bims-celmim Biomed News
on Cellular and mitochondrial metabolism
Issue of 2026–01–18
fifteen papers selected by
Marc Segarra Mondejar, AINA
  1. Purinosomes and mitochondria: Dynamic interactomes at the crossroads of metabolism, cell structure, and disease.
  2. Inactive ryanodine receptors sustain lysosomal availability for autophagy by promoting ER-lysosomal contact site formation.
  3. Hexokinase detachment from mitochondria drives the Warburg effect to support compartmentalized ATP production.
  4. Nucleotide metabolic rewiring enables NLRP3 inflammasome hyperactivation in obesity.
  5. HEPES in Cell Culture Alters the Multi-Omics Profile Exhibited by Gaucher Disease Fibroblasts.
  6. Coenzyme A mitigates cystine-deprivation-induced ferroptosis by suppressing the iron-starvation response.
  7. AMPK at the interface of nutrient sensing, metabolic flux and energy homeostasis.
  8. Metabolic adaptation in CD4+ T cells promotes immunosuppression in sepsis.
  9. Cancer might evade immune defences by stealing mitochondria.
  10. Heterogeneity, dynamics and organelle interactions of lipid droplets.
  11. Integrated proteomics and metabolomics reveal phytosesquiterpene lactones inhibit TNBC cell activity by depleting ATP synthesis and reprogramming primary metabolism.
  12. Nvj3 regulates Dga1-mediated triacylglycerol synthesis and lipid droplet formation at ER contact sites.
  13. Mitochondrial fission during mitophagy requires both inner and outer mitofissins.
  14. Autophagy Cholesterol Axis Remodeling Supports Malignant Progression and Chemoresistance in Glioma.
  15. Self-Calibrating Ratiometric DNA Aptamer Probe for Quantitative ATP Imaging in Living Cells.
  1. Pharmacol Res. 2026 Jan 10. pii: S1043-6618(26)00011-3. [Epub ahead of print]224 108096
    Purinosomes and mitochondria: Dynamic interactomes at the crossroads of metabolism, cell structure, and disease.
    Andrea Mancini, Elisabetta Betterini, Matteo Bordi, Francesco Cecconi.
      Mitochondria are central hubs of cellular metabolism, integrating nutrient catabolism, ATP production, redox balance, and biosynthetic precursor supply. Recent work has revealed that their influence extends beyond canonical bioenergetics to include intimate connections with cytosolic multi-enzyme assemblies. Among these, the purinosome, the complex dedicated to de novo purine biosynthesis, has emerged as a paradigmatic example of how metabolic pathways achieve efficiency through spatial and functional coupling. This Review highlights the dynamic interplay between purinosomes and mitochondria. We describe how mitochondrial metabolism supplies key substrates, including aspartate, glycine, and formate, while oxidative phosphorylation provides the ATP required for nucleotide synthesis. We discuss how purinosomes assemble through liquid-liquid phase separation, position near mitochondria in response to energetic stress, and act as adaptive metabolic hubs that sense and integrate growth and nutrient signals. Finally, we examine how disruption of this mitochondrion-purinosome axis contributes to disease, from rare neurodevelopmental disorders to cancer and neurodegeneration.
    Keywords:  Cancer biology; Metabolons; Mitochondria metabolism; Nucleotide metabolism; Organelle contact sites; Purine synthesis
    DOI:  https://doi.org/10.1016/j.phrs.2026.108096
  2. Nat Commun. 2026 Jan 15.
    Inactive ryanodine receptors sustain lysosomal availability for autophagy by promoting ER-lysosomal contact site formation.
    Tim Vervliet, Jens Loncke, Marko Sever, Karan Ahuja, Chris Van den Haute, Tomas Luyten, Grace E Stutzmann, Catherine Verfaillie, Tihomir Tomašič, Geert Bultynck.
