bims-mitdis Biomed News
on Mitochondrial disorders
Issue of 2025–06–29
sixty papers selected by
Catalina Vasilescu, Helmholz Munich



  1. J Cell Biol. 2025 Aug 04. pii: e202408025. [Epub ahead of print]224(8):
      Mutations in the E3 ubiquitin ligase Parkin gene have been linked to early onset Parkinson's disease. Besides many other roles, Parkin is involved in clearance of damaged mitochondria via mitophagy-a process of particular importance in dopaminergic neurons. Upon mitochondrial damage, Parkin accumulates at the outer mitochondrial membrane and is activated, leading to ubiquitination of many mitochondrial substrates and recruitment of mitophagy effectors. While the activation mechanisms of autoinhibited Parkin have been extensively studied, it remains unknown how Parkin recognizes its substrates for ubiquitination. Here, we characterize a conserved region in the flexible linker between the Ubl and RING0 domains of Parkin, which is indispensable for Parkin interaction with the mitochondrial GTPase Miro1. Our results may explain fast kinetics of Miro1 ubiquitination by Parkin in recombinant assays and provide a biochemical explanation for Miro1-dependent Parkin recruitment to the mitochondrial membrane observed in cells. Our findings are important for understanding mitochondrial homeostasis and may inspire new therapeutic avenues for Parkinson's disease.
    DOI:  https://doi.org/10.1083/jcb.202408025
  2. Front Cardiovasc Med. 2025 ;12 1582219
      Mitochondrial depletion syndrome (MTDPS) is a heterogeneous group of genetic disorders characterized by a significant reduction in mitochondrial DNA (mtDNA) copy number, leading to the impaired mitochondrial function. The pathogenesis of MTDPS includes impaired mtDNA replication, damaged nucleotide metabolism and dysregulated mitochondrial dynamics. Due to its high energy demands, the heart is sensitive to the mitochondrial dysfunction. And the energy deficiency caused by the MTDPS contributes to the development of the mitochondrial cardiomyopathy. In this review, we summarize the cardiac phenotypes in the MTDPS, and the role of the mitochondrial injury in the myocardial damage. In specific, the association of the MTDPS-causing genes and their cardiac phenotypes are detailed. Moreover, the current treatment strategies for MTDPS are summarized. This review aims to integrate the current knowledge on the MTDPS and its cardiac phenotypes in order to provide insights for the further research and the clinic management.
    Keywords:  cardiomyopathy; mitochondrial DNA depletion syndrome; mitochondrial damage; mitochondrial dynamics; mitochondrial dysfunction; mtDNA replication; nucleotide metabolism
    DOI:  https://doi.org/10.3389/fcvm.2025.1582219
  3. Mol Cell. 2025 Jun 20. pii: S1097-2765(25)00472-1. [Epub ahead of print]
      Mitochondrial small open reading frame (ORF)-encoded microproteins (SEPs) are key regulators and components of the electron transport chain (ETC). Although ETC complex I assembly is tightly coupled to nutrient availability, including serine, the coordinating mechanism remains unknown. A genome-wide CRISPR screen targeting SEPs revealed that deletion of the LINC00493-encoded microprotein SMIM26 sensitizes cells to one-carbon restriction. SMIM26 interacts with mitochondrial serine transporters SFXN1/2 and the mitoribosome, forming a functional triad that facilitates translation of the complex I subunit mt-ND5. SMIM26 loss impairs serine import, reduces folate intermediates, and disrupts key mitochondrial tRNA modifications (τm5U and τm5s²U), resulting in ND5 translation failure and complex I deficiency. SMIM26 deletion is embryonic lethal in mice and impedes tumor growth in a xenograft model of folate-dependent acute myeloid leukemia. These findings define SMIM26 as a critical integrator of one-carbon flux and complex I biogenesis and establish a paradigm for localized mitochondrial translation through transporter-ribosome interactions.
    Keywords:  complex I; electron transport chain; micropeptides; mitochondria; mitochondrial translation; one-carbon pathway; oxidative phosphorylation; small ORF-encoded peptides
    DOI:  https://doi.org/10.1016/j.molcel.2025.05.033
  4. Nat Commun. 2025 Jun 25. 16(1): 5388
      S-adenosylmethionine (SAM) is the principal methyl donor in cells and is essential for mitochondrial gene expression, influencing RNA modifications, translation, and ribosome biogenesis. Using direct long-read RNA sequencing in mouse tissues and embryonic fibroblasts, we show that processing of the mitochondrial ribosomal gene cluster fails in the absence of mitochondrial SAM, leading to an accumulation of unprocessed precursors. Proteomic analysis of ribosome fractions revealed these precursors associated with processing and assembly factors, indicating stalled biogenesis. Structural analysis by cryo-electron microscopy demonstrated that SAM-dependent methylation is required for peptidyl transferase centre formation during mitoribosome assembly. Our findings identify a critical role for SAM in coordinating mitoribosomal RNA processing and large subunit maturation, linking cellular methylation potential to mitochondrial translation capacity.
    DOI:  https://doi.org/10.1038/s41467-025-60977-x
  5. Nucleic Acid Ther. 2025 Jun 18.
      We present a general method for in-cellulo delivery of 2'-O-methyl (2'-OMe) RNA oligonucleotides (oligos) to mitochondria for antisense applications, with potential for implementation in other mitochondrial DNA (mtDNA)-targeted therapies. Exosomes, which are nanoscale, naturally occurring extracellular vesicles (EVs), have been employed for biotechnology applications in oligonucleotide delivery in recent years. We discovered that exosomes from fetal bovine serum (FBS) can be used as a simple and biologically compatible delivery agent of 2'-OMe RNA antisense oligonucleotides to cellular mitochondria, leading to target protein knockdown. While most RNA interference and antisense mechanisms occur in the cytoplasm or nucleus, the need for mitochondrial targeting has become increasingly apparent. Mitochondrial disease describes a variety of currently incurable syndromes that especially affect organs requiring significant energy including the muscles, heart, and brain. Many of these syndromes result from mutations in mtDNA, which codes for the 13 proteins of the oxidative phosphorylation system and are thus often implicated in inherited metabolic disorders.
    Keywords:  2′-OMe RNA; antisense oligonucleotides; exosome-based delivery; extracellular vesicles; fetal bovine serum; mitochondrial localization
    DOI:  https://doi.org/10.1089/nat.2024.0067
  6. Exp Mol Med. 2025 Jun 24.
      Barth syndrome (BTHS) is an ultrarare, infantile-onset, X-linked recessive mitochondrial disorder that primarily affects males, owing to mutations in TAFAZZIN, which catalyzes the remodeling of cardiolipin, a mitochondrial phospholipid required for oxidative phosphorylation. Mitochondrial transplantation is a novel technique to treat mitochondrial dysfunction by delivering healthy mitochondria to diseased cells or tissues. Here we explored the possibility of using stem-cell-derived cardiomyocytes as a source of mitochondrial transplantation to treat BTHS. We established induced pluripotent stem (iPS) cells from healthy individuals and from patients with BTHS and differentiated them into cardiomyocytes. The iPS-cell-differentiated cardiomyocytes (CMs) derived from patients with BTHS exhibited less expression of cardiomyocytes markers, such as α-SA, cTnT and cTnI, and smaller cell size than normal iPS-cell-derived CMs. Multielectrode array analysis revealed that BTHS CMs exhibited shorter beat period and longer field potential duration than normal CMs. In addition, mitochondrial morphology and function were impaired and mitophagy was decreased in BTHS CMs compared with normal CMs. Transplantation of mitochondria isolated from normal CMs induced mitophagy in BTHS CMs, mitigated mitochondrial dysfunction and promoted mitochondrial biogenesis. Furthermore, mitochondrial transplantation stimulated cardiac maturation and alleviated cardiac arrhythmia of BTHS CMs. These results suggest that normal CMs are useful for allogeneic transplantation in the treatment of mitochondrial diseases, including BTHS.
    DOI:  https://doi.org/10.1038/s12276-025-01472-7
  7. Clin Transl Med. 2025 Jun;15(6): e70385
       BACKGROUND: OXA1L is crucial for mitochondrial protein insertion and assembly into the inner mitochondrial membrane, and its variants have been recently linked to mitochondrial encephalopathy. However, the definitive pathogenic link between OXA1L variants and mitochondrial diseases as well as the underlying pathogenesis remains elusive.
    METHODS: In this study, we identified bi-allelic variants of c.620G>T, p.(Cys207Phe) and c.1163_1164del, p.(Val388Alafs*15) in OXA1L gene in a mitochondrial myopathy patient using whole exome sequencing. To unravel the genotype-phenotype relationship and underlying pathogenic mechanism between OXA1L variants and mitochondrial diseases, patient-specific human-induced pluripotent stem cells (hiPSC) were reprogrammed and differentiated into myotubes, while OXA1L knockout human immortalised skeletal muscle cells (IHSMC) and a conditional skeletal muscle knockout mouse model was generated using clustered regularly interspaced short palindromic repeats/Cas9 genomic editing technology.
    RESULTS: Both patient-specific hiPSC differentiated myotubes and OXA1L knockout IHSMC showed combined mitochondrial respiratory chain defects and oxidative phosphorylation (OXPHOS) impairments. Notably, in OXA1L-knockout IHSMC, transfection of wild-type human OXA1L but not truncated mutant form rescued the respiratory chain defects. Moreover, skeletal muscle conditional Oxa1l knockout mice exhibited OXPHOS deficiencies and skeletal muscle morphofunctional abnormalities, recapitulating the phenotypes of mitochondrial myopathy. Further functional investigations revealed that impaired OXPHOS resulting of OXA1L deficiency led to elevated reactive oxygen species production, which possibly activated the nuclear factor kappa B signalling pathway, triggering cell apoptosis.
