bims-celmim Biomed News
on Cellular and mitochondrial metabolism
Issue of 2025–11–30
25 papers selected by
Marc Segarra Mondejar, AINA
  1. Neuronal glycolytic reprogramming drives lethality via accelerated aging in a Drosophila model of tauopathy.
  2. Spatial Partitioning of Core Glycolysis Enables Tissue-Specific Metabolic Programs In Vivo.
  3. Sec14L6 is a phosphoinositide transporter that regulates phosphoinositide homeostasis and biogenesis of lipid droplets.
  4. Disruption of the PIKfyve complex unveils an adaptive mechanism to promote lysosomal repair and mitochondrial homeostasis.
  5. High-Resolution Spatiotemporal Mapping of Cerebral Metabolism During Middle-Cerebral-Artery Occlusion/Reperfusion Progression: Preliminary Insights.
  6. Neuronal compartmentalization results in "impoverished" axonal mitochondria.
  7. Imaging Cellular Metabolic Rewiring with SuMMIT-SRS.
  8. Macropinocytosis enables metabolic recycling of extracellular DNA in cancer cells.
  9. MICU2 controls mitochondrial calcium signaling and migration in neurons during development.
  10. Mitochondrial NAD+ drives liver regeneration.
  11. Isoform-specific regulation of PKM by acetylation.
  12. CYFIP1 governs the development of cortical axons by modulating calcium availability.
  13. The histone modification regulator, SIN3, plays a role in the cellular response to changes in glycolytic flux.
  14. Cellular metabolism biosensing: From extracellular microenvironment to intracellular physiology monitoring.
  15. ER exit sites mediated by the COPII adaptor sec24D selectively recruit lipid raft-preferring proteins for rapid ER export.
  16. MitoLandscape, a semi-automated pipeline for subcellular localization and quantification of mitochondria.
  17. The luminal domain region of Seipin/Fld1 is dispensable for establishing functional ER sites for lipid droplet biogenesis.
  18. All-Optical Multimodal Mapping of Single Cell-Type-Specific Metabolic Activities via REDCAT.
  19. Homeostatic regulation of intrinsic lipid curvature in eukaryotic cells.
  20. Visualizing intraorganellar ultrastructures, dynamics, and interactions with open-access background-free Lock-in-SIM.
  21. pH-dependent regulation in SLC38A9.
  22. Multi-omic analysis reveals lipid dysregulation associated with mitochondrial dysfunction in parkinson's disease brain.
  23. A surrogate barrier model for high-throughput blood-brain barrier permeability prediction: integrating LLC-PK1-MOCK/MDR1 Cells and lysosomal trapping correction.
  24. Homocysteine disrupts lysosomal function by V-ATPase inhibition.
  25. Modulation of the PGRMC1/NLRP7/HLA-C axis by autophagy is linked to both spontaneous preterm birth and gestational choriocarcinoma.
  1. bioRxiv. 2025 Oct 27. pii: 2025.10.27.684860. [Epub ahead of print]
    Neuronal glycolytic reprogramming drives lethality via accelerated aging in a Drosophila model of tauopathy.
    Richa Gupta, Hope McGinnis, Elham Rastegari, Matthew S Price, Kartik Venkatachalam.
      Neurometabolic dysfunction is a hallmark of Alzheimer's disease (AD) and tauopathies. Whether these changes drive pathology or represent compensatory, protective responses remains unresolved. Here, we demonstrate that human tau induces Warburg-like metabolism in Drosophila neurons, characterized by coordinated upregulation of glycolytic enzymes and lactate dehydrogenase that mirrors metabolic signatures in human AD. Despite intact mitochondrial oxidative phosphorylation, tau -expressing fly neurons preferentially utilize glycolysis for ATP production and operate with diminished metabolic reserve. Crucially, this metabolic reprogramming drives rather than protects against pathology as genetic suppression of glycolysis or lactate dehydrogenase completely rescued tau-induced lethality. Further, Gompertz mortality analysis revealed that hyperactive glycolysis in tau neurons drives premature lethality by accelerating biological aging rate without affecting baseline mortality. Collectively, these findings establish aberrant neuronal glycolysis as a cause rather than a consequence of tau pathology, and demonstrate that sustained glycolytic metabolism in mature neurons exacts a specific cost in the form of accelerated aging.
    DOI:  https://doi.org/10.1101/2025.10.27.684860
  2. bioRxiv. 2025 Nov 10. pii: 2025.11.09.687253. [Epub ahead of print]
    Spatial Partitioning of Core Glycolysis Enables Tissue-Specific Metabolic Programs In Vivo.
    Ian J Gonzalez, Aaron D Wolfe, Benjamin Clark, Michael Hanna, Quishi Sun, Snusha Ravikumar, Anastasia Tsives, Sarah E Emerson, Stephan Siebel, Richard Kibbey, Daniel Colón-Ramos.
      Tissues exhibit metabolic heterogeneity that tailors metabolism to their physiological demands. How the conserved pathways of metabolism achieve metabolic heterogeneity is not well understood, particularly in vivo. We established a system in Caenorhabditis elegans to investigate tissue-specific requirements for glucose 6-phosphate isomerase (GPI-1), a conserved glycolytic enzyme that also regulates the pentose phosphate pathway (PPP). Using CRISPR-Cas9 genome editing, we found that gpi-1 knockout animals display germline defects consistent with impaired PPP, and somatic defects consistent with impaired glycolysis. We discovered that two GPI-1 isoforms are differentially expressed and localized: GPI-1A is expressed in most tissues, where it displays cytosolic localization, whereas GPI-1B is primarily expressed in the germline, where it localizes to subcellular foci near the endoplasmic reticulum. GPI-1B expression alone is sufficient to maintain wild type levels of reproductive fitness, but insufficient to reconstitute wild-type glycolytic dynamics. Our findings uncover isoform-specific, spatially-compartmentalized functions of GPI-1 that underpin tissue-specific anabolic and catabolic metabolism in vivo , underscoring roles for subcellular localization in achieving tissue-specific metabolic flux.
