bims-proteo Biomed News
on Proteostasis
Issue of 2025–06–29
fifty papers selected by
Eric Chevet, INSERM
  1. Transferrin receptor controls both autophagosome formation and closure via phosphatidylinositol 3-phosphate synthesis.
  2. PLAA/UFD-3 regulates P-bodies through its intrinsic disordered domain.
  3. Quantitative Proteomic Analysis Reveals JMJD6 and DNAJB11 as Endogenous Substrates of E3 Ligase RFFL.
  4. Structure of fungal tRNA ligase Trl1 with RNA reveals conserved substrate-binding principles.
  5. Harnessing the Evolution of Proteostasis Networks to Reverse Cognitive Dysfunction.
  6. Activation of Unfolded Protein Response Pathway in Malignancies: Interplay with Extracellular Matrix and Targeting Perspectives.
  7. Azoacetylene Electrophiles for Proteome-wide Ligand and Target Discovery: Supporting Targeted Protein Degradation.
  8. HAX1 mediates SARS-CoV-2 spike-triggered unfolded protein response in host cells.
  9. The triad interaction of ULK1, ATG13, and FIP200 is required for ULK complex formation and autophagy.
  10. Recent advances in chemical proteomics for protein profiling and targeted degradation.
  11. Construction of PROTAC-Mediated Ternary Complex Structure Distribution Profiles Using Extensive Conformational Search.
  12. The expanding repertoire of ESCRT functions in cell biology and disease.
  13. Stalled translation on transcripts cleaved by RNase L activates signaling important for innate immunity.
  14. External strain on the plasma membrane is relayed to the endoplasmic reticulum by membrane contact sites and alters cellular energetics.
  15. Sequestration of ribosome biogenesis factors in HSV-1 nuclear aggregates revealed by spatially resolved thermal profiling.
  16. Genetic perturbation of cellular homeostasis regulates Integrated Stress Response signalling to control Drosophila hematopoiesis.
  17. SEL1L-HRD1-mediated ERAD in mammals.
  18. TNIP1 and Autophagy Receptors regulate STING Signaling.
  19. AIMP3 maintains cardiac homeostasis by regulating the editing activity of methionyl-tRNA synthetase.
  20. Global kinetic model of lipid-induced α-synuclein aggregation and its inhibition by small molecules.
  21. Autophagy-targeted NBR1-p62/SQSTM1 complexes promote breast cancer metastasis by sequestering ITCH.
  22. HSP70 is a chaperone for IL-33 activity in chronic airway disease.
  23. A substrate-interacting region of Parkin directs ubiquitination of the mitochondrial GTPase Miro1.
  24. Endoplasmic reticulum stress responses in anticancer immunity.
  25. PDZ-directed substrate recruitment is the primary determinant of specific 4E-BP1 dephosphorylation by PP1-Neurabin.
  26. A host organelle integrates stolen chloroplasts for animal photosynthesis.
  27. Microaggrephagy: an ESCRT-I-PTPN23-dependent pathway for MAPT/tau aggregate clearance.
  28. LRBA deficiency impairs autophagy and contributes to enhanced antigen presentation and T-cell dysregulation.
  29. Acidosis attenuates the hypoxic stabilization of HIF-1α by activating lysosomal degradation.
  30. Unifying Perspectives on the Activity and Genotypic Targeting of Pharmacological Chaperones.
  31. Loss of STIM2, but not of STIM1, drives colorectal cancer metastasis through metabolic reprogramming and the ATF4 ER stress pathway.
  32. A small-molecule VHL molecular glue degrader for cysteine dioxygenase 1.
  33. Design and Semisynthesis of Ubiquitin Extension Probes for Structural Analysis of Cullin1-Mediated Substrate Polyubiquitination.
  34. Enhanced Protein-Protein Interaction Discovery via AlphaFold-Multimer.
  35. VIRMA-mediated m6A modification regulates forebrain formation through modulating ribosome biogenesis.
  36. The Vps13-like protein BLTP2 regulates phosphatidylethanolamine levels to maintain plasma membrane fluidity and breast cancer aggressiveness.
  37. Methods to accelerate PROTAC drug discovery.
  38. Ribosomal RNA transcription regulates splicing through ribosomal protein RPL22.
  39. Visualizing stress granule dynamics with an RNA guanine quadruplex targeted ruthenium(ii) peptide conjugate.
  40. Dietary control of peripheral adipose storage capacity through membrane lipid remodelling.
  41. Isoform-specific function of NSD3 in DNA replication stress confers resistance to PARP inhibitors in prostate cancer.
  42. Sphingomyelins in mosquito saliva reconfigure skin lipidome to promote viral protein levels and enhance transmission of flaviviruses.
  43. Enhancing autophagy by redox regulation extends lifespan in Drosophila.
  44. Mislocalization of nucleic acids is a convergent and targetable mechanism in Alzheimer's disease and frontotemporal dementia.
  45. ChemPerturb-seq screen identifies a small molecule cocktail enhancing human beta cell survival after subcutaneous transplantation.
  46. BAG2 releases SAMD4B upon sensing of arginine deficiency to promote tumor cell survival.
  47. A lysosomal surveillance response to stress extends healthspan.
  48. Profiling antigen-binding affinity of B cell repertoires in tumors by deep learning predicts immune-checkpoint inhibitor treatment outcomes.
  49. The human autophagy-initiating complexes ULK1C and PI3KC3-C1.
  50. Design of combination therapeutics from protein response to drugs in ovarian cancer cells.
  1. Dev Cell. 2025 Jun 13. pii: S1534-5807(25)00327-2. [Epub ahead of print]
    Transferrin receptor controls both autophagosome formation and closure via phosphatidylinositol 3-phosphate synthesis.
    Claudia Puri, So Jung Park, Lidia Wrobel, David C Rubinsztein.
      Autophagosome formation involves multiple sequential steps that need to be coordinated and linked. Here, we describe in mammalian cells that the transferrin receptor (TfR) links LC3 family conjugation to phagophore membranes, an early step in autophagosome biogenesis, with subsequent autophagosome closure. TfR depletion impairs autophagic flux and its overexpression stimulates this catabolic process in an iron-independent manner. TfR is ubiquitinated by the ubiquitin ligase MARCH8 in the RAB11A-LC3B-positive membranes that are conjugated by LC3 family members from which phagophores emanate. Ubiquitinated TfR recruits the VPS34 component VPS15, enabling phosphatidylinositol 3-phosphate (PI(3)P) synthesis on nascent autophagosome membranes. This PI(3)P is not only important for LC3-lipid conjugation but also for subsequent phagophore closure, where TfR-dependent PI(3)P recruits the endosomal sorting complexes required for transport (ESCRT) complex. This TfR activity occurs after endocytosis of iron-containing transferrin, its canonical function, as TfR only binds VPS15 after iron detachment from transferrin that is enabled by pH lowering in the endocytic compartment.
    Keywords:  ESCRT complex; PI(3)P; autophagosome closure; autophagy; transferrin receptor
    DOI:  https://doi.org/10.1016/j.devcel.2025.05.016
  2. Proc Natl Acad Sci U S A. 2025 Jul;122(26): e2427250122
    PLAA/UFD-3 regulates P-bodies through its intrinsic disordered domain.
    Alakananda Das, Yanping Qiu, Trevor J Wolf, Ella Brissett, Jaehyoung Cho, Heenam Park, Eugene C Chen, Tsui-Fen Chou, Paul W Sternberg.
      Regulation of proteome homeostasis is crucial for the survival and adaptation to changing environments for all species. In eukaryotes, this process is finely tuned through regulation at the level of transcription, translation, protein modification, and protein degradation. The phospholipase A2 activating protein (PLAA) is present in all eukaryotes and believed to be a key player in ubiquitin-dependent protein sorting and degradation via its interactions with ubiquitin and/or the AAA+ ATPase, valosin-containing protein (VCP/p97). PLAA's molecular targets and interaction network remain unclear. We used Caenorhabditis elegans and unbiased proteome-scale approaches to investigate neuronal specific interactors of the C. elegans PLAA ortholog UFD-3 (ubiquitin fusion degradation 3), its effect on ubiquitinated proteins, and global protein expression changes in an ufd-3 mutant. We found that PLAA may play a unique role in cytoplasmic messenger ribonucleic acid (mRNA) processing bodies (P-bodies). Using biochemical analysis in vitro and fluorescence imaging in C. elegans, we show that UFD-3 directly interacts with the mRNA decapping complex regulatory subunit DCAP-1. UFD-3's intrinsic disordered region (IDR), which contains conserved amino acid motifs, is important for the recruitment of DCAP-1 to P-bodies. Finally, we show that loss of the IDR does not affect UFD-3's role in sorting ubiquitinated proteins through the multivesicular body pathway. Collectively, our results suggest that UFD-3's role in P-bodies is distinct from its role in the ubiquitin-dependent protein degradation pathway and the IDR is only critical for UFD-3-regulated P-bodies pathways. Thus, PLAA/UFD-3 might regulate the proteome via two distinct pathways: ubiquitinated protein turnover, as well as mRNA regulation through P-bodies.
    Keywords:  intrinsically disordered reg; mating behavior; protein localization; proteomics; proximity labeling
    DOI:  https://doi.org/10.1073/pnas.2427250122
  3. J Proteome Res. 2025 Jun 26.
    Quantitative Proteomic Analysis Reveals JMJD6 and DNAJB11 as Endogenous Substrates of E3 Ligase RFFL.
    Nikhil Dev Narendradev, Soumitra Marathe, Sabyasachi Baboo, Daniel B McClatchy, Jolene K Diedrich, Parul Jain, Rahul Purwar, John R Yates, Srinivasa Murty Srinivasula.
      The ubiquitin-proteasome system contributes to protein quality control, involving E3 ligases that ubiquitinate proteins and leading to their degradation. The dysregulation of protein degradation results in the abnormal accumulation of proteins and is implicated in the pathology of diverse diseases, making targeted protein degradation a promising therapeutic strategy. Here, we focus on RFFL, an endosome-associated RING E3 ligase involved in mitochondrial homeostasis and the clearance of misfolded cystic fibrosis transmembrane conductance regulator proteins. Using label-free quantitative mass spectrometry based proteomics for interactome and differential expression analyses, we systematically investigated and identified putative substrates of RFFL. For more confident identification, we performed these analyses on three cell lines that we generated: an RFFL knockout cell line generated using CRISPR/Cas9, another cell line rescuing RFFL expression when complemented with KO cells with stably expressing RFFL cDNA, and wild-type cells. We validated JMJD6 and DNAJB11 as substrates of endogenous RFFL, providing orthogonal validation and confidence in our screening approach. We demonstrated that RFFL ubiquitinates and degrades JMJD6 and DNAJB11 via the proteasomal pathway using in vivo assays. Interestingly, we also discovered a hitherto unknown role of RFFL in lipid metabolism. Collectively, this study provides the first comprehensive and unbiased analysis of RFFL substrates employing multiple complementary approaches.