      Lysosomal and endoplasmic reticulum (ER) Ca2+ release mutually influence each other's functions. Recent work revealed that ER-located ryanodine receptor(s) (RyR(s)) Ca2+ release channels suppress autophagosome turnover by the lysosomes. In familial Alzheimer's disease, inhibiting RyR hyperactivity restored autophagic flux by normalizing lysosomal vacuolar H+-ATPase (vATPase) levels. However, the mechanisms by which RyRs control lysosomal function and how this involves the vATPase remain unknown. Here, we show that RyRs interact with the ATP6v0a1 subunit of the vATPase, contributing to ER-lysosomal contact site formation. This interaction suppresses RyR-mediated Ca²⁺ release, leading to reduced lysosomal exocytosis. Pharmacological inhibition of RyR activity mimics these effects on lysosomal exocytosis. Retaining lysosomes inside cells via RyR inhibition increases ER-lysosomal contact site formation, rendering lysosomes more available for autophagic flux. In summary, these findings establish RyR/ATP6v0a1 complexes as ER-lysosomal tethers that dynamically and Ca2+ dependently regulate the intracellular availability of lysosomes to participate in autophagic flux.
    DOI:  https://doi.org/10.1038/s41467-025-68054-z
  3. Nat Metab. 2026 Jan 16.
    Hexokinase detachment from mitochondria drives the Warburg effect to support compartmentalized ATP production.
    Kimberly S Huggler, Kyle M Flickinger, Matthew H Forsberg, Carlos A Mellado Fritz, Gavin R Chang, Meghan F McGuire, Christian M Capitini, Jason R Cantor.
      Hexokinase (HK) catalyses the phosphorylation of glucose to glucose 6-phosphate, marking the first step of glucose metabolism. Most cancer cells co-express two homologous HK isoforms, HK1 and HK2, which can each bind the outer mitochondrial membrane (OMM). CRISPR screens performed across hundreds of cancer cell lines indicate that both isoforms are dispensable for growth in conventional culture media. By contrast, HK2 deletion impaired cell growth in human plasma-like medium. Here we show that this conditional HK2 dependence can be traced to the subcellular distribution of HK1. Notably, OMM-detached (cytosolic) rather than OMM-docked HK supports cell growth and aerobic glycolysis (the Warburg effect), an enigmatic phenotype of most proliferating cells. We show that under conditions promoting increased translocation of HK1 to the OMM, HK2 is required for cytosolic HK activity to sustain this phenotype, thereby driving sufficient glycolytic ATP production. Our results reveal a basis for conditional HK2 essentiality and suggest that demand for compartmentalized ATP synthesis explains why cells engage in aerobic glycolysis.
    DOI:  https://doi.org/10.1038/s42255-025-01428-1
  4. Science. 2026 Jan 15. 391(6782): eadq9006
    Nucleotide metabolic rewiring enables NLRP3 inflammasome hyperactivation in obesity.
    Danhui Liu, Chuanli Zhou, Xiaochen Wang, Zhou Luo, Ruiyao Xu, Shanshan Huo, Lina Guo, Xuemei Luo, Shuhan Yang, Arielle Click, Janiece Vancil, Paola Barajas, Victor Mijares, Hamid Baniasadi, Nan Yan, Jan Rehwinkel, Dustin C Hancks, Elizabeth H Chen, Shuang Liang, Zhenyu Zhong.
      Obesity is a major disease risk factor due to obesity-associated hyperinflammation. We found that obesity induced Nod-like receptor pyrin domain-containing 3 (NLRP3) inflammasome hyperactivation and excessive interleukin (IL)-1β production in macrophages by disrupting SAM and HD domain-containing protein 1 (SAMHD1), a deoxynucleoside triphosphate (dNTP) hydrolase crucial for nucleotide balance. This caused aberrant accumulation of dNTPs, which can be transported into mitochondria, and initiated mitochondrial DNA (mtDNA) neosynthesis, which increased the presence of oxidized mtDNA and triggered NLRP3 hyperactivation. Deletion of SAMHD1 promoted NLRP3 hyperactivation in cells isolated from zebrafish, mice, and humans. SAMHD1-deficient mice showed elevated circulating IL-1β, insulin resistance, and metabolic dysfunction-associated steatohepatitis. Blocking dNTP mitochondrial transport prevented NLRP3 hyperactivation in macrophages from obese patients and SAMHD1-deficient mice. Our study revealed that obesity by inhibiting SAMHD1 rewired macrophage nucleotide metabolism, thereby triggering NLRP3 inflammasome hyperactivation to drive disease progression.