    DOI:  https://doi.org/10.1101/2025.11.09.687253
  3. Nat Commun. 2025 Nov 26. 16(1): 10518
    Sec14L6 is a phosphoinositide transporter that regulates phosphoinositide homeostasis and biogenesis of lipid droplets.
    Tiantian Zhou, Juan Xiong, Xuewen Hu, Yuanjiao Du, Anbing Shi, Weiping Chang, Jichao Qin, Lin Deng, Wei-Ke Ji.
      Lipid droplets (LDs) are evolutionarily conserved organelles essential for cellular metabolism. They form and grow at the endoplasmic reticulum (ER), requiring lipid transfer between these compartments, yet the underlying molecular mechanisms remain elusive. We identify Sec14L6, a unique Sec14 family member, as a lipid transporter regulating phosphoinositide (PIP) homeostasis and LD biogenesis, promoting adipogenic differentiation of mesenchymal stem cells. Sec14L6 directly binds the LD biogenesis factor ACSL3, which facilitates the association of Sec14L6 with LD surface. Furthermore, the ER membrane protein PGRMC1 recruits Sec14L6 to the ER. Targeted lipidomics revealed profound PIP dysregulation in Sec14L6-KO cells: LDs accumulated phosphoinositide-4-phosphate (PI4P) and PI(4,5)P₂, while these PIPs were reduced within the ER. In vitro assays demonstrated that Sec14L6 transports PI4P and PI(4,5)P₂. Sec14L6 knockout significantly impaired LD formation; this defect was rescued by wild-type Sec14L6, but not by lipid-transfer-deficient mutants. Our study reveals an essential role for Sec14L6 in PIP homeostasis and promotes LD biogenesis through lipid transfer between the ER and LDs.
    DOI:  https://doi.org/10.1038/s41467-025-65540-2
  4. Nat Commun. 2025 Nov 28. 16(1): 10761
    Disruption of the PIKfyve complex unveils an adaptive mechanism to promote lysosomal repair and mitochondrial homeostasis.
    Candice Kutchukian, Maria Casas, Rose E Dixon, Eamonn J Dickson.
      Lysosomes are essential organelles that regulate cellular homeostasis through complex membrane interactions. Phosphoinositide lipids play critical roles in orchestrating these functions by recruiting specific proteins to organelle membranes. The PIKfyve/Fig4/Vac14 complex regulates PI(3,5)P₂ metabolism, and intriguingly, while loss-of-function mutations cause neurodegeneration, acute PIKfyve inhibition shows therapeutic potential in neurodegenerative disorders. We demonstrate that PIKfyve/Fig4/Vac14 dysfunction triggers a compensatory response where reduced mTORC1 activity leads to ULK1-dependent trafficking of ATG9A and PI4KIIα from the TGN to lysosomes. This increases lysosomal PI(4)P, facilitating cholesterol and phosphatidylserine transport at ER-lysosome contacts to promote membrane repair. Concurrently, elevated lysosomal PI(4)P recruits ORP1L to ER-lysosome-mitochondria three-way contacts, enabling PI(4)P transfer to mitochondria that drives ULK1-dependent fragmentation and increased respiration. These findings reveal a role for PIKfyve/Fig4/Vac14 in coordinating lysosomal repair and mitochondrial homeostasis, offering insights into cellular stress responses.
    DOI:  https://doi.org/10.1038/s41467-025-65798-6
  5. Biomolecules. 2025 Nov 06. pii: 1558. [Epub ahead of print]15(11):
    High-Resolution Spatiotemporal Mapping of Cerebral Metabolism During Middle-Cerebral-Artery Occlusion/Reperfusion Progression: Preliminary Insights.
    Zhongcheng Yuan, Minhao Xu, Mingze Lu, Guancheng Wang, Jingyuan Ma, Sitong Ding, Haoan Wu, Yu Zhang, Ming Ma.
      Ischemia-reperfusion is a rapidly evolving cascade that involves a variety of metabolic shifts whose precise timing and sequential order are still poorly understood. Clarifying these dynamics is critical for understanding the core injury trajectory of stroke and for refining time-delimited therapeutic interventions. More broadly, continuous in situ monitoring of the middle-cerebral-artery occlusion process at the system level has not yet been achieved. Here, we report the first single-subject high-resolution spatiotemporal resolution metabolic maps of the ultra-early phase of ischemic stroke in a rodent model. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) imaging mapped a metabolic abnormality area in the ischemic hemisphere that propagates from the striatum to the cortex. Microdialysis probes were then stereotaxically implanted within this metabolic abnormality area, capturing 10,429 metabolites that resolved into 16 temporally distinct trajectories aligned with probe insertion, ischemic injury, and reperfusion injury. Analysis of specific metabolic pathways mainly revealed that the delayed clearance of metabolic waste (urea and tryptamine) during early reperfusion, the transient attenuation of the citrate-to-oxaloacetate buffering gradient within the TCA cycle, and the accumulation of extracellular branched-chain amino acids all play crucial roles in shaping the injury trajectory. Simultaneously, the depletion of cellular repair mechanisms (pyrimidine synthesis) in the early phase of reperfusion also warrants our attention. These findings provide novel insights into the molecular basis and mechanisms of ischemia-reperfusion and offer a comprehensive resource for further investigation.
