bims-proteo Biomed News
on Proteostasis
Issue of 2025–09–14
38 papers selected by
Eric Chevet, INSERM
  1. Dynamic regulation of the oxidative stress response by the E3 ligase TRIP12.
  2. Coordination of autophagosome closure and release by the Alzheimer's disease-associated protein BIN1.
  3. ATG16L1 controls mammalian vacuolar proton ATPase.
  4. α-Synuclein aggregates inhibit ESCRT-III through sequestration and collateral degradation.
  5. E3 ubiquitin ligases in signaling, disease, and therapeutics.
  6. Thyroidal expression of ER molecular chaperone GRP170 is required for efficient TSH-mediated thyroid hormone synthesis.
  7. BLTP3A is associated with membranes of the late endocytic pathway and is an effector of CASM.
  8. The E3 ubiquitin ligase SIAH1 targets a bacterial pore-forming toxin to facilitate xenophagy.
  9. Protocol for monitoring mRNA translation and degradation in human cell-free lysates.
  10. A novel role for the E2F transcription factor and the ER stress sensor IRE1 in cytoplasmic DNA accumulation.
  11. Biomolecular condensates of ATG18 reshape ER for autophagy in plants.
  12. Inhibition of the UFD-1-NPL-4 complex triggers an aberrant immune response in Caenorhabditis elegans.
  13. The E3 ligase MEX3B forms a tripartite complex with Rest and Hotair to determine the proliferative capacity of neural progenitor cells.
  14. Mammalian mitohormesis: from mitochondrial stressors to organismal benefits.
  15. Rapid UPF1 depletion illuminates the temporal dynamics of the NMD-regulated human transcriptome.
  16. Loss of CHMP2A implicates an ordered assembly of ESCRT-III proteins during cytokinetic abscission.
  17. Giant KASH proteins and ribosomes establish distinct cytoplasmic biophysical properties in vivo.
  18. The hexosamine biosynthetic pathway drives tumor immune evasion via translational control of PD-L1 at the elongation level.
  19. Bi-handed assembly chaperones regulate protein complex assembly through an intramolecular handover mechanism.
  20. Genome-wide mapping of RNA-protein associations through sequencing.
  21. TurboID-based mapping of organelle membrane protein interactomes with digitonin-permeabilization.
  22. Urolithin a modulates inter-organellar communication via calcium-dependent mitophagy to promote healthy ageing.
  23. EMC2 promotes triple negative breast cancer growth by protecting FDFT1 from endoplasmic reticulum associated degradation to impair ferroptosis susceptibility.
  24. Evaluation of machine learning-assisted directed evolution across diverse combinatorial landscapes.
  25. A rapid imaging-based screen for induced-proximity degraders identifies a potent degrader of oncoprotein SKP2.
  26. Dynein-2 requires HSP90 chaperone activity to ensure robust retrograde IFT and ciliogenesis.
  27. The citrullinating enzyme PADI4 governs progenitor cell proliferation and translation in developing hair follicles.
  28. Activity-Based Ubiquitin Probes Capture the Sulfenylated State of Deubiquitinases.
  29. TOM1 G307D variant impairs interaction with TOLLIP, autophagosome-lysosome fusion and regulation of innate immunity.
  30. Computational design of sequence-specific DNA-binding proteins.
  31. The IRE1/XBP1s Axis Regulates Innate Immune Responses in Conventional Dendritic Cells During ZIKV Infection.
  32. Overexpression of ATase1 and ATase2 disrupts the secretome and causes a progeria phenotype.
  33. Structural basis for LZTR1 recognition of RAS GTPases for degradation.
  34. Molecular exaptation by the integrin αI domain.
  35. S-nitrosylation of pVHL regulates β2 adrenergic receptor function.
  36. Paying attention to attention: High attention sites as indicators of protein family and function in language models.
  37. Smg5 enhances oligodendrocyte differentiation via nonsense-mediated mRNA decay of Hnrnpl variant transcripts.
  38. Amplifying antigen-induced cellular responses with proximity labelling.
  1. Cell Rep. 2025 Sep 09. pii: S2211-1247(25)01033-2. [Epub ahead of print]44(9): 116262
    Dynamic regulation of the oxidative stress response by the E3 ligase TRIP12.
    Andrew J Ingersoll, Devlon M McCloud, Jenny Y Hu, Michael Rape.
      Centered on the transcription factor NRF2 and its E3 ligase CUL3KEAP1, the oxidative stress response protects cells from damage by reactive oxygen species (ROS). Increasing ROS inhibits CUL3KEAP1 to stabilize NRF2 and elicit antioxidant gene expression, while cells recovering from stress rapidly turn over NRF2 again to prevent reductive stress and oxeiptosis-dependent death. How cells reinitiate NRF2 degradation after ROS have been cleared remains poorly understood. Here, we identify the essential E3 ligase TRIP12 as a crucial component of the oxidative stress response. TRIP12 is a ubiquitin chain elongation factor that cooperates with CUL3KEAP1 to ensure robust NRF2 degradation. In this manner, TRIP12 accelerates stress response silencing as oxidative stress is being resolved but limits NRF2 activation during stress. The need for dynamic control of NRF2 degradation therefore comes at the cost of diminished stress signaling, suggesting that TRIP12 inhibition could be used to treat degenerative pathologies characterized by ROS accumulation.
    Keywords:  CP: Cancer; CP: Molecular biology; CUL3; KEAP1; NRF2; ROS; TRIP12; oxidative stress response; proteasome; reactive oxygen species; ubiquitin
    DOI:  https://doi.org/10.1016/j.celrep.2025.116262
  2. Cell Rep. 2025 Sep 09. pii: S2211-1247(25)01037-X. [Epub ahead of print]44(9): 116266
    Coordination of autophagosome closure and release by the Alzheimer's disease-associated protein BIN1.
    Jennifer E Palmer, Claudia Puri, Ryan S O'Rourke, Sung Min Son, Chun Sang, Yu-Wen Alvin Huang, David C Rubinsztein.
      Autophagosome closure by the endosomal sorting complex required for transport (ESCRT) complex is a prerequisite for their dynamin 2 (DNM2)-dependent release from the recycling endosome and subsequent lysosomal clearance. However, the mechanism that coordinates autophagosome closure and release is unknown. We identified that the Alzheimer's disease-associated protein bridging integrator 1 (BIN1) is a critical mediator of this coordination. Prior to autophagosome closure, BIN1 is held at autophagosomes by ESCRT-III and inhibits DNM2. Once the autophagosome has closed and ESCRT-III disassembles, BIN1 is released, removing the inhibition of DNM2. This mechanism provides insight into the functional consequences of increased BIN1 expression, as this occurs in microglia with Alzheimer's disease risk-associated polymorphisms. We find that the overexpression of BIN1 microglial isoforms inhibits DNM2-mediated autophagosome release and autophagic clearance. This provides a coherent explanation for the increased Alzheimer's disease risk associated with BIN1, as impaired microglial autophagy alters phagocytosis and is associated with microglial senescence and neuroinflammation.
    Keywords:  Alzheimer’s disease; BIN1; CP: Cell biology; CP: Neuroscience; DNM2; ESCRT-III; autophagy; microglia
    DOI:  https://doi.org/10.1016/j.celrep.2025.116266
  3. J Cell Biol. 2025 Oct 06. pii: e202503166. [Epub ahead of print]224(10):
    ATG16L1 controls mammalian vacuolar proton ATPase.
    Thabata L A Duque, Masroor Paddar, Einar Trosdal, Ruheena Javed, Lee Allers, Michal H Mudd, Prithvi Akepati, Soumya R Mishra, Michelle Salemi, Brett Phinney, Shawn B Bratton, Thomas Wileman, Vojo Deretic.
      The mechanisms governing mammalian proton pump V-ATPase function are of fundamental and medical interest. The assembly and disassembly of cytoplasmic V1 domain with the membrane-embedded V0 domain of V-ATPase is a key aspect of V-ATPase localization and function. Here, we show that the mammalian protein ATG16L1, primarily appreciated for its role in canonical autophagy and in noncanonical membrane atg8ylation processes, controls V-ATPase. ATG16L1 knockout elevated V-ATPase activity, increased V1 presence on endomembranes, and increased the number of acidified intracellular compartments. ATG16L1's ability to efficiently bind V-ATPase was required for its inhibitory role in endolysosomal acidification and for control of Mycobacterium tuberculosis infection in mice. These findings uncover a hitherto unappreciated role of ATG16L1 in regulating V-ATPase, a key pump governing acidification and functionality of the endolysosomal system along with its physiological roles.
