bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2025–05–11
29 papers selected by
Kıvanç Görgülü, Technical University of Munich
  1. Activation of lysosomal iron triggers ferroptosis in cancer.
  2. Loss of lipid asymmetry facilitates plasma membrane blebbing by decreasing membrane lipid packing.
  3. Neural crossroads of pancreatic cancer: how nociceptors drive tumor progression and immune evasion.
  4. Key Mechanisms in Lysosome Stability, Degradation and Repair.
  5. Effect of a scramblase activator upon lipid and probe scrambling and membrane domain formation in HEK 293T cells.
  6. The Curated Cancer Cell Atlas provides a comprehensive characterization of tumors at single-cell resolution.
  7. Aneuploidy generates enhanced nucleotide dependency and sensitivity to metabolic perturbation.
  8. The many faces of membrane tension for biomembranes and vesicles.
  9. MALT1 inhibitor MI-2 induces ferroptosis by direct targeting of GPX4.
  10. Autophagy in cancer and protein conformational disorders.
  11. Exploring the sensitivities of experimental techniques to various types of membrane asymmetry using atomistic simulations.
  12. Reduced Palmitoylation of SQSTM1/p62 in Huntington Disease Is Associated With Impaired Autophagy.
  13. Lipid-degrading small molecule kills cancer cells by ferroptosis.
  14. P4HA1 mediates hypoxia-induced invasion in human pancreatic cancer organoids.
  15. Sympathetic activation of white adipose tissue recruits neutrophils to limit energy expenditure.
  16. AlphaFold2 SLiM screen for LC3-LIR interactions in autophagy.
  17. Annexin A5 controls VDAC1-dependent mitochondrial Ca2+ homeostasis and determines cellular susceptibility to apoptosis.
  18. Detecting ribosome collisions with differential rRNA fragment analysis in ribosome profiling data.
  19. Asymmetric membrane properties through a protein lens.
  20. Pattern formation of lipid domains in bilayer membranes.
  21. PLA2G15 is a BMP hydrolase and its targeting ameliorates lysosomal disease.
  22. The tumor suppressor HNRNPK induces p53-dependent nucleolar stress to drive ribosomopathies.
  23. All-at-once spatial proteome profiling of complex tissue context with single-cell-type resolution by proximity proteomics.
  24. Persistent pseudopod splitting is an effective chemotaxis strategy in shallow gradients.
  25. Protein-induced membrane asymmetry modulates OMP folding kinetics and stability.
  26. m6A alters ribosome dynamics to initiate mRNA degradation.
  27. Lipid-GPCR interactions in an asymmetric plasma membrane model.
  28. Nonequilibrium asymmetry in the living cell membrane.
  29. Biochemical investigation of LC3/GABARAP-ligand interaction as an important quality measure for LC3/GABARAP-targeting small molecules: addendum to the guidelines (4th edition).
  1. Nature. 2025 May 07.
    Activation of lysosomal iron triggers ferroptosis in cancer.
    Tatiana Cañeque, Leeroy Baron, Sebastian Müller, Alanis Carmona, Ludovic Colombeau, Antoine Versini, Stéphanie Solier, Christine Gaillet, Fabien Sindikubwabo, Julio L Sampaio, Marie Sabatier, Eikan Mishima, Armel Picard-Bernes, Laurène Syx, Nicolas Servant, Bérangère Lombard, Damarys Loew, Jiashuo Zheng, Bettina Proneth, Leishemba K Thoidingjam, Laurence Grimaud, Cameron S Fraser, Krystina J Szylo, Emma Der Kazarian, Caroline Bonnet, Emmanuelle Charafe-Jauffret, Christophe Ginestier, Patricia Santofimia-Castaño, Matias Estaras, Nelson Dusetti, Juan Lucio Iovanna, Antonio Sa Cunha, Gabriella Pittau, Pascal Hammel, Dimitri Tzanis, Sylvie Bonvalot, Sarah Watson, Vincent Gandon, Aditya Upadhyay, Derek A Pratt, Florêncio Porto Freitas, José Pedro Friedmann Angeli, Brent R Stockwell, Marcus Conrad, Jessalyn M Ubellacker, Raphaël Rodriguez.
      Iron catalyses the oxidation of lipids in biological membranes and promotes a form of cell death called ferroptosis1. Defining where this chemistry occurs in the cell can inform the design of drugs capable of inducing or inhibiting ferroptosis in various disease-relevant settings. Genetic approaches have revealed suppressors of ferroptosis2-4; by contrast, small molecules can provide spatiotemporal control of the chemistry at work5. Here we show that the ferroptosis inhibitor liproxstatin-1 exerts cytoprotective effects by inactivating iron in lysosomes. We also show that the ferroptosis inducer RSL3 initiates membrane lipid oxidation in lysosomes. We designed a small-molecule activator of lysosomal iron-fentomycin-1-to induce the oxidative degradation of phospholipids and ultimately ferroptosis. Fentomycin-1 is able to kill iron-rich CD44high primary sarcoma and pancreatic ductal adenocarcinoma cells, which can promote metastasis and fuel drug tolerance. In such cells, iron regulates cell adaptation6,7 while conferring vulnerability to ferroptosis8,9. Sarcoma cells exposed to sublethal doses of fentomycin-1 acquire a ferroptosis-resistant cell state characterized by the downregulation of mesenchymal markers and the activation of a membrane-damage response. This phospholipid degrader can eradicate drug-tolerant persister cancer cells in vitro and reduces intranodal tumour growth in a mouse model of breast cancer metastasis. Together, these results show that control of iron reactivity confers therapeutic benefits, establish lysosomal iron as a druggable target and highlight the value of targeting cell states10.
    DOI:  https://doi.org/10.1038/s41586-025-08974-4
  2. Proc Natl Acad Sci U S A. 2025 May 13. 122(19): e2417145122
    Loss of lipid asymmetry facilitates plasma membrane blebbing by decreasing membrane lipid packing.
    Hong-Yin Wang, Alissa Rumin, Milka Doktorova, Daryna Sputay, Sze Ham Chan, Ann M Wehman, Kandice R Levental, Ilya Levental.