    DOI:  https://doi.org/10.1126/science.adq9006
  5. J Cell Biochem. 2026 Jan;127(1): e70080
    HEPES in Cell Culture Alters the Multi-Omics Profile Exhibited by Gaucher Disease Fibroblasts.
    Eleonore M Corazolla, Bauke V Schomakers, Maria M Trętowicz, Jill Hermans, Michel van Weeghel, Frédéric M Vaz, Mia L Pras-Raves, Karen Ghauharali-van der Vlugt, Femke S Beers-Stet, Susanna M I Goorden, Judith Jansen-Meijer, Georges E Janssens, Carla E M Hollak, Riekelt H Houtkooper, André B P van Kuilenburg.
      Lysosomal function can be affected by components in cell culture. This in turn may influence cellular metabolism and, consequently, research and diagnostics outcomes. One such component is the commonly used pH buffer 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES). HEPES specifically impacts the trafficking of the lysosomal enzyme glucocerebrosidase, which is deficient in Gaucher disease (GD). Understanding how HEPES affects cellular models of GD is essential, since glucocerebrosidase is central to diagnostic testing and the investigation of GD pathophysiology. Therefore, we examined the broader effects of HEPES on cultured fibroblasts from individuals with GD and healthy controls. We cultured dermal fibroblasts of eight adults with GD and seven healthy age- and sex-matched controls. The cells were cultured in two culture media, Ham's F10 and DMEM, both with and without HEPES. We assessed glucocerebrosidase enzyme activity and sphingolipid concentrations using a quantitative UPLC-MS/MS method. Additionally, we conducted multi-omics analyses, consisting of lipidomics, metabolomics and proteomics, to explore the broader impact of HEPES in cell culture on fibroblasts. Glucocerebrosidase activity in cell lysates increased after HEPES exposure in both GD and control fibroblasts, to an extent that may influence diagnostic outcomes. In GD fibroblasts, substrate accumulation was absent and not altered by HEPES exposure. GD fibroblasts exhibited a multi-omics profile largely overlapping with healthy controls and lacking the typical pathological features associated with GD in other cell types, such as mitochondrial dysfunction, dysregulated autophagy, disruption of intracellular calcium homeostasis, ER stress and chronic oxidative stress. In addition, the multi-omics profile was altered by HEPES, however in a non-specific manner. In conclusion, HEPES influences fibroblasts in culture, both from healthy controls and from patients with GD. Furthermore, GD fibroblasts lack a specific disease-related profile. This renders cultured fibroblasts unsuitable for studying pathophysiological processes in GD. Culturing GD fibroblasts with HEPES may compromise the reliability of diagnostics.
    Keywords:  Gaucher disease; HEPES; cell culture techniques; multi‐omics
    DOI:  https://doi.org/10.1002/jcb.70080
  6. FEBS J. 2026 Jan 16.
    Coenzyme A mitigates cystine-deprivation-induced ferroptosis by suppressing the iron-starvation response.
    Mingjun Tan, Yunzhan Li, Lei Sang, Min Wang, Qin Li, Zekun Li, Dongxiao Sun, Xiuchao Wang, Shengyu Yang.
