bims-proteo Biomed News
on Proteostasis
Issue of 2026–10–04
fifty-nine papers selected by
Eric Chevet, INSERM
  1. RNF185 destabilizes specific membrane proteins that span all ERAD branches.
  2. Cotranslational membrane protein biogenesis by an EMC-bound translocon.
  3. Conformational diversity of collided ribosomes determines stress signaling.
  4. Atg39-dependent nucleophagy and Atg40-dependent ER-phagy require the Apq12/Brr6/Brl1 complex.
  5. Excess Met1-linked ubiquitination leads to solid aggregate formation.
  6. RABC1 regulates ER-phagy to maintain er homeostasis in plants.
  7. Targeted extracellular degradation of LRP8 promotes ferroptosis in cancer cells.
  8. Listeria promotes systemic infection by targeting the host deubiquitinase CYLD.
  9. Distinct roles for TANGO1S domains in maintaining ER-Golgi architecture.
  10. ZNFX1, an immunoregulatory RNA helicase and E3 ubiquitin ligase, assembles into pleiomorphic polymers.
  11. Substrate Binding Relieves Autoinhibition of the Mammalian GCN2 Kinase.
  12. A redox switch on CDC48 regulates peroxisomal function via controlling peroxisome-associated protein degradation.
  13. Mitochondria-ER-lipid droplet crosstalk regulates metabolic adaptation to shear stress via VMP1.
  14. Single residues within the YTHDF intrinsically disordered region control m 6 A-dependent mRNA decay.
  15. NAT10 governs development through the Xrp1-mediated stress response rather than its RNA acetylation activity.
  16. FBXL21 regulates diurnal proteostasis in skeletal muscle by targeting DNAJB6 and client proteins.
  17. A lipid transfer-dependent feedback loop activates ATG9A compartments in autophagy initiation.
  18. Proteostasis as a Therapeutic Backbone: Proteasome Inhibitors and Emerging Regimens.
  19. Ubiquitin chain editing promotes p97/VCP-dependent removal of RPA from DNA.
  20. Collided ribosomes are rescued by the endonuclease Rae1 and trans- translation.
  21. A functional screen for bacterial ubiquitin regulation identifies an unusual Pseudomonas aeruginosa E3 ligase.
  22. A Versatile DNA-Encoded Library Platform for the Discovery of Highly Cooperative Chemical Inducers of Proximity.
  23. A TBK1/ULK1 signalling axis couples lysosomal stress to TFEB activation.
  24. Pathogenic mutations in ATAD3A cause dysregulation of RagC/D-TFEB axis and disrupt lysosomal homeostasis.
  25. GPCR antagonism via rewiring of receptor trafficking and degradation.
  26. More than the 'master regulator of cytosolic proteostasis': new roles for HSF1 in promoting tissue health.
  27. The Cytoplasmic MicroRNA-Production Complex Regulates MicroRNA Sorting to Extracellular Vesicles During Synaptogenesis.
  28. A small molecule increases mycobacterial translation fidelity without a fitness cost by targeting ribosomal protein S5.
  29. Redox activation of ATG5 licenses autophagy upon nutrient restriction.
  30. Chondroitin sulfate shapes the unfolded protein response in osteosarcoma to sustain cancer cell survival during ER stress.
  31. Protein rheostasis: quality control in metastable proteomes.
  32. RBP-Driven RNA sorting and local translation establish the molecular identities of individual sensory axons.
  33. Tracking cellular biomolecular condensate dynamics under proteostatic stress with middle-down phosphoproteomics.
  34. Enzyme Activation of Triggerable Polymeric Lysosome Targeting Chimeras.
  35. Targeted Protein Degradation via the Ubiquitin-proteasome System: Emerging Advances and Therapeutic Challenges of PROTACs.
  36. Polymeric alpha-1 antitrypsin perturbs Paneth cell proteostasis and exacerbates intestinal inflammation.
  37. Structural basis of diverse substrate recruitment by the HIV-1 Vpr-hijacked CRL4(VprBP) E3 ligase.
  38. Substrate Profiling of RNF216 Uncovers a Translation-Linked OTUD4 Regulatory Axis.
  39. Pulling and gripping: Proteasomal translocation pathway elements differentially contribute to substrate unfolding.
  40. EZH2 Inhibition Induces a Metabolic Stress Response Sensitizing Triple-Negative Breast Cancer to Glutaminase Targeting.
  41. Structural Mechanisms of DNAJC13 Dimeric Assembly and InsP6 binding in Recycling Endosome Regulation.
  42. Nuclear LC3 Interactome Profiling Identifies Clathrin Heavy Chain as a Mediator of Nuclear LC3 Translocation in Trabecular Meshwork Cells.
  43. A novel mechanism of ubiquitin-charging a bivalent E2:RING E3 complex.
  44. Protocol for ribosome-protected fragment isolation from rare leukemia cell populations.
  45. TROP2-Targeting Chimeras (TRTACs) for Tumor Cell-Selective Membrane Protein Degradation and Enhanced Drug Delivery.
  46. Ubiquitin-Dependent Protein Quality Control in Polyglutamine Diseases.
  47. BAG3 coordinates astrocytic proteostasis of Alzheimer's disease-linked proteins via proteasome, autophagy, and retromer interactions.
  48. Integrated stress response activation in combination PG3 and cisplatin-treated TP53 -mutated head and neck squamous carcinoma cells induces potent apoptotic response.
  49. D-allose reprogrammes the mitophagy-mtDNA-cGAS-STING-ADM2 axis to restore nucleus pulposus cell homeostasis in intervertebral disc degeneration.
  50. The barcoded binding assay: Multiplexed profiling of chaperone binding selectivity for hydrophobic substrates.
  51. Desulfinylation of the deubiquitinase USP7 by SRXN1 attenuates oxidative acute liver injury.
  52. CAPZ regulates endosomal-exosomal trafficking of plasma membrane PD-L1 independently of canonical autophagy.
  53. Proteostasis and ageing dissidence.
  54. Toxoplasma gondii GRA66 prevents premature egress driven by the host phospholipase RARRES3, independently of RNF213.
  55. Tandem RNA-binding domain architecture drives PKR activation through an intramolecular interface.
  56. Biphasic SMN condensates safeguard stress-induced cytoplasmic mislocalization of splicing RNPs.
  57. Variant-Specific Rewiring of Lysosomal Positioning by STARD9 Produces Opposite Cholesterol Trafficking Outcomes.
  58. Stepwise-coordinated polymeric nanoplatform enables traceless cytosolic protein delivery and autophagy blockade for enhanced tumor therapy.
  59. ULK1-dependent autophagy initiation promotes MHC-I degradation and immune evasion in HPV-positive head and neck cancer.
  1. bioRxiv. 2026 Sep 24. pii: 2026.09.23.753758. [Epub ahead of print]
    RNF185 destabilizes specific membrane proteins that span all ERAD branches.
    Haruka Chino, Isabella M Ruiz, Joshua H Corbo, Joao A Paulo, Sichen Shao.
      The endoplasmic reticulum (ER) is the primary site of eukaryotic membrane protein synthesis and quality control, which largely relies on ER-associated degradation (ERAD) to eliminate aberrant nascent proteins. In yeast, two ubiquitin ligases target proteins for ERAD based on aberrancies in substrate transmembrane (ERAD-M), lumenal (ERAD-L), or cytosolic (ERAD-C) domains. How an expanded repertoire of mammalian ERAD factors selects substrates across these classifications is unclear. Here, we show that the human ER-resident RNF185 ubiquitin ligase complex destabilizes a small but specific set of membrane proteins that span all three ERAD branches. Comparisons of three single-pass membrane proteins destabilized by RNF185 identify ERAD-M features in misoriented CHST10, ERAD-C features in unassembled SRPRB, and N-linked glycosylation-dependent ERAD-L features in ATP1B2. Our findings identify RNF185-destabilized membrane proteins with distinct aberrancies that collectively encompass all canonical ERAD substrate classifications and unexpectedly diverse quality control defects.
    DOI:  https://doi.org/10.64898/2026.09.23.753758
  2. Nat Struct Mol Biol. 2026 Sep 28.
    Cotranslational membrane protein biogenesis by an EMC-bound translocon.
    Madeline G Rollins, Josephine Tang, Yu Wan, Arunkumar Sundaram, Haoxi Wu, Qianru Li, Juliette Fedry, Ramanujan S Hegde, Zhe Ji, Robert J Keenan.
      Secretory and membrane protein biogenesis occurs at ribosome-bound translocons in the endoplasmic reticulum (ER) membrane. Translocons consist of the Sec61 protein-conducting channel and various maturation factors including chaperones, modification enzymes and membrane insertases, which assemble dynamically in response to the biogenesis needs of a nascent polypeptide. In human cells, nascent multipass membrane proteins utilize the ER membrane protein complex (EMC) and the multipass translocon (MPT), but how these factors are deployed during biogenesis is unknown. Here we combine selective ribosome profiling, cryo-electron tomography and site-specific photocrosslinking to reveal the substrate range, timing and mechanism of EMC recruitment. We show that multipass proteins initially engage the MPT for insertion and chaperoning, with EMC being recruited when substrates begin to exceed the cytosolic or intramembrane capacity of MPT. Substrate-engaged EMC is observed at the interface between the MPT's protected membrane cavity and the bulk ER membrane, where it is poised for insertion, chaperoning and assembly. These results show how EMC and MPT cooperate at the ribosome-bound translocon to facilitate multipass membrane protein biogenesis.
    DOI:  https://doi.org/10.1038/s41594-026-01889-2
  3. bioRxiv. 2026 Sep 10. pii: 2026.06.16.730951. [Epub ahead of print]
    Conformational diversity of collided ribosomes determines stress signaling.
    Jeffrey J Li, Timo Denk, Matthias Thoms, Allen R Buskirk, Roland Beckmann, Rachel Green.
      Ribosome collisions trigger pathways that clear stalled ribosomes, and when sufficiently abundant, the integrated stress response (ISR) through GCN2 and the ribotoxic stress response (RSR) through ZAK. The inhibitors anisomycin (ANS), emetine (EME), and didemnin B (DDB) are commonly used to induce collisions in studying these responses. Here, we demonstrate that these drugs induce distinct signatures: ANS and DDB potently activate ZAK whereas EME does not. We define transcriptional programs induced by these inhibitors, where collisions induce the RSR and general inhibition of translation leads to an RSR-independent response. Surprisingly, we find that collisions induced by EME, unlike ANS, are not cleared by ASCC3. The cryo-EM structure of human disomes stalled by EME reveals its mechanism of inhibition and a conformation distinct from ANS-stalled disomes. These differences in collision geometry explain the different outcomes in quality control and signaling activation, showing how ribosome stalling events can yield distinct cellular responses.