    Keywords:  MALDI-MS imaging; TCA cycle; branched-chain amino acids; ischemia–reperfusion injury; microdialysis; spatiotemporal metabolomics
    DOI:  https://doi.org/10.3390/biom15111558
  6. bioRxiv. 2025 Oct 29. pii: 2025.10.27.684882. [Epub ahead of print]
    Neuronal compartmentalization results in "impoverished" axonal mitochondria.
    Adrian Marti Pastor, Liam S C Lewis, Stephan Müller, Pedro Guedes-Dias, Lorena Olifiers, Polina Pelkonen, Almudena Del Rio Martin, Caroline Fecher, Nikolai P Skiba, Ying Hao, Antoneta Gavoci, Laura Trovo, Mehmet Akif Aktas, Hariharan M Mahadevan, Angelika B Harbauer, Howard M Bomze, Vadim Y Arshavsky, Monika S Brill, Romain Cartoni, Stefan F Lichtenthaler, Sidney M Gospe, Thomas Misgeld.
      Mitochondria differ depending on their location within a neuron. Morphological heterogeneity between somatic, dendritic, and axonal mitochondria is well established. Emerging evidence suggests that further specialization is needed to meet the unique demands of different neuronal compartments. However, the molecular and functional diversity of mitochondria within a neuron remains poorly understood. Here, we utilized proteomics in MitoTag mice to profile somatodendritic and axonal mitochondria across four distinct neuron types, thereby generating a compendium of intracellular mitochondrial diversity. Combining proteomics, functional, and immunofluorescence analyses, we demonstrated that axonal mitochondria are not defined by the presence of unique proteins, but rather by the selective loss or preservation of specific pathways compared to their somatodendritic counterparts. This results in "impoverished" axonal mitochondria, which are characterized by diminished mtDNA expression and impaired oxidative phosphorylation yet retain other pathways, such as fatty acid metabolism. Bioinformatic analyses of multiomic data identified local translation as one mechanism underlying compartment-specific diversity. Together, these findings provide a comprehensive in vivo framework for understanding mitochondrial specialization across neuronal compartments.
    DOI:  https://doi.org/10.1101/2025.10.27.684882
  7. bioRxiv. 2025 Oct 13. pii: 2025.10.10.681769. [Epub ahead of print]
    Imaging Cellular Metabolic Rewiring with SuMMIT-SRS.
    Yajuan Li, Zhi Li, Yuhan Li, Kevin Rhine, Sural Ranamukhaarachchi, Shuo Qin, Hongje Jang, Jorge Villazon, Zhiliang Bai, Stephanie I Fraley, Gene W Yeo, Rong Fan, Lingyan Shi.
      Cells dynamically rewire their metabolic pathways in response to physiological and pathological cues. Such plasticity is particularly critical in neurons, stem cells, cancer cells, and immune cells, where biosynthetic demands can shift rapidly. However, current metabolic imaging techniques using isotope labeling typically track only one metabolite at a time, limiting their ability to capture the rapid dynamics of complex metabolic networks including coordinated precursor utilization, crosstalk, and turnover. Here, we present Subcellular Multiplexed Metabolic Isotope Tracing Stimulated Raman Scattering microscopy (SuMMIT-SRS), a platform that enables simultaneous visualization of multiple metabolic dynamics at subcellular resolution. By exploiting the distinct vibrational signatures of carbon-deuterium bonds derived from multiple deuterated amino acids, lipids, and monosaccharide tracers, SuMMIT-SRS maps co-regulated DNA, RNA, protein, and lipid synthesis at the same time and resolves various individual amino acid-mediated metabolic pathways within intact cells and tissues. We demonstrate SuMMIT's broad utility across Drosophila fat body tissue and developing brain, tumor organoids, aged human neurons, and mouse liver, capturing cell type-specific metabolic rewiring under genetic and pathological perturbations. This approach extends SRS to multiplexed isotope tracing, offering a powerful tool to uncover dynamic and complex biosynthesis programs in development, health, and disease.
    Keywords:  Metabolic rewiring; SRS; lipid; metabolism; multiplex; optical imaging; protein
    DOI:  https://doi.org/10.1101/2025.10.10.681769
  8. bioRxiv. 2025 Oct 23. pii: 2025.10.22.684010. [Epub ahead of print]
    Macropinocytosis enables metabolic recycling of extracellular DNA in cancer cells.
    Wen-Hsuan Yang, Olivia T Stamatatos, Evdokia Michalopoulou, Ava Sann, Paolo Cifani, Linda Van Aelst, Justin R Cross, Tse-Luen Wee, Michael J Lukey.
      Avid nutrient consumption is a metabolic hallmark of cancer and leads to regional depletion of key metabolites within the tumor microenvironment (TME). Cancer cells consequently employ diverse strategies to acquire the fuels needed for growth, including bulk uptake of the extracellular medium by macropinocytosis. Here, we show that breast and pancreatic cancer cells macropinocytically internalize extracellular DNA (exDNA), an abundant component of the TME, and deliver it to lysosomes for degradation. This provides a supply of nucleotides that sustains growth when de novo biosynthesis is impaired by glutamine restriction or pharmacological blockade. Mechanistically, this process is dependent on the non-redundant lysosomal equilibrative nucleoside transporter SLC29A3 (ENT3), which mediates the export of nucleosides from the lysosomal lumen into the cytosol. Accordingly, genetic ablation of SLC29A3 or pharmacological disruption of lysosomal function prevents exDNA scavenging and potently sensitizes breast tumors to antimetabolite chemotherapy in vivo . These findings reveal a previously unrecognized nutrient acquisition pathway through which cancer cells recycle exDNA into metabolic building blocks and highlight SLC29A3 as a mediator of metabolic flexibility and a potential target to improve chemotherapy response.