    Keywords:  E3 ligase; RFFL; interactome; proteomics; substrates
    DOI:  https://doi.org/10.1021/acs.jproteome.5c00086
  4. Nat Struct Mol Biol. 2025 Jun 25.
    Structure of fungal tRNA ligase Trl1 with RNA reveals conserved substrate-binding principles.
    Sandra Köhler, Jürgen Kopp, Satyabrata Maiti, Janusz M Bujnicki, Jirka Peschek.
      RNA ligases play a vital role in RNA processing and maturation, including tRNA splicing, RNA repair and the unfolded protein response (UPR). In fungi and plants, the tripartite tRNA ligase Trl1 catalyzes the joining of TSEN-cleaved pre-tRNA exon halves. Trl1 also functions as ligase in the non-conventional HAC1 mRNA splicing during the UPR. The final ligation step is performed by the N-terminal adenylyltransferase domain (ligase; LIG). The spatial arrangement of the exon ends during the ligation reaction has remained elusive. Here we report the crystal structure of Chaetomium thermophilum Trl1-LIG in complex with a tRNA-derived substrate. Our structure represents a snapshot of the activated RNA intermediate and defines the conserved substrate-binding interface. The underlying enzyme-substrate interplay reveals a substrate-binding principle shared by adenylyltransferases. Moreover, we identify the determinants of RNA end specificity as well as the specific roles of Trl1-LIG's subdomains during ligase activation, substrate binding and phosphoryl transfer.
    DOI:  https://doi.org/10.1038/s41594-025-01589-3
  5. bioRxiv. 2025 Mar 02. pii: 2025.02.28.640897. [Epub ahead of print]
    Harnessing the Evolution of Proteostasis Networks to Reverse Cognitive Dysfunction.
    Lucas C Reineke, Ping Jun Zhu, Udit Dalwadi, Sean W Dooling, Yuwei Liu, I-Ching Wang, Sara Young-Baird, James Okoh, Santosh Kumar Kuncha, Hongyi Zhou, Akshara Kannan, Hyekyung Park, Nicolas A Debeaubien, Tristan Croll, D John Lee, Christopher Arthur, Thomas E Dever, Peter Walter, Jin Chen, Adam Frost, Mauro Costa-Mattioli.
      The integrated stress response (ISR) is a highly conserved network essential for maintaining cellular homeostasis and cognitive function. Here, we investigated how persistent ISR activation impacts cognitive performance, primarily focusing on a PPP1R15B R658C genetic variant associated with intellectual disability. By generating a novel mouse model that mimics this human condition, we revealed that this variant destabilizes the PPP1R15B•PP1 phosphatase complex, resulting in chronic ISR activation, impaired protein synthesis, and deficits in long-term memory. Importantly, we found that the cognitive and synaptic deficits in Ppp1r15b R658C mice are directly due to ISR activation. Leveraging insights from evolutionary biology, we characterized DP71L, a viral orthologue of PPP1R15B, through detailed molecular and structural analyses, uncovering its mechanism of action as a potent pan-ISR inhibitor. Remarkably, we found that DP71L not only buffers cognitive decline associated with a wide array of conditions-including Down syndrome, Alzheimer's disease and aging-but also enhances long-term synaptic plasticity and memory in healthy mice. These findings highlight the promise of utilizing evolutionary insight to inform innovative therapeutic strategies.
    DOI:  https://doi.org/10.1101/2025.02.28.640897
  6. Cancers (Basel). 2025 Jun 13. pii: 1972. [Epub ahead of print]17(12):
    Activation of Unfolded Protein Response Pathway in Malignancies: Interplay with Extracellular Matrix and Targeting Perspectives.
    Eleftherios N Athanasopoulos, Angeliki Natsiou, Maria Kyriazopoulou, Dimitra Manou, Achilleas D Theocharis, Vassiliki T Labropoulou.
      Malignant cells exhibit elevated rates of protein synthesis and secretion to facilitate tumor growth, proliferation, and tumorigenesis. Upon malignant transformation, the endoplasmic reticulum (ER) experiences stress due to the accumulation of unfolded or misfolded proteins in the ER lumen, lack of nutrient availability and overall hostile tumor microenvironment conditions. The demand for regulated protein turnover and proteostasis reinstatement results in the activation of the unfolded protein response (UPR) pathway for cellular adaptation and survival. The UPR machinery utilizes the BiP chaperone and three ER-bound sensors, PERK, IRE1, and ATF6, to substantiate signal transduction and orchestrate gene expression associated with protein folding, degradation and recycling, inflammation, autophagy, and programmed cell death. The pleiotropic function of UPR emerges as a central mediator for tumor progression, especially in multiple myeloma and glioblastoma pathologies. Numerous studies have recently pointed out that communication of the extracellular matrix (ECM) with surrounding tumor cells dictates in part UPR activity and vice versa. In the context of this dynamic interplay, ER stress and UPR mechanisms have been proposed as potential targets to elicit novel and effective therapeutic approaches in clinical trials.
    Keywords:  endoplasmic reticulum stress; extracellular matrix; glioblastoma; multiple myeloma; signaling; unfolded protein response
    DOI:  https://doi.org/10.3390/cancers17121972
  7. J Med Chem. 2025 Jun 25.
    Azoacetylene Electrophiles for Proteome-wide Ligand and Target Discovery: Supporting Targeted Protein Degradation.
    Jiayi Du, Shengrong Li, Fang Xu, Zehong Lin, Tongyi Hong, Yifang Li, Zeyi Deng, Yi Tan, Zhengqiu Li.
      Covalent probes integrated with chemical proteomics have been an efficient method for disclosing new druggable targets and E3 ubiquitin ligases supporting targeted protein degradation. However, a large fraction of the proteome including E3 ligases remains inaccessible with existing electrophiles. In this work, we developed a new reactive warhead, terminal azoacetylene, which can be generated by in situ desilylation for proteome profiling under cellular conditions. A series of uncharacterized targets and E3 ubiquitin ligases were covalently engaged. Fragment-based ligand discovery (FBLD) showed that the azoacetylene-containing fragments can covalently bind a series of essential protein hits at the active sites such as C130 of TUFM probably modulating the protein functions. Incorporation of this warhead into BRD4 targeting inhibitor JQ1 led to generation of novel small molecular degraders that degrade BRD4 without inducing the hook effect. This provides a new method for ligand and target discovery, as well as the development of new types of small molecular degraders.
    DOI:  https://doi.org/10.1021/acs.jmedchem.5c00229
  8. FEBS J. 2025 Jun 26.
    HAX1 mediates SARS-CoV-2 spike-triggered unfolded protein response in host cells.
    Fang Zhu, Xiangpeng Sheng, Fan Yang, Xuechen Wang, Cong Yang, Jin Ren, Chengcheng Wang, Ronggui Hu.
      SARS-CoV-2 continues to evolve with enhanced transmissibility, a feature primarily mediated by its spike (S) protein. While expression of the S protein in human cells can induce the accumulation of reactive oxygen species (ROS), the regulatory mechanisms governing this process remain poorly understood. Here, we identify the human protein HCLS1-associated protein X-1 (HAX1) as a key regulator that mitigates SARS-CoV-2S-induced ROS accumulation. A genome-wide screen revealed HAX1 as a binding partner of the SARS-CoV-2S protein in mammalian cells. HAX1 specifically interacts with the S1 subunit of S, and its deficiency effectively abolishes S-induced activation of endoplasmic reticulum (ER) stress responses, including the unfolded protein response (UPR). Notably, HAX1-dependent UPR activation is unique to SARS-CoV-2S and certain variants and is not triggered by other UPR inducers. Loss of HAX1 markedly exacerbates SARS-CoV-2S-induced ROS accumulation and mitochondrial dysfunction. Collectively, our findings uncover a previously unrecognized mechanism by which S modulates host stress responses and establish HAX1 as a host factor involved in SARS-CoV-2-related processes.
    Keywords:  ER stress; HAX1; SARS‐CoV‐2; Spike; unfolded protein response
    DOI:  https://doi.org/10.1111/febs.70163
  9. Elife. 2025 06 24. pii: RP101531. [Epub ahead of print]13
    The triad interaction of ULK1, ATG13, and FIP200 is required for ULK complex formation and autophagy.
    Yutaro Hama, Yuko Fujioka, Hayashi Yamamoto, Noboru Mizushima, Nobuo N Noda.
      In mammals, autophagosome formation, a central event in autophagy, is initiated by the ULK complex comprising ULK1/2, FIP200, ATG13, and ATG101. However, the structural basis and mechanism underlying the ULK complex assembly have yet to be fully clarified. Here, we predicted the core interactions organizing the ULK complex using AlphaFold, which proposed that the intrinsically disordered region of ATG13 engages the bases of the two UBL domains in the FIP200 dimer via two phenylalanines and also binds the tandem microtubule-interacting and transport domain of ULK1, thereby yielding the 1:1:2 stoichiometry of the ULK1-ATG13-FIP200 complex. We validated the predicted interactions by point mutations and demonstrated direct triad interactions among ULK1, ATG13, and FIP200 in vitro and in cells, wherein each interaction was additively important for autophagic flux. These results indicate that the ULK1-ATG13-FIP200 triadic interaction is crucial for autophagosome formation and provides a structural basis and insights into the regulation mechanism of autophagy initiation in mammals.
    Keywords:  ATG13; AlphaFold; FIP200; ULK complex; autophagy; cell biology; human
    DOI:  https://doi.org/10.7554/eLife.101531
  10. Curr Opin Chem Biol. 2025 Jun 26. pii: S1367-5931(25)00037-7. [Epub ahead of print]87 102605
    Recent advances in chemical proteomics for protein profiling and targeted degradation.
    Peng-Kai Liu, Zicong Wang, Lingjun Li.
      Chemical proteomics has emerged as a powerful approach to decipher protein function, interactions, and targeted degradation pathways in complex biological systems. Recent advances in chemical labeling strategies, including activity-based protein profiling (ABPP), proximity labeling (PL), and proteolysis-targeting chimeras (PROTACs), have facilitated a deeper understanding of protein function and interaction networks. First, ABPP employs covalent probes to selectively label active enzymes, uncovering functional proteomics and drug-target interactions. Innovations such as PhosID-ABPP and streamlined cysteine ABPP have improved site-specific quantification and throughput, enabling proteome-wide analysis of enzyme activity and small-molecule interactions. Second, PL enables the characterization of transient protein-protein interactions using enzymatic or chemically triggered approaches. Advances including TurboID and TransitID enhanced the spatiotemporal resolution of PL. Third, PROTACs expand the scope of targeted protein degradation by leveraging the ubiquitin-proteasome system. Collectively, we highlight recent advancements in integrating mass spectrometry (MS) with these methodologies in the field of chemical proteomics.
    Keywords:  Activity-based protein profiling (ABPP); Chemical proteomics; Drug discovery; Mass spectrometry; Protein–protein interactions; Proteolysis-targeting chimeras (PROTACs); Proximity labeling (PL)
    DOI:  https://doi.org/10.1016/j.cbpa.2025.102605
  11. J Chem Inf Model. 2025 Jun 23.
    Construction of PROTAC-Mediated Ternary Complex Structure Distribution Profiles Using Extensive Conformational Search.