    DOI:  https://doi.org/10.64898/2026.06.16.730951
  4. Proc Natl Acad Sci U S A. 2026 Oct 06. 123(40): e2620353123
    Atg39-dependent nucleophagy and Atg40-dependent ER-phagy require the Apq12/Brr6/Brl1 complex.
    Dongmei Liu, Shuliang Chen, Hua Yuan, Susan Ferro-Novick, Peter Novick.
      Autophagic degradation of parts of the nucleus (nucleophagy) and endoplasmic reticulum (ER-phagy) utilizes the selective autophagy receptors, Atg39 and Atg40, that link their target structures to the autophagy machinery. Here, we show that a complex of three proteins, Apq12, Brr6, and Brl1, is needed for both Atg39-dependent nucleophagy and Atg40-dependent ER-phagy. Apq12, Brr6, and Brl1 each have two transmembrane domains flanking a luminal region that contains an amphipathic helix. The Apq12/Brr6/Brl1 complex has been implicated in membrane remodeling during nuclear pore formation. While autophagosomal flux is unaffected by the loss of Apq12, Brr6, or Brl1, vacuolar delivery of outer and inner nuclear membrane proteins (Hmg1, Nvj1, and Src1), pan-ER proteins (Sec63 and Per33), and an ER tubule junction protein (Lnp1) is blocked. In apq12Δ cells, Atg39 and Atg40 receptors concentrate in puncta and associate with the autophagic machinery; however, their delivery to the vacuole is blocked. Atg39 puncta extend into the cytoplasm but fail to detach from the nuclear envelope (NE), suggesting a defect in membrane scission. Coimmunoprecipitation and crosslinking analysis indicate that Atg39 and Atg40 interact with the Apq12/Brr6/Brl1 complex. We propose that the membrane remodeling activity of the Apq12/Brr6/Brl1 complex is needed to release fragments of the NE and ER so they can be sequestered within autophagosomes for degradation.
    Keywords:  Apq12; Brl1; Brr6; ER-phagy; nucleophagy
    DOI:  https://doi.org/10.1073/pnas.2620353123
  5. EMBO J. 2026 Sep 28.
    Excess Met1-linked ubiquitination leads to solid aggregate formation.
    Stephanie Kaypee, Minori Miyasaka, Takuto Nakajima, Hiromi Nishimura, Jun-Ichi Sakamaki, Masaaki Komatsu, Fumiyo Ikeda.
      The ubiquitin ligase HOIL-1 regulates the formation of Met1-linked (linear) ubiquitin chains through its coordination with the E3 ligase HOIP within the Linear Ubiquitin Chain Assembly Complex (LUBAC). While HOIP-dependent Met1-linked ubiquitination is well established in inflammation and immunity, the physiological importance of its quantitative control remains unclear. Here, we show that cells expressing catalytically inactive HOIL-1 accumulate increased α-synuclein, tau, and amyloid-β aggregates. This is associated with defective late-stage autophagic flux, characterized by impaired delivery of p62-positive aggregates to lysosomes. In parallel, p62 bodies undergo a biophysical transition from dynamic, liquid-like condensates to rigid, solid-like structures. Elevation of Met1-linked ubiquitin chains, either through HOIL-1 inactivation or depletion of the Met1-specific deubiquitinase OTULIN, phenocopies these defects. Together, our findings identify HOIL-1 as a key regulator of aggregate clearance and proteostasis through quantitative control of Met1-linked ubiquitination.
    DOI:  https://doi.org/10.1038/s44318-026-00909-7
  6. Autophagy. 2026 Sep 28.
    RABC1 regulates ER-phagy to maintain er homeostasis in plants.
    Shuai Hu, Xiaohui Ma, Zhengjia Wang, Jinbo Shen.
      Endoplasmic reticulum (ER)-phagy is an important ER quality-control pathway that selectively removes misfolded or unfolded proteins and damaged ER membranes through autophagic degradation in the vacuole. Recent studies have identified small guanosine triphosphatases (GTPases) as important regulators of the autophagy pathway. However, how they couple stress signals to ER-phagy remains poorly understood in plants. Our recent work demonstrated that RABC1 regulates ER-phagy under dithiothreitol (DTT)-, tunicamycin (TM)-, and heat-induced ER stress through interaction with the exocyst complex component SEC5A. Upon ER stress, green fluorescent protein (GFP)-RABC1 was recruited to the ATG8e-positive autophagosomes from the ER and Golgi in root cells, whereas autophagic flux and calnexin 1 (CNX1)-mRFP-tagged ER membranes turnover were impaired in the rabc1 mutant. Together, our findings support a model in which RABC1 acts as a molecular switch that interacts with distinct effectors to regulate autophagy under different stress conditions.
    Keywords:  Autophagy; Er-phagy; RABC1; SEC5A; er stress
    DOI:  https://doi.org/10.1080/15548627.2026.2739788
  7. Proc Natl Acad Sci U S A. 2026 Oct 06. 123(40): e2616933123
    Targeted extracellular degradation of LRP8 promotes ferroptosis in cancer cells.
    Fangzhu Zhao, Alex Inague, Trenton M Peters-Clarke, Yifei Chen, Snehal D Ganjave, Yun Zhang, Kun Miao, Zi Yao, Yan Wu, Madison K C Seto, Kevin K Leung, James A Olzmann, James A Wells.
      Tumor reliance on antioxidant defenses creates a vulnerability to ferroptosis, yet strategies to therapeutically disable these systems remain limited. Here, we identify targeted degradation of the selenium uptake receptor lipoprotein receptor-related protein 8 (LRP8) as an effective approach to decrease the abundance of the ferroptosis-protective enzyme glutathione peroxidase 4 (GPX4). Using bispecific cytokine receptor-targeting chimeras (KineTACs) that couple LRP8 to cytokine receptor internalization pathways, we selectively direct LRP8 to the lysosome for degradation. LRP8 degradation reduces the abundance of several selenoproteins, including GPX4, lowering the cellular threshold for lipid peroxidation and sensitizing cancer cells to ferroptosis. These findings establish receptor-mediated selenium uptake as a critical, targetable node in ferroptosis resistance and demonstrate that extracellular protein degradation can be leveraged to reprogram intracellular translational dependencies in cancer cells. More broadly, this work provides a framework for exploiting nutrient acquisition pathways to overcome therapy resistance.
    Keywords:  LRP8; antibody engineering; ferroptosis; targeted protein degradation
    DOI:  https://doi.org/10.1073/pnas.2616933123
  8. Nat Commun. 2026 Aug 31. pii: 10351. [Epub ahead of print]17(1):
    Listeria promotes systemic infection by targeting the host deubiquitinase CYLD.
    Dustin A Ammendolia, Cara J Ellison, Yuelin Zheng, Etienne Coyaud, Estelle M N Laurent, Bing-Ru Yan, Bri Manning, Anna K Waldmann, Joel M J Tan, Scott Frendo-Cumbo, Carina Lyons, Ji-Young Youn, Brian Raught, Keith Ireton, Darren E Higgins, Paul R Elliott, John H Brumell.
      Listeria monocytogenes (Lm) is an intracellular pathogen that can cause life-threatening systemic infections. Internalin C (InlC) is a secreted factor required for full Lm virulence in systemic models of infection by mechanisms that remain unclear. Here, we show that InlC binds to CYLD, a host deubiquitinase and regulator of innate immunity. Structurally, we reveal the LRR domain of InlC binds the CAP-Gly2 domain of CYLD, independently of InlC's binding site for other host target proteins. Lm strains harboring amino acid substitutions in InlC that selectively disrupt binding to CYLD were examined. We demonstrate that after Lm accesses the host cytosol, InlC recruits CYLD to ubiquitin-positive bacteria. Recruitment of CYLD was dependent on the host E3 ligase RNF213, a major initiator of ubiquitin-mediated defenses. Furthermore, InlC-CYLD binding contributed to Lm virulence in mice. Together, these findings reveal how a secreted bacterial factor promotes the recruitment of a host deubiquitinase to cytosolic bacteria in response to ubiquitin-mediated defenses.
    DOI:  https://doi.org/10.1038/s41467-026-77063-5
  9. J Cell Sci. 2026 Oct 02. pii: jcs.265001. [Epub ahead of print]
    Distinct roles for TANGO1S domains in maintaining ER-Golgi architecture.
    Elizabeth A Lawrence, Lorna Hodgson, Judith Mantell, M Esther Prada-Sanchez, Chrissy L Hammond, David J Stephens, Nicola L Stevenson.
      The endoplasmic reticulum (ER)-Golgi interface is a dynamic trafficking hub maintained in part by TANGO1, a scaffolding protein that coordinates proteins and membranes at ER exit sites (ERES). TANGO1 has two isoforms: TANGO1L, which has a lumenal SH3 domain, and TANGO1S, which lacks this domain but retains the transmembrane and cytoplasmic coiled-coil (CC), TEER, and PRD domains common to both forms. We showed previously that loss of both isoforms disrupts ER-Golgi organization more severely than TANGO1L loss alone, indicating TANGO1S is functional and can compensate. Here we dissect the role of each TANGO1 cytoplasmic domain in maintaining secretory pathway organisation by expressing TANGO1S domain-deletion mutants in TANGO1L-/S- knockout cells. We show that TANGO1 loss causes cis-Golgi vesiculation that cannot be rescued by TANGO1S. TANGO1S re-expression also failed to rescue COPII and Sec16 recruitment in this cell line. Meanwhile, the TEER domain is essential for the organisation of the ER, and the TEER, CC2 and PRD domain are required for a defined ERGIC. This study represents an advance towards a domain-level resolution of TANGO1S function.
    Keywords:  ERES; ERGIC; Endoplasmic reticulum; Golgi; TANGO1
    DOI:  https://doi.org/10.1242/jcs.265001
  10. Mol Cell. 2026 Oct 01. pii: S1097-2765(26)00623-4. [Epub ahead of print]86(19): 4018-4031.e4
    ZNFX1, an immunoregulatory RNA helicase and E3 ubiquitin ligase, assembles into pleiomorphic polymers.
    Katerina Naydenova, Thomas Mund, Matthew C J Yip, Ceara M Harper, Kendra E Leigh, Jacqueline Hankinson, Keith B Boyle, Alexander Heatley, Elsje G Otten, Valeria Lulla, Yorgo Modis, Felix Randow.