      Membrane blebs have important roles in cell migration, apoptosis, and intercellular communication through extracellular vesicles (EVs). While plasma membranes (PM) typically maintain phosphatidylserine (PS) on their cytoplasmic leaflet, most blebs have PS exposed on their outer leaflet, revealing that loss of steady-state lipid asymmetry often accompanies PM blebbing. How these changes in PM lipid organization regulate membrane properties and affect bleb formation remains unknown. We confirmed that lipid scrambling through the scramblase TMEM16F is essential for chemically induced membrane blebbing across cell types, with the kinetics of PS exposure being tightly coupled to the kinetics of bleb formation. Measurement of lipid packing with environment-sensitive probes revealed that lipid scrambling changes the physical properties of the PM, reducing lipid packing and facilitating the bilayer bending required for bleb formation. Accordingly, reducing lipid packing of the PM through cholesterol extraction, elevated temperature, or treatment with biological amphiphiles promoted blebbing in the absence of TMEM16F. Consistent with these cellular observations, blebbing in Caenorhabditis elegans embryos measured via EV production was significantly reduced by depleting the TMEM16-homolog ANOH-2. Our findings suggest that changing membrane biophysical properties by lipid scrambling is an important contributor to the formation of blebs and EVs and potentially other cellular processes involving PM deformation.
    Keywords:  extracellular vesicle; lipid asymmetry; lipid scrambling; membrane blebbing; membrane stiffness
    DOI:  https://doi.org/10.1073/pnas.2417145122
  3. Cell Res. 2025 May 07.
    Neural crossroads of pancreatic cancer: how nociceptors drive tumor progression and immune evasion.
    Carmen Mota Reyes, Helmut Friess, Ihsan Ekin Demir.
      
    DOI:  https://doi.org/10.1038/s41422-025-01124-5
  4. Mol Cell Biol. 2025 May 09. 1-13
    Key Mechanisms in Lysosome Stability, Degradation and Repair.
    Rui Zhang, Marc A Vooijs, Tom Gh Keulers.
      Lysosomes are organelles that play pivotal roles in macromolecule digestion, signal transduction, autophagy, and cellular homeostasis. Lysosome instability, including the inhibition of lysosomal intracellular activity and the leakage of their contents, is associated with various pathologies, including cancer, neurodegenerative diseases, inflammatory diseases and infections. These lysosomal-related pathologies highlight the significance of factors contributing to lysosomal dysfunction. The vulnerability of the lysosomal membrane and its components to internal and external stimuli make lysosomes particularly susceptible to damage. Cells are equipped with mechanisms to repair or degrade damaged lysosomes to prevent cell death. Understanding the factors influencing lysosome stabilization and damage repair is essential for developing effective therapeutic interventions for diseases. This review explores the factors affecting lysosome acidification, membrane integrity, and functional homeostasis and examines the underlying mechanisms of lysosomal damage repair. In addition, we summarize how various risk factors impact lysosomal activity and cell fate.
    Keywords:  ESCRT; Lysosome stabilization; ROS; lipid peroxidation; lysophagy; lysosomal membrane permeabilization
    DOI:  https://doi.org/10.1080/10985549.2025.2494762
  5. Faraday Discuss. 2025 May 09.
    Effect of a scramblase activator upon lipid and probe scrambling and membrane domain formation in HEK 293T cells.
    Shinako Kakuda, Erwin London.
      Lipid asymmetry, a difference between the lipid composition of the inner and outer leaflets (monolayers) of a membrane, is a characteristic of mammalian plasma membranes. In artificial lipid vesicles, asymmetry can either suppress or induce the formation of coexisting ordered and disordered lipid domains depending on lipid composition. In mammalian plasma membrane preparations, loss of asymmetry induces the formation of ordered domains. In this report, we studied the effect of a scramblase activator, the ionophore BrA23187 (BrA) plus Ca2+, upon ordered domain (lipid raft) formation in human embryonic kidney 293T cells. Addition of BrA induced a decrease in FRET between the plasma membrane outer leaflet probe TMADPH, which partly associates with ordered domains, and ODRB, which localizes largely in liquid disordered domains. This is consistent with the formation of coexisting ordered and disordered domains in the plasma membrane. In addition, upon BrA addition, the plasma membrane outer leaflet probe Pro12A exhibited a decrease in the generalized polarization (GP) suggesting a decrease in outer leaflet membrane order, perhaps due to a decrease in outer leaflet cholesterol However, there are other explanations for these observations. To test if BrA induced scrambling of fluorescent membrane probes, which would complicate interpretation of the experiments described above, we measured the effect of BrA upon extractability of outer leaflet probes with MβCD (in most cases, MβCD was more effective for extraction than BSA). These experiments showed that at most a small fraction of probes migrate to the inner leaflet upon addition of BrA. Other experiments raise the possibility that BrA binding to membranes may directly influence ordered domain formation and properties or alter fluorescence by direct interactions with TMADPH, and thus not reflect changes in domain formation.
    DOI:  https://doi.org/10.1039/d4fd00211c
  6. Nat Cancer. 2025 May 08.
    The Curated Cancer Cell Atlas provides a comprehensive characterization of tumors at single-cell resolution.
    Michael Tyler, Avishai Gavish, Chaya Barbolin, Roi Tschernichovsky, Rouven Hoefflin, Michael Mints, Sidharth V Puram, Itay Tirosh.
      Recent years have seen a rapid proliferation of single-cell cancer studies, yet most of these studies profiled few tumors, limiting their statistical power. Combining data and results across studies holds great promise but also involves various challenges. We recently began to address these challenges by curating a large collection of cancer single-cell RNA-sequencing datasets, leveraging it for systematic analyses of tumor heterogeneity. Here we greatly extend this repository to 124 datasets for over 40 cancer types, together comprising 2,836 samples, with improved data annotations, visualizations and exploration. Using this vast cohort, we generate an updated map of recurrent expression programs in malignant cells and systematically quantify context-dependent gene expression and cell-cycle patterns across cell types and cancer types. These data, annotations and analysis results are all freely available for exploration and download through the Curated Cancer Cell Atlas, a central community resource that opens new avenues in cancer research.
    DOI:  https://doi.org/10.1038/s43018-025-00957-8
  7. Genes Dev. 2025 May 05.
    Aneuploidy generates enhanced nucleotide dependency and sensitivity to metabolic perturbation.
    Rayna Y Magesh, Arshia N Kaur, Faith N Keller, Abdulrazak Frederick, Tenzin Tseyang, John A Haley, Alejandra M Rivera-Nieves, Anthony C Liang, David A Guertin, Jessica B Spinelli, Stephen J Elledge, Emma V Watson.