    DOI:  https://doi.org/10.1101/2025.10.22.684010
  9. Cell Rep. 2025 Nov 20. pii: S2211-1247(25)01355-5. [Epub ahead of print]44(12): 116583
    MICU2 controls mitochondrial calcium signaling and migration in neurons during development.
    Elena Berezhnaya, Benjamín Cartes-Saavedra, Raghavendra Singh, Macarena Rodríguez-Prados, Orly Reiner, Fowzan S Alkuraya, György Hajnóczky.
      Neurological disorders are linked to mitochondrial dysfunction and calcium overload. Mitochondrial calcium uptake is mediated by the mitochondrial calcium uniporter (mtCU), regulated by MICU1, which can be either homodimerized or heterodimerized with MICU2 or MICU3. Though MICU2 is scarce in the adult brain, MICU2 loss in patients leads to a neurodevelopmental disorder. We hypothesized that MICU2 is required for developmental calcium signaling and neuronal migration. MICU2 is present in the developing mouse brain but disappears by maturation, contrasting with other mtCU subunits that increase. MICU2 loss in mice does not affect cytoplasmic calcium but augments the mitochondrial matrix calcium rise in primary cortical neurons, leading to neuronal overmigration in the cortex and behavioral changes at 2 but not 12 months. Consistently, mitochondrial calcium uptake is not significantly affected in the adult animal cortex. MICU2-deficient patient fibroblasts copy the mitochondria-confined calcium alteration in developing neurons. Thus, MICU2 is important during neurodevelopment, likely by regulating the mtCU, and is eliminated by brain maturation.
    Keywords:  CP: cell biology; CP: neuroscience; MCU; MICU2; MICU3; anxiety; brain development; calcium signaling; mitochondria; neurodevelopmental disorders; neurons; radial migration
    DOI:  https://doi.org/10.1016/j.celrep.2025.116583
  10. Nat Metab. 2025 Nov 24.
    Mitochondrial NAD+ drives liver regeneration.
      
    DOI:  https://doi.org/10.1038/s42255-025-01414-7
  11. Proc Natl Acad Sci U S A. 2025 Dec 02. 122(48): e2527086122
    Isoform-specific regulation of PKM by acetylation.
    Dariia Pavlenko, Joaquin Tamargo-Azpilicueta, Hila Nudelman, Yuval Ankri, Anat Shahar, Irene Díaz-Moreno, Eyal Arbely.
      Pyruvate kinase (PK) is a crucial glycolytic protein involved in vital cellular processes ranging from cell proliferation to immune responses. The activity and functions of PK are tightly regulated by diverse mechanisms, including posttranslational Nϵ-lysine acetylation. Although previous studies have explored the impact of acetylation on selected lysine residues within the M2 isoform of PK (PKM2), a more comprehensive selection of acetylation sites and their respective effects on both PKM2 and the highly homologous PKM1 isoform is lacking. Here, we describe the structural, functional, and regulatory effects of site-specific acetylation on an expanded set of conserved lysines in PKM2 and selected lysines in PKM1. To study homogeneously acetylated proteins, we genetically encoded the incorporation of acetylated lysine into PKM variants expressed in bacteria and cultured mammalian cells. Our integrated biochemical, structural, and computational approach revealed K115 acetylation as an inhibitory modification in both PKM1 and PKM2 that stabilizes a closed active site conformation of the proteins. We also show that, in contrast to K115 acetylation, previously reported acetylation of K305 inhibits PKM2 but has no effect on the activity and oligomerization of PKM1. These findings propose the existence of both uniform and isoform-specific regulatory mechanisms of PKM, mediated by acetylation.
    Keywords:  deacetylation; genetic code expansion; glycolysis; metabolic regulation; posttranslational modifications
    DOI:  https://doi.org/10.1073/pnas.2527086122
  12. Nat Commun. 2025 Nov 28. 16(1): 10764
    CYFIP1 governs the development of cortical axons by modulating calcium availability.
    Carlotta Ricci, Maëllie Julie Midroit, Federico Caicci, Tilmann Achsel, Nuria Domínguez-Iturza, Claudia Bagni.
      The human CYFIP1 gene is linked to Autism Spectrum Disorder (ASD) and Schizophrenia (SCZ), both associated with brain connectivity defects and corpus callosum abnormalities. Previous studies demonstrated that Cyfip1-heterozygous mice exhibit diminished bilateral functional connectivity and callosal defects-resembling observations in ASD and SCZ patients. Here, we demonstrate that CYFIP1 is crucial for cortical axonal development and identify insufficient calcium uptake as the pivotal mechanism. In vivo, Cyfip1 heterozygosity delays callosal axon growth and arborization. Additionally, Cyfip1-deficient cortical neurons and axons have reduced intracellular calcium, along with impaired mitochondria morphology, activity, and motility. Mechanistically, CYFIP1 binds and stabilises the mRNA of specific voltage-gated calcium channel subunits, explaining the decreased calcium concentration in Cyfip1+/- cells. Notably, elevating intracellular calcium rescues delayed axonal growth and mitochondrial defects in Cyfip1-deficient neurons. These findings highlight that, by regulating mRNA metabolism, CYFIP1 ensures proper callosal development, offering insights into brain connectivity disruptions underlaying neurodevelopmental disorders.