      NFX1-type zinc-finger-containing 1 (ZNFX1) is an SF1-family RNA helicase essential for innate immunity. Patients with ZNFX1 mutations experience recurrent infections, yet the underlying mechanism remains unclear. We determined cryo-electron microscopy (cryo-EM) structures of RNA-bound and RNA-free ZNFX1 and revealed auto-inhibition of the helicase-associated ATPase through a regulatory insertion occluding the RNA-binding groove. ZNFX1 also functions as a bi-catalytic E3 ubiquitin ligase, containing an RZ finger homologous to RNF213 and a previously unidentified Miz-like domain, which cooperate in catalyzing ubiquitylation. Ubiquitin chains enhance E3 activity further. Patient mutations demonstrate that the helicase/ATPase, E3 ligase, and the rigid zinc-finger stalk connecting them are required for function. ZNFX1 can assemble into structured, pleiomorphic polymers fostering trans-auto-ubiquitylation; understanding this reveals a mechanism that controls ZNFX1 stability and may facilitate RNA sequestration in stress granules and antiviral activity. These findings establish ZNFX1 as a multifunctional enzyme in immunity that couples RNA sensing to ubiquitin signaling and assembles into higher-order structures for signal amplification.
    Keywords:  E3 ubiquitin ligases; RNA helicases; innate immunity
    DOI:  https://doi.org/10.1016/j.molcel.2026.09.002
  11. bioRxiv. 2026 Sep 13. pii: 2026.09.09.750423. [Epub ahead of print]
    Substrate Binding Relieves Autoinhibition of the Mammalian GCN2 Kinase.
    Marco A Catipovic, Rachel Green.
      Cells respond to amino acid starvation and translational stress through the integrated stress response (ISR) kinase GCN2. Current models maintain that GCN2 senses these stresses either by binding deacylated tRNAs that accumulate during amino acid starvation or by recognizing ribosome collisions that arise from perturbation of translation elongation. As tRNA charging is inherently coupled to translation elongation, assessing the relative contributions of these two pathways has proven difficult. To fully separate the roles of these potential agonists, we reconstituted the regulation of GCN2 in vitro using recombinant proteins. GCN2 has low basal activity, which is stimulated by the addition of ribosomes or the ribosomal P-stalk. By contrast, deacylated tRNAs fail to activate GCN2 both in vitro and in vivo and instead counteract activation by the ribosome and inhibit GCN2 auto- and substrate phosphorylation. This inhibition of autophosphorylation requires the presence of GCN2's substrate, eIF2α, suggesting that the substrate plays a role in kinase regulation. Indeed, eIF2α binding to a newly defined autoinhibitory loop in the GCN2 kinase domain relieves its repression and stimulates autophosphorylation of the kinase's activation loop. Together, these data redefine the role of tRNAs in the regulation of GCN2 and reveal a mechanism novel to kinases in general by which substrate binding regulates activation loop phosphorylation.
    DOI:  https://doi.org/10.64898/2026.09.09.750423
  12. Nat Commun. 2026 Aug 26. pii: 10238. [Epub ahead of print]17(1):
    A redox switch on CDC48 regulates peroxisomal function via controlling peroxisome-associated protein degradation.
    Jialong Li, Yandong Wang, Ronghui Pan, Rongcheng Lin.
      Peroxisomes are single-membrane-bound organelles essential for diverse metabolic reactions and cellular redox homeostasis, yet the contribution of ubiquitin-proteasome system to peroxisomal biology remains unclear. Here, we demonstrate that the AAA-ATPase complex comprising Cell Division Cycle48 (CDC48), Nuclear Protein Localization4 (NPL4) and Ubiquitin Fusion Degradation1 (UFD1) is indispensable for peroxisomal biogenesis and physiological function in Arabidopsis. We identify the peroxisomal membrane peroxin PEX22 as a direct substrate of the CDC48 complex and show that this complex promotes ubiquitin-dependent PEX22 turnover. Genetic analyses place CDC48 complex upstream of PEX22 in controlling peroxisomal biogenesis and activity. Moreover, H₂O₂‑triggered Cys271 oxidation represses CDC48 ATPase activity, stabilizing PEX22 via slowed degradation; nucleoredoxin NRX1 reduces oxidized CDC48 to recover its function. Consistently, transgenic plants harboring the redox-insensitive CDC48-C271S variant display accelerated PEX22 turnover and enhanced susceptibility to oxidative stress. Collectively, our findings establish the CDC48 complex as a putative H₂O₂ sensor that governs ubiquitin-mediated peroxisome-associated protein degradation (PexAD), enabling fine-tuning of peroxisomal performance in plant development and upon environmental stress.
    DOI:  https://doi.org/10.1038/s41467-026-77146-3
  13. Curr Biol. 2026 Oct 02. pii: S0960-9822(26)01198-X. [Epub ahead of print]
    Mitochondria-ER-lipid droplet crosstalk regulates metabolic adaptation to shear stress via VMP1.
    Maria Laura Rojas, Maribel Lara Corona, Carla Alemany, Juliane Da Graça, Nicolas Goudin, Ivan Nemazanyy, Ida Chiara Guerrera, Aurore Claude-Taupin, Etienne Morel.
      Kidney tubular epithelial cells adapt to physiological urinary flow through rapid metabolic remodeling,1,2 but the mechanisms coordinating this response remain poorly understood. Shear stress promotes lipid catabolism and mitochondrial activity in these cells,3,4 but how changes in mitochondrial dynamics contribute to this metabolic adaptation remains poorly understood.5,6,7,8,9 Here, we show that physiological shear stress rapidly remodels mitochondrial morphology in kidney epithelial cells in vitro and in the zebrafish pronephros, characterized by the emergence of a distinct pool of donut-shaped mitochondria. This remodeling is accompanied by a transient stabilization of mitochondria-endoplasmic reticulum (ER) contact sites (MERCs), occurring independently of any increase in overall ER volume. Using split-TurboID proximity labeling and mass spectrometry, we detected subtle changes in the molecular environment of MERCs during shear stress, including increased proximity of proteins implicated in lipid transfer and membrane contact-site biology. We further show that shear stress promotes the formation of ER-lipid droplet (LD)-mitochondria contact sites and facilitates the local transfer of fatty acids from LDs to mitochondria. This lipid transfer requires vacuolar membrane protein 1 (VMP1), a component of membrane contact sites, whose depletion perturbs LDs and compromises metabolic adaptation to shear stress. Together, our findings identify ER-LD-mitochondria contact sites as dynamic platforms that coordinate lipid transfer and mitochondrial remodeling during the early adaptation of kidney epithelial cells to physiological shear stress, highlighting membrane contact sites as important components of the cellular response to mechanical forces.
    Keywords:  contact sites; endoplasmic reticulum; kidney epithelial cells; lipid droplets; metabolic adaptation; mitochondria; shear stress; zebrafish
    DOI:  https://doi.org/10.1016/j.cub.2026.09.021
  14. bioRxiv. 2026 Sep 22. pii: 2026.09.18.752746. [Epub ahead of print]
    Single residues within the YTHDF intrinsically disordered region control m 6 A-dependent mRNA decay.
    Benedetta Ricci, Danika Sommer, Daymieri Narvaez, Ruqi Zhang, Jacob Geri, Sara Zaccara.
      Intrinsically disordered regions (IDRs) mediate protein interactions, condensate partitioning, and regulatory control in RNA-binding proteins, yet the residue-level logic underlying IDR function has remained difficult to define using conventional fragment- and truncation-based approaches. Here, we apply cytosine and adenine base-editor screens to map residue-level regulation within the intrinsically disordered region of YTHDF2, a cytoplasmic reader that couples m 6 A to mRNA decay. By tiling base edits across YTHDF2 in a YTHDF1/YTHDF3-null background, we identify IDR residues that regulate YTHDF2-dependent cellular fitness. Interestingly, we identified residues that limit YTHDF2 decay activity. Their mutation disrupts endogenous protein-protein interactions and generates hyperactive YTHDF2 variants that promote m 6 A-dependent mRNA decay and alter recruitment to cytoplasmic RNA granules, without affecting intrinsic m 6 A binding or protein stability. These findings define the residue-level regulatory logic of the YTHDF2 IDR and establish base-editor screening as a strategy to uncover functional mechanisms encoded within IDR regions.
    DOI:  https://doi.org/10.64898/2026.09.18.752746
  15. Sci Adv. 2026 Oct 02. 12(40): eaeg0399
    NAT10 governs development through the Xrp1-mediated stress response rather than its RNA acetylation activity.
    Lin Wang, Zhao Xue, Shuyang Gao, Yanhua Wang, Houguang Lu, Bo Peng, Tianyi Fan, Ying Cao, Jian-Quan Ni, Guoliang Lu, Jinzhong Lin, Dong Yan.
      N4-acetylcytidine (ac4C) is installed by N-acetyltransferase 10 (NAT10) and represents the only known acetylation mark on messenger RNA. Since NAT10 also acetylates transfer RNA, ribosomal RNA (rRNA), and proteins, its in vivo molecular mechanisms remain elusive. Here, we demonstrate that knockdown of Drosophila NAT10 induces an eye-to-antenna transformation, c-Jun amino-terminal kinase activation, and cell apoptosis. NAT10 facilitates ac4C modification in rRNA, and its loss impairs rRNA processing and ribosomal assembly. Depletion of NAT10 activates the integrated stress response, ultimately leading to reduced global protein synthesis. Crucially, Xrp1 plays a key role in the stress response and its ablation rescues most NAT10 loss-of-function defects and transcriptomic alterations. We also performed acetylated RNA immunoprecipitation and sequencing (acRIP-seq) on control and NAT10 knockdown flies. Furthermore, a catalytically deficient form of NAT10, which is unable to mediate ac4C acetylation, completely rescues the lethality of NAT10 mutants. Collectively, these findings establish that NAT10's primary developmental function stems from its roles in ribosome biogenesis and Xrp1 activation, which are independent of its RNA acetylation activity.
    DOI:  https://doi.org/10.1126/sciadv.aeg0399
  16. EMBO Rep. 2026 Sep 30.
    FBXL21 regulates diurnal proteostasis in skeletal muscle by targeting DNAJB6 and client proteins.
    Ji Ye Lim, Jaebok Wi, Marvin Wirianto, Chorong Han, Sun Young Kim, Jane Nguyen, Sehyun Jung, Kristin Eckel-Mahan, Sung Yun Jung, Karyn A Esser, Zheng Chen, Seung-Hee Yoo.
