bims-cagime 2025-08-03 papers

bims-cagime

Biomed News

on Cancer, aging and metabolism

Issue of 2025–08–03
37 papers selected by
Kıvanç Görgülü, Technical University of Munich

Senescence-associated lysosomal dysfunction impairs cystine deprivation-induced lipid peroxidation and ferroptosis.
Macropinocytosis maintains CAF subtype identity under metabolic stress in pancreatic cancer.
HIF-independent oxygen sensing via KDM6A regulates ferroptosis.
Tracking Intracellular Labile Iron with a Genetically Encoded Fluorescent Reporter System Based on Protein Stability.
Alterations in Lipid Saturation Trigger Remodeling of the Outer Mitochondrial Membrane.
Michael Patrick Sheetz, 1946-2025, a devotee of and major contributor to membrane and cytoskeletal biology.
Mitochondrial glutathione import enables breast cancer metastasis via integrated stress response signaling.
Proteomic analyses identify targets, pathways, and cellular consequences of oncogenic KRAS signaling.
ATXN3 regulates lysosome regeneration after damage by targeting K48-K63-branched ubiquitin chains.
Metabolic adaptations in cancer progression.
Therapeutic targeting of syndecan-1 axis overcomes acquired resistance to KRAS-targeted therapy in gastrointestinal cancers.
Dual-Anchoring Fluorescent Probes Enable Facile Recognition of Pyroptosis through Dynamic Tracking of Plasma Membrane.
De novo pyrimidine biosynthesis inhibition synergizes with BCL-XL targeting in pancreatic cancer.
Directed evolution of LaccID for cell surface proximity labeling and electron microscopy.
Apoptotic cells promote circulating tumor cell survival and metastasis.
The source of dietary fat influences anti-tumour immunity in obese mice.
Circulating tumor cells: Blood-based detection, molecular biology, and clinical applications.
Reconstitution of BNIP3/NIX-mitophagy initiation reveals hierarchical flexibility of the autophagy machinery.
NRAC controls CD36-mediated fatty acid uptake in adipocytes and lipid clearance in vivo.
Cathartocytosis: Jettisoning of cellular material during reprogramming of differentiated cells.
Cmem Builder: An Automated Tool for Curved Membrane Construction in Molecular Dynamics Simulations.
Cholesterol modulates membrane elasticity via unified biophysical laws.
Expression of IMPACT curtails metabolic plasticity and augments NK cell killing to abrogate metastatic growth.
Conservation and divergence of metabolic phenotypes between patient tumours and matched xenografts.
PRMT5 Identified as a Viable Target for Combination Therapy in Preclinical Models of Pancreatic Cancer.
The minimal chemotactic cell.
Multiomics analysis of GSTP1 knockdown pancreatic cancer cells reveals key regulators of redox and metabolic homeostasis.
Robust consensus nuclear and cell segmentation.
Small Molecule Promoters of Endogenous Lipid Droplet Accumulation Drive Lysophagy.
ACLY inhibition promotes tumour immunity and suppresses liver cancer.
iLOV3.0 Fluorescent Probes for Monitoring ROS Changes in Mammalian Cells.
A Fluorescence-Based Flippase Assay to Monitor Lipid Transport by Drs2-Cdc50.
Six-Minute Walk Test Is Superior to Grip Strength as a Marker of Functional Recovery During Cancer Cachexia Rehabilitation.
Adherent cells undergo rate-softening mediated by actomyosin kinetics.
A simple, accurate method for the measurement of lysosomal activity.
Optimal sensing through phase separation.
Targeted Degradation Technologies Utilizing Autophagy.

Nat Commun. 2025 Jul 29. 16(1): 6617

Senescence-associated lysosomal dysfunction impairs cystine deprivation-induced lipid peroxidation and ferroptosis.

Tze Mun Loo, Xiangyu Zhou, Yoko Tanaka, Sho Sugawara, Shota Yamauchi, Hiroko Kawasaki, Yuta Matsuoka, Yuki Sugiura, Shinya Sakuma, Yoko Yamanishi, Satoshi Yotsumoto, Kosuke Dodo, Yoshitaka Shirasaki, Takashi Kamatani, Akiko Takahashi.

Senescent cells, characterized by irreversible cell cycle arrest and inflammatory factor secretion, promote various age-related pathologies. Senescent cells exhibit resistance to ferroptosis, a form of iron-dependent cell death; however, the underlying mechanisms remain unclear. Here, we discovered that lysosomal acidity was crucial for lipid peroxidation and ferroptosis induction by cystine deprivation. In senescent cells, lysosomal alkalinization causes the aberrant retention of ferrous iron in lysosomes, resulting in resistance to ferroptosis. Treatment with the V-ATPase activator EN6 restored lysosomal acidity and ferroptosis sensitivity in senescent cells. A similar ferroptosis resistance mechanism involving lysosomal alkalinization was observed in pancreatic cancer cell lines. EN6 treatment prevented pancreatic cancer development in xenograft and Kras mutant mouse models. Our findings reveal a link between lysosomal dysfunction and the regulation of ferroptosis, suggesting a therapeutic strategy for the treatment of age-related diseases.

DOI: https://doi.org/10.1038/s41467-025-61894-9
Cancer Cell. 2025 Jul 15. pii: S1535-6108(25)00271-5. [Epub ahead of print]

Macropinocytosis maintains CAF subtype identity under metabolic stress in pancreatic cancer.

Yijuan Zhang, Li Ling, Rabi Murad, Swetha Maganti, Ambroise Manceau, Hannah A Hetrick, Madelaine Neff, Cheska Marie Galapate, Shea F Grenier, Florent Carrette, Karen Duong-Polk, Anindya Bagchi, David A Scott, Yoav Altman, Jennifer L Hope, Andrew M Lowy, Linda M Bradley, Cosimo Commisso.

  Pancreatic ductal adenocarcinoma (PDAC) tumors are glutamine deficient, and both tumor cells and cancer-associated fibroblasts (CAFs) rely on this amino acid to maintain fitness and induce macropinocytosis as an adaptive response. CAFs play a critical role in sculpting the tumor microenvironment, yet how adaptations to metabolic stress impact the stromal architecture remains elusive. In this study, we find that macropinocytosis sustains the myCAF phenotype under glutamine limitation by preventing inflammatory reprogramming. Our data demonstrate that metabolic stress induces an intrinsic inflammatory CAF (iCAF) program through MEK-ERK signaling. We find that blocking macropinocytosis in vivo promotes myCAF-to-iCAF transitions, remodeling the tumor stroma. Importantly, stromal remodeling driven by macropinocytosis inhibition-including iCAF enrichment, collagen reduction, immune cell infiltration, and vascular expansion-sensitizes PDAC tumors to immunotherapy and chemotherapy. Our findings reveal that inhibiting macropinocytosis promotes an inflammatory, less fibrotic tumor microenvironment that can be leveraged to improve therapeutic responses in PDAC.

Keywords:  CAF heterogeneity; chemotherapy; drug delivery; immunotherapy; macropinocytosis; metabolic stress; pancreatic cancer; plasticity; stromal architecture; tumor microenvironment

DOI:  https://doi.org/10.1016/j.ccell.2025.06.021
Mol Cell. 2025 Jul 15. pii: S1097-2765(25)00580-5. [Epub ahead of print]

HIF-independent oxygen sensing via KDM6A regulates ferroptosis.

Alexander M Minikes, Pei Liu, Hua Wang, Jiachen Hu, Hanan Alwaseem, Yueming Li, Xuejun Jiang.

  Ferroptosis, a metabolic cell death process driven by iron-dependent phospholipid peroxidation, is implicated in various pathologies, including cancer. While metabolic factors such as glucose, lipids, and multiple amino acids have all been demonstrated to modulate ferroptosis, the role of oxygen, another fundamental metabolic component, in ferroptosis is not fully understood. Here, we show that cells acclimated to a low oxygen environment develop marked resistance to ferroptosis, and this resistance is independent of canonical oxygen-sensing pathway mediated by prolyl hydroxylases (PHDs) and HIF transcription factors. Instead, hypoxia suppresses ferroptosis by inhibiting KDM6A, a tumor suppressor and oxygen-dependent histone demethylase, leading to reduced expression of its transcriptional targets, including lipid metabolic enzymes ACSL4 and ETNK1, thus rewiring cellular phospholipid profile to a ferroptosis-resistant state. Relevant to cancer, pharmacological inhibition of the oncogenic histone methyltransferase EZH2, which opposes KDM6A activity, restored ferroptosis sensitivity of xenograft bladder tumor tissues harboring KDM6A mutation.

