bims-proteo Biomed News
on Proteostasis
Issue of 2026–01–18
71 papers selected by
Eric Chevet, INSERM
  1. tRNA synthetase activity is required for stress granule and P-body assembly.
  2. A non-canonical role for UPR ER during heat stress in C. elegans.
  3. Proteasomal proteolysis in p62 condensates directs tumor suppression or growth depending on their subcellular localization.
  4. 1-deoxysphingolipids dysregulate membrane properties and cargo trafficking in the early secretory pathway.
  5. VAPA at the inner nuclear membrane affects nuclear lamins and nuclear morphology.
  6. K48-ubiquitin-dependent proteases cut-up post-ER proteins.
  7. A GluN2B disease-associated variant promotes the degradation of NMDA receptors via autophagy.
  8. Systematic Characterization of Cancer-Associated SPOP Mutants Reveals Novel and Reprogrammable Degradative Activities.
  9. The cotranslational cycle of the ribosome-bound Hsp70 homolog Ssb.
  10. SNX-3 confers lysosomal fusion-competence to sustain basal autophagy.
  11. Atypical tetracyclines promote longevity and ferroptotic neuroprotection via translation attenuation.
  12. Dominant-negative TP53 mutations potentiated by the HSF1-regulated proteostasis network.
  13. Quality control and signaling pathways at stalled ribosomes.
  14. Native Mass Spectrometry Analysis of Cullin RING Ubiquitin E3 Ligase Complexes in the Context of Targeted Protein Degradation.
  15. ATG9A-dependent, LC3-independent autophagy curbs the immune system to protect against disease.
  16. Improved protein binder design using β-pairing targeted RFdiffusion.
  17. Cryo-EM structure of the human COP1-DET1 ubiquitin ligase complex.
  18. The Ubiquitin-Proteasome Pathway Mediates Selective Degradation of Unloaded Argonaute Proteins in C. elegans.
  19. Nvj3 regulates Dga1-mediated triacylglycerol synthesis and lipid droplet formation at ER contact sites.
  20. Inhibition of mTOR Enhances the Efficacy of Proteasome-Dependent Targeted Protein Degradation Approaches.
  21. The E3 ubiquitin ligase mechanism specifying target-directed microRNA degradation.
  22. Inactive ryanodine receptors sustain lysosomal availability for autophagy by promoting ER-lysosomal contact site formation.
  23. Unveiling the neuroprotective power of mitochondrial transfer in orofacial inflammatory pain through ER membrane remodeling.
  24. FGF1 ameliorates hepatic steatosis through acute activation of the unfolded protein response and VLDL production.
  25. Plant rhabdovirus glycoprotein activates unfolded protein response-mediated antiviral ER-phagy in insect vectors.
  26. Poly(ADP-ribose) glycohydrolase enforces p21 degradation via dePARylation to promote gastric cancer progression.
  27. Chronic stress disrupts the network among stress granules, P-bodies, and motor proteins.
  28. Eukaryotic initiation factor eIF3 facilitates loading of eukaryotic release factor eRF1 or suppressor tRNA to the ribosome.
  29. Structural and mechanistic analysis of covalent ligands targeting the RNA-binding protein NONO.
  30. LAMP1 and LAMP2A localise to axonal organelles with distinct motility dynamics and partially overlapping molecular signatures in human neurons.
  31. N1-Methylpseudouridine directly modulates translation dynamics.
  32. ClpA and ClpAP-Catalyzed Unfolding and Translocation are Differentially Coupled to ATP Binding.
  33. Bridging single-molecule and genome-wide studies of cellular mRNA translation.
  34. Hydrogen/deuterium exchange mass spectrometry analysis of ribosome-nascent chain complexes to study protein biogenesis at the peptide level.
  35. Parkin Deficiency Impairs ER-Mitochondria Associations and calcium homeostasis via IP3R-Grp75-VDAC1 Complex.
  36. Muscle meets Lysosomes: emerging strategies in muscular dystrophy.
  37. Expanding roles of N-glycosylation in the endoplasmic reticulum.
  38. Arsenic disrupts autophagosome-lysosome fusion in a zinc dependent manner across multiple human skin and lung cells lines.
  39. Enterococcus faecalis redox metabolism activates the unfolded protein response to impair wound healing.
  40. Small molecule splicing modulators that disrupt O-GlcNAc homeostasis.
  41. FBXO11 suppression rewires an NPM1-centered interactome influencing the progression of myelodysplastic syndrome.
  42. Allosteric PROTACs: Expanding the Horizon of Targeted Protein Degradation.
  43. Organelle dysfunction and TNT-mediated aggregate spreading in neurodegeneration.
  44. Stress Granules as a Central Hub Linking Organelle Stress, Aging, and Neurodegeneration.
  45. PAR-Driven Condensation Maintains Stalled Replication Fork Stability.
  46. Predicting epistasis across proteins by structural logic.
  47. NRF1 is upregulated by docosahexaenoic acid to ameliorate MASH through the inhibition of ER stress.
  48. The ALS-associated E425K mutation uncouples DNAJC7 from the Hsp70 chaperone cycle.
  49. Multilayer Host Engineering of Saccharomyces cerevisiae to Enhance Cricket Paralysis Virus (CrPV) Internal Ribosome Entry Site Mediated Translation.
  50. Generating ER-TRG and CA-ER-TRG Knock-in Mice and Quantitative In Vivo Imaging of ER-phagy.
  51. Loss of SEL1L or Hrd1 increases intrinsic LOX levels in HEK293 cells independently of proteasomal degradation.
  52. Manipulation of the Unfolded Protein Response by Intracellular Bacterial Pathogens: Mechanisms of ER Hijacking and Therapeutic Implications.
  53. MYO5A-mediated stabilization promotes the acquisition of fusion competence in sealed autophagosomes.
  54. Degradation of the TEAD•YAP/TAZ Transcription Factor Complex by Heterobifunctional Small Molecules that Bind to the TEAD Allosteric Lipid Pocket.
  55. Limited proteolysis-coupled mass spectrometry captures proteome-wide protein structural alterations and biomolecular condensation in living cells.
  56. Functionally coupled ion channels begin co-assembling at the start of their synthesis.
  57. Amino acid restriction sensitizes lung cancer cells to ferroptosis via GCN2-dependent activation of the integrated stress response.
  58. Alleviation of ER stress via targeted genetic engineering enhances recombinant ovalbumin secretion in Saccharomyces cerevisiae.
  59. Benchmarking co-folding methods to predict the structures of covalent protein-ligand complexes.
  60. Comprehensive mapping of RNA modification dynamics and crosstalk via deep learning and nanopore direct RNA-sequencing.
  61. A bacterial translation activator with an intrinsically disordered RNA-binding region.
  62. Extracellular vesicle-based targeted protein degradation platform for multiple extracellular proteins.
  63. Enhancement of bZIP60 function through C-terminal region translated after splicing in Arabidopsis.
  64. Piezo1 dictates K+ homeostasis through coordinated regulation of the ubiquitin ligase Kelch-like 3 in RBCs and the kidney.
  65. Chemical Proteomics Identifies Ketogenesis-Mediated Cysteine Modifications Regulating Redox Function.
  66. Strategic variations in sarbecovirus and merbecovirus Nsp1 linker regions for translation inhibition.
  67. LYTACs and other extracellular targeted protein degradation TACnologies: guiding principles and potential clinical prospects.
  68. Proteolysis-targeting Chimera efficacy prediction using a deep-learning-QSP model.
  69. Differentiation in the human urothelia is defined by distinct alternative polyadenylation.
  70. A shared binding interface controls Vps13 organelle-specific targeting independently of its vacuolar protein sorting function.
  71. Candida albicans-induced ubiquitination of EGFR reveals novel host-fungal interaction pathways.
  1. Genes Dev. 2026 Jan 14.
    tRNA synthetase activity is required for stress granule and P-body assembly.
    Max Baymiller, Noah S Helton, Benjamin Dodd, Stephanie L Moon.
      Translation elongation defects activate the integrated stress response (ISR), but whether and how ribosome stalls are cleared to enable mRNA release for ribonucleoprotein (RNP) granule assembly remain unclear. We show that blocking tRNA aminoacylation generates persistent uncollided ribosome stalls that inhibit stress granule and P-body assembly despite robust ISR activation. Collided ribosomes are rapidly cleared by ZNF598-dependent ribosome-associated quality control within 4 h, while uncollided stalls resist clearance and persist for >16 h. Puromycin releases persistent stalls and restores RNP granule formation. The block in stress granule assembly is generalizable across tRNA synthetase inhibitors and amino acid deprivation. Therefore, stress granules represent signal integrators reporting translation elongation status when initiation is suppressed. Our findings reveal that translation quality control pathways selectively clear collided ribosomes, establish that translation elongation stress uncouples RNP granule assembly from the ISR, and suggest that tolerating uncollided stalls may be adaptive for cotranslational processes essential for cellular function.
    Keywords:  P-bodies; halofuginone; integrated stress response; ribosome collisions; ribosome-associated quality control; stress granules; tRNA synthetase; translation elongation
    DOI:  https://doi.org/10.1101/gad.353535.125
  2. bioRxiv. 2026 Jan 09. pii: 2026.01.08.698479. [Epub ahead of print]
    A non-canonical role for UPR ER during heat stress in C. elegans.
    Athena Alcala, Toni Castro Torres, Rebecca Aviles Barahona, Phillip A Frankino, Ryo Higuchi-Sanabria, Gilberto Garcia.
      Organisms rely on coordinated stress responses to maintain cellular homeostasis. Perhaps the best-known example of multiple stress inputs converging onto a single response is the integrated stress response (ISR), which reduces global translation under various stress conditions to reduce the protein folding burden of the cell. Similarly, most stress responses generally involve coordination of additional protein homeostasis (proteostasis) pathways, including increased expression of chaperones to refold proteins, as well as activation of clearance mechanisms, such as autophagy and the ubiquitin proteosome system. Our study investigates how heat stress can influence coordinated activation of both cytosolic and ER chaperones, exploring bidirectional cross talk between canonical activators of the cytosolic heat-shock response (HSR) and the unfolded protein response of the ER (UPR ER ). Using robust transcriptional reporters in the C. elegans model system, we explore a non-canonical activation of the UPR ER under heat stress by the coordinated effects of XBP-1 and HSF-1. We further investigate inter tissue communications of stress whereby neuronal or glial activation of the UPR ER can result in heterotypic enhancement of the HSR in peripheral and can increase thermotolerance. This work highlights the complex convergence of cellular stress responses, a phenomenon that may reflect a general strategy wherein localized stress can activate numerous proteostasis pathways to prevent whole cell and whole organism damage.
    Article Summary: A reductionist approach to studying cellular stress responses is critical for dissecting specific molecular and genetic drivers of stress response. However, stress responses are often convergent and overlapping, and these single input and output studies may miss their complex interplay. Many studies have revealed the intricate coordination of stress responses, including the ability of seemingly organelle-specific stress responses, like mitochondrial stress responses, to directly influence cytosolic and ER health. Our study adds to this growing field by describing a unique, bidirectional crosstalk between the cytosolic and ER stress pathways, highlighting systemic coordination of stress resilience.
    DOI:  https://doi.org/10.64898/2026.01.08.698479
  3. Proc Natl Acad Sci U S A. 2026 Jan 20. 123(3): e2529422123
    Proteasomal proteolysis in p62 condensates directs tumor suppression or growth depending on their subcellular localization.
    Chen Lulu-Shimron, Zhiwen Luo, Vera Brekhman, Lina Huang, Ido Livneh, Hidetaka Kosako, Victoria Cohen-Kaplan, Aaron Ciechanover.
      p62/SQSTM1 generates liquid-liquid phase-separated condensates that participate in diverse processes, including protein quality control (PQC) and autophagy. Nuclear p62 condensates were shown to act as ubiquitin- and proteasome-mediated degradation hubs, whereas the involvement of cytoplasmic condensates in this pathway has remained unclear. Here, we show that cytoplasmic p62 condensates serve as a hub for proteasomal degradation that displays distinct substrate preferences compared with nuclear condensates. Specifically, cytoplasmic condensates mediate accelerated degradation of the tumor suppressor p53 through recruitment MDM2, its E3 ligase, while nuclear condensates are selectively enriched with USP7, a deubiquitinating enzyme (DUB) that stabilizes p53. Immunohistochemical analysis of human tissues reveal that p62 in healthy tissues is largely localized to the nucleus, whereas in the corresponding malignant tissues, it is largely in the cytosol, which is correlated with reduced p53 abundance in tumors. Nuclear p62 condensates also promote the degradation of oncogenic c-Myc, underscoring compartment-specific differences in protein turnover. Experiments in cancer cells and xenografts demonstrate that cytoplasmic p62 condensates drive tumor growth, whereas nuclear p62 condensates suppress it. Moreover, condensate formation rather than p62 expression alone is required for both enhanced proteolytic activity and tumor growth modulation. Proteomic analysis reveals that nuclear p62, unlike its cytosolic counterpart, is linked to enrichment of proteins associated with apoptosis, p53 stabilization, DNA damage response, and cellular senescence-all related to tumor suppression. These findings establish that p62 condensates provide compartment-specific regulation of ubiquitin and proteasomal degradation and suggest that manipulating their localization or affecting their dynamics can offer different therapeutic opportunities.
    Keywords:  c-Myc; cancer; p53; p62 condensates; ubiquitin–proteasome system
    DOI:  https://doi.org/10.1073/pnas.2529422123
  4. Cell Chem Biol. 2026 Jan 15. pii: S2451-9456(25)00399-X. [Epub ahead of print]33(1): 45-58.e8
    1-deoxysphingolipids dysregulate membrane properties and cargo trafficking in the early secretory pathway.
    Yi-Ting Tsai, Nicolas-Frédéric Lipp, Olivia Seidel, Riya Varma, Aurelie Laguerre, Kristina Solorio-Kirpichyan, Adrian M Wong, Roberto J Brea, Grace H McGregor, Thekla Cordes, Neal K Devaraj, Lars Kuerschner, Sonya Neal, Christian M Metallo, Itay Budin.
      1-Deoxysphingolipids are non-canonical sphingolipids linked to several diseases, yet their cellular effects are poorly understood. Here, we utilize lipid chemical biology approaches to investigate the role of 1-deoxysphingolipid metabolism on the properties and functions of secretory membranes. We applied organelle-specific bioorthogonal labeling to visualize the subcellular distribution of metabolically tagged sphingolipids. We observed that 1-deoxysphingolipids are retained in the endoplasmic reticulum (ER) and specifically in ER exit sites (ERESs), suggesting that they do not efficiently sort into vesicular carriers. Cell lines expressing disease-associated variants of serine palmitoyl-CoA transferase accumulated 1-deoxysphingolipids, which were accompanied by a reduction in ER membrane fluidity and enlargement of ERES. We found that the rates of membrane protein release from the ER were altered in response to 1-deoxysphingolipid metabolism in a manner dependent on the protein's affinity for ordered or disordered membranes. The dysregulation of sphingolipid metabolism can thus alter secretory membrane properties and affect protein trafficking.
    Keywords:  ceramides; endoplasmic reticulum; membrane fluidity; sphingolipids; trafficking
    DOI:  https://doi.org/10.1016/j.chembiol.2025.12.006
  5. J Cell Sci. 2026 Jun 15. pii: jcs264298. [Epub ahead of print]139(12):
    VAPA at the inner nuclear membrane affects nuclear lamins and nuclear morphology.
    Inés Rodríguez-González, David Kohlhause, Christof Lenz, Henning Urlaub, Christiane Spillner, Ralph H Kehlenbach.
      Vesicle-associated membrane protein-associated protein A (VAPA) is a protein of the endoplasmic reticulum (ER) and a component of several membrane contact sites (MCSs). We show here that VAPA also localizes to the inner nuclear membrane (INM), in close proximity to nuclear lamins, INM proteins and nucleoporins. Using our proteomics approach 'rapamycin- and APEX-dependent identification of proteins by SILAC' (RAPIDS), we identified several nuclear proximity partners of VAPA, including emerin, different LAP2 isoforms, lamin A/C and Nup153. Depletion of VAPA in various cellular systems resulted in reduced nuclear lamin levels and aberrant nuclear morphology, including the formation of membrane invaginations and tunnels. Furthermore, histone acetylation levels were altered. Our data suggest that VAPA has distinct nuclear functions, in addition to its established role as an ER organizer.