    Genki Kudo, Takumi Hirao, Ryuhei Harada, Yasuteru Shigeta, Takatsugu Hirokawa, Ryunosuke Yoshino.
      Proteolysis-targeting chimeras (PROTACs) are heterobifunctional small molecules that recruit E3 ubiquitin ligases to a target protein and induce its ubiquitination by forming a ternary complex. However, the structural dynamics underlying the complex formation and degradation efficiency remain unclear. In this study, we attempted an extensive conformational search using the parallel cascade selection molecular dynamics (PaCS-MD) and outlier flooding (OFLOOD) method for PROTACs differing in linker length. Markov state models revealed that while all PROTACs share a common low free-energy state, their structural distribution profiles differ significantly. These results suggest that linker-dependent conformational distribution profiles modulate degradation activity and cooperativity, offering mechanistic insights into rational PROTAC design.
    DOI:  https://doi.org/10.1021/acs.jcim.5c00102
  12. Nature. 2025 Jun 25.
    The expanding repertoire of ESCRT functions in cell biology and disease.
    James H Hurley, Alyssa N Coyne, Marta Miączyńska, Harald Stenmark.
      The endosomal sorting complex required for transport (ESCRT) is a multicomplex machinery comprising proteins that are conserved from bacteria to humans and has diverse roles in regulating the dynamics of cellular membranes. ESCRT functions have far-reaching consequences for cell biological processes such as intracellular traffic, membrane repair, cell signalling, metabolic regulation, cell division and genome maintenance. Here we review recent insights that emphasize the pathophysiological consequences of ESCRT dysfunctions, including infections, immune disorders, cancers and neurological diseases. We highlight the possibilities of using our knowledge about ESCRT structures and functions for drug discovery.
    DOI:  https://doi.org/10.1038/s41586-025-08950-y
  13. bioRxiv. 2025 Jun 11. pii: 2025.06.10.658914. [Epub ahead of print]
    Stalled translation on transcripts cleaved by RNase L activates signaling important for innate immunity.
    Agnes Karasik, Grant D Jones, Nicholas R Guydosh.
      RNase L is an endonuclease that responds to infections by cleaving most host- and pathogen-derived single-stranded RNAs. This widespread RNA cleavage can lead to death of the infected cell via the ribotoxic stress response (RSR). An ongoing challenge is to understand how RNase L's endonuclease activity triggers cell death to benefit the host. To address this question, we used nanopore-based long-read sequencing to show that 3' mRNA fragments in the cell were not fully degraded after RNase L activation and that these fragments were translated by ribosomes. We further asked whether ribosomes on mRNA fragments stall when they reach 3' ends created by RNase L. We used ribosome profiling to capture footprints protected by these ribosomes, which can be identified by their short length (15-18 nt). We found that RNase L activation increased the number of stalled ribosomes at RNase L cleavage sites. Loss of the ribosome rescue factor PELO increased the number of short footprints derived from stalled ribosomes and augmented the RSR. Our work therefore establishes a role for fragmented mRNA in causing ribosome stalling that promotes innate immunity via the RSR.
    Highlights: Activation of RNase L leads to accumulation and translation of mRNA fragmentsRibosomes stall at the 3' end of the RNase L cleaved mRNA fragmentsPELO rescues ribosomes stalled due to RNase L activation.
    DOI:  https://doi.org/10.1101/2025.06.10.658914
  14. Sci Adv. 2025 Jun 27. 11(26): eads6132
    External strain on the plasma membrane is relayed to the endoplasmic reticulum by membrane contact sites and alters cellular energetics.
    Ziming Chen, Peilin Chen, Jiayue Li, Euphemie Landao-Bassonga, John Papadimitriou, Junjie Gao, Delin Liu, Andrew Tai, Jinjin Ma, David Lloyd, Brendan F Kennedy, Ming Hao Zheng.
      Mechanotransduction is essential for living cells to adapt to their extracellular environment. However, it is unclear how the biophysical adaptation of intracellular organelles responds to mechanical stress or how these adaptive changes affect cellular homeostasis. Here, using the tendon cell as a mechanosensitive cell type within a bioreactor, we show that the tension of the plasma membrane (PM) and the endoplasmic reticulum (ER) adaptively increases in response to repetitive external stimuli. Depletion of stromal interaction molecule 1 (STIM1), the highest expressed PM-ER tether protein, interfered with mechanotransduction from the PM to the ER, and affected the ER tension. We found that an optimized mechanical strain increased ER tension in a homeostatic manner, but excessive strain resulted in ER expansion, as well as activating ER stress. Last, we showed that changes in ER tension were linked with ER-mitochondria interactions and associated with cellular energetics and function. Together, these findings identify a PM-ER mechanotransduction mechanism that dose-dependently regulates cellular metabolism.
    DOI:  https://doi.org/10.1126/sciadv.ads6132
  15. Sci Adv. 2025 Jun 27. 11(26): eadw6814
    Sequestration of ribosome biogenesis factors in HSV-1 nuclear aggregates revealed by spatially resolved thermal profiling.
    Peter J Metzger, Tavis J Reed, Krystal K Lum, Jordy F Botello, Lifei Jiang, Clifford P Brangwynne, Olga G Troyanskaya, Ileana M Cristea.
      Viruses exploit host cell reliance on compartmentalization to facilitate their replication. Herpes simplex virus type 1 (HSV-1) modulates the subcellular localization of host proteins to suppress immune activation, license viral gene expression, and achieve translational shutoff. To spatially resolve dynamic protein-protein interaction (PPI) networks during infection with an immunostimulatory HSV-1 strain, we integrated nuclear/cytoplasmic fractionation with thermal proximity coaggregation analysis (N/C-TPCA). The resulting expanded depth and spatial resolution of PPIs charted compartment-specific assemblies of protein complexes throughout infection. We find that a broader suite of host chaperones than previously anticipated exhibits nuclear recruitment to form condensates known as virus-induced chaperone-enriched (VICE) domains. Monitoring protein and RNA constituents and ribosome activity, we establish that VICE domains sequester ribosome biogenesis factors from ribosomal RNA, accompanying a cell-wide defect in ribosome supply. These findings highlight infection-driven VICE domains as nodes of translational remodeling and demonstrate the utility of N/C-TPCA to study dynamic biological contexts.
    DOI:  https://doi.org/10.1126/sciadv.adw6814
  16. Biol Open. 2025 Jun 17. pii: bio.062046. [Epub ahead of print]
    Genetic perturbation of cellular homeostasis regulates Integrated Stress Response signalling to control Drosophila hematopoiesis.
    Kishalay Ghosh, Rohit Krishnan Iyer, Saloni Sood, Mohamed Sabeelil Islam, Jyotsana G Labad, Rohan Jayant Khadilkar.
      Aging results in a decline in cellular and molecular functions. One of the hallmarks of aging is stem cell exhaustion which impacts self-renewal and differentiation. We employ the Drosophila larval lymph gland (LG) to investigate the impact of genetic perturbation of cellular homeostasis on hematopoiesis. LG consists of a Posterior Signalling Center (PSC) - a stem cell niche that maintains Medullary Zone (MZ) prohemocytes whereas Cortical Zone (CZ) consists of differentiated hemocytes. We employ over-activation of Toll or Imd pathway for disrupting cellular homeostasis whereas over-express Foxo or Atg8 for balancing it. Genetic perturbation of cellular homeostasis displays hallmarks of aging. Induction of Toll or Imd pathway locally and systemically leads to a decreased niche size and increased differentiation whereas Foxo or Atg8 over-expression shows an opposite trend. We show that the Integrated Stress Response (ISR) pathway is induced upon Toll or Imd over-activation and LG's with ISR perturbation show increased hemocyte differentiation. Genetic epistasis shows that ectopic over-expression of ISR components upon Imd activation can rescue hematopoietic defects. Overall, our study explores how genetic perturbation of cellular homeostasis can impact hematopoiesis. Our research has implications in understanding how abrogation of cellular homeostatic mechanisms may lead to onset of malignancies.
    Keywords:   Drosophila ; Hematopoiesis; Homeostasis; Signalling; Stem cells
    DOI:  https://doi.org/10.1242/bio.062046
  17. Nat Cell Biol. 2025 Jun 25.
    SEL1L-HRD1-mediated ERAD in mammals.
    Huilun Helen Wang, Ida Biunno, Shengyi Sun, Ling Qi.
      Endoplasmic reticulum-associated degradation (ERAD) is a critical quality control mechanism responsible for eliminating misfolded or unassembled proteins. It maintains endoplasmic reticulum homeostasis, ensures a proper folding environment and regulates substrate protein levels. Following its discovery in the late 1980s and early 1990s, research on ERAD in mammals-particularly that mediated by the conserved protein complex comprising suppressor/enhancer of Lin-12-like protein 1-like (SEL1L) and HMG-CoA reductase degradation protein 1 (HRD1)-has advanced substantially over the past decade. SEL1L-HRD1-mediated ERAD is now recognized as a fundamental process in mammals that governs various physiological functions largely in a substrate-specific manner. In humans, mutations in this complex have been causally linked to ERAD-associated neurodevelopmental disorders with onset in infancy (ENDI) and ENDI-agammaglobulinaemia. This Review highlights the SEL1L-HRD1-mediated ERAD pathway, exploring its machinery, molecular mechanism and physiological relevance and potential therapeutic strategies targeting this system.
    DOI:  https://doi.org/10.1038/s41556-025-01690-1
  18. bioRxiv. 2025 Apr 23. pii: 2025.04.21.649822. [Epub ahead of print]
    TNIP1 and Autophagy Receptors regulate STING Signaling.
    Eric N Bunker, Tara D Fischer, Peng-Peng Zhu, François Le Guerroué, Richard J Youle.
      Activation of the cGAS-STING pathway stimulates innate immune signaling as well as LC3B lipidation and ubiquitylation at Golgi-related vesicles upon STING trafficking. Although ubiquitylation at these subcellular sites has been associated with regulating NF-κB-related innate immune signaling, the mechanisms of Golgi-localized polyubiquitin chain regulation of immune signaling is not well understood. We report here that the ubiquitin- and LC3B-binding proteins, TNIP1 and autophagy receptors p62, NBR1, NDP52, TAX1BP1, and OPTN associate with STING-induced ubiquitin and LC3B-labeled vesicles, and that p62 and NBR1 act redundantly in spatial clustering of the LC3B-labeled vesicles in the perinuclear region. We also find that while TBK1 kinase activity is not required for the recruitment of TNIP1 and the autophagy receptors, it also plays a role in sequestration of the LC3B-labeled vesicles. The ubiquitin binding domains, rather than the LC3B-interacting regions, of TNIP1 and OPTN are specifically important for their recruitment to Ub/LC3B-associated perinuclear vesicles, while OPTN is also recruited through a TBK1-dependent mechanism. Functionally, we find that TNIP1 and OPTN play a role in STING-mediated innate immune signaling, with TNIP1 acting as a significant negative regulator of both NF-κB- and Interferon-mediated gene expression. Together, these results highlight autophagy-independent mechanisms of autophagy receptors and TNIP1 with unanticipated roles in regulating STING-mediated innate immunity.