Keywords:  ACSL4; ETNK1; KDM6A; KMT2D; bladder cancer; cancer therapy; ferroptosis; hypoxia; lipid metabolism; oxygen sensing

DOI:  https://doi.org/10.1016/j.molcel.2025.07.001
ACS Sens. 2025 Aug 01.

Tracking Intracellular Labile Iron with a Genetically Encoded Fluorescent Reporter System Based on Protein Stability.

Ali Akyol, Şeyma Çimen, Benjamin Gottschalk, Yusuf C Erdoğan, Anna Lischnig, Amy Barton Alston, Reinaldo Digigow, Beat Flühmann, Emrah Eroğlu, Wolfgang F Graier, Roland Malli.

  Here, we developed IronFist, a genetically encoded fluorescent reporter that enables dynamic tracking of labile ferrous ions (Fe2+) in live cells. IronFist is a bicistronic system combining the iron-sensitive hemerythrin-like (Hr) domain from the F-box and leucine-rich repeat protein 5 (FBXL5), fused to the bright fluorescent protein (FP) mNeonGreen, alongside mCherry as a reference FP signal. When labile iron levels are low, Hr-mNeonGreen undergoes ubiquitination and degradation, leading to a low green-to-red fluorescence ratio. Conversely, as cytosolic Fe2+ levels rise and Fe2+ ions bind to Hr, the green fluorescence is stabilized, increasing the IronFist ratio signal. Using IronFist for end point measurements, we observed that most cells maintain low basal labile iron levels. However, upon treatment with iron(II) sulfate or iron carbohydrate nanoparticles, we detected significant elevations in the cellular labile iron pool (LIP). Cells responded faster and more strongly to iron(II) sulfate, whereas responses to iron carbohydrate nanoparticles were slower and weaker. Time-lapse imaging further revealed substantial cell-to-cell heterogeneity in iron handling. We conclude that IronFist fills a critical methodological gap in assessing cellular iron homeostasis and related pathologies by enabling high-content tracking of iron dynamics at the single-cell level.

Keywords:  Hr domain; fluorescence microscopy; iron carbohydrate nanoparticles; iron reporter; iron-dependent degron; labile iron pool

DOI:  https://doi.org/10.1021/acssensors.5c01165
Mol Biol Cell. 2025 Jul 30. mbcE25010033

Alterations in Lipid Saturation Trigger Remodeling of the Outer Mitochondrial Membrane.

Sara Wong, Katherine R Bertram, Sai Sangeetha Balasubramaniam, Nidhi Raghuram, Thomas Knight, J Alan Maschek, James E Cox, Adam L Hughes.

Lipid saturation is a key determinant of membrane function and organelle health, with changes in saturation triggering adaptive quality control mechanisms to maintain membrane integrity. Among cellular membranes, the mitochondrial outer membrane (OMM) is an important interface for many cellular functions, but how lipid saturation impacts OMM function remains unclear. Here, we show that increased intracellular unsaturated fatty acids (UFAs) remodel the OMM by promoting the formation of multilamellar mitochondrial-derived compartments (MDCs), which sequester proteins and lipids from the OMM. These effects depend on the incorporation of UFAs into membrane phospholipids, suggesting that changes in membrane bilayer composition mediate this process. Furthermore, elevated UFAs impair the assembly of the OMM protein translocase (TOM) complex, with unassembled TOM components captured into MDCs. Collectively, these findings suggest that alterations in phospholipid saturation may destabilize OMM protein complexes and trigger an adaptive response to sequester excess membrane proteins through MDC formation.

DOI: https://doi.org/10.1091/mbc.E25-01-0033
Mol Biol Cell. 2025 Aug 01. 36(8): fe1

Michael Patrick Sheetz, 1946-2025, a devotee of and major contributor to membrane and cytoskeletal biology.

Linda J Kenney, Ronald D Vale, James A Spudich.

Michael P. Sheetz (1946-2025) advanced the field of mechanobiology through his creative experiments, new methodologies, and keen insights. His research touched many fields of cell biology, including membrane biophysics, motor proteins, the cytoskeleton, cell migration, and cellular senescence. In addition to his research, Sheetz was a leader who built vibrant academic departments and institutes and advanced the careers of many trainees.

DOI: https://doi.org/10.1091/mbc.E25-05-0208
Cancer Discov. 2025 Jul 31.

Mitochondrial glutathione import enables breast cancer metastasis via integrated stress response signaling.

Hsi-Wen Yeh, Nicole Lauren DelGaudio, Beste Uygur, Alon Millet, Artem Khan, Gokhan Unlu, Michael Xiao, Rebecca C Timson, Caifan Li, Kerem Ozcan, Karl W Smith, Luiza Martins Nascentes Melo, Gabriele Allies, Olca Basturk, Albert Sickmann, Erol C Bayraktar, Richard Possemato, Alpaslan Tasdogan, Kivanc Birsoy.

Cancer cells require substantial metabolic adaptations to metastasize to distant organs, but the metabolites essential for successful colonization remain poorly defined. Here, we used a mitochondrial metabolomics approach to compare primary and metastatic breast cancer cells. This analysis revealed accumulation of mitochondrial glutathione (GSH) during lung metastasis, driven by elevated expression of SLC25A39, a mitochondrial GSH transporter. Loss of SLC25A39 impairs metastatic colonization in genetic screens, cell line models, and patient-derived xenografts, without affecting primary tumor growth. Mitochondrial GSH import is specifically required during early colonization and functions independently of its canonical antioxidant role. CRISPR activation screens identified ATF4, a stress-induced transcription factor, as a bypass mechanism that restores metastatic potential in SLC25A39-deficient cells. Mechanistically, SLC25A39 is required for optimal ATF4 activation during metastasis and under hypoxia, linking mitochondrial GSH availability to integrated stress response signaling. These findings identify mitochondrial GSH as a necessary and limiting metabolite for metastatic progression.

DOI: https://doi.org/10.1158/2159-8290.CD-24-1556
Sci Signal. 2025 Jul 29. 18(897): eadt6552

Proteomic analyses identify targets, pathways, and cellular consequences of oncogenic KRAS signaling.

Nicole Kabella, Florian P Bayer, Konstantinos Stamatiou, Miriam Abele, Amirhossein Sakhteman, Yun-Chien Chang, Vinona Wagner, Antje Gabriel, Johannes Krumm, Maria Reinecke, Melanie Holzner, Michael Aigner, Matthew The, Hannes Hahne, Florian Bassermann, Christina Ludwig, Paola Vagnarelli, Bernhard Kuster.

Mutations that activate the small GTPase KRAS are a frequent genetic alteration in cancer, and drug discovery efforts have led to inhibitors that block KRAS activity. We sought to better understand oncogenic KRAS signaling and the cytostatic effects of drugs that target this system. We performed proteomic analyses to investigate changes in protein abundance and posttranslational modifications in inhibitor-treated human KRAS-mutant pancreatic (KRAS G12C and G12D) and lung cancer (KRAS G12C) cells. The inhibitors used target these mutant forms of KRAS, the downstream effectors MEK and ERK, and the upstream regulators SHP2 and SOS1. Comparisons of phosphoproteomes between cell lines revealed a core KRAS signaling signature and cell line-specific signaling networks. In all cell lines, phosphoproteomes were dominated by different degrees of autonomous, oncogenic KRAS activity. Comparison of phosphoproteomes after short and long drug exposures revealed the temporal dynamics of KRAS-MEK-ERK axis inhibition that resulted in cell cycle exit. This transition to a quiescent state occurred in the absence of substantial proteome remodeling but included broad changes in protein phosphorylation and ubiquitylation. The collective data reveal insights into oncogenic KRAS signaling, place many additional proteins into this functional context, and implicate cell cycle exit as a mechanism by which cells evade death upon KRAS signaling inhibition.

DOI: https://doi.org/10.1126/scisignal.adt6552
EMBO J. 2025 Jul 29.

ATXN3 regulates lysosome regeneration after damage by targeting K48-K63-branched ubiquitin chains.

Maike Reinders, Bojana Kravic, Pinki Gahlot, Sandra Koska, Johannes van den Boom, Nina Schulze, Sophie Levantovsky, Stefan Kleine, Markus Kaiser, Yogesh Kulathu, Christian Behrends, Hemmo Meyer.