    Keywords:  Endoplasmic reticulum; Inner nuclear membrane; Nuclear envelope; Nuclear tunnels; Nucleus; VAPA
    DOI:  https://doi.org/10.1242/jcs.264298
  6. Nat Commun. 2026 Jan 13.
    K48-ubiquitin-dependent proteases cut-up post-ER proteins.
    Annabel Y Minard, Stanley Winistorfer, Liping Yu, Robert C Piper.
      Polyubiquitin chains, linked via K48 or K63 of ubiquitin, direct membrane proteins in the secretory system to distinct degradative fates. However, it's unclear whether these linkage isomers are functionally interchangeable. Here we show that for post-endoplasmic reticulum (ER) proteins, K63-linked polyubiquitination induces sorting into multivesicular bodies (MVBs) and lysosomal degradation. In contrast, K48-linked polyubiquitination induces shearing from the membrane and proteasomal degradation. This process involves two ubiquitin-dependent proteases: Ddi1, a conserved cytosolic ubiquilin that generates fragments from soluble and membrane proteins, and Rbd2, an intramembrane rhomboid protease that produces lumenal fragments from membrane proteins at Golgi/endosomes and the vacuolar membrane. Ddi1's catalytic core, the HDD-RVP domain, is sufficient for ubiquitin-dependent proteolysis. It binds ubiquitin directly and its activity is enhanced by auxiliary ubiquitin binding domains: an atypical UBL domain and a UBA domain. These findings demonstrate that polyubiquitin chains linked by different residues encode distinct degradative fates for post-ER proteins, and reveal two proteases that target ubiquitinated integral membrane proteins in a process we call CUT-UP (Cleavage of Ubiquitinated Targets by Ubiquitin-dependent Proteases).
    DOI:  https://doi.org/10.1038/s41467-026-68367-7
  7. J Biol Chem. 2026 Jan 10. pii: S0021-9258(26)00017-7. [Epub ahead of print] 111147
    A GluN2B disease-associated variant promotes the degradation of NMDA receptors via autophagy.
    Taylor M Benske, Marnie P Williams, Pei-Pei Zhang, Adrian J Palumbo, Ting-Wei Mu.
      N-methyl-D-aspartate receptors (NMDARs) are essential for excitatory neurotransmission, and missense mutations can severely disrupt their function. Pathogenic variants often lead to proteostasis defects, including improper folding, impaired assembly, and reduced trafficking to the plasma membrane, ultimately compromising the physiological function of NMDARs and thereby contributing to neurological diseases. However, mechanisms by which the proteostasis network recognizes and degrades aggregated, misfolded, and trafficking-deficient pathogenic NMDARs remain poorly understood. Here, we demonstrate that the R519Q GluN2B variant is retained in the endoplasmic reticulum (ER) and fails to traffic to the cell surface to form functional NMDARs. Pharmacological and genetic inhibition of autophagy resulted in the accumulation of this variant, indicating that it is degraded by the autophagy-lysosomal proteolysis pathway. Since GluN2B subunit has a cytosolic LC3-interacting region (LIR) motif, disruption of the LIR motif via mutagenesis similarly impairs the autophagic clearance of this variant. Furthermore, we demonstrate that this variant is recognized by the ER-phagy receptor CCPG1 and that the LIR domain plays a facilitative role in strengthening this interaction. Our results provide a novel molecular mechanism for the ER-to-lysosome associated degradation of NMDAR variants and identify a pathway for targeted therapeutic intervention for neurological disorders with dysfunctional NMDARs.
    Keywords:  CCPG1; ER-phagy; LC3-interacting region (LIR); NMDAR; proteostasis
    DOI:  https://doi.org/10.1016/j.jbc.2026.111147
  8. Chembiochem. 2026 Jan;27(1): e202500914
    Systematic Characterization of Cancer-Associated SPOP Mutants Reveals Novel and Reprogrammable Degradative Activities.
    Alana G Caldwell, Harshil Parmar, Xiaokang Jin, Chen Zhou, Xiaoyu Zhang.
      Speckle-type POZ protein (SPOP) functions as the substrate adaptor of the Cullin3-RING ligase complex and is recurrently mutated in multiple cancer types. Among these, F102C and F133L are frequent prostate cancer mutations within the substrate-binding domain, yet their biochemical consequences remain incompletely understood. Using quantitative proteomics, we show that SPOP-F133L, unlike SPOP-F102C, retains degradative activity toward the nuclear basket proteins NUP153 and TPR, indicating substrate-dependent loss-of-function. Moreover, SPOP-F133L induces partial down-regulation of p53 through a Cullin-RING ligase-dependent, post-translational mechanism, revealing a potential neo-substrate relationship. Finally, we demonstrate that both SPOP-F102C and SPOP-F133L support targeted protein degradation in an engineered cellular system. These findings define the degradative capacities of SPOP mutants and highlight opportunities to repurpose these variants as mutant-selective E3 ligases for therapeutic applications.
    Keywords:  E3 ligase; protein–protein interactions; proteomics; recurrent mutation; targeted protein degradation
    DOI:  https://doi.org/10.1002/cbic.202500914
  9. Nat Commun. 2026 Jan 16.
    The cotranslational cycle of the ribosome-bound Hsp70 homolog Ssb.
    Ying Zhang, Lorenz Grundmann, Leonie Vollmar, Julia Schimpf, Volker Hübscher, Mohd Areeb, Irina Grishkovskaya, Anna Moddemann, Kerstin Werner, Thorsten Hugel, David Haselbach, Sabine Rospert.
      Coupling of ribosomal translation with cotranslational protein folding is essential for cellular homeostasis. In eukaryotes, Hsp70 and its J-domain cochaperone, the heterodimeric ribosome-associated complex (RAC), are central to this process; however, mechanistic insights into the coordination of Hsp70 function with translation remain limited. Here, we present two cryo-EM structures of the ribosome-bound yeast Hsp70 Ssb, identifying Rpl25/uL23 as the ribosomal binding site and revealing its interaction with a model nascent chain. Together with detailed biochemical and mutational analyses, these structures enable us to delineate the intricate RAC-dependent cycle, which positions the substrate binding domain of Ssb-ATP close to the tunnel exit to receive nascent chains. This arrangement allows Ssb to undergo substantial conformational changes upon ATP hydrolysis without steric clashes with the ribosome, while the substrate binding domain of Ssb, now anchored by the tightly bound nascent chain, remains close to the tunnel exit.
    DOI:  https://doi.org/10.1038/s41467-025-67685-6
  10. Cell Mol Life Sci. 2026 Jan 15.
    SNX-3 confers lysosomal fusion-competence to sustain basal autophagy.
    Qiaoju Kang, Zhenyu Liu, Lianyan Xiang, Sai Yang, Ping Yi, Rongying Zhang.
      Autophagy, the process for recycling cytoplasm in the lysosome, relies on tightly regulated membrane trafficking. During autophagy, autophagosomes either fuse with endosomes generating amphisomes and then lysosomes, or directly fuse with lysosomes, in both cases generating autolysosomes that degrade their contents. It remains unclear whether specific mechanisms or conditions determine these alternate routes. Here, we demonstrate that the endosomal regulator SNX3 specifically regulates basal autophagy under nutrient-adequate conditions in both Caenorhabditis elegans (C. elegans) and cultured mammalian cells. In C. elegans, SNX-3 depletion elevates autophagy independently of the UNC-51/ULK1 complex and leads to the accumulation of both autophagosomes and amphisomes, which consequently impairs the clearance of autophagic cargo, including SQST-1/p62 and protein aggregates. Mechanistically, SNX-3 depletion differentially regulates the machineries required for autophagosome-lysosome fusion. In snx-3 mutants, the Q-SNARE components SYX-17 and SNAP-29 translocate to autophagosomes, where they assemble with the endosomal R-SNAREs VAMP-7 and VAMP-8 to promote amphisome formation. Conversely, loss of SNX-3 impairs the lysosomal delivery of VAMP-8 and RAB-7, both essential for autophagosome/amphisome-lysosome fusion, thereby generating fusion-incompetent lysosomes. However, starvation restores the lysosomal fusion capability compromised by snx-3 depletion. Our findings reveal that autophagosome-lysosome fusion is preferentially regulated by nutrient status, and identify an endosomal regulator that tunes membrane trafficking with changing autophagy demands.
    Keywords:  Amphisome; Autophagosome–lysosome fusion; Basal autophagy; RAB-7; SNARE
    DOI:  https://doi.org/10.1007/s00018-025-06074-0
  11. bioRxiv. 2026 Jan 11. pii: 2026.01.09.698733. [Epub ahead of print]
    Atypical tetracyclines promote longevity and ferroptotic neuroprotection via translation attenuation.
    Khalyd J Clay, Manuel Sanchez-Alavez, Ian Newman, Na Na, Ana P Verduzco Espinoza, Alan To, Shannon Saad, Hollis T Cline, Michael Petrascheck.
      Preclinical and clinical studies have reported neuroprotective and geroprotective effects of tetracyclines that are independent of their antibiotic activity, but the underlying mechanisms remain unclear. Here, we systematically profile widely used tetracyclines, including impurities and degradation products, and identify translation attenuation as the shared driver of their neuroprotective and longevity-promoting effects, independent of classical tetracycline mechanisms. Instead, we uncover two mechanistically distinct classes of tetracyclines. Mitochondrial-targeting tetracyclines (MitoTets), exemplified by doxycycline, inhibit the mitochondrial ribosome and attenuate cytosolic translation through activation of the Integrated Stress Response (ISR). In contrast, atypical tetracyclines such as 4-epiminocycline and 12-aminominocycline act as cytosolic-targeting tetracyclines (CytoTets), directly inhibiting the cytosolic ribosome, bypassing the ISR, and protecting neurons from ferroptotic cell death. CytoTets are non-antibiotic, brain-penetrant, and neuroprotective in mouse and human neurons, establishing the tetracyclines as a tunable chemical scaffold for selectively targeting translation in aging and neurodegeneration.
    Highlights: The tetracyclines broadly attenuate translation in multiple eukaryotic modelsTranslation attenuation results from both ISR-dependent and ISR-independent mechanismsDiscovery of atypical, cytosolic targeting tetracyclines (CytoTETs) that protect from ferroptosis ISR-independentlyCytoTETs inhibit translation and are neuroprotective in human-derived neurons and mouse hippocampus.
    DOI:  https://doi.org/10.64898/2026.01.09.698733
  12. Mol Cell. 2026 Jan 14. pii: S1097-2765(25)00987-6. [Epub ahead of print]
    Dominant-negative TP53 mutations potentiated by the HSF1-regulated proteostasis network.
    Stephanie Halim, Rebecca M Sebastian, Kristi E Liivak, Jessica E Patrick, Tiffani Hui, David R Amici, Andrew O Giacomelli, Paulina Rios, Vincent L Butty, William C Hahn, Francisco J Sánchez-Rivera, Marc L Mendillo, Yu-Shan Lin, Matthew D Shoulders.
      Protein mutational landscapes are shaped by how amino acid substitutions affect stability and folding or aggregation kinetics. These properties are modulated by cellular proteostasis networks. Heat shock factor 1 (HSF1) is the master regulator of cytosolic and nuclear proteostasis. Chronic HSF1 activity upregulation is a hallmark of cancer cells, potentially because upregulated proteostasis factors facilitate the acquisition and maintenance of oncogenic mutations. Here, we assess how HSF1 activation influences mutational trajectories by which p53 can escape cytotoxic pressure from nutlin-3, an inhibitor of the p53 regulator mouse double minute 2 homolog (MDM2). HSF1 activation broadly increases the fitness of dominant-negative p53 substitutions, particularly non-conservative, biophysically unfavorable amino acid changes within buried regions of the p53 DNA-binding domain. These findings demonstrate that HSF1 activation reshapes the oncogenic mutational landscape by preferentially supporting the emergence and persistence of biophysically disruptive, cancer-associated p53 substitutions, linking proteostasis network activity directly to oncogenic evolution.
    Keywords:  HSP70; HSP90; cancer evolution; chaperones; deep mutational scanning; heat shock factor I; mutational buffering; p53; protein folding; proteostasis
    DOI:  https://doi.org/10.1016/j.molcel.2025.12.013
  13. Exp Mol Med. 2026 Jan 15.
    Quality control and signaling pathways at stalled ribosomes.
    Weili Denyse Chang, Young-Jun Choe.
      Aberrant mRNAs can arise from errors in RNA processing or from various physicochemical insults. Ribosomes translating such faulty mRNAs may stall, producing incomplete and potentially toxic polypeptides. These aberrant translation products are eliminated by the ribosome-associated quality control pathway. Ribosome stalling also leads to ribosome collisions, which can activate signaling pathways that enable cells to adapt to stress or determine cell fate. Here, in this Review, we summarize the molecular mechanisms of ribosome stalling and the associated quality control and signaling pathways, and discuss their implications in disease and therapeutics.
    DOI:  https://doi.org/10.1038/s12276-025-01623-w
  14. bioRxiv. 2026 Jan 07. pii: 2025.12.04.692288. [Epub ahead of print]
    Native Mass Spectrometry Analysis of Cullin RING Ubiquitin E3 Ligase Complexes in the Context of Targeted Protein Degradation.
    Louise M Sternicki, Charlotte Crowe, Lianne H E Wieske, Alessio Ciulli, Sally-Ann Poulsen.
      The binding of PROTACs to their partner ubiquitin E3 ligase (E3) and a protein of interest (POI) is critical for PROTAC development and validation. Characterisation of PROTAC complexes by cryo-electron microscopy and X-ray crystallography is not always feasible, especially where species may be transient and protein structures may not resolve due to flexible domains or intrinsically disordered regions. More routine biophysical methods with broader applicability to varied samples is desirable to support the rapidly expanding targeted protein degradation field. The majority of PROTACs in development and in the clinic act through a Cullin RING E3 Ligase (CRL) of which the pentameric von Hippel-Lindau (VHL) Cullin 2 RING E3 complex (CRL2 VHL ) is the foundational example. Native mass spectrometry (nMS) can be used to characterise protein complexes but has not previously been used to characterise a full E3 or any E3-E2 interactions. Here, we show that CRL2 VHL is amenable to characterisation by nMS and its interactions with the other protein components integral to the targeted protein degradation mechanism can be observed. Specifically, we characterise binary, ternary and higher order complexes that comprise CRL2 VHL , including the multiprotein systems of POI-PROTAC-CRL2 VHL , CRL2 VHL -E2-Ub, and POI-PROTAC-CRL2 VHL -E2-Ub, all of which are essential in facilitating productive POI ubiquitination and degradation. We benchmarked the nMS with two POI examples (BRD4 BD2 and KRAS) with their respective PROTACs (MZ1 and ACBI3) and were able to observe all relevant complexes across both systems. We anticipate that our findings will open avenues for nMS to integrate as an alternative experimental method enabling scalable characterisation of the intricate high mass multiprotein interactions central to the PROTAC mechanism of action.
    DOI:  https://doi.org/10.64898/2025.12.04.692288
  15. Autophagy Rep. 2026 ;5(1): 2614147
    ATG9A-dependent, LC3-independent autophagy curbs the immune system to protect against disease.
    Dario Priem, Mathieu Jm Bertrand.
      Selective autophagy is generally believed to require the conjugation of microtubule associated protein 1 light chain 3 (LC3) proteins (or other autophagy-related 8 [ATG8] family members) on the inner phagophore leaflet to enable the recruitment of cargo-bound selective autophagy receptors. However, this paradigm is challenged by the discovery that cytosolic cargoes can still be selectively targeted by phagophores even in the absence of LC3 proteins. In a recent study published in Immunity, we discovered that ATG9A-dependent, LC3-independent autophagy facilitates the degradation of multiple inflammatory signaling complexes to prevent an inflammatory skin disease.
    Keywords:  ATG9A; LC3-independent autophagy; STING; TNF; ZBP1; cGAS; cell death; inflammation; inflammatory skin disease; nucleic acid immunity
    DOI:  https://doi.org/10.1080/27694127.2026.2614147
  16. Nat Commun. 2026 Jan 10.
    Improved protein binder design using β-pairing targeted RFdiffusion.
    Isaac Sappington, Martin Toul, David S Lee, Stephanie A Robinson, Inna Goreshnik, Clara McCurdy, Tung Ching Chan, Nic Buchholz, Buwei Huang, Dionne Vafeados, Mariana Garcia-Sanchez, Nicole Roullier, Matthias Glögl, Christopher J Kim, Joseph L Watson, Susana Vázquez Torres, Koen H G Verschueren, Kenneth Verstraete, Cynthia S Hinck, Melisa Benard-Valle, Brian Coventry, Jeremiah Nelson Sims, Green Ahn, Xinru Wang, Andrew P Hinck, Timothy P Jenkins, Hannele Ruohola-Baker, Steven M Banik, Savvas N Savvides, David Baker.