    DOI:  https://doi.org/10.1083/jcb.202503166
  4. Mol Cell. 2025 Sep 10. pii: S1097-2765(25)00707-5. [Epub ahead of print]
    α-Synuclein aggregates inhibit ESCRT-III through sequestration and collateral degradation.
    Cole S Sitron, Victoria A Trinkaus, Ana Galesic, Maximilian Garhammer, Patricia Yuste-Checa, Ulrich Dransfeld, Dennis Feigenbutz, Jiuchun Zhang, Larysa Ivashko, Irina Dudanova, J Wade Harper, F Ulrich Hartl.
      α-Synuclein aggregation is a hallmark of Parkinson's disease and related synucleinopathies. Extracellular α-synuclein fibrils enter naive cells via endocytosis, followed by transit into the cytoplasm to seed endogenous α-synuclein aggregation. Intracellular aggregates sequester numerous proteins, including subunits of the endosomal sorting complexes required for transport (ESCRT)-III system for endolysosome membrane repair, but the toxic effects of these events remain poorly understood. Using cellular models and in vitro reconstitution, we found that α-synuclein fibrils interact with a conserved α-helix in ESCRT-III proteins. This interaction sequesters ESCRT-III subunits and triggers their proteasomal destruction in a process of "collateral degradation." These twin mechanisms deplete the available ESCRT-III pool, initiating a toxic feedback loop. The ensuing loss of ESCRT function compromises endolysosome membranes, thereby facilitating escape of aggregate seeds into the cytoplasm, facilitating a "second wave" of templated aggregation and ESCRT-III sequestration. We suggest that collateral degradation and the triggering of self-perpetuating systems are general mechanisms of sequestration-induced proteotoxicity.
    Keywords:  CHMP2B; ESCRT; ESCRT-III; Parkinson’s disease; aggregation; lysosome; protein aggregate spreading; proteostasis; sequestration; α-synuclein
    DOI:  https://doi.org/10.1016/j.molcel.2025.08.022
  5. Trends Biochem Sci. 2025 Sep 11. pii: S0968-0004(25)00186-0. [Epub ahead of print]
    E3 ubiquitin ligases in signaling, disease, and therapeutics.
    Pirouz Ebadi, Caleb M Stratton, Shaun K Olsen.
      The ubiquitin-proteasome system (UPS) is a central regulator of protein turnover and signaling, with E3 ubiquitin ligases conferring substrate specificity and chain-type control. Recent advances have revealed new mechanistic classes of E3 ligases and expanded our understanding of their roles in disease, including cancer, neurodegeneration, and immune dysfunction. These insights have fueled the development of targeted protein degradation strategies that harness the UPS to eliminate disease-associated proteins. Approaches such as proteolysis-targeting chimeras (PROTACs), molecular glues, and antibody-based degraders are broadening the druggable proteome. Despite this progress, key challenges remain, including limited E3 ligase diversity, difficulties in degrader delivery, and resistance mechanisms. This review outlines recent advances in E3 ligase biology and therapeutic degradation, emphasizing opportunities to expand and refine UPS-targeted interventions.
    Keywords:  E3 ubiquitin ligase; PROTAC; molecular glues; targeted protein degradation; ubiquitin; ubiquitin–proteasome system
    DOI:  https://doi.org/10.1016/j.tibs.2025.07.009
  6. JCI Insight. 2025 Sep 09. pii: e191837. [Epub ahead of print]10(17):
    Thyroidal expression of ER molecular chaperone GRP170 is required for efficient TSH-mediated thyroid hormone synthesis.
    Xiaohan Zhang, Crystal Young, Xiao-Hui Liao, Samuel Refetoff, Stephanie M Mutchler, Jeffrey L Brodsky, Teresa M Buck, Peter Arvan.
      Intracellular trafficking of secretory and membrane proteins from the endoplasmic reticulum (ER) to the cell surface, via the secretory pathway, is crucial to the differentiated function of epithelial tissues. In the thyroid gland, a prerequisite for such trafficking is proper protein folding in the ER, assisted by an array of ER molecular chaperones. One of the most abundant of these chaperones, Glucose-Regulated-Protein-170 (GRP170, encoded by Hyou1), is a noncanonical hsp70-like family member. Thyroid follicular epithelial cells abundantly express GRP170, but the role of this abundant ER chaperone in thyrocytes remains unknown. Here, we have examined the effect of inducible Pax8-specific (thyroid and kidney) deficiency of GRP170 in mice, in parallel with siRNA-treated PCCL3 (rat) thyrocytes for knockdown of GRP170. Thyrocyte-specific loss of GRP170 in vivo triggers primary hypothyroidism with a deficient thyroidal response to Thyroid-Stimulating Hormone (TSH). In addition, knockdown of GRP170 in PCCL3 thyrocytes inhibits the folding and forward trafficking of TSH receptors to the cell surface. Taken together, our findings suggest that GRP170 contributes to the conformational maturation of TSH receptors and thyroid gland responsiveness to TSH, which is required for proper regulation of thyroid hormone synthesis.
    Keywords:  Cell biology; Endocrinology; Protein traffic; Thyroid disease
    DOI:  https://doi.org/10.1172/jci.insight.191837
  7. EMBO J. 2025 Sep 11.
    BLTP3A is associated with membranes of the late endocytic pathway and is an effector of CASM.
    Michael G Hanna, Hely O Rodriguez Cruz, Kenshiro Fujise, Yumei Wu, C Shan Xu, Song Pang, Zhuonging Li, Mara Monetti, Pietro De Camilli.
      Recent studies have identified a family of rod-shaped proteins thought to mediate lipid transfer at intracellular membrane contacts by a bridge-like mechanism. We show that one such protein, bridge-like lipid transfer protein 3A (BLTP3A)/UHRF1BP1 binds VAMP7 vesicles via its C-terminal region, and anchors them to lysosomes via its chorein domain-containing N-terminal region binding to Rab7. Upon lysosome damage, BLTP3A-positive vesicles rapidly (within minutes) dissociate from lysosomes. Lysosome damage is known to activate the CASM (Conjugation of ATG8 to Single Membranes) pathway, leading to lipidation and lysosomal recruitment of mammalian ATG8 (mATG8) proteins. We find that this process drives the reassociation of BLTP3A with damaged lysosomes via an interaction of its LIR motif with mATG8 which coincides with a dissociation from the vesicles. Our findings reveal that BLTP3A is an effector of CASM, potentially as part of a mechanism to help repair or minimize lysosome damage.
    Keywords:  BLTP3; LC3; Lysosome; Rab45; Urate Crystals
    DOI:  https://doi.org/10.1038/s44318-025-00543-9
  8. Cell Rep. 2025 Sep 09. pii: S2211-1247(25)01028-9. [Epub ahead of print]44(9): 116257
    The E3 ubiquitin ligase SIAH1 targets a bacterial pore-forming toxin to facilitate xenophagy.
    Min Wu, Xin Hu, Junpei Iibushi, Kazunori Murase, Takashi Nozawa, Ichiro Nakagawa.
      Xenophagy, one form of selective autophagy, recognizes and eliminates the invading pathogen through the ubiquitination of bacterial surface components. Xenophagy is initiated by the damage of bacteria-surrounding endosomes by bacterial toxins; however, whether the host targets these xenophagy-inducible secretory factors to recognize bacteria remains unclear. Here, we report that E3 ligase SIAH1 recognizes and ubiquitinates streptolysin O (SLO), a pore-forming toxin secreted by group A Streptococcus (GAS). SIAH1 specifically recognizes the PSVP motif in SLO and mediates K48-linked polyubiquitination at Lys464. SIAH1 depletion significantly reduced GAS ubiquitination, impaired autophagosome formation, and enhanced bacterial survival. Studies with ATG16L1 and FIP200 knockout cell lines suggested that anti-GAS defense involves sequential deployment of the LC3-associated phagocytosis-like (LAP-like) process followed by canonical autophagy, and SIAH1 coordinates these pathways. Our findings reveal SIAH1's crucial role in bacterial toxin recognition and demonstrate a mechanism in which bacterial virulence factors themselves become targets of host xenophagy machinery.