  The cellular response to lysosomal damage involves fine-tuned mechanisms of membrane repair, lysosome regeneration and lysophagy, but how these different processes are coordinated is unclear. Here we show in human cells that the deubiquitinating enzyme ATXN3 helps restore integrity of the lysosomal system after damage by targeting K48-K63-branched ubiquitin chains on regenerating lysosomes. We find that ATXN3 is required for lysophagic flux after lysosomal damage but is not involved in the initial phagophore formation on terminally damaged lysosomes. Instead, ATXN3 is recruited to a distinct subset of lysosomes that are decorated with phosphatidylinositol-(4,5)-bisphosphate and that are not yet fully reacidified. There, ATXN3, along with its partner VCP/p97, targets and turns over K48-K63-branched ubiquitin conjugates. ATXN3 thus facilitates degradation of a fraction of LAMP2 via microautophagy to regenerate the lysosomal membrane and to thereby reestablish degradative capacity needed also for completion of lysophagy. Our findings identify a key role of ATXN3 in restoring lysosomal function after lysosomal membrane damage and uncover K48-K63-branched ubiquitin chain-regulated regeneration as a critical element of the lysosomal damage stress response.

Keywords:  Autophagy; Lysosome; Membrane; Stress Response; Ubiquitin

DOI:  https://doi.org/10.1038/s44318-025-00517-x
Physiol Rev. 2025 Jul 28.

Metabolic adaptations in cancer progression.

Yiming Peng-Winkler, Sarah-Maria Fendt.

  Cancer cells reprogram their metabolism as they travel to distant organs to establish metastases, the leading cause of cancer-related mortality. While the metabolic state of primary tumors has been extensively studied, the specific metabolic alterations associated with metastases have only recently garnered significant attention. The metabolic dependencies that arise during the metastatic cascade, along with the adaptive metabolic shifts required for growth in a new microenvironment, present promising therapeutic targets. In this review, we provide an overview of cancer metabolism, followed by a detailed exploration of the metabolic changes occurring at each stage of metastasis and within common organs of metastatic spread. Lastly, we examine the potential and challenges of targeting metabolic pathways in cancer therapy.

Keywords:  Cancer; Metabolism; Metabolism-based therapy; Metastasis; Organ microenvironment

DOI:  https://doi.org/10.1152/physrev.00037.2024
Cell Rep Med. 2025 Jul 22. pii: S2666-3791(25)00326-X. [Epub ahead of print] 102253

Therapeutic targeting of syndecan-1 axis overcomes acquired resistance to KRAS-targeted therapy in gastrointestinal cancers.

Madelaine S Theardy, Mitsunobu Takeda, Alexey Sorokin, Shuaitong Chen, Zecheng Yang, Xiaofei Wang, Preeti Kanikarla, Oluwadara Coker, Phuoc Nguyen, Yongkun Wei, Jun Yao, Liang Yan, Yanqing Jin, Yiming Cai, Masakatsu Paku, Ziheng Chen, Kara Z Li, Francesca Citron, Hideo Tomihara, Sisi Gao, Angela K Deem, Jun Zhao, Huamin Wang, Samir Hanash, Ronald A DePinho, Anirban Maitra, Giulio F Draetta, Haoqiang Ying, Scott Kopetz, Wantong Yao.

  The therapeutic benefit of recently developed mutant KRAS (KRAS∗) inhibitors remains limited by the rapid onset of resistance. Here, we aim to delineate mechanisms underlying acquired resistance and identify actionable targets for overcoming this clinical challenge. Previously, we identified syndecan-1 (SDC1) as a key effector for pancreatic cancer progression whose surface expression is driven by KRAS∗. By leveraging both pancreatic and colorectal cancer models, we show that surface SDC1 expression initially diminishes upon KRAS∗ inhibition but recovers in tumor cells that bypass KRAS∗ dependency. Mechanistically, we reveal that YAP1 activation drives the recovery of SDC1 surface localization to enhance macropinocytosis-mediated nutrient salvaging and activation of multiple receptor tyrosine kinases for tumor maintenance, promoting resistance to KRAS∗-targeted therapy. Overall, our study provides a strong rationale for targeting the YAP-SDC1 axis to overcome resistance to KRAS∗ inhibition, thereby revealing promising therapeutic opportunities for improving the clinical outcome of patients with KRAS∗-mutated cancers.

Keywords:  KRAS inhibitor; colorectal cancer; macropinocytosis; pancreatic cancer; syndecan; therapy resistance

DOI:  https://doi.org/10.1016/j.xcrm.2025.102253
Anal Chem. 2025 Aug 01.

Dual-Anchoring Fluorescent Probes Enable Facile Recognition of Pyroptosis through Dynamic Tracking of Plasma Membrane.

Junjun Pan, Aohui Peng, Shuyi Jiang, Xin Peng, Chuixi Kong, Jin Zhou, Hui Feng, Zhigang Jin, Zhaosheng Qian.

Pyroptosis is a pro-inflammatory form of programmed cell death mediated by the gasdermin protein family, often leading to potentially fatal systemic inflammation and associated with severe diseases like cancer. However, there remains a lack of simple yet effective tools for identifying and monitoring pyroptosis in real time. Herein, we report dual-anchoring fluorescent probes (SPD-R) designed for long-term tracking of the plasma membrane and demonstrate their utility in the identification and real-time monitoring of pyroptosis over hours. The identification of pyroptosis heavily relies on the dynamic changes of the plasma membrane during its distinct stages, making the key feature of these probes their ability to maintain long-term anchoring to the plasma membrane. In the designed SPD-R probes, three positive charges and a rigid steric hindrance unit were incorporated to enhance probe impermeability, while long alkyl chains at both ends were introduced to promote interactions with the lipid bilayer of the plasma membrane. These probes exhibit strong labeling capacity across various cell types and achieve up to several hours of stable anchoring on the plasma membrane, enabling real-time tracking of the entire pyroptosis process induced by LPS. The SPD-R probes successfully label and track significant increases in cell volume and notable bubbling during typical pyroptosis, providing a straightforward approach for pyroptosis recognition. Furthermore, this method was extended to the fast screening of Chinese medicines that induce cancer cell pyroptosis, with Hedyotis diffusa serving as an example.

DOI: https://doi.org/10.1021/acs.analchem.5c02560
Nat Commun. 2025 Jul 30. 16(1): 6987

De novo pyrimidine biosynthesis inhibition synergizes with BCL-XL targeting in pancreatic cancer.

Huan Zhang, Naiara Santana-Codina, Qijia Yu, Clara Poupault, Claudia Campos, Xingping Qin, Nicole Sindoni, Marina Ciscar, Aparna Padhye, Miljan Kuljanin, Junning Wang, Matthew J Dorman, Peter Bross, Andrew J Aguirre, Stephanie K Dougan, Kristopher A Sarosiek, Joseph D Mancias.

Oncogenic KRAS induces metabolic rewiring in pancreatic ductal adenocarcinoma (PDAC) characterized, in part, by dependency on de novo pyrimidine biosynthesis. Pharmacologic inhibition of dihydroorotate dehydrogenase (DHODH), an enzyme in the de novo pyrimidine synthesis pathway, delays pancreatic tumor growth; however, limited monotherapy efficacy suggests that compensatory pathways may drive resistance. Here, we use an integrated metabolomic, proteomic and in vitro and in vivo DHODH inhibitor-anchored genetic screening approach to identify compensatory pathways to DHODH inhibition (DHODHi) and targets for combination therapy strategies. We demonstrate that DHODHi alters the apoptotic regulatory proteome thereby enhancing sensitivity to inhibitors of the anti-apoptotic BCL2L1 (BCL-XL) protein. Co-targeting DHODH and BCL-XL synergistically induces apoptosis in PDAC cells and patient-derived organoids. The combination of DHODH inhibition with Brequinar and BCL-XL degradation by DT2216, a proteolysis targeting chimera (PROTAC), significantly inhibits PDAC tumor growth. These data define mechanisms of adaptation to DHODHi and support combination therapy targeting BCL-XL in PDAC.

DOI: https://doi.org/10.1038/s41467-025-61242-x
Nat Chem Biol. 2025 Aug 01.

Directed evolution of LaccID for cell surface proximity labeling and electron microscopy.

Song-Yi Lee, Heegwang Roh, David Gonzalez-Perez, Mason R Mackey, Daniel Hoces, Colleen N McLaughlin, Chang Lin, Stephen R Adams, Khanh Nguyen, Keun-Young Kim, David J Luginbuhl, Liqun Luo, Namrata D Udeshi, Steven A Carr, Rogelio A Hernández-López, Mark H Ellisman, Miguel Alcalde, Alice Y Ting.