      The labile iron pool in the cell is required for ferroptosis, a form of regulated cell death resulting from excessive lipid peroxidation and membrane damage. Glutathione (GSH) is critical for lipid-peroxide scavenging, and cysteine is the rate-limiting amino acid in GSH synthesis. Cysteine metabolism intricately intertwines with iron metabolism, either directly by participating in assembly of the iron-sulfur cluster or indirectly through the pantothenate pathway and coenzyme A (CoA) synthesis. However, the regulation of iron homeostasis in cystine (Cys2)-deprivation-induced ferroptosis is poorly understood. Here, we show that Cys2 deprivation promotes ferroptosis, at least in part, by activating the iron-starvation response (ISR), and CoA can mitigate ferroptosis by suppressing the ISR. Mechanistically, Cys2 deprivation promotes the oxidation of cytosolic iron-sulfur clusters to activate the ISR; CoA and related small-molecule thiols in the pantothenate pathway suppress the ISR and ferroptosis by preventing the oxidation of iron-sulfur clusters in Cys2-deprived cells. Our findings provide important insight into the regulation of the ISR in Cys2-deprivation-induced ferroptosis, and show that CoA can protect cells from Cys2-deprivation-induced ferroptosis by suppressing the ISR.
    Keywords:  Coenzyme A; cysteine; cystine‐deprivation; ferroptosis; iron‐starvation response; iron–sulfur cluster; pantothenate pathway
    DOI:  https://doi.org/10.1111/febs.70411
  7. Nat Metab. 2026 Jan 12.
    AMPK at the interface of nutrient sensing, metabolic flux and energy homeostasis.
    Tyler K T Smith, Logan K Townsend, William J Smiles, Jonathan S Oakhill, Morgan D Fullerton, Gregory R Steinberg.
      The orchestration of cellular metabolism requires the integration of signals related to energy stores and nutrient availability through multiple overlapping mechanisms. AMP-activated protein kinase (AMPK) is a pivotal energy sensor that responds to reductions in adenylate charge; however, studies over the past decade have also positioned AMPK as a key integrator of nutrient-derived signals that coordinate metabolic function. This Review highlights recent advances in our understanding of how AMPK senses nutrients and regulates metabolic activity across tissues, timescales and cell types. These effects are mediated through the phosphorylation of substrates involved in metabolite trafficking, mitochondrial function, autophagy, transcription, ubiquitination, proliferation and cell survival pathways, including ferroptosis. Particular attention is given to the role of AMPK in the pathophysiology of obesity, type 2 diabetes, metabolic dysfunction-associated steatotic liver disease, cardiovascular and renal diseases, neurodegenerative disorders and cancer. Collectively, these findings reinforce AMPK as a central metabolic node that aligns cellular behaviour with energetic demand. Continued investigation into its nutrient-sensing mechanisms holds promise for identifying new strategies to restore metabolic balance in disease.
    DOI:  https://doi.org/10.1038/s42255-025-01442-3
  8. Nat Immunol. 2026 Jan 15.
    Metabolic adaptation in CD4+ T cells promotes immunosuppression in sepsis.
    Nicole M Chapman, Hongbo Chi.
      
    DOI:  https://doi.org/10.1038/s41590-025-02403-4
  9. Nature. 2026 Jan 14.
    Cancer might evade immune defences by stealing mitochondria.
    Laura Dattaro.
      
    Keywords:  Cancer; Immunology; Metabolism
    DOI:  https://doi.org/10.1038/d41586-026-00123-9
  10. Nat Rev Mol Cell Biol. 2026 Jan 15.
    Heterogeneity, dynamics and organelle interactions of lipid droplets.
    W Mike Henne, Sarah Cohen.
      Lipid droplets (LDs) are emerging as key factors in cellular physiology, with roles beyond energy storage, including metabolic homeostasis, signalling and development. Together with a growing list of functions, diverse LD populations are being identified in different tissue types as well as within the context of single cells. Here we summarize recent work highlighting LD diversity from three perspectives: their lipid and protein compositional heterogeneity; differences in abundance, size and spatial organization within cells; and the diverse contacts they form with other organelles, all of which contribute to LD function. We also discuss tools and approaches used to visualize LD heterogeneity, the role of LDs in pathophysiology and disease, and open questions in the field.