    Keywords:  CP: Microbiology; E3 ligase; LC3-associated phagocytosis; SIAH 1; group A Streptococcus; streptolysin O; ubiquitin; xenophagy
    DOI:  https://doi.org/10.1016/j.celrep.2025.116257
  9. STAR Protoc. 2025 Sep 10. pii: S2666-1667(25)00479-4. [Epub ahead of print]6(3): 104073
    Protocol for monitoring mRNA translation and degradation in human cell-free lysates.
    Nino Schwaller, Evangelos D Karousis.
      Cell-free translation is a powerful tool for studying protein synthesis. Here, we present a protocol for monitoring mRNA translation and degradation using a human cell-free translation system. The protocol includes instructions for preparing translation-competent lysates via dual centrifugation, generating and capping reporter mRNAs compatible with luciferase assays, performing cell-free translation reactions, and assessing mRNA stability by northern blotting. The protocol supports comparative analysis of translation efficiency and mRNA decay for various 5' UTRs and viral RNA elements. For complete details on the use and execution of this protocol, please refer to Gurzeler et al.1 and Bäumlin et al.2.
    Keywords:  Biotechnology and bioengineering; Molecular Biology; Molecular/Chemical probes; cell Biology; cell culture; gene Expression
    DOI:  https://doi.org/10.1016/j.xpro.2025.104073
  10. Genetics. 2025 Sep 11. pii: iyaf190. [Epub ahead of print]
    A novel role for the E2F transcription factor and the ER stress sensor IRE1 in cytoplasmic DNA accumulation.
    Arghya Das, Yining Li, Yiting Fan, Nam-Sung Moon.
      The E2F family of transcription factors are key regulators of the cell cycle in all metazoans. While they are primarily known for their role in cell cycle progression, E2Fs also play broader roles in cellular physiology, including the maintenance of exocrine tissue homeostasis. However, the underlying mechanisms that render exocrine cells particularly sensitive to E2F deregulation remain poorly understood. The Drosophila larval salivary gland (SG), like its mammalian counterpart, is an exocrine tissue that produces large quantities of "glue proteins" in the endoplasmic reticulum (ER). Here, we show that E2F activity is important for the exocrine function of the Drosophila SG. The loss of de2f1b, an alternatively spliced isoform of Drosophila E2F1, leads to elevated DNA damage and accumulation of cytoplasmic DNA (cytoDNA) in the SGs. Surprisingly, we found that IRE1, a key sensor of the unfolded protein response, is required for ER homeostasis during development that is critical for preventing cytoDNA accumulation in the SG. Importantly, we found evidence demonstrating that IRE1 activity is attenuated in de2f1b-deficient SGs, contributing to ER dysfunction and cytoDNA accumulation. Together, these findings reveal an unanticipated link between ER homeostasis and cytoDNA processing and offer mechanistic insights into why exocrine tissues are particularly vulnerable to E2F deregulation.
    Keywords:  Cytoplasmic DNA; Drosophila; E2F; Endocycle; Endoplasmic reticulum; IRE1; Unfolded Protein Response
    DOI:  https://doi.org/10.1093/genetics/iyaf190
  11. Dev Cell. 2025 Sep 11. pii: S1534-5807(25)00533-7. [Epub ahead of print]
    Biomolecular condensates of ATG18 reshape ER for autophagy in plants.
    Yang Shao, Xunzheng Li, Benhui Shi, Songyang Wang, Zisheng Luo, Yanqun Xu, Baolei Li, Shuqing Feng, Li Liang, Huanquan Zheng, Jiaqi Sun.
      Autophagosomes originate from and maintain association with the endoplasmic reticulum (ER) during their formation, yet how these processes are molecularly coordinated in plants remains poorly understood. Here, we demonstrate that Arabidopsis autophagy-related protein 18a (ATG18a), a key organizer of early autophagosome formation, undergoes phase separation to form biomolecular condensates on the ER membrane, which progress from highly mobile droplets to stable ring-like structures, while the ER is reshaped. We discovered that ATG18a condensates work together with ROOT HAIR DEFECTIVE3 (RHD3), an ER membrane-shaping protein, with RABC1 serving as a molecular linker between them. Importantly, RABC1 facilitates both RHD3 assembly necessary for the formation of ring-like ER structures and its interaction with ATG18a condensates. These findings reveal a mechanism whereby biomolecular condensates work together with membrane-shaping proteins to reshape specialized membrane domains through wetting interactions, providing an insight into autophagosome formation in plant stress responses.
    Keywords:  ATG18; RABC1; RHD3; autophagosomes; autophagy; biomolecular condensates; plant response to stresses; salt stress; the ER; wetting interaction
    DOI:  https://doi.org/10.1016/j.devcel.2025.08.013
  12. Elife. 2025 Sep 10. pii: RP94310. [Epub ahead of print]13
    Inhibition of the UFD-1-NPL-4 complex triggers an aberrant immune response in Caenorhabditis elegans.
    Rajneesh Rao, Alejandro Aballay, Jogender Singh.
      The UFD-1 (ubiquitin fusion degradation 1)-NPL-4 (nuclear protein localization homolog 4) heterodimer is involved in extracting ubiquitinated proteins from several plasma membrane locations, including the endoplasmic reticulum. This heterodimer complex helps in the degradation of ubiquitinated proteins via the proteasome with the help of the AAA+ATPase CDC-48. While the ubiquitin-proteasome system is known to have important roles in maintaining innate immune responses, the role of the UFD-1-NPL-4 complex in regulating immunity remains elusive. In this study, we investigate the role of the UFD-1-NPL-4 complex in maintaining Caenorhabditis elegans innate immune responses. Inhibition of the UFD-1-NPL-4 complex activates an aberrant immune response that reduces the survival of the wild-type worms on the pathogenic bacterium Pseudomonas aeruginosa despite diminishing colonization of the gut with the bacterium. This aberrant immune response improves the survival of severely immunocompromised worms on pathogenic bacteria but is detrimental on nonpathogenic bacteria. Transcriptomics studies reveal that the GATA transcription factor ELT-2 mediates the aberrant immune response upon inhibition of the UFD-1-NPL-4 complex. Collectively, our findings show that inhibition of the UFD-1-NPL-4 complex triggers an aberrant immune response that is detrimental to immunocompetent worms under infection conditions but can be advantageous for immunocompromised worms.
    Keywords:  C. elegans; ERAD; GATA transcription factor; colonization; genetics; genomics; immune response; immunology; inflammation; ubiquitin-proteasome system
    DOI:  https://doi.org/10.7554/eLife.94310
  13. Open Biol. 2025 Sep;15(9): 250164
    The E3 ligase MEX3B forms a tripartite complex with Rest and Hotair to determine the proliferative capacity of neural progenitor cells.
    Kamakshi Garg, Gourav Sharma, Sarbani Samaddar, Sourav Banerjee.
      E3 ubiquitin ligases regulate the cellular proteome via proteasome-dependent protein degradation; however, there exist limited studies outlining their non-canonical functions. RNA-binding ubiquitin ligases (RBULs) represent a subset of E3 ligases that harbour RNA-binding domains, making them uniquely positioned to function as both RNA-binding proteins and E3 ligases. Our initial microarray screen for E3 ligases from mouse cortical neural progenitor cells identified MEX3B, a known RNA-binding ubiquitin ligase, to be differentially expressed. Here, we characterize the non-canonical role of MEX3B in the context of neural proliferation. We find that MEX3B is significantly reduced following the differentiation of neural progenitor cells (NPCs). The knockdown of MEX3B blocks the proliferative state of NPCs and leads to the enhancement of neurite length and dendrite branching. We observed that MEX3B regulates the stability of Rest mRNA in proliferative NPCs. Mechanistically, MEX3B interacts with Rest mRNA and the lncRNA Hotair to form a tripartite complex in the presence of basic fibroblast growth factor (bFGF). Loss of Hotair disrupts this complex; conversely, MEX3B RNAi significantly reduces Hotair abundance. Rest mRNA levels remain unaffected by Hotair knockdown, suggesting that the latter acts as a scaffold to facilitate bFGF-dependent MEX3B-Rest interaction in the MEX3B-Rest-Hotair tripartite axis. Our study demonstrates an RNA-driven post-transcriptional mechanism underlying NPC proliferation.
    Keywords:  MEX3B; REST; RNA-binding ubiquitin ligase; long non-coding RNA; neural progenitor cells
    DOI:  https://doi.org/10.1098/rsob.250164
  14. EMBO J. 2025 Sep 08.
    Mammalian mitohormesis: from mitochondrial stressors to organismal benefits.