Enzymes that oxidize aromatic substrates have been harnessed for cell-based technologies including proximity labeling and electron microscopy; however, they are associated with drawbacks such as the need for toxic H2O2. Here, we explore multicopper oxidases (laccases) as a new enzyme class for proximity labeling and electron microscopy in mammalian cells. LaccID was generated through 11 rounds of directed evolution from an ancestral fungal laccase and catalyzes one-electron oxidation of diverse aromatic substrates using O2 instead of toxic H2O2. Surprisingly, we found that LaccID is selectively active at the surface plasma membrane of both living and fixed cells. We use LaccID proximity labeling and mass spectrometry to map the changing surface proteome of T cells that engage with tumor cells through antigen-specific T cell receptors. In addition, we use LaccID as a genetically encodable tag for EM visualization of cell surface features in mammalian cell culture and in the fly brain. Our study paves the way for future cell-based applications of LaccID.

DOI: https://doi.org/10.1038/s41589-025-01973-6
Commun Biol. 2025 Jul 29. 8(1): 1121

Apoptotic cells promote circulating tumor cell survival and metastasis.

Cassidy E Hagan, Valerie M Sheehan, Charles M Rezanka, Meijie A Li, Laura Martínez-Escardó, Kirsteen J Campbell, Annelise G Snyder, Mark Headley, Andrew Oberst.

During tumor progression and especially following cytotoxic therapy, cell death of both tumor and stromal cells is widespread. Despite clinical observations that high levels of apoptotic cells correlate with poorer patient outcomes, the physiological effects of dying cells on tumor progression remain incompletely understood. Here, we report that circulating apoptotic cells robustly enhance tumor cell metastasis to the lungs. Using intravenous metastasis models, we observed that the presence of apoptotic cells, but not cells dying by other mechanisms, supports circulating tumor cell (CTC) survival following arrest in the lung vasculature. Apoptotic cells promote CTC survival by recruiting platelets to the forming metastatic niche. Apoptotic cells externalize the phospholipid phosphatidylserine to the outer leaflet of the plasma membrane, which we found increased the activity of the coagulation initiator Tissue Factor, thereby triggering the formation of platelet clots that protect proximal CTCs. Inhibiting the ability of apoptotic cells to induce coagulation by knocking out Tissue Factor, blocking phosphatidylserine, or administering the anticoagulant heparin abrogated the pro-metastatic effect of apoptotic cells. This work demonstrates a previously unappreciated role for apoptotic cells in facilitating metastasis by establishing CTC-supportive emboli, and suggests points of intervention that may reduce the pro-metastatic effect of apoptotic cells.

DOI: https://doi.org/10.1038/s42003-025-08541-7
Nat Metab. 2025 Jul 25.

The source of dietary fat influences anti-tumour immunity in obese mice.

Britta Kunkemoeller, Hannah Prendeville, Claire McIntyre, Ayantu Temesgen, Rόisín M Loftus, Conghui Yao, Lydia Dyck, Linda V Sinclair, Christina Rollings, Aaron Douglas, Gerard Pernes, Kathleen A J Mitchelson, Cathal Harmon, Mathilde Raverdeau, Ross Ward, Harry Kane, Jaclyn Kline, Katie L O'Brien, Martin Brennan, Frances Smith, Brenneth Stevens, Helen M Roche, Ed C Lavelle, David K Finlay, Doreen A Cantrell, Edward T Chouchani, Susan Kaech, Evanna L Mills, Marcia Haigis, Lydia Lynch.

Obesity increases the risk of many cancers and impairs the anti-tumour immune response. However, little is known about whether the source or composition of dietary fat affects tumour growth or anti-tumour immunity in obesity. Here, we show that high-fat diets (HFDs) derived from lard, beef tallow or butter accelerate tumour growth in a syngeneic model of melanoma, but HFDs based on coconut oil, palm oil or olive oil do not, despite equivalent obesity. Using butter-based and palm oil-based HFDs as examples, we find that these dietary fat sources differentially regulate natural killer and CD8 T cell infiltration and function within the tumour microenvironment, governed by distinct effects on the plasma metabolome and intracellular metabolism. We identify diet-related lipid intermediates, namely long-chain acylcarnitine species, as immunosuppressive metabolites enriched in mice fed butter compared to palm oil HFD. Together, these results highlight the significance of diet in maintaining a healthy immune system and suggest that modifying dietary fat may improve cancer outcomes in obesity.

DOI: https://doi.org/10.1038/s42255-025-01330-w
Cancer Cell. 2025 Jul 23. pii: S1535-6108(25)00314-9. [Epub ahead of print]

Circulating tumor cells: Blood-based detection, molecular biology, and clinical applications.

Charles S Dai, Avanish Mishra, Jon Edd, Mehmet Toner, Shyamala Maheswaran, Daniel A Haber.

  Circulating tumor cells (CTCs) are cancer cells, shed from primary tumors or metastases into the bloodstream. The first non-invasive "liquid biopsy" for cancer monitoring, CTCs have been largely surpassed by circulating tumor DNA (ctDNA) for clinical applications, given the ease of DNA sequencing without specialized cell isolation methods. However, emerging rare cell capture technologies that can process larger blood volumes and enable advanced single-cell analyses may enhance the range and potential of CTC-based biomarkers. CTCs are increasingly valuable for assessing tumor heterogeneity, guiding protein biomarker-driven cancer immune therapies, and assessing heterogeneous drug resistance, as well as for detecting minimal disease. CTCs, thus, remain central to understanding cancer dissemination and are poised to offer complementary diagnostic roles in the application of minimally invasive liquid biopsies for cancer. Here, we review recent advances in the study of these rare circulating cancer cells and discuss current limitations and future directions.

Keywords:  cancer metastasis; circulating tumor cells; ex vivo cancer models; liquid biopsy biomarkers; minimal residual disease; molecular diagnostics; rare cell technologies; tumor heterogeneity

DOI:  https://doi.org/10.1016/j.ccell.2025.07.008
Nat Cell Biol. 2025 Jul 25.

Reconstitution of BNIP3/NIX-mitophagy initiation reveals hierarchical flexibility of the autophagy machinery.

Elias Adriaenssens, Stefan Schaar, Annan S I Cook, Jan F M Stuke, Justyna Sawa-Makarska, Thanh Ngoc Nguyen, Xuefeng Ren, Martina Schuschnig, Julia Romanov, Grace Khuu, Louise Uoselis, Michael Lazarou, Gerhard Hummer, James H Hurley, Sascha Martens.

Selective autophagy is a lysosomal degradation pathway that is critical for maintaining cellular homeostasis by disposing of harmful cellular material. Although the mechanisms by which soluble cargo receptors recruit the autophagy machinery are becoming increasingly clear, the principles governing how organelle-localized transmembrane cargo receptors initiate selective autophagy remain poorly understood. Here we demonstrate that the human transmembrane cargo receptors can initiate autophagosome biogenesis not only by recruiting the upstream FIP200/ULK1 complex but also via a WIPI-ATG13 complex. This latter pathway is employed by the BNIP3/NIX receptors to trigger mitophagy. Additionally, other transmembrane mitophagy receptors, including FUNDC1 and BCL2L13, exclusively use the FIP200/ULK1 complex, whereas FKBP8 and the ER-phagy receptor TEX264 are capable of utilizing both pathways to initiate autophagy. Our study defines the molecular rules for initiation by transmembrane cargo receptors, revealing remarkable flexibility in the assembly and activation of the autophagy machinery, with important implications for therapeutic interventions.

DOI: https://doi.org/10.1038/s41556-025-01712-y
EMBO J. 2025 Aug 01.

NRAC controls CD36-mediated fatty acid uptake in adipocytes and lipid clearance in vivo.

Inderjeet Singh, Yasuhiro Onogi, Filipe Menezes, Dina Khasanova, Lingru Kang, Chenxi Wang, Julio Ruiz-Trave, Sapna Sharma, Ahmed Khalil, Valentin K Reichenbach, Yingzi Shi, Andrew Flatley, Xiaocheng Yan, Andreas Israel, Nathalia R V Dragano, Juan Antonio Aguilar-Pimentel, Anne Hoffmann, Adhideb Ghosh, Falko Noé, Christian Wolfrum, Sebastian Cucuruz, Ann-Christine König, Ingo Burtscher, Stefanie M Hauck, Heiko Lickert, Susanna M Hofmann, Regina Feederle, Sonja C Schriever, Rene Hernandez-Bautista, Gencer Sancar, Alberto Cebrian-Serrano, Igor Tetko, Helmut Fuchs, Valérie Gailus-Durner, Matthias Blüher, Martin Hrabě de Angelis, Siegfried Ussar.