      Circadian regulation of proteostasis, a key determinant of muscle health, remains poorly understood. Here, we identify DNAJB6, an Hsp40 (DnaJ) co-chaperone, as a substrate of the circadian E3 ligase FBXL21. FBXL21 mediates the ubiquitination-dependent proteasomal degradation of both DNAJB6 and its client proteins, including Desmin. In contrast, myopathy-causing mutations of DNAJB6 confer resistance to FBXL21-directed degradation. Fbxl21 KO C2C12 cells display aberrant Desmin accumulation, and show aggravated cytoplasmic accumulation of TDP-43, another DNAJB6 client protein, in response to heat shock. Under timed exercise as a physiological stressor, WT mice display robust diurnal rhythms in the levels of stress granule markers (G3BP1 and FUS) and TDP-43 as a function of exercise timing. In contrast, the Fbxl21 hypomorph Psttm mutant mice show elevated expression of these proteins without exercise, which is exacerbated under exercise-induced stress conditions. Importantly, these abnormalities are rescued by skeletal muscle-specific FBXL21 expression. Our study elucidates a novel diurnal regulatory mechanism of skeletal muscle proteostasis via FBXL21 as a chaperone-linked E3 ligase, highlighting the FBXL21-DNAJB6 axis as a potential therapeutic target for myopathies.
    DOI:  https://doi.org/10.1038/s44319-026-00922-1
  17. Nat Commun. 2026 08 31. pii: 10374. [Epub ahead of print]17(1):
    A lipid transfer-dependent feedback loop activates ATG9A compartments in autophagy initiation.
    Elisabeth Holzer, Justyna Sawa-Makarska, Daniel Bernklau, Julia Romanov, Martina Schuschnig, Sascha Martens.
      Autophagy degrades cellular material by sequestering it within autophagosomes, which form de novo from precursors called phagophores. Phagophore assembly and expansion require ATG9A-positive seed compartments, the lipid transfer protein ATG2A, and the class III phosphatidylinositol 3-phosphate kinase complex I (PI3KC3-C1). PI3KC3-C1 synthesizes phosphatidylinositol 3-phosphate (PI3P), a key lipid that drives downstream processes for phagophore expansion, including ATG8 lipidation. We find that ATG9A compartments contain only traces of phosphatidylinositol (PI), likely insufficient for efficient PI3P production or recruitment of PI3P-binding effectors. Nevertheless, ATG2A is recruited to these compartments and mediates lipid transfer, including PI, into them. Remarkably, even without detectable PI3P, ATG9A compartments are direct substrates for ATG8 lipidation, and ATG8 proteins themselves enhance ATG2A-mediated lipid transfer. In cells, ATG2A is essential for the appearance of PI3P on ATG9A compartments. Our findings support a model in which a lipid transfer-driven feedback loop activates ATG9A compartments for phagophore expansion.
    DOI:  https://doi.org/10.1038/s41467-026-77368-5
  18. Cancer J. 2026 Sep-Oct 01;32(5):pii: e00852. [Epub ahead of print]32(5):
    Proteostasis as a Therapeutic Backbone: Proteasome Inhibitors and Emerging Regimens.
    Yan Song, Poy Theprungsirikul, Annamaria Gulla, Kenneth C Anderson.
      Multiple myeloma cells are uniquely dependent on proteostasis due to their exceptionally high immunoglobulin synthesis and protein turnover, rendering them hypersensitive to disruptions in protein degradation. Proteasome inhibitors targeting the 20 S catalytic core exploit this vulnerability and remain a therapeutic cornerstone in newly diagnosed and relapsed/refractory settings, anchoring both triplet and quadruplet regimens that incorporate anti-CD38 monoclonal antibodies, immunomodulatory agents, antibody-drug conjugates, and nuclear export inhibitors. Beyond the 20 S core, emerging targets, including the 19 S regulatory particle, deubiquitinating enzymes, and the E1-E2-E3 ubiquitination cascade, offer opportunities to intensify proteotoxic stress and overcome resistance. Targeted protein degradation platforms, including CELMoDs, proteolysis targeting chimeras, molecular glues, and proteasome cap-recruiting chimeras, further expand the therapeutic repertoire. Compensatory stress responses involving autophagy, the unfolded protein response, and the bone marrow microenvironment modulate therapeutic vulnerability. Collectively, these advances position proteostasis not as a single druggable target but as an integrated therapeutic network amenable to rational, biomarker-informed combination strategies in multiple myeloma.
    Keywords:  Proteasome inhibitor; multiple myeloma; proteostasis; ubiquitin-proteasome system
    DOI:  https://doi.org/10.1097/PPO.0000000000000852
  19. Mol Cell. 2026 Oct 01. pii: S1097-2765(26)00621-0. [Epub ahead of print]86(19): 3897-3914.e18
    Ubiquitin chain editing promotes p97/VCP-dependent removal of RPA from DNA.
    Matthew Drake, Xianzhen Zhou, Alexandra McStea, Benjamin Foster, Kay Yi Chong, Martin Attwood, Leslye Roca Burgos, Laura Bennett, Sneha Anand, Merve Kaplan, Jessica J Lu, Vaishnavi Ravikumar, Marjorie Fournier, Elena Navarro-Guerrero, Michael Ranes, Weston B Struwe, Christopher Staples, Daniel Ebner, Ian Gibbs-Seymour.
      DNA repair requires dynamic control of proteins on single-stranded DNA (ssDNA), yet how ubiquitin signaling regulates ssDNA-bound factors remains poorly understood. Here, we identify in human cells a ubiquitin chain-editing mechanism that promotes extraction of replication protein A (RPA) from ssDNA. We show that RPA stimulates the deubiquitinase ZUP1 and enhances its activity toward K63 linkages within ubiquitin chains. In response to DNA damage, RPA is modified with branched K48-K63 ubiquitin chains. ZUP1 selectively removes K63 linkages from these chains, remodeling the ubiquitin signal on RPA. We show that ZUP1-mediated editing of branched ubiquitin chains promotes p97/VCP-dependent removal of ubiquitinated RPA from ssDNA. Loss of ZUP1 causes accumulation of branched ubiquitin chains on RPA and pathological RPA trapping on ssDNA, resulting in elevated ssDNA signaling and genome instability. Together, our findings identify ubiquitin chain editing as a mechanism that controls RPA dynamics on ssDNA and enables p97/VCP-mediated protein extraction during DNA repair.
    Keywords:  DNA repair; DUBs; RFWD3; RPA; VCP; ZUP1; deubiquitinases; genome stability; p97; ssDNA; ubiquitin
    DOI:  https://doi.org/10.1016/j.molcel.2026.09.004
  20. bioRxiv. 2026 Sep 24. pii: 2026.09.23.753862. [Epub ahead of print]
    Collided ribosomes are rescued by the endonuclease Rae1 and trans- translation.
    Daniel D Tetreault, Katrina Callan, Cassidy R Prince, Heather A Feaga.
      Ribosome stalling is a major problem in all domains of life. When a ribosome stalls, trailing ribosomes may catch up to and collide with the stalled ribosome, depleting protein synthesis capacity. Here, we describe a novel pathway used by Gram-positive bacteria to rescue ribosome collisions. We used the ATPase defective ABCF protein YdiF(EQ 2 ) to induce ribosome stalling and collisions in Bacillus subtilis . Ribosome profiling (Ribo-seq) of YdiF(EQ 2 )-expressing cells revealed that collided ribosomes are enriched for tmRNA, a functional RNA involved in trans -translation. We confirmed that tmRNA tagging activity is globally increased upon expression of any ATPase defective ABCF as well as in cells treated with the collision-inducing antibiotic erythromycin, suggesting this is a generalizable mechanism to rescue stalled and collided ribosomes. The global increase in tmRNA tagging that occurred in response to both erythromycin and YdiF(EQ2) induced collisions was dependent on the Rae1 endonuclease. Loss of trans -translation in cells experiencing widespread ribosome collisions leads to a severe fitness defect, consistent with the importance of this pathway in rescuing ribosomes stalled on truncated mRNAs that result from ribosome collisions. Altogether, our work supports a model in which Rae1 cleaves mRNA on collided ribosomes, thereby generating a truncated mRNA substrate for trans -translation and leading to rescue and recycling of the collided ribosomes. We term this mechanism C ollision- A ssociated R ae1-induced trans- Translation (CART). CART broadens the repertoire of tools that bacteria use to manage ribosome collisions.
    Significance: Prolonged ribosome stalling leads to ribosome collisions, which are rescued by specialized factors. While ribosome collisions have been extensively studied in eukaryotes, our understanding of collision rescue in bacteria is in its infancy. Data described here are the first to directly show that tmRNA mediates rescue of collided ribosomes in a Gram-positive bacterium, Bacillus subtilis . This pathway is analogous to what occurs in model organisms such as Escherichia coli and Saccharomyces cerevisiae , but relies on an unrelated nuclease, Rae1. Since B. subtilis and E. coli are on opposite ends of the bacterial phylogenetic tree, and since Rae1 is broadly conserved in bacteria, our findings suggest that mRNA cleavage arose convergently in distantly related bacteria as a strategy to rescue ribosome collisions. Convergent evolution of these pathways highlights the importance of rescuing collided ribosomes in all organisms. Moreover, insights into ribosome rescue in E. coli and B. subtilis can guide studies of ribosome rescue in bacteria with intermediary phylogenetic relatedness to these two model organisms.
    DOI:  https://doi.org/10.64898/2026.09.23.753862
  21. EMBO J. 2026 Sep 28.
    A functional screen for bacterial ubiquitin regulation identifies an unusual Pseudomonas aeruginosa E3 ligase.
    Cameron G Roberts, Supender Kaur, Shun-Je Bhark, Aaron J Ogden, Michael E Divine, Gus D Warren, Donghoon Kang, Natalia V Kirienko, Paul P Geurink, Monique P C Mulder, Ernesto S Nakayasu, Jason E McDermott, Joshua N Adkins, Alejandro Aballay, Jonathan N Pruneda.
      Bacterial pathogens have adapted secreted effector proteins that hijack host ubiquitination to gain control over host responses. These effectors can exhibit, for example, E3 ligase or deubiquitinase activities, often without any homology to eukaryotic regulators. Such convergence in function poses a challenge to the discovery of additional ubiquitin-targeted virulence factors. Pseudomonas aeruginosa, for example, is a pervasive opportunistic pathogen with numerous mechanisms of virulence, yet no described E3 ligases or deubiquitinases. We have developed a workflow to functionally screen natively secreted bacterial effectors for ubiquitin regulatory activities. After benchmarking this approach on Salmonella Typhimurium, Enteropathogenic Escherichia coli, and Shigella flexneri, we identified a cryptic E3 ligase secreted by P. aeruginosa, which we have termed Pseudomonas Ub ligase 1 (PUL-1). PUL-1 resembles none of the other E3 ligases previously established in or outside of the eukaryotic system. Importantly, in an animal model of P. aeruginosa infection, PUL-1 ligase activity plays an important role in regulating virulence. Thus, functional identification of ubiquitin-targeted effectors can expand our appreciation of ubiquitin regulation during bacterial pathogenesis.