    Amanda L Gunawan, Irene Liparulo, Andreas Stahl.
      A variety of stressors, including environmental insults, pathological conditions, and transition states, constantly challenge cells that, in turn, activate adaptive responses to maintain homeostasis. Mitochondria have pivotal roles in orchestrating these responses that influence not only cellular energy production but also broader physiological processes. Mitochondria contribute to stress adaptation through mechanisms including induction of the mitochondrial unfolded protein response (UPRmt) and the integrated stress response (ISR). These responses are essential for managing mitochondrial proteostasis and restoring cellular function, with each being tailored to specific stressors and cellular milieus. While excessive stress can lead to maladaptive responses, mitohormesis refers to the beneficial effects of low-level mitochondrial stress. Initially studied in invertebrates and cell cultures, recent research has expanded to mammalian models of mitohormesis. In this literature review, we describe the current landscape of mammalian mitohormesis research and identify mechanistic patterns that result in local, systemic, or interorgan mitohormesis. These investigations reveal the potential for targeting mitohormesis for therapeutic benefit and can transform the treatment of diseases commonly associated with mitochondrial stress in humans.
    Keywords:  Integrated Stress Response; Mammalian Models; Mitochondrial Retrograde Signaling; Mitochondrial Unfolded Protein Response (UPRmt); Mitohormesis
    DOI:  https://doi.org/10.1038/s44318-025-00549-3
  15. Mol Cell. 2025 Sep 10. pii: S1097-2765(25)00700-2. [Epub ahead of print]
    Rapid UPF1 depletion illuminates the temporal dynamics of the NMD-regulated human transcriptome.
    Volker Boehm, Damaris Wallmeroth, Paul O Wulf, Oliver Popp, Luiz Gustavo Teixeira Alves, Lucie Reinecke, Maximilian Riedel, Emanuel Wyler, Marek Franitza, Kerstin Becker, Karina Polkovnychenko, Simone Del Giudice, Nouhad Benlasfer, Philipp Mertins, Markus Landthaler, Niels H Gehring.
      The RNA helicase UPF1 shapes the transcriptome as the core factor of nonsense-mediated mRNA decay (NMD). The essential role of UPF1 in human cells has impeded efforts to delineate its directly regulated transcripts and molecular function. To investigate the effects of rapid UPF1 depletion, we engineered human cell lines with endogenous UPF1 fused to conditional degron tags. Temporal-resolution transcriptomic analyses identified direct target mRNAs, consisting predominantly of NMD substrates that are mostly stabilized within hours of UPF1 depletion. By integrating long-read sequencing and ribosome profiling data, we defined the consolidated NMD-regulated human transcriptome (NMDRHT), uncovering previously unannotated transcripts and establishing alternative splicing as a major contributor of NMD-targeted mRNAs. Additionally, we identified non-canonical NMD events that lack indication of being driven by other UPF1-dependent degradation routes. Our work refines the role of the post-transcriptional regulator UPF1 and introduces an experimentally validated NMD-regulated transcriptome as a navigable resource at https://nmdrht.uni-koeln.de.
    Keywords:  NMD; RNA surveillance; UPF1; alternative splicing; degron; long-read sequencing; mRNA degradation; nonsense-mediated mRNA decay; transcriptome; translation
    DOI:  https://doi.org/10.1016/j.molcel.2025.08.015
  16. Mol Biol Cell. 2025 Sep 10. mbcE25060279
    Loss of CHMP2A implicates an ordered assembly of ESCRT-III proteins during cytokinetic abscission.
    Nikita Kamenetsky, Dikla Nachmias, Suman Khan, Ori Avinoam, Itay Hazan, Alexander Upcher, Natalie Elia.
      The ESCRT machinery mediates membrane remodeling in fundamental cellular processes including cytokinesis, endosomal sorting, nuclear envelope reformation, and membrane repair. Membrane constriction and scission is driven by the filament-forming ESCRT-III complex and the AAA-ATPase VPS4. While ESCRT-III-driven membrane scission is generally established, the mechanisms governing the assembly and coordination of its twelve mammalian isoforms in cells remain poorly understood. Here, we examined the spatial organization and interdependence of ESCRT-III subunits during mammalian cytokinetic abscission by depleting CHMP2A, a core ESCRT-III component. Using live cell imaging, structured illumination microscopy (SIM) and correlative light-electron microscopy (CLEM), we show that CHMP2A knockout cells display a significant delay-but not failure-in abscission, accompanied by distinct mislocalization phenotypes across ESCRT-III subunits. While IST1 and CHMP2B were minimally disrupted, CHMP4B, CHMP3, and CHMP1B display progressively severe organization defects at the abscission site. Dual-protein imaging reveals disrupted coordination between ESCRT-III subunits in individual CHMP2A-deficient cells, supporting an ordered assembly of ESCRT-III subunits in cytokinetic abscission. Together, our findings provide the first in vivo evidence for hierarchical assembly of ESCRT-III subunits during ESCRT-mediated membrane remodeling and identify CHMP2A as a key organizer of ESCRT-III architecture essential for timely membrane abscission. [Media: see text] [Media: see text].
    DOI:  https://doi.org/10.1091/mbc.E25-06-0279
  17. Sci Adv. 2025 Sep 12. 11(37): eadx0952
    Giant KASH proteins and ribosomes establish distinct cytoplasmic biophysical properties in vivo.
    Xiangyi Ding, Hongyan Hao, Daniel Elnatan, Patrick Neo Alinaya, Shilpi Kalra, Abby Kaur, Sweta Kumari, Liam J Holt, G W Gant Luxton, Daniel A Starr.
      Understanding how cells control their biophysical properties during development remains a fundamental challenge. While macromolecular crowding affects multiple cellular processes in single cells, its regulation in living animals remains poorly understood. Using genetically encoded multimeric nanoparticles for in vivo rheology, we found that Caenorhabditis elegans tissues maintain mesoscale properties that differ from those observed across diverse systems, including bacteria, yeast species, and cultured mammalian cells. We identified two conserved mechanisms controlling particle mobility: Ribosome concentration, a known regulator of cytoplasmic crowding, works in concert with a previously unknown function for the giant KASH (Klarsicht/ANC-1/SYNE homology) protein ANC-1 in providing structural constraints through associating with the endoplasmic reticulum. These findings reveal mechanisms by which tissues establish and maintain distinct mesoscale properties, with implications for understanding cellular organization across species.
    DOI:  https://doi.org/10.1126/sciadv.adx0952
  18. Cell Rep. 2025 Sep 10. pii: S2211-1247(25)01020-4. [Epub ahead of print]44(9): 116249
    The hexosamine biosynthetic pathway drives tumor immune evasion via translational control of PD-L1 at the elongation level.
    Bo Liu, Yingying Wu, Anyi Xiang, Yiyi Yu, Shushu Song, Ying Zhang, Jianxin Gu, Haojie Lu, Ping Xu, Fenglin Liu, Yuanyuan Ruan.
      Cancer cells reprogram cellular energetics to drive tumorigenesis and escape immunosurveillance. Nevertheless, how this is molecularly connected remains largely undefined. The hexosamine biosynthetic pathway (HBP) serves as a critical metabolic node in cancer cells that provides the basis for protein glycosylation. Herein, we show that HBP flux inhibition by knocking out its rate-limiting enzyme GFAT1 suppressed tumor growth and stimulated cytotoxic CD8+ T lymphocyte infiltration in a colorectal cancer model. GFAT1 induced the expression of the immune checkpoint PD-L1 at the translational level by bypassing signal peptide-mediated translation elongation arrest. Proteomic and glycoproteomic screening indicated that GFAT1 facilitated the N-linked glycosylation and protein expression of integrin α2/α3 subunits, leading to FAK activation and elongation factor eEF1A2 upregulation. Pharmacological inhibition of HBP noticeably enhanced the efficacy of immune checkpoint blockade in vivo. Together, these findings unravel how immune checkpoint proteins are manipulated by metabolic dysregulation, which can be exploited as metabolic vulnerability for improving immunotherapies.
    Keywords:  CP: Cancer; PD-L1; glycosylation; hexosamine biosynthetic pathway; immune evasion; protein synthesis
    DOI:  https://doi.org/10.1016/j.celrep.2025.116249
  19. Sci Adv. 2025 Sep 12. 11(37): eadw9158
    Bi-handed assembly chaperones regulate protein complex assembly through an intramolecular handover mechanism.
