconda and PyPI. As such, we provide a valuable resource for advancing the understanding of epigenetic modifications in health and disease.
    Keywords:  1000 Genomes; Epigenetics; Methylation; Modifications; ONT; Visualization
    DOI:  https://doi.org/10.1186/s12859-025-06106-3
  58. Nat Metab. 2025 Mar 25.
    A BCAA-haem axis regulates brown fat function.
      
    DOI:  https://doi.org/10.1038/s42255-025-01266-1
  59. Sci Adv. 2025 Mar 28. 11(13): eadu7334
    Systematic high-throughput evaluation reveals FrCas9's superior specificity and efficiency for therapeutic genome editing.
    Rui Tian, Xun Tian, Meiying Yang, Yuping Song, Tingting Zhao, Chaoyue Zhong, Wei Zhu, Ping Zhou, Zhiqiang Han, Zheng Hu.
      CRISPR-Cas9 systems have revolutionized genome editing, but the off-target effects of Cas9 limit its use in clinical applications. Here, we systematically evaluate FrCas9, a variant from Faecalibaculum rodentium, for cell and gene therapy (CGT) applications and compare its performance to SpCas9 and OpenCRISPR-1. OpenCRISPR-1 is a CRISPR system synthesized de novo using large language models (LLMs) but has not yet undergone systematic characterization. Using AID-seq, Amplicon sequencing, and GUIDE-seq, we assessed the on-target activity and off-target profiles of these systems across multiple genomic loci. FrCas9 demonstrated higher on-target efficiency and substantially fewer off-target effects than SpCas9 and OpenCRISPR-1. Furthermore, TREX2 fusion with FrCas9 reduced large deletions and translocations, enhancing genomic stability. Through screening of 1903 sgRNAs targeting 21 CGT-relevant genes using sequential AID-seq, Amplicon sequencing, and GUIDE-seq analysis, we identified optimal sgRNAs for each gene. Our high-throughput screening platform highlights FrCas9, particularly in its TREX2-fused form, as a highly specific and efficient tool for precise therapeutic genome editing.
    DOI:  https://doi.org/10.1126/sciadv.adu7334
  60. Lancet Neurol. 2025 Apr;pii: S1474-4422(25)00073-0. [Epub ahead of print]24(4): 285-286
    The Genetic Epidemiology of Parkinson's Disease view - Authors' reply.
    all the authors
      
    DOI:  https://doi.org/10.1016/S1474-4422(25)00073-0
  61. STAR Protoc. 2025 Mar 21. pii: S2666-1667(25)00106-6. [Epub ahead of print]6(2): 103700
    Protocol for generating transgene-free naive human induced pluripotent stem cells from somatic cells using modified Sendai viral system.
    Kaho Washizu, Shinya Yamanaka, Akira Kunitomi.
      Sendai virus (SeV) vector represents a powerful tool for generating naive and primed human induced pluripotent stem cells (iPSCs) from somatic cells. Here, we present a protocol for the generation of transgene-free naive human iPSCs from human dermal fibroblasts (HDFs) and human peripheral mononuclear cells (PBMCs) using a modified SeV vector system. We describe steps for thawing the HDFs or PBMCs, reseeding HDFs, SeV vector infection, reseeding the SeV-infected HDFs or PBMCs on an irradiated mouse embryonic fibroblast (iMEF) plate, switching to t2iLGö+Y medium, passaging the generated naive iPSCs, removing the SeV vectors, and cryopreserving the naive iPSCs. For complete details on the use and execution of this protocol, please refer to Kunitomi et al.1,2.
    Keywords:  Biotechnology and bioengineering; Cell biology; Cell culture; Stem cells
    DOI:  https://doi.org/10.1016/j.xpro.2025.103700
  62. Free Radic Biol Med. 2025 Mar 22. pii: S0891-5849(25)00179-0. [Epub ahead of print]
    Reductive Stress and Mitochondrial Dysfunction: The Hidden Link in Chronic Disease.
    Joseph Mercola.
      Conventional theories of oxidative stress have long focused on the deleterious consequences of excessive reactive oxygen species (ROS) formation. However, growing evidence reveals that an overload of reducing equivalents-termed reductive stress-may be equally pivotal in driving mitochondrial dysfunction and chronic disease. In this paradigm, abnormally high concentrations of NADH and NADPH create an electron "traffic jam" in the mitochondrial electron transport chain (ETC), leading to partial inhibition or reverse electron flow at upstream complexes. Paradoxically, this hyper-reduced environment promotes ROS generation by increasing electron leakage to molecular oxygen, thereby intensifying oxidative damage to lipids, proteins, and mitochondrial DNA. This review explores the intertwined nature of reductive and oxidative stress, showing how a surplus of reducing equivalents can potentiate metabolic derangements in conditions such as type 2 diabetes, nonalcoholic fatty liver disease, and neurodegenerative disorders. We discuss common drivers of reductive overload, including chronic hyperglycemia, high-fat diets, and specific dietary patterns-particularly those enriched in polyunsaturated omega-6 fatty acids-that inundate mitochondria with electron donors. We also highlight emerging evidence that targeted assessment of redox biomarkers (e.g., lactate:pyruvate, β-hydroxybutyrate:acetoacetate ratios) can provide clinically relevant indicators of reductive stress. Finally, we examine how novel therapeutic strategies can address the underlying reductive imbalance, from rational nutrient modulation to pharmacologic interventions that restore NAD+ levels or optimize ETC flux. Recognizing reductive stress as a critical inflection point in mitochondrial pathophysiology underscores the need for a refined redox framework, one that moves beyond conventional oxidative paradigms to embrace the full spectrum of redox dysregulation in chronic degenerative disease.
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2025.03.029
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