    DOI:  https://doi.org/10.1038/s44318-026-00917-7
  22. Adv Sci (Weinh). 2026 Sep 27. e77819
    A Versatile DNA-Encoded Library Platform for the Discovery of Highly Cooperative Chemical Inducers of Proximity.
    Bingqi Tong, Sunny A Tang, Yifan Deng, Zhihan Nan, Alison X Gao, Gregory A Michaud, Simone Bonazzi, Frédéric Berst, Frédéric J Zécri, Shuang Liu, William J Gibson, Stuart L Schreiber.
      Chemical-induced proximity, particularly via molecular glues, represents a rapidly advancing therapeutic paradigm. Expanding the scope of proximity-based therapeutics requires generalizable discovery platforms that can be easily tailored to diverse presenter proteins. Here, we report a versatile DNA-encoded library (DEL) strategy designed to identify highly cooperative chemical inducers of proximity (CIPs) for multiple presenter proteins. By leveraging a novel precursor library constructed via on-DNA strained allene cycloadditions, we generated three distinct CIP-DELs biased toward Cereblon (CRBN), VHL, and FKBP12. Screening the CRBN-focused library with BRD9 identified B67b, a novel compound that mediates ternary complex formation between CRBN and BRD9 with nanomolar potency and strong molecular glue-like cooperativity (α >300). B67b engages BRD9 in a CRBN-dependent manner, a characteristic of classical molecular glues, and its high cooperativity is driven by the synergistic contribution of all its structural and stereochemical components. This work establishes a scalable, generalizable framework for repurposing generic DELs into presenter-specific screening tools, offering a powerful approach to accelerate the discovery of CIP therapeutics for a broad range of targets.
    Keywords:  DNA‐encoded library; drug discovery; induced proximity; molecular glue; ternary complex cooperativity
    DOI:  https://doi.org/10.1002/advs.77819
  23. Nature. 2026 Sep 30.
    A TBK1/ULK1 signalling axis couples lysosomal stress to TFEB activation.
    Alessandra Esposito, Ibrahim Bayramoglu, Chiara Varriale, Maria Pia Cavaliere, Maria Elsa Brunetti, Rossella Agostinis, Rachel M Jansen, Dan A Tudorica, Yongjia Duan, Martina Spacci, Alessia Calcagnì, Carolin D Strobl, Verena Passerini, Fabrizio Andreone, Jlenia Monfregola, Julia Richter, Francesca Sacco, Wolfram Klapper, Oliver Weigert, Andrea Ballabio, Roberto Zoncu, James H Hurley, Joern Dengjel, Gennaro Napolitano.
      Lysosomal adaptation to environmental changes is critical for cellular and metabolic homeostasis and requires coordination by the mTORC1 kinase, which conveys nutritional and stress signals into distinct, substrate-specific outputs1,2. The FLCN-FNIP complex (FLCN:FNIP) serves as a crucial regulator of lysosomal function by selectively controlling the ability of mTORC1 to inhibit transcription factor EB (TFEB), a master regulator of catabolic programs and a known oncogene3. Yet how FLCN:FNIP activity is regulated has remained unclear. Here we identify a nutrient-independent lysosomal signalling pathway that regulates FLCN through v-ATPase-driven recruitment of TBK1 or ULK1 (TBK1/ULK1) to lysosomes, via the TAX1BP1 adaptor. This enables TBK1/ULK1-mediated FNIP1 phosphorylation at S296, resulting in inhibition of FLCN and nuclear translocation of TFEB. Recurrent ATP6V1B2 v-ATPase mutations, found in patients with follicular lymphoma, constitutively activate this pathway, leading to hyperactivation of TFEB and follicular lymphoma proliferation. Our work uncovers a lysosomal signalling pathway that is critical for lysosomal adaptation and tumorigenesis.
    DOI:  https://doi.org/10.1038/s41586-026-11093-3
  24. bioRxiv. 2026 Sep 25. pii: 2026.09.23.753918. [Epub ahead of print]
    Pathogenic mutations in ATAD3A cause dysregulation of RagC/D-TFEB axis and disrupt lysosomal homeostasis.
    Mathew B McDougal, Abigail Sandoval, Mike Kinter, Antrix Jain, Sukyeong Lee, Sung Yun Jung, Wan Hee Yoon.
      We previously discovered that a de novo variant p.R528W in ATAD3A , encoding a mitochondrial membrane-anchored protein, causes a human neurological syndrome. While ATAD3A mutations induce aberrant lysosomal expansion accompanied by undigested material in the lysosomes, how mutant ATAD3A disrupts lysosomal homeostasis and whether this contributes to neurodevelopmental defects remain unknown. Here we show that pathogenic ATAD3A p.R528W expression disrupts the mTORC1-TFEB axis as revealed by dysregulation of mTORC1 substrate phosphorylation, TFEB nuclear localization, and CLEAR gene activation associated with lysosomal biogenesis. ATAD3A binds to lysosome-localized Rag C/D GTPases, which constitute a platform for TFEB recruitment, with pathogenic variants increasing this association and thereby decreasing lysosomal localization of Rag GTPases. Importantly, overexpression of RagC or RagD restores TFEB phosphorylation in human cells expressing p.R528W, and RagC- D overexpression or TFEB/Mitf knockdown rescues lysosomal expansion and neurodevelopmental defects in Drosophila . These data indicate that disrupted Rag GTPase recruitment to lysosomes and subsequent aberrant TFEB/Mitf activation contribute to neurodevelopmental and lysosomal phenotypes caused by pathogenic mutations in ATAD3A . Our work reveals a novel role for the mitochondrial resident protein ATAD3A in modulating lysosomal homeostasis through regulation of the mTORC1-TFEB axis, providing a mechanistic link between impaired mitochondrial and lysosomal homeostasis in a neurodevelopmental disorder.
    DOI:  https://doi.org/10.64898/2026.09.23.753918
  25. Nature. 2026 Sep 30.
    GPCR antagonism via rewiring of receptor trafficking and degradation.
    Kaitlin Rhee, Lawrence Shue, Akimasa Adachi, Pengwei Sun, James Osei-Owusu, Dingjingyu Zhou, Aoxing Cheng, Yi Ran Xu, Qingyue Li, Apoorva Baluapuri, Edward P Harvey, Karen Adelman, Meredith A Skiba, Bianxiao Cui, Jun R Huh, Andrew C Kruse, Xin Zhou.
      G-protein-coupled receptors (GPCRs) represent one of the most important yet incompletely addressed classes of therapeutic targets1. Here we report a strategy for functional GPCR antagonism through bispecific antibody-mediated endocytosis and lysosomal degradation. GPCR-TfR1 targeting chimeras (GTACs) achieve potent and selective downregulation of multiple GPCRs, including BILF1, RXFP1 and CCR6-viral, cancer and immune targets that have been difficult to drug2-4. GTACs lead to complete inhibition of receptor signalling, including constitutive signalling, with more than one to two orders of magnitude greater potency than conventional antibody antagonists. Using protein engineering and multicolour live-cell imaging, we establish a context-dependent degrader design and explain the cellular mechanisms, with broad relevance for degrader technology. The GTAC platform establishes induced endocytosis and rewiring protein trafficking as a model for therapeutic GPCR modulation.
    DOI:  https://doi.org/10.1038/s41586-026-11088-0
  26. Philos Trans R Soc Lond B Biol Sci. 2026 Oct 01. pii: 20250270. [Epub ahead of print]381(1960):
    More than the 'master regulator of cytosolic proteostasis': new roles for HSF1 in promoting tissue health.
    John Labbadia.
      Maintaining protein homeostasis (proteostasis) is crucial for long-term tissue health. This requires the action of stress response pathways and protein quality control mechanisms that act within or across different sub-cellular compartments to preserve proteome integrity. Within the cytosol/nucleus, the loss of proteostasis induces a transcriptional programme known as the heat-shock response (HSR) through activation of heat-shock factor 1 (HSF1). The HSR rapidly elevates levels of molecular chaperones, co-chaperones and protein degradation factors that restore proteostasis in the cytosol/nucleus. As a result, the ability of HSF1 to promote tissue health has long been attributed to its capacity to safeguard the cytosolic/nucleosolic proteome. However, over the past 15 years, it has become apparent that HSF1 activity is also intimately coupled with the biogenesis and maintenance of other organelles, including mitochondria, peroxisomes, the endoplasmic reticulum, lysosomes and chloroplasts. This suggests that HSF1 promotes tissue health in plants and animals through mechanisms beyond the maintenance of cytosolic/nucleosolic proteostasis. In this opinion piece, I will discuss advances in our understanding of the interplay between HSF1 and organelle homeostasis and make the case that the existing model for the relationship between HSF1 and tissue health should be expanded to encompass these additional roles. This article is part of the Theo Murphy meeting issue 'ProteostaSys: a systems view of proteostasis'.
    Keywords:  HSF1; healthy ageing; organelle homeostasis; proteostasis; stress responses
    DOI:  https://doi.org/10.1098/rstb.2025.0270
  27. Adv Sci (Weinh). 2026 Sep 30. e78067
    The Cytoplasmic MicroRNA-Production Complex Regulates MicroRNA Sorting to Extracellular Vesicles During Synaptogenesis.
    Yuzhou Zeng, Selina Luhmann, Aditi Saha, Marta Orlando, Haider Sami, Timo Glatter, Anna Antoniou.
      Extracellular vesicles (EVs) have emerged as novel players of cell communication, in part via the transfer of microRNA (miRNA) cargo to recipient cells. Although several RNA-binding proteins (RBPs) were shown to shuttle miRNAs to EVs, how they are loaded with miRNA is unknown. This study demonstrates that the core accessory protein of the cytoplasmic miRNA production machinery TRBP interacts with several EV-RBPs and differentially regulates miRNA secretion in small EVs. TRBP-regulated EV-miRNAs were highly enriched in a Syncrip-recognition sequence motif. Moreover, we show that Syncrip associates with TRBP via its C-terminal domain at the surface of the endoplasmic reticulum (ER), a nucleation site of the miRNA production machinery. The recruitment of Syncrip to ER-associated TRBP was regulated by ER-endosome membrane contact sites (MCS), whereby increased MCS formation promotes Syncrip loading to endosomes as well as Argonaute 2-positive RNA granules. Functionally, this pathway is important for the regulation of synapse formation in developing neurons, as expression of miRNA- or TRBP-binding deficient mutants of Syncrip differentially regulated synapse formation in EV-donor and recipient neurons. Overall, this work highlights an important role of TRBP as a gatekeeper for microRNA sorting, which could be crucial for the fine-tuning of neuronal circuits during postnatal development.