  Adipose tissue is a central organiser of systemic lipid homeostasis and a pharmacological target in obesity, orchestrating cellular responses to environmental cues. Nutritionally regulated adipose and cardiac enriched protein (NRAC) is a small adipocyte-specific transmembrane protein with unknown function. Here, we show that Nrac directly interacts with scavenger receptor CD36 via its first transmembrane domain. Forming a complex with CD36 and caveolin-1 under low extracellular fatty acid (FA) concentrations, NRAC modulates CD36-dependent fatty acid uptake in adipocytes. Upon increase in extracellular FA levels, NRAC is ubiquitinated and internalised, leading to CD36's dissociation from caveolin-1 and clathrin-mediated endocytosis. This results in increased fatty acid uptake into fat cells, adipocyte hypertrophy, increased fat mass and elevated lipid clearance from the blood in chow-diet-fed mice. Finally, human NRAC expression and the intronic SNP rs12878589 are associated with body fat distribution and obesity. Together, these findings reveal a novel regulatory mechanism by which adipocytes sense and respond to extracellular fatty acid availability to fine-tune lipid uptake and storage at cellular and organismal level.

Keywords:  Adipose Tissue; CD36; Clathrin-mediated Endocytosis; Fatty Acid Uptake; Hypertrophy

DOI:  https://doi.org/10.1038/s44318-025-00520-2
Cell Rep. 2025 Jul 29. pii: S2211-1247(25)00841-1. [Epub ahead of print]44(8): 116070

Cathartocytosis: Jettisoning of cellular material during reprogramming of differentiated cells.

Jeffrey W Brown, Xiaobo Lin, Gabriel Anthony Nicolazzi, Xuemei Liu, Thanh Nguyen, Megan D Radyk, Joseph Burclaff, Jason C Mills.

  Injury causes differentiated cells to undergo massive reprogramming to become proliferative and repair tissue via paligenosis. Gastric chief cells use paligenosis to reprogram into progenitor-like spasmolytic-polypeptide-expressing metaplasia (SPEM) cells. Stage 1 of paligenosis is the downscaling of mature cell architecture via a process involving lysosomes. Here, we notice that sulfated glycoproteins are not only digested during paligenosis but also excreted into the gland. Various genetic and pharmacological approaches show that endoplasmic reticulum membranes and secretory granule cargo are also excreted and that the process proceeds in parallel with but is mechanistically independent of autophagy. Three-dimensional light and electron microscopy demonstrated that excretion occurs via unique, complex, multi-chambered invaginations of the apical plasma membrane. As this lysosome-independent cell cleansing process does not seem to have been priorly described, we termed it "cathartocytosis." Cathartocytosis allows a cell to rapidly eject excess material without waiting for autophagic and lysosomal digestion, providing for efficient cellular downscaling.

Keywords:  CP: Cell biology; Das-1; EPG5; Rab7; secretion; sulfated mucins

DOI:  https://doi.org/10.1016/j.celrep.2025.116070
J Chem Theory Comput. 2025 Jul 28.

Cmem Builder: An Automated Tool for Curved Membrane Construction in Molecular Dynamics Simulations.

Jiabin Guo, Kin Lei, Jixing Liu, Henry Hy Tong, Yun Lyna Luo, Wei Han, Shu Li.

Membrane curvature is a fundamental property of biological membranes, driving essential processes such as endocytosis, vesicle formation, and mechanotransduction. Molecular dynamics (MD) simulations have become a powerful approach for studying curved membrane systems, providing atomistic insights into curvature-driven phenomena and protein-membrane interactions. However, online platforms like CHARMM-GUI and CGMD focus on constructing flat bilayers or vesicles and lack support for generating curved membranes with defined geometries. Local tools, while more flexible, often do not incorporate protein-specific curvature features, such as those from the Orientations of Proteins in Membranes (OPM) database, which are critical for accurately modeling protein-lipid interactions in curved environments. To address these limitations, we developed Cmem Builder, a novel and user-friendly web server for automating the generation of curved lipid membranes and membrane-protein complexes for coarse-grained (CG) MD simulations using the MARTINI force field. Cmem Builder specializes in generating Z-axis symmetric curved membrane shapes, supports curvature profiles derived from OPM database or custom geometries, allows extensive control over lipid composition, and ensures lipid placement through geometric sampling. The tool has been successfully applied to classical curved membrane systems, including Piezo1 and BAR proteins, as well as plasma membranes with asymmetric lipid compositions, demonstrating its accuracy and efficiency. In total, Cmem Builder provides a robust and accessible platform for exploring the complex dynamics of curved membrane systems. The tool is freely available at https://cmembuilder.com.

DOI: https://doi.org/10.1021/acs.jctc.5c00467
Nat Commun. 2025 Jul 31. 16(1): 7024

Cholesterol modulates membrane elasticity via unified biophysical laws.

Teshani Kumarage, Sudipta Gupta, Nicholas B Morris, Fathima T Doole, Haden L Scott, Laura-Roxana Stingaciu, Sai Venkatesh Pingali, John Katsaras, George Khelashvili, Milka Doktorova, Michael F Brown, Rana Ashkar.

Cholesterol and lipid unsaturation underlie a balance of opposing forces that features prominently in adaptive cell responses to diet and environmental cues. These competing factors have resulted in contradictory observations of membrane elasticity across different measurement scales, requiring chemical specificity to explain incompatible structural and elastic effects. Here, we demonstrate that - unlike macroscopic observations - lipid membranes exhibit a unified elastic behavior in the mesoscopic regime between molecular and macroscopic dimensions. Using nuclear spin techniques and computational analysis, we find that mesoscopic bending moduli follow a universal dependence on the lipid packing density regardless of cholesterol content, lipid unsaturation, or temperature. Our observations reveal that compositional complexity can be explained by simple biophysical laws that directly map membrane elasticity to molecular packing associated with biological function, curvature transformations, and protein interactions. The obtained scaling laws closely align with theoretical predictions based on conformational chain entropy and elastic stress fields. These findings provide unique insights into the membrane design rules optimized by nature and unlock predictive capabilities for guiding the functional performance of lipid-based materials in synthetic biology and real-world applications.

DOI: https://doi.org/10.1038/s41467-025-62106-0
Cancer Discov. 2025 Jul 31.

Expression of IMPACT curtails metabolic plasticity and augments NK cell killing to abrogate metastatic growth.

Surajit Sinha, Abir Kumar Panda, Rodrigo Xavier das Neves, Zeribe C Nwosu, Ke Xu, Elke van Beek, Priyanka P Desai, Sivasish Sindiri, Sruthi Chempati, Kirsten Remmert, Billel Gasmi, Linda Bojmar, Constantinos Zambirinis, Alexander J Rossi, Reed I Ayabe, Michael M Wach, James D McDonald, Samantha M Ruff, Emily A Verbus, Areeba Saif, Alyssa V Eade, Carolina M Larrain, Lindsay R Friedman, Shreya Gupta, Alok Ranjan, Martha E Teke, Tahsin M Khan, Tracey Pu, Amber Leila Sarvestani, Carrie E Ryan, Jacob T Lambdin, Kenneth Luberice, Stephanie N Gregory, Stephanie C Lux, Hanna Hong, Allen J Luna, Imani A Alexander, Sarfraz R Akmal, Shahyan U Rehman, Ashley Rainey, Todd D Prickett, Vishal N Koparde, Samantha Sevilla, Skyler A Kuhn, King Chan, Zhonghe Sun, Nina Bubunenko, Eileen Li, Cathleen Hannah, Geneti Gaga, Thorkell Andresson, Margaret C Cam, Xiaolin Wu, Lisa M Jenkins, Andrew M Blakely, Jeremy L Davis, Giorgio Trinchieri, Pankaj K Singh, James C Yang, Marina Pasca di Magliano, Costas A Lyssiotis, Michael B Yaffe, Ethan M Shevach, Jonathan M Hernandez.

Given the propensity of aggressive epithelial tumors to form hepatic metastases, we performed an in vivo cDNA screen using the mouse liver and KRASG12D/TP53R273H pancreatic cells to identify the RNA binding protein GCN1 as integral component of hepatic outgrowth. RNAi experiments reveal that GCN1 triggers the ISR to activate serine, folate, and methionine biosynthetic pathways together with amino acid transporters, which act in concert to facilitate acquisition of metabolites and to restore redox homeostasis. Alongside activation of the ISR, we found that GCN1 also functions in the nucleus where it interacts with HNRNPK to suppress the expression of MHC-I molecules, and natural killer (NK) ligands. Intriguingly, we identified IMPACT as an endogenous competitive inhibitor of GCN1 that blocks both ISR-dependent metabolic control and disrupts HNRNPK interaction. In doing so, IMPACT enhances tumor immunogenicity to unleash NK cell killing, in addition to sensitizing metastatic tumor cells to immune checkpoint blockade (ICB).