    CONCLUSIONS: Together, our findings reinforce the genotype-phenotype association between OXA1L variations and mitochondrial diseases and further delineate the potential molecular mechanisms of how OXA1L deficiency causes skeletal muscle deficits in mitochondrial myopathy.
    KEYPOINTS: OXA1L gene bi-allelic variants cause mitochondrial myopathy. OXA1L deficiency results in combined mitochondrial respiratory chain defects and OXPHOS impairments. OXA1L deficiency leads to elevated ROS production, which may activate the NF-κB signalling pathway, disturbing myogenic gene expression and triggering cell apoptosis.
    Keywords:  NF‐κB signalling pathway; OXA1L; mitochondrial myopathy; oxidative phosphorylation; reactive oxygen species
    DOI:  https://doi.org/10.1002/ctm2.70385
  8. Biochimie. 2025 Jun 25. pii: S0300-9084(25)00128-2. [Epub ahead of print]
      Mitochondria contain their own circular genome (mtDNA), which encodes essential components of the oxidative phosphorylation (OXPHOS) system. Mitochondrial DNA transcription is a unique and relatively simple process, requiring a specialized transcription machinery that consists of a RNA polymerase (POLRMT), two transcription factors (TFAM and TFB2M), and an elongation factor (TEFM). During transcription, a non-canonical initiating nucleotide (NCIN) can be incorporated as the first nucleotide, serving as a 5' cap. Mitochondrial transcription produces large polycistronic transcripts, which are subsequently processed by ribonucleases to generate individual messenger RNAs (mt-mRNAs), ribosomal RNAs (mt-rRNAs), and transfer RNAs (mt-tRNAs). This review will specifically focus on the maturation and regulation of mt-mRNAs. Following their release from the primary transcript, mt-mRNAs undergo various post-transcriptional modifications, including methylation, pseudouridylation, and polyadenylation. These modifications play a crucial role in determining mt-mRNAs fate by influencing their stability, translation efficiency, and overall mitochondrial function. Additionally, the spatial organization of these processes within mitochondrial RNA granules (MRGs) suggests a compartmentalized system for mitochondrial gene regulation, ensuring precise coordination between transcription, processing, and translation. A deeper understanding of these post-transcriptional modifications provides valuable insights into mitochondrial gene expression and its broader impact on cellular metabolism.
    Keywords:  LRPPRC/SLIRP; Mitochondrial mRNA; RNA degradation; mitochondrial RNA granule; post-transcriptional modification
    DOI:  https://doi.org/10.1016/j.biochi.2025.06.015
  9. J Biochem. 2025 Jun 20. pii: mvaf037. [Epub ahead of print]
      Mitochondria are intracellular organelles originating from intracellular symbiotic bacteria that play essential roles in life activities such as energy production, metabolism, Ca2+ storage, signal transduction, and cell death. Mitochondria also function as hubs for host defense against harmful stimuli such as infection and inflammation control. However, when cells are exposed to stress, mitochondrial homeostasis is disrupted, and mitochondrial DNA (mtDNA) can leak into the cytoplasm or extracellular space. Leaked mtDNA activates innate immune sensors, causing severe inflammation and contributing to the pathogenesis of human diseases. In this review, we summarize the mechanisms by which mtDNA leaks from the mitochondria and subsequently induces inflammation. We also review the relationship between mtDNA leakage and human diseases.
    Keywords:  human diseases; innate immune response; mitochondria quality control; mitochondrial DNA; mtDNA leakage
    DOI:  https://doi.org/10.1093/jb/mvaf037
  10. Curr Biol. 2025 Jun 23. pii: S0960-9822(25)00576-7. [Epub ahead of print]35(12): R595-R597
      von der Malsburg et al. introduce the mitochondrial contact site and cristae organizing system, a complex that localises to the inner mitochondrial membrane at crista junctions and stabilises these curved membrane domains.
    DOI:  https://doi.org/10.1016/j.cub.2025.05.001
  11. Spectrochim Acta A Mol Biomol Spectrosc. 2025 Jun 19. pii: S1386-1425(25)00886-8. [Epub ahead of print]343 126579
      Mitochondrial cristae are the site of intracellular biochemical reactions that play important roles in many life activities, and the destruction of mitochondrial cristae will lead to mitochondrial dysfunction inducing related diseases. Therefore, the visualization of mitochondria cristae is of great significance for physiopathology and medicine. Herein, we reported a group of cyanine-based fluorescent probes (Cy-OH and Cy-Cl) for tagging mitochondrial cristae under STED microscopy. Cy-OH and Cy-Cl could gather in the inner mitochondrial membrane via electrostatic interaction. And the ability of Cy-OH to target mitochondria depending on mitochondrial membrane potential (MMP), while Cy-Cl could tightly immobilize in mitochondria benefiting by the reaction between its chloromethyl group and mitochondrial proteins, regardless of MMP. Moreover, Cy-Cl reacted fully with mitochondrial proteins during the staining process and exhibited superior performance than some commercial mitochondrial trackers. Importantly, profiting from the high photostability and high brightness of the two probes, they could clearly depict the mitochondrial cristae at a concentration of 200 nM under STED microscopy. We believe that Cy-OH and Cy-Cl could serve as vigoroso tools to study mitochondrial cristae changes in various physiological and pathological processes.
    Keywords:  High photostability; MMP-independent; Mitochondrial cristae; Near-infrared; STED
    DOI:  https://doi.org/10.1016/j.saa.2025.126579
  12. PLoS Biol. 2025 Jun;23(6): e3003207
      Mutations in the mitochondrial genome can cause maternally inherited diseases, cancer, and aging-related conditions. Recent technological progress now enables the creation and correction of mutations in the mitochondrial genome, but it remains relatively unknown how patients with primary mitochondrial disease can benefit from this technology. Here, we demonstrate the potential of the double-stranded DNA deaminase toxin A-derived cytosine base editor (DdCBE) to develop disease models and therapeutic strategies for mitochondrial disease in primary human cells. Introduction of the m.15150G > A mutation in liver organoids resulted in organoid lines with varying degrees of heteroplasmy and correspondingly reduced ATP production, providing a unique model to study functional consequences of different levels of heteroplasmy of this mutation. Correction of the m.4291T > C mutation in patient-derived fibroblasts restored mitochondrial membrane potential. DdCBE generated sustainable edits with high specificity and product purity. To prepare for clinical application, we found that mRNA-mediated mitochondrial base editing resulted in increased efficiency and cellular viability compared to DNA-mediated editing. Moreover, we showed efficient delivery of the mRNA mitochondrial base editors using lipid nanoparticles, which is currently the most advanced non-viral in vivo delivery system for gene products. Our study thus demonstrates the potential of mitochondrial base editing to not only generate unique in vitro models to study these diseases, but also to functionally correct mitochondrial mutations in patient-derived cells for future therapeutic purposes.
    DOI:  https://doi.org/10.1371/journal.pbio.3003207
  13. Nat Struct Mol Biol. 2025 Jun 25.
      The failure to clear dysfunctional mitochondria, cell death and inflammation have been linked in neurodegenerative disease, but their relationship and role in these conditions is not fully understood. Loss of Vps13d prevents clearance of mitochondria, and mutations in human VPS13D have been associated with neurological movement disorders. To investigate the relationship between mitochondrial health, inflammation and neurodegeneration, we created a conditional Vps13d-knockout mouse. Loss of Vps13d in excitatory neurons resulted in behavioral changes and neurodegeneration. Vacuolar protein sorting 13D (VPS13D) deficiency also caused mitochondrial ultrastructural defects and dysfunction in neurons followed by gasdermin E processing, cyclic GMP-AMP synthase (cGAS)-stimulator of interferon response cGAMP interactor (STING) signaling, microglial activation and cell death. Gasdermin E localization with mitochondria in Vps13d-mutant neurons was required for elevated extracellular mitochondrial DNA that promoted activation of microglia. Depletion of microglia suppressed cell death and behavioral phenotypes but not mitochondrial changes in the neuron-specific Vps13d-knockout model, indicating that microglia promote cell death in this model of neurodegenerative disease.
    DOI:  https://doi.org/10.1038/s41594-025-01602-9
  14. Biomolecules. 2025 May 28. pii: 785. [Epub ahead of print]15(6):
      Frataxin is a component of the iron-sulfur (Fe-S) cluster assembly complex in mitochondria, and deficiency is associated with Friedreich ataxia (FA). The yeast homolog Yfh1 resembles and cross-complements with its human equivalent, and frataxin bypass scenarios are of particular interest because they may point to strategies for treating FA. Here, we describe frataxin/Yfh1 bypass by overexpression of Rsm22, an assembly factor for the mitochondrial ribosome. Rsm22 overexpression in Yfh1-depleted yeast cells restored critical processes in mitochondria, including Fe-S cluster assembly, lipoic acid synthesis, iron homeostasis, and heme synthesis, to a significant extent. Formation of cytoplasmic Fe-S proteins was also restored, suggesting recovery of the mitochondrial ability to generate the (Fe-S)int intermediate that is exported from mitochondria and is utilized for cytoplasmic Fe-S cluster assembly. Importantly, an essential component of the mitochondrial iron-sulfur cluster machinery, namely ferredoxin, was virtually absent in mitochondria lacking Yfh1, but it was recovered with Rsm22 overexpression. Interestingly, ferredoxin overexpression could offset some of the effects of Yfh1 depletion. Ferredoxin has recently been shown to bind to the cysteine desulfurase protein Nfs1 at the same site as Yfh1, in a conserved arginine patch on Nfs1, such that ferredoxin binding at this site may confer frataxin-bypass activity.