    Keywords:  RNA granule; RNA‐binding protein; extracellular vesicle; microRNA sorting; organelle contact sites; synapse formation
    DOI:  https://doi.org/10.1002/advs.78067
  28. mBio. 2026 Oct 02. e0205626
    A small molecule increases mycobacterial translation fidelity without a fitness cost by targeting ribosomal protein S5.
    Jie Wu, Swarnava Chaudhuri, Sarah Caldwell Feid, Hemant Joshi, Miaomiao Pan, Qingwen Kawaji, Gang Liu, Babak Javid.
      A central dogma of molecular biology is the "speed-accuracy trade-off," where ribosomes must slow down to ensure accurate protein synthesis. In mycobacteria, a high basal level of mistranslation at glutamine and asparagine codons, caused by an indirect tRNA aminoacylation pathway, promotes tolerance to the antibiotic rifampicin. While pharmacologically increasing translational fidelity is a promising strategy to combat antibiotic tolerance, the underlying mechanisms remain poorly understood. Here, we screened 9,000 synthetic compounds and identified benzo[d]isoxazole-4,7-diones as a novel chemical class that reduces mycobacterial mistranslation. Medicinal chemistry optimization yielded a lead compound, 9787, with superior potency in decreasing mistranslation and reversing rifampicin tolerance. Using competitive chemical proteomics, we identified the 30S ribosomal protein S5 (RpS5) as the specific cellular target. Remarkably, compound 9787 enhances translational fidelity at concentrations that do not measurably impact the overall rate of protein synthesis. Our findings challenge the universality of the speed-accuracy trade-off, demonstrating that fidelity can be improved independently of translation speed. This work reveals that the ribosomal small subunit is a druggable target for modulating translational quality control, and introduces a new strategy for combating antibiotic-tolerant bacteria without the associated fitness cost of slowed translation.IMPORTANCEA fundamental principle in molecular biology holds that ribosomes face a trade-off between translation speed and accuracy: going faster means making more errors, while maintaining high fidelity requires slowing down. This study challenges that paradigm by identifying a small molecule that increases translational accuracy in mycobacteria without affecting the rate of protein synthesis. The compound targets ribosomal protein S5 and specifically reduces errors arising from physiologically mischarged tRNAs-a quality control problem distinct from the well-studied codon-anticodon mismatches. This form of mistranslation contributes to antibiotic tolerance in tuberculosis, making it a potential therapeutic target. Our findings reveal unexpected flexibility in how ribosomes maintain translation quality and suggest that pharmacologically increasing fidelity without the fitness cost of slowed protein synthesis may be an attractive strategy for combating antibiotic-tolerant bacteria.
    Keywords:  antibiotic tolerance; medicinal chemistry; mistranslation; mycobacteria; small molecule; translational fidelity
    DOI:  https://doi.org/10.1128/mbio.02056-26
  29. bioRxiv. 2026 Sep 24. pii: 2026.09.23.753803. [Epub ahead of print]
    Redox activation of ATG5 licenses autophagy upon nutrient restriction.
    Sanghee Shin, Ekaterina D Korobkina, Claudia Lennicke, Nils Burger, Bingsen Zhang, Haopeng Xiao, Yaoyu Wang, Jonathan J Petrocelli, Shelley M Wei, David Wood, Yu Lei, Xiaomei Zhang, Maria C Perez-Matos, Muneeb A Sharif, Jiuchun Zhang, Helena M Cochemé, William B Mair, Edward T Chouchani.
      Dietary restriction (DR) protects against metabolic disease, extends lifespan, and is associated with remodeling of tissue reactive oxygen species (ROS). ROS control biological adaptation through reversible oxidation of protein cysteines, yet the targets of DR-initiated redox signaling are unknown. Here we generate OxiDR, a tissue-resolved atlas of the cysteine redox proteome that quantifies oxidation state under DR. Rather than oxidizing the proteome broadly, DR selectively targets a high-amplitude set of cysteines in a tissue-specific manner, allowing systematic classification of biological processes subject to DR-mediated redox regulation. Among the cysteines most highly oxidized upon DR is Cys19 of the core autophagy protein ATG5. We show oxidation of Cys19 is required for ATG5-mediated autophagosome formation and for autophagy triggered by nutrient restriction in human cells and mice. Reversible oxidation of this cysteine promotes ATG5 binding to ATG10, thus forming the ATG5-ATG12 conjugate that lipidates LC3B/ATG8 and matures the autophagosome. In mice, loss of this redox switch prevents effective initiation of autophagy upon nutrient restriction, resulting in gross tissue pathology and rapid onset of mortality. The autophagic response to nutrient restriction is thus gated by oxidation of a single cysteine.
    DOI:  https://doi.org/10.64898/2026.09.23.753803
  30. Cell Death Dis. 2026 Aug 01. pii: 840. [Epub ahead of print]17(1):
    Chondroitin sulfate shapes the unfolded protein response in osteosarcoma to sustain cancer cell survival during ER stress.
    Maj Sofie Ørum-Madsen, Jessica Pihl, Thomas M Clausen, Charlotte Spliid, Anne-Chloé Dhez, Hai-Feng Zhang, Yen-Hsi Chen, Elena Ethel Vidal-Calvo, Tobias Gustavsson, Michael Lizardo, Irina Nelepcu, Joey Lo, Jeffrey D Esko, Htoo Zarni Oo, Alberto Delaidelli, Mette Ørskov Agerbæk, Anne Steinø, Nader Al Nakouzi, Ali Salanti, Poul Sorensen, Mads Daugaard.
      Tumors must adapt to high levels of endoplasmic reticulum (ER) stress to sustain tumor growth and metastases. The chaperone GRP78 (BiP/HSPA5) is a key component of the unfolded protein response (UPR) and essential for ER stress management and adaptive signaling supporting pro-survival UPR activities. Here, we report that oncofetal chondroitin sulfate (CS) glycosaminoglycans are required for ER stress adaptation in osteosarcoma. When osteosarcoma cells encounter ER stress, they upregulate 4-O-sulfated CS at the expense of other glycosaminoglycans leading to a reconfiguration of the glycocalyx in favor of an oncofetal CS subtype. Genetic ablation of the CS synthesis pathway impairs the UPR by preventing osteosarcoma cells from mounting GRP78 expression in response to ER stress. CS deficiency makes osteosarcoma cells hypersensitive to inhibition of GRP78 under both ambient and ER stress conditions, and acute ER stress drives CS-defective osteosarcoma cells into an apoptotic cell death that can be rescued by re-instating CS 4-O-sulfation capacity. This has direct implications for the metastatic progression of osteosarcoma, whereby oncofetal CS protects dissociated osteosarcoma cells from anoikis to allow pulmonary colonization in mice. Our data identify CS glycosaminoglycans as a critical component of the UPR that permits osteosarcoma cells to manage ER stress.
    DOI:  https://doi.org/10.1038/s41419-026-09151-9
  31. Philos Trans R Soc Lond B Biol Sci. 2026 Oct 01. pii: 20250279. [Epub ahead of print]381(1960):
    Protein rheostasis: quality control in metastable proteomes.
    Michele Vendruscolo.
      Protein homeostasis is often described as the capacity of cellular quality-control systems to maintain proteome function by favouring functional protein states. Yet many proteins can populate multiple states, including native conformations, liquid-like condensed assemblies, and aggregated states, reflecting the metastability of the proteome. As a framework for understanding how cells preserve proteome function under such conditions, we discuss protein rheostasis as the system that regulates thermodynamic driving forces and kinetic barriers to control the flux between alternative states over time. Framing proteome maintenance in terms of rheostatic control over the multiple states helps rationalize how ageing, stress, and mutations redistribute populations towards condensed and aggregated states by eroding kinetic buffering capacity, and it suggests therapeutic opportunities that restore control by tuning the transitions between metastable states. This article is part of the Theo Murphy meeting issue 'ProteostaSys: a systems view of proteostasis'.
    Keywords:  liquid–liquid phase separation; protein aggregation; protein homeostasis; protein misfolding
    DOI:  https://doi.org/10.1098/rstb.2025.0279
  32. Nat Commun. 2026 08 31. pii: 10345. [Epub ahead of print]17(1):
    RBP-Driven RNA sorting and local translation establish the molecular identities of individual sensory axons.
    Elizabeth S Silagi, Ezechukwu Nduka, Jesus Zuniga Paiz, Maria F Pazyra-Murphy, Shamsuddin A Bhuiyan, William Renthal, Rosalind A Segal.
      Neurons extend long axons that traverse distinct microenvironments, yet how these subcellular compartments acquire and maintain specialized identities remains unclear. Here, we use spatial translatomics to define the local translatomes of somatosensory dorsal root ganglion (DRG) neurons. Translating Ribosome Affinity Purification and RNA sequencing (TRAP-seq) reveal thousands of mRNAs preferentially translated within central axons, peripheral axons, or DRG somata, establishing compartment-specific translational programs. Many of these transcripts encode ion channels and neurotransmitter receptors that may confer distinct electrophysiological and regenerative properties to each axon. Integration of the TRAP-seq data with published RNA-seq identify locally translated components that change following neuropathic injury and could thereby adjust neuronal activity. We identify RNA regulons coordinated by RNA-binding proteins (RBPs) SFPQ and SRSF10, which preferentially bind and traffic mRNAs to peripheral or central axons, respectively. These findings indicate that RBP-guided RNA sorting and local translation enable the establishment and dynamic local modulation of somatosensory function.
    DOI:  https://doi.org/10.1038/s41467-026-77192-x
  33. bioRxiv. 2026 Sep 23. pii: 2026.09.22.753649. [Epub ahead of print]
    Tracking cellular biomolecular condensate dynamics under proteostatic stress with middle-down phosphoproteomics.
    Boomathi Pandi, Dominic C M Ng, Peyton Schaal, Lorena Alamillo, Shaonil Binti, Edward Lau, Maggie P Y Lam.
      Coordination of biological function requires the partition of cellular components including into biomolecular condensates, but an overall landscape of how protein compartmentalize into higher-order assemblies under stress is still emerging. We apply proteome-wide solubility profiling to compare the compositions of NP-40-insoluble proteins and their phosphorylation status, using a new mass spectrometry-based hybrid bottom-up and chemical middle-down proteomics approach to analyze the solubility behavior of 8,740 proteins and 31,647 phosphopeptides under normal and ER stress conditions. Cell stress induces a pervasive differential partition of proteins in and out of detergent- insoluble cellular compartments. This differential partition is partially orthogonal to stress-induced abundance changes and comprises both phosphorylation-dependent and phosphorylation-independent mechanisms. Whereas phosphorylation-independent partition changes involve largely secretory pathway proteins and implicate higher-order assemblies of chaperones and clients, phosphorylation-based partitions suggest a dynamic rearrangement of biomolecular condensate compositions across cytoplasmic and nuclear ribonucleoprotein assemblies. The accumulation of serine/arginine rich (SR) proteins and other annotated nuclear speckle members in the condensate-rich proteome fractions emerges as a central feature of stress-induced remodeling. Our results establish global solubility dynamics as an integral component of proteome stress response and implicates broad involvements of splice factor spatial reorganization as a prominent facet of ER stress response.