      Designing proteins that bind with high affinity to hydrophilic protein target sites remains a challenging problem. Here we show that RFdiffusion can be conditioned to generate protein scaffolds that form geometrically matched extended β-sheets with target protein edge β-strands in which polar groups on the target are complemented with hydrogen bonding groups on the design. We use this approach to design binders against edge-strand target sites on KIT, PDGFRɑ, ALK-2, ALK-3, FCRL5, NRP1, and α-CTX, and obtain higher (pM to mid nM) affinities and success rates than unconditioned RFdiffusion. Despite sharing β-strand interactions, designs have high specificity, reflecting the precise customization of interacting β-strand geometry and additional designed binder-target interactions. A binder-KIT co-crystal structure is nearly identical to the design model, confirming the accuracy of the design approach. The ability to robustly generate binders to the hydrophilic interaction surfaces of exposed β-strands considerably increases the range of computational binder design.
    DOI:  https://doi.org/10.1038/s41467-025-67866-3
  17. Nat Commun. 2026 Jan 15. 17(1): 543
    Cryo-EM structure of the human COP1-DET1 ubiquitin ligase complex.
    Shan Wang, Fei Teng, Goran Stjepanovic, Feng Rao, Ming-Yuan Su.
      Ubiquitin modifications regulate fundamental cellular activities by modulating protein stability and function. The ubiquitin ligase COP1, which is present across species from plants to humans, plays a crucial role in the ubiquitination of developmental transcription factors. While COP1 can function independently, it can also be incorporated into CULLIN4-RING ubiquitin ligase (CRL4) complexes through the DET1 adaptor protein. Despite its biological significance, the structural and functional mechanisms of COP1 and DET1-containing complexes remains poorly understood. Here we present the cryo-electron microscopy structures of human COP1 in complex with DDB1-DDA1-DET1 and Ube2e2, revealing an inactive stacked assembly state. Co-expression with COP1 substrates including c-Jun or ETS2 disrupts this configuration, inducing a conformational rearrangement into a distinct dimeric state that allows substrate access. Structural modelling identifies the spatial organization of COP1 WD40 domains where substrate recruits. DET1 serves as a structural scaffold, bridging COP1 and Ube2e2 to initiate potential ubiquitin addition on substrates, while DDB1 recruits the CULLIN4-RBX1 complex to facilitate Ube2d3-mediated ubiquitin chain elongation. These results reveal the dynamic interplay between the structural states of the CRL4DET1-COP1 E3 ligase complex and its substrate specific activation mechanism, offering mechanistic insights into ubiquitination regulation and a basis for future studies on E3 ligase dynamics.
    DOI:  https://doi.org/10.1038/s41467-026-68375-7
  18. Genetics. 2026 Jan 13. pii: iyag007. [Epub ahead of print]
    The Ubiquitin-Proteasome Pathway Mediates Selective Degradation of Unloaded Argonaute Proteins in C. elegans.
    Jenny M Zhao, Dieu An H Nguyen, Diego Cervantes, Brandon Vong, Carolyn M Phillips.
      Argonaute proteins are essential effectors of small RNA-mediated gene regulation, yet the extent to which their stability depends on small RNA loading remains poorly understood. In Caenorhabditis elegans, we systematically disrupted the small RNA binding capacity of multiple Argonaute proteins to assess their stability in the absence of small RNA partners. We found that while most Argonautes remain stable when unable to bind small RNAs, a subset, including PRG-1, HRDE-1, and PPW-2, exhibited markedly reduced protein levels. Focusing on the PIWI-clade Argonaute PRG-1, we show that its destabilization occurs post-translationally and is independent of mRNA expression or translational efficiency. Instead, unbound PRG-1 is targeted for degradation by the ubiquitin-proteasome system. Additionally, the failure to load piRNAs disrupts PRG-1 localization to perinuclear germ granules. We further identify the E3 ubiquitin ligase EEL-1 as a factor contributing to the degradation of unloaded PRG-1. These findings uncover a critical role for small RNA loading in maintaining the stability and localization of a subset of Argonaute proteins, and reveal a quality control mechanism that selectively eliminates unbound PRG-1 to preserve germline regulatory fidelity.
    DOI:  https://doi.org/10.1093/genetics/iyag007
  19. bioRxiv. 2026 Jan 09. pii: 2026.01.05.695145. [Epub ahead of print]
    Nvj3 regulates Dga1-mediated triacylglycerol synthesis and lipid droplet formation at ER contact sites.
    Daniel Adebayo, Eseiwi Obaseki, Scott Miller, Marwa Aboumourad, Zaid Saadeh, Zachary Pomicter, Lindsey Kreinbring, Mohamad Rahal, Kashvi Vasudeva, Garrett Frost, Reema Smadi, Hanaa Hariri.
      Lipid droplet (LD) biogenesis is essential for lipid homeostasis during nutrient stress, yet how lipid intermediates are spatially organized to support efficient triacylglycerol (TAG) synthesis remains unclear. Here, we identify Nvj3 as a nutrient-responsive regulator that links diacylglycerol (DAG) availability to TAG synthesis and LD formation at the endoplasmic reticulum (ER). Nvj3 is induced by glucose depletion and recruited to LD-associated ER domains. Loss of Nvj3 causes neutral lipid accumulation under steady state conditions but delays TAG synthesis under acute inducible metabolic transitions. Using controlled TAG induction systems, we show that Nvj3 is required to couple Dga1-dependent TAG synthesis to LD formation. In the absence of Nvj3, TAG accumulates but remains inefficiently packaged into LDs. Consistent with this defect, nvj3Δ cells exhibit altered phospholipid remodeling and mislocalization of DAG away from ER domains during starvation. Together, these findings establish Nvj3 as an organizer of lipid availability during metabolic stress and suggest that spatial control of DAG is a key determinant of LD biogenesis.
    Summary: This study identifies Nvj3 as a spatial organizer of Dga1-dependent lipid droplet formation. Nvj3 promotes proper diacylglycerol positioning, and enables efficient triacylglycerol synthesis during metabolic stress. We propose that Nvj3 regulates lipid flux through spatial compartmentalization of diacylglycerol at membrane contact site-associated ER domains.
    DOI:  https://doi.org/10.64898/2026.01.05.695145
  20. Cancer Res. 2026 Jan 13.
    Inhibition of mTOR Enhances the Efficacy of Proteasome-Dependent Targeted Protein Degradation Approaches.
    Yang Liu, Yifeng Sun, Tian-Yu Song, Si-Yuan Ni, Yi-Sheng Pu, Yuan-Chen Chang, Wenrong Zhou, Zhen-Gang Peng, Jiajia Liu, Jun Shi, Wei Lu, Yong Cang.
      Targeted protein degradation (TPD) approaches, including molecular glue degraders (MGDs) and proteolysis targeting chimeras (PROTACs), overcome traditional occupancy-based inhibitor limitations and facilitate therapeutic development against "undruggable" disease-causing proteins. However, resistance to TPD is common, highlighting the need to further understand the driving mechanisms to improve treatment efficacy. Here, we identified a critical role of mTOR signaling in regulating PROTAC and MGD efficacy in vitro and in vivo. Activation or inhibition of mTOR respectively diminished or enhanced degradation efficacy of all proteasome-dependent TPD modalities tested. Mechanistically, mTOR inhibition suppressed de novo protein synthesis, thus creating a synthetic vulnerability by depleting replenishment of proteins targeted by PROTACs or MGDs. When applied to myeloma, mTOR inhibitors restored sensitivity to pomalidomide, one of the best characterized MGDs, in resistant cell lines and reduced malignant plasma cells in relapsed/refractory patients. This study reveals a clinically translatable strategy combining approved mTOR inhibitors with MGDs or PROTACs to enhance the therapeutic index of targeted protein degradation.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-25-3941
  21. bioRxiv. 2026 Jan 05. pii: 2026.01.05.697729. [Epub ahead of print]
    The E3 ubiquitin ligase mechanism specifying target-directed microRNA degradation.
    Jakob Farnung, Elena Slobodyanyuk, Peter Y Wang, Lianne W Blodgett, Daniel H Lin, Susanne von Gronau, Brenda A Schulman, David P Bartel.
      MicroRNAs (miRNAs) associate with Argonaute (AGO) proteins to form complexes that down-regulate target RNAs, including mRNAs from most human genes 1-3 . Within each complex, the miRNA pairs to target mRNAs to specify their repression, and AGO provides effector function while also protecting the miRNA from cellular nucleases 2-5 . Although much has been learned about this mode of posttranscriptional gene regulation, less is known about how the miRNAs themselves are regulated. In one such regulatory pathway, unusual miRNA targets called "trigger" RNAs reverse the canonical regulatory logic and instead down-regulate microRNAs 6-21 . This target- d irected m iRNA d egradation (TDMD) is thought to require a c ullin- R ING E3 ligase (CRL) because it depends on the cullin protein CUL3 and other ubiquitylation components, including the BC-box protein ZSWIM8 (ref. 22,23). ZSWIM8 is required for murine perinatal viability and for destabilization of most short-lived miRNAs, but is otherwise poorly understood 23-25 . Here, we demonstrate that a human AGO-miRNA- trigger complex selectively binds ZSWIM8 for CUL3-mediated polyubiquitylation of the AGO protein within this complex. Cryogenic electron-microscopy (cryo-EM) analyses show how ZSWIM8 recognizes the distinct AGO2 and miRNA-trigger conformations shaped by pairing of the miRNA to the trigger. For example, this pairing extracts the miRNA from a binding pocket within AGO2, allowing the pocket to be captured by ZSWIM8, and it directs the trigger RNA along a distinct trajectory to be also recognized by ZSWIM8. These results biochemically establish AGO binding and polyubiquitylation as the key regulatory step of TDMD, define a unique CRL class, and reveal generalizable RNA-RNA, RNA-protein, and protein-protein interactions that specify the ubiquitin-mediated degradation of AGO with exquisite selectivity. The substrate features recognized by the E3 ubiquitin ligase do not conform to a conventional degron 26-28 , but rather establish a two-RNA-factor authentication mechanism specifying a protein ubiquitylation substrate.
    DOI:  https://doi.org/10.64898/2026.01.05.697729
  22. Nat Commun. 2026 Jan 15.
    Inactive ryanodine receptors sustain lysosomal availability for autophagy by promoting ER-lysosomal contact site formation.
    Tim Vervliet, Jens Loncke, Marko Sever, Karan Ahuja, Chris Van den Haute, Tomas Luyten, Grace E Stutzmann, Catherine Verfaillie, Tihomir Tomašič, Geert Bultynck.
      Lysosomal and endoplasmic reticulum (ER) Ca2+ release mutually influence each other's functions. Recent work revealed that ER-located ryanodine receptor(s) (RyR(s)) Ca2+ release channels suppress autophagosome turnover by the lysosomes. In familial Alzheimer's disease, inhibiting RyR hyperactivity restored autophagic flux by normalizing lysosomal vacuolar H+-ATPase (vATPase) levels. However, the mechanisms by which RyRs control lysosomal function and how this involves the vATPase remain unknown. Here, we show that RyRs interact with the ATP6v0a1 subunit of the vATPase, contributing to ER-lysosomal contact site formation. This interaction suppresses RyR-mediated Ca²⁺ release, leading to reduced lysosomal exocytosis. Pharmacological inhibition of RyR activity mimics these effects on lysosomal exocytosis. Retaining lysosomes inside cells via RyR inhibition increases ER-lysosomal contact site formation, rendering lysosomes more available for autophagic flux. In summary, these findings establish RyR/ATP6v0a1 complexes as ER-lysosomal tethers that dynamically and Ca2+ dependently regulate the intracellular availability of lysosomes to participate in autophagic flux.
    DOI:  https://doi.org/10.1038/s41467-025-68054-z
  23. Cell Rep. 2026 Jan 13. pii: S2211-1247(25)01581-5. [Epub ahead of print]45(1): 116809
    Unveiling the neuroprotective power of mitochondrial transfer in orofacial inflammatory pain through ER membrane remodeling.
    Chen Li, Yike Li, Fei Liu, Boyao Lu, Shiyang Ye, Dexin Zhu, Muyun Wang, Junyu Chen, Cheng Zhou, Chunjie Li, Yanyan Zhang, Jiefei Shen.
      Neuro-glial mitochondrial transfer critically sustains neuronal function in disease. While this transfer reshapes inflammatory microenvironments, its pathological mechanisms in peripheral inflammatory pain remain uncharacterized, impeding targeted interventions. Here, employing primary satellite glial cells (SGCs)-trigeminal ganglion neurons (TGNs) co-culture models, we demonstrate that, during acute inflammation, SGCs transfer functional mitochondria to injured TGNs via tunneling nanotubes and free mitochondrial uptake. Inflammatory stress impairs mitophagy, leading to dysfunctional mitochondrial accumulation and heightened neuronal hyperexcitability. Mitochondria from SGCs restore mitophagic flux and enhance mitochondrial-endoplasmic reticulum (ER) contact sites, thereby facilitating calcium exchange and homeostasis while reducing neuronal hyperexcitability. Critically, Atl1 knockout and overexpression mice models reveal that ATL1-driven ER restructuring initiates autophagosome formation during mitophagy and regulates early-stage autophagic progression. Taken together, our findings uncover a neuroprotective axis wherein glial mitochondrial donation safeguards neurons, directly nominating mitochondrial dynamics for therapeutic intervention in orofacial inflammatory pain.
    Keywords:  ATL1; CP: cell biology; CP: neuroscience; endoplasmic reticulum; inflammatory pain; mitochondrial transplantation; mitophagy; trigeminal ganglion
    DOI:  https://doi.org/10.1016/j.celrep.2025.116809
  24. JHEP Rep. 2026 Feb;8(2): 101660
    FGF1 ameliorates hepatic steatosis through acute activation of the unfolded protein response and VLDL production.
    Tim van Zutphen, Dicky Struik, Weilin Liu, Sihao Liu, Benan Pelin Sermikli, Justina C Wolters, Henkjan J Verkade, Annette R Atkins, Michael Downes, Ronald M Evans, Johan W Jonker.
       Background & Aims: Metabolic dysfunction-associated steatotic liver disease (MASLD) is a serious chronic liver disease with limited therapeutic options. Fibroblast growth factor (FGF) analogs show promising therapeutic benefits for MASLD, yet the underlying mechanisms remain incompletely understood. Here, we studied the mechanism underlying the anti-steatotic properties of FGF1, the prototype member of the FGF family.
    Methods: The effect of FGF1 was studied in human and rodent hepatocytes and in obese mouse models exhibiting acute or chronic endoplasmic reticulum (ER) stress characteristic of MASLD. Metabolic analysis and proteomics were applied to evaluate liver physiology, ER stress and signaling.
    Results: We show that FGF1 reduces hepatic triglyceride (TG) levels in obese mice (51%, p <0.01, n = 8) via acute stimulation of very-low-density lipoprotein (VLDL, 3.9-fold, p <0.01, n = 8) secretion in an ER stress-dependent manner. This anti-steatotic effect was independent of adipose FGF receptor 1, which is required for the glucose-lowering effect of FGF1. Mechanistically, activation of the unfolded protein response (UPR), resulting in stabilization of apolipoprotein B (ApoB, 1.8-fold, p <0.01, n = 8), the main structural protein component of atherogenic lipoprotein particles, was identified as the key mechanism by which FGF1 drives VLDL secretion. Post-translational control of ApoB by FGF1 was potentiated by pre-existing ER stress. FGF1 stimulated major regulators of protein synthesis, and during ER stress, all three branches of the UPR were activated. In ER stress-primed lean mice, FGF1 adopted novel TG secretion activity (2.2-fold, p <0.05, n = 6). Conversely, alleviation of ER stress in obese mice suppressed FGF1-stimulated VLDL-TG production (49%, n = 11, p <0.05).
    Conclusion: These results define ER stress-dependent modulation of VLDL secretion as a mechanism underlying the anti-steatotic activity of FGF1. Targeting the FGF-UPR pathway may thus have therapeutic potential for treating MASLD.