DOI: https://doi.org/10.1158/2159-8290.CD-24-1055
Nat Metab. 2025 Jul 29.

Conservation and divergence of metabolic phenotypes between patient tumours and matched xenografts.

Aparna D Rao, Ling Cai, Marelize Snyman, Rachel E Walsdorf, Xiangyi Li, Sophia N Wix, Gabrielle Gard, Ariel B Brown, Juliana Kim, Joao Santos Patricio, Sarah Muh, Misty Martin Sandoval, Lauren G Zacharias, Kristen A Heimdal, Wen Gu, Jade Homsi, Brittny Tillman, Rohit Sharma, Travis W Vandergriff, Ashley Solmonson, Brandon Faubert, Thomas P Mathews, Sean J Morrison, Ralph J DeBerardinis, Jennifer G Gill.

Patient-derived xenografts (PDXs) are frequently used as preclinical models, but their recapitulation of tumour metabolism in patients has not been closely examined. We developed a parallel workflow to analyse [U-13C]glucose tracing and metabolomics data from patient melanomas and matched PDXs. Melanomas from patients have substantial TCA cycle labelling, similar to levels in human brain tumours. Although levels of TCA cycle labelling in PDXs were similar to those in the original patient tumours, PDXs had higher labelling in glycolytic metabolites. Through metabolomics, we observed consistent alterations of 100 metabolites among PDXs and patient tumours that reflected species-specific differences in diet, host physiology and microbiota. Despite these differences, most of nearly 200 PDXs retained a 'metabolic fingerprint' largely durable over six passages and often traceable back to the patient tumour of origin. This study identifies both high- and low-fidelity metabolites in the PDX model system, providing a resource for cancer metabolism researchers.

DOI: https://doi.org/10.1038/s42255-025-01338-2
Biomolecules. 2025 Jun 30. pii: 948. [Epub ahead of print]15(7):

PRMT5 Identified as a Viable Target for Combination Therapy in Preclinical Models of Pancreatic Cancer.

Xiaolong Wei, William J Kane, Sara J Adair, Sarbajeet Nagdas, Denis Liu, Todd W Bauer.

  Pancreatic cancer is the third leading cause of cancer-related death in the US. First-line chemotherapy regimens for pancreatic ductal adenocarcinoma (PDAC) include FOLFIRINOX or gemcitabine (Gem) with or without paclitaxel (Ptx); however, 5-year survival with these regimens remains poor. Previous work has demonstrated protein arginine methyltransferase 5 (PRMT5) to be a promising therapeutic target in combination with Gem for the treatment of PDAC; however, these findings have yet to be confirmed in relevant preclinical models of PDAC. To test the possibility of PRMT5 as a viable therapeutic target, clinically relevant orthotopic and metastatic patient-derived xenograft (PDX) mouse models of PDAC growth were utilized to evaluate the effect of PRMT5 knockout (KO) or pharmacologic inhibition on treatment with Gem alone or Gem with Ptx. Primary endpoints included tumor volume, tumor weight, or metastatic tumor burden as appropriate. The results showed that Gem-treated PRMT5 KO tumors exhibited decreased growth and were smaller in size compared to Gem-treated wild-type (WT) tumors. Similarly, the Gem-treated PRMT5 KO metastatic burden was lower than the Gem-treated WT metastatic burden. The addition of a PRMT5 pharmacologic inhibitor to Gem and Ptx therapy resulted in a lower final tumor weight and fewer metastatic tumors. The depletion of PRMT5 results in increased DNA damage in response to Gem and Ptx treatment. Thus, PRMT5 genetic depletion or inhibition in combination with Gem-based therapy improved the response in primary and metastatic PDAC in clinically relevant mouse models, suggesting that PRMT5 is a viable therapeutic target for combination therapy in PDAC.

Keywords:  DNA damage; PRMT5; combination therapy; gemcitabine; orthotopic PDX mouse model; paclitaxel; pancreatic cancer; therapeutic target

DOI:  https://doi.org/10.3390/biom15070948
Sci Adv. 2025 Jul 25. 11(30): eadx9364

The minimal chemotactic cell.

Bárbara Borges-Fernandes, Azzurra Apriceno, Andres Arango-Restrepo, Safa Almadhi, Subhadip Ghosh, Joe Forth, Jorge Pedro López-Alonso, Iban Ubarretxena-Belandia, José Miguel Rubi, Lorena Ruiz-Pérez, Ian Williams, Giuseppe Battaglia.

The movement of cells and microorganisms in response to chemical gradients, chemotaxis, is fundamental to the evolution of myriad biological processes. In this work, we demonstrate that even the simplest cell-like structures are capable of chemotactic navigation. By encapsulating enzymes within lipid vesicles that incorporate a minimal number of membrane pores, we reveal that a solitary vesicle can actively propel itself toward an enzyme substrate gradient. Specifically, vesicles loaded with either glucose oxidase or urease and embedded with corresponding transmembrane proteins were tracked within a microfluidic device under a controlled substrate gradient. Our findings establish that a system comprising only an encapsulated enzyme and a single transmembrane pore is sufficient to initiate chemotaxis. This proof-of-concept model underscores the minimalistic yet powerful nature of cellular navigation mechanisms, providing a previously unknown perspective on the origins and evolution of chemotactic behavior in biological systems.

DOI: https://doi.org/10.1126/sciadv.adx9364
Biol Open. 2025 Jul 28. pii: bio.061986. [Epub ahead of print]

Multiomics analysis of GSTP1 knockdown pancreatic cancer cells reveals key regulators of redox and metabolic homeostasis.

Jenna N Duttenhefner, Rahul R Singh, Katherine Schmidt, Katie M Reindl.

  Glutathione S transferase pi-1 (GSTP1) is a detoxification enzyme essential for oxidative homeostasis. In cancer, GSTP1 has been implicated in tumorigenicity, cell cycle progression, and chemoresistance. While GSTP1 depletion has been associated with decreased cancer growth in various models, the mechanism remains poorly understood. This study investigates GSTP1 as a therapeutic target for pancreatic ductal adenocarcinoma (PDAC) using inducible knockdown models. We demonstrate that GSTP1 loss disrupts redox balance, impairs cell survival, and induces metabolic adaptations. Multiomics analysis characterized the global impact of inducible GSTP1 knockdown on the transcriptome and proteome of PDAC cells, identifying 550 differentially expressed genes and 62 proteins. Notably, 43 of these showed consistent regulation at both the mRNA and protein levels. We identify dysregulation of key stress response proteins, including dimethylarginine dimethylaminohydrolase 1 (DDAH1), involved in nitric oxide metabolism, and protein disulfide isomerase A6 (PDIA6), which maintains protein homeostasis. The interplay between GSTP1, DDAH1, and PDIA6 highlights the complexity of redox regulation in pancreatic cancer and suggests that targeting GSTP1 may offer a new therapeutic approach for PDAC.

Keywords:  GSTP1; Multiomics analysis; Pancreatic ductal adenocarcinoma; Redox homeostasis; Therapeutic targets

DOI:  https://doi.org/10.1242/bio.061986
Front Genet. 2025 ;16 1547788

Robust consensus nuclear and cell segmentation.

Melis O Irfan, Eduardo A González-Solares, Tristan Whitmarsh, Alireza Molaeinezhad, Mohammad Al Sa'd, Claire M Mulvey, Marta Páez Ribes, Atefeh Fatemi, Dario Bressan, Nicholas A Walton.

  Cell segmentation is a crucial step in numerous biomedical imaging endeavors-so much so that the community is flooded with publicly available, state-of-the-art segmentation techniques ready for out-of-the-box use. Assessing the strengths and limitations of each method on a tissue sample set and then selecting the optimal method for each research objective and input image are time-consuming and exacting tasks that often monopolize the resources of biologists, biochemists, immunologists, and pathologists, despite not being the primary goal of their research projects. In this work, we present a segmentation software wrapper, coined CellSampler, which runs a selection of established segmentation methods and then combines their individual segmentation masks into a single optimized mask. This so-called "uber mask" selects the best of the established masks across local neighborhoods within the image, where both the neighborhood size and the statistical measure used to define what qualifies as "best" are user-defined.