    Keywords:  METTL17; Rsm22; Yfh1; ferredoxin; frataxin; iron–sulfur proteins; mitochondria; yeast
    DOI:  https://doi.org/10.3390/biom15060785
  15. J Inherit Metab Dis. 2025 Jul;48(4): e70053
    University of Washington Center for Rare Disease Research
      Mitochondriopathies are a diverse group of disorders caused by disruption of typical mitochondrial function. Heterogenous in nature, many of these disorders arise due to variants in genes encoding key mitochondrial proteins involved in transcription and translation of mitochondrial machinery. One such gene, VARS2, encodes a mitochondrial aminoacyl-tRNA synthetase that catalyzes the attachment of valine to its cognate tRNA molecule. Bi-allelic variants in VARS2 have been linked to several forms of mitochondrial encephalopathies or cardiomyoencephalopathies. While associated clinical phenotypes vary, they can include developmental delays, axial hypotonia, limb spasticity, and epilepsy. Here, we describe three additional clinical cases of VARS2-related mitochondriopathy with sequencing-confirmed variants in VARS2 that illustrate the phenotypic variability of this disorder. These include three novel variants: Lys225Glu, Cys281Tyr, and Leu732Cysfs*29. We further assess the pathogenicity and severity of the effects of the variants underlying these cases in a Xenopus model of disease through assaying both cardiac function and brain size. In addition, we use this model of VARS2 loss of function to assess the therapeutic potential of previously proposed amino acid supplementation. Through this approach, we demonstrate that the beneficial effects of valine supplementation in VARS2 mitochondriopathy may be dependent on residual enzyme activity.
    Keywords:   Xenopus tropicalis ; VARS2 ; aminoacyltransferase; mitochondria; valine
    DOI:  https://doi.org/10.1002/jimd.70053
  16. bioRxiv. 2025 May 03. pii: 2025.04.29.650984. [Epub ahead of print]
      Purifying selection that limits the transmission of harmful mitochondrial DNA (mtDNA) mutations has been observed in both human and animal models. Yet the precise mechanism underlying this process remains undefined. Here, we present a highly specific and efficient in situ imaging method capable of visualizing mtDNA variants that differ by only a few nucleotides at single-molecule resolution in Drosophila ovaries. Using this method, we revealed that selection primarily occurs within a narrow developmental window during germline cysts differentiation. At this stage, the proportion of the deleterious mtDNA variant decreases without a reduction in its absolute copy number. Instead, the healthier mtDNA variant replicates more frequently, thereby outcompeting the co-existing deleterious variant. These findings provide direct evidence that mtDNA selection is driven by replication competition rather than active elimination processes, shedding light on a fundamental yet previously unresolved mechanism governing mitochondrial genome transmission.
    DOI:  https://doi.org/10.1101/2025.04.29.650984
  17. Eur J Hum Genet. 2025 Jun 27.
      Primary Coenzyme Q10 (CoQ10) deficiencies are a group of clinically heterogenous mitochondrial disorders that result from defects in CoQ10 biosynthesis. Their diagnosis is complicated by the absence of pathognomonic signs and poor genotype-phenotype correlations. Pathogenic variants in the COQ9 gene are a rare cause of CoQ10 deficiency: few cases have been reported, and the clinical presentation was described as a very severe multisystemic disorder with neonatal onset, ultimately leading to premature death. Through exome sequencing, we identified a novel homozygous splicing variant c.73 G > A in the COQ9 gene (NG_027696.1, NM_020312.4) in two adult siblings who presented with pure spastic paraplegia with onset in childhood. mRNA analysis from different tissues of one of the siblings revealed that this variant alters COQ9 splicing, resulting in undetectable levels of COQ9 and COQ7 proteins and reduced concentrations of CoQ10 in muscle and fibroblasts. Additionally, the accumulation of 6-demethoxycoenzyme Q10, the substrate of COQ7, was observed in both plasma and fibroblasts. Furthermore, fibroblast proliferation rate was reduced when enhancing the mitochondrial metabolism by replacing glucose with galactose in the culture medium, and was rescued by the addition of exogenous CoQ10, suggesting a therapeutic avenue for these patients. Altogether, we report here the first example of hereditary spastic paraplegia related to a mutation of the COQ9 gene that expands the spectrum of clinical manifestations and opens new therapeutic opportunities.
    DOI:  https://doi.org/10.1038/s41431-025-01895-w
  18. Int J Mol Sci. 2025 Jun 07. pii: 5481. [Epub ahead of print]26(12):
      Recent advances in flow cytometry facilitate the detection of subcellular components, such as organelles and vesicles. Fluorescence-activated mitochondria sorting (FAMS) is a flow cytometry-based technique that allows for quantitative analysis and sorting of mitochondria as individual organelles from various tissues and in vitro cell culture. This manuscript details three novel applications of this technique to study mitochondrial function on an organelle-specific level, which is not possible with other approaches. Specifically, we detail the further development and versatility of this nanoscaled flow cytometry approach, including assays to quantitatively assess mitochondrial subpopulations, mitochondrial protein translocation, and both free-floating and EV-encapsulated secreted mitochondria. We demonstrate a multi-parameter quantitative assay for the analysis of mitochondrial autophagy using antibodies targeting the proteins PINK1 and Parkin corresponding to ΔΨM and further show how these can be assessed for mtDNA content on a single organelle level. Further, we establish parameters for the size and surface marker-based analysis of EVs, many of which contain identifiable and respiring mitochondria, as well as free-floating respiratory-competent mitochondria. These results display the versatility of nanoscaled flow cytometry in terms of both sample input and target organelle and provide an important methodological means for the quantitative assessment of mitochondrial features.
    Keywords:  extracellular vesicle sorting; flow cytometry; fluorescence-activated mitochondria sorting; mitochondria; organelle sorting
    DOI:  https://doi.org/10.3390/ijms26125481
  19. Protein Sci. 2025 Jul;34(7): e70207
      In eukaryotes, cellular respiration takes place in the cristae of mitochondria. The mitochondrial inner membrane protein Mic60, a core component of the mitochondrial contact site and cristae organizing system, is crucial for the organization and stabilization of crista junctions and its associated functions. While the C-terminal Mitofilin domain of Mic60 is necessary for cellular respiration, the sequence determinants for this function have remained unclear. Here, we used ancestral sequence reconstruction to generate Mitofilin ancestors up to and including the last opisthokont common ancestor (LOCA). We found that yeast-lineage derived Mitofilin ancestors as far back as the LOCA rescue respiration. By comparing Mitofilin ancestors, we identified four residues sufficient to explain the respiratory difference between yeast- and animal-derived Mitofilin ancestors. Our results provide a foundation for investigating the conservation of Mic60-mediated cristae junction interactions.
    Keywords:  LOCA; MICOS; Mic60; ancestral sequence reconstruction; cellular respiration; cristae; mitochondria
    DOI:  https://doi.org/10.1002/pro.70207
  20. Mitochondrion. 2025 Jun 19. pii: S1567-7249(25)00056-X. [Epub ahead of print] 102059
      MTERF3, a negative regulator of mtDNA transcription, was first identified in 2007.Recent studies have revealed the pivotal role of MTERF3 throughout the entire lifecycle of mtDNA. However, no disease phenotypes have been linked to this gene till now. Genetic testing was performed on two unrelated families. Mitochondrial respiration and OXPHOS complex activity were assessed in patient-derived fibroblasts. An MTERF3 knockdown HEK293 cell line was generated, followed by rescue experiments with wild-type and mutant MTERF3. Two patients mainly presented with developmental delay. Genetic testing identified compound heterozygous variants c.635dup p.(Asn212Lysfs*7) and c.1055C > T p.(Pro352Leu) in Patient 1, and a homozygous variant c.943A > Gp.(Met315Val) in Patient 2. Patient's fibroblasts and MTERF3 knockdown cells showed impaired mitochondrial respiration and reduced levels of OXPHOS complexes I, III, and IV. Transcription of MT-ND5, ND6, COII, and COIII was reduced, while other mitochondrial genes were upregulated. Wild-type MTERF3 expression restored these defects, but the variant Pro352Leu from patient failed to rescue mitochondrial dysfunction. This study identifies a novel mitochondrial disease phenotype and establishes the first association with MTERF3, expanding the mitochondrial disease spectrum and offering insights into the clinical relevance of the MTERF family.
    Keywords:  MTERF3; Mitochondrial disease; Novel mutation; OXPHOS complex; mtDNA transcription
    DOI:  https://doi.org/10.1016/j.mito.2025.102059
  21. Nitric Oxide. 2025 Jun 20. pii: S1089-8603(25)00059-X. [Epub ahead of print]158 67-75
      3-mercaptopyruvate sulfurtransferase (MPST) is an enzyme implicated in the generation of the gasotransmitter hydrogen sulfide (H2S). Unlike, the other two H2S-synthesizing enzymes cystathionine gamma lyase (CSE) and cystathionine beta synthase (CBS), MPST is found in the mitochondria. However, the mechanisms through which MPST gains access to the mitochondria and its exact localization within this organelle remain unclear. Using immunogold electron microscopy staining, we localized MPST on the inner mitochondrial membrane. To study the pathway of mitochondrial entry for MPST, pharmacological inhibitors of different components of the translocase of outer/inner membrane were used. In line with the observation that ΜPST is found on the inner mitochondrial membrane, inhibition of TIM23 blocked MPST mitochondrial entry. Generation of N-terminally truncated forms of ΜPST did not interfere with the ability of the enzyme to gain access into the mitochondria, suggesting that an N-terminal pre-sequence does not mediate MPST mitochondrial entry. In agreement to this finding, cytosolic and mitochondrial MPST had a similar molecular weight. Interestingly, N-terminally deleted MPST exhibited reduced expression levels, indicating that this part of the enzyme is required for protein stability. Molecular dynamics simulations confirmed that deletion of the first 39 amino acids of the enzyme destabilizes the protein. Our findings reveal that MPST is present on the inner mitochondrial membrane and that its entry into mitochondria does not involve the N-terminus of the protein.