    DOI:  https://doi.org/10.1101/2025.04.21.649822
  19. Nat Cardiovasc Res. 2025 Jun 25.
    AIMP3 maintains cardiac homeostasis by regulating the editing activity of methionyl-tRNA synthetase.
    Anindhya S Das, Charles P Rabolli, Colton R Martens, Han-Kai Jiang, Yingshen Zhang, Aubree A Zimmer, Kevin Lin, Kedryn K Baskin, Juan D Alfonzo, Federica Accornero.
      In mammals, nine aminoacyl tRNA synthetases (ARSs) and three auxiliary proteins (ARS-interacting multifunctional proteins 1-3 (AIMP1-3)) form the multisynthetase complex (MSC), a molecular hub that provides a subset of aminoacylated tRNAs to the ribosome and partakes in translation-independent signaling. Knowledge of the role of AIMPs in organ physiology is currently limited. AIMP3 (also known as EEF1E1) was proposed to anchor methionyl tRNA synthetase (MetRS) in the complex and regulate protein synthesis through translation initiation and elongation. Here we show that a cardiomyocyte-specific conditional knockout of AIMP3 in mice leads to lethal cardiomyopathy. MetRS localization, aminoacylation efficiency and global protein synthesis were unaffected in our model, suggesting an alternative mechanism for the pathology. We found that AIMP3 is essential for homocysteine editing by MetRS, a reaction that is necessary for the maintenance of translation fidelity. Homocysteine accumulation induced reactive oxygen species production, protein aggregation, mitochondrial dysfunction, autophagy and ultimately cell death.
    DOI:  https://doi.org/10.1038/s44161-025-00670-w
  20. Proc Natl Acad Sci U S A. 2025 Jul;122(26): e2422427122
    Global kinetic model of lipid-induced α-synuclein aggregation and its inhibition by small molecules.
    Alisdair Stevenson, Roxine Staats, Alexander J Dear, David Voderholzer, Jesper E Dreier, Georg Meisl, Raphael Guido, Tuomas P J Knowles, Céline Galvagnion, Alexander K Buell, Michele Vendruscolo, Thomas C T Michaels.
      The aggregation of α-synuclein into amyloid fibrils is a hallmark of Parkinson's disease. This process has been shown to directly involve interactions between proteins and lipid surfaces when the latter are present. Despite this importance, the molecular mechanisms of lipid-induced amyloid aggregation have remained largely elusive. Here, we present a global kinetic model to describe lipid-induced amyloid aggregation of α-synuclein. Using this framework, we find that α-synuclein fibrils form via a two-step primary nucleation mechanism and that lipid molecules are directly involved in both the nucleation and fibril elongation steps, giving rise to lipid-protein coaggregates. To illustrate the applicability of this kinetic approach to drug discovery, we identify the mechanism of action of squalamine, a known inhibitor of lipid-induced α-synuclein aggregation, revealing that this small molecule reduces the rate of lipid-dependent primary nucleation. Our work will likely guide the rational design of α-synuclein aggregation inhibitors.
    Keywords:  Parkinson’s disease; aggregation inhibitors; lipid-induced amyloid aggregation
    DOI:  https://doi.org/10.1073/pnas.2422427122
  21. Nat Cell Biol. 2025 Jun 27.
    Autophagy-targeted NBR1-p62/SQSTM1 complexes promote breast cancer metastasis by sequestering ITCH.
    Gourish Mondal, Hugo Gonzalez, Timothy Marsh, Andrew M Leidal, Ariadne Vlahakis, Pravin R Phadatare, Sofı́a Bustamante Eguiguren, Michael Bruck, Akul Naik, Mark Jesus M Magbanua, Laura A Huppert, Arun P Wiita, Jeroen P Roose, Jennifer M Rosenbluth, Jayanta Debnath.
      Autophagy deficiency in breast cancer promotes metastasis through the accumulation of the autophagy cargo receptor NBR1. Here we show that autophagy normally suppresses breast cancer metastasis by enabling the clearance of NBR1-p62/SQSTM1 complexes that instruct p63-mediated pro-metastatic basal differentiation programmes. When autophagy is inhibited, the autophagy cargo receptors NBR1 and p62/SQSTM1 accumulate within biomolecular condensates in cells, which drives basal differentiation in both mouse and human breast cancer models. Mechanistically, these NBR1-p62/SQSTM1 complexes sequester ITCH, a ubiquitin ligase that degrades and negatively regulates p63 in breast cancer cells, thereby stabilizing and activating p63. Accordingly, mutant forms of NBR1 unable to sequester ITCH into NBR1-p62/SQSTM1 complexes do not promote basal differentiation and metastasis in vivo. Overall, our findings illuminate how proteostatic defects arising in the setting of therapeutic autophagy inhibition modulate epithelial lineage fidelity and metastatic progression.
    DOI:  https://doi.org/10.1038/s41556-025-01689-8
  22. JCI Insight. 2025 Jun 24. pii: e193640. [Epub ahead of print]
    HSP70 is a chaperone for IL-33 activity in chronic airway disease.
    Omar A Osorio, Heather E Raphael, Colin E Kluender, Ghandi F Hassan, Lucy S Cohen, Deborah F Steinberg, Ella Katz-Kiriakos, Morgan D Payne, Ethan M Luo, Jamie L Hicks, Derek E Byers, Jennifer Alexander-Brett.
      IL-33 is a key driver of type-2 inflammation and implicated in pathology of COPD and asthma. However, the mechanism for IL-33 secretion and regulation in the context of chronic airway disease is poorly understood. We previously reported an airway disease-associated isoform IL-33Δ34 that escapes nuclear sequestration and is tonically secreted from epithelial cells. Here, we describe how this IL-33Δ34 isoform interacts with HSP70 within cells and is targeted to secretory organelles through coordinated binding to phosphatidylserine (PS), and delivered to compartments for unconventional protein secretion (CUPS). Once secreted, extracellular HSP70 (eHSP70) in complex with IL-33Δ34 stabilizes cytokine by inhibiting oxidation and degradation, which results in enhanced IL-33Δ34-receptor binding and activity. We further find evidence that IL-33 along with mediators of the proteostasis network HSP70, HSP90 and the Chaperonin Containing TCP1 (CCT) complex are dysregulated in human chronic airway disease. This phenomenon is reflected in the differential extracellular vesicle (EV) proteome in bronchial wash from COPD and asthma samples, which could mark disease activity and potentiate IL-33 function. This study confirms proteostasis intermediates, chiefly HSP70, as a chaperone for non-canonical IL-33 secretion and activity that may be amenable for therapeutic targeting in the chronic airway diseases COPD and asthma.
    Keywords:  COPD; Chaperones; Cytokines; Immunology; Pulmonology
    DOI:  https://doi.org/10.1172/jci.insight.193640
  23. J Cell Biol. 2025 Aug 04. pii: e202408025. [Epub ahead of print]224(8):
    A substrate-interacting region of Parkin directs ubiquitination of the mitochondrial GTPase Miro1.
    Joanna Koszela, Anne Rintala-Dempsey, Giulia Salzano, Viveka Pimenta, Outi Kamarainen, Mads Gabrielsen, Aasna L Parui, Gary S Shaw, Helen Walden.
      Mutations in the E3 ubiquitin ligase Parkin gene have been linked to early onset Parkinson's disease. Besides many other roles, Parkin is involved in clearance of damaged mitochondria via mitophagy-a process of particular importance in dopaminergic neurons. Upon mitochondrial damage, Parkin accumulates at the outer mitochondrial membrane and is activated, leading to ubiquitination of many mitochondrial substrates and recruitment of mitophagy effectors. While the activation mechanisms of autoinhibited Parkin have been extensively studied, it remains unknown how Parkin recognizes its substrates for ubiquitination. Here, we characterize a conserved region in the flexible linker between the Ubl and RING0 domains of Parkin, which is indispensable for Parkin interaction with the mitochondrial GTPase Miro1. Our results may explain fast kinetics of Miro1 ubiquitination by Parkin in recombinant assays and provide a biochemical explanation for Miro1-dependent Parkin recruitment to the mitochondrial membrane observed in cells. Our findings are important for understanding mitochondrial homeostasis and may inspire new therapeutic avenues for Parkinson's disease.
    DOI:  https://doi.org/10.1083/jcb.202408025
  24. Nat Rev Cancer. 2025 Jun 24.
    Endoplasmic reticulum stress responses in anticancer immunity.
    Sung-Min Hwang, Shiun Chang, Paulo C Rodriguez, Juan R Cubillos-Ruiz.
      The endoplasmic reticulum (ER) has a central role in processes essential for mounting effective and durable antitumour immunity; this includes regulating protein synthesis, folding, modification and trafficking in immune cells. However, the tumour microenvironment imposes hostile conditions that disrupt ER homeostasis in both malignant and infiltrating immune cells, leading to chronic activation of the unfolded protein response (UPR). Dysregulated ER stress responses have emerged as critical modulators of cancer progression and immune escape, influencing the initiation, development and maintenance of antitumour immunity. In this Review, we examine how tumour-induced ER stress reshapes the functional landscape of immune cells within the tumour microenvironment. We highlight recent discoveries demonstrating how ER stress curtails endogenous antitumour immunity and reduces the efficacy of immunotherapies. Furthermore, we underscore novel therapeutic strategies targeting ER stress sensors or UPR components to restore immune function and enhance cancer immunotherapy outcomes. Together, this provides a comprehensive overview of the interplay between ER stress responses and antitumour immunity, emphasizing the potential of UPR-targeted interventions to improve immune control of cancer.
    DOI:  https://doi.org/10.1038/s41568-025-00836-5
  25. Elife. 2025 Jun 23. pii: RP103403. [Epub ahead of print]13
    PDZ-directed substrate recruitment is the primary determinant of specific 4E-BP1 dephosphorylation by PP1-Neurabin.
    Roman O Fedoryshchak, Karim El-Bouri, Dhira Joshi, Stephane Mouilleron, Richard Treisman.
      Phosphoprotein phosphatase 1 (PP1) relies on association with PP1-interacting proteins (PIPs) to generate substrate-specific PIP/PP1 holoenzymes, but the lack of well-defined substrates has hindered elucidation of the mechanisms involved. We previously demonstrated that the Phactr1 PIP confers sequence specificity on the Phactr1/PP1 holoenzyme by remodelling the PP1 hydrophobic substrate groove. Phactr1 defines a group of 'RVxF-ΦΦ-R-W' PIPs that all interact with PP1 in a similar fashion. Here, we use a PP1-PIP fusion approach to address sequence specificity and identify substrates of the RVxF-ΦΦ-R-W family PIPs. We show that the four Phactr proteins confer identical sequence specificities on their holoenzymes. We identify the 4E-BP and p70 S6K translational regulators as substrates for the Neurabin/Spinophilin PIPs, implicated in neuronal plasticity, pointing to a role for their holoenzymes in mTORC1-dependent translational control. Biochemical and structural experiments show that in contrast to the Phactrs, substrate recruitment and catalytic efficiency of the PP1-Neurabin and PP1-Spinophilin fusions is primarily determined by substrate interaction with the PDZ domain adjoining their RVxF-ΦΦ-R-W motifs, rather than by recognition of the remodelled PP1 hydrophobic groove. Thus, even PIPs that interact with PP1 in a similar manner use different mechanisms to ensure substrate selectivity.