    Jingyi Wu, Chun Wan, Yuan Tian, Yan Ouyang, Harrison Puscher, Suzhao Li, Qian Yin, Jingshi Shen.
      A critical yet challenging step in protein complex assembly is the formation of a dimeric intermediate that serves as a seed for incorporating additional subunits. We hypothesized that this step could be facilitated by "bi-handed" chaperones that recognize two different subunits through distinct domains (hands). However, whether such chaperones exist remained unknown. Here, we identify AAGAB as a bona fide bi-handed chaperone. AAGAB uses its C-terminal domain (CTD) to bind the α subunit and its GTPase-like domain (GD) to bind the σ2 subunit of the AP2 adaptor complex, a central player in membrane trafficking. AAGAB first recruits α via its CTD; σ2 then joins through interaction with α, forming a conformationally immature α:σ2 hemicomplex at the CTD. This hemicomplex is subsequently transferred to the GD via a GD:σ2 binding interface, accompanied by conformational maturation. These findings establish AAGAB as the founding member of a bi-handed chaperone family and reveal an intramolecular handover mechanism that underlies their mode of action.
    DOI:  https://doi.org/10.1126/sciadv.adw9158
  20. Nat Biotechnol. 2025 Sep 09.
    Genome-wide mapping of RNA-protein associations through sequencing.
    Zhijie Qi, Shuanghong Xue, Junchen Chen, Wenxin Zhao, Kara Johnson, Xingzhao Wen, John Lalith Charles Richard, Pei Lin, Sheng Zhong.
      RNA-protein interactions critically regulate gene expression and cellular processes, yet their comprehensive mapping remains challenging due to their structural diversity. We introduce PRIM-seq (protein-RNA interaction mapping by sequencing), a method for concurrent de novo identification of RNA-binding proteins and their associated RNAs. PRIM-seq generates unique chimeric DNA sequences by proximity ligation of RNAs with protein-linked DNA barcodes, which are subsequently decoded through sequencing. We apply PRIM-seq to two human cell lines and construct a human RNA-protein association network (HuRPA), encompassing >350,000 associations involving ~7,000 RNAs and ~11,000 proteins, including 2,610 proteins that each interact with at least 10 distinct RNAs. We experimentally validate the tumorigenesis-associated lincRNA LINC00339, the RNA with the highest number of protein associations in HuRPA, as a protein-associated RNA. We further validate the RNA-associating abilities of chromatin-conformation regulators SMC1A, SMC3 and RAD21, as well as the metabolic enzyme PHGDH. PRIM-seq enables systematic discovery and prioritization of RNA-binding proteins and their targets without gene- or protein-specific reagents.
    DOI:  https://doi.org/10.1038/s41587-025-02780-z
  21. Biophys Rep. 2025 Aug 31. 11(4): 219-231
    TurboID-based mapping of organelle membrane protein interactomes with digitonin-permeabilization.
    Yang Sun, Lan Yang, Jingzi Zhang, Pu Tian, Shue Chen, Lei Fang, Zhi Hong.
      Protein-protein interactions at organelle membranes bridge organelles in close proximity, facilitating regulated metabolite exchange and maintaining cellular homeostasis. Enzyme-catalyzed proximity labeling (PL) has been widely used to uncover the molecular composition of these interactions, but excessive labeling of irrelevant cytosolic proteins complicates data analysis. To address this, we developed a streamlined protocol that combines the TurboID system with digitonin-permeabilization to efficiently map protein interactions at organelle membranes in live mammalian cells. Digitonin selectively permeabilizes the plasma membrane, removing cytosolic proteins while preserving the integrity of inner membranes like the ER and mitochondria. This approach enhances spatial resolution in proteo-mic analysis, enabling a more precise map for protein interactome. Using this method, we successfully achieved proximal labeling of ER-localized proteins REEP1 and REEP6 to decipher their interaction networks, demonstrating its applicability for studying membrane-associated interactions with greater clarity and reduced contamination.
    Keywords:  Digitonin-permeabilization; Organelle interactomes; Protein–protein interactions; Proximity labeling; REEP1/REEP6
    DOI:  https://doi.org/10.52601/bpr.2025.240051
  22. Autophagy. 2025 Sep 13.
    Urolithin a modulates inter-organellar communication via calcium-dependent mitophagy to promote healthy ageing.
    Antonis Roussos, Katerina Kitopoulou, Fivos Borbolis, Christina Ploumi, Despoina D Gianniou, Zhiquan Li, Haijun He, Eleni Tsakiri, Helena Borland, Ioannis K Kostakis, Martina Samiotaki, Ioannis P Trougakos, Vilhelm A Bohr, Konstantinos Palikaras.
      Mitochondrial dysfunction and impaired mitophagy are hallmarks of aging and age-related pathologies. Disrupted inter-organellar communication among mitochondria, endoplasmic reticulum (ER), and lysosomes, further contributes to cellular dysfunction. While mitophagy has emerged as a promising target for neuroprotection and geroprotection, its potential to restore age-associated defects in organellar crosstalk remains unclear. Here, we show that mitophagy deficiency deregulates the morphology and homeostasis of mitochondria, ER and lysosomes, mirroring age-related alterations. In contrast, urolithin A (UA), a gut-derived metabolite and potent mitophagy inducer, restores inter-organellar communication via calcium signaling, thereby, promoting mitophagy, healthspan and longevity. Our multi-omic analyses reveal that UA reorganizes ER, mitochondrial and lysosomal networks, linking inter-organellar dynamics to mitochondrial quality control. In C. elegans, UA induces calcium release from the ER, enhances lysosomal activity, and drives DRP-1/DNM1L/DRP1-mediated mitochondrial fission, culminating in efficient mitophagy. Calcium chelation abolishes UA-induced mitophagy, blocking its beneficial impact on muscle function and lifespan, underscoring the critical role of calcium signaling in UA's geroprotective effects. Furthermore, UA-induced calcium elevation activates mitochondrial biogenesis via UNC-43/CAMK2D and SKN-1/NFE2L2/Nrf2 pathways, which are both essential for healthspan and lifespan extension. Similarly, in mammalian cells, UA increases intracellular calcium, enhances mitophagy and mitochondrial metabolism, and mitigates stress-induced senescence in a calcium-dependent manner. Our findings uncover a conserved mechanism by which UA-induced mitophagy restores inter-organellar communication, supporting cellular homeostasis and organismal health.
    Keywords:  Calcium; ER; cellular senescence; geroprotection; lysosome; mitochondria
    DOI:  https://doi.org/10.1080/15548627.2025.2561073
  23. Oncogene. 2025 Sep 10.
    EMC2 promotes triple negative breast cancer growth by protecting FDFT1 from endoplasmic reticulum associated degradation to impair ferroptosis susceptibility.
    Xinrui Dong, Huijuan Dai, Linli Yao, Yanping Lin, Yaohui Wang, Ye Li, Xueli Zhang, Liheng Zhou, Jinsong Lu, Wenjin Yin.
      Cholesterol biosynthesis is more activated in triple negative breast cancer (TNBC) than in other subtype breast cancer and plays essential role in facilitating TNBC. However, the regulatory network and how cholesterol biosynthesis contribute to TNBC development and progression are not well elucidated. Here, we found that reticulum membrane protein complex 2 (EMC2) is highly expressed in TNBC and predicts short survival of patients. In vitro and in vivo experiments displayed that EMC2 could promote TNBC growth. We also displayed that EMC2 could increase intracellular cholesterol biosynthesis by regulating Farnesyltransferase 1 (FDFT1) expression. Mechanistically, we validated that EMC2 interacted with heat shock protein 90(HSP90) to sustain FDFT1 protein quality and correctly located in the ER membrane through protecting it from endoplasmic reticulum associated degradation (ERAD). Furthermore, EMC2 decreased TNBC cell ferroptosis susceptibility through elevating intracellular cholesterol contents. Collectively, our findings shed a new insight that EMC2 is critical for boosting cholesterol biosynthesis and ferroptosis resistance. Targeting EMC2 could be a promising novel therapeutic target for TNBC treatment.
    DOI:  https://doi.org/10.1038/s41388-025-03545-3
  24. Cell Syst. 2025 Sep 10. pii: S2405-4712(25)00220-0. [Epub ahead of print] 101387
    Evaluation of machine learning-assisted directed evolution across diverse combinatorial landscapes.