    DOI:  https://doi.org/10.64898/2026.09.22.753649
  34. bioRxiv. 2026 Sep 07. pii: 2026.09.03.749226. [Epub ahead of print]
    Enzyme Activation of Triggerable Polymeric Lysosome Targeting Chimeras.
    Badal Singh, Priyanath Das, Aniket Majee, Adam Tajdi, Anirudh Deverajan, Allison Lomeli, Amelia Talluri, Ranit Dutta, S Thayumanavan.
      Targeted protein degradation (TPD) has been highly effective for intracellular targets, but extending this approach to extracellular and membrane-bound proteins remains difficult because most extracellular TPD (eTPD) strategies depend on ligand-targeting receptors (LTRs) whose expression and recycling vary across tissues. Existing LTR-independent, multivalent platforms expand the scope of eTPDs by eliminating this dependence on LTRs and enabling cancer-selective designs. Layering in an additional degree of selectivity through endogenously activatable triggers, such as overexpressed enzymes, can enhance the tissue tropism of the LTR-independent platforms and widen therapeutic window. Here we report triggerable polymeric lysosome targeting chimeras (tPolyTACs), which combine antibody-defined targeting with locally triggered covalent capture. tPolyTACs conjugate monoclonal antibodies to phosphatase-cleavable substrates that, upon engagement of endogenous cell-surface phosphatases, unmask a reactive quinone methide electrophile, covalently trapping the target complex and driving its clathrin-mediated internalization and autolysosomal degradation. We show tPolyTAC-mediated degradation of the membrane proteins EGFR, PD-L1, and cMET, and demonstrate that para-substituted electrophiles outperform ortho-substituted analogues likely due to more favorable active-site positioning. These results establish tPoly-TACs as a modular, covalent, enzyme-responsive platform that resolves the efficiency-selectivity trade-off limitations in extracellular degradation.
    DOI:  https://doi.org/10.64898/2026.09.03.749226
  35. Mini Rev Med Chem. 2026 Sep 28.
    Targeted Protein Degradation via the Ubiquitin-proteasome System: Emerging Advances and Therapeutic Challenges of PROTACs.
    Somnath D Bhinge, Ganesh Gadekar, Vrushabh Patil, Ramling Mali, Bahubali Patil.
       INTRODUCTION/OBJECTIVE: Proteolysis-targeting chimeras (PROTACs) are an emerging therapeutic technology in medicinal chemistry that induces selective degradation of target proteins through the ubiquitin-proteasome system. This review aims to summarize recent advances in PROTAC design, mechanisms of action, therapeutic applications, clinical development, and emerging strategies for overcoming current limitations.
    METHODS: A structured literature search was conducted using PubMed, Scopus, Web of Science, and Google Scholar for studies published between January 2010 and December 2025. Search terms included "PROTACs," "proteolysis-targeting chimeras," "targeted protein degradation," "E3 ligase," "protein degradation," and "PROTAC clinical trials." Studies addressing PROTAC design, mechanisms, medicinal chemistry, therapeutic applications, and clinical development were included. Articles unrelated to targeted protein degradation, duplicate publications, non-relevant studies, and publications lacking sufficient scientific information were excluded.
    RESULTS: The literature indicates that PROTACs can achieve efficient and selective degradation of disease-associated proteins, offering advantages over conventional occupancy-based inhibitors. Advances in E3 ligase recruitment, linker optimization, bifunctional degrader design, photoactivatable PROTACs, and dual-target degraders have substantially expanded the potential of this platform. Several PROTAC candidates have progressed into clinical development, supporting their translational potential.
    DISCUSSION: Despite significant progress, challenges remain, including poor solubility, limited oral bioavailability, pharmacokinetic constraints, tissue-specific delivery, and potential off-target effects. Computational modeling, personalized medicine, and improved drug-delivery systems may facilitate more rational and efficient PROTAC development.
    CONCLUSION: PROTACs represent a promising approach for targeted protein degradation and may expand the druggable proteome. Continued advances in medicinal chemistry, pharmacology, computational design, and delivery technologies are essential for translating PROTACs into effective clinical therapies.
    Keywords:  E3 ligase; PROTAC; drug discovery; medicinal chemistry; neurodegenerative diseases.; oncology; pharmacokinetics; targeted protein degradation; ubiquitin-proteasome system
    DOI:  https://doi.org/10.2174/0113895575486076260907143722
  36. Signal Transduct Target Ther. 2026 Sep 29. pii: 414. [Epub ahead of print]11(1):
    Polymeric alpha-1 antitrypsin perturbs Paneth cell proteostasis and exacerbates intestinal inflammation.
    Francesco Annunziata, Felipe Dos Santos Matos, Irene Relvini, Jing Lu, Giovanna Federico, Claudia D'Agostino, Valentina Schiano, Veronica Maffia, Bruno Maria Custode, Gaetano Raiola, Barbara De Marino, Rossella De Cegli, Eugenio Del Prete, Elena Polishchuk, Carmen Ambrosio, Debora Compare, Gerardo Nardone, Francesco Neri, Nunzia Pastore.
      Alpha-1 antitrypsin (AAT) is a serine protease inhibitor that protects tissue from neutrophil elastase and other proteases, particularly in the lung. Mutations in SERPINA1, including the Z mutation, lead to AAT deficiency (AATD), characterized by reduced circulating AAT and increased risk of pulmonary emphysema, liver cirrhosis, and hepatocellular carcinoma. Beyond these well-characterized manifestations, AATD has been associated with panniculitis, rheumatoid arthritis, and glomerulonephritis. Emerging evidence has also suggested a link between AATD and inflammatory bowel diseases (IBDs), although experimental validation is lacking. In this study, we demonstrate that PiZ transgenic mice expressing the polymer-forming ATZ display increased susceptibility to dextran sodium sulfate (DSS)-induced colitis, accompanied by marked Paneth cell abnormalities. The accumulation of polymeric ATZ in Paneth cells is associated with the endoplasmic reticulum (ER) stress response, impaired lysosomal clearance, altered association of Lysozyme-1 (Lyz1) with LC3-containing compartments, and increased Lyz1 secretion. These intestinal alterations were accompanied by changes in microbiota composition, whereas DSS exposure and exogenous lysozyme administration were associated with aggravated intestinal and hepatic pathology. Pharmacological inhibition of ER stress restored crypt homeostasis and normalized Lyz1 secretion. Human Pi*ZZ iPSC-derived intestinal organoids similarly showed ATZ polymer accumulation in secretory epithelial cells and transcriptional alterations involving ER protein processing and epithelial homeostasis. In addition, polymeric ATZ was detected in ileal crypts from a single individual with AATD and intestinal disease. Together, our data reveal a Paneth cell-intrinsic ER stress mechanism linking ATZ accumulation to gut epithelial dysfunction, highlighting a previously underexplored role of the gut-liver axis in AATD.
    DOI:  https://doi.org/10.1038/s41392-026-02925-9
  37. PLoS Pathog. 2026 Oct;22(10): e1014610
    Structural basis of diverse substrate recruitment by the HIV-1 Vpr-hijacked CRL4(VprBP) E3 ligase.
    Dianhong Wang, Weijia Ding, Jingwei Xu, Ruofan Li, Tamino Cairoli, Qi Liu, Yiying Zhu, Ye Xiang.
      HIV-1 hijacks the host CUL4-RING E3 ligase (CRL4) complex to trigger the degradation of various restriction factors. The viral accessory protein Vpr is the key protein that recruits diverse substrates to the receptor VprBP for proteasomal degradation. However, the underlying molecular mechanism remains poorly understood. Here, we characterize the molecular architecture of the DDB1-VprBP substrate recognition unit alone and in complex with different Vpr-substrates. Our results pinpoint that the LisH domain of VprBP mediates the DDB1-VprBP dimerization. Of note, the Armadillo-like (ARM-like) domains of VprBP adopt either an "up" or "down" conformation that may assist in accommodating Vpr-substrates of divergent stoichiometries and sizes. When bound to the small Vpr substrate UNG2, the two "up" ARM-like domains wrap around and directly interact with two symmetrical UNG2 molecules. Intriguingly, the ARM-like domains adopt a "down" conformation when bound to the larger TET2 fragment. Our structures highlight that the conformational dynamics of VprBP could enable the DDB1-VprBP complex to mediate the degradation of diverse Vpr-bound host factors. These findings could facilitate the structure-guided development of CRL4(VprBP)-based targeted protein degradation.
    DOI:  https://doi.org/10.1371/journal.ppat.1014610
  38. bioRxiv. 2026 Sep 21. pii: 2026.08.26.747332. [Epub ahead of print]
    Substrate Profiling of RNF216 Uncovers a Translation-Linked OTUD4 Regulatory Axis.
    Wei Wei, Ruochuan Liu, Jing Zhang, Shu Liu, Antoinette J Charles, Devansh G Asati, Zachary D Allen, Delance Wright, Kangli Peng, Ellissa Krekeler, Nima Mosammaparast, Jun Yin, Angela M Mabb.
      Mutations in the E3 Ubiquitin (Ub) ligase RNF216 cause Gordon Holmes syndrome (GHS), a neurodegenerative disorder accompanied by neuroendocrine disruption. We developed an orthogonal ubiquitin transfer (OUT) platform to capture RNF216 substrates in neuronal cells and identified OTUD4, a deubiquitinating enzyme (DUB) mutated in GHS, and FMRP, a neuronal-enriched translational repressor. RNF216 predominantly synthesizes K6-linked Ub chains on OTUD4 to induce its degradation, forming donut-shaped structures in neurons. In return, OTUD4 removes the ubiquitination of RNF216 and FMRP. Analysis of RNF216 substrates revealed biological functions regulating protein synthesis, a shared function of the OTUD4-RNF216 substrate interaction network. Indeed, RNF216 expression increased protein synthesis rates while Rnf216 deletion decreased dendritic development in neurons. Overall, our findings show that RNF216 and OTUD4 balance rates of protein synthesis and degradation and suggest GHS-related mutations in RNF216 or OTUD4 may offset this balance, triggering neurodegeneration.
    Highlights: Orthogonal Ubiquitin Transfer cascade identifies RNF216 substrates in neuronal cellsThe deubiquitinating enzyme OTUD4 is a RNF216 substrateRNF216 controls rates of protein synthesis and degradationRNF216 and OTUD4 operate as a catalytic pair.