    Impact and implications: Fibroblast growth factors show therapeutic potential in both preclinical models and clinical trials for treating metabolic dysfunction-associated steatotic liver disease, a highly prevalent condition with limited treatment options. Identifying the mechanisms underlying their anti-steatotic effects may accelerate clinical development. Our finding that triglyceride secretion is the major driver of the anti-steatotic action of FGF1, together with the involvement of an adaptive unfolded protein response, provides deeper insight into the therapeutic potential of this pathway. These results also highlight possible implications for liver physiology and for the circulating lipoprotein profile, with relevance for both efficacy and safety considerations.
    Keywords:  ER stress; Fibroblast growth factor 1; Unfolded Protein Response; VLDL secretion
    DOI:  https://doi.org/10.1016/j.jhepr.2025.101660
  25. PLoS Pathog. 2026 Jan 16. 22(1): e1013888
    Plant rhabdovirus glycoprotein activates unfolded protein response-mediated antiviral ER-phagy in insect vectors.
    Siyu Chen, Yu Cheng, Yupeng Tang, Jian Zhang, You Li, Dongsheng Jia, Hongyan Chen, Taiyun Wei.
      Although viral infection-induced endoplasmic reticulum autophagy (ER-phagy) is well characterized in mammalian systems, the mechanisms underlying arbovirus-triggered ER-phagy in insect vectors remain poorly understood. This study demonstrates that rice stripe mosaic virus (RSMV), a cytorhabdovirus transmitted by leafhopper vectors, activates the unfolded protein response (UPR) to induce ER-phagy as an antiviral defense mechanism. During viral assembly in the ER lumen, RSMV glycoprotein (G) disrupts the interaction between ER chaperone BiP and ER kinase PERK, leading to the release of PERK to activate subsequent signaling cascade. This ultimately activates the transcription factor ATF4, which regulates the expression of the autophagy-related gene ATG8, thereby linking the UPR to autophagy. Mechanistically, RSMV assembly promotes the formation of ER-derived amorphous inclusions that recruit ATG8 through interaction with ER-phagy receptor Sec62. This process culminates in the sequestration of both viral particles and ER fragments into autophagosomes, initiating ER-phagy triggered by viral infection. Functional studies confirmed that microinjection of RSMV G activates both the UPR and ER-phagy, while knockdown of PERK, ATF4, ATG8, or Sec62 significantly enhances viral accumulation, underscoring their essential antiviral roles. Our findings reveal a conserved nature of UPR-induced ER-phagy across vertebrate and invertebrate systems, advancing our understanding of arbovirus-vector interactions and antiviral defense mechanisms.
    DOI:  https://doi.org/10.1371/journal.ppat.1013888
  26. J Clin Invest. 2026 Jan 15. pii: e195538. [Epub ahead of print]
    Poly(ADP-ribose) glycohydrolase enforces p21 degradation via dePARylation to promote gastric cancer progression.
    Yangchan Hu, Qimei Bao, Yixing Huang, Yan Wang, Xin Zhao, Junjun Nan, Yuxin Meng, Mingcong Deng, Yuancong Li, Zirui Zhuang, Hanyi He, Dan Zu, Yuke Zhong, Chunkai Zhang, Bing Wang, Ran Li, Yanhua He, Qihan Wang, Min Liu, John A Tainer, Yin Shi, Xiangdong Cheng, Ji Jing, Zu Ye.
      Dysregulation of cell cycle checkpoints is a cancer hallmark with ubiquitination controlled protein stability playing pivotal roles. Although p21, a key cyclin-dependent kinase inhibitor, is tightly regulated by ubiquitin-mediated degradation, the key upstream modulators of its ubiquitination remain incompletely defined. Here, we identify poly(ADP-ribose) glycohydrolase (PARG) as a regulator of p21 stability in gastric cancer (GC) cells. We show that PARG expression is markedly upregulated in GC tissues and correlates with poor patient prognosis. Functional assays revealed that genetic depletion of PARG triggers G2/M phase arrest and impairs GC cell proliferation. Mechanistically, we demonstrate that PARG loss enhances p21 PARylation, which disrupts its association with E3 ubiquitin ligase, thereby reducing K48-linked ubiquitination and leading to p21 protein stabilization. Moreover, we identify lysine residues K161 and K163 as critical sites for PARG-mediated regulation of p21 ubiquitination. Our findings reveal a post-translational regulatory axis in which PARG governs cell cycle progression by modulating the PARylation-dependent ubiquitination of p21. These results broaden the understanding of p21 regulation in cancer and highlight PARG as a potential therapeutic target for GC treatment.
    Keywords:  Cell biology; Drug screens; Gastric cancer; Gastroenterology; Oncology; Ubiquitin-proteosome system
    DOI:  https://doi.org/10.1172/JCI195538
  27. bioRxiv. 2026 Jan 11. pii: 2026.01.09.698697. [Epub ahead of print]
    Chronic stress disrupts the network among stress granules, P-bodies, and motor proteins.
    Yuichiro Adachi, Masashi Masuda, Yutaka Taketani, Paul J Anderson, Pavel V Ivanov.
      Stress granules (SGs) are dynamic RNA granules that rapidly form in response to various stresses concurrent with mRNA translation shutdown, contributing to cellular adaptation and disease pathogenesis. While SG assembly and disassembly under acute stress have been extensively characterized, how SGs behave under chronic stress remains poorly understood. We previously reported that chronic stress preconditioning inhibits the earliest steps of SG assembly via translation-dependent mechanisms. In contrast, the regulation of SG maturation under chronic stress has not yet been investigated. Here, we show that chronic stress decreases SG size by disrupting the MYH9-dependent myosin crosstalk with core SG nucleator G3BP1. This defect leads to impaired SG and processing body (PB) docking as well, limiting the biogenesis of early SGs. Furthermore, chronic stress reduces expression of the SG nucleator UBAP2L, required for SG-PB docking, thus exacerbating these deficiences. In summary, chronic stress disrupts the myosin-SG-PB network and inhibits SG maturation in a translation-independent manner.
    Summary: Chronic stress disrupts the MYH9 network with G3BP1, which inhibits stress granule (SG) assembly. It also decreases UBAP2L levels, which inhibits SG and processing body docking. Disrupted MYH9 network further inhibits this docking, thus blocking maturation of SGs.
    DOI:  https://doi.org/10.64898/2026.01.09.698697
  28. Nucleic Acids Res. 2026 Jan 05. pii: gkaf1372. [Epub ahead of print]54(1):
    Eukaryotic initiation factor eIF3 facilitates loading of eukaryotic release factor eRF1 or suppressor tRNA to the ribosome.
    Ekaterina Shuvalova, Alexey Shuvalov, Walaa Al Sheikh, Alexandr Klishin, Nikita Biziaev, Elena Alkalaeva.
      Eukaryotic translation initiation factor eIF3 plays a pivotal role in 48S preinitiation complex assembly and ribosomal scanning. It binds simultaneously to the 40S ribosomal subunit and the eIF4F cap-binding complex, which, through its interaction with poly(A)-binding protein (PABP), facilitates closed-loop mRNA structure formation. PABP also interacts with the eukaryotic release factor eRF3, thereby co-localizing initiation and release factors, and suggesting potential functional crosstalk. Using a reconstituted mammalian translation system, we demonstrate that eIF3 significantly enhances translation termination. Specifically, eIF3 promotes the loading of eRF1 into the ribosomal A site, accelerating the GTPase activity of eRF3 and the rate of peptide release. We also show that eIF3 facilitates the binding of suppressor or near-cognate tRNAs to stop codons to enable readthrough and continued elongation. These findings establish a conserved, direct role of eIF3 in regulation both translation termination and stop codon readthrough, a mechanism particularly relevant within closed-loop mRNA structures and during upstream open reading frame translation.
    DOI:  https://doi.org/10.1093/nar/gkaf1372
  29. Cell Chem Biol. 2026 Jan 13. pii: S2451-9456(25)00426-X. [Epub ahead of print]
    Structural and mechanistic analysis of covalent ligands targeting the RNA-binding protein NONO.
    Garrett L Lindsey, Thomas K Hockley, Alejandro Villa Gomez, Andrew C Marshall, William R Brothers, Colin T Finney, Jacob Gross, Archa H Fox, Gene W Yeo, Bruno Melillo, Charles S Bond, Benjamin F Cravatt.
      RNA-binding proteins (RBPs) play important roles in mRNA transcription, processing, and translation. Chemical tools are lacking for RBPs, which has hindered efforts to perturb and understand RBP function in cells. We previously described a chloroacetamide compound (R)-SKBG-1 that covalently binds the RBP NONO and stabilizes its interactions with mRNAs, leading to transcriptional remodeling and suppression of cancer cell growth. Here, we report the crystal structure of an (R)-SKBG-1:NONO complex, which confirms covalent modification of cysteine-145 at a pocket proximal to the RNA-binding interface of the protein. We show that this pocket can also be targeted by a lower reactivity chlorofluoroacetamide analog (R, R)-GL-373, which retains the pharmacological properties of (R)-SKBG-1, including blockade of estrogen receptor expression in breast cancer cells, while displaying much greater proteome-wide selectivity. Our findings thus show that NONO can be targeted by covalent ligands with high specificity to pharmacologically suppress pro-tumorigenic gene products in cancer cells.
    Keywords:  NONO; RNA-binding proteins; activity-based protein profiling; chemical proteomics; chlorofluoroacetamide; covalent; cysteine; stereochemistry; transcriptome
    DOI:  https://doi.org/10.1016/j.chembiol.2025.12.010
  30. J Cell Sci. 2026 Jan 15. pii: jcs.264466. [Epub ahead of print]
    LAMP1 and LAMP2A localise to axonal organelles with distinct motility dynamics and partially overlapping molecular signatures in human neurons.
    Reem Abouward, Alya Masoud Abdelhafid, Oscar G Wilkins, Song-Yi Lee, Fairouz Ibrahim, Mark Skehel, Alice Ting, Nicol Birsa, Jernej Ule, Giampietro Schiavo.
      LAMP1 and LAMP2A are abundant proteins of late endosomal/lysosomal compartments that are often used interchangeably to label what is assumed to be the same organelle population, potentially obscuring distinct physiological roles. Here, we characterised the axonal transport dynamics of LAMP1- and LAMP2A-positive compartments in human iPSC-derived cortical neurons. We found that LAMP1-positive organelles move slower in the retrograde direction, pause more frequently, and display a broader anterograde velocity distribution than LAMP2A-positive vesicles, indicating distinct trafficking behaviours. Co-transport analysis revealed that approximately 65% of motile LAMP-positive organelles carry both markers, with higher co-transport in the retrograde direction. To explore molecular differences underlying these behaviours, we performed proximity labelling using full-length LAMP1 or LAMP2A fused to the light-activated biotin ligase, LOV-Turbo. This approach revealed largely overlapping interactomes, with LAMP2A-associated proteins forming a subset of the LAMP1 interactome and showing an enrichment for synaptic vesicle-related proteins. We further validated ZFYVE16 as a novel interactor of both compartments. Together, our findings indicate that LAMP1- and LAMP2A- positive organelles share overlapping molecular identities but represent functionally distinct axonal populations with divergent transport dynamics.
    Keywords:  Axonal transport; Endosomes; Lysosomes; Proximity labelling; Synaptic vesicles
    DOI:  https://doi.org/10.1242/jcs.264466
  31. Nature. 2026 Jan 14.
    N1-Methylpseudouridine directly modulates translation dynamics.
    Batsheva Rozman, Karin Broennimann, K Shanmugha Rajan, Aharon Nachshon, Chiranjeet Saha, Tamar Arazi, Vishnu Mohan, Tamar Geiger, Clayton J Wollner, Justin M Richner, Eric Westhof, Ada Yonath, Anat Bashan, Noam Stern-Ginossar.
      The considerable success of mRNA vaccines against SARS-CoV-2 has underscored the potential of synthetic mRNA as a transformative biomedical technology1. A critical feature of this approach is the incorporation of the modified nucleoside N1-methylpseudouridine (m1Ψ), which enhances antigen expression while reducing immunogenicity2-5. However, a comprehensive understanding of how m1Ψ influences translation remains incomplete. Here we use ribosome profiling at the subcodon resolution to show that m1Ψ increases ribosome density on synthetic mRNAs, leading to higher protein production independent of innate immune activation or eIF2α phosphorylation. We find that m1Ψ directly slows ribosome movement in defined sequence contexts while simultaneously promoting translation initiation. Structural studies using cryo-electron microscopy reveal that m1Ψ alters interactions within the ribosomal decoding centre, providing a mechanistic basis for slowed elongation. Furthermore, by introducing synonymous recoding that disrupts the modification-mediated changes in elongation, we show that the m1Ψ-dependent enhancement of protein output is modulated by codon composition, and that m1Ψ impact is strongest in mRNAs containing non-optimal codons with uridines at the wobble position. Together, these findings demonstrate that m1Ψ directly modulates translation dynamics, thereby increasing protein yield from synthetic mRNAs in specific sequence contexts.
    DOI:  https://doi.org/10.1038/s41586-025-09945-5
  32. Biophys J. 2026 Jan 09. pii: S0006-3495(26)00007-X. [Epub ahead of print]
    ClpA and ClpAP-Catalyzed Unfolding and Translocation are Differentially Coupled to ATP Binding.
    Liana Islam, Jaskamaljot Kaur Banwait, Nasib Karl Maluf, Aaron L Lucius.
      Proteome maintenance is underpinned by molecular motors from the AAA+ superfamily. E. coli ClpA is a representative AAA+ motor that associates with the tetradecameric serine protease ClpP forming the ATP-dependent protease, ClpAP. ClpA, unfolds substrates targeted for degradation and translocates them into the central channel of ClpP where the substrate is degraded. However, when ClpA is not associated with ClpP, the motor uses its unfolding activity to noncovalently remodel protein substrates. Although a large body of work exists on the mechanisms of ClpAP-catalyzed protein unfolding and degradation, much less is known about the mechanisms of protein remodeling reactions. In fact, there is a dearth of mechanistic information to complement the emerging static structural information on many AAA+ family members that remodel proteins without covalent modification. Here we report results from single-turnover stopped-flow experiments to interrogate the ClpA-catalyzed mechanisms of protein unfolding, both alone and when associated with ClpP. To this end, we used substrates containing tandem repeats of the Titin I27 domain. We show that both ClpA and ClpAP catalyze cooperative protein unfolding of the Titin I27 domain in a single kinetic step. This cooperative unfolding is followed by repeated rounds of translocation on the newly unfolded polypeptide. At saturating ATP, ClpA and ClpAP catalyze protein unfolding and translocation at (12.0 ± 0.4) aa s-1 and (33.2 ± 1.1) aa s-1, respectively. By examining the complete ATP-dependence of the reaction, we have deconvoluted the elementary rate constants for unfolding and translocation from the overall rate. At saturating ATP, the translocation rate constant is approximately 8 and 24-fold faster than the unfolding rate constant for ClpA and ClpAP, respectively. Furthermore, the unfolding rate constant for ClpAP is ∼3 fold faster than for ClpA alone. This indicates fundamental differences in the unfolding mechanisms between ClpA alone and ClpA associated with ClpP.
    DOI:  https://doi.org/10.1016/j.bpj.2026.01.007
  33. RNA. 2026 Jan 12. pii: rna.080824.125. [Epub ahead of print]
    Bridging single-molecule and genome-wide studies of cellular mRNA translation.
    Adam Koch, Kotaro Tomuro, Taisei Wakigawa, Tatsuya Morisaki, Shintaro J Iwasaki, Timothy J Stasevich.
      The translation of mRNA is a tightly regulated, energy-intensive process that drives cellular diversity. Understanding its control requires tools that can capture behavior across scales. Over the past two decades, two complementary techniques have emerged that have transformed our understanding of mRNA translation within cells: ribosome profiling (Ribo-Seq) and live, single-molecule imaging. Ribo-Seq provides genome-wide, codon-level maps of ribosome positions, revealing pause sites, novel open reading frames, and global translation efficiencies. In contrast, live, single-molecule imaging visualizes translation on individual mRNAs in living cells, uncovering heterogeneous initiation, elongation, pausing, and spatial organization in real time. Together, these methods offer complementary strengths - molecular breadth versus temporal and spatial precision - but are rarely applied in tandem. Here, we review their principles, key discoveries, and recent innovations that are bringing them closer together, including endogenous tagging, higher-throughput imaging, absolute calibration, and spatially resolved footprinting. Integrating these approaches promises a unified, multiscale view of translation that connects the dynamics of individual ribosomes to genome-wide patterns of protein synthesis.
    DOI:  https://doi.org/10.1261/rna.080824.125
  34. Nat Protoc. 2026 Jan 12.
    Hydrogen/deuterium exchange mass spectrometry analysis of ribosome-nascent chain complexes to study protein biogenesis at the peptide level.