    Keywords:  4E-BP1; Neurabin; PIP; Phactr; biochemistry; chemical biology; human; mTORC1; molecular biophysics; phosphatase; structural biology
    DOI:  https://doi.org/10.7554/eLife.103403
  26. Cell. 2025 Jun 21. pii: S0092-8674(25)00637-3. [Epub ahead of print]
    A host organelle integrates stolen chloroplasts for animal photosynthesis.
    Corey A H Allard, Angus B Thies, Rishav Mitra, Patric M Vaelli, Olivia D Leto, Brittany L Walsh, Elise M J Laetz, Martin Tresguerres, Amy S Y Lee, Nicholas W Bellono.
      Eukaryotic life evolved over a billion years ago when ancient cells engulfed and integrated prokaryotes to become modern mitochondria and chloroplasts. Sacoglossan "solar-powered" sea slugs possess the ability to acquire organelles within a single lifetime by selectively retaining consumed chloroplasts that remain photosynthetically active for nearly a year. The mechanism for this "animal photosynthesis" remains unknown. Here, we discovered that foreign chloroplasts are housed within novel, host-derived organelles we term "kleptosomes." Kleptosomes use ATP-sensitive ion channels to maintain a luminal environment that supports chloroplast photosynthesis and longevity. Upon slug starvation, kleptosomes digest stored chloroplasts for additional nutrients, thereby serving as a food source. We leveraged this discovery to find that organellar retention and digestion of photosynthetic cargo has convergently evolved in other photosynthetic animals, including corals and anemones. Thus, our study reveals mechanisms underlying the long-term acquisition and evolutionary incorporation of intracellular symbionts into organelles that support complex cellular function.
    Keywords:  cell biology; endosymbiosis; evolution; kleptoplasty; organellar ion channels; photosynthetic animal
    DOI:  https://doi.org/10.1016/j.cell.2025.06.003
  27. Autophagy. 2025 Jun 26.
    Microaggrephagy: an ESCRT-I-PTPN23-dependent pathway for MAPT/tau aggregate clearance.
    Shoshiro Hirayama, Shigeo Murata.
      The clearance mechanisms for ubiquitinated protein aggregates, such as MAPT/tau in neurodegenerative diseases, remain incompletely understood, particularly regarding the role of microautophagy. To identify mediators of this process, we performed an unbiased genome-wide CRISPR knockout screen using cells propagating MAPT/tau repeat domain (MAPT/tauRD) aggregates. This screen identified the ESCRT-I complex and the accessory protein PTPN23 as essential for the clearance of ubiquitinated MAPT/tauRD aggregates via a microautophagy-dependent pathway, operating independently of macroautophagy and chaperone-mediated autophagy. We designate this pathway "microaggrephagy". Mechanistically, microaggrephagy involves the recognition of polyubiquitinated aggregates by the ESCRT-I subunit TSG101, with PTPN23 acting as an adaptor bridging ESCRT-I and ESCRT-III to facilitate microautophagic engulfment. Furthermore, a disease-associated mutation in the ESCRT-I component UBAP1 disrupts its interaction with PTPN23 and impairs MAPT/tau clearance, implicating dysfunction of this pathway in neurodegenerative pathogenesis. These findings establish microaggrephagy as a distinct cellular mechanism for degrading pathological protein aggregates, provide a molecular basis for its function, and suggest potential therapeutic targets for proteinopathies.
    Keywords:  ESCRT-I complex; MAPT/tau repeat domain (mapt/taurd); PTPN23; microaggrephagy; microautophagy; protein aggregates
    DOI:  https://doi.org/10.1080/15548627.2025.2525866
  28. EMBO Rep. 2025 Jun 23.
    LRBA deficiency impairs autophagy and contributes to enhanced antigen presentation and T-cell dysregulation.
    Elena Sindram, Marie-Celine Deau, Laure-Anne Ligeon, Pablo Sanchez-Martin, Sigrun Nestel, Sophie Jung, Stefanie Ruf, Pankaj Mishra, Michele Proietti, Stefan Günther, Kathrin Thedieck, Eleni Roussa, Angelika Rambold, Christian Münz, Claudine Kraft, Bodo Grimbacher, Laura Gámez-Díaz.
      Reduced autophagy is associated with the aberrant humoral response observed in lipopolysaccharide-responsive beige-like anchor protein (LRBA) deficiency; however, the molecular mechanisms and their impact on T-cell responses remain poorly understood. We identify two novel LRBA interactors, phosphoinositide 3-kinase regulatory subunit 4 (PIK3R4) and FYVE And Coiled-Coil Domain Autophagy Adaptor 1 (FYCO1), which each play key roles in autophagy. PIK3R4 facilitates the production of phosphatidylinositol-3 phosphate (PI(3)P) that promotes autophagosome formation and autophagosome-lysosome fusion, whereas FYCO1 supports autophagosome movement. LRBA-knockout (KO) cells show impaired PI(3)P production, reduced autophagosome-lysosome fusion, accumulation of enlarged autophagosomes, and decreased cargo degradation. In line with the role of autophagy as a major degradation system for MHC-II loading and antigen presentation, we observe increased numbers of MHC class II and LC3 vesicles, along with enhanced antigen presentation in absence of LRBA, resulting in a higher production of proinflammatory cytokines from T cells in vitro. Our work suggests a novel biological role of LRBA controlling antigen presentation and T-cell responses by positively regulating autophagy, which may contribute to T-cell immune dysregulation observed in LRBA-deficient patients.
    Keywords:  Autophagy; FYCO1; Immune Dysregulation; LRBA; PIK3R4
    DOI:  https://doi.org/10.1038/s44319-025-00504-7
  29. J Cell Biol. 2025 Aug 04. pii: e202409103. [Epub ahead of print]224(8):
    Acidosis attenuates the hypoxic stabilization of HIF-1α by activating lysosomal degradation.
    Bobby White, Zhenyi Wang, Matthew Dean, Johanna Michl, Natalia Nieora, Sarah Flannery, Iolanda Vendrell, Roman Fischer, Alzbeta Hulikova, Pawel Swietach.
      Hypoxia-inducible factors (HIFs) mediate cellular responses to low oxygen, notably enhanced fermentation that acidifies poorly perfused tissues and may eventually become more damaging than adaptive. How pH feeds back on hypoxic signaling is unclear but critical to investigate because acidosis and hypoxia are mechanistically coupled in diffusion-limited settings, such as tumors. Here, we examined the pH sensitivity of hypoxic signaling in colorectal cancer cells that can survive acidosis. HIF-1α stabilization under acidotic hypoxia was transient, declining over 48 h. Proteomic analyses identified responses that followed HIF-1α, including canonical HIF targets (e.g., CA9, PDK1), but these did not reflect a proteome-wide downregulation. Enrichment analyses suggested a role for lysosomal degradation. Indeed, HIF-1α destabilization was blocked by inactivating lysosomes, but not proteasome inhibitors. Acidotic hypoxia stimulated lysosomal activity and autophagy via mammalian target of rapamycin complex I (mTORC1), resulting in HIF-1α degradation. This response protects cells from excessive acidification by unchecked fermentation. Thus, alkaline conditions are permissive for at least some aspects of HIF-1α signaling.
    DOI:  https://doi.org/10.1083/jcb.202409103
  30. J Biol Chem. 2025 Jun 17. pii: S0021-9258(25)02225-2. [Epub ahead of print] 110375
    Unifying Perspectives on the Activity and Genotypic Targeting of Pharmacological Chaperones.
    Austin Tedman, Muskan Goel, Sohan Shah, Jonathan P Schlebach.
      Several diseases of protein misfolding can now be treated with an emerging class of therapeutics known as pharmacological chaperones, pharmacochaperones, or correctors. These small molecules exploit the universal thermodynamic coupling between ligand binding and protein folding to suppress conformational defects that disrupt protein homeostasis. While the mechanistic basis of their activity is quite simple in theory, their nuanced proteostatic effects can vary depending on the intrinsic properties of their target proteins and the cellular context. Deviations in activity are especially pronounced across panels of pathogenic variants of the target protein. In this perspective, we explore the factors that shape the potency of pharmacochaperones and the intrinsic sensitivity of different target proteins in relation to various theoretical considerations and experimental observations. We discuss how emerging technologies have provided general insights into the molecular basis of the variant-specific effects of certain pharmacochaperones. We also highlight ongoing efforts to identify existing drugs that stabilize misfolded variants and to repurpose them as pharmacochaperones. Finally, we discuss how the chaperone activity of current drugs could potentially contribute to complex pharmacology and deviations in therapeutic efficacy across patient cohorts. Together, these principles provide a coherent framework that may help guide the discovery and precision targeting of next generation pharmacochaperones for both current and new targets involved in proteostasis diseases. An audio recording of this work is included in the supplement and can be freely streamed here.
    DOI:  https://doi.org/10.1016/j.jbc.2025.110375
  31. Sci Signal. 2025 Jun 24. 18(892): eads6550
    Loss of STIM2, but not of STIM1, drives colorectal cancer metastasis through metabolic reprogramming and the ATF4 ER stress pathway.
    Trayambak Pathak, J Cory Benson, Martin T Johnson, Ping Xin, Ahmed Emam Abdelnaby, Vonn Walter, Walter A Koltun, Gregory S Yochum, Nadine Hempel, Mohamed Trebak.
      The large amounts of calcium (Ca2+) stored in the endoplasmic reticulum (ER) and the controlled release of this Ca2+ store into the cytosol regulate many cellular functions, and altered ER Ca2+ homeostasis induces ER stress. Stromal-interacting molecules 1 and 2 (STIM1/2) are homologous ER-resident Ca2+ sensors that synergistically activate cytosolic Ca2+ influx through Orai channels to promote Ca2+-dependent changes in gene expression and ER Ca2+ refilling. Here, we demonstrated that reduced abundance of STIM2, but not that of STIM1, was associated with poor prognosis in colorectal cancer (CRC). STIM2-deficient CRC cells showed enhanced ER Ca2+ content in a manner dependent on the ER Ca2+ pump SERCA2, increased expression of genes associated with protein translation, and transcriptional and metabolic rewiring. STIM2 deficiency in CRC xenografts led to increased tumor size, invasion, and metastasis. STIM2 loss activated the expression of genes involved in ER stress responses in a manner dependent on the chaperone BiP and the transcription factor ATF4 and independent of Orai channels. These results suggest that loss of STIM2 may inform CRC prognosis.