    DOI:  https://doi.org/10.64898/2026.08.26.747332
  39. Protein Sci. 2026 Nov;35(11): e70814
    Pulling and gripping: Proteasomal translocation pathway elements differentially contribute to substrate unfolding.
    Edwin R Ragwan, Kristi M Pham, Arushi Palta, Destiny N McWilliams, Daniel A Kraut.
      Eukaryotic proteomes are regulated by the ubiquitin-proteasome system. Unwanted proteins are tagged with ubiquitin, which is recognized by the 26S proteasome for degradation. To enter the proteasome's 20S core particle and be hydrolyzed into peptides, the substrate must be unfolded and threaded through the 19S regulatory particle. Tyrosine "aromatic paddles" of the regulatory particle's Rpt motor subunits unfold and translocate the substrate by pulling it toward the core particle. We previously showed that for substrates degraded from their N-termini, polyglycine tracts inserted before a stable domain impair the proteasome's ability to grip or unfold the substrate at multiple points along the translocation pathway. Herein we compare unfolding from the N- and C-termini of model substrates. We find that a combination of local substrate structural elements, grip sequence, and possibly fundamental asymmetry of the proteasomal unfolding machinery that favors N-terminal unfolding affect the proteasome's ability to unfold substrates. Although the basic unfolding and translocation mechanism is conserved regardless of substrate orientation, we find differences in the ability of polyglycine tracts to disrupt unfolding from the N versus the C terminus. Finally, comparison of the rates of unfolding versus substrate release shows that different regions along the translocation pathway employ different mechanisms to facilitate unfolding and translocation. Interactions with aromatic paddles primarily increase the rate of substrate unfolding, while interactions predicted to take place with the core particle α-ring N-termini primarily decrease the rate of premature release of partially degraded substrates.
    Keywords:  ATP‐dependent protease; motor protein; proteasome; protein unfolding; translocation; ubiquitin‐proteasome system
    DOI:  https://doi.org/10.1002/pro.70814
  40. Cancer Res. 2026 Oct 02.
    EZH2 Inhibition Induces a Metabolic Stress Response Sensitizing Triple-Negative Breast Cancer to Glutaminase Targeting.
    Lucas Porras, Marina Carrière, Ann-Sophie Gironne, Hugo Philippeau, Gabriel Alzial, Elise Quadri, Jeanne Cormery, Yousef Alaessa, Anie Monast, Faustine Gorse, Myriame St-Arnaud, Mariana De Sa Tavares Russo, Sylvie Nathalie Christine Mader, Daina Z Avizonis, Sébastien Lemieux, Morag Park, Geneviève Deblois.
      EZH2, the catalytic subunit of the histone methyltransferase complex PRC2, is overexpressed and associated with poor prognosis in triple-negative breast cancer (TNBC). Although EZH2 inhibition significantly alters chromatin landscapes and gene expression, it has limited impact on the growth of TNBC models, suggesting adaptive compensatory mechanisms. Here, we demonstrated that EZH2 inhibition causes the accumulation of misfolded proteins and double-stranded RNA (dsRNA), triggering an essential integrated stress response (ISR) through PKR and PERK activation. By inducing ISR-mediated ATF4, EZH2 inhibition enhanced amino acid flux and promoted glutaminolysis to support TNBC cell survival. Pharmacological targeting of this metabolic axis with a glutaminase inhibitor in combination with EZH2 inhibition significantly impaired TNBC cell proliferation and tumor growth. These findings reveal a stress-driven metabolic adaptation that sustains TNBC survival upon EZH2 blockade and highlight inhibition of this pathway as a strategy to enhance the efficacy of EZH2 inhibitors in TNBC.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-26-0801
  41. bioRxiv. 2026 Jul 29. pii: 2026.07.28.740790. [Epub ahead of print]
    Structural Mechanisms of DNAJC13 Dimeric Assembly and InsP6 binding in Recycling Endosome Regulation.
    Tao Fu, Chan Lee, Frances V Hundley, Joao A Paulo, J Wade Harper.
      The balance of plasma membrane protein degradation and recycling during endocytosis is regulated, in part, by the large J-domain-containing protein DNAJC13/RME-8 and WASH complexes, which function together with Retromer to support cargo trafficking into recycling endosomes. Despite extensive cellular and biochemical studies, the structural basis and proteomic landscape of DNAJC13 function remain elusive. Here, we find that DNAJC13 forms an unexpected antiparallel homodimeric architecture involving distinctive symmetrical interactions between composite IWN1 and α-solenoid ARM2 domains in each protomer. Additionally, two PH-like domains of unknown function (PHL2 and PHL3), adjacent to the PI(3)P-binding PHL1 domain, form an unanticipated composite, positively charged pocket occupied by InsP6, as visualized structurally and verified by mass spectrometry. Proteomic profiling of DNAJC13-associated endosomes revealed enrichment of recycling endosomal components and WASH complexes. Mutations disrupting the dimer interface disable recruitment of WASH complexes to endosomes and result in elongated endosomal tubulation. Disruption of InsP6 binding impairs DNAJC13 binding to PI(3)P-containing vesicles in vitro and Transferrin-positive endosomes in cells. We further demonstrate that DNAJC13 dimerization and InsP6 binding promote melanin production during melanosome maturation, a process known to require recycling endosomes. This work provides a structural and mechanistic framework for understanding DNAJC13 function in recycling endosome control.
    DOI:  https://doi.org/10.64898/2026.07.28.740790
  42. bioRxiv. 2026 Sep 23. pii: 2026.09.22.753497. [Epub ahead of print]
    Nuclear LC3 Interactome Profiling Identifies Clathrin Heavy Chain as a Mediator of Nuclear LC3 Translocation in Trabecular Meshwork Cells.
    Myoung Sup Shim, Chien-Chich Chou, Mi Sun Sung, Aleks Grimsrud, Vaibhav Desikan, Nikolai Skiba, Paloma B Liton.
      Although MAP1LC3B/LC3B (LC3) is best known as a cytoplasmic marker of autophagosome biogenesis, a substantial pool of LC3 resides in the nucleus and shuttles dynamically between nuclear and cytoplasmic compartments, yet the composition and regulation of this nuclear interactome remain poorly defined. Using mass spectrometry-based proteomic profiling of nuclear GFP-LC3 immunoprecipitates from primary human trabecular meshwork (TM) cells, a mechanosensitive ocular cell type, we identified a reproducible nuclear LC3 interactome enriched for proteins containing LC3-interacting region (LIR) and expanded LIR (xLIR) motifs and reported nuclear localization. Among these, clathrin heavy chain (CLTC) emerged as a previously unrecognized nuclear LC3 partner that localizes to the nucleus and colocalizes with nuclear LC3 puncta. CLTC depletion reduced basal LC3-II levels, consistent with a role in autophagosome biogenesis, and markedly impaired nuclear LC3 accumulation induced by both nuclear export blockade with leptomycin B and cyclic mechanical stretch, without altering total CLTC abundance, indicating that CLTC actively promotes LC3 nuclear translocation rather than serving as a passive scaffold. Strikingly, mechanical stress-induced nuclear LC3 trafficking, but not its basal component, was selectively impaired in TM cells derived from glaucoma patients, despite comparable CLTC levels, pointing to a defect in coupling this trafficking pathway to mechanotransduction rather than in the core transport machinery itself. Together, these findings establish nuclear LC3 trafficking as an actively regulated, CLTC-dependent process linked to cytoskeletal and vesicular machinery, and implicate its dysregulation in mechanically stressed glaucomatous cells, providing a framework for understanding how autophagy intersects with nuclear homeostasis, mechanotransduction, and glaucoma pathogenesis.
    DOI:  https://doi.org/10.64898/2026.09.22.753497
  43. bioRxiv. 2026 Sep 09. pii: 2026.09.04.749512. [Epub ahead of print]
    A novel mechanism of ubiquitin-charging a bivalent E2:RING E3 complex.
    Samuel Perreault, Ayush Mistry, Edwin Antony, Mark Hedglin.
      Protein ubiquitination results from a cascade of enzyme interactions that transfer ubiquitin from one covalent bond to another. First, an E1 is activated by attaching ubiquitin to itself through a thioester bond. Next, in E2 charging, activated E1 interacts with an E2 and transfers ubiquitin to a thioester bond on the E2. Finally, an E3 mediates the transfer of ubiquitin from a charged E2 to an isopeptide bond on a protein target. RING E3s, the largest E3 family in eukaryotes, function as scaffolds rather than enzymes and facilitate direct transfer of ubiquitin from a charged E2 to a protein target by simultaneously engaging both. All RING E3s engage an E2 at an interface that overlaps with the E1-binding site and several form stable, bivalent complexes with an E2 through additional binding surfaces. How E2 charging proceeds within these complexes remains unclear. Here, we utilize human Rad6(Rad18) 2 as a model bivalent E2:RING E3 complex to delineate the interplay of protein•protein interactions among the primary human E1 (Uba1), an E2 (Rad6), and a RING E3 (Rad18) during E2 charging. Collectively, the results reveal a novel mechanism that is not directed by ubiquitin thioester "affinity switches." Rather, interactions of Rad18 with Rad6 slow the chemistry step and all preceding steps of the Uba1 catalytic cycle via competitive inhibition but accelerate release of charged Rad6 from Uba1, ultimately stimulating Rad6 charging overall. To the best of our knowledge, this represents the first example of a RING E3 stimulating Uba1-dependent charging of an E2.
    DOI:  https://doi.org/10.64898/2026.09.04.749512
  44. STAR Protoc. 2026 Sep 26. pii: S2666-1667(26)00514-9. [Epub ahead of print]7(4): 104861
    Protocol for ribosome-protected fragment isolation from rare leukemia cell populations.
    Christina Mayerhofer, Ernst Mayerhofer, Dan Li, Trine Kristiansen, Tobias Feuchtinger, David T Scadden.
      Malignant cells under chemotherapy stress alter protein translation, necessitating sensitive methods to profile rare, surviving populations in vivo. We present a protocol for profiling low-input acute myeloid leukemia samples using an optimized ribosome profiling approach. We describe steps for lysing cell inputs, digesting unshielded RNA, enriching ribosome footprints, and gel-based size selection. We then detail procedures for constructing sequencing-ready libraries and using a computational pipeline to align and quantify both protected fragments and matching transcriptomes for reproducible translation analysis. For complete details on the use and execution of this protocol, please refer to Mayerhofer et al.1.
    Keywords:  Cancer; Cell Biology; Metabolism
    DOI:  https://doi.org/10.1016/j.xpro.2026.104861
  45. Adv Sci (Weinh). 2026 