    Alžběta Roeselová, Aleksandra Pajak, Thomas E Wales, Grant A Pellowe, Svend Kjær, John R Engen, David Balchin.
      Nascent proteins begin to fold during their synthesis, while still attached to the ribosome. The dynamic nature of ribosome-nascent chain complexes (RNCs) poses a challenge for conventional structural biology approaches, limiting our understanding of dynamic cotranslational events. Hydrogen-deuterium exchange mass spectrometry (HDX-MS) is a powerful label-free technique for studying the conformational equilibria and refolding of full-length proteins with peptide resolution. However, the large size of the ribosome and the need for stable, highly homogeneous samples have hindered the application of HDX-MS to RNCs. Here we present a strategy for analysing conformational dynamics and interactors of Escherichia coli RNCs using HDX-MS. High-quality RNCs are obtained through the gentle lysis of high-density cultures expressing uniformly stalled ribosomes, followed by ultracentrifugation and tag-based affinity purification. Peptide-resolution information on protein conformational dynamics is obtained by pulse deuterium labeling, quenching with an RNA-compatible low pH buffer and offline digestion with pepsin. Extensive data analysis with use of specific internal controls allows for the confident assignment of mass spectra to specific peptides, ensuring good coverage of the nascent chain and ribosomal proteins. This method provides a valuable complement to existing structural techniques such as cryo-electron microscopy and nuclear magnetic resonance, and enables detailed characterization of large, partially structured nascent chains and their interactions with the ribosomal proteins and molecular chaperones. The protocol takes 1-3 months, from sample preparation and data acquisition to data analysis, and requires standard expertise in cloning and protein purification and intermediate expertise in HDX-MS.
    DOI:  https://doi.org/10.1038/s41596-025-01279-w
  35. Int J Biol Sci. 2026 ;22(2): 731-749
    Parkin Deficiency Impairs ER-Mitochondria Associations and calcium homeostasis via IP3R-Grp75-VDAC1 Complex.
    Nai-Jia Xue, Yi Liu, Zhi-Hao Lin, Wen-Hao Huang, Feng Zhang, Ran Zheng, Xiao-Li Si, Lu-Yan Gu, Yi Fan, Jia-Li Pu, Bao-Rong Zhang.
      Disruption of mitochondria-associated endoplasmic reticulum membranes (MAMs) and calcium homeostasis has been implicated in the pathogenesis of Parkinson's disease (PD). Parkin, a PD-associated E3 ubiquitin ligase, has been shown to regulate MAM integrity and calcium dynamics. However, the mechanisms of Parkin recruitment and its substrate specificity have not been well understood. This investigation has demonstrated that loss of Parkin enhances ER-mitochondria associations and leads to excessive calcium flux in MAM, resulting in abnormal mitochondrial permeability transition pore (mPTP) opening and decreased cell viability. Further, Parkin physically interacts with IP3R-Grp75-VDAC1 complex at ER-mitochondria contact sites, where it is recruited by IP3R-mediated calcium flux and mitophagy. More importantly, Parkin deficiency leads to the accumulation of IP3R levels, particularly in MAM region. In addition, Parkin fine-tunes the stability of the complex and ubiquitinates IP3R for degradation via the ubiquitin-proteasomal system, ensuring suitable calcium transfer. Taken together, our study reveals a novel role of Parkin in regulating ER-mitochondria contacts, providing insights into PD pathogenesis and potential therapeutic strategies targeting MAMs.
    Keywords:  IP3R; Parkin; calcium; mitochondria-associated ER membrane; ubiquitination
    DOI:  https://doi.org/10.7150/ijbs.121759
  36. Autophagy. 2026 Jan 14. 1-3
    Muscle meets Lysosomes: emerging strategies in muscular dystrophy.
    Abbass Jaber, David Israeli.
      Duchenne muscular dystrophy (DMD) is caused by the loss of DMD (dystrophin), leading to sarcolemmal fragility and progressive muscle degeneration. Although adeno-associated viral (AAV) microdystrophin (µDMD) therapies have advanced clinically, their benefits remain partial, highlighting the need to identify secondary cellular defects that limit therapeutic efficacy. In our recent study, we demonstrated that lysosomal dysfunction is a conserved, intrinsic, and persistent feature of DMD pathology. Using mouse, canine, and human dystrophic muscle, we show marked lysosomal membrane permeabilization (LMP), impaired acidification, defective proteolysis, and inefficient membrane repair, all hallmarks of compromised lysosomal integrity. Cholesterol accumulation within dystrophic myofibers further exacerbates these defects, linking lipid dysregulation to lysosomal injury and accelerated muscle degeneration. We find macroautophagy/autophagy impairment in DMD stems in part from reduced autophagosome-lysosome fusion, reframing autophagy failure as a downstream consequence of lysosomal damage. µDMD gene therapy only partially corrects these abnormalities and does not fully restore lysosomal stability. In contrast, combining µDMD with the lysosome-activating disaccharide trehalose produces synergistic benefits, improving muscle strength, architecture, and molecular signatures beyond either treatment alone. These findings position lysosomal dysfunction as a central driver of DMD pathophysiology and support therapeutic strategies that pair gene restoration with lysosomal enhancement.Abbreviation: AAV: adeno-associated virus; DAGC: DMD-associated glycoprotein complex; DMD: Duchenne muscular dystrophy; FDA: Food and Drug Administration; LMP: lysosome membrane permeabilization; MTOR: mechanistic target of rapamycin kinase; µDMD: microdystrophin.
    Keywords:  Autophagy; Duchenne muscular dystrophy; galectin-3; lysosome; microdystrophin
    DOI:  https://doi.org/10.1080/15548627.2026.2615985
  37. Trends Cell Biol. 2026 Jan 14. pii: S0962-8924(25)00265-X. [Epub ahead of print]
    Expanding roles of N-glycosylation in the endoplasmic reticulum.
    Mengxiao Ma, Rajat Rohatgi.
      N-linked glycosylation in the endoplasmic reticulum (ER), catalyzed by two oligosaccharyltransferase (OST) complexes, has long been viewed as a constitutive post-translational modification. Recent discoveries suggest that OST complexes play a much more plastic and directive role in regulating ER processes. Here, we review this work and focus on one specific mechanism that uses N-glycosylation to regulate the stability of the ER chaperone HSP90B1. This degradative process regulates the cell-surface abundance of multiple signaling receptors that are HSP90B1 clients: toll-like receptors, WNT receptors, and growth factor receptors. This unusual system enables the status of ER-based processes to influence the sensitivity of cells to extracellular signals, with implications for tissue growth and development, inflammation, and immune function.
    Keywords:  chaperone; endoplasmic reticulum; glycosylation; oligosaccharyltransferase; signaling receptor; translation
    DOI:  https://doi.org/10.1016/j.tcb.2025.12.001
  38. Arch Toxicol. 2026 Jan 14.
    Arsenic disrupts autophagosome-lysosome fusion in a zinc dependent manner across multiple human skin and lung cells lines.
    Rachel V Goff, Sidimohamed Elmoustapha, Shelia D Thomas, Mayukh Banerjee.
      Chronic environmental arsenic exposure causes skin and lung cancers, but the molecular mechanisms are poorly understood. We identified that chronic trivalent inorganic arsenite (iAs) exposure at population relevant 100 nM concentration activates autophagy while suppressing downstream protein degradation. Here, we studied the mechanisms by which environmental iAs exposure uncouples autophagy activation from autophagic protein degradation across human skin and lung cell lines. During autophagy, zinc regulates the critical autophagosome-lysosome fusion (ALF) step connecting initiation to final protein degradation. iAs disrupts zinc dependent processes. Thus, we hypothesized that iAs suppresses autophagy by compromising ALF. We demonstrate that environmental 100 nM iAs exposure specifically targets the ALF step of autophagy to suppress autophagic protein degradation across multiple skin and lung cell line models. We show that iAs suppresses ALF in a zinc dependent manner. Physiological zinc supplementation (1 μM) prevented and rescued against iAs-induced suppression of ALF and autophagic protein degradation in the short and long-term. Our work provides a framework to understand and further investigate the precise molecular mechanisms by which chronic environmental iAs exposure disrupts global protein degradation, thereby inducing proteotoxicity across multiple target tissues and contributing to the observed proteome-wide differential expression patterns during multi-organ carcinogenesis.
    Keywords:  Arsenic; Autophagosome–lysosome fusion; Autophagy; Protein degradation; Zinc
    DOI:  https://doi.org/10.1007/s00204-025-04230-w
  39. Sci Adv. 2026 Jan 16. 12(3): eaeb5297
    Enterococcus faecalis redox metabolism activates the unfolded protein response to impair wound healing.
    Aaron Ming Zhi Tan, Cenk Celik, Stella Yue Ting Lee, Mark Veleba, Caroline S Manzano, Rahim M K Abdul, Guillaume Thibault, Kimberly A Kline.
      Enterococcus faecalis is an opportunistic pathogen that thrives in biofilm-associated infections and delays wound healing, yet how it impairs host tissue responses is unclear. Here, we identified extracellular electron transport (EET) as a previously unrecognized source of reactive oxygen species (ROS) in E. faecalis and showed that this activity directly triggers the unfolded protein response (UPR) in epithelial cells and delays epithelial cell migration. ROS detoxification with catalase suppressed E. faecalis-induced UPR and rescued epithelial cell migration, while exogenous hydrogen peroxide was sufficient to restore UPR activation in EET-deficient strains. UPR disruption by pharmacological inhibition also impaired cell migration, highlighting a critical role for UPR homeostasis in wound repair. Our findings establish EET as a virulence mechanism that links bacterial redox metabolism to host cell stress and impaired repair, offering previously unidentified avenues for therapeutic intervention in chronic infections.
    DOI:  https://doi.org/10.1126/sciadv.aeb5297
  40. Nat Commun. 2026 Jan 12.
    Small molecule splicing modulators that disrupt O-GlcNAc homeostasis.
    Steven S Cheng, Alison C Mody, Amedeo Vetere, Ashwin Govindan, Yewon Lee, Danielle E Khost, Rohan Narayan, Timothy B Sackton, Nicholas K Conrad, Bridget K Wagner, Christina M Woo.
      O-Linked N-acetylglucosamine (O-GlcNAc) is a nucleocytoplasmic post-translational modification that is tightly regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). Dysregulation of O-GlcNAc in human disease has motivated efforts to therapeutically modulate O-GlcNAc. Drug repurposing efforts can accelerate these campaigns and unveil how clinically relevant compounds and pathways intersect with O-GlcNAc. Here we report the results of three parallel drug repurposing screens against the O-GlcNAc cycling enzymes in cells and in vitro that reveal kinase inhibitors GSK690693 and Y-33075 act as splicing modulators that disrupt O-GlcNAc homeostasis and simultaneously downregulate OGT and OGA. These effects are independent of their respective annotated targets, AKT and ROCK, and are distinct from OGT and OGA inhibitors and similar kinase inhibitors. Evaluation of a panel of splicing modulators revealed three additional potent compounds (OTS964, indisulam, GNF2133) that similarly downregulate OGT and OGA with distinct splicing profiles. These findings reveal previously unobserved splicing modulator chemotypes and approaches to disrupt O-GlcNAc homeostasis.
    DOI:  https://doi.org/10.1038/s41467-025-68271-6
  41. J Clin Invest. 2026 Jan 16. pii: e193636. [Epub ahead of print]136(2):
    FBXO11 suppression rewires an NPM1-centered interactome influencing the progression of myelodysplastic syndrome.
    Madeline Niederkorn, Lavanya Bezavada, Anitria Cotton, Lance E Palmer, Lahiri Konada, Trent Hall, Vishwajeeth R Pagala, Jinbin Zhai, Zuo-Fei Yuan, Yingxue Fu, Jacob A Steele, Shilpa Narina, Andrew Schild, Chengzhou Wu, Sarah Aminov, Michael Schieber, Erin McGovern, Aaron B Taylor, Sandeep Gurbuxani, Peng Xu, Peng Ji, Laura J Janke, Anthony A High, Guolian Kang, Shondra M Pruett-Miller, Mitchell Weiss, Amit Verma, Raajit K Rampal, John D Crispino.
      Myelodysplastic syndromes (MDSs) are malignant hematopoietic stem and progenitor cell (HSPC) disorders that lead to ineffective blood production with poor outcomes. We previously showed that F-box only protein 11 (FBXO11) is downregulated in MDS, and here we report how this event contributes to disease progression. Integration of multiomics data revealed that the SCF-FBXO11 complex regulates spliceosome and ribosome components in a nucleophosmin 1 (NPM1)-centric network. FBXO11 facilitates the ubiquitylation of NPM1, whereby deletion of FBXO11 results in the reorganization of NPM1 and a de-repression of alternative splicing. Label-free total quantitative proteomics demonstrated that the FBXO11-NPM1 interactome was markedly downregulated in cells from patients with CD34+ MDS. In addition, we discovered that MYC was evicted from the FBXO11 promoter by TLR2 activation, revealing that it was a MYC target gene and explaining why FBXO11 expression was decreased in MDS. In MDS mouse models, genetic ablation of Fbxo11 exacerbated neutropenia concomitant with a profound decrease in NPM1 protein levels. Finally, we discovered rare mutations in FBXO11, which mapped to a previously unstudied functional intrinsically disordered region (IDR) in the N-terminus responsible for binding NPM1. These data support a model in which FBXO11 rewires RNA binding and ribosomal subnetworks through ubiquitylation of NPM1, ultimately restricting MDS progression.
    Keywords:  Hematology; Leukemias; Oncology; Ubiquitin-proteosome system
    DOI:  https://doi.org/10.1172/JCI193636
  42. J Am Chem Soc. 2026 Jan 13.
    Allosteric PROTACs: Expanding the Horizon of Targeted Protein Degradation.
    Aileen Frost, Suzanne O'Connor, Alessio Ciulli.
      Proteolysis-targeting chimeras (PROTACs) have transformed the concept of chemical intervention in biological systems by co-opting the ubiquitin-proteasome system to selectively degrade proteins. A key promise of this modality is that proximity alone─not inhibition─is required, allowing binding anywhere on the protein surface to trigger degradation. Yet despite this conceptual freedom, most PROTACs to date have been built from orthosteric inhibitors. The use of allosteric or functionally silent ligands remains a largely untapped opportunity. In this Perspective, we spotlight pioneering efforts in allosteric PROTAC design and explore how such strategies could unlock improved outcomes for target selectivity, efficacy, and resistance management while also modulating physicochemical properties to enhance in vivo performance. We further discuss the practical and conceptual challenges and the advances needed to make allosteric targeting a mainstream strategy in the design of protein degraders and other proximity-inducing molecules.
    DOI:  https://doi.org/10.1021/jacs.5c14840
  43. Physiology (Bethesda). 2026 Jan 16.
    Organelle dysfunction and TNT-mediated aggregate spreading in neurodegeneration.
    Valentine Thomas, Chiara Zurzolo.
      Organelle dysfunction is a central hallmark of neurodegenerative diseases (NDs), which are characterized by the pathological accumulation of misfolded proteins capable of inducing aggregation in healthy cells. This process generates a self-perpetuating cycle of protein misfolding and spreading across interconnected neuronal networks. In this review, we provide an integrated overview of organelle alterations associated with major NDs, emphasizing the pivotal roles of lysosomes, mitochondria, and the endoplasmic reticulum (ER) at the crossroads of proteostasis, metabolism, and stress signaling. We examine how defects in these organelles create conditions that favor aggregate formation and cellular vulnerability, with a focus on α-synuclein and Tau, the main aggregating proteins in Parkinson's and Alzheimer's diseases, respectively. We then explore mechanisms of intercellular protein transfer, highlighting the emerging role of tunneling nanotubes (TNTs). We discuss how organelle status influences TNT formation and cargo selection, and how TNTs may act as conduits for the propagation of pathogenic aggregates. Finally, we summarize the downstream consequences of TNT-mediated transfer in recipient cells, including alterations in the autophagy-lysosomal pathway, TFEB-dependent transcription, mitochondrial stress responses, calcium homeostasis, and inflammatory or senescent signaling. Together, these insights underscore the intertwined roles of organelle dysfunction and TNT-mediated communication in driving the progression of NDs and suggest new therapeutic avenues aimed at restoring organelle function and limiting aggregate spread.