    DOI:  https://doi.org/10.1126/scisignal.ads6550
  32. Nat Chem Biol. 2025 Jun 24.
    A small-molecule VHL molecular glue degrader for cysteine dioxygenase 1.
    Antonin Tutter, Dennis Buckley, Andrei A Golosov, Xiaolei Ma, Wei Shu, Daniel J J McKay, Veronique Darsigny, Dustin Dovala, Rohan Beckwith, Jonathan Solomon, Pasupuleti Rao, Lei Xu, Aleem Fazal, Andreas Lingel, Charles Wartchow, Jennifer S Cobb, Amanda Hachey, Jennifer Tullai, Gregory A Michaud.
      The von Hippel-Lindau tumor suppressor gene product (pVHL) is an E3 ligase substrate receptor that binds proline-hydroxylated hypoxia-inducible factor HIF1α, leading to its ubiquitin-dependent degradation. By using protein arrays, we identified a small molecule that binds the HIF1α-binding pocket on pVHL and functions as a molecular glue degrader of the neosubstrate cysteine dioxygenase (CDO1) by recruiting it into the VHL-Cullin-RING E3 ligase complex and leading to its selective degradation. The CDO1-binding region involved in VHL recruitment was characterized through a combination of mutagenesis and protein-protein docking coupled with molecular-dynamics-based solvation analysis. The X-ray structure of the ternary complexes of VHL, CDO1 and degrader molecules confirms the binding region prediction and provides atomic insights into key molecular glue interactions.
    DOI:  https://doi.org/10.1038/s41589-025-01936-x
  33. J Am Chem Soc. 2025 Jun 25.
    Design and Semisynthesis of Ubiquitin Extension Probes for Structural Analysis of Cullin1-Mediated Substrate Polyubiquitination.
    Chuntong Li, Fangyu Zhao, Chong Zuo, Liying Zhang, Yangwode Jing, Xu Li, Guo-Chao Chu, Luyu Shi, Yingyue Zhang, Han Wang, Shuzhe Sun, Maoshen Sun, Huasong Ai, Lu-Jun Liang, Jinghong Li.
      Chemical trapping strategies have recently emerged as powerful approaches for investigating the structural dynamics of E3 ligase-catalyzed substrate ubiquitination. However, current ubiquitination-derived probes are limited to studying substrate mono- or diubiquitination events. Probes capable of investigating how E3 ligases accommodate E2-Ub conjugates and ubiquitinated substrates to generate longer ubiquitin chains remain unexplored. In this work, we report the development of two Cullin1 E3 ligase (CRL1)-dependent probes, Extension ProbeUb2 and Extension ProbeUb4, which mimic transient intermediates formed during CRL1-catalyzed K48-linked diubiquitin and tetraubiquitin chain formation on substrate p27. Notably, a chemoenzymatic semisynthetic strategy was devised to generate Extension ProbeUb4, involving the enzymatic conjugation of a preformed K48-linked diubiquitin to a synthetic Ub-p27-degron construct using the E2 conjugating enzyme UBE2K. Both Extension ProbeUb2 and Extension ProbeUb4 formed stable complexes with N8-CRL1Skp1/Skp2/Cks1 (comprising neddylated Cullin1-Rbx1 and the substrate receptor complex Skp1-Skp2-Cks1), facilitating structural analysis by chemical cross-linking mass spectrometry (CX-MS) and cryo-electron microscopy (cryo-EM). Our results indicate the presence of multiple distinct conformations of the catalytic module (comprising the RING domain of Rbx1, CDC34-Ub, and the acceptor ubiquitin) within the di- and tetraubiquitination complexes, while the conformation of the Cullin1-Skp1-Skp2-Cks1 subunit remains unchanged. In conclusion, this work expands the toolkit available for chemical trapping strategies and provides advanced insights into CRL-catalyzed substrate polyubiquitination.
    DOI:  https://doi.org/10.1021/jacs.5c06399
  34. bioRxiv. 2024 Feb 21. pii: 2024.02.19.580970. [Epub ahead of print]
    Enhanced Protein-Protein Interaction Discovery via AlphaFold-Multimer.
    Ah-Ram Kim, Yanhui Hu, Aram Comjean, Jonathan Rodiger, Stephanie E Mohr, Norbert Perrimon.
      Accurately mapping protein-protein interactions (PPIs) is critical for elucidating cellular functions and has significant implications for health and disease. Conventional experimental approaches, while foundational, often fall short in capturing direct, dynamic interactions, especially those with transient or small interfaces. Our study leverages AlphaFold-Multimer (AFM) to re-evaluate high-confidence PPI datasets from Drosophila and human. Our analysis uncovers a significant limitation of the AFM-derived interface pTM (ipTM) metric, which, while reflective of structural integrity, can miss physiologically relevant interactions at small interfaces or within flexible regions. To bridge this gap, we introduce the Local Interaction Score (LIS), derived from AFM's Predicted Aligned Error (PAE), focusing on areas with low PAE values, indicative of the high confidence in interaction predictions. The LIS method demonstrates enhanced sensitivity in detecting PPIs, particularly among those that involve flexible and small interfaces. By applying LIS to large-scale Drosophila datasets, we enhance the detection of direct interactions. Moreover, we present FlyPredictome, an online platform that integrates our AFM-based predictions with additional information such as gene expression correlations and subcellular localization predictions. This study not only improves upon AFM's utility in PPI prediction but also highlights the potential of computational methods to complement and enhance experimental approaches in the identification of PPI networks.
    DOI:  https://doi.org/10.1101/2024.02.19.580970
  35. Sci Adv. 2025 Jun 27. 11(26): eadq9643
    VIRMA-mediated m6A modification regulates forebrain formation through modulating ribosome biogenesis.
    Min Wu, Xiaoli Wu, Haifeng Sun, Wen Wang, Leyi Zhang, Xia Liu, Yifan Zhang, Xinning Zhang, Jun Liu, Bin Shen, Tao Zhou.
      N6-Methyladenosine (m6A) modification plays crucial roles in tissue development and homeostasis. However, the mechanisms underlying cellular adaptation of m6A modification and their impact on protein synthesis machinery remain unclear. VIRMA, the largest and evolutionarily conserved core of the m6A methyltransferase complex, is highly expressed in the embryonic brain and various cancers. Here, we demonstrate that VIRMA-mediated m6A modification is essential for active ribosome biogenesis. VIRMA depletion destabilizes the entire writer complex and reduces m6A levels, leading to decreased proliferation and increased apoptosis of neural progenitor/stem cells, ultimately causing severe forebrain developmental defects. Mechanistically, VIRMA depletion impairs ribosome biogenesis by inhibiting mRNA decay, triggering a p53-dependent stress response and compromising global protein synthesis. These findings extend to some cancer cells, suggesting a potential conservation of this mechanism. Overall, our study reveals the critical role of m6A in adapting protein synthesis machinery during brain development.
    DOI:  https://doi.org/10.1126/sciadv.adq9643
  36. Nat Cell Biol. 2025 Jun 27.
    The Vps13-like protein BLTP2 regulates phosphatidylethanolamine levels to maintain plasma membrane fluidity and breast cancer aggressiveness.
    Subhrajit Banerjee, Stephan Daetwyler, Xiaofei Bai, Morgane Michaud, Juliette Jouhet, Derk Binns, Shruthi Madhugiri, Emma Johnson, Chao-Wen Wang, Reto Fiolka, Alexandre Toulmay, William A Prinz.
      Lipid transport proteins (LTPs) facilitate non-vesicular lipid exchange between cellular compartments and have critical roles in lipid homeostasis. A recently identified family of bridge-like LTPs (BLTPs) is thought to form lipid-transporting conduits between organelles. One of these, BLTP2, is conserved across species but its function is not known. Here we show that BLTP2 regulates plasma membrane (PM) fluidity by increasing phosphatidylethanolamine (PE) levels in the PM. BLTP2 localizes to endoplasmic reticulum (ER)-PM contact sites, and transports PE in vivo, suggesting it drives PE movement from ER to PM. We find that BLTP2 works in parallel with another pathway that regulates intracellular PE distribution and PM fluidity. BLTP2 expression correlates with breast cancer aggressiveness. We found that BLTP2 facilitates growth of a triple negative breast cancer cell line and sustains its aggressiveness in an in vivo model of metastasis, suggesting maintenance of PM fluidity by BLTP2 may be critical for tumorigenesis in humans.
    DOI:  https://doi.org/10.1038/s41556-025-01672-3
  37. Biochem J. 2025 Jun 25. pii: BCJ20243018. [Epub ahead of print]482(13):
    Methods to accelerate PROTAC drug discovery.
    Jeyan Osman, Philip E Thompson, Manuela Jörg, Martin J Scanlon.
      Proteolysis-targeting chimeras (PROTACs) represent a novel and promising modality for probing biological systems, elucidating pharmacological mechanisms, and identifying potential therapeutic leads. The field has made significant strides, as demonstrated by the growing number of PROTACs advancing to clinical trials. Despite this progress, the development of PROTACs faces significant challenges, which is partially due to the heterobivalent nature of this class of molecules. PROTACs must simultaneously bind to a protein of interest and an E3 ubiquitin ligase. This means PROTACs are significantly larger and more complex than conventional small molecules. This complexity impacts their design and synthesis, requiring strategic approaches to create libraries of PROTACs with various combinations of target ligands, linkers, and E3 ligase-recruiting elements. To fully realise the potential of this innovative technology, there is a need for novel approaches to accelerate the development of PROTACs. This review focuses on three critical areas to accelerate PROTAC development: appropriate target selection, modular chemical synthesis, and high-throughput biological evaluation. By appropriate selection of target proteins for degradation, optimizing synthesis methods to handle the complexity of PROTAC molecules, and employing robust high-throughput biological assays to evaluate PROTAC activity, researchers in academia and industry have streamlined the development and increased the success rate of PROTAC-based discovery programmes.
    Keywords:  DNA-encoded library; click chemistry; direct-to-biology; proteolysis-targeting chimeras; solid-phase synthesis
    DOI:  https://doi.org/10.1042/BCJ20243018
  38. Cell Chem Biol. 2025 Jun 13. pii: S2451-9456(25)00173-4. [Epub ahead of print]
    Ribosomal RNA transcription regulates splicing through ribosomal protein RPL22.
    Wenjun Fan, Hester Liu, Gregory C Stachelek, Asma Begum, Catherine E Davis, Tony E Dorado, Glen Ernst, William C Reinhold, Busra Ozbek, Qizhi Zheng, Angelo M De Marzo, N V Rajeshkumar, James C Barrow, Marikki Laiho.