    DOI:  https://doi.org/10.1038/s41580-025-00945-x
  11. Sci Rep. 2026 Jan 14.
    Integrated proteomics and metabolomics reveal phytosesquiterpene lactones inhibit TNBC cell activity by depleting ATP synthesis and reprogramming primary metabolism.
    Jeng-Yuan Shiau, Han-Jung Huang, Kyoko Nakagawa-Goto, Lie-Fen Shyur.
      Phytosesquiterpene lactones deoxyelephantopin (DET) and its derivative DETD-35 are reported to induce oxidative stress towards inhibiting triple-negative breast cancer (TNBC) cell activities. This study aimed to elucidate how DET and DETD-35 affect mitochondrial function and systemic metabolism in TNBC cells. DET and DETD-35 promoted mitochondrial superoxide production by upregulating expression of SOD1 and SOD2, induced permeability transition pore opening, and attenuated intracellular ATP levels. Neither compound interfered with mitochondrial respiration/bioenergetics in normal mammary MCF-10A cells. Comparative mitochondrial proteome and bioinformatic analyses showed significant deregulation of proteins related to the oxidative phosphorylation, depolarization of mitochondria, and apoptosis signaling in DET- or DETD-35-treated TNBC cells, and primary metabolomics revealed that both compounds deregulated metabolites dynamics and the corresponding metabolic pathways in TNBC cells. Knockdown of the PRKCA gene/protein involved in inducing mitochondrial toxicity in TNBC cells reversed cytotoxicity, apoptosis, and the levels of several metabolites induced by DET or DETD-35 in the cancer cells. Integrated Pearson's correlation and IPA network analyses of differentially expressed proteins and metabolites revealed the networks of ATP synthesis, energy homeostasis, and respiration, depolarization, and transmembrane potential in mitochondria highly correlated to the compound effects. Notable, DET/DETD-35 inhibited mitochondrial ATPase activity, and molecular modeling further predicted the binding sites of either compound with ATP synthase at the subunits α/β and c/a interfaces. The overexpression of ATP synthase-related proteins ATP5A1 and ATP5C1 in the tumor microenvironment of MDA-MB-231 xenograft mice were also significantly suppressed by DET and DETD-35 treatments. In summary, this study identifies DETD-35 and DET as novel ATPase inhibitors which are attributed to disrupting mitochondrial biogenetics and cellular metabolism and networking in TNBC cells.
    Keywords:  ATP synthase; Breast cancer; Metabolic reprogramming; Mitochondrial dysfunction; Mitochondrial proteome; Sesquiterpene lactone
    DOI:  https://doi.org/10.1038/s41598-026-35194-1
  12. bioRxiv. 2026 Jan 09. pii: 2026.01.05.695145. [Epub ahead of print]
    Nvj3 regulates Dga1-mediated triacylglycerol synthesis and lipid droplet formation at ER contact sites.
    Daniel Adebayo, Eseiwi Obaseki, Scott Miller, Marwa Aboumourad, Zaid Saadeh, Zachary Pomicter, Lindsey Kreinbring, Mohamad Rahal, Kashvi Vasudeva, Garrett Frost, Reema Smadi, Hanaa Hariri.
      Lipid droplet (LD) biogenesis is essential for lipid homeostasis during nutrient stress, yet how lipid intermediates are spatially organized to support efficient triacylglycerol (TAG) synthesis remains unclear. Here, we identify Nvj3 as a nutrient-responsive regulator that links diacylglycerol (DAG) availability to TAG synthesis and LD formation at the endoplasmic reticulum (ER). Nvj3 is induced by glucose depletion and recruited to LD-associated ER domains. Loss of Nvj3 causes neutral lipid accumulation under steady state conditions but delays TAG synthesis under acute inducible metabolic transitions. Using controlled TAG induction systems, we show that Nvj3 is required to couple Dga1-dependent TAG synthesis to LD formation. In the absence of Nvj3, TAG accumulates but remains inefficiently packaged into LDs. Consistent with this defect, nvj3Δ cells exhibit altered phospholipid remodeling and mislocalization of DAG away from ER domains during starvation. Together, these findings establish Nvj3 as an organizer of lipid availability during metabolic stress and suggest that spatial control of DAG is a key determinant of LD biogenesis.