    Keywords:  neurodegeneration; organelle dysfunction; protein aggregates; tunneling nanotubes
    DOI:  https://doi.org/10.1152/physiol.00048.2025
  44. BMB Rep. 2026 Jan 12. pii: 6696. [Epub ahead of print]
    Stress Granules as a Central Hub Linking Organelle Stress, Aging, and Neurodegeneration.
    Hyun-Ji Ham, Jin-A Lee.
      Stress granules (SGs) are dynamic cytoplasmic assemblies composed of RNAs and proteins that form in response to cellular stress, serving to halt translation and protect cellular integrity. In neurons, SGs mediate adaptive, pro-survival responses to acute stress; however, their dysregulation has been increasingly associated with both aging and neurodegenerative diseases. Aging neurons frequently exhibit changes in SG dynamics - with an increased propensity to form SGs while displaying reduced efficiency in their clearance - resulting in persistent granules that can facilitate the accumulation of pathological protein aggregates (e.g., TDP-43 or tau). Aberrant SG formation and defective clearance mechanisms are implicated in the pathogenesis of key neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), Alzheimer's disease (AD), and Parkinson's disease (PD). Recent findings have shown that SGs interface with organelles such as lysosomes, mitochondria, and the endoplasmic reticulum, utilizing autophagic and other protein quality-control mechanisms for clearance. As these clearance pathways progressively decline with age, SGs can transition from promoting cellular adaptation to contributing to cellular dysfunction. In this mini-review, we examine how aging influences SG biology, detail the role of SGs in neurodegenerative diseases, and discuss emerging mechanistic insights and therapeutic strategies aimed at modulating SG dynamics in the context of brain aging.
  45. bioRxiv. 2026 Jan 08. pii: 2026.01.07.698279. [Epub ahead of print]
    PAR-Driven Condensation Maintains Stalled Replication Fork Stability.
    Lei Zhang, Zeyu Zhang, Timothy R O'Leary, Arkadi Shwartz, Guoyun Kao, Patrick R Griffin, Yong Zhang.
      Poly(ADP-ribose) (PAR) is a nucleic acid-like heterogeneous polymer in nature. Recently, it was found to engage in liquid-liquid phase separation (LLPS), generating condensates as an emerging class of subcellular structures with pivotal functions in response to stimuli. As a post-translational modification catalyzed by PAR polymerases (PARPs), PAR is known to modulate many key events in cells. However, its involvement in biomolecular condensation remains elusive. Through an imaging-based screening of small molecules with diverse biological activities, we here discovered that PAR undergoes LLPS upon inhibiting proteasome in different types of cells, resulting in co-condensation of PAR with proteasome and ubiquitin chains in nucleus. This unprecedented co-condensation is dependent on PARP2 not PARP1 and requires K6-linked ubiquitylation. PAR is shown for the first time to directly interact with ubiquitin chains. Notably, stalled DNA replication forks arose from proteasome inhibition are co-localized with PAR-proteasome-ubiquitin chain condensates. By attenuating replication and stabilizing stalled replication forks, PAR-proteasome-ubiquitin chain condensates sustain genomic integrity under proteasomal stress. This work demonstrates a self-protective mechanism in stressed cells and provides fundamental understanding of PAR condensation in cell biology.
    DOI:  https://doi.org/10.64898/2026.01.07.698279
  46. Proc Natl Acad Sci U S A. 2026 Jan 20. 123(3): e2516291123
    Predicting epistasis across proteins by structural logic.
    Michelle Tang, Gareth A Cromie, Anowarul Kabir, Martin S Timour, Julee Ashmead, Russell S Lo, Nathaniel Corley, Frank DiMaio, Hiroki Morizono, Ljubica Caldovic, Nicholas Ah Mew, Andrea Gropman, Amarda Shehu, Aimée M Dudley.
      Accurately predicting the phenotypic consequences of genetic variation is a major challenge for precision medicine. The problem is exacerbated by epistatic interactions, nonadditive effects between genetic variants that produce unexpected phenotypes. Here, we explore an understudied form of positive epistasis: intragenic complementation, in which pairs of loss-of-function variants restore near wild-type protein function. Using mutational scanning in yeast, we identify thousands of such interactions in a clinically important enzyme, human argininosuccinate lyase (ASL). Restoration of protein function is not due to the biochemical properties of the substituted amino acids, but rather to a structural feature of the protein, the active site assembly. We develop a machine learning algorithm that uses protein language model embeddings to predict intragenic complementation in ASL with 99.6% accuracy. Additionally, the model trained on ASL generalizes to a structurally related but sequence-divergent enzyme, fumarase, with accuracy over 90%. Our findings reveal a structural basis for this form of epistasis and provide a predictive framework that could extend to at least 4% of human proteins.
    Keywords:  epistasis; machine learning; variant effects
    DOI:  https://doi.org/10.1073/pnas.2516291123
  47. Cell Death Dis. 2026 Jan 16. 17(1): 47
    NRF1 is upregulated by docosahexaenoic acid to ameliorate MASH through the inhibition of ER stress.
    Mengchi Lin, Hongtao Zhang, Shuai Chen, Jie Zhang, Chenxi Tang, Xin Song, Jiaming Zhou, Zixin Xu, Yali Mu, Hang Zeng, Changqing Yang, Chaohui Yu, Chengfu Xu.
      Despite the high prevalence of metabolic dysfunction-associated steatohepatitis (MASH), the number of effective therapeutic targets is limited due to a vague understanding of its intricate pathogenesis. In this study, we reported that the expression of nuclear factor erythroid-derived 2-related factor 1 (NRF1), an endoplasmic reticulum (ER) membrane-bound transcription factor that governs the expression of proteasome subunit genes, was significantly reduced in liver tissues from MAFLD patients and from mice fed a high-fat diet (HFD) for 20 weeks. Liver-specific overexpression of NRF1 in mice markedly ameliorated HFD-driven hepatic steatosis, liver injury and inflammation. Elevated NRF1 expression restored the function of the proteasome, facilitating the degradation of unfolded and nonfunctioning proteins, thereby mitigating ER stress and reducing oxidative stress. Moreover, docosahexaenoic acid (DHA) was found to increase NRF1 expression, contributing to the amelioration of MASH. Mechanistically, DHA inhibited the ubiquitination of NRF1 via the cytoplasmic E3 ligases FBW7 and HRD1 at the ER membrane, thereby preventing its degradation. Liver-specific knockdown of NRF1 abrogated the protective effect of DHA on HFD-driven MASH in mice. Together, our findings underscore the pivotal role of NRF1 in the DHA-mediated amelioration of MASH and suggest that NRF1 is a potential therapeutic target for MASH management.
    DOI:  https://doi.org/10.1038/s41419-025-08139-1
  48. FEBS J. 2026 Jan 14.
    The ALS-associated E425K mutation uncouples DNAJC7 from the Hsp70 chaperone cycle.
    Bar Elmaleh, Ofrah Faust, Rina Rosenzweig.
      DNAJC7, a member of the J-domain protein (JDP/Hsp40) family, plays a key role in protein homeostasis by regulating Hsp70 activity and preventing protein aggregation. Mutations in DNAJC7 have been linked to amyotrophic lateral sclerosis (ALS); yet, the molecular mechanisms by which these variants impair chaperone function remain poorly understood. DNAJC7 is a conserved chaperone featuring both a canonical J-domain, essential for Hsp70 activation, and three TPR domains, which serve as protein-protein binding interfaces. Here, we investigate the structural and functional consequences of the ALS-associated E425K mutation located within the conserved J-domain. Using NMR spectroscopy, we show that although the E425K mutation does not alter the structure of the protein, it significantly disrupts the conserved J-domain-Hsp70 interaction. We further identify a second Hsp70-binding interface within the TPR domains, which interacts with the C-terminal EEVD motif of Hsp70. This TPR-EEVD interaction is preserved in the E425K mutant but cannot compensate for the loss of J-domain binding or restore DNAJC7-dependent Hsp70 activation. Functionally, we show that the TPR domains of DNAJC7 directly bind TDP-43 and prevent its aggregation and that this holdase activity is retained in the E425K mutant. However, the mutant fails to support client transfer to Hsp70 and the subsequent Hsp70-mediated substrate refolding. Together, these findings demonstrate that DNAJC7 requires coordinated action of both J-domain and TPRs to regulate Hsp70 function and that disruption of J-domain-mediated activation uncouples DNAJC7 from the Hsp70 cycle, providing a mechanistic basis for its dysfunction in ALS.
    Keywords:  ALS; DNAJC7; Hsp70; Molecular chaperones; NMR spectroscopy
    DOI:  https://doi.org/10.1111/febs.70395
  49. ACS Synth Biol. 2026 Jan 17.
    Multilayer Host Engineering of Saccharomyces cerevisiae to Enhance Cricket Paralysis Virus (CrPV) Internal Ribosome Entry Site Mediated Translation.
    Zeyu Cao, Xiaotong Zou, Koichi Ito, Kei Endo.
      Internal ribosome entry sites (IRESs) provide compact RNA elements for noncanonical translation and hold promise as building blocks for RNA-based regulation in synthetic biology. However, the cricket paralysis virus (CrPV) IRES shows very low activity in Saccharomyces cerevisiae, limiting its broader utility despite extensive structural and biochemical studies. Here we report a yeast engineering strategy that enhances CrPV IRES-mediated translation by combining host modifications at three mechanistically distinct levels: translation initiation, tRNA modification, and mRNA stability. A reporter-based screen revealed host factors that influence IRES activity and uncovered a trade-off between IRES stimulation and maintenance of cap-dependent translation required for growth. Stepwise integration of nonsense-mediated decay deficiency, a tad3 temperature-sensitive allele, and wild-type eIF4E overexpression yielded a strain with up to an order-of-magnitude increase in reporter output compared with that of the parental strain. These results establish a proof-of-principle framework for host engineering of noncanonical translation.
    Keywords:  Cricket paralysis virus IRES; Translational control; Yeast engineering; mRNA stability; tRNA modification
    DOI:  https://doi.org/10.1021/acssynbio.5c00744
  50. Bio Protoc. 2026 Jan 05. 16(1): e5559
    Generating ER-TRG and CA-ER-TRG Knock-in Mice and Quantitative In Vivo Imaging of ER-phagy.
    Mengyuan Zhang, Yu Zhang, Yongjuan Sang, Qiming Sun.
      ER-phagy, a selective autophagy process crucial for maintaining cellular homeostasis by targeting the endoplasmic reticulum (ER), has been challenging to study in vivo due to the lack of suitable spatiotemporal quantification tools. Existing methods like electron microscopy, biochemical assays, and in vitro reporters lack resolution, scalability, or physiological relevance. Here, we present a detailed protocol for generating two transgenic mouse models: ER-TRG (constitutively expressing an ER lumen-targeting tandem RFP-GFP tag) and CA-ER-TRG (Cre-recombinase-activated ER-TRG). Additionally, we outline procedures for quantitative imaging of ER-phagy in vivo, covering tissue preparation, confocal microscopy, and signal analysis. This protocol offers a robust and reproducible tool for investigating ER-phagy dynamics across various tissues, developmental stages, and pathophysiological conditions, facilitating both fundamental and translational research. Key features • Enables live, single-cell resolution imaging of ER-phagy dynamics across intact tissues in mice. • Features a Cre-recombinase-activated knock-in model (CA-ER-TRG) for spatiotemporally controlled ER-phagy studies in specific cell types. • Quantifies ER-phagy flux via pH-sensitive RFP-GFP signal ratiometry and lysosomal co-localization in vivo.
    Keywords:  CA-ER-TRG mice; ER-TRG mice; ER-phagy; In vivo imaging; Quantitative analysis
    DOI:  https://doi.org/10.21769/BioProtoc.5559
  51. Mol Biol Rep. 2026 Jan 10. 53(1): 272
    Loss of SEL1L or Hrd1 increases intrinsic LOX levels in HEK293 cells independently of proteasomal degradation.
    Ryoichi Murase, Izumi Nagae, Genki Kato, Maiko Yasui, Mutsumi Kawasumi, Mahmoud Kandeel, Fumiaki Sato, Kentaro Oh-Hashi.
       BACKGROUND: We previously established HEK293 cells deficient in SEL1L, a key component of endoplasmic reticulum-associated protein degradation (ERAD), and attempted to identify factors affected by ERAD dysfunction through comprehensive MS analysis. Among the factors increased by SEL1L-deficiency, lysyl oxidase (LOX) which plays a role in extracellular matrix cross-linking was involved.
    METHODS AND RESULTS: To elucidate the regulatory mechanisms of LOX mRNA and protein, we applied several ERAD-deficient cells and drug treatments. In SEL1L-deficient cells, LOX mRNA expression levels were observed to increase approximately twofold, and the increase in high molecular weight preproLOX protein was more remarkable in each SEL1L deletion. SEL1L-deficiency also slightly increased the amount of secreted LOX protein cleaved by proteases. Increased proproLOX was also observed in cells lacking Hrd1 that associates with SEL1L. Interestingly, LOX protein was hardly increased in cells deficient for EDEM2 and TXNDC11, which are involved in mannose trimming of N-glycosylated proteins. Since ERAD failure may induce sustained ER stress, we examined the effects of three ER stress inducers on LOX expression. Changes in LOX mRNA following 6-h treatment with each reagent was negligible compared to the major ER stress-inducible mRNA. In contrast, long-term treatment with thapsigargin and tunicamycin increased intracellular LOX protein, but significantly decreased secreted LOX protein. Finally, we examined the effects of proteasome and lysosome inhibitors on LOX expression and, unexpectedly, each reagent hardly increased LOX mRNA or protein levels.
    CONCLUSIONS: These results suggest that expression of intrinsic LOX protein is regulated by SEL1L and Hrd1 in a ubiquitin-proteasome-independent manner.
    Keywords:  ER stress; ERAD; Hrd1; LOX; SEL1L
    DOI:  https://doi.org/10.1007/s11033-025-11388-0
  52. FASEB J. 2026 Jan 31. 40(2): e71441
    Manipulation of the Unfolded Protein Response by Intracellular Bacterial Pathogens: Mechanisms of ER Hijacking and Therapeutic Implications.
    Enhui Dai, Dongjie Sun, Yanxiao Zhao, Mengtao Zhang, Yifan Wu, Jiabo Ding.
      The unfolded protein response (UPR) is a cellular stress response mechanism that maintains endoplasmic reticulum (ER) homeostasis through three signaling pathways mediated by IRE1α, PERK, and ATF6 sensors. While UPR's role in viral infections has been well documented, recent studies indicate that intracellular bacterial pathogens have evolved specific mechanisms to hijack UPR signaling for survival and replication. This review examines UPR manipulation strategies employed by major bacterial pathogens, including Brucella, Mycobacterium tuberculosis, Legionella, and Salmonella. These pathogens utilize effector proteins that target specific UPR components: Brucella effectors VceC, BspB, TcpB, and BspL interact with ER chaperones and ERAD machinery; M. tuberculosis proteins Rv0297, ESAT-6, HBHA, and CdhM disrupt calcium homeostasis and alter ER morphology; Legionella Lpg0519 activates atypical ATF6 signaling; and bacterial toxins including cholera toxin bind IRE1α structural motifs for pathway activation. The molecular basis of UPR manipulation includes direct protein-protein interactions, calcium signaling interference, ER morphological disruption, and transcriptional program modulation. Bacterial hijacking of UPR pathways affects ER-phagy processes and host immune responses, facilitating intracellular survival. UPR pathway components serve as potential targets for host-directed therapy against persistent and drug-resistant infections. Small molecule modulators targeting IRE1α kinase activity, PERK inhibitors, and ATF6 pathway regulators may complement conventional antimicrobial approaches. Characterization of these host-pathogen interactions provides insights for developing therapeutic strategies that target bacterial dependencies on cellular stress responses.
    Keywords:  bacterial infection; effector proteins; endoplasmic reticulophagy; immune response; therapeutic strategy; unfolded protein response
    DOI:  https://doi.org/10.1096/fj.202503547R
  53. EMBO J. 2026 Jan 15.
    MYO5A-mediated stabilization promotes the acquisition of fusion competence in sealed autophagosomes.
    Akshaya Nambiar, René Martin, Kamakshi Tomar, Hans-Joachim Knölker, Sandhya P Koushika, Subramaniam K, Ravi Manjithaya.