    DOI:  https://doi.org/10.1038/s41467-025-65801-0
  13. PLoS One. 2025 ;20(11): e0335411
    The histone modification regulator, SIN3, plays a role in the cellular response to changes in glycolytic flux.
    Imad Soukar, Anindita Mitra, Linh Vo, Melanie Rofoo, Miriam L Greenberg, Lori A Pile.
      Epigenetic regulation and metabolism are connected. Epigenetic regulators, like the SIN3 complex, affect the expression of a wide range of genes, including those encoding metabolic enzymes essential for central carbon metabolism. The idea that epigenetic modifiers can sense and respond to metabolic flux by regulating gene expression has long been proposed. In support of this cross-talk, we provide data linking SIN3 regulatory action on a subset of metabolic genes with the cellular response to changes in metabolic flux. Furthermore, we show that loss of SIN3 is linked to decreases in mitochondrial respiration and the cellular response to mitochondrial and glycolytic stress. Data presented here provide evidence that SIN3 is important for the cellular response to metabolic flux change.
    DOI:  https://doi.org/10.1371/journal.pone.0335411
  14. Biosens Bioelectron. 2025 Nov 21. pii: S0956-5663(25)01127-3. [Epub ahead of print]295 118250
    Cellular metabolism biosensing: From extracellular microenvironment to intracellular physiology monitoring.
    Tao Liang, Chenlei Gu, Jiaru Fang, Dongxin Xu, Zhen Wang, Ciro Chiappini, Ning Hu.
      Metabolism is crucial for the growth, development, and life activities of organisms. Microsensors designed to monitor cellular metabolism are essential for investigation in cell culture. Electrochemical and optical sensors are dominantly applied to analyze classic metabolites in extracellular microenvironments, such as protons, oxygen, glucose, pyruvate, lactate, and reactive oxygen/nitrogen species. These methodologies facilitate the label-free, noninvasive, and continuous recording of transient effects, providing insights beyond static images and endpoint assays. This comprehensive review aims to critically evaluate and discuss advances in cellular metabolic monitoring, particularly focusing on the utilization of integrated biosensors in cell culture and detection systems, which are commonly termed as microphysiological systems. Initially, metabolic pathways are analyzed to elucidate the roles of key metabolites related to acidification, respiration, energy metabolism, and transient reactive species. Subsequently, this review comprehensively discusses sensing analytes, mechanisms, systems, cultures, and applications in advanced multimodal biosensing systems, including commercial products, as documented in references. Finally, current challenges and future directions associated with the development of more advanced metabolic biosensing systems are delineated to enhance their sensing capabilities, and meet the emerging clinical and scientific demands.
    Keywords:  Cellular metabolites; Extracellular microenvironment monitoring; Metabolic pathways; Microphysiological systems; Multimodal biosensing
    DOI:  https://doi.org/10.1016/j.bios.2025.118250
  15. Nat Commun. 2025 Nov 27. 16(1): 10694
    ER exit sites mediated by the COPII adaptor sec24D selectively recruit lipid raft-preferring proteins for rapid ER export.
    Ivan Castello-Serrano, Shikha Dagar, Rossana Ippolito, Kandice R Levental, Ilya Levental.
      The determinants of sub-cellular trafficking for many membrane proteins are poorly understood. Lipid-driven membrane nanodomains known as lipid rafts have been widely implicated in post-Golgi traffic, but their involvement in protein sorting in the endoplasmic reticulum has not been widely considered. To assess the role of membrane domains in the early secretory pathway, we use the Retention Using Selective Hooks system to synchronize and quantitatively assess trafficking rates and destinations of model proteins with tunable raft affinities. We find that raft-preferring constructs exit the ER faster than raft-excluded and have distinct preferences for ER exit sites marked by specific isoforms of sec24 cargo adaptors. Namely, raft-excluded cargo localizes to sec24A-positive sites while raft-preferring cargo localizes to sec24D ERES, dependent on p24-family cargo adapters TMED2/10. Finally, sec24D, but not sec24A, ERES accumulate a fluorescent cholesterol analog. These observations suggest that association with raft-like domains affects protein export from the ER.
    DOI:  https://doi.org/10.1038/s41467-025-65726-8
  16. Front Cell Dev Biol. 2025 ;13 1668779
    MitoLandscape, a semi-automated pipeline for subcellular localization and quantification of mitochondria.
    Enrico Negri, Virginia Fernández, Víctor Borrell.
      The precise characterization of mitochondrial morphology and subcellular localization provides crucial insights into cellular metabolic states and developmental fates. However, accurately analyzing mitochondria in cells with complex morphologies remains challenging, particularly within intact tissues where current methodologies lack sufficient resolution and specificity. Here we introduce MitoLandscape, an innovative pipeline tailored for comprehensive mitochondrial analysis at single-cell resolution in the developing nervous system. MitoLandscape integrates Airyscan super-resolution microscopy, semi-automated segmentation (leveraging ImageJ and 3DSlicer), machine-learning-driven pixel classification (ilastik), and a modular custom Python script for robust and versatile analysis. Using graph-based representations derived from manual annotations and binary mitochondrial images, MitoLandscape efficiently extracts detailed morphological parameters from distinct subcellular compartments, applicable from cells with simple morphologies to complex neuronal architectures. Additionally, the pipeline quantifies mitochondrial distribution relative to specific cellular landmarks, such as nucleus or soma. We validated MitoLandscape in vitro and in neural tissue, demonstrating its capability to precisely and reliably map mitochondrial features in diverse biological contexts. MitoLandscape represents a powerful, user-friendly, and highly adaptable solution for investigating mitochondrial biology in cell and developmental research.