      Ribosome biosynthesis is a cancer vulnerability targeted by inhibiting RNA polymerase I (Pol I) transcription. We developed specific Pol I inhibitors that activate a ribotoxic stress pathway to uncover drivers of sensitivity. Integrating multi-omics and drug response data from a large cancer cell panel, we found that RPL22 frameshift mutations confer Pol I inhibitor sensitivity. Mechanistically, RPL22 interacts directly with 28S rRNA and mRNA splice junctions, acting as a splicing regulator. RPL22 deficiency, intensified by 28S rRNA sequestration, promotes splicing of its paralog RPL22L1 and the p53 negative regulator MDM4. Both chemical and genetic inhibition of rRNA synthesis broadly remodel mRNA splicing controlling hundreds of targets. Notably, RPL22-dependent alternative splicing is reversed by Pol I inhibition, revealing a non-canonical ribotoxic stress-initiated tumor suppressive pathway. This study uncovers a robust mechanism linking rRNA synthesis activity to splicing, coordinated by the ribosomal protein RPL22.
    Keywords:  MDM4; RPL22; RPL22L1; cancer; nucleolus; rRNA synthesis; ribosome biogenesis; small-molecule; splicing; therapeutics
    DOI:  https://doi.org/10.1016/j.chembiol.2025.05.012
  39. RSC Chem Biol. 2025 Jun 19.
    Visualizing stress granule dynamics with an RNA guanine quadruplex targeted ruthenium(ii) peptide conjugate.
    Rhianne C Curley, Lorcan Holden, Tia E Keyes.
      Stress granules (SGs) are membraneless ribonucleoprotein assemblies that form in response to cellular stress. They have been linked to cell survival and cancer progression, though many questions remain regarding their structure, function and therapeutic potential. Live-cell fluorescence imaging is key to advancing understanding of SGs, but there are very few small-molecule probes reported that selectively image these organelles. RNA G-quadruplex (rG4) folding is believed to play a role in initiation of SG formation. Thus, to create a probe for SGs, we conjugated a G4 binding domain peptide from RNA helicase associated with AU-rich element (RHAU) to a luminescent [Ru(bpy)2(PIC-COOH)]2+, Ru-RHAU. Ru-RHAU is designed to target rG4s and thus SGs in live cells. Studies in cellulo demonstrate that Ru-RHAU can induce SG formation in a concentration and time dependent manner and immunolabelling confirmed the complex remains associated with rG4s in the SGs. The SG stimulation is attributed to stabilization of rG4 by Ru-RHAU consistent with rG4's role in SG formation. Ru-RHAU shows low cytotoxicity under imaging conditions, facilitating prolonged observation in live cells. Interestingly, under more intense photoirradiation, Ru-RHAU induces phototoxicity through an apoptotic pathway. Ru-RHAU is a versatile tool for probing SG dynamics and function in cellular stress responses and has heretofore unconsidered potential in phototherapeutic applications targeting SGs.
    DOI:  https://doi.org/10.1039/d5cb00008d
  40. Nat Metab. 2025 Jun 27.
    Dietary control of peripheral adipose storage capacity through membrane lipid remodelling.
    Marcus J Tol, Yuta Shimanaka, Alexander H Bedard, Jennifer Sapia, Liujuan Cui, Mariana Colaço-Gaspar, Peter Hofer, Alessandra Ferrari, Kevin Qian, John P Kennelly, Stephen D Lee, Yajing Gao, Xu Xiao, Jie Gao, Julia J Mack, Thomas A Weston, Kevin J Williams, Baolong Su, Calvin Pan, Aldons J Lusis, Daniel P Pike, Alex Reed, Natalia Milosevich, Benjamin F Cravatt, Makoto Arita, Stephen G Young, David A Ford, Rudolf Zechner, Stefano Vanni, Peter Tontonoz.
      Genetic and dietary cues are known drivers of obesity, yet how they converge at the molecular level is incompletely understood. Here we show that PPARγ supports hypertrophic expansion of adipose tissue via transcriptional control of LPCAT3, an endoplasmic reticulum (ER)-resident O-acyltransferase that selectively enriches diet-derived omega-6 polyunsaturated fatty acids (n-6 PUFAs) in the membrane lipidome. In mice fed a high-fat diet, lowering membrane n-6 PUFA levels through genetic or dietary interventions results in aberrant adipose triglyceride (TG) turnover, ectopic fat deposition and insulin resistance. Additionally, we detail a non-canonical adaptive response in 'lipodystrophic' Lpcat3-/- adipose tissues that engages a futile lipid cycle to increase metabolic rate and offset lipid overflow to ectopic sites. Live-cell imaging, lipidomics and molecular dynamics simulations reveal that adipocyte LPCAT3 activity enriches n-6 arachidonate in the phosphatidylethanolamine (PE)-dense ER-lipid droplet interface. Functionally, this localized PE remodelling optimizes TG storage by driving the formation of large droplets that exhibit greater resistance to adipose TG lipase activity. These findings highlight the PPARγ-LPCAT3 axis as a mechanistic link between dietary n-6 PUFA intake, adipose expandability and systemic energy balance.
    DOI:  https://doi.org/10.1038/s42255-025-01320-y
  41. Mol Cell. 2025 Jun 19. pii: S1097-2765(25)00504-0. [Epub ahead of print]
    Isoform-specific function of NSD3 in DNA replication stress confers resistance to PARP inhibitors in prostate cancer.
    Shouhai Zhu, Huanyao Gao, Dan Jiang, Guijie Guo, Jing Hou, Yiqun Han, Chao Zhang, Xiaoping Hu, Shreya Indulkar, Jake A Kloeber, Qi Hu, Yanxia Jiang, Xiangyu Zeng, Yaobin Ouyang, Jing Lu, Ping Yin, Kuntian Luo, Jinzhou Huang, Zheming Wu, Bin Chen, Huaping Xiao, Sonja Dragojevic, Hongran Qin, Xiang Zhou, Jian Jin, Xinyi Tu, Liewei Wang, Zhenkun Lou.
      Poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) have been approved for the treatment of metastatic castration-resistant prostate cancer (mCRPC) in patients with deleterious BRCA1/2 alterations. Although this marks a significant milestone, intrinsic or acquired therapy resistance remains a major challenge that limits clinical efficacy. Here, we demonstrate that dysregulated ubiquitination and turnover by the cullin 3 (CUL3)ZBTB2 E3 ligase complex induce the upregulation of the short isoform of nuclear-receptor-binding SET domain protein 3 (NSD3) (NSD3S), which confers PARPi resistance in prostate cancer cells and patient-derived mCRPC samples. Mechanistically, ATR drives the localization of NSD3S at stalled replication forks, where it antagonizes the PTIP-dependent recruitment of the MRE11 nuclease, thereby protecting nascent DNA from extensive degradation and ensuring fork stabilization. Importantly, pharmacological degradation of NSD3S using an NSD3-targeting proteolysis-targeting chimera (PROTAC) efficiently enhances PARPi sensitivity in both cell-line-derived xenograft and patient-derived xenograft (PDX) mouse models. These findings establish NSD3S as a key determinant of PARPi toxicity in mCRPC.
    Keywords:  CUL3; MRE11; NSD3; PARP inhibitor; PTIP; prostate cancer; proteolysis-targeting chimera; replication fork
    DOI:  https://doi.org/10.1016/j.molcel.2025.06.004
  42. Cell Metab. 2025 Jun 20. pii: S1550-4131(25)00295-5. [Epub ahead of print]
    Sphingomyelins in mosquito saliva reconfigure skin lipidome to promote viral protein levels and enhance transmission of flaviviruses.
    Hacène Medkour, Lauryne Pruvost, Elliott F Miot, Xiaoqian Gong, Virginie Vaissayre, Mihra Tavadia, Pascal Boutinaud, Justine Revel, Atitaya Hitakarun, Wannapa Sornjai, Jim Zoladek, Duncan R Smith, Sébastien Nisole, Esther Nolte-'t Hoen, Justine Bertrand-Michel, Dorothée Missé, Guillaume Marti, Julien Pompon.
      Many flaviviruses with high pandemic potential are transmitted through mosquito bites. While mosquito saliva is essential for transmission and represents a promising pan-flaviviral target, there is a dearth of knowledge on salivary metabolic transmission enhancers. Here, we show that extracellular vesicle (EV)-derived sphingomyelins in mosquito saliva reconfigure the human cell lipidome to increase viral protein levels, boosting skin infection and enhancing transmission for flaviviruses. Lipids within internalized mosquito EVs enhance infection in fibroblast and immune human primary cells for multiple flaviviruses. Mosquito EV lipids selectively increase viral translation by inhibiting infection-induced endoplasmic reticulum (ER)-associated degradation of viral proteins. Infection enhancement solely results from the sphingomyelins within salivary mosquito EVs that augment human cell sphingomyelin concentration. Finally, EV-lipid co-inoculation exacerbates disease severity in vivo in mouse transmission assays. By discovering and elucidating how metabolic components of mosquito saliva promote transmission of flaviviruses, our study unveils lipids as a new category of targets against vectored transmission.
    Keywords:  ERAD; Orthoflavivirus; West Nile; Zika; bite; dengue; extracellular vesicles; lipids; transmission; unfolded protein response
    DOI:  https://doi.org/10.1016/j.cmet.2025.05.015
  43. Nat Commun. 2025 Jun 25. 16(1): 5379
    Enhancing autophagy by redox regulation extends lifespan in Drosophila.
    Claudia Lennicke, Ivana Bjedov, Sebastian Grönke, Katja E Menger, Andrew M James, Jorge Iván Castillo-Quan, Lucie A G van Leeuwen, Andrea Foley, Marcela Buricova, Jennifer Adcott, Alex Montoya, Holger B Kramer, Pavel V Shliaha, Angela Logan, Filipe Cabreiro, Michael P Murphy, Linda Partridge, Helena M Cochemé.
      Dysregulation of redox homeostasis is implicated in the ageing process and the pathology of age-related diseases. To study redox signalling by H2O2 in vivo, we established a redox-shifted model by manipulating levels of the H2O2-degrading enzyme catalase in Drosophila. Here we report that ubiquitous over-expression of catalase robustly extends lifespan in females. As anticipated, these flies are strongly resistant to a range of oxidative stress challenges, but interestingly are sensitive to starvation, which could not be explained by differences in levels of energy reserves. This led us to explore the contribution of autophagy, which is an important mechanism for organismal survival in response to starvation. We show that autophagy is essential for the increased lifespan by catalase upregulation, as the survival benefits are completely abolished upon global autophagy knock-down. Furthermore, using a specific redox-inactive knock-in mutant, we highlight the in vivo role of a key regulatory cysteine residue in Atg4a, which is required for the lifespan extension in our catalase model. Altogether, these findings confirm the redox regulation of autophagy in vivo as an important modulator of longevity.
    DOI:  https://doi.org/10.1038/s41467-025-60603-w
  44. Cell Rep. 2025 Jun 24. pii: S2211-1247(25)00638-2. [Epub ahead of print]44(7): 115867
    Mislocalization of nucleic acids is a convergent and targetable mechanism in Alzheimer's disease and frontotemporal dementia.
    Christy Hung, Miriam Llorian, Koustav Pal, Frederick J Livesey, Jyoti Choudhary, Theodoros I Roumeliotis, Rickie Patani.