      Autophagy requires precise regulation of autophagosome-lysosome fusion, yet the molecular details of this process remain incompletely understood. Here, we identify the class V myosin MYO5A as a critical regulator of autophagic flux. The genetic or pharmacological inhibition of MYO5A in Saccharomyces cerevisiae, mammalian cells, or Caenorhabditis elegans blocked autophagic flux by preventing autophagosome-lysosome fusion. MYO5A facilitates the maturation of autophagosomes into fusion-competent intermediates as its loss altered the localization of fusion machinery on autophagosomes and reduced the pool of stationary autophagosomes, a step that proved critical for subsequent fusion with lysosomes. Domain mapping and targeted mutagenesis revealed that two LIR motifs (PAYRVL and QAYIGL) within the coiled-coil and globular tail domains of MYO5A mediate its direct interaction with LC3 on autophagosomes. Live imaging in mammalian cells and C. elegans added support for this role, revealing how MYO5A regulates autophagic flux to ensure fusion. Together, these findings establish MYO5A as a regulator of autophagy and highlight its potential as a target for fine-tuning autophagic flux.
    Keywords:  Actomyosin Dynamics; Autophagic Flux; Autophagosome–Lysosome Fusion; MYO5A; Unconventional Myosins
    DOI:  https://doi.org/10.1038/s44318-025-00686-9
  54. bioRxiv. 2026 Jan 11. pii: 2026.01.09.698724. [Epub ahead of print]
    Degradation of the TEAD•YAP/TAZ Transcription Factor Complex by Heterobifunctional Small Molecules that Bind to the TEAD Allosteric Lipid Pocket.
    I-Ju Yeh, Mona K Ghozayel, Khuchtumur Bum-Erdene, Samy O Meroueh.
      Central kinases of the Hippo tumor suppressor pathway phosphorylate the transcriptional coactivators YAP and TAZ to sequester them in the cytoplasm. In cancer, Hippo pathway kinases have reduced activity, leading to translocation of YAP and TAZ into the nucleus where they engage TEADs and other transcription factors. Here, we explore whether heterobifunctional small molecules that bind to the TEAD allosteric lipid-binding pocket can degrade the TEAD•YAP/TAZ complex. We design and synthesize heterobifunctional molecules that consist of flufenamic acid analogs that bind to the allosteric TEAD lipid pocket, a long and flexible linker, and thalidomide to engage E3 ubiquitin ligase component cereblon. The bifunctional compounds promote ternary complex formation in biochemical assays and mammalian cells but exhibited modest degradation of TEAD, YAP, and TAZ in cancer cells. Methyl ester analogs of these compounds led to substantial proteasomal degradation of the TEAD•YAP/TAZ complex in cancer cells. This work provides a strategy for depletion of nuclear YAP and TAZ and for exploration of their TEAD-dependent and TEAD-independent activities in vivo .
    DOI:  https://doi.org/10.64898/2026.01.09.698724
  55. Mol Syst Biol. 2026 Jan 12.
    Limited proteolysis-coupled mass spectrometry captures proteome-wide protein structural alterations and biomolecular condensation in living cells.
    Franziska Elsässer, Roberta Florea, Felix Räsch, Mostafa Zedan, Nesli-Ece Sen, Tim Pflästerer, Tatjana Kleele, Robbie Loewith, Karsten Weis, Natalie de Souza, Paola Picotti.
      The function of a protein is determined by its structure, which may change dynamically in response to post-translational modifications, interaction with other molecules, or environmental factors like temperature. Limited proteolysis-coupled mass spectrometry (LiP-MS) captures such structural alterations on a proteome-wide scale via the detection of altered protease susceptibility patterns of proteins. However, this technique has so far required cell lysis, which exposes proteins to non-native conditions and can disrupt labile interactions such as those occurring within biomolecular condensates. To study protein structures directly within cells, we developed in-cell LiP-MS. We optimized conditions for introduction of proteinase K into human cells using electroporation and validated that intracellular cleavage occurs. In-cell LiP-MS captured the known binding of rapamycin to FKBP1A within the cell. Moreover, it detected global protein structural alterations upon sodium arsenite treatment and captured the structural dynamics of hundreds of proteins from biomolecular condensates with peptide level resolution and within live human cells. The data allowed monitoring of structural alterations of individual sites on the involved proteins, such as known RNA-binding and intrinsically-disordered regions, and dissected the timing of the different events. We detected known (G3BP1) and novel structural alterations of proteins from stress granules as well as from nuclear speckles and validated alteration of nuclear speckles by fluorescence microscopy and of the protein SERBP1 by polysome profiling. Our dataset further provides a resource describing the structural changes of human proteins in response to a cellular stress leading to biomolecular condensation and pinpoints structurally altered regions. Comparison of LiP-based structural fingerprints before and after cell lysis revealed which human proteins are susceptible to structural change upon cell lysis, therefore guiding the design of future experiments requiring native protein structures.
    Keywords:  Biomolecular Condensation; LiP-MS; Nuclear Speckles; Stress Granules; Structural Proteomics
    DOI:  https://doi.org/10.1038/s44320-025-00182-6
  56. Elife. 2026 Jan 15. pii: RP106791. [Epub ahead of print]14
    Functionally coupled ion channels begin co-assembling at the start of their synthesis.
    Roya Pournejati, Jessica M Huang, Michael Ma, Claudia M Moreno, Oscar Vivas.
      Calcium binding to BK channels lowers BK activation threshold, substantiating functional coupling with calcium-permeable channels. This coupling requires close proximity between different channel types, and the formation of BK-CaV1.3 hetero-clusters at nanometer distances exemplifies this unique organization. To investigate the structural basis of this interaction, we tested the hypothesis that BK and CaV1.3 channels assemble before their insertion into the plasma membrane. Our approach incorporated four strategies: (1) detecting interactions between BK and CaV1.3 proteins inside the cell, (2) identifying membrane compartments where intracellular hetero-clusters reside, (3) measuring the proximity of their mRNAs, and (4) assessing protein interactions at the plasma membrane during early translation. These analyses revealed that a subset of BK and CaV1.3 transcripts are spatially close in micro-translational complexes, and their newly synthesized proteins associate within the endoplasmic reticulum (ER) and Golgi. Comparisons with other proteins, transcripts, and randomized localization models support the conclusion that BK and CaV1.3 hetero-clusters form before their insertion at the plasma membrane.
    Keywords:  BK channel; cell biology; clustering; functional coupling; molecular biophysics; none; rat insulinoma cell line; structural biology; structural coupling; voltage-gated Ca channel
    DOI:  https://doi.org/10.7554/eLife.106791
  57. Redox Biol. 2025 Dec 24. pii: S2213-2317(25)00501-4. [Epub ahead of print]90 103988
    Amino acid restriction sensitizes lung cancer cells to ferroptosis via GCN2-dependent activation of the integrated stress response.
    Viktor Antonsson Garellick, Nadia Gul, Parvin Horrieh, Dyar Mustafa, Angana A H Patel, Martin Dankis, Samantha W Alvarez, Johanna Berndtson, Saeed Mahdavi, Maria Schwarz, Andreas Persson, Fikret Zahirovic, Clotilde Wiel, Volkan I Sayin, Per Lindahl.
      Lung cancer cells are vulnerable to iron-dependent oxidation of phospholipids leading to ferroptosis, a process countered by glutathione peroxidase-4 that converts lipid hydroperoxides to lipid alcohols using glutathione as reducing agent. Since ferroptosis-inducing agents are in clinical development, identifying modifiers of ferroptosis susceptibility is warranted. Here, we investigate the impact of amino acids on susceptibility to buthionine sulfoximine (BSO), a glutamate-cysteine ligase inhibitor that blocks biosynthesis of glutathione. We found that reduced amounts of amino acids other than cysteine increased the sensitivity to BSO and other ferroptosis-inducing agents, in a panel of mouse and human lung cancer cells, without affecting glutathione production. Activation of the amino acid sensor protein GCN2 and the integrated stress response lowered the threshold for lipid peroxidation by promoting ATF4-dependent mitochondrial respiration and reactive oxygen species leakage from the electron transport chain under glutathione depletion. The finding provides new insights into lung cancer metabolism and raises the possibility of using amino acid restricted diets in combination with ferroptosis-inducing agents as cancer therapies.
    Keywords:  Amino acids; Ferroptosis; Glutathione; Integrated stress response; Lung cancer; Mitochondrial respiration
    DOI:  https://doi.org/10.1016/j.redox.2025.103988
  58. Food Sci Biotechnol. 2026 Jan;35(2): 391-397
    Alleviation of ER stress via targeted genetic engineering enhances recombinant ovalbumin secretion in Saccharomyces cerevisiae.
    Eun Bi Yoon, Kyoung Chan Jin, Yu Jeung Lim, Joong-Hyuck Auh, Hyun Gyu Choi, Sun-Ki Kim.
      Enhancing the secretion of heterologous proteins in Saccharomyces cerevisiae is often hindered by endoplasmic reticulum (ER) stress and inefficiencies in intracellular trafficking. To address these limitations, we systematically evaluated 14 genetic modifications associated with the secretory pathway to improve recombinant ovalbumin (OVA) production. Deletion of PAH1, a negative regulator of phospholipid biosynthesis, resulted in the highest OVA secretion (5.68 mg/L), representing a 74% improvement over the background strain-likely due to increased ER membrane biogenesis. Similarly, knockout of GOS1, a Golgi-to-ER SNARE protein, enhanced secretion, possibly by reducing retrograde trafficking and limiting ER retention of secretory cargo. In contrast, disruption of endosome-to-Golgi transport via VPS5 deletion resulted in intracellular accumulation of OVA and complete secretion failure. These findings suggest that alleviating ER stress and modulating vesicular trafficking are key strategies for enhancing protein secretion in yeast, offering a functional basis for rational host strain engineering.
    Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-025-02044-1.
    Keywords:  ER stress; Ovalbumin; Saccharomyces cerevisiae; Vesicular trafficking
    DOI:  https://doi.org/10.1007/s10068-025-02044-1
  59. Acta Pharmacol Sin. 2026 Jan 12.
    Benchmarking co-folding methods to predict the structures of covalent protein-ligand complexes.
    Tong-Han Zhang, Jin-Tao Zhu, Zhi-Xian Huang, Juan Xie, Jian-Feng Pei, Lu-Hua Lai.
      Targeted covalent inhibitors (TCIs) are emerging as a new modality in drug discovery because of their strong binding affinity and prolonged target engagement. However, the rational design of TCIs remains a significant challenge and is hindered by the lack of methods that accurately predict the structures of covalent protein-ligand complexes. Recent advances in co-folding approaches have made substantial strides in modeling complex biomolecular structures. Despite significant progress, their performance profiles for predicting the structures of covalent protein-ligand complexes remain largely unexplored because of the absence of rigorous benchmarks. Here, we introduce CoFD-Bench, a comprehensive benchmark dataset comprising 218 recently resolved covalent complexes designed to systematically evaluate both classical docking methods (AutoDock-GPU, CovDock, and GNINA) and deep learning co-folding models (AlphaFold3 (AF3), Chai-1, and Boltz-1x). Our results demonstrate that co-folding methods achieve superior ligand RMSD accuracy and protein-ligand interaction recovery. However, their performance markedly declines for novel pocket-ligand pairs. In contrast, classical docking methods exhibit stable but modest performance, which is primarily limited by target conformations. Furthermore, computational efficiency evaluations show that co-folding methods are slower than classical approaches, posing challenges for large-scale predictions. We also reveal that AF3 has the potential to identify native covalent residues through noncovalent co-folding, with a ligand RMSD comparable to that of covalent co-folding. These findings offer a possible route to explore covalent binding without prior specification of reactive residues, which are often unknown in real-world scenarios. Our study provides crucial insights and new opportunities for future co-folding-based TCI design, informing future model applications and improvements. CoFD-Bench offers rigorous evaluation criteria, diverse docking scenarios, and various methodological baselines, positioning it as an important benchmark for future model development and assessment.
    Keywords:  benchmarking; co-folding methods; covalent complex prediction; targeted covalent inhibitors
    DOI:  https://doi.org/10.1038/s41401-025-01721-5
  60. Nat Commun. 2026 Jan 14.
    Comprehensive mapping of RNA modification dynamics and crosstalk via deep learning and nanopore direct RNA-sequencing.
    Han Dong, Yongsheng Gao, Zhengyi Cai, Yi Li, Xing Li, Fangqing Zhao, Jinyang Zhang.
      Despite the extensive studies of individual RNA modifications, the lack of methods to detect multiple modification types simultaneously has left the global epitranscriptomic landscape and its underlying crosstalk largely unexplored. Here, we present ORCA (Omni-RNA modification Characterization and Annotation), a deep learning framework that enables comprehensive mapping of RNA modification landscape using nanopore direct RNA sequencing. ORCA employs domain adversarial learning to detect and quantify a wide range of modifications by leveraging mixed stoichiometry-driven signal and sequence variability between modified and unmodified nucleotides. It also incorporates a transfer learning module for accurate annotation of modification types with minimal prior knowledge. Applying ORCA to multiple human cell lines reveals widespread, isoform-specific modification patterns, as well as intricate cooperative and competitive interactions among neighboring modification sites. This approach substantially expands the repertoire of known RNA modification sites and elucidates their spatial organization, revealing the emerging roles of RNA modifications in splicing regulation. ORCA thus provides an unbiased and generalizable framework for decoding RNA modification dynamics and their regulatory complexity across diverse biological contexts.
    DOI:  https://doi.org/10.1038/s41467-026-68419-y
  61. Proc Natl Acad Sci U S A. 2026 Jan 20. 123(3): e2519770123
    A bacterial translation activator with an intrinsically disordered RNA-binding region.
    Pallabi Basu, Elizabeth A Farland, James C Charity, Kade A Townsend, Simon L Dove.
      Bacterial RNA-binding proteins (RBPs) that control the translation of multiple transcripts act largely as negative regulators. Here, we report the identification and characterization of a positive regulator of translation (called PhaF) in the opportunistic pathogen Pseudomonas aeruginosa. Using CLIP-seq and CLAP-seq we identify upward of 50 transcripts targeted by PhaF. We demonstrate that PhaF acts to stimulate the translation of target mRNAs by binding upstream of the Shine-Dalgarno sequence using one or more of the multiple KPAA motifs located in an intrinsically disordered region of the protein. Importantly, we show that PhaF plays a key physiological role in P. aeruginosa through its translational control of the pslA transcript required for exopolysaccharide synthesis and biofilm formation. Our findings uncover an activator of translation in bacteria that binds target transcripts using an RNA-binding region reminiscent of those that are prominent in eukaryotic RBPs.
    Keywords:  IDR; PhaF; Pseudomonas aeruginosa; PslA; post-transcriptional regulator
    DOI:  https://doi.org/10.1073/pnas.2519770123
  62. EMBO Mol Med. 2026 Jan 12.
    Extracellular vesicle-based targeted protein degradation platform for multiple extracellular proteins.
    Bide Tong, Xiaoguang Zhang, Dingchao Zhu, Yulei Wang, Junyu Wei, Zixuan Ou, Huaizhen Liang, Hanpeng Xu, Zhengdong Zhang, Jie Lei, Xingyu Zhou, Di Wu, Yu Song, Kun Wang, Xiaobo Feng, Lei Tan, Zhiwei Liao, Cao Yang.
      Targeted protein degradation (TPD) is an emerging therapeutic approach that enables the degradation of undruggable targets via intracellular degradation systems. Extracellular vesicles (EVs) have shown potential to act as next-generation TPD platforms. However, the molecular mechanism underlying their degradation remains unknown, which restricts their application in TPD. In this study, we found that the autophagy-mediated lysosomal pathway was the major route by which EVs were degraded. MAP1LC3B recognized the LIR motifs of SQSTM1 and induced the degradation of EVs in the autophagy pathway. Based on the EV degradation mode, we developed an EV-based targeted protein degradation platform (EVTPD) using EVs loaded with the LIR motif of SQSTM1 as a degradation signal. Additionally, target protein-binding domains were integrated into the EVTPD to capture target proteins. EVTPD selectively degraded extracellular proteins without requiring receptors on target cells. Furthermore, dual-targeting EVTPD effectively degraded both TNF-α and IL-1β and exhibited potent anti-inflammatory effects in rat and goat models of intervertebral disc degeneration. This study has established a modular EV-based TPD strategy with multi-targeting potential.
    Keywords:  Autophagy; Extracellular Vesicles; Inflammation; Intervertebral Disc Degeneration; Targeted Protein Degradation
    DOI:  https://doi.org/10.1038/s44321-025-00371-8
  63. Plant Biotechnol (Tokyo). 2025 Dec 25. 42(4): 383-388
    Enhancement of bZIP60 function through C-terminal region translated after splicing in Arabidopsis.