    Francesca-Zhoufan Li, Jason Yang, Kadina E Johnston, Emre Gürsoy, Yisong Yue, Frances H Arnold.
      Various machine learning-assisted directed evolution (MLDE) strategies have been shown to identify high-fitness protein variants more efficiently than typical directed evolution approaches. However, limited understanding of the factors influencing MLDE performance across diverse proteins has hindered optimal strategy selection for wet-lab campaigns. To address this, we systematically analyzed multiple MLDE strategies, including active learning and focused training using six distinct zero-shot predictors, across 16 diverse protein fitness landscapes. By quantifying landscape navigability with six attributes, we found that MLDE offers a greater advantage on landscapes that are more challenging for directed evolution, especially when focused training is combined with active learning. Despite varying levels of advantage across landscapes, focused training with zero-shot predictors leveraging distinct evolutionary, structural, and stability knowledge sources consistently outperforms random sampling for both binding interactions and enzyme activities. Our findings provide practical guidelines for selecting MLDE strategies for protein engineering. A record of this paper's transparent peer review process is included in the supplemental information.
    Keywords:  combinatorial mutagenesis; directed evolution; epistasis; fitness prediction; machine learning; protein engineering; zero-shot predictor
    DOI:  https://doi.org/10.1016/j.cels.2025.101387
  25. Nat Biotechnol. 2025 Sep 10.
    A rapid imaging-based screen for induced-proximity degraders identifies a potent degrader of oncoprotein SKP2.
    Yankai Chu, Shishuang Chen, Mingyang Yang, Yin Chen, Huiling Fang, Pinyu Huang, Yueru Xie, Chi Sun, Yun Chen, Baoding Zhang, Li Li, Hongchen Mu, Ding Song, Wentao Cheng, Chao Wang, Wei Jiang, Xiaolan Xu, Zhengjin He, Shuo Chen, Mingxian Liu, Jingchuan Ma, Man Yang, Jiaqi Cao, Jing Gao, Jiali Shen, Lulu Zhang, Yu Bai, Zheyi Liu, Jingyao Chen, Siqi Dai, Yi Arial Zeng, Yun Zhao, Hu Zhou, Chong Chen, Huanwei Ru, Li Tan, Ximin Chi, Fangjun Wang, Daming Gao, Moubin Lin, Xianming Deng, Hai Jiang.
      Targeted protein degraders hold potential as therapeutic agents to target conventionally 'undruggable' proteins. Here, we develop a high-throughput screen, DEath FUSion Escaper (DEFUSE), to identify small-molecule protein degraders. By conjugating the protein of interest to a fast-acting triggerable death protein, this approach translates target protein degradation into a cell survival phenotype to illustrate the presence of degraders. Using this method, we discovered a small molecule (SKPer1) that triggers degradation of the oncoprotein SKP2 and specifically kills SKP2-expressing cancer cells. Mechanistically, SKPer1 acts as an induced-proximity degrader by inducing interaction between SKP2 and an E3 ligase, STUB1, resulting in SKP2 ubiquitination and degradation. SKPer1 exhibits substantial tumour suppression with good safety profiles in vivo. We further show that a sequence of ten amino acids from SKP2 can serve as a versatile degradation tag.
    DOI:  https://doi.org/10.1038/s41587-025-02793-8
  26. J Cell Sci. 2025 Sep 11. pii: jcs.264034. [Epub ahead of print]
    Dynein-2 requires HSP90 chaperone activity to ensure robust retrograde IFT and ciliogenesis.
    Tiago J Dantas, Diogo M Abreu, Maria J G De-Castro, Ana R G De-Castro, Noopur V Khobrekar, Sónia A Rocha, Carla M C Abreu.
      The microtubule motor dynein-2 is responsible for retrograde intraflagellar transport (IFT), a process critical for cilia assembly and cilium-dependent signaling. Mutations in genes encoding dynein-2 subunits interfere with ciliogenesis and are among the most frequent causes of skeletal ciliopathies. Despite its importance, little is known regarding dynein-2 assembly and regulation. Here, we identify the molecular HSP90 chaperone as a critical regulator of dynein-2 complex stability and function. Pharmacological inhibition of HSP90 causes a severe decrease in the levels of dynein-2 subunits, without detectable alterations in cytoplasmic dynein-1 and the anterograde IFT kinesin-2 motor KIF3A. Consistent with disrupted dynein-2 function, HSP90 inhibition progressively disrupts retrograde IFT and severely impairs ciliogenesis. We demonstrate that HSP90 associates with the dynein-2 complex, promoting its assembly and stabilization. These results establish dynein-2 as a novel HSP90 client and provide important mechanistic insights into the regulation of dynein-2 assembly.
    Keywords:  Cilia; Intraflagellar transport; Molecular chaperone; Molecular motors
    DOI:  https://doi.org/10.1242/jcs.264034
  27. Sci Adv. 2025 Sep 12. 11(37): eadx4511
    The citrullinating enzyme PADI4 governs progenitor cell proliferation and translation in developing hair follicles.
    Kim Vikhe Patil, Nil Campamà Sanz, Kylie Hin-Man Mak, Wei Yang, Karl Annusver, Jasson Makkar, David Grommisch, Li Lei, Priyanka Sharma, Ryan R Driskell, Maria Kasper, Maria Genander.
      Posttranslational protein modifications have emerged as a mechanism regulating progenitor cell state transitions during tissue formation. Herein, we exploit the stereotyped hair follicle development to delineate the function of PADI4, an enzyme converting peptidylarginine to citrulline. Single-cell sequencing places Padi4 in both progenitor and differentiated hair lineage cells and indicates that PADI4 acts to repress transcription during hair follicle development. We establish PADI4 as a negative regulator of proliferation, acting on LEF1-positive hair shaft committed progenitor cells. Mechanistically, PADI4 citrullinates proteins associated with mRNA processing and ribosomal biogenesis, and lack of PADI4 promotes protein synthesis and ribosomal RNA transcription in vivo. Characterizing key translational effectors, we demonstrate that PADI4 citrullinates the translational repressor 4E-BP1 and reveal a cross-talk between PADI4 activity and 4E-BP1 phosphorylation. This work sheds light on how posttranslational modifications affect progenitor cell states and tissue formation.
    DOI:  https://doi.org/10.1126/sciadv.adx4511
  28. Angew Chem Int Ed Engl. 2025 Sep 07. e202512311
    Activity-Based Ubiquitin Probes Capture the Sulfenylated State of Deubiquitinases.
    Zian Chen, Guorui Li, Jiahao Zhang, Xiaoyu Xu, Qipeng Yan, Qingyu Wang, Qinfeng Zhang, Jing Huang.
      Activity-based ubiquitin probes (Ub-ABPs) are powerful tools for studying the functional landscape of deubiquitinases (DUBs). While most existing Ub probes have focused on examining the native state of DUBs, oxidative stress, especially in cancer and inflammatory contexts, can oxidize the catalytic cysteine of DUBs, significantly altering their activity. Here, we developed three novel ubiquitin-based activity probes (Ub-ABPs) to selectively trap the sulfenylated form of deubiquitinases (DUB-SOH). These probes employ ubiquitin as the recognition element and incorporate distinct warheads: an electrophilic norbornene moiety (Biotin-Ub-NMA) or dimedone-derived cyclic C-nucleophiles (Biotin-Ub-PRD and Biotin-Ub75-DYn-2), enabling covalent capture of oxidized cysteine residues. Of these, Biotin-Ub-PRD and Biotin-Ub75-DYn-2 successfully labeled DUB-SOH, highlighting the importance of proper probe-substrate interaction for effective trapping. Optimization of the ubiquitin length showed that the Ub74 variant displayed enhanced affinity toward DUB-SOH. Biotin-Ub74-DYn-2 enabled enrichment and identification of DUB-SOH targets via immunocapture and label-free quantitative proteomics. Collectively, these sulfenic acid-targeting Ub-ABPs represent versatile tools for elucidating redox-dependent DUB regulation, with potential applications in understanding redox dysregulation in disease contexts.
    Keywords:  Activity‐based ubiquitin probes; Deubiquitinase; Post‐translational modification; Redox signalling; Sulfenylation
    DOI:  https://doi.org/10.1002/anie.202512311
  29. Dis Model Mech. 2025 Sep 12. pii: dmm.052140. [Epub ahead of print]
    TOM1 G307D variant impairs interaction with TOLLIP, autophagosome-lysosome fusion and regulation of innate immunity.