      Nucleocytoplasmic transport defects are observed in Alzheimer's disease (AD) and frontotemporal dementia (FTD). Here, we assess mRNA nucleocytoplasmic localization by performing transcriptome-wide profiling on nuclear and cytoplasmic fractions of human iPSC-derived cortical neurons from healthy individuals compared to those with familial AD or FTD. We find that AD- and FTD-causing mutations induce significant changes in mRNA nucleocytoplasmic distribution. We additionally observe the redistribution of mitochondria-related transcripts across AD and FTD neurons. The significantly increased mitochondrial RNA (mtRNA) in the cytosol of AD and FTD mutant neurons raised the possibility of leakage, which motivated us to investigate mtDNA leakage. We reveal abnormal cytoplasmic accumulation of mtDNA in AD and FTD cortical neurons together with evidence of mitochondrial aberrance. Importantly, mislocalisation of nucleic acids, mitochondrial dysfunction and cGAS-STING activation can be ameliorated through VCP D2 ATPase inhibition.
    Keywords:  Alzheimer's disease; CP: Molecular biology; CP: Neuroscience; VCP inhibition; cGAS-STING; frontotemporal dementia; mRNA; mitochondrial DNA; mitochondrial RNA; mitochondrial dysfunction; neurodegeneration; nucleocytoplasmic redistribution
    DOI:  https://doi.org/10.1016/j.celrep.2025.115867
  45. Cell Stem Cell. 2025 Jun 19. pii: S1934-5909(25)00224-3. [Epub ahead of print]
    ChemPerturb-seq screen identifies a small molecule cocktail enhancing human beta cell survival after subcutaneous transplantation.
    J Jeya Vandana, Jiajun Zhu, Alice Maria Giani, Tuo Zhang, Lauretta A Lacko, Dongliang Leng, D Leland Taylor, Brian N Lee, Zhaowei Han, Tiancheng Jiao, Yuanhao Huang, Meiqi Zhao, Xinyi Liu, Angie Chi Nok Chong, Dongxiang Xue, Zihe Meng, Jenny Z Xiang, Chendong Pan, Wei Wang, Ali Naji, Todd Evans, Jie Liu, Francis S Collins, Chengyang Liu, Shuibing Chen.
      Traditional chemical screens have focused on a single assay per screen, making them labor intensive and costly. Here, we combined a chemical screen with single-cell RNA sequencing (scRNA-seq) to perform Chemical Perturb-seq (ChemPerturb-seq), enabling a systematic analysis of the molecular changes of human beta cells upon individual small molecule treatments. Using this platform, we performed an in vivo barcoded screen and discovered a small molecule cocktail, including beta-lipotropin 61-91, insulin growth factor-1, and prostaglandin E2, with which preconditioning human beta cells and primary islets significantly enhanced function and survival when transplanted subcutaneously to female, but not to male, mice. We identified two additional molecules, serotonin and histamine, that promote islet function when transplanted subcutaneously to male mice using ChemPerturb-seq. Such small molecule cocktails could be applied to improve the current FDA-approved islet transplantation procedure. Finally, we developed an artificial intelligence (AI)-powered website, ChemPerturbDB, which provides user-friendly open access analysis of the extensive ChemPerturb-seq dataset.
    Keywords:  ChemPerturb-seq; beta-lipotropin 61-91; histamine; insulin growth factor-1; islet transplantation; primary islets; prostaglandin E2; serotonin
    DOI:  https://doi.org/10.1016/j.stem.2025.06.002
  46. Mol Cell. 2025 Jun 16. pii: S1097-2765(25)00499-X. [Epub ahead of print]
    BAG2 releases SAMD4B upon sensing of arginine deficiency to promote tumor cell survival.
    Meng-Ying Chen, Chun-Yu Sun, Ru Zhao, Xiao-Lin Guan, Miao-Lin Li, Fan Zhang, Zhi-Han Wan, Jing-Xuan Feng, Miao Yin, Qun-Ying Lei, Qi-Xiang Ma.
      Cells possess numerous metabolite sensors that detect essential nutrients for growth, with many directly binding to metabolites and responding to their levels. Given the vital role of arginine in various physiological and pathological processes, we hypothesized that there may be undiscovered sensors that detect arginine deficiency. Through a series of unbiased screening strategies in human cancer cell line models, we identified Bcl2-associated athanogene (BAG) family molecular chaperone regulator 2 (BAG2) as an arginine sensor, which could directly bind to arginine at the glutamine residue 167 (Q167). Upon arginine deficiency, BAG2 releases sterile alpha motif domain-containing protein 4B (SAMD4B), leading to β-catenin degradation to stabilize ATF4 protein, enhancing cell survival. When arginine is abundant, a strengthened binding between BAG2 and SAMD4B prevents β-catenin degradation, activating the Wnt/β-catenin pathway to support cell growth. Overall, our findings uncover an arginine-sensing pathway consisting of BAG2 and SAMD4B that promotes cancer cell adaptation to nutritional stress.
    Keywords:  BAG2; SAMD4B; arginine; cancer; metabolism; metabolite sensing; metabolite sensor
    DOI:  https://doi.org/10.1016/j.molcel.2025.05.035
  47. Nat Cell Biol. 2025 Jun 26.
    A lysosomal surveillance response to stress extends healthspan.
    Terytty Yang Li, Arwen W Gao, Rendan Yang, Yu Sun, Yuxuan Lei, Xiaoxu Li, Lin Chen, Yasmine J Liu, Rachel N Arey, Kimberly Morales, Raya B Liu, Wenzheng Wang, Ang Zhou, Tong-Jin Zhao, Weisha Li, Amélia Lalou, Qi Wang, Tanes Lima, Riekelt H Houtkooper, Johan Auwerx.
      Lysosomes are cytoplasmic organelles central for the degradation of macromolecules to maintain cellular homoeostasis and health. However, how lysosomal activity can be boosted to counteract ageing and ageing-related diseases remains elusive. Here we reveal that silencing specific vacuolar H+-ATPase subunits (for example, vha-6), which are essential for intestinal lumen acidification in Caenorhabditis elegans, extends lifespan by ~60%. This longevity phenotype can be explained by an adaptive transcriptional response typified by induction of a set of transcripts involved in lysosomal function and proteolysis, which we termed the lysosomal surveillance response (LySR). LySR activation is characterized by boosted lysosomal activity and enhanced clearance of protein aggregates in worm models of Alzheimer's disease, Huntington's disease and amyotrophic lateral sclerosis, thereby improving fitness. The GATA transcription factor ELT-2 governs the LySR programme and its associated beneficial effects. Activating the LySR pathway may therefore represent an attractive mechanism to reduce proteotoxicity and, as such, potentially extend healthspan.
    DOI:  https://doi.org/10.1038/s41556-025-01693-y
  48. Nat Cancer. 2025 Jun 27.
    Profiling antigen-binding affinity of B cell repertoires in tumors by deep learning predicts immune-checkpoint inhibitor treatment outcomes.
    Bing Song, Kaiwen Wang, Saiyang Na, Jia Yao, Farjana J Fattah, Alexandra L Martin, Mitchell S von Itzstein, Donghan M Yang, Jialiang Liu, Yaming Xue, Chaoying Liang, Yuzhi Guo, Indu Raman, Chengsong Zhu, Jonathan E Dowell, Jade Homsi, Sawsan Rashdan, Shengjie Yang, Mary E Gwin, Tuoqi Wu, David Hsiehchen, Yvonne Gloria-McCutchen, Catherine Pei-Ju Lu, Prithvi Raj, Xiao-Chen Bai, Jun Wang, Jose Conejo-Garcia, Yang Xie, Junzhou Huang, David E Gerber, Tao Wang.
      The capability to profile the landscape of antigen-binding affinities of a vast number of antibodies (B cell receptors, BCRs) will provide a powerful tool to reveal biological insights. However, experimental approaches for detecting antibody-antigen interactions are costly and time-consuming and can only achieve low-to-mid throughput. In this work, we developed Cmai (contrastive modeling for antigen-antibody interactions) to address the prediction of binding between antibodies and antigens that can be scaled to high-throughput sequencing data. We devised a biomarker based on the output from Cmai to map the antigen-binding affinities of BCR repertoires. We found that the abundance of tumor antigen-targeting antibodies is predictive of immune-checkpoint inhibitor (ICI) treatment response. We also found that, during immune-related adverse events (irAEs) caused by ICI, humoral immunity is preferentially responsive to intracellular antigens from the organs affected by the irAEs. We used Cmai to construct a BCR-based irAE risk score, which predicted the timing of the occurrence of irAEs.
    DOI:  https://doi.org/10.1038/s43018-025-01001-5
  49. J Biol Chem. 2025 Jun 19. pii: S0021-9258(25)02241-0. [Epub ahead of print] 110391
    The human autophagy-initiating complexes ULK1C and PI3KC3-C1.
    Minghao Chen, James Hurley.
      The unc-51-like kinase complex (ULK1C) and the class III phosphatidylinositol 3-kinase complex I (PI3KC3-C1) are the key regulators of macroautophagy initiation. Understanding the assembly and coordination of these two complexes is essential for deciphering their cellular regulation and targeting them for therapeutic enhancement. This review highlights recent advances in our understanding of the structural organization and activation mechanisms of ULK1C and PI3KC3-C1 at the molecular level and discusses their roles within the protein interaction network governing autophagy initiation.
    DOI:  https://doi.org/10.1016/j.jbc.2025.110391
  50. bioRxiv. 2025 May 02. pii: 2025.04.28.651139. [Epub ahead of print]
    Design of combination therapeutics from protein response to drugs in ovarian cancer cells.
    Alexandra Franz, Ciyue Shen, Fabian Coscia, Kenneth Munroe, Lea Charaoui, Anil Korkut, Matthias Mann, Augustin Luna, Chris Sander.
      High-grade serous ovarian cancer (HGSOC) remains the most lethal gynecologic malignancy and novel treatment approaches are needed. Here, we used unbiased quantitative protein mass spectrometry to assess the cellular response profile to drug perturbations in ovarian cancer cells for the rational design of potential combination therapies. Analysis of the perturbation profiles revealed proteins responding across several drug perturbations (called frequently responsive below) as well as drug-specific protein responses. The frequently responsive proteins included proteins that reflected general drug resistance mechanisms such as changes in drug efflux pumps. Network analysis of drug-specific protein responses revealed known and potential novel markers of resistance, which were used to rationalize the design of anti-resistance drug pairs. We experimentally tested the anti-proliferative effects of 12 of the proposed drug combinations in 6 HGSOC cell lines. Drug combinations tested with additive or synergistic effects are plausible candidates for overcoming or preventing resistance to single agents; these include several combinations that were synergistic (with PARPi, MEKi, and SRCi). Additionally, we observed 0.05-0.11 micromolar response to GPX4 inhibitors as single agents in the OVCAR-4 cell line. We propose several drug combinations as potential therapeutic candidates in ovarian cancer, as well as GPX4 inhibitors as single agents.
    DOI:  https://doi.org/10.1101/2025.04.28.651139
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