      Despite the general detriment of aneuploidy to cellular fitness, >90% of solid tumors carry an imbalanced karyotype. This existing paradox and the molecular responses to aneuploidy remain poorly understood. Here, we explore these cellular stresses and unique vulnerabilities of aneuploidy in human mammary epithelial cells (HMECs) enriched for breast cancer-associated copy number alterations (CNAs). To uncover the genetic dependencies specific to aneuploid cells, we conducted a comprehensive, genome-wide CRISPR knockout screen in isogenic aneuploid and diploid HMEC lines. Our study reveals that aneuploid HMECs exhibit an increased reliance on pyrimidine biosynthesis and mitochondrial oxidative phosphorylation genes and demonstrate heightened fitness advantages upon loss of tumor suppressor genes. Using an integrative multiomic analysis, we confirmed nucleotide pool insufficiency as a key contributor to widespread cellular dysfunction in aneuploid HMECs with net copy number gain. Although diploid cells can switch seamlessly between pyrimidine synthesis and salvage, cells with increased chromosomal content exhibit p53 activation and S-phase arrest when relying on salvage alone, alongside increased sensitivity to DNA-damaging chemotherapeutics. This work advances our understanding of the consequences of aneuploidy and uncovers potential avenues for patient stratification and therapeutic intervention based on tumor ploidy.
    Keywords:  CRISPR; aneuploidy; cancer; genomics; metabolism
    DOI:  https://doi.org/10.1101/gad.352512.124
  8. Faraday Discuss. 2025 May 06.
    The many faces of membrane tension for biomembranes and vesicles.
    Reinhard Lipowsky.
      Membrane tension is generated by external forces and constraints that affect the membrane's surface area. Many studies have been pursued to elucidate this intuitive concept. Here, we focus on biomimetic model systems, for which the notion of membrane tension can be elaborated in a quantitative manner. Bottom-up, these membrane systems are planar bilayers, unilamellar nanovesicles, and giant unilamellar vesicles (GUVs). For planar bilayers and nanovesicles, we need to distinguish the mechanical bilayer tension within the whole bilayer from the mechanical leaflet tensions within the individual bilayer leaflets. In addition, the fluctuation spectrum of the bilayer defines the bilayer's fluctuation tension. These different tensions can be determined by molecular dynamics simulations. On the micron-scale, GUV membranes as observed by light microscopy, experience both a mechanical and a curvature-elastic tension, as follows from the theory of curvature elasticity. The curvature-elastic tension represents a material parameter, whereas the mechanical tension is a kind of hidden variable that depends on the size and shape of the membrane as demonstrated for multispherical shapes of GUVs. Furthermore, the excess membrane area stored in the shape fluctuations is again governed by a fluctuation tension. Many membrane systems exhibit characteristic tension thresholds. Examples include, the shapes of tubular membranes and the morphological responses of vesicle membranes to condensate droplets.
    DOI:  https://doi.org/10.1039/d4fd00184b
  9. Proc Natl Acad Sci U S A. 2025 May 20. 122(20): e2507997122
    MALT1 inhibitor MI-2 induces ferroptosis by direct targeting of GPX4.
    Eikan Mishima, Thomas J O'Neill, Kai P Hoefig, Deng Chen, Gesine Behrens, Bernhard Henkelmann, Junya Ito, Kiyotaka Nakagawa, Vigo Heissmeyer, Marcus Conrad, Daniel Krappmann.
      
    DOI:  https://doi.org/10.1073/pnas.2507997122
  10. FEBS Lett. 2025 May 08.
    Autophagy in cancer and protein conformational disorders.
    Sergio Attanasio.
      Autophagy is a catabolic process by which cells maintain cellular homeostasis through the degradation of dysfunctional cytoplasmic components, such as toxic misfolded proteins and damaged organelles, within the lysosome. It is a multistep process that is tightly regulated by nutrient, energy, and stress-sensing mechanisms. Autophagy plays a pivotal role in various biological processes, including protein and organelle quality control, defense against pathogen infections, cell metabolism, and immune surveillance. As a result, autophagy dysfunction is linked to a variety of pathological conditions. The role of autophagy in cancer is complex and dynamic. Depending on the context, autophagy can have both tumor-suppressive and pro-tumorigenic effects. In contrast, its role is more clearly defined in protein conformational disorders, where autophagy serves as a mechanism to reduce toxic protein aggregation, thereby improving cellular homeostasis. Because autophagy-based therapies hold promising potential for the treatment of cancer and protein conformational disorders, this review will highlight the latest findings and advancements in these areas.
    Keywords:  RAS‐ and RAF‐induced cancer; alpha‐1 antitrypsin deficiency; autophagy; autophagy inhibitors; clinical trials; neurodegenerative disorders; pancreatic ductal adenocarcinoma; protein misfolding; proteinopathies
    DOI:  https://doi.org/10.1002/1873-3468.70061
  11. Faraday Discuss. 2025 May 08.
    Exploring the sensitivities of experimental techniques to various types of membrane asymmetry using atomistic simulations.
    Frederick A Heberle, Milka Doktorova.
      Biological membranes have two leaflets that can differ in both lipid composition and total lipid abundance. These different types of asymmetries play a major role in determining the biophysical properties of the membrane; however, they have proven challenging to assay experimentally even in simpler model systems. Molecular dynamics simulations offer the means for detailed computational investigation of systematically varied interleaflet lipid distributions, but opportunities for critical validation with wet lab experiments are scarce. To help address this problem, here we use atomistic simulations of asymmetric bilayers to generate synthetic experimental data and thus investigate the sensitivity of various approaches to changes in relative lipid composition, number, and cholesterol distribution. Contrary to trends in symmetric bilayers, the simulations showed a decrease in lipid packing with increasing cholesterol in differentially stressed asymmetric bilayers, with more pronounced changes in the more loosely packed leaflet. Representative experimental data computed from the simulation trajectories indicated that the detection of asymmetry-induced changes in leaflet properties should be possible with environment-sensitive fluorescent probes and NMR observables, but may require optimization of sample preparation conditions. On the other hand, small-angle scattering data are already experimentally accessible and can reveal differential leaflet packing densities through a model-free analysis. We further show that computationally generated cryo-EM intensity profiles are highly sensitive to phospholipid imbalance between membrane leaflets. Together, these findings provide a roadmap for developing targeted applications of the in vitro techniques and obtaining experimental data critical for validating computationally derived principles related to membrane asymmetry.
    DOI:  https://doi.org/10.1039/d4fd00200h
  12. FASEB J. 2025 May 15. 39(9): e70549
    Reduced Palmitoylation of SQSTM1/p62 in Huntington Disease Is Associated With Impaired Autophagy.