    Heljä K M Lång, Tiffany G Roach, Maarit Hölttä, Salla Keskitalo, Markku Varjosalo, Kaarina Heiskanen, Megan V Collins, Mikko R J Seppänen, Daniel G S Capelluto, Elina Ikonen, Samppa J Ryhänen.
      A recently described G307D variant of the endosomal adaptor protein TOM1 causes severe early-onset multiorgan autoimmunity and combined immunodeficiency. By combining biophysical, biochemical, and cell culture experiments, we show that the variant causes a defect in the interaction between TOM1 and TOLLIP, another adaptor protein involved in cargo trafficking and regulation of innate immunity. The G307D variant impairs the ability of TOM1 to reduce TOLLIP's phosphatidylinositol 3-phosphate binding, an important regulatory mechanism for cargo trafficking commitment for both proteins. Our experiments using TOM1 G307D patient cells suggest that the variant affects autophagy seen as an aggravated response to amino acid starvation and accumulation of autophagosomes due to autophagosome-lysosome fusion defect. In addition, inflammatory pathways showed excessive activation in TOM1 G307D patient cells. Our data suggest that the interaction between TOM1 and TOLLIP has a role in the regulation of the human immune system and highlight the importance of fundamental cellular functions, such as cargo trafficking, in controlling immune responses. Our study also provides insights into the caveats of immunomodulatory and stem cell therapies in patients with TOM1 pathogenic variants.
    Keywords:  Autophagy; Inborn errors of immunity; TOLLIP; TOM1
    DOI:  https://doi.org/10.1242/dmm.052140
  30. Nat Struct Mol Biol. 2025 Sep 12.
    Computational design of sequence-specific DNA-binding proteins.
    Cameron J Glasscock, Robert J Pecoraro, Ryan McHugh, Lindsey A Doyle, Wei Chen, Olivier Boivin, Beau Lonnquist, Emily Na, Yuliya Politanska, Hugh K Haddox, David Cox, Christoffer Norn, Brian Coventry, Inna Goreshnik, Dionne Vafeados, Gyu Rie Lee, Raluca Gordân, Barry L Stoddard, Frank DiMaio, David Baker.
      Sequence-specific DNA-binding proteins (DBPs) have critical roles in biology and biotechnology and there has been considerable interest in the engineering of DBPs with new or altered specificities for genome editing and other applications. While there has been some success in reprogramming naturally occurring DBPs using selection methods, the computational design of new DBPs that recognize arbitrary target sites remains an outstanding challenge. We describe a computational method for the design of small DBPs that recognize short specific target sequences through interactions with bases in the major groove and use this method to generate binders for five distinct DNA targets with mid-nanomolar to high-nanomolar affinities. The individual binding modules have specificity closely matching the computational models at as many as six base-pair positions and higher-order specificity can be achieved by rigidly positioning the binders along the DNA double helix using RFdiffusion. The crystal structure of a designed DBP-target site complex is in close agreement with the design model and the designed DBPs function in both Escherichia coli and mammalian cells to repress and activate transcription of neighboring genes. Our method provides a route to small and, hence, readily deliverable sequence-specific DBPs for gene regulation and editing.
    DOI:  https://doi.org/10.1038/s41594-025-01669-4
  31. FASEB J. 2025 Sep 15. 39(17): e71025
    The IRE1/XBP1s Axis Regulates Innate Immune Responses in Conventional Dendritic Cells During ZIKV Infection.
    Mónica Guzmán-Rodríguez, Tomás Hernández-Díaz, Paula Lisboa, Javier López-Schettini, Sofia Sanhueza, Lisette Leyton, Takao Iwawaki, Ricardo Soto-Rifo, Fabiola Osorio.
      Zika virus (ZIKV) is a mosquito-borne flavivirus causing a major epidemic in the Americas in 2015. Dendritic cells (DCs) are leukocytes with key antiviral functions, but their role in ZIKV infection remains under investigation. While most studies have focused on the monocyte-derived subtype of DCs, less is known about conventional dendritic cells (cDCs), essential for the orchestration of antiviral adaptive immunity. This study investigates the mechanisms by which cDCs respond to ZIKV for antiviral cytokine production. Here, using murine cultures, we demonstrate that ZIKV infection and not detection of ZIKV-infected dead cells activates cDCs by inducing type I interferons (IFN-I) and proinflammatory cytokines. Furthermore, ZIKV-infected cDCs markedly activated the IRE1/XBP1s axis of the unfolded protein response (UPR). Flow cytometry analysis indicates that among cDCs, type 1 cDCs (cDC1s) are responsible for ZIKV detection. Functionally, genetic loss of XBP1s curtailed expression of the costimulatory molecule CD86 and the production of IFN-I and proinflammatory cytokines by cDCs, without exhibiting increased susceptibility to ZIKV infection. These effects are attributable to perturbations in the IRE1/XBP1s axis and not due to overcompensation of PERK or IRE1 kinase signaling. Finally, tissue resident cDCs also exhibit susceptibility to infection, potentially establishing these cells as ZIKV targets in vivo. These findings underscore a critical role for the IRE1/XBP1s pathway in fine-tuning cDC activation to ZIKV, linking viral recognition to cDC functional maturation and opening new avenues for exploring UPR pathways targeting cDCs in the context of flavivirus infections.
    Keywords:  IRE1; XBP1; Zika virus; cDC1; dendritic cells; unfolded protein response
    DOI:  https://doi.org/10.1096/fj.202501186R
  32. Life Sci Alliance. 2025 Dec;pii: e202503378. [Epub ahead of print]8(12):
    Overexpression of ATase1 and ATase2 disrupts the secretome and causes a progeria phenotype.
    Tzu-Lin Cheng, Feixuan Wu, Md Ezazul Haque, Abigail R Thiel, Danqing Wang, Jeffrey J Helgager, Lingjun Li, Luigi Puglielli.
      Nε-lysine acetylation in the lumen of the ER requires two acetyltransferases, ATase1/NAT8B and ATase2/NAT8. They are type II membrane proteins and belong to the larger GNAT superfamily of acetyltransferases. Their enzymatic activity is tightly coupled to the import of acetyl-CoA in the lumen of the ER by AT-1/SLC33A1. Gene duplication events involving 3q25.31 (harboring AT-1/SLC33A1) and 2p13.1 (harboring ATase1/NAT8B and ATase2/NAT8) are associated with autism spectrum disorder with intellectual disability and progeria-like dysmorphism. Here, we report the generation and phenotypic characterization of mice with systemic overexpression of ATase1 (ATase1 sTg) and ATase2 (ATase2 sTg). Overexpression of either ATase at conception was found to be lethal while overexpression at birth was found to cause a progeria-like phenotype that included skin alterations, lordokyphosis, reduced bone density, sarcopenia, splenomegaly, adenomegaly, and systemic inflammation. The phenotype of ATase1 sTg mice displayed incomplete penetrance, while the phenotype of ATase2 sTg displayed full penetrance and was more severe. Mechanistically, the phenotype was linked to altered dynamics of the secretory pathway with defects affecting the quality of the secretome.
    DOI:  https://doi.org/10.26508/lsa.202503378
  33. Science. 2025 Sep 11. 389(6765): 1112-1117
    Structural basis for LZTR1 recognition of RAS GTPases for degradation.
    Srisathiyanarayanan Dharmaiah, Daniel A Bonsor, Stephanie P Mo, Alvaro Fernandez-Cabrera, Albert H Chan, Simon Messing, Matthew Drew, Martha Vega, Dwight V Nissley, Dominic Esposito, Pau Castel, Dhirendra K Simanshu.
      The RAS family of small guanosine triphosphatases (GTPases) are tightly regulated signaling molecules that are further modulated by ubiquitination and proteolysis. Leucine Zipper-like Transcription Regulator 1 (LZTR1), a substrate adapter of the Cullin-3 RING E3 ubiquitin ligase, binds specific RAS GTPases and promotes their ubiquitination and proteasomal degradation. We present structures of LZTR1 Kelch domains bound to RIT1, MRAS, and KRAS, revealing interfaces that govern RAS isoform selectivity and nucleotide specificity. Biochemical and structural analyses of disease-associated Kelch domain mutations revealed three types of alterations: impaired substrate interaction, loop destabilization, and blade-blade repulsion. In cellular and mouse models, mutations disrupting substrate binding phenocopied LZTR1 loss, underscoring its substrate specificity. These findings define RAS recognition mechanisms by LZTR1 and suggest a molecular glue strategy to degrade oncogenic KRAS.