    Keywords:  3-mercaptopyruvate sulfurtransferase; Cristae; Mitochondria; TIM/TOM; TIM23
    DOI:  https://doi.org/10.1016/j.niox.2025.06.004
  22. Trends Biochem Sci. 2025 Jun 20. pii: S0968-0004(25)00130-6. [Epub ahead of print]
      
    DOI:  https://doi.org/10.1016/j.tibs.2025.05.010
  23. Mol Cell. 2025 Jun 24. pii: S1097-2765(25)00500-3. [Epub ahead of print]
      Apoptosis-inducing factor 1 (AIFM1) is a flavoprotein essential for mitochondrial function and biogenesis. Its interaction with MIA40/CHCHD4, the central component of the mitochondrial disulfide relay, accounts for some, but not all, aspects of AIFM1 function. We provide a high-confidence AIFM1 interactome that elucidates functional partners within the mitochondrial intermembrane space. We found that AIFM1 binding to adenylate kinase 2 (AK2), an essential enzyme that maintains cellular adenine nucleotide pools, depends on the AK2 C-terminal domain. High-resolution cryoelectron microscopy (cryo-EM) and biochemical analyses showed that both MIA40 and AK2A bind the AIFM1 C-terminal β-sheet domain. Their binding enhances NADH oxidoreductase activity by locking an active dimer conformation and, in the case of MIA40, affecting the cofactor-binding site. The AIFM1-AK2A interaction is important during mitochondrial respiration because AIFM1 serves as a recruiting hub within the IMS, regulating mitochondrial bioenergetic output by creating hotspots of metabolic enzymes.
    Keywords:  AIFM1; AK2; ATP; ATP transport; MIA40/CHCHD4; MICOS; metabolism; mitochondria
    DOI:  https://doi.org/10.1016/j.molcel.2025.05.036
  24. Autophagy. 2025 Jun 26.
      Lipophagy engulfs lipid droplets and delivers them to lysosomes for degradation. We found that lipophagy levels were low in most fly tissues, except for the prothoracic gland (PG) during larval development. Therefore, we performed a small-scale screening in the PG to identify regulators of lipophagy. We discovered that the loss of nmd, a gene encoding a mitochondrial AAA-ATPase, led to developmental failure and reduced lipophagy in the PG. Further studies indicated that nmd was not only required for lipophagy but also essential for general macroautophagy/autophagy in both PG and fat body tissues. Autophagy was induced but blocked at the autophagosome-lysosome fusion stage upon nmd reduction. Additionally, nmd interacted with mitochondrial protein import machinery, such as Tom20, Tom40, and the import cargo, such as Idh. Loss of nmd decreased protein import into mitochondria. Similar to the loss of nmd, reduction of Tom20 or Tom40 also resulted in reduced lipophagy in the PG. In adult flies, reducing nmd expression in the eyes caused lipid droplet accumulation and severe degeneration during aging. Overexpression of bmm, a triglyceride lipase, reduced lipid droplets in the eye but did not rescue the eye degeneration caused by the reduction of nmd.
    Keywords:  Drosophila; lipophagy; mitochondrial protein import; neuronal homeostasis; nmd; prothoracic gland
    DOI:  https://doi.org/10.1080/15548627.2025.2522124
  25. Antioxidants (Basel). 2025 Jun 16. pii: 741. [Epub ahead of print]14(6):
      Ethylmalonic encephalopathy (EE) is a serious metabolic disorder that usually appears in early childhood development and the effects are seen primarily in the brain, gastrointestinal tract, and peripheral vessels. EE is caused by pathogenic variants in the gene that encodes the ETHE1 protein, and its main features are high levels of acidic compounds in body fluids and decreased activity of the mitochondrial complex IV, which limits energy production in tissues that require a large supply of energy. ETHE1 is a mitochondrial sulfur dioxygenase that plays the role of hydrogen sulfide (H2S) detoxification, and, when altered, it leads to the accumulation of this gaseous molecule due to its deficient elimination. In this article, we characterised the pathophysiology of ETHE1 deficiency in cellular models, fibroblasts, and induced neurons, derived from a patient with a homozygous pathogenic variant in ETHE1. Furthermore, we evaluated the effect of the activation of the mitochondrial unfolded protein response (mtUPR) on the mutant phenotype. Our results suggest that mutant fibroblasts have alterations in ETHE1 protein expression levels, associated with elevated levels of H2S and protein persulfidation, mitochondrial dysfunction, iron/lipofuscin accumulation, and oxidative stress. We also identified a cocktail of compounds consisting of pterostilbene, nicotinamide, riboflavin, thiamine, biotin, lipoic acid, and L-carnitine that improved the cellular and metabolic alterations. The positive effect of the cocktail was dependent on sirtuin 3 activation (SIRT3) and was also confirmed in induced neurons obtained by direct reprogramming. In conclusion, personalised precision medicine in EE using patient-derived cellular models can be an interesting approach for the screening and evaluation of potential therapies. In addition, the activation of the SIRT3 axe of mtUPR is a promising therapeutic strategy for rescuing ETHE1 pathogenic variants.
    Keywords:  ETHE1; H2S; SIRT3; bioenergetics; ethylmalonic encephalopathy; mitochondrial diseases; mtUPR; protein persulfidation
    DOI:  https://doi.org/10.3390/antiox14060741
  26. Nat Aging. 2025 Jun 27.
      Heteroplasmic pathogenic mitochondrial DNA (mtDNA) mutations are key drivers of mitochondrial diseases, yet their tissue-specific and cell-specific accumulation patterns during aging and the mechanistic links to pathology remain poorly understood. In this study, we employed DddA-derived cytosine base editor technology to generate three mouse models harboring distinct pathogenic mitochondrial tRNA mutations. These mutations exhibited age-dependent accumulation in the kidneys, leading to severe kidney defects that well recapitulate human mitochondrial kidney disease. Mitochondrial single-cell assay for transposase-accessible chromatin with sequencing (mtscATAC-seq) revealed unique heteroplasmy dynamics across different kidney cell types: podocytes exhibited a positive selection for mutant mtDNA, whereas tubular epithelial cells displayed neutral drift of mutations during aging. Integrative analyses combining mtscATAC-seq, single-cell RNA sequencing and spatially enhanced resolution omics sequencing further identified molecular changes in high-mutant defective cells, including increased AP-1 family transcription factor activity, tubular epithelial cell proliferation and immune activation, which contribute to disease progression. Our study underscores the importance of kidney function monitoring in patients with mitochondrial disease, particularly in older adults, and establishes robust preclinical models to facilitate the development of therapeutic strategies.
    DOI:  https://doi.org/10.1038/s43587-025-00909-y
  27. Exp Mol Med. 2025 Jun 26.
      Somatic cell reprogramming into human induced pluripotent stem cells entails significant intracellular changes, including modifications in mitochondrial metabolism and a decrease in mitochondrial DNA copy number. However, the mechanisms underlying this decrease in mitochondrial DNA copy number during reprogramming remain unclear. Here we aimed to elucidate these underlying mechanisms. Through a meta-analysis of several RNA sequencing datasets, we identified genes responsible for the decrease in mitochondrial DNA. We investigated the functions of these identified genes and assessed their regulatory mechanisms. In particular, the expression of the thymidine kinase 2 gene (TK2), located in the mitochondria and required for mitochondrial DNA synthesis, is decreased in human pluripotent stem cells as compared with its expression in somatic cells. TK2 was significantly downregulated during reprogramming and markedly upregulated during differentiation. Collectively, this decrease in TK2 levels induces a decrease in mitochondrial DNA copy number and contributes to shaping the metabolic characteristics of human pluripotent stem cells. However, contrary to our expectations, treatment with a TK2 inhibitor impaired somatic cell reprogramming. These results suggest that decreased TK2 expression may result from metabolic conversion during somatic cell reprogramming.
    DOI:  https://doi.org/10.1038/s12276-025-01476-3
  28. Mol Cell. 2025 Jun 24. pii: S1097-2765(25)00507-6. [Epub ahead of print]
      ATP fuels crucial cellular processes and is obtained mostly by oxidative phosphorylation (OXPHOS) at the inner mitochondrial membrane. While significant progress has been made in mechanistic understanding of ATP production, critical aspects surrounding its substrate supply logistics are poorly understood. We identify an interaction between mitochondrial apoptosis-inducing factor 1 (AIFM1) and adenylate kinase 2 (AK2) as gatekeeper of ATP synthase. This interaction is NADH dependent and influenced by glycolysis, linking it to the cell's metabolic state. Genetic interference with AIFM1/AK2 association impedes the ability of Caenorhabditis elegans animals to handle altered metabolic rates and nutrient availability. Together, the results imply AIFM1 as a cellular NADH sensor, placing AK2 next to the OXPHOS complexes for local ADP regeneration as the substrate for ATP synthesis. This metabolic signal relay balances ATP synthase substrate supply against ATP conservation, enabling cells to adapt to fluctuating energy availability, with possible implications for AIFM1-related mitochondrial diseases.