    F Abrar, M C Davies, Y Alshehabi, A Kumar, A Dang, Y T N Nguyen, J Collins, N S Caron, J S Choudhary, S S Sanders, M O Collins, M R Hayden, D D O Martin.
      Disruption of autophagy has emerged as a common feature in many neurodegenerative diseases. Autophagy is a membrane-dependent pathway that requires many key regulators to quickly localize on and off membranes during induction, promoting membrane fusion. Previously, our bioinformatic approaches have shown that autophagy and Huntington disease (HD) are enriched in palmitoylated proteins. Palmitoylation involves the reversible addition of long-chain fatty acids to promote membrane binding. Herein, we show that inhibition of palmitoylation regulates the abundance of several key regulators of autophagy and leads to a partial block of autophagic flux. We confirm that the autophagy receptor SQSTM1/p62 (sequestosome 1) is palmitoylated and directed to the lysosome. Importantly, we report that SQSTM1 palmitoylation is significantly reduced in HD patient and mouse model brains. This finding reveals a novel mechanism contributing to the generation of empty autophagosomes previously seen in HD models and patient-derived cells.
    DOI:  https://doi.org/10.1096/fj.202401781R
  13. Nature. 2025 May 07.
    Lipid-degrading small molecule kills cancer cells by ferroptosis.
    Mike Lange, James A Olzmann.
      
    Keywords:  Cancer; Cell biology
    DOI:  https://doi.org/10.1038/d41586-025-01337-z
  14. Cancer Res Commun. 2025 May 07.
    P4HA1 mediates hypoxia-induced invasion in human pancreatic cancer organoids.
    Bernat Navarro-Serer, Maria F Wissler, Brandi K Glover, Michael G Lerner, Harsh H Oza, Vania Wang, Hidur Knutsdottir, Fatemeh Shojaeian, Kathleen Noller, Saravana Gowtham Baskaran, Sarah Hughes, Alana M Weaver, Daniel Wilentz, Oluwatobiloba Olayemi, Joel S Bader, Elana J Fertig, Daniele M Gilkes, Laura D Wood.
      Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive malignancy with dismal prognosis. PDAC develops in a hypoxic environment in which cells adapt and activate processes to allow survival under low oxygen conditions, some of which may enhance the ability of cancer cells to invade locally or metastasize distantly. Using human PDAC organoids, we determined that hypoxia consistently enhanced invasion across 11 patient-derived models. Using RNA-sequencing of hypoxic invasive organoids (compared with matched invasive normoxic organoids from the same patients), we identified prolyl 4-hydroxylase subunit alpha 1 (P4HA1) as a potential regulator of PDAC invasion in hypoxia. Leveraging publicly available datasets from human tissue, we determined that P4HA1 is more highly expressed in PDAC compared to normal pancreatic tissue and that high P4HA1 expression correlates with poor patient prognosis. To further interrogate the role of P4HA1 in invasion of hypoxic patient-derived organoids, we quantified invasion in organoids modified to knockdown and or overexpress P4HA1, demonstrating that P4HA1 is necessary for hypoxia-enhanced invasion and sufficient to increase invasion in normoxia in PDAC organoids. Our results identify P4HA1 as a driver of PDAC organoid invasion in hypoxia.
    DOI:  https://doi.org/10.1158/2767-9764.CRC-24-0025
  15. Res Sq. 2025 Apr 16. pii: rs.3.rs-6414640. [Epub ahead of print]
    Sympathetic activation of white adipose tissue recruits neutrophils to limit energy expenditure.
    Seunghwan Son, Cindy Xu, Janice Jang, Maddox Dinh, Yuliya Skorobogatko, Haipeng Fu, Joseph M Valentine, Garam An, Wei Ying, Ruth T Yu, Michael Downes, Ronald M Evans, Alan R Saltiel.
      Adipose tissue maintains energy homeostasis by storing lipids during nutrient surplus and releasing them through lipolysis in times of energy demand. While lipolysis is essential for short term metabolic adaptation, prolonged metabolic stress requires adaptive changes that preserve energy reserves. Here, we report that β-adrenergic activation of adipocytes induces a transient and depot-specific infiltration of neutrophils into white adipose tissue (WAT), particularly in lipid-rich visceral WAT. Neutrophil recruitment requires the stimulation of both lipolysis and p38 MAPK activation in adipocytes. Recruited neutrophils locally secrete IL-1β, which suppresses lipolysis and limits excessive energy expenditure. Neutrophil depletion or blockade of IL-1β production increased lipolysis, leading to reduced WAT mass upon repeated β3-adrenergic stimulation. Together, these findings reveal an unexpected role of neutrophil-derived IL-1β in preserving lipid stores during metabolic stress, highlighting a physiological function of innate immune cells in maintaining energy homeostasis.
    DOI:  https://doi.org/10.21203/rs.3.rs-6414640/v1
  16. Autophagy. 2025 May 04. 1-21
    AlphaFold2 SLiM screen for LC3-LIR interactions in autophagy.
    Jan Felix Maximilian Stuke, Gerhard Hummer.