    Keywords:  computational biology; machine learning; morphology; neurodevelopment; organelle; super-resolution
    DOI:  https://doi.org/10.3389/fcell.2025.1668779
  17. Nat Commun. 2025 Nov 27. 16(1): 10642
    The luminal domain region of Seipin/Fld1 is dispensable for establishing functional ER sites for lipid droplet biogenesis.
    Amit Khatri, Ritika Chaudhary, R Mankamna Kumari, Vineet Choudhary.
      Lipid droplet (LD) biogenesis occurs in the endoplasmic reticulum (ER), the mechanisms of which is not completely known. Seipin (Fld1 in yeast) is a crucial ER membrane protein that defines LD biogenesis sites. Here, we show that truncated seipin, Fld1-∆LR in yeast, and the human equivalent hSeipin-∆LR, mutants lacking the conserved luminal domain region (LR), functionally complement the LD biogenesis defect of fld1∆ mutants. Fld1-∆LR foci colocalize with factors: Nem1, Ldb16, Pex30 and Yft2, which are important for LD biogenesis and these sites become enriched in diacylglycerol upon stimulation of LD formation. Fld1-∆LR forms a homo-oligomeric complex facilitated by protein-protein interactions. We show that mutating the 31st proline abrogates the functioning of Fld1-∆LR. We demonstrate the critical regulatory role of LR of seipin in partitioning triacylglycerol into LDs. We conclude that LR of seipin is dispensable for establishing functional ER sites to recruit proteins for LD biogenesis.
    DOI:  https://doi.org/10.1038/s41467-025-65645-8
  18. bioRxiv. 2025 Oct 11. pii: 2024.11.07.622511. [Epub ahead of print]
    All-Optical Multimodal Mapping of Single Cell-Type-Specific Metabolic Activities via REDCAT.
    Yajuan Li, Zhaojun Zhang, Archibald Enninful, Negin Farzad, Presha Rajbhandari, Jungmin Nam, Xiaoyu Qin, Jorge Villazon, Anthony A Fung, Hongje Jang, Zhiliang Bai, Nancy R Zhang, Brent R Stockwell, Rong Fan, Mina L Xu, Zongming Ma, Lingyan Shi.
      Metabolism underlies cell growth, survival, and function, yet its activities vary widely across cell types and tissue environments. Spatially resolving these processes in situ at single-cell resolution is essential to advance our understanding of cellular function and tissue physiology in health and disease. However, existing approaches are limited by either destructive workflows, insufficient spatial resolution and biochemical specificity, or lack of direct linkage to cell identity. Here, we present Raman Enhanced Delineation of Cell Atlases in Tissues (REDCAT), a multimodal all-optical platform that integrates stimulated Raman scattering, autofluorescence redox imaging, second harmonic generation, and high-plex immunofluorescence to co-map metabolic activities and cell types within the same tissue section. REDCAT achieves subcellular resolution profiling of protein, lipid, redox, and nuclear acid metabolism, together with extracellular matrix composition, in both FFPE and fresh-frozen human tissues. Applied to normal lymph nodes, REDCAT delineated distinct redox and lipid remodeling programs across germinal center B-cell zones and immune subsets, highlighting cell-type-specific metabolic specialization. In lymphoma, it revealed profound metabolic reprogramming, including extensive lipid accumulation, nuclear metabolic heterogeneity, and a transitional metabolic state associated with transformation from chronic lymphocytic leukemia to diffuse large B-cell lymphoma, thereby illuminating tumor evolution in situ . In human liver, REDCAT resolved cell-type-specific lipid droplet diversity and zonation-dependent nuclear metabolic gradients, uncovering new principles of spatial metabolic organization. By directly linking cell identity with spatial metabolic states at single-cell or subcellular resolution, REDCAT establishes a broadly applicable framework for studying immune function, tumor progression, and tissue physiology, and offers a new path to deciphering the metabolic basis of health and disease.
    DOI:  https://doi.org/10.1101/2024.11.07.622511
  19. bioRxiv. 2025 Oct 14. pii: 2025.10.13.682232. [Epub ahead of print]
    Homeostatic regulation of intrinsic lipid curvature in eukaryotic cells.
    Daniel Milshteyn, Jacob R Winnikoff, Elida Kocharian, Aaron M Armando, Edward A Dennis, Peter R Girguis, Itay Budin.
      Cell membranes are composed of both bilayer-supporting and non-bilayer phospholipids, with the latter's negative intrinsic curvature aiding in membrane trafficking and the dynamics of membrane proteins. Phospholipid metabolism has long been recognized to maintain membrane fluidity, but whether it also acts to maintain the function of high-curvature lipids is not resolved. Here, we find that cells grown under hydrostatic pressure - used to artificially reduce lipid curvature - maintain lipidome curvature through metabolic acclimation. We first observed that manipulation of the lipidome curvature via the phosphatidylethanolamine (PE) to phosphatidylcholine (PC) ratio affects high-pressure growth and viability of yeast independently of membrane fluidity. In wild-type cells, X-ray scattering measurements revealed an increased propensity for lipid extracts to form non-lamellar phases after extended pressure incubations. Unexpectedly, this change in phase behavior was not due to increased levels of PE, but of phosphatidylinositol (PI), the only major phospholipid class whose curvature had not been previously characterized. We found that PI is a non-bilayer lipid, with a negative curvature intermediate to that of PE and PC. Accounting for PI, mean lipidome curvature was defended in response to pressure by two distantly related yeasts. Lipidome curvature also responded to pressure in a human cancer cell line through ether phospholipid metabolism and chain remodeling, but not in bacterial cells. These findings indicate that eukaryotic phospholipid metabolism uses diverse mechanisms to maintain curvature frustration in cell membranes.