    Summary: This study identifies Nvj3 as a spatial organizer of Dga1-dependent lipid droplet formation. Nvj3 promotes proper diacylglycerol positioning, and enables efficient triacylglycerol synthesis during metabolic stress. We propose that Nvj3 regulates lipid flux through spatial compartmentalization of diacylglycerol at membrane contact site-associated ER domains.
    DOI:  https://doi.org/10.64898/2026.01.05.695145
  13. EMBO Rep. 2026 Jan 13.
    Mitochondrial fission during mitophagy requires both inner and outer mitofissins.
    Kentaro Furukawa, Tatsuro Maruyama, Yuji Sakai, Shun-Ichi Yamashita, Keiichi Inoue, Tomoyuki Fukuda, Nobuo N Noda, Tomotake Kanki.
      Mitophagy maintains mitochondrial homeostasis through the selective degradation of damaged or excess mitochondria. Recently, we identified mitofissin/Atg44, a mitochondrial intermembrane space-resident fission factor, which directly acts on lipid membranes and drives mitochondrial fission required for mitophagy in yeast. However, it remains unclear whether mitofissin is sufficient for mitophagy-associated mitochondrial fission and whether other factors act from outside mitochondria. Here, we identify a mitochondrial outer membrane-resident mitofissin-like microprotein required for mitophagy, and we name it mitofissin 2/Mfi2 based on the following results. Overexpression of an N-terminal Atg44-like region of Mfi2 induces mitochondrial fragmentation and partially restores mitophagy in atg44Δ cells. Mfi2 binds to lipid membranes and mediates membrane fission in a cardiolipin-dependent manner in vitro, demonstrating its intrinsic mitofissin activity. Coarse-grained molecular dynamics simulations further support the stable interaction of Mfi2 with cardiolipin-containing bilayers. Genetic analyses reveal that Mfi2 and the dynamin-related protein Dnm1 independently facilitate mitochondrial fission during mitophagy. Thus, Atg44 and Mfi2, two mitofissins with distinct localizations, are required for mitophagy-associated mitochondrial fission.
    Keywords:  Atg44; Mfi2; Mitochondrial Fission; Mitofissin; Mitophagy
    DOI:  https://doi.org/10.1038/s44319-025-00689-x
  14. bioRxiv. 2026 Jan 07. pii: 2026.01.06.697885. [Epub ahead of print]
    Autophagy Cholesterol Axis Remodeling Supports Malignant Progression and Chemoresistance in Glioma.
    Shahla Shojaei, Amir Barzegar Behrooz, Kianoosh Naghibzadeh, João Basso, Javad Alizadeh, Tania Dehesh, Roham Sabei, Bhavya Bhushan, Mehdi Eshraghi, Simone C da Silva Rosa, Courtney Clark, Mateusz Marek Tomczyk, Laura Cole, Grant Hatch, Vernon W Dolinsky, Vinith Yathindranath, Donald Miller, Christopher Pasco, Sanjiv Dhingra, Abhay Srivastava, Amir Ravandi, Rui Vitorino, Stevan Pecic, Negar Azarpira, Mahmoud Aghaei, Saeid Ghavami.