      In selective macroautophagy/autophagy, cargo recruitment is mediated by MAP1LC3/LC3-interacting regions (LIRs)/Atg8-family interacting motifs (AIMs) in the cargo or cargo receptor proteins. The binding of these motifs to LC3/Atg8 proteins at the phagophore membrane is often modulated by post-translational modifications, especially phosphorylation. As a challenge for computational LIR predictions, sequences may contain the short canonical (W/F/Y)XX(L/I/V) motif without being functional. Conversely, LIRs may be formed by non-canonical but functional sequence motifs. AlphaFold2 has proven to be useful for LIR predictions, even if some LIRs are missed and proteins with thousands of residues reach the limits of computational feasibility. We present a fragment-based approach to address these limitations. We find that fragment length and phosphomimetic mutations modulate the interactions predicted by AlphaFold2. Systematic fragment screening for a range of target proteins yields structural models for interactions that AlphaFold2 and AlphaFold3 fail to predict for full-length targets. We provide guidance on fragment choice, sequence tuning, LC3 isoform effects, and scoring for optimal LIR screens. Finally, we also test the transferability of this general framework to SUMO-SIM interactions, another type of protein-protein interaction involving short linear motifs (SLiMs).Abbreviations: 2-HP-LIR: ncLIR binding either or both HPs with non-canonical residues; AIM: Atg8-family interacting motif; ap. LIR: antiparallel LIR; A.t.; Arabidopsis thaliana; AT5G06830/C53 (A.t.): CDK5RAP3-like protein; Atg8/ATG8: autophagy related 8, in yeast and plants, respectively; ATG8CL: ATG8C-like of Solanum tuberosum (potato); ATG8E: ATG8e of A.t.; Av. num. of contacts: average number of heavy atom contacts; BCL2: BCL2 apoptosis regulator; BNIP3: BCL2 interacting protein 3; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CALR: calreticulin; can. LIR: canonical LIR; CDF: cumulative distribution function; CDK5RAP3/C53 (H.s.): CDK5 regulatory subunit associated protein 3; [DE]W[DE]-LIR: TRIM5-like ncLIR; DSK2A: ubiquitin domain-containing protein DSK2a; FUNDC1: FUN14 domain containing 1; GABARAP: GABA type A receptor-associated protein; HP0/1/2: hydrophobic pocket 0/1/2; HP0-LIR: ncLIR engaging HP0; H.s.; Homo sapiens; lcLIR: low-confidence LIR (ncLIR not similar to previously characterized ncLIRs); LDS: LIR-docking site; LIR: LC3-interacting region; LO score: length-weighted fraction of occurrence score; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MD: molecular dynamics; MEFV/pyrin: MEFV innate immunity regulator, pyrin; minPAE: minimum PAE; MSA: multiple sequence alignment; ncLIR: non-canonical LIR; NPC: nuclear pore complex; Nup159: nucleoporin 159; NUP214: nucleoporin 214; OPTN: optineurin; other@LDS: other interaction proximal to the LIR-docking site; PAE: predicted aligned error; PDCD6IP: programmed cell death 6 interacting protein; PDF: probability distribution function; pLDDT: predicted local-distance difference test; PLEKHM1: pleckstrin homology and RUN domain containing M1; PTM: post-translational modification; sAIM: shuffled AIM (ncLIR with shuffled motif); seq.: sequence; SIM: SUMO-interacting motif; SLiM: short linear motif; SMN1/SMN: survival of motor neuron 1, telomeric; ST: phosphomimetic; STBD1: starch binding domain 1; STK3: serine/threonine kinase 3; SUMO: small ubiquitin like modifier; TBC1D2/TBC1D2A: TBC1 domain family member 2; TEX264: testis expressed 264, ER-phagy receptor; TRIM5/TRIM5α: tripartite motif-containing protein 5; UDS: UIM-docking site; UIM: ubiquitin-interacting motif; UIMC1/RAP80: ubiquitin interaction motif containing 1; ULK1: unc-51 like autophagy activating kinase 1; ULK2: unc-51 like autophagy activating kinase 2; WT: wild type.
    Keywords:  AIM; Atg8; SUMO-SIM interaction; phosphorylation; prediction; selective autophagy receptor
    DOI:  https://doi.org/10.1080/15548627.2025.2493999
  17. EMBO J. 2025 May 09.
    Annexin A5 controls VDAC1-dependent mitochondrial Ca2+ homeostasis and determines cellular susceptibility to apoptosis.
    Furkan E Oflaz, Alexander I Bondarenko, Michael Trenker, Markus Waldeck-Weiermair, Benjamin Gottschalk, Eva Bernhart, Zhanat Koshenov, Snježana Radulović, Rene Rost, Martin Hirtl, Johannes Pilic, Aditya Karunanithi Nivedita, Adlet Sagintayev, Gerd Leitinger, Bent Brachvogel, Susanne Summerauer, Varda Shoshan-Barmatz, Roland Malli, Wolfgang F Graier.
      Annexin A5 (AnxA5) is a Ca2+-dependent phospholipid-binding protein associated with the regulation of intracellular Ca2+ homeostasis. However, the precise role of AnxA5 in controlling mitochondrial Ca2+ signaling remains elusive. Here, we introduce a novel function of AnxA5 in regulating mitochondrial Ca2+ signaling. Our investigation revealed that AnxA5 localizes at and in the mitochondria and orchestrates intermembrane space Ca2+ signaling upon high Ca2+ elevations induced by ER Ca2+ release. Proximity ligation assays and co-immunoprecipitation revealed a close association but no direct contact of AnxA5 with the voltage-dependent anion channel (VDAC1) in the outer mitochondrial membrane (OMM). In single-cell mitochondrial Ca2+ measurements and electrophysiological recordings, AnxA5 was found to enhance Ca2+ flux through the OMM by promoting the Ca2+-permeable state of VDAC1. By modulating intermembrane space Ca2+ signaling, AnxA5 shapes mitochondrial ultrastructure and influences the dynamicity of the mitochondrial Ca2+ uniporter. Furthermore, by controlling VDAC1's oligomeric state, AnxA5 is protective against cisplatin and selenite-induced apoptotic cell death. Our study uncovers AnxA5 as an integral regulator of VDAC1 in physiological and pathological conditions.
    Keywords:  Annexin-A5; Apoptotic Cell Death; Intermembrane Space Ca2⁺ Signaling; VDAC1 Ca2+ Permeability
    DOI:  https://doi.org/10.1038/s44318-025-00454-9
  18. NAR Genom Bioinform. 2025 Jun;7(2): lqaf045
    Detecting ribosome collisions with differential rRNA fragment analysis in ribosome profiling data.
    Edwin Sakyi Kyei-Baffour, Jitske Bak, Joana Silva, William J Faller, Ferhat Alkan.
      It has become clear in recent years that ribosomes regularly stall during translation. Such translation impairment has many causes, including exposure to ribotoxic stress agents, the presence of specific RNA structures or sequences, or a shortage of amino acids or translation factors. If they are not resolved, stalled ribosomes can lead to ribosome collisions that are continuously surveilled by various sensor proteins. This in turn initiates a cascade of signalling events that can change the physiology and behaviour of cells. However, measuring changes in collision abundance has proved challenging, and as a result, the importance of collision-mediated biological responses is still unclear. Here, we show that computational analyses of standard ribosome profiling (Ribo-seq) data enable the prediction of changes in ribosome collisions between conditions. This is achieved by using the known 3D structure of collided ribosomes to define the ribosomal RNA (rRNA) positions that are differentially digested by RNases during the Ribo-seq protocol. Comparison of the relative rRNA reads at these positions allows the relative quantification of collisions between samples, an approach we call differential ribosome collisions by Analysis of rRNA Fragments (dricARF). When applied to public datasets across multiple organisms, our approach detects changes in collision events with unprecedented accuracy and sensitivity. In addition to providing supplementary evidence for ribosome collisions, our tool has the potential to uncover novel biological processes that are mediated by them. dricARF is available as part of the ARF R package and can be accessed through https://github.com/fallerlab/ARF.