    Keywords:  AIFM1; AK2; ATP synthesis; OXPHOS; adenylate kinase 2; apoptosis-inducing factor 1; cell signaling; crosslinking mass spectrometry; energy metabolism; mitochondria; mitochondrial; oxidative phosphorylation; protein structure; protein-protein interaction
    DOI:  https://doi.org/10.1016/j.molcel.2025.06.007
  29. Psychoneuroendocrinology. 2025 Jun 06. pii: S0306-4530(25)00229-X. [Epub ahead of print]179 107506
      Human blood contains cell-free mitochondrial DNA (cf-mtDNA) that dynamically increases in concentration in response to acute mental stress. Like other neuroendocrine stress markers, we previously found that cf-mtDNA is also detectable in saliva, calling for studies examining saliva cf-mtDNA reactivity to mental stress. In the present study, participants from the MiSBIE (Mitochondrial Stress, Brain Imaging, and Epigenetics) study (n = 68, 66 % women), were exposed to a brief socio-evaluative stressor, which induced a striking 280 % or 2.8-fold increase in saliva cf-mtDNA concentration within 10 min (g=0.55, p < 0.0001). In blood drawn concurrently with saliva sampling, stress increased cf-mtDNA by an average 32 % at 60 min in serum (g=0.20), but not in anticoagulated plasma where cf-mtDNA decreased by 19 % at 60 min (g=0.25). Examining the influence of mitochondrial health on cf-mtDNA reactivity in participants with rare mitochondrial diseases (MitoD), we report that a subset of MitoD participants exhibit markedly blunted saliva cf-mtDNA stress reactivity, suggesting that bioenergetic defects within mitochondria may influence the magnitude of saliva, and possibly blood cf-mtDNA responses. Our results document robust saliva cf-mtDNA stress reactivity and provide a methodology to examine the psychobiological regulation of cell-free mitochondria in future studies.
    Keywords:  Acute psychological stress; Cell-free mitochondrial DNA (cf-mtDNA); Energy; Mitochondrion; Repeated measures; Saliva
    DOI:  https://doi.org/10.1016/j.psyneuen.2025.107506
  30. Biochim Biophys Acta Bioenerg. 2025 Jun 17. pii: S0005-2728(25)00029-5. [Epub ahead of print]1866(4): 149563
      Multisubunit Mrp (multiple resistance and pH adaptation) type sodium proton antiporters are indispensable for the growth of alkali and salt tolerant bacteria and archaea. They share sequence and structural similarity with the membrane domain of respiratory complex I, a key mitochondrial enzyme. The molecular mechanism of complex I and Mrp antiporters has remained largely unknown and is the subject of intense debate. Here, by combining site-directed mutagenesis with large-scale molecular dynamics simulations, we explore the conformational dynamics of a key histidine residue in the MrpA subunit of the antiporter. We show that point mutations perturbing the conformational mobility of the histidine sidechain directly affect the transport activity of the antiporter. We identify that protonation state variations in conserved lysine residues around the histidine drive hydrogen bonding rearrangements and hydration changes coupled to sidechain and backbone conformational dynamics. Finally, we develop detailed and testable mechanistic models of proton transfer in Mrp antiporter and complex I, in which the histidine switch functions as a unique gating element.
    Keywords:  Alkali tolerant bacteria; Biological energy conversion; Histidine tautomer; Hydrogen bonding; Molecular dynamics; Molecular gating; Protonation states
    DOI:  https://doi.org/10.1016/j.bbabio.2025.149563
  31. FASEB J. 2025 Jun 30. 39(12): e70776
      Mitochondrial glucose metabolism is critical for glucose-stimulated insulin secretion and glucose homeostasis in pancreatic β cells. We previously showed that KCNH6, a voltage-dependent potassium (Kv) channel, participated regulation of insulin secretion in pancreatic β cells, however, its role in mitochondrial metabolism remains unclear. Since we recently found that KCNH6 distributed in mitochondria, in this study, we investigated the role of KCNH6 in regulating mitochondrial function in pancreatic β cells by using a β cell-specific knockout (KCNH6-βKO) mouse model. Proteomics analysis of islets indicated that multiple proteins involved in mitochondrial metabolism were dysregulated in islets of KCNH6-βKO mice. Additionally, KCNH6-deficient β cells exhibited damaged mitochondria morphology and oxidative respiration dysfunction, which manifested as decreased glucose-induced ATP production, elevated NADH/NAD+ ratio and ROS levels. Impaired mitochondrial metabolism in βKO islets were significantly alleviated after the re-expression of KCNH6. Mechanistically, a physical interaction between KCNH6 and complex I assembly subunit Ndufa13 was detected, providing direct evidence of KCNH6's ability to regulate mitochondrial function. These results suggested that KCNH6 could be a promising therapeutic target for improving energy metabolism in β cells.
    Keywords:  KCNH6; Ndufa13; glucose metabolism; mitochondrial metabolism; oxidative phosphorylation; proteomics
    DOI:  https://doi.org/10.1096/fj.202500707RR
  32. Genome Biol. 2025 Jun 25. 26(1): 179
       BACKGROUND: Experimental data from functional assays have a critical role in interpreting the impact of genetic variants. Assay data must be unambiguously mapped to a reference genome to make it accessible, but it is often reported relative to assay-specific sequences, complicating downstream use and integration of variant data across resources. To make multiplexed assays of variant effect (MAVE) data more broadly available to the research and clinical communities, the Atlas of Variant Effects Alliance mapped MAVE data from the MaveDB community database to human reference sequences, creating an extensive set of machine-readable homology mappings that are incorporated into widely used human genomics applications.
    RESULTS: Here, we map approximately 9.0 million individual protein and nucleotide variants in MaveDB to the human genome, describing the examined variants with respect to human reference sequences while preserving the data provenance of the original MAVE sequences. We then disseminate the results to major genomic resources including the Genomics 2 Proteins Portal, UCSC Genome Browser, Ensembl Variant Effect Predictor, and DECIPHER platform. Within these applications, MAVE variants can now be visualized and integrated with other relevant clinical and biological data, making additional knowledge available when performing variant interpretation and conducting other research activities.
    CONCLUSIONS: Mapping MAVE variants to human reference sequences and sharing the mapped dataset with several key human genomics applications enables a new and diverse set of applications for MAVE data. This study provides increased access to functional data that can assist in clinical variant interpretation pipelines and enable biomedical research and discovery.
    Keywords:  Deep mutational scanning; Functional assay; Genomic medicine; Genomics; Global Alliance for Genomics and Health; Massively parallel reporter assays; Multiplexed assays of variant effect; Variation representation specification
    DOI:  https://doi.org/10.1186/s13059-025-03647-x
  33. Biochem J. 2025 Jun 25. pii: BCJ20253133. [Epub ahead of print]482(13):
      Axonal transport is crucial for neuronal health and function, facilitating the delivery of newly synthesized material from the soma via anterograde transport and the removal of aged proteins and damaged organelles for degradation via retrograde transport. Emerging evidence links Parkinson's disease (PD)-causing mutations in the leucine-rich repeat kinase 2 (LRRK2) gene to dysfunctional axonal transport. Pathogenic LRRK2 mutations induce increased LRRK2 kinase activity, leading to the hyperphosphorylation of RAB proteins, which are key regulators of intracellular trafficking and transport. Here, we review the current literature on how LRRK2 affects the axonal transport of different cargoes, focusing on synaptic vesicle precursors, mitochondria, and autophagosomes. We further discuss how LRRK2 influences cytoskeletal dynamics and how it affects vesicle trafficking at the Golgi, which may indirectly contribute to its effect on axonal transport. This review summarizes our current understanding of how pathogenic LRRK2 hyperactivation disrupts axonal transport and how this may be linked to the neurodegeneration of PD.
    Keywords:  LRRK2; Parkinson’s disease; RAB GTPases; axonal transport
    DOI:  https://doi.org/10.1042/BCJ20253133
  34. PLoS Comput Biol. 2025 Jun;21(6): e1013090
      Mitochondrial (MT) mutations serve as natural genetic markers for inferring clonal relationships using single cell sequencing data. However, the fundamental challenge of MT mutation-based lineage tracing is automated identification of informative MT mutations. Here, we introduced an open-source computational algorithm called "MitoTracer", which accurately identified clonally informative MT mutations and inferred evolutionary lineage from scRNA-seq or scATAC-seq samples. We benchmarked MitoTracer using the ground-truth experimental lineage sequencing data and demonstrated its superior performance over the existing methods measured by high sensitivity and specificity. MitoTracer is compatible with multiple single cell sequencing platforms. Its application to a cancer evolution dataset revealed the genes related to primary BRAF-inhibitor resistance from scRNA-seq data of BRAF-mutated cancer cells. Overall, our work provided a valuable tool for capturing real informative MT mutations and tracing the lineages among cells.
    DOI:  https://doi.org/10.1371/journal.pcbi.1013090
  35. Cell Rep. 2025 Jun 24. pii: S2211-1247(25)00638-2. [Epub ahead of print]44(7): 115867
      Nucleocytoplasmic transport defects are observed in Alzheimer's disease (AD) and frontotemporal dementia (FTD). Here, we assess mRNA nucleocytoplasmic localization by performing transcriptome-wide profiling on nuclear and cytoplasmic fractions of human iPSC-derived cortical neurons from healthy individuals compared to those with familial AD or FTD. We find that AD- and FTD-causing mutations induce significant changes in mRNA nucleocytoplasmic distribution. We additionally observe the redistribution of mitochondria-related transcripts across AD and FTD neurons. The significantly increased mitochondrial RNA (mtRNA) in the cytosol of AD and FTD mutant neurons raised the possibility of leakage, which motivated us to investigate mtDNA leakage. We reveal abnormal cytoplasmic accumulation of mtDNA in AD and FTD cortical neurons together with evidence of mitochondrial aberrance. Importantly, mislocalisation of nucleic acids, mitochondrial dysfunction and cGAS-STING activation can be ameliorated through VCP D2 ATPase inhibition.