    Yuji Iwata, Hiroyuki Mizoguchi, Nozomu Koizumi.
      The unfolded protein response (UPR) is a central regulatory pathway that ensures the proper function of the endoplasmic reticulum (ER) through efficient protein folding and quality control. In Arabidopsis, bZIP60 mRNA is activated by an IRE1-mediated unconventional splicing that excises a 23-nucleotide intron, resulting in the spliced form (bZIP60s mRNA) that encodes the active bZIP60 transcription factor lacking a transmembrane domain. In this study, we investigated the functional role of the spliced form-specific C-terminal extension, hereafter referred to as ORF2. Transient expression assays in Arabidopsis mesophyll protoplasts demonstrated that full-length bZIP60s potently activates the BiP3 promoter compared to a truncated variant lacking ORF2. Fusion of ORF2 to transcription factors unrelated to the UPR did not enhance their transcriptional potency, underscoring its specialized role in the context of bZIP60s. Furthermore, mutation in a conserved nuclear localization signal within ORF2 decreased promoter activation by bZIP60s. Fusion of ORF2 to GFP enhanced the nuclear localization of GFP. Our results suggest that ORF2 is critical for the full transcriptional activity of bZIP60s to ensure an efficient UPR.
    Keywords:  Arabidopsis thaliana; bZIP transcription factor; endoplasmic reticulum; nuclear localization signal; unfolded protein response
    DOI:  https://doi.org/10.5511/plantbiotechnology.25.0603a
  64. Proc Natl Acad Sci U S A. 2026 Jan 20. 123(3): e2513222123
    Piezo1 dictates K+ homeostasis through coordinated regulation of the ubiquitin ligase Kelch-like 3 in RBCs and the kidney.
    Kenichi Ishizawa, Ken Kaseda, Wataru Fujii, Yoshihiro Tomomitsu, Daigoro Hirohama, Osamu Yamazaki, Yuta Kochi, Shigeru Shibata.
      The maintenance of potassium (K+) balance is a fundamental biological process involving multiple tissues. However, the roles of intertissue crosstalk in K+ homeostasis remain poorly understood. Here, we demonstrate that the mechanosensor Piezo1 dictates extracellular K+ homeostasis by orchestrating the ubiquitin ligase Kelch-like 3 (KLHL3) activity in red blood cells (RBCs; erythrocytes) and the kidney. Genetic variants within KLHL3 with expression quantitative trait locus effects are associated with altered RBC parameters, and CRISPR-generated KLHL3 knock-in (KLHL3-KI) mice carrying a nonphosphorylatable Ala substitution at its activation site (Ser433) reveal that KLHL3 regulates erythrocyte volume by modulating with-no-lysine 1 (WNK1). In wild-type, but not in KLHL3-KI, erythrocytes, Piezo1 activates KLHL3 through Ser433 dephosphorylation, reducing WNK1 abundance and intracellular K+ content-a physiologically adaptive response to hypo-osmotic stress. KLHL3-KI mice exhibit hyperkalemia and reduced fractional K+ excretion, accompanied by elevated WNK levels and reduced renal outer medullary K+ (ROMK) abundance in collecting ducts of the kidney. Single-cell transcriptomics confirm coexpression of Piezo1 and KLHL3 in these segments, where Piezo1 regulates WNK abundance through KLHL3-Ser433 dephosphorylation. In human genetic studies of 200,367 UK Biobank participants, the PIEZO1 missense variant rs563555492 (p.L2277M) is independently associated with lower urinary K+. Piezo1-mediated WNK1 regulation is abolished in human kidney cells expressing Piezo1L2277M. Causal role of Piezo1 in regulating K+ excretion and ROMK was confirmed in vivo. These findings identify Piezo1-KLHL3 interaction as a key intertissue signaling mechanism between erythrocytes and the kidney that governs K+ homeostasis, and suggest this pathway as a therapeutic target for dyskalemia.
    Keywords:  electrolyte homeostasis; interorgan communication; ubiquitin proteasome system
    DOI:  https://doi.org/10.1073/pnas.2513222123
  65. Angew Chem Int Ed Engl. 2026 Jan 16. e19830
    Chemical Proteomics Identifies Ketogenesis-Mediated Cysteine Modifications Regulating Redox Function.
    Yuan-Fei Zhou, Ling Zhang, Zhuoyi L Niu, Xin Wang, Alejandro Storper, Ryan Hunt, Yingming Zhao, Nima Sharifi, Zhipeng A Wang.
      All the studies of ketogenesis-dependent post-translational modifications (PTMs), notably those mediated by ketone bodies, β-hydroxybutyrate (Bhb) and acetoacetate (Acac), have focused on lysine acylations. However, given the chemically diverse and reactive nature of metabolites generated, it remains unclear whether non-lysine modifications can also happen. Here, we develop an acetoacetate-alkyne (Acac-alkyne) chemical probe that enables efficient metabolic labeling, robust fluorescent visualization, and site-specific identification of Acac-modified proteins. By combining chemical proteomics with open-search strategy, we showed that Acac induces previously uncharacterized cysteine modifications in mammalian cells. Notably, cysteine crotonation (Ccr) is validated by employing both probe-based and standard peptide-based co-elution assays. Metabolic pathway tracing further identifies BDH1 and ECHS1 as key enzymes that generate Ccr formation. We further demonstrate that Ccr at PRDX3 C229 site impairs dimerization and redox activity, linking this newly discovered modification to the regulation of cellular reactive oxygen species. Together, these findings establish ketone metabolism as a novel source of cysteine modifications and provide an alternative mechanistic pathway to explain the profound biological effects of ketone bodies.
    Keywords:  Cysteine modifications; Ketone body; Protein modifications; Proteomics; Redox regulation
    DOI:  https://doi.org/10.1002/anie.202519830
  66. Nucleic Acids Res. 2026 Jan 14. pii: gkag017. [Epub ahead of print]54(2):
    Strategic variations in sarbecovirus and merbecovirus Nsp1 linker regions for translation inhibition.
    Ruixi Yan, Mingbo Wu, Xiangyu Ge, Qianqian Jin, Moyu Wang, Haolong Zhou, Yan Li, Yue Wang, Shuai Yuan.
      Nonstructural protein 1 (Nsp1) is a key virulence factor of coronaviruses, and its stable binding to the 40S ribosomal mRNA entry channel facilitates multiple functions, including suppression of host immune responses and degradation of host mRNA. To understand the structural basis of the conserved protein across viral lineages, we determined the cryo-EM structures of Nsp1-40S complexes of four coronaviruses from wild animals. Our results show that all Nsp1 proteins engage the mRNA entry channel via their C-terminal domain (CTD), but do not fully restrict the rotational mobility of the 40S head, which retains ∼5° of movement and repositions the Nsp1 linker region. Comparative analysis revealed distinct patterns in the linker regions connecting the N- and CTDs. Sarbecovirus Nsp1 contains a longer linker, whereas the merbecovirus Nsp1 adopts a shorter linker that navigates structural constraints more readily. Functionally, we find that linker length correlates with translation inhibition efficiency, suggesting a structural tuning mechanism. Additionally, variations in linker and helix 1 of the CTD among different lineages may serve as molecular markers for viral classification. Together, our results provide a comparative structural framework for understanding how coronavirus Nsp1 proteins modulate host translation and reflect evolutionary adaptations in ribosome engagement.
    DOI:  https://doi.org/10.1093/nar/gkag017
  67. Drug Discov Today. 2026 Jan 12. pii: S1359-6446(26)00006-1. [Epub ahead of print] 104601
    LYTACs and other extracellular targeted protein degradation TACnologies: guiding principles and potential clinical prospects.
    Björn Niebel, Wanyin Chen, Kaori Mukai, Christophe Henry.
      Lysosome-targeting chimeras (LYTACs) have emerged as a powerful modality in the field of extracellular targeted protein degradation (eTPD). The proximity-inducing mode of action of LYTACs can be applied to a growing number of lysosome-targeting receptors (LTRs) and membrane-bound E3 ligases to degrade extracellular proteins. In this review, we highlight preceding eTPD approaches and discuss in depth the plethora of newly identified LTRs that can be exploited in a LYTAC molecule. To provide guidance in the fast-growing LYTAC field, we elaborate on parameters to assess the preclinical validation of the various TACnologies, highlight opportunities for engineering catalytic LYTACs, and finish with pharmacokinetic (PK) considerations.
    Keywords:  LYTAC; extracellular targeted protein degradation; preclinical development
    DOI:  https://doi.org/10.1016/j.drudis.2026.104601
  68. J Cheminform. 2026 Jan 12.
    Proteolysis-targeting Chimera efficacy prediction using a deep-learning-QSP model.
    Sungwoo Goo, Jina Kim, Soyoung Lee, Sangkeun Jung, Jung-Woo Chae, Jae-Mun Choi, Hwi-Yeol Yun.
      This study presents an integrated computational modeling framework combining deep learning and Quantitative Systems Pharmacology (QSP) to predict the efficacy of PROTAC (PROteolysis Targeting Chimera) molecules. PROTACs have emerged as promising therapeutics for targeted protein degradation (TPD), offering significant advantages in addressing proteins that traditional small-molecule inhibitors cannot target. However, experimental evaluation of PROTAC efficacy is hindered by extensive variability in molecular configurations, necessitating efficient computational prediction methods. The proposed model integrates binding affinity predictions from DeepCalici, a convolutional neural network-based deep learning model, with a mechanistic QSP Hook model to estimate key pharmacodynamic parameters, notably half-maximal degradation concentration(DC50) and maximal degradation(Dmax). This study utilized curated experimental data from PROTAC-DB, including experimentally validated DC50 and Dmax values. The dissociation constants (Kd) between PROTAC molecules and their protein targets (POI) or E3 ligases were predicted using DeepCalici and, then incorporated into the Hook model. To enhance the prediction accuracy, a supplementary deep neural network adjusted the hook model parameters based on chemical and biochemical features. The integrated modeling approach achieved a strong predictive performance for DC50, demonstrating its practical value in prioritizing effective PROTAC candidates. However, the predictions for Dmax were less accurate, likely reflecting the variability in the experimental conditions not captured in the current dataset. This study highlights the critical importance of comprehensive structural data for accurate modeling of PROTAC efficacy and suggests future improvements using standardized experimental data. Such integrative modeling approaches promise to accelerate the discovery and optimization of PROTAC therapeutics.
    Keywords:  Deep learning; PROteolysis TArgeting Chimera; Quantitative systems pharmacology
    DOI:  https://doi.org/10.1186/s13321-026-01152-2
  69. Cell Rep. 2026 Jan 13. pii: S2211-1247(25)01611-0. [Epub ahead of print]45(1): 116839
    Differentiation in the human urothelia is defined by distinct alternative polyadenylation.
    Ninh B Le, Surbhi Sona, Briana Santo, Yi Zhang, Rosie Ou, R Allen Schweickart, Veena Kochat, William I Padron, Kunal Rai, Shih-Han Lee, Joo Mi Yi, Oliver Wessely, Byron H Lee, Angela H Ting.
      Distinct epithelial cell states arise during differentiation, but mechanisms generating transcriptomic diversity among them remain poorly defined. The human ureter urothelium contains basal progenitor, intermediate cells, and terminally differentiated umbrella cells. Prior single-cell RNA sequencing revealed similar global gene expression profiles across these states, raising the question of how distinct identities emerge. Here, we show that alternative cleavage and polyadenylation (APA) introduces a major layer of transcriptomic diversity during urothelial differentiation, largely independent of changes in mRNA levels. Analysis of 13,544 urothelial cells identified hundreds of differentiation-associated APA events. Single-cell imaging revealed spatially specific APA patterns, and reporter assays demonstrated gene- and context-dependent control of protein expression by alternative 3' untranslated regions (3' UTRs), consistent with in situ protein patterns. Conserved motifs in APA-regulated 3' UTRs, including transcription factor binding sites and Alu elements, suggest mechanisms for polyadenylation site selection. Our study establishes APA as a key contributor to transcriptomic complexity in the human urothelium.
    Keywords:  APA; CP: Genomics; CP: Molecular biology; alternative cleavage and polyadenylation; human urothelium; mRNA isoform; poly(A); polyadenylation; single-cell RNA sequencing; single-cell spatial gene expression; urothelial differentiation
    DOI:  https://doi.org/10.1016/j.celrep.2025.116839
  70. Mol Biol Cell. 2026 Jan 15. mbcE25080384
    A shared binding interface controls Vps13 organelle-specific targeting independently of its vacuolar protein sorting function.
    Kevin Ryan Jeffers, Samantha Katarzyna Dziurdzik, Michael Davey, Jordan Faith Drotsky, Elizabeth Conibear.
      Yeast vacuolar protein sorting 13 (Vps13) is a bridge-like transporter that directs lipid flow between membranes at organelle contact sites. Vps13 targeting relies on organelle-specific adaptors containing proline-X-proline (PxP) motifs, which compete for binding to the Vps13 adaptor-binding (VAB) domain. Though a VAB-PxP interface has been identified for the mitochondrial adaptor Mcp1, whether other adaptors use identical binding mechanisms is unknown. Moreover, not every Vps13 function is connected to a known PxP adaptor, suggesting other adaptors may exist. Here, we validate the significance of the shared VAB-PxP interface by showing that mutations within this region inhibit both adaptor binding and Vps13 membrane targeting in vivo. Using predictive modeling, we demonstrate that while adaptors share a common Vps13-binding interface, slight differences between these interfaces may contribute to preferential binding and adaptor competition. Notably, we find that the VPS pathway functions independently of the PxP motif binding site. Our results indicate that Vps13 likely employs a non-PxP adaptor mechanism in this pathway, yet the structural integrity of the VAB domain remains essential for proper pathway function.
    DOI:  https://doi.org/10.1091/mbc.E25-08-0384
  71. mBio. 2026 Jan 12. e0344825
    Candida albicans-induced ubiquitination of EGFR reveals novel host-fungal interaction pathways.
    Léa Lortal, James S Griffiths, Emily L Priest, Alexander Kempf, Olivia K A Paulin, Nicole O Ponde, Antzela Tsavou, Don N Wickramasinghe, Andrew Donkin, Claire M Lyon, Olivia W Hepworth, Jonathan P Richardson, Julian R Naglik.
      Candida albicans causes severe mucosal and systemic infections, with hypha formation playing a key role in its virulence. Hyphal invasion via endocytosis is mediated predominantly through interactions between Als3p and the epidermal growth factor receptor (EGFR). Subsequent EGFR activation by candidalysin, a hyphal-secreted cytolytic peptide toxin encoded by the ECE1 gene, induces receptor signaling and immune responses. While EGFR ubiquitination critically regulates receptor trafficking and signaling, its involvement during C. albicans infection has remained unexplored. Here, we demonstrate that C. albicans induces EGFR ubiquitination, leading to altered trafficking and lysosomal degradation in an ECE1- and ALS3-dependent manner. This correlates with changes in EGFR ligand expression, adaptor recruitment, and protein ubiquitination in oral epithelial cells. In a mouse model of oropharyngeal candidiasis, wild-type C. albicans and ece1Δ/Δ and als3Δ/Δ mutant strains were found to differentially regulate Egfr expression, ubiquitin pathway-associated genes, and protein ubiquitination. Furthermore, conditional EGFR knockout was protective during infection. Together, our findings reveal that C. albicans infection modulates the host ubiquitin system, including direct effects on EGFR, highlighting a novel aspect of host-fungal interactions.IMPORTANCECandida albicans is a common fungal pathogen that causes both mucosal infections, such as thrush, and life-threatening systemic diseases. A key step in infection is the fungus invading epithelial tissues and activating the host epidermal growth factor receptor (EGFR). We discovered that C. albicans alters how EGFR is regulated by inducing its ubiquitination, a modification that leads to receptor degradation. This process depends on two major fungal virulence factors: the adhesin Als3p and Ece1p, the polypeptide that contains the candidalysin toxin. The fungus also broadly increases protein ubiquitination in oral epithelial cells. In a mouse model of oral infection, loss of EGFR in epithelial tissues reduced disease severity, suggesting that the receptor helps the fungus establish infection. These findings reveal a previously unrecognized strategy by which C. albicans manipulates protein ubiquitination and regulation in epithelial cells, offering new insights into fungal pathogenesis and potential therapeutic approaches that target host pathways.
    Keywords:  Candida albicans; candidalysin; epidermal growth factor receptor; oropharyngeal candidiasis; ubiquitin
    DOI:  https://doi.org/10.1128/mbio.03448-25
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