    DOI:  https://doi.org/10.1093/nargab/lqaf045
  19. Faraday Discuss. 2025 May 06.
    Asymmetric membrane properties through a protein lens.
    Joseph H Lorent, Angela Cabrera-Jojoa, Kandice R Levental, Ilya Levental, Edward Lyman.
      Plasma membranes are asymmetric, with each monolayer presenting specific lipid compositions and biophysical properties. Transmembrane domains (TMDs) of single-pass transmembrane proteins (spTMPs) have adapted their physico-chemical properties to these asymmetric constraints. In this study, we analysed the structural features of such TMDs across the tree of life to obtain information about their interaction with asymmetric membrane bilayers and predict species-specific membrane properties. We observed that TMDs in the plasma membranes of all eukaryotic species possess asymmetries in lipid accessible surface area (ASA), hydrophobicity, aromaticity and charge. Bacteria deviate from this trend, with strong differences between bacterial clades. Notably, TMDs in the Golgi and the endoplasmic reticulum of eukaryotic species display inverted profiles for accessible surface area, hydrophobicity and aromaticity compared to their plasma-membrane counterparts. To determine how well TMD profiles reflect average membrane properties, we performed molecular dynamics simulations of a spTMP in an asymmetric lipid bilayer whose composition approximates the human plasma membrane. The simulated spTMP was chosen to represent the average TMD properties of the human proteome. We compared the electron density profiles of the simulated asymmetric membrane to the average TMD profiles derived from the human proteome and observed that phospholipid acyl-chain density overlapped very well with TMD hydrophobicity, and phosphate group density with TMD charge. The profiles of phospholipid unsaturation in the acyl chains overlapped well with the average location of TMD phenylalanines in the cytoplasmic leaflet, while there was additional accumulation of large hydrophobic and aromatic residues in the membrane midplane, which had low acyl-chain density. This study reveals the complementarity of membrane and TMD properties in asymmetric membranes, suggesting that the properties of TMDs can be used to make predictions about the properties of their solvating membranes.
    DOI:  https://doi.org/10.1039/d4fd00199k
  20. Soft Matter. 2025 May 09.
    Pattern formation of lipid domains in bilayer membranes.
    Qiwei Yu, Andrej Košmrlj.
      Phase separation plays an important role in spatial organization and material distribution of biological membranes, which are essential for crucial biological functions ranging from signaling and stress response to vesicle trafficking. Domains arising from demixing of molecules coarsen indefinitely unless growth is arrested at a finite size by additional mechanisms (e.g., membrane elasticity). The resulting finite-size domains self-organize into regular patterns such as stripes and dots, which are called modulated phases. Here, we examine the size and morphology of lipid domains with a minimal theoretical model that considers both the elastic deformation of the membrane and the chemical interactions between lipids, which are coupled by a preferred membrane curvature that depends on the local lipid composition. Microscopically, the coupling is caused by an asymmetry between leaflets which emerges after extra lipids (e.g., DPPC) are introduced to the outer leaflet. The additional lipid partitions preferentially to domains where it is enriched, creating a preferred curvature that depends on local composition. We use an amplitude expansion to determine the domain size and morphology of patterns that minimize the total free energy, which is validated by numerical simulations and compared against experiments in synthetic model membranes and cell-derived membranes. The morphology of patterns varies with membrane lipid composition following a complex morphological diagram, which is in good agreement with experiments. The domain size decreases monotonically with a membrane bending modulus but can be non-monotonic with surface tension. Our results offer testable predictions, such as pattern hysteresis upon cycling external stimuli, diverse pattern morphology near critical points, and non-monotonic dependence of the domain size on osmotic pressure, which motivate future experiments. The presented theoretical framework is generally applicable to pattern formation on deformable surfaces.
    DOI:  https://doi.org/10.1039/d5sm00276a
  21. Nature. 2025 May 07.
    PLA2G15 is a BMP hydrolase and its targeting ameliorates lysosomal disease.
    Kwamina Nyame, Jian Xiong, Hisham N Alsohybe, Arthur P H de Jong, Isabelle V Peña, Ricardo de Miguel, Thijn R Brummelkamp, Guido Hartmann, Sebastian M B Nijman, Matthijs Raaben, Judith A Simcox, Vincent A Blomen, Monther Abu-Remaileh.
      Lysosomes catabolize lipids and other biological molecules, maintaining cellular and organismal homeostasis. Bis(monoacylglycero)phosphate (BMP), a major lipid constituent of intralysosomal vesicles, stimulates lipid-degrading enzymes and is altered in various human conditions, including neurodegenerative diseases1,2. Although lysosomal BMP synthase was recently discovered3, the enzymes mediating BMP turnover remain elusive. Here we show that lysosomal phospholipase PLA2G15 is a physiological BMP hydrolase. We further demonstrate that the resistance of BMP to lysosomal hydrolysis arises from its unique sn2, sn2' esterification position and stereochemistry, as neither feature alone confers resistance. Purified PLA2G15 catabolizes most BMP species derived from cell and tissue lysosomes. Furthermore, PLA2G15 efficiently hydrolyses synthesized BMP stereoisomers with primary esters, challenging the long-held thought that BMP stereochemistry alone ensures resistance to acid phospholipases. Conversely, BMP with secondary esters and S,S stereoconfiguration is stable in vitro and requires acyl migration for hydrolysis in lysosomes. Consistent with our biochemical data, PLA2G15-deficient cells and tissues accumulate several BMP species, a phenotype reversible by supplementing wild-type PLA2G15 but not its inactive mutant. Targeting PLA2G15 reduces the cholesterol accumulation in fibroblasts of patients with Niemann-Pick disease type C1 and significantly ameliorates disease pathologies in Niemann-Pick disease type C1-deficient mice, leading to an extended lifespan. Our findings established the rules governing BMP stability in lysosomes and identified PLA2G15 as a lysosomal BMP hydrolase and a potential target for therapeutic intervention in neurodegenerative diseases.
    DOI:  https://doi.org/10.1038/s41586-025-08942-y
  22. J Clin Invest. 2025 May 08. pii: e183697. [Epub ahead of print]
    The tumor suppressor HNRNPK induces p53-dependent nucleolar stress to drive ribosomopathies.