    Keywords:  Alzheimer's disease; CP: Molecular biology; CP: Neuroscience; VCP inhibition; cGAS-STING; frontotemporal dementia; mRNA; mitochondrial DNA; mitochondrial RNA; mitochondrial dysfunction; neurodegeneration; nucleocytoplasmic redistribution
    DOI:  https://doi.org/10.1016/j.celrep.2025.115867
  36. Nature. 2025 Jun 25.
      
    Keywords:  Cancer; Cell biology; Metabolism; Neuroscience
    DOI:  https://doi.org/10.1038/d41586-025-01941-z
  37. bioRxiv. 2025 Apr 07. pii: 2025.02.20.639106. [Epub ahead of print]
      The Voltage Dependent Anion Channel (VDAC) is the most ubiquitous protein in the mitochondrial outer membrane. This channel facilitates the flux of water-soluble metabolites and ions like calcium across the mitochondrial outer membrane. Beyond this canonical role, VDAC has been implicated, through interactions with protein partners, in several cellular processes such as apoptosis, calcium signaling, and lipid metabolism. There are three VDAC isoforms in mammalian cells, VDAC 1, 2, and 3, with varying tissue-specific expression profiles. From a biophysical standpoint, all three isoforms can conduct metabolites and ions with similar efficiency. However, isoform knockouts (KOs) in mice lead to distinct phenotypes, which may be due to differences in VDAC isoform interactions with partner proteins. To understand the functional role of each VDAC isoform within a single cell type, we created functional KOs of each isoform in HeLa cells and performed a comparative study of their metabolic activity and proteomics. We found that each isoform KO alters the proteome differently, with VDAC3 KO dramatically downregulating key members of the electron transport chain (ETC) while shifting the mitochondria into a glutamine-dependent state. Importantly, this unexpected dependence of mitochondrial function on the VDAC3 isoform is not compensated by the more ubiquitously expressed VDAC1 and VDAC2 isoforms. In contrast, VDAC2 KO did not affect respiration but upregulated ETC components and decreased key enzymes in the glutamine metabolic pathway. VDAC1 KO specifically reduced glycolytic activity linked to decreased hexokinase localization to mitochondria. These results reveal non-redundant roles of VDAC isoforms in cancer cell metabolic adaptability.
    DOI:  https://doi.org/10.1101/2025.02.20.639106
  38. Redox Biol. 2025 Jun 14. pii: S2213-2317(25)00245-9. [Epub ahead of print]85 103732
      Sarcopenia is the age-related degeneration of skeletal muscle, resulting in loss of skeletal muscle tone, mass, and quality. Skeletal muscle is a source of systemic metabolites and macromolecules important for neuronal health, function, and healthy neuronal aging. Age-related loss of skeletal muscle might result in decreased metabolite and macromolecule availability, resulting in reduced neuronal function or increased susceptibility to unhealthy aging and neurodegenerative diseases. We aimed to identify muscle metabolite candidates that regulate healthy aging. C57BL/6J mice were aged to young adult (4 months) and old age (25 months) and skeletal muscle was collected. Age-related muscle loss was confirmed by reduced muscle mass, muscle fiber degeneration, reduced myosin intensity, in addition to a metabolic shift and increased DNA damage in skeletal muscle. Using a low molecular weight enriched metabolomics protocol, we assessed the metabolic profile of skeletal muscle from young adult and old age mice and identified 20 metabolites that were significantly changed in aged muscle. These metabolite candidates were tested in C. elegans assays of lifespan, healthspan, muscle, and mitochondrial morphology under normal and stressed conditions. We identified four metabolite candidates (beta-alanine, 4-guanidinobutanoic acid, 4-hydroxyproline, pantothenic acid) that, when supplemented in C. elegans provided robust gero- and mitochondrial protection. These candidates also affected life-, and health- span in C. elegans models of amyotrophic lateral sclerosis (ALS) and Duchenne muscular dystrophy (DMD). Our findings support that aging muscle can be used to identify novel metabolite modulators of lifespan and health and may show promise for future treatments of neurodegenerative and neuromuscular disorders.
    Keywords:  Aging; C. elegans; Metabolomics; Mitochondria; Sarcopenia
    DOI:  https://doi.org/10.1016/j.redox.2025.103732
  39. Trends Endocrinol Metab. 2025 Jun 24. pii: S1043-2760(25)00128-6. [Epub ahead of print]
      Heme has remarkable functions in mitochondrial energetics. Recently, Duerre et al. found that branched-chain amino acids (BCAA) are preferentially utilized for heme biosynthesis to facilitate mitochondrial thermogenesis in brown fat. Disrupting heme biosynthesis shifts the metabolic fate of BCAAs toward histone propionylation, inhibiting the transcription of thermogenic genes.
    Keywords:  branched-chain amino acids; brown adipose tissue; heme synthesis; histone propionylation; mitochondria
    DOI:  https://doi.org/10.1016/j.tem.2025.06.005
  40. Methods Mol Biol. 2025 ;2952 369-410
      The mapping of genotypes to phenotypes is a cornerstone of genetics, critical for understanding disease mechanisms and advancing precision medicine. The advent of next-generation sequencing (NGS) technologies has enabled the generation of extensive genomic datasets, yet the complexity and scale of these data demand innovative analytical approaches. Artificial intelligence (AI) has emerged as a transformative tool, integrating genotype and phenotype data, uncovering intricate patterns, and driving advancements in diagnosis, therapy, and research.AI applications in phenotype-genotype mapping span various machine learning and deep learning techniques. Supervised learning methods, such as Support Vector Machines (SVMs), Random Forests, and Gradient Boosting, predict variant pathogenicity and classify genetic risks by leveraging curated datasets. Unsupervised approaches, including k-Means clustering and hierarchical clustering, uncover hidden patterns in data, enabling the identification of disease subtypes and novel associations. Dimensionality reduction techniques like Principal Component Analysis (PCA) and t-Distributed Stochastic Neighbor Embedding (t-SNE) simplify high-dimensional genomic data for analysis and visualization. Neural networks, including Convolutional and Recurrent Neural Networks (CNNs and RNNs), excel at extracting insights from complex datasets like gene expression profiles and genomic sequences. These methodologies have found applications in rare disease diagnosis, drug discovery, and risk assessment for complex diseases. AI tools integrate genetic and phenotypic data to prioritize pathogenic variants, significantly improving diagnostic yields for unresolved cases. Multi-omic data integration, incorporating genomics, transcriptomics, and proteomics, offers a holistic perspective on genotype-phenotype relationships. In drug discovery, AI identifies therapeutic targets and predicts drug efficacy, accelerating the development of precision treatments.Despite its potential, challenges persist. Data heterogeneity, limited interpretability of AI models, privacy concerns, and insufficient datasets for rare diseases impede broader implementation. To address these issues, AI frameworks incorporate data standardization, explainability techniques like SHAP and LIME, federated learning for secure collaborative research, and data augmentation methods such as transfer learning and GANs. Future directions include the integration of multi-omic data, advanced explainable AI for clinical adoption, and the expansion of federated learning to facilitate cross-institutional collaborations. By bridging the gap between genotype and phenotype, AI-driven methodologies are transforming clinical genomics and personalized medicine. This chapter explores the methodologies, applications, challenges, and future prospects of AI in phenotype-genotype mapping, highlighting its pivotal role in advancing genetic research and improving healthcare outcomes.
    Keywords:  Artificial intelligence; Genetic disorders; Graph Neural Networks; Human Phenotype Ontology; Next-generation sequencing; Polygenic Risk Scores
    DOI:  https://doi.org/10.1007/978-1-0716-4690-8_21
  41. Mol Cells. 2025 Jun 22. pii: S1016-8478(25)00071-8. [Epub ahead of print] 100247
      Neurodegenerative diseases involve toxic protein aggregation. Recent evidence suggests that biomolecular phase separation, a process in which proteins and nucleic acids form dynamic, liquid-like condensates, plays a key role in this aggregation. Optogenetics, originally developed to control neuronal activity with light, has emerged as a powerful tool to investigate phase separation in living systems. This is achieved by fusing disease-associated proteins to light-sensitive oligomerization domains, enabling researchers to induce or reverse condensate formation with precise spatial and temporal control. This review highlights how optogenetic systems such as OptoDroplet are being used to dissect the mechanisms of neurodegenerative disease. We examine how these tools have been applied in models of neurodegenerative diseases such as amyotrophic lateral sclerosis, Alzheimer's, Parkinson's, and Huntington's disease. These studies implicate small oligomeric aggregates as key drivers of toxicity and highlight new opportunities for therapeutic screening. Finally, we discuss advances in light-controlled dissolution of condensates and future directions for applying optogenetics to combat neurodegeneration. By enabling precise, dynamic control of protein phase behavior in living systems, optogenetic approaches provide a powerful framework for elucidating disease mechanisms and informing the development of targeted therapies.
    Keywords:  Biomolecular phase separation; Neurodegenerative diseases; OptoDroplet; Optogenetics; Protein aggregation; neurodegeneration
    DOI:  https://doi.org/10.1016/j.mocell.2025.100247
  42. FASEB J. 2025 Jun 30. 39(12): e70739
      Barth syndrome (BTHS) is a rare disease caused by mutations in the tafazzin gene that affects the heart and muscles; however, to date, no clinically effective drugs are available. In BTHS, mitochondrial function is reduced owing to changes in cardiolipin metabolism. We developed mitochonic acid 5 (MA-5), a small-molecule compound that increases ATP levels, improves mitochondrial dynamics, and is effective in treating mitochondrial and muscle diseases. Therefore, this study examined the effectiveness of MA-5 in treating BTHS. The mitochondrial functions of four isolated BTHS skin fibroblasts were examined. Human BTHS induced pluripotent stem cell (iPSC) were differentiated into myoblasts and cardiolipin metabolism and mitochondrial functions were analyzed. RNA-seq was performed to clarify the metabolic changes. Using a Drosophila melanogaster model of BTHS, the effects of MA-5 on motor performance and cardiac phenotype were examined. MA-5 improved mitochondrial function and reduced cell death due to oxidative stress in skin fibroblasts of patients with BTHS. MA-5 promoted ATP production and reduced oxidative stress in human BTHS iPS cell-derived myoblasts. RNA-seq analysis revealed that MA-5 alleviated endoplasmic reticulum stress in BTHS cells. Administration of MA-5 to BTHS Drosophila improved locomotor ability and tachycardia observed in patients with BTHS. Protein interaction analyses suggested colocalization of ATPase and the MA-5-binding protein mitofilin. These data suggested that MA-5 improves BTHS dysfunction and may serve as a novel therapeutic agent for BTHS.