    DOI:  https://doi.org/10.1126/science.adv7088
  34. Sci Adv. 2025 Sep 12. 11(37): eadx9567
    Molecular exaptation by the integrin αI domain.
    Jeremy A Hollis, Matthew C Chan, Harmit S Malik, Melody G Campbell.
      Integrins bind ligands between their alpha (α) and beta (β) subunits and transmit signals through conformational changes. Early in chordate evolution, some α subunits acquired an "inserted" (I) domain that expanded integrin's ligand-binding repertoire but obstructed the ancestral ligand pocket, seemingly blocking conventional integrin activation. Here, we compare cryo-electron microscopy structures of apo and ligand-bound states of the I domain-containing αEβ7 integrin and the I domain-lacking α4β7 integrin to illuminate how the I domain intrinsically mimics an extrinsic ligand to preserve integrin function. We trace the I domain's evolutionary origin to an ancestral collagen-collagen interaction domain, identifying an ancient molecular exaptation that facilitated integrin activation immediately upon I domain insertion. Our analyses reveal the evolutionary and biochemical basis of expanded cellular communication in vertebrates.
    DOI:  https://doi.org/10.1126/sciadv.adx9567
  35. Proc Natl Acad Sci U S A. 2025 Sep 16. 122(37): e2515326122
    S-nitrosylation of pVHL regulates β2 adrenergic receptor function.
    Zachary W Grimmett, Hiroki Hayashi, Thomas M Raffay, Justin Lin, Richard T Premont, Jonathan S Stamler.
      The β2-adrenergic receptor (β2AR), a prototype G protein-coupled receptor, controls cardiopulmonary function underpinning O2 delivery. Abundance of the β2AR is canonically regulated by G protein-coupled receptor kinases and β-arrestins, but neither controls constitutive receptor levels, which are dependent on ambient O2. Basal β2AR expression is instead regulated by the prolyl hydroxylase/pVHL-E3 ubiquitin ligase system, explaining O2 responsivity. Interplay between O2 and nitric oxide (NO, a potent bronchodilator) is central to cardiopulmonary function. Here, we demonstrate that pVHL-mediated β2AR degradation is counteracted by NO, revealing pVHL control of pulmonary function. NO S-nitrosylates Cys77 in human pVHL (cognate to mouse Cys43), which induces binding of the E3 ubiquitin ligase c-Cbl to degrade pVHL, thereby increasing β2AR expression. pVHL-C43S mutant mice refractory to S-nitrosylation exhibit decreases in β2AR signaling and increases in airway tone. Thus, pVHL controls adrenergic pulmonary function and contributes to bronchodilation by NO. Our findings suggest therapeutic approaches to asthma and obstructive airway disease.
    Keywords:  S-nitrosothiol; nitric oxide; pVHL ubiquitin ligase; posttranslational modification; redox signaling
    DOI:  https://doi.org/10.1073/pnas.2515326122
  36. PLoS Comput Biol. 2025 Sep 12. 21(9): e1013424
    Paying attention to attention: High attention sites as indicators of protein family and function in language models.
    Gowri Nayar, Alp Tartici, Russ B Altman.
      Protein Language Models (PLMs) use transformer architectures to capture patterns within protein primary sequences, providing a powerful computational representation of the amino acid sequence. Through large-scale training on protein primary sequences, PLMs generate vector representations that encapsulate the biochemical and structural properties of proteins. At the core of PLMs is the attention mechanism, which facilitates the capture of long-range dependencies by computing pairwise importance scores across residues, thereby highlighting regions of biological interaction within the sequence. The attention matrices offer an untapped opportunity to uncover specific biological properties of proteins, particularly their functions. In this work, we introduce a novel approach, using the Evolutionary Scale Modelling (ESM) , for identifying High Attention (HA) sites within protein primary sequences, corresponding to key residues that define protein families. By examining attention patterns across multiple layers, we pinpoint residues that contribute most to family classification and function prediction. Our contributions are as follows: (1) we propose a method for identifying HA sites at critical residues from the middle layers of the PLM; (2) we demonstrate that these HA sites provide interpretable links to biological functions; and (3) we show that HA sites improve active site predictions for functions of unannotated proteins. We make available the HA sites for the human proteome. This work offers a broadly applicable approach to protein classification and functional annotation and provides a biological interpretation of the PLM's representation.
    DOI:  https://doi.org/10.1371/journal.pcbi.1013424
  37. J Neurosci. 2025 Sep 10. pii: e0371252025. [Epub ahead of print]
    Smg5 enhances oligodendrocyte differentiation via nonsense-mediated mRNA decay of Hnrnpl variant transcripts.
    Min Jiang, Conghui Li, Binghua Xie, Tangliang Li, Zhong-Min Dai, Mengsheng Qiu.
      Nonsense-mediated mRNA decay (NMD) is a conserved RNA surveillance mechanism that degrades transcripts with premature termination codons (PTCs) and finetunes gene expression by targeting RNA transcripts with other NMD inducing features. This study demonstrates that conditional knockout of Smg5, a key NMD component, in oligodendrocyte lineage cells disrupts the degradation of PTC-containing transcripts, including aberrant variants of the RNA-binding protein Hnrnpl The loss of SMG5 in both sexes of mice impaired oligodendrocyte differentiation, reduced myelin gene expression, and led to thinner myelin sheaths and compromised motor function in mice. Mechanistically, HNRNPL was shown to regulate the alternative splicing of myelin-associated genes Mag and Nfasc, and promote oligodendrocyte differentiation. These findings reveal that SMG5-mediated NMD ensures RNA processing fidelity essential for proper oligodendrocyte development and CNS myelination.Significance Statement The deletion of Smg5 in oligodendrocyte lineage cells disrupts terminal differentiation by impairing nonsense-mediated mRNA decay (NMD). This NMD deficiency dysregulates Hnrnpl expression, revealing that Smg5-dependent post-transcriptional control of Hnrnpl is essential for oligodendrocyte maturation. Mechanistically, HNRNPL protein promotes the splicing of Mag pre-mRNA, enabling production of the Large-MAG, a critical effector of central nervous system myelin sheath formation. Thus, the Smg5-Hnrnpl-Mag axis constitutes a previously unrecognized regulatory pathway indispensable for CNS myelination.
    DOI:  https://doi.org/10.1523/JNEUROSCI.0371-25.2025
  38. Nature. 2025 Sep 10.
    Amplifying antigen-induced cellular responses with proximity labelling.
    Shuojun Li, Yinghui Men, Zihan Wang, Yingcheng Wu, Hao Sun, Mingyang Yin, Xinrui Fan, Guiyun Deng, Zhicheng Yang, Tiange Yang, Yudian Xiao, Hu Zhou, Guangchuan Wang, Jia Fan, Chenqi Xu, Qiang Gao, Shuo Han.
      Antigen-induced clustering of cell surface receptors, including T cell receptors and Fc receptors, represents a widespread mechanism in cell signalling activation1,2. However, most naturally occurring antigens, such as tumour-associated antigens, stimulate limited receptor clustering and on-target responses owing to insufficient density3-5. Here we repurpose proximity labelling6, a method used to biotinylate and identify spatially proximal proteins, to amplify designed probes as synthetic antigen clusters on the cell surface. We develop an in vivo proximity-labelling technology controlled by either red light or ultrasound to covalently tag fluorescein probes at high density near a target antigen. Using T cell receptors as an example, we demonstrate that the amplified fluorescein effectively clusters and directs a fluorescein-binding bispecific T cell engager to induce enhanced T cell activation and cytotoxicity. Noninvasive, tissue-selective labelling in multiple syngeneic mouse tumour models produces potent immune responses that rapidly eradicate treated tumours. Efficient cell lysis further promotes epitope spreading to induce systemic immunity against untreated distal lesions and immune memory against rechallenge. Thus, proximity-labelling chemistry holds promise as a generalized strategy to manipulate antigen-dependent receptor function and cell states.
    DOI:  https://doi.org/10.1038/s41586-025-09518-6
This page is being maintained by Thomas Krichel. It was last updated on 2025‒09‒14 at 6:30.