    Pedro Aguilar-Garrido, María Velasco-Estévez, Miguel Ángel Navarro-Aguadero, Alvaro Otero-Sobrino, Marta Ibañez-Navarro, Miguel Ángel Marugal, María Hernández-Sánchez, Prerna Malaney, Ashley Rodriguez, Oscar Benitez, Xiaorui Zhang, Marisa Jl Aitken, Alejandra Ortiz-Ruiz, Diego Megias, Manuel Pérez-Martínez, Gadea Mata, Jesús Gomez, Miguel Lafarga, Orlando Dominguez, Osvaldo Graña-Castro, Eduardo Caleiras, Pilar Ximenez-Embun, Marta Isasa, Paloma J de Andrés, Sandra Rodriguez-Perales, Raul Torres-Ruiz, Enrique Revilla, Rosa María García-Martín, Daniel Azorín, Josune Zubicaray, Julian Sevilla, Oleksandra Sirozh, Vanesa Lafarga, Joaquín Martinez-Lopez, Sean M Post, Miguel Gallardo.
      The nucleolus is a membraneless organelle and an excellent stress sensor. Any changes in its architecture or composition lead to nucleolar stress, resulting in cell cycle arrest and interruption of ribosomal activity, critical factors in aging and cancer. In this study, we identified and described the pivotal role of the RNA-binding protein (RBP) HNRNPK in ribosome and nucleolar dynamics. We developed an in vitro model of endogenous HNRNPK overexpression and an in vivo mouse model of ubiquitous HNRNPK overexpression. These models showed disruptions in translation and caused alterations in the nucleolar structure, resulting in p53-dependent nucleolar stress, cell cycle arrest, senescence, and bone marrow failure phenotype, similar to what is observed in patients with ribosomopathies. Together, our findings identify HNRNPK as a master regulator of ribosome biogenesis (RiBi) and nucleolar homeostasis through p53, providing a new perspective on the orchestration of nucleolar integrity, ribosome function and cellular senescence.
    Keywords:  Aging; Cellular senescence; Hematology; Hematopoietic stem cells; Mouse models; Stem cells
    DOI:  https://doi.org/10.1172/JCI183697
  23. Cell Syst. 2025 May 05. pii: S2405-4712(25)00124-3. [Epub ahead of print] 101291
    All-at-once spatial proteome profiling of complex tissue context with single-cell-type resolution by proximity proteomics.
    Yiheng Mao, Yuan Li, Zhendong Zheng, Yanfen Xu, Mi Ke, An He, Fuchao Liang, Keren Zhang, Xi Wang, Weina Gao, Ruijun Tian.
      Spatial proteomics enables in-depth mapping of tissue architectures, mostly achieved by laser microdissection-mass spectrometry (LMD-MS) and antibody-based imaging. However, trade-offs among sampling precision, throughput, and proteome coverage still limit the applicability of these strategies. Here, we propose proximity labeling for spatial proteomics (PSPro) by combining precise antibody-targeted biotinylation and efficient affinity purification for all-at-once cell-type proteome capture with sub-micrometer resolution from single tissue slice. With fine-tuned labeling parameters, PSPro shows reliable performance in benchmarking against flow cytometry- and LMD-based proteomic workflows. We apply PSPro to tumor and spleen slices, enriching thousands of proteins containing known markers from ten cell types. We further incorporate LMD into PSPro to facilitate comparison of cell subpopulations from the same tissue slice, revealing spatial proteome heterogeneity of cancer cells and immune cells in pancreatic tumor. Collectively, PSPro converts the traditional "antibody-epitope" paradigm to an "antibody-cell-type proteome" for spatial biology in a user-friendly manner. A record of this paper's transparent peer review process is included in the supplemental information.
    Keywords:  affinity purification-mass spectrometry; laser microdissection; proximity labeling; spatial proteomics; tissue slice
    DOI:  https://doi.org/10.1016/j.cels.2025.101291
  24. Proc Natl Acad Sci U S A. 2025 May 13. 122(19): e2502368122
    Persistent pseudopod splitting is an effective chemotaxis strategy in shallow gradients.
    Albert Alonso, Julius B Kirkegaard, Robert G Endres.
      Single-cell organisms and various cell types use a range of motility modes when following a chemical gradient, but it is unclear which mode is best suited for different gradients. Here, we model directional decision-making in chemotactic amoeboid cells as a stimulus-dependent actin recruitment contest. Pseudopods extending from the cell body compete for a finite actin pool to push the cell in their direction until one pseudopod wins and determines the direction of movement. Our minimal model provides a quantitative understanding of the strategies cells use to reach the physical limit of accurate chemotaxis, aligning with data without explicit gradient sensing or cellular memory for persistence. To generalize our model, we employ reinforcement learning optimization to study the effect of pseudopod suppression, a simple but effective cellular algorithm by which cells can suppress possible directions of movement. Different pseudopod-based chemotaxis strategies emerge naturally depending on the environment and its dynamics. For instance, in static gradients, cells can react faster at the cost of pseudopod accuracy, which is particularly useful in noisy, shallow gradients where it paradoxically increases chemotactic accuracy. In contrast, in dynamics gradients, cells form de novo pseudopods. Overall, our work demonstrates mechanical intelligence for high chemotaxis performance with minimal cellular regulation.
    Keywords:  chemotaxis; mechanical intelligence; physical limits; pseudopods; reinforcement learning
    DOI:  https://doi.org/10.1073/pnas.2502368122
  25. Faraday Discuss. 2025 May 08.
    Protein-induced membrane asymmetry modulates OMP folding kinetics and stability.
    Jonathan M Machin, Neil A Ranson, Sheena E Radford.
      Biological membranes are asymmetric structures, with asymmetry arising from differences in lipid identity in each leaflet of the bilayer, as well as non-uniform distribution of lipids and small molecules in the membrane. Proteins can also induce and modulate membrane asymmetry based on their shape, sequence and interactions with lipids. How membrane asymmetry affects macromolecular behaviour is poorly understood because of the complexity of natural membrane systems, and difficulties in creating relevant asymmetric bilayer systems in vitro. Here, we present a method exploiting the efficient, unidirectional folding of the transmembrane β-barrel outer membrane protein, OmpA, to create asymmetric proteoliposomes with protein-induced dipoles of known direction (arising from sequence variation engineered into the OmpA loops). We then characterise the folding kinetics and stability of different OmpA variants into these proteoliposomes. We find that both the primary sequence of the folding OmpA and the dipole of the membrane into which folding occurs play an important role for modulating the rate of folding. Critically, we find that by complementarily matching the charge on the folding protein to the membrane dipole it is possible to enhance both the folding kinetics and the stability of the folded OmpA. The results hint at how cells might exploit loop charge in membrane-embedded proteins to manipulate membrane environments for adaptation and survival.