    Keywords:   Drosophila ; ATP; Barth syndrome; cardiolipin; iPS; mitochondria
    DOI:  https://doi.org/10.1096/fj.202401856RRR
  43. Nature. 2025 Jun 27.
      
    Keywords:  Cell biology; Machine learning; Medical research
    DOI:  https://doi.org/10.1038/d41586-025-02011-0
  44. Nat Rev Mol Cell Biol. 2025 Jun 23.
      Intracellular membrane contact sites (MCSs) between organelles have crucial roles in cellular signalling and homeostasis. These sites, which are often disrupted in pathological conditions, enable the exchange of ions, lipids and metabolites between membrane-bound compartments, helping cells adapt to varying physiological conditions. Specific tether proteins and complexes stabilize these interactions and mediate responses to different intracellular or extracellular stimuli. The study of MCSs has progressed in recent years, owing to the development of new methods such as genetically encoded reporter constructs, advanced imaging techniques, including super-resolution microscopy and electron tomography, and proteomic approaches based on mass spectrometry. These tools have enabled unprecedented visualization and quantification of organelle interactions, as well as identification of the molecular players involved. This Expert Recommendation aims to define and map the 'organelle contactome', describing key proteins involved in contact site formation and the roles of MCSs in cellular function. We also explore contact site dynamics and detail advantages and disadvantages of the methodologies for studying them. Importantly, we consolidate open questions in contact site research and discuss challenges and limitations of the current experimental approaches.
    DOI:  https://doi.org/10.1038/s41580-025-00864-x
  45. Nature. 2025 Jun;642(8069): 864-866
      
    Keywords:  Machine learning; Medical research; Policy
    DOI:  https://doi.org/10.1038/d41586-025-01946-8
  46. Nat Genet. 2025 Jun 23.
      Understanding the disease risk of genetic variants is fundamental to precision medicine. Estimates of penetrance-the probability of disease for individuals with a variant allele-rely on disease-specific cohorts, clinical testing and emerging electronic health record (EHR)-linked biobanks. These data sources, while valuable, each have limitations in quality, representativeness and analyzability. Here, we provide a historical account of the currently accepted pathogenicity classification system and data available in ClinVar, a public archive that aggregates variant interpretations but lacks detailed data for accurate penetrance assessment, highlighting its oversimplification of disease risk. We propose an integrative Bayesian framework that unifies pathogenicity and penetrance, leveraging both functional and real-world evidence to refine risk predictions. In addition, we advocate for enhancing ClinVar with the inclusion of high-priority phenotypes, age-stratified data and population-based cohorts linked to EHRs. We suggest developing a community repository of population-based penetrance estimates to support the clinical application of genetic data.
    DOI:  https://doi.org/10.1038/s41588-025-02212-3
  47. Protein Sci. 2025 Jul;34(7): e70190
      The LRRK2 gene is a key contributor to the genetic risk of Parkinson's disease, and a priority drug target for the disorder. Leucine Rich Repeat Kinase 2, the protein product of LRRK2, is a multidomain enzyme implicated in a range of cellular processes-including endolysosomal trafficking and damage response. Based on the report that truncation and structural variants resulting in loss of LRRK2 protein are observed in human populations, genomic sequence repositories were queried for coding variants affecting key catalytic residues in LRRK2-resulting in the identification of three variants (K1347E, K1347R, and T1348P) predicted to ablate the capacity of LRRK2 to bind GTP. Biochemical and cellular characterization of these variants confirmed loss of GTP binding, as well as reduced or loss of kinase activity. These data demonstrate the presence of rare coding enzymatic loss-of-function variants in humans, with implications for our understanding of LRRK2 as a driver of disease and as a drug target.
    Keywords:  GTPase; LRRK2; Parkinson's; leucine rich repeat kinase 2; lysosomes
    DOI:  https://doi.org/10.1002/pro.70190
  48. PLoS One. 2025 ;20(6): e0326249
      Low cardiorespiratory fitness (CRF) is a well-established risk factor for cardiovascular disease (CVD) and all-cause mortality. Since CRF is largely genetically determined, understanding the genetic influences on CRF might reveal the protective mechanisms of high CRF. One gene found to be associated with CRF is COX7A2L. COX7A2L is a mitochondrial supercomplex assembly factor, but its role in cellular metabolism remains a topic of discussion. We hypothesized that COX7A2L could play a role in cellular respiration in cardiomyocytes, affecting cardiac function and CRF. To determine the effect of COX7A2L on cardiomyocyte function, we overexpressed and knocked down COX7A2L in human AC16 cardiomyocytes and performed MTT assays and Seahorse XF Cell Mito Stress Tests to assess cell viability and mitochondrial function. For the mitochondrial function measurements, we stimulated the cells with isoproterenol to investigate if the effect of altering COX7A2L levels would be larger under simulated increased energy demand. Overexpression and knockdown were validated using sandwich ELISA. Our findings showed that altering COX7A2L expression in human AC16 cardiomyocytes did not significantly affect cell viability or mitochondrial function. Further research is necessary to determine whether COX7A2L influences cardiomyocyte function and CRF.
    DOI:  https://doi.org/10.1371/journal.pone.0326249
  49. Mol Cell Endocrinol. 2025 Jun 24. pii: S0303-7207(25)00157-1. [Epub ahead of print] 112606
      Targeted metabolomics and ELISAs shown that Mdivi-1 treatment increased the levels of steroid hormones (progesterone and estradiol) in the supernatants of KGN cell culture medium. The purpose of this study was to explore the mechanism of Mdivi-1 promoting steroid hormone synthesis in granulosa cells (GCs). In vitro experiments revealed that Mdivi-1 did not affect the total protein expression of Drp1 in KGN cells or human luteinized GCs but increased Drp1 Ser637 phosphorylation, reduced Drp1 Ser616 phosphorylation, inhibited Drp1 mitochondrial translocation, and upregulated mitochondrial fusion proteins, promoting mitochondrial fusion. In terms of energy production, Mdivi-1 increased the expression of mitochondrial complexes I and V and the ATP concentration in GCs, increasing the energy supply for steroidogenesis. Mdivi-1 exposure significantly increased the expression and mitochondrial localization of StAR and CYP11A1 in the steroid production pathway of GCs. Further in vivo experiments demonstrated that, compared with the controls, Mdivi-1 treatment significantly increased the levels of Drp1 Ser637, StAR and CYP11A1 in ovarian tissue and the serum levels of progesterone and estradiol. Taken together, these findings suggest that Mdivi-1 induces mitochondrial fusion by increasing Drp1 phosphorylation at Ser637 and weakening the interaction between Drp1 and mitochondria. Moreover, mitochondrial fusion increases the cellular bioenergetics and the expression of StAR and CYP11A1 as well as their mitochondrial localization, thereby enhancing the activity of steroidogenesis in GCs.
    Keywords:  Drp1 phosphorylation; Granulosa cells; Mdivi-1; Mitochondrial dynamics; Steroidogenesis
    DOI:  https://doi.org/10.1016/j.mce.2025.112606
  50. Nat Med. 2025 Jun 27.
      With aging, deviation of human blood counts from their normal range accompanies the transition from health to disease. Hematopoietic stem and progenitor cells (HSPCs) deliver life-long multi-lineage output, but their variation across healthy humans with aging, and their diagnostic utility, haven't been characterized in depth thus far. To address this, we introduced an HSPC reference model using single-cell RNA profiling of circulating CD34+ HSPCs from 148 healthy age- and sex-diverse individuals. We characterized physiological circulating HSPC composition, showed that age-related myeloid bias is predominant in older men and defined age-related transcriptional signatures in lymphoid progenitors. We further demonstrated the potential of this resource to facilitate the diagnosis of myelodysplastic syndrome (MDS) from peripheral blood without bone marrow sampling, defining classes of patients with MDS and abnormal lymphocyte, basophil or granulocyte progenitor frequencies. Our resource provides insights into HSPC reference ranges across the lifespan and has the potential to facilitate the clinical applications of single-cell genomics in hematology.
    DOI:  https://doi.org/10.1038/s41591-025-03716-5
  51. Nature. 2025 Jun 25.
      
    Keywords:  Cancer; Medical research; Neuroscience
    DOI:  https://doi.org/10.1038/d41586-025-01718-4
  52. Blood Adv. 2025 Jun 24. pii: bloodadvances.2024015009. [Epub ahead of print]
      Tracking transfused platelets is important to evaluate platelet transfusion efficiency. Traditionally, corrected count increments were used; however, quantitative polymerase chain reaction-based methods have recently been developed. As both these methods have some limitations, we developed a new method based on next-generation sequencing (NGS) of platelet mitochondrial DNA (mtDNA). We identified several single nucleotide variant markers by sequencing the entire mtDNA region of platelets, and used NGS to estimate the proportion of each platelet unit. This method was validated using mixed platelets obtained from different donors at various ratios. We confirmed the applicability of this method in patients who received platelet transfusions using pre- and post-transfusion samples from 12 patients with hematological malignancies. The method showed good linearity (r² > 0.99 in the range of mixing ratios from 1:1 to 1:50) in the platelet-mixing experiment. In addition, platelet tracking in patients who received transfusions was feasible using this method. Furthermore, it was possible to track individual platelets in patients who received a single platelet transfusion and in those who received multiple transfusions, including a patient who received five platelet transfusions. Hence, this NGS-based platelet-tracking method can be used for patients with various conditions.