Keywords:  bioinformatics; computer vision; imaging mass cytometry; multiplexed imaging; single-cell segmentation

DOI:  https://doi.org/10.3389/fgene.2025.1547788
Angew Chem Int Ed Engl. 2025 Jul 26. e202504595

Small Molecule Promoters of Endogenous Lipid Droplet Accumulation Drive Lysophagy.

Sai Kumari Vechalapu, Santhosh Duraisamy, Sathyapriya Senthil, Rakesh Kumar, Deepa Ajnar, Apparao Draksharapu, Suresh Kumar, Dharmaraja Allimuthu.

  Lipid droplets (LDs) are central in regulating metabolism in stress-induced conditions, including one triggered by nutrient deprivation. Genetic manipulation of the lipid metabolic network or supplementation of a high-fat diet/oleic acid (OA) are the traditional routes for voluntarily triggering LD formation in cells and animals to study the role of LDs in disease biology. We developed a new screening platform for identifying small molecule-based LD inducers, which identified linoleic acid (LOA, diunsaturated fatty acid) as a better tool than OA (monounsaturated fatty acid) in promoting LD formation in cells. Subsequent screening and validation discovered a novel heterocyclic compound and respective iron complex for promoting a rapid organization of endogenous lipids into droplets by mimicking desaturase function and modulating oxidative lipid metabolism. Notably, our mass spectral lipidomics analysis presented the overproduction of phosphatidylcholines and small triglycerides (TG), a hallmark of LDs. We uncovered that the abrupt levels of LD formation induced by our molecules promoted a unique cell death mechanism, lysophagy in cancer cells, to prevent their proliferation, movement, and colonization. Collectively, our work introduces new small molecules as powerful tools for reliably promoting LD accumulation in cells, a promising tool for studying the role in health and disease.

Keywords:  High‐content imaging; Lipid droplet; Lipidomics; Lysophagy; Small molecules

DOI:  https://doi.org/10.1002/anie.202504595
Nature. 2025 Jul 30.

ACLY inhibition promotes tumour immunity and suppresses liver cancer.

Jaya Gautam, Jianhan Wu, James S V Lally, Jamie D McNicol, Russta Fayyazi, Elham Ahmadi, Daniela Carmen Oniciu, Spencer Heaton, Roger S Newton, Sonia Rehal, Dipankar Bhattacharya, Fiorella Di Pastena, Binh Nguyen, Celina M Valvano, Logan K Townsend, Suhrid Banskota, Battsetseg Batchuluun, Maria Joy Therese Jabile, Alice Payne, Junfeng Lu, Eric M Desjardins, Naoto Kubota, Evangelia E Tsakiridis, Bejal Mistry, Alex Aganostopoulos, Vanessa Houde, Ann Dansercoer, Koen H G Verschueren, Savvas N Savvides, Joanne A Hammill, Ksenia Bezverbnaya, Paola Muti, Theodoros Tsakiridis, Wenting Dai, Lei Jiang, Yujin Hoshida, Mark Larché, Jonathan L Bramson, Scott L Friedman, Kenneth Verstraete, Dongdong Wang, Gregory R Steinberg.

Immunosuppressive tumour microenvironments are common in cancers such as metabolic dysfunction-associated steatohepatitis (MASH)-driven hepatocellular carcinoma (HCC) (MASH-HCC)1-3. Although immune cell metabolism influences effector function, the effect of tumour metabolism on immunogenicity is less understood4. ATP citrate lyase (ACLY) links substrate availability and mitochondrial metabolism with lipid biosynthesis and gene regulation5-7. Although ACLY inhibition shows antiproliferative effects in various tumours, clinical translation has been limited by challenges in inhibitor development and compensatory metabolic pathways8-12. Here, using a mouse model of MASH-HCC that mirrors human disease, genetic inhibition of ACLY in hepatocytes and tumours reduced neoplastic lesions by over 70%. To evaluate the therapeutic potential of this pathway, a novel small-molecule ACLY inhibitor, EVT0185 (6-[4-(5-carboxy-5-methyl-hexyl)-phenyl]-2,2-dimethylhexanoic acid), was identified via phenotypic screening. EVT0185 is converted to a CoA thioester in the liver by SLC27A2 and structural analysis by cryo-electron microscopy reveals that EVT0185-CoA directly interacts with the CoA-binding site of ACLY. Oral delivery of EVT0185 in three mouse models of MASH-HCC dramatically reduces tumour burden as monotherapy and enhances efficacy of current standards of care including tyrosine kinase inhibitors and immunotherapies. Transcriptomic and spatial profiling in mice and humans linked reduced tumour ACLY with increases in the chemokine CXCL13, tumour-infiltrating B cells and tertiary lymphoid structures. The depletion of B cells blocked the antitumour effects of ACLY inhibition. Together, these findings illustrate how targeting tumour metabolism can rewire immune function and suppress cancer progression in MASH-HCC.

DOI: https://doi.org/10.1038/s41586-025-09297-0
Bioconjug Chem. 2025 Jul 29.

iLOV3.0 Fluorescent Probes for Monitoring ROS Changes in Mammalian Cells.

Shuhui Zhang, Chongyuan Ye, Linyun Guo, Xiaokun Jian, Long Hua, Tangyuan Xie, Tian Huang, Gengsheng Cao.

Monitoring global ROS levels in mammalian cells is essential for capturing redox dynamics that influence both normal physiology and disease progression. Here, we mutated the fluorescent protein iLOV to become more sensitive to ROS in mammalian cells. This mutation enabled the long-term dynamic monitoring of ROS changes in mammalian cells. We confirmed that the mutated iLOV3.0 protein is more sensitive to redox reactions than the iLOV1.0 protein. The iLOV3.0 probe is broadly applicable to mammalian cells, capable of targeting organelles, and exhibits time- and dose-dependent responses to oxidants and reductants while also monitoring ROS changes caused by cellular nutrient stress and alterations in redox-related enzymes. Importantly, the iLOV3.0 probe can track ROS fluctuations during iPSC induction and viral invasion. In summary, this study developed a reactive oxygen species monitoring tool based on the iLOV protein that can be used for mammalian cell imaging. This probe directly utilizes the photophysical properties of iLOV to respond to ROS and undergo fluorescence signal changes without requiring exogenous cofactors. This method is expected to make up for the limitations of existing ROS monitoring tools and provide a new technical means for the dynamic study of the redox states in living cells.

DOI: https://doi.org/10.1021/acs.bioconjchem.5c00208
Bio Protoc. 2025 Jul 20. 15(14): e5393

A Fluorescence-Based Flippase Assay to Monitor Lipid Transport by Drs2-Cdc50.

Inja M Van Der Linden, Sara Abad Herrera, Cédric Montigny, Guillaume Lenoir, Thomas Günther Pomorski, Huriye D Uzun.

  Flippases, a functionally distinct group of transmembrane proteins that flip lipids from the extracellular or luminal side to the cytosolic side of biological membranes, are key players in many important physiological processes, such as membrane trafficking and cellular signaling. To study the function of these membrane proteins under chemically defined conditions, reconstituting them into artificial vesicles is a crucial and effective approach. There are various methods for protein reconstitution involving different detergents and detergent removal techniques to integrate membrane proteins into artificial vesicles. In this protocol, we describe the reconstitution of the yeast flippase complex Drs2-Cdc50, which translocates phosphatidylserine across membranes of the trans-Golgi network at the expense of ATP hydrolysis. The flippase complex is incorporated into liposomes using a zwitterionic detergent, followed by detergent removal via dialysis-a gentle and effective strategy that helps preserve protein function. To evaluate the activity of the reconstituted flippase complex, two complementary assays are employed: (1) a fluorescence-based quenching assay to measure lipid transport; and (2) an ATPase assay using an ATP-regenerating system to measure ATP hydrolysis. Together, these methods provide a robust platform for analyzing the functional reconstitution of Drs2-Cdc50 in a defined membrane environment. Key features • Provides a gentle reconstitution approach via detergent removal by dialysis. • Measures ATPase activity using an NADH-coupled assay with an ATP-regenerating system. • Assesses lipid flippase activity with a sodium dithionite-based assay with fluorescent lipid derivatives. • Provides a well-defined experimental setup for direct characterization of lipid flippases.