    DOI:  https://doi.org/10.1101/2025.10.13.682232
  20. Nat Commun. 2025 Nov 28. 16(1): 10765
    Visualizing intraorganellar ultrastructures, dynamics, and interactions with open-access background-free Lock-in-SIM.
    Wenjie Liu, Meng Zhang, Wenbin Zhu, Shunyu Xie, Jinfeng Zhang, Shuhao Qian, Zhiyi Liu, Xiu Zheng, Qiuyuan Fang, Wei Yang, Yi Wang, Dazhao Zhu, Jianjie Dong, Xu Liu, Youhua Chen, Cuifang Kuang, Yu-Hui Zhang, Lothar Schermelleh.
      Structured illumination microscopy (SIM) is a powerful method for fast and gentle live-cell super-resolution imaging. However, its susceptibility to reconstruction artifacts from out-of-focus blur and background imposes substantial barriers to analyze the dynamics of densely packed volumetric intraorganellar ultrastructures that are typically in a size range of SIM's spatial resolution. To address this limitation, we have developed Lock-in-SIM, an open-access two-dimensional SIM framework that eliminates background and maximizes the recovery of sub-diffraction information with the highest possible frequency extraction. By leveraging the intrinsic modulation differences of volumetric sample structures, Lock-in-SIM enables efficient optical sectioning, extends imaging depth, and improves data fidelity and quantifiability. We demonstrate the superiority of Lock-in-SIM by visualizing various challenging intraorganellar ultrastructures in live cells. Our investigations uncover mechanisms of mitochondrial fission and endoplasmic reticulum-lysosome interactions and provide insights into the intricate yet highly regulated structural remodeling of organelles.
    DOI:  https://doi.org/10.1038/s41467-025-65805-w
  21. bioRxiv. 2025 Oct 25. pii: 2025.10.24.684471. [Epub ahead of print]
    pH-dependent regulation in SLC38A9.
    Xuelang Mu, Ampon Sae Her, Tamir Gonen.
      Cells rely on precise metabolic control to adapt to environmental cues. The mechanistic target of rapamycin complex 1 (mTORC1) senses nutrient availability, with amino acids serving as key signals. Lysosomes, which act as nutrient recycling centers, maintain amino acid homeostasis by breaking down macromolecules and releasing amino acids for cellular use. SLC38A9, a lysosomal amino acid transporter, functions as both a transporter and a sensor in the mTORC1 pathway. Here, we investigated whether SLC38A9 activity is regulated by pH. We show that arginine uptake by SLC38A9 is pH-dependent, and that the histidine residue His544 serves as the pH sensor. Mutating His544 abolishes the pH dependence of arginine uptake without impairing overall transport activity, indicating that His544 is not directly involved in substrate binding. Instead, protonation or deprotonation of His544 appears to influence transport through SLC38A9. To explore this mechanism, we compared two structures of SLC38A9 that we determined, one at high pH and one at low pH, and proposed a working model for pH-induced activation. These findings highlight the role of local ionic changes in modulating lysosomal transporters and underscore the intricate regulatory mechanisms that govern SLC38A9 function and, ultimately, mTORC1 signaling.
    DOI:  https://doi.org/10.1101/2025.10.24.684471
  22. Nat Commun. 2025 Nov 25. 16(1): 10490
    Multi-omic analysis reveals lipid dysregulation associated with mitochondrial dysfunction in parkinson's disease brain.
    Jenny Hällqvist, Christina E Toomey, Rui Pinto, Tomas Baldwin, Ivan Doykov, Anna Wernick, Mesfer Al Shahrani, James R Evans, Joanne Lachica, Simon Pope, Simon Heales, Simon Eaton, Kevin Mills, Sonia Gandhi, Wendy E Heywood.
      Parkinson's disease (PD) is an increasingly prevalent neurodegenerative disorder, largely sporadic in origin, with limited understanding of age- and region-specific lipid alterations in the human brain. Dysregulation of glycosphingolipid catabolism has been implicated in PD, yet comprehensive spatiotemporal profiling remains sparse. Here, we performed targeted lipidomics across eight anatomically distinct brain regions in post-mortem controls, mid-stage, and late-stage PD cases using high-precision tissue dissection. Each region displayed distinct lipid signatures, with several age-associated alterations-most notably in hexosylceramides, including glucosylceramide. In PD, glycosphingolipids were reduced in subcortical regions but elevated in cortical regions, particularly gangliosides, HexCer, and Hex2Cer, accompanied by increased sphingolipids and decreased phospholipids. The most pronounced mid-stage changes occurred in the putamen, where very long chain ceramide species and plasmalogen PE decreased, then normalising in late-stage disease. Lyso-phosphatidylcholine increased progressively throughout PD progression. Integrating proteomic data, we observed sphingomyelin levels associated with PD-related proteins, while dysregulated mitochondrial function correlated with antioxidant plasmalogens, long-chain ceramides, lyso-phosphatidylcholine, and HexCer in the putamen. These findings highlight region- and stage-specific lipid alterations in PD and their potential convergence with mitochondrial dysfunction.