    DOI:  https://doi.org/10.1182/bloodadvances.2024015009
  53. Med Sci (Basel). 2025 Jun 09. pii: 75. [Epub ahead of print]13(2):
      Background: Next-generation sequencing (NGS), particularly whole-exome sequencing (WES), has become a powerful diagnostic tool for rare genetic conditions. However, its success rate varies based on the underlying genetic etiology and the population studied. Methods: This retrospective study evaluated the diagnostic yield of NGS in a cohort of 137 pediatric patients with suspected rare genetic disorders in Bulgaria, a setting where such testing is not reimbursed and must be self-funded. The patients underwent either WES or targeted gene panel testing based on clinical presentation, family history, and genetic evaluation. Results: The overall diagnostic yield was 45.99%, with WES achieving 51.25% and targeted testing achieving 38.60%. The highest yield was observed in patients presenting with both dysmorphic features and neurodevelopmental delays (62.5%), while the lowest was observed among those with isolated neurodevelopmental issues (10%). A significant portion of the identified variants (35.9%) were novel. Eight patients were diagnosed with copy number variants (CNVs) detected only through WES. Conclusions: Our findings illustrate the value of WES as a first-line test and highlight the impact of deep phenotyping on diagnostic success. This study also emphasizes the need for a population-specific reference genome and equal access to genomic diagnostics in all European countries.
    Keywords:  diagnostic yield; genetic counseling; next-generation sequencing; rare disease; success rate
    DOI:  https://doi.org/10.3390/medsci13020075
  54. Antioxidants (Basel). 2025 May 22. pii: 618. [Epub ahead of print]14(6):
      Genetics and mitochondrial (mt) dysfunction contribute to metabolic dysfunction-associated steatotic liver disease (MASLD). Recently, we demonstrated that the co-presence of PNPLA3, TM6SF2 and MBOAT7 polymorphisms predisposes to disease progression in MASLD patients and that their deletion triggers mt maladaptation in vitro. Here, we deepened the impact of the silencing of these genes on mt dynamism and respiration by reintroducing TM6SF2 and/or MBOAT7 wild-type proteins in deleted cells through lentiviral infection. Since hepatic mt bioenergetics is impaired in MASLD, in the attempt to identify a non-invasive signature, we then compared the enzymatic mt activity of seahorses, which was assessed in liver biopsies and peripheral blood mononuclear cells (PBMCs) of biopsy-proven MASLD patients (n = 44; Discovery cohort) stratified according to the number of the three at-risk variants (3NRV). Concerning the in vitro results, the rescue of MBOAT7 and/or TM6SF2 wild-type proteins resulted in the assembly of spaghetti-shaped mitochondria with improved oxidative phosphorylation (OXPHOS) capacity. In the Discovery cohort, the hepatic bioenergetic profile fully reflected that in PBMCs and was impaired especially in 3NRV carriers. A lowered serum respiration rate was confirmed in noninvasively assessed MASLD (n = 45; Fibroscan-MASLD cohort), while it did not change in unrelated liver disease patients (n = 45). In summary, we firstly demonstrated that mt circulating respirometry reflects that in liver and is specific in defining genetic MASLD.
    Keywords:  MASLD; genetics; mitochondria; non-invasive biomarker
    DOI:  https://doi.org/10.3390/antiox14060618
  55. Nature. 2025 Jun 25.
      
    Keywords:  Genomics; Machine learning; Proteomics
    DOI:  https://doi.org/10.1038/d41586-025-01998-w
  56. Genetics. 2025 Jun 25. pii: iyaf122. [Epub ahead of print]
      Neurons maintain their morphology over prolonged periods of adult life with limited regenerative capacity. Among the various factors that shape neuronal morphology, lipids function as membrane components, signaling molecules, and regulators of synaptic plasticity. Here, we tested genes involved in phospholipid biosynthesis and identified their roles in axon regrowth and maintenance. CEPT-2 and EPT-1 are enzymes catalyzing the final steps in the de novo phospholipid synthesis (Kennedy) pathway. Loss of function mutants of cept-2 or ept-1 show reduced axon regrowth and failure to maintain axon morphology. We demonstrate that CEPT-2 is required cell-autonomously to prevent age-related axonal morphology defects. We further investigated genetic interactions of cept-2 or ept-1 with dip-2, a conserved regulator of lipid metabolism that affects axon morphology maintenance and regrowth after injury. Loss of function in dip-2 led to suppression of axon regrowth defects observed in either cept-2 or ept-2 mutants, suggesting that DIP-2 acts to counterbalance phospholipid synthesis. Our findings reveal the genetic regulation of lipid metabolism as critical for axon maintenance following injury and during aging.
    Keywords:  Kennedy pathway; aging; axon degeneration; axon regeneration; phosphatidylcholine; phosphatidylethanolamine
    DOI:  https://doi.org/10.1093/genetics/iyaf122
  57. Gen Physiol Biophys. 2025 Jul;44(4): 275-287
      The aim of our work was to study impact of tauroursodeoxycholic acid (TUDCA), 4-phenylbutyric acid (PBA) and their combination on mitochondrial functions and morphology. TUDCA, PBA and their combination have a significant impact on mitochondrial respiration. Although both TUDCA and PBA are considered to be chemical chaperones influencing endoplasmic reticulum (ER) stress, they affect mitochondrial respiration in a specific way. While TUDCA decreases ROUTINE, maximal, succinate-driven maximal, ATP-coupled and leak respirations; PBA increases spare respiratory capacity (SRC). Combination of TUDCA with PBA increases ROUTINE, maximal, succinate driven maximal and ATP-coupled respirations and SRC. TUDCA, PBA and their combination exhibits positive impact on mitochondria elongation and do not induce expression of proteins involved in mitochondrial fusion and unfolded protein response. Our results do not indicate the impact of either TUDCA or PBA on ER stress since pre-treatment of the cells with either TUDCA or PBA does not significantly affect tunicamycin-induced expression of HRD1, GRP78 and SEL1L. The impact of PBA and combination of TUDCA with PBA on mitochondrial functions might be associated with their possible neuroprotective effects. Although TUDCA exhibits positive effect on inner mitochondrial membrane, the possible neuroprotective effect of TUDCA might involve mechanism distinct from modification of mitochondrial functions.
    DOI:  https://doi.org/10.4149/gpb_2025017
  58. Ann Hum Genet. 2025 Jun 27. e12606
      We review three areas of human genetics that have been developed in the past few decades, in which statistical innovation has made a crucial contribution with recent important advances and the potential for further rapid progress. The first topic is the development of mathematical models for the genealogy underlying samples of genome-wide genetic data. Coalescent theory emerged in the 1980s, leaped ahead in the past decade and is now burgeoning into new application areas in population, evolutionary and medical genetics. The second is the development of statistical methods for genome-wide association studies which has made great strides over two decades, including exciting recent developments for association testing based on coalescent theory and improved methods for trait prediction. Finally, we review the statistical ideas that helped resolve the controversies surrounding the introduction of forensic DNA profiling in the early 1990s. Big advances in interpretation of the predominant autosomal DNA profiles have set a benchmark for other areas of forensic science, but the statistical assessment of uniparentally inherited profiles (derived from the mitochondrial DNA or the Y chromosome) remains unsatisfactory.
    Keywords:  ancestral recombination graph; coalescent models; complex traits; forensic DNA profiles; genetic epidemiology; genome‐wide association studies; heritability
    DOI:  https://doi.org/10.1111/ahg.12606
  59. Elife. 2025 Jun 23. pii: RP99986. [Epub ahead of print]13
      Hypoxia is an important physiological stress causing nerve injuries and several brain diseases. However, the mechanism of brain response to hypoxia remains unclear, thus limiting the development of interventional strategies. This study conducted combined analyses of single-nucleus transcriptome sequencing and extracellular vesicle transcriptome sequencing on hypoxic mouse brains, described cell-cell communication in the brain under hypoxia from intercellular and extracellular dimensions, confirmed that hemoglobin mRNA was transferred from non-neuronal cells to neurons, and eventually expressed. Then we further explored the role of exosomal hemoglobin transfer in vitro, using human-derived cell lines, and clarified that hypoxia promoted the transfer and expression of exosomal hemoglobin between endothelial cells and neurons. We found the vital function of exosomal hemoglobin to protect against neurological injury by maintaining mitochondrial homeostasis in neurons. In conclusion, this study identified a novel mechanism of 'mutual aid' in hypoxia responses in the brain, involving exosomal hemoglobin transfer, clarified the important role of exosomal communication in the process of brain stress response, and provided a novel interventional perspective for hypoxia-related brain diseases.
    Keywords:  brain-derived extracellular vesicles; hemoglobin; hypoxia; mitochondrial homeostasis; mouse; nerve injury; neuroscience
    DOI:  https://doi.org/10.7554/eLife.99986
  60. bioRxiv. 2025 Apr 30. pii: 2025.04.30.651504. [Epub ahead of print]
      Overexpression of RNase H1, a ribonuclease that degrades RNA:DNA hybrids and R-loops, can suppress genome instability phenotypes in a range of maladaptive conditions. This has been interpreted to suggest that genotoxic co-transcriptional R-loops arise under these conditions and are resolved by RNase H1. Here, we manipulated RNase H1 levels using conditional knockout and overexpression models in primary murine B cells and mapped the resulting genomic R-loop landscapes. Rnaseh1 deletion resulted in a dramatic loss of mitochondrial replication and compromised B cell responses, consistent with a critical mitochondrial function for RNase H1. Genome-wide R-loops were, however, not significantly affected. More surprisingly, overexpressing active nuclear RNase H1 did not lead to significant reduction of R-loop levels or change their distribution. These results were confirmed using a human cell line in which active, nuclear RNase H1 can be induced. Our findings indicate that co-transcriptional R-loops are not efficiently resolved by RNase H1 and suggest that the identity of the RNA/DNA hybrids at the root of the genome instability phenotypes suppressed by RNase H1 may need to be re-interpreted.
    DOI:  https://doi.org/10.1101/2025.04.30.651504