Keywords:  Dialysis; Flippase; Floppase; Large unilamellar vesicles; Lipid flipping; Lipid transport; NADH-coupled ATPase assay; Reconstitution

DOI:  https://doi.org/10.21769/BioProtoc.5393
J Cachexia Sarcopenia Muscle. 2025 Aug;16(4): e70024

Six-Minute Walk Test Is Superior to Grip Strength as a Marker of Functional Recovery During Cancer Cachexia Rehabilitation.

Addison Barber, Amber Willbanks, Kathryn Abplanalp, Christopher W Lewis, Ben Binder-Markey, Prakash Jayabalan, Richard L Lieber, Ishan Roy.

   BACKGROUND: Decline in functional independence is a defining event of cancer cachexia, and attempts at creating cachexia-specific therapies have largely failed because of the inability to identify treatments that improve functional capacity. This may be, in part, due to a lack of outcomes that are appropriate and sensitive enough to detect functional recovery. Grip strength is a frequently used outcome measure in cachexia clinical studies; however, the use of gait-based measures is now emerging. These two outcome measures have never been directly compared in the same cohort of cachexia patients regarding their ability to measure and relationship to functional independence. We hypothesize that gait-based measures more comprehensively act as a proxy measure for functional independence related to cachexia.
METHODS: In a retrospective cohort study of 485 cancer patients with a range of cachexia severity and related functional decline who required care at a single-centre inpatient rehabilitation facility (IRF), we assessed the six-minute walk test (6MWT) and hand grip strength (hGS) as proxy measures for functional capacity. Functional capacity is defined as mobility and activities of daily living (ADLs), is quantified by measures of functional independence and referred to here as the Total Motor Score. Cachexia patients were identified primarily using the Fearon et al. consensus criteria, with secondary identification by the Weight Loss Grading Scale (WLGS), Prognostic Nutritional Index (PNI) and neutrophil-to-lymphocyte ratio (NLR). Primary outcomes were change/gain in Total Motor Score, IRF discharge destination (e.g., homebound status or need for care facility) and 6-month survival.
RESULTS: The presence of cachexia in this cohort was 63%. This cohort was 52% male. Mean age was 63 ± 0.63 (SEM) years. Multivariate linear regression demonstrated that change in 6MWT (p < 0.0001) but not hGS (p = 0.084) correlated with Total Motor Score gain after controlling for age, disease burden, cancer type, previous cancer treatment and baseline motor function as covariates. Area under the curve analysis revealed that change in 6MWT (p < 0.0001, AUC = 0.77) was a stronger predictor of Total Motor Score gain than hGS (p = 0.0016, AUC = 0.59). In a multivariate logistic regression model, discharge from IRF to home with independence was predicted by change in 6MWT (p = 0.0007) but not hGS (p = 0.8075). Six-month survival post-rehabilitation was predicted by change in 6MWT (p = 0.0345) but not hGS (p = 0.9025) in a multivariate Cox proportional hazards model.
CONCLUSIONS: Multiple analytical approaches to our data set demonstrate that changes in 6MWT are better associated with cachexia-related outcomes and should be included in future cachexia studies.

Keywords:  cachexia; grip strength; physical function; six‐minute walk test

DOI:  https://doi.org/10.1002/jcsm.70024
Biophys J. 2025 Jul 25. pii: S0006-3495(25)00461-8. [Epub ahead of print]

Adherent cells undergo rate-softening mediated by actomyosin kinetics.

Samuel F Boland, Juan E Abrahante, Patrick W Alford.

Emerging studies suggest that a wide range of chronic diseases can be linked to prior physical trauma and, in some cases, to the supraphysiological deformation rates experienced by cells during injury. However, the mechanical behavior of cells during these deformations is poorly understood. Here, we studied the strain rate dependent mechanics of vascular smooth muscle cells over rates spanning five orders of magnitude, from physiological to supraphysiological. We find that cells deformed at increasing rates undergo substantial rate-softening in tension but have no rate-dependence when returned to zero strain. This reversible rate-softening is mediated by actin-myosin binding kinetics. Further, we find that at supraphysiological strain rates, cells experience actin-myosin binding mediated disruption of contractile force and alteration of gene expression. Our results suggest a mechanism by which cells shield themselves from excessive forces through cytoskeletal relaxation that loses efficacy at high strain rates like those experienced during mechanical trauma.

DOI: https://doi.org/10.1016/j.bpj.2025.07.026
Methods. 2025 Jul 24. pii: S1046-2023(25)00163-X. [Epub ahead of print]

A simple, accurate method for the measurement of lysosomal activity.

Yoshito Iwai, Yuriko Furuya, Yuji Oguro, Keiji Yamamoto.

  Lysosomes are responsible for the degradation of intra- and extracellular components and are thus essential for the quality control of proteins and organelles. Lysosomal dysfunction leads to lysosomal storage diseases, and it is therefore important to identify which types of stress cause functional abnormalities. Lysosomal function is generally evaluated by measuring the enzyme activity of lysosomes with fluorescent dyes. However, fluorescence microscopy can lead to different outcomes due to variations in the field of view, the analysis software used, and the parameter settings. We therefore developed a method that uses only a microplate reader and DQ Green BSA, a dye that emits fluorescence upon lysosomal degradation, to ascertain lysosomal activity. HEK293 cells were treated with DQ Green BSA with or without bafilomycin A1 and lysates extracted using radioimmunoprecipitation buffer. Fluorescence intensities and protein concentrations in the cell lysates were then measured using a microplate reader and the bicinchoninic acid method, respectively, and the fluorescence intensity divided by the protein concentration. Results indicated a significant lysosome inhibitor-induced dose-dependent decrease in the lysosomal activity. The Z'-factor of 0.77 obtained using the proposed method is a significant improvement over the - 0.06 obtained using the conventional method. The versatility of the method was evaluated with different cell types, cell lysis buffers, inhibitors, and protease substrates, with results suggesting that the method works regardless of the cells or reagents used, indicating the relative simplicity and accuracy of the proposed method as compared to the currently utilized method.

Keywords:  Autophagy-lysosomal pathway; Cell lysate; Experimental design; Fluorescent protein; Lysosomal activity; Microplate reader

DOI:  https://doi.org/10.1016/j.ymeth.2025.07.008
ArXiv. 2025 Jul 25. pii: arXiv:2507.19021v1. [Epub ahead of print]

Optimal sensing through phase separation.

Henry Alston, Mason Rouches, Arvind Murugan, Aleksandra M Walczak, Thierry Mora.

Biomolecular condensates form on timescales of seconds in cells upon environmental or compositional changes. Condensate formation is thus argued to act as a mechanism for sensing such changes and quickly initiating downstream processes, such as forming stress granules in response to heat stress and amplifying cGAS enzymatic activity upon detection of cytosolic DNA. Here, we show that phase separation allows cells to discriminate small concentration differences on finite, biologically relevant timescales. We propose optimal sensing protocols, which use the sharp onset of phase separation. We show how, given experimentally measured rates, cells can achieve rapid and robust sensing of concentration differences of 1% on a timescale of minutes, offering an alternative to classical biochemical mechanisms.
Int J Mol Sci. 2025 Jul 08. pii: 6576. [Epub ahead of print]26(14):

Targeted Degradation Technologies Utilizing Autophagy.

Zeyu Zhou, Jiaming Liang, Binghua Cheng, Yanyan Li, Wenjie Zhou, Hui Tian, Wenli Shi, Ke Liu, Lijing Fang, Hongchang Li, Ximing Shao.

  Targeted degradation technologies, primarily referring to targeted protein degradation, have emerged as promising drug discovery strategies. In contrast to traditional "occupancy-driven" inhibition approaches, these technologies ingeniously leverage the cell's endogenous degradation mechanisms to achieve specific elimination of disease-causing targets. Autophagy, a highly conserved cellular clearance pathway, possesses broad substrate recognition capabilities, enabling degradation of not only individual proteins but also protein aggregates, damaged organelles, and invading pathogens. Given these characteristics, researchers are actively exploring the application of autophagy mechanisms in targeted degradation technologies. Herein, we summarize recent advances in autophagy-dependent degradation approaches, including autophagosome tethering compounds (ATTEC), autophagy-targeting chimeras (AUTAC), autophagy-targeting Chimera (AUTOTAC), chaperone-mediated autophagy (CMA)-based methods, nanotechnology-based strategies, and the newly introduced autophagy-induced antibody (AUTAB) technique, highlighting their mechanisms, advantages, and potential applications in treating tumors, neurodegenerative diseases, and other challenging conditions.

Keywords:  autophagy; drug discovery; lysosome; targeted protein degradation

DOI:  https://doi.org/10.3390/ijms26146576