    DOI:  https://doi.org/10.1038/s41467-025-65489-2
  23. Drug Deliv. 2025 Dec 31. 32(1): 2585612
    A surrogate barrier model for high-throughput blood-brain barrier permeability prediction: integrating LLC-PK1-MOCK/MDR1 Cells and lysosomal trapping correction.
    Juanwen Hu, Xue Jiang, Cong Li, Qiannan Zhang, Xia Wu, Wenpeng Zhang, Xiaomei Zhuang.
      To mitigate risks in central nervous system (CNS) drug development, we established a high-throughput in vitro blood-brain barrier (BBB) model using LLC-PK1-MOCK and LLC-PK1-MDR1 cells in a Transwell system, aiming to replicate in vivo brain distribution and elucidate permeability mechanisms. Model integrity was assessed via transepithelial electrical resistance (TEER) and efflux functionality using control drugs (atenolol, digoxin). Bidirectional transport studies of 41 compounds quantified permeability (Papp), efflux ratios (ER), and recoveries, while in vivo brain distribution parameters (Kp,uu,brain) were derived from literature and rat studies. The model demonstrated critical BBB features: tight junction integrity (TEER > 70 Ω·cm2), P-gp efflux activity (digoxin ER = 5.10 ~ 17.12), and discrimination of passive diffusion (63.41% of drugs) from transporter-mediated mechanisms (19.5% P-gp substrates). A training set of 20 randomly selected drugs revealed a robust correlation between MDR1-derived Papp(A-B) and Kp,uu,brain (R = 0.8886), with the remaining 21 compounds validating predictive accuracy (≤2-fold error). Four alkaloids exhibiting low recovery (<80%) due to lysosomal trapping were corrected using Bafilomycin A1, aligning their permeability with in vivo outcomes. These results position the LLC-PK1-MOCK/MDR1 model as a reliable surrogate tool for early CNS drug screening, enabling rapid prioritization of candidates based on BBB penetration potential. Its integration into preclinical workflows promises to accelerate the development of therapeutics for neurological disorders.
    Keywords:  CNS drug; LLC-PK1-MDR1 cells; blood-brain barrier; surrogate barrier model
    DOI:  https://doi.org/10.1080/10717544.2025.2585612
  24. J Cell Biol. 2026 Jan 05. pii: e202503081. [Epub ahead of print]225(1):
    Homocysteine disrupts lysosomal function by V-ATPase inhibition.
    Yang Yang, Qianjin Kong, Chaolian Liu, Fengyang Wang, Meijiao Li, Shalan Li, Yuehui Shi, Leonard Krall, Xin Wang, Shan He, Kai Jiang, Xuna Wu, Mei Yang, Chonglin Yang.
      Lysosomes are degradation and signaling organelles central to metabolic homeostasis. It remains unclear whether and how harmful metabolites compromise lysosome function in the etiopathology of metabolic disorders. Combining Caenorhabditiselegans and mouse models, we demonstrate that homocysteine, an intermediate in methionine-cysteine metabolism and the cause of the life-threatening disease homocystinuria, disrupts lysosomal functions. In C. elegans, mutations in cystathionine β-synthase cause strong buildup of homocysteine and developmental arrest. We reveal that homocysteine binds to and homocysteinylates V-ATPase, causing its inhibition and consequently impairment of lysosomal degradative capacity. This leads to enormous enlargement of lysosomes with extensive cargo accumulation and lysosomal membrane damage in severe cases. Cbs-deficient mice similarly accumulate homocysteine, displaying abnormal or damaged lysosomes reminiscent of lysosomal storage diseases in multiple tissues. These findings not only uncover how a metabolite can damage lysosomes but also establish lysosomal impairment as a critical contributing factor to homocystinuria and homocysteine-related diseases.
    DOI:  https://doi.org/10.1083/jcb.202503081
  25. Proc Natl Acad Sci U S A. 2025 Dec 02. 122(48): e2509798122
    Modulation of the PGRMC1/NLRP7/HLA-C axis by autophagy is linked to both spontaneous preterm birth and gestational choriocarcinoma.
    Qin Li, Huaxiao Yu, Dongyi Ling, Jie Zhou, Shanshan Zhen, Yangyang Li, Lijuan Wang, Zetong Zheng, Wenjian Cen, Hongyi Gao, Hui Chen, Jack L Strominger, Ziming Du.
      Spontaneous preterm birth (SPTB) and gestational choriocarcinoma are both associated with complex physiological processes that significantly impact maternal health. While the molecular mechanisms underlying SPTB and gestational choriocarcinoma remain poorly understood, emerging evidence suggests that immune regulation plays a crucial role in both conditions. In this study, we revealed that progesterone regulates autophagy via the noncanonical progesterone receptor membrane component 1 (PGRMC1), which then modulates NLRP7 levels, thereby impacting HLA-C expression in the JEG3 cell line, an extravillous trophoblast (EVT) model. Furthermore, a significant positive correlation between NLRP7 and HLA-C expression was observed in EVTs from placental tissues and choriocarcinoma samples. In cases of SPTB, we found both reduced expression of NLRP7 and HLA-C in EVTs. Similarly, in gestational choriocarcinoma samples, we observed significantly lower expression levels of NLRP7 and HLA-C, further suggesting a shared immune evasion mechanism. These findings not only provide insights into the molecular mechanisms underlying both SPTB and choriocarcinoma but also identify the progesterone-driven NLRP7/HLA-C axis as a promising target for therapeutic intervention, offering strategies for improving outcomes in both conditions.
    Keywords:  HLA-C; NLRP7; choriocarcinoma; spontaneous preterm birth
    DOI:  https://doi.org/10.1073/pnas.2509798122
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