    DOI:  https://doi.org/10.1039/d4fd00180j
  26. Cell. 2025 May 05. pii: S0092-8674(25)00455-6. [Epub ahead of print]
    m6A alters ribosome dynamics to initiate mRNA degradation.
    Shino Murakami, Anthony O Olarerin-George, Jianheng Fox Liu, Sara Zaccara, Ben Hawley, Samie R Jaffrey.
      Degradation of mRNA containing N6-methyladenosine (m6A) is essential for cell growth, differentiation, and stress responses. Here, we show that m6A markedly alters ribosome dynamics and that these alterations mediate the degradation effect of m6A on mRNA. We find that m6A is a potent inducer of ribosome stalling, and these stalls lead to ribosome collisions that form a unique conformation unlike those seen in other contexts. We find that the degree of ribosome stalling correlates with m6A-mediated mRNA degradation, and increasing the persistence of collided ribosomes correlates with enhanced m6A-mediated mRNA degradation. Ribosome stalling and collision at m6A is followed by recruitment of YTHDF m6A reader proteins to promote mRNA degradation. We show that mechanisms that reduce ribosome stalling and collisions, such as translation suppression during stress, stabilize m6A-mRNAs and increase their abundance, enabling stress responses. Overall, our study reveals the ribosome as the initial m6A sensor for beginning m6A-mRNA degradation.
    Keywords:  N(6)-methyladenosine; TimeLapse-seq; adaptive response; amino acid depletion; m(6)A; mRNA decay; mRNA degradation; mRNA stability; ribosome collision; ribosome stall
    DOI:  https://doi.org/10.1016/j.cell.2025.04.020
  27. Faraday Discuss. 2025 May 08.
    Lipid-GPCR interactions in an asymmetric plasma membrane model.
    Jingjing Ji, Edward Lyman.
      We report simulations and analysis of the A2A adenosine receptor in its fully active state, in two different membrane environments. The first is a model in which the lipids are distributed asymmetrically according to recent lipidomics, simulations, and biophysical measurements, which together establish the distribution of lipids and cholesterol between the two leaflets. The second is the symmetrized version, which captures the membrane state following loss of lipid asymmetry. By comparing lipid-protein interactions between these two cases we show that solvation by phosphatidyl serine (PS) is insensitive to the loss of asymmetry-an abundance of positively charged sidechains around the cytoplasmic side of the receptor enriches solvation by PS in both membrane states. Cholesterol interactions are sensitive to the loss of asymmetry, with the abundance of cholesterol in the exoplasmic leaflet driving long-lived cholesterol interactions in the asymmetric state. However, one cholesterol interaction site on helix 6 is observed in both cases, and was also observed in earlier work with different membrane models, supporting its identification as a bona fide cholesterol binding site.
    DOI:  https://doi.org/10.1039/d4fd00210e
  28. Faraday Discuss. 2025 May 08.
    Nonequilibrium asymmetry in the living cell membrane.
    Ajay Kumar Bansal, Madan Rao.
      We will discuss how sustained nonequilibrium processes operating at the plasma membrane (PM) determine the dynamical organisation (both lateral and transverse) of lipids, their maintenance and control, under physiological conditions. These nonequilibrium processes include active contractile stresses arising from the inevitable interaction of the inner leaflet of the PM with the adjoining actomyosin cortex, and active flipping of specific lipids. Recently, we showed that the inner leaflet phosphatidylserine (PS) interacts with the actomyosin cortex and engages in a strong transbilayer coupling across the leaflets. Here we develop an active Flory-Huggins theory for the mesoscale segregation of liquid-ordered (lo)-liquid-disordered (ld) domains in an asymmetric membrane bilayer, that incorporates both active contractile stresses at the inner leaflet and transbilayer coupling across the leaflets. The interplay between chemical potential gradients, transbilayer coupling and active stresses drives a rich pattern of mesoscale lo domains - static, strongly fluctuating and moving active emulsions - even at temperatures beyond the equilibrium phase transition temperature. We study conditions under which domain registry and slippage could be observed. We end with a discussion on the role of active flippases on PS in maintaining the active mesoscale organisation.
    DOI:  https://doi.org/10.1039/d4fd00207e
  29. Autophagy. 2025 May 09. 1-5
    Biochemical investigation of LC3/GABARAP-ligand interaction as an important quality measure for LC3/GABARAP-targeting small molecules: addendum to the guidelines (4th edition).
    Martin P Schwalm, Christopher Lenz, Krishna Saxena, Daniel J Klionsky, Ewgenij Proschak, Stefan Knapp.
      Targeted protein degradation (TPD) represents a new therapeutic modality that allows the targeting of proteins that are considered undruggable by conventional small molecules. While TPD approaches via the ubiquitin-proteasome system are well established and validated, additional degradation pathways still require rigorous characterization. Here, we focus on macroautophagy/autophagy tethering compounds, a class of small molecules, designed to recruit cargo to LC3/GABARAP proteins for subsequent autophagosome-dependent degradation. We provide guidance for the biophysical and structural characterization of small molecule modulators for studying LC3/GABARAP-ligand interactions. In addition, we discuss potential limitations of autophagy-based TPD systems and emphasize the need for rigorous quality control in the development of LC3/GABARAP-targeting small molecules.Abbreviations: DSF: differential scanning fluorimetry; FP: fluorescence polarization; FRET: Förster/fluorescence resonance energy transfer; HTRF: homogeneous time-resolved fluorescence; ITC: isothermal titration calorimetry; LIR: LC3-interacting region; MGs: molecular glues; NMR: nuclear magnetic resonance; PROTACs: PROteolysis-TArgeting Chimeras; SPR: surface plasmon resonance; TPD: targeted protein degradation; TR-FRET: time-resolved Förster/fluorescence resonance energy transfer; UPS: ubiquitin-proteasome system.
    Keywords:  ATTECs; AUTACs; Atg8; GABARAP; LC3
    DOI:  https://doi.org/10.1080/15548627.2025.2498506
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