      Glioma progression and resistance to temozolomide (TMZ) remain major clinical challenges. Here, we investigated whether dysregulated autophagy and cholesterol metabolism are coordinately remodeled during glioma progression and TMZ resistance. Tissue microarray analysis of astrocytoma and glioblastoma specimens revealed progressive autophagosome accumulation, reflected by increased LC3β puncta, coupled with impaired autophagic flux compared with adjacent normal brain tissue. These alterations intensified with tumor grade and were associated with upregulation of farnesyl diphosphate synthase (FDPS), linking malignant progression to cholesterol pathway remodeling. TMZ-resistant (R) glioblastoma cells exhibited epithelial-to-mesenchymal transition, mitotic quiescence, and mitochondrial remodeling consistent with a therapy-tolerant phenotype. Bioenergetic profiling demonstrated reduced respiratory reserve, diminished ATP-linked respiration, and elevated proton leak, indicating constrained metabolic flexibility. In parallel, impaired autophagy flux was associated with suppression of de novo cholesterol synthesis and transcriptional downregulation of SREBP-2 and LDL-R. Comprehensive lipidomic profiling revealed marked cholesterol metabolic reprogramming in R cells, characterized by accumulation of specific cholesteryl esters, including CE 22:5, CE 22:6, CE 22:4, and CE 20:4, despite reduced cholesterol biosynthesis. Pharmacologic inhibition of the mevalonate pathway with simvastatin significantly altered cholesteryl ester profiles but failed to restore autophagy flux or sensitize R cells to TMZ-induced apoptosis, even under combined TMZ-simvastatin treatment.
    Lay Abstract: As gliomas progress from astrocytoma to glioblastoma, autophagy becomes dysregulated and cholesterol metabolism is rewired. This coordinated remodeling supports tumor survival, metabolic plasticity, and resistance to temozolomide therapy.
    Highlights: Autophagy flux blockade intensifies during progression from astrocytoma to glioblastomaDysregulated autophagy is coupled to altered cholesterol metabolism in malignant gliomasTMZ-resistant glioblastoma cells undergo epithelial-to-mesenchymal transition and mitotic quiescenceResistant cells exhibit constrained bioenergetic capacity and mitochondrial remodelingImpaired autophagy suppresses de novo cholesterol synthesis and lipid recyclingLipidomics reveals accumulation of long-chain cholesteryl esters in TMZ-resistant cellsStatin-based cholesterol inhibition fails to resensitize glioblastoma cells to temozolomide.
    DOI:  https://doi.org/10.64898/2026.01.06.697885
  15. Chemistry. 2026 Jan 14. e03293
    Self-Calibrating Ratiometric DNA Aptamer Probe for Quantitative ATP Imaging in Living Cells.
    Yunsong Xu, Kunihiko Morihiro, Rui Cong, Samruddhi Maheshwar Patil, Idoia Wille, Akimitsu Okamoto.
      Accurate monitoring of adenosine triphosphate (ATP)-the universal energy currency of cells-is essential for elucidating cellular metabolism and disease progression. However, the high basal concentration of intracellular ATP (1-10 mM) and variable probe uptake during imaging have hampered the development of reliable fluorescent sensors. Although DNA aptamer-based probes provide excellent selectivity, conventional turn-on designs often lack internal calibration, and Förster resonance energy transfer-based ratiometric probes typically exhibit limited signal changes. We report a ratiometric DNA duplex sensor comprising a Cy5-labeled ATP aptamer and a thiazole orange (TO)-labeled exciton-controlled hybridization-sensitive fluorescent oligonucleotide (ECHO) probe. Upon ATP binding, the aptamer structure switches and releases the reporter strand, resulting in a pronounced decrease in TO fluorescence while the Cy5 signal remains constant. Rational insertion of a polythymidine spacer effectively suppressed undesired TO-to-Cy5 energy transfer, enabling a reliable ratiometric Cy5/ECHO readout. The sensor operates robustly across physiological ATP concentrations, exhibits high nucleotide selectivity and satisfactory serum stability, and shows minimal cytotoxicity. Live-cell flow cytometry and confocal imaging further confirmed that cancer cells displayed significantly higher Cy5/ECHO ratios than normal fibroblasts. This internally self-calibrating aptamer sensor thus provides a powerful platform for intracellular ATP imaging and cancer diagnostics.
    Keywords:  ATP; DNA; aptamers; fluorescence; ratiometric sensors
    DOI:  https://doi.org/10.1002/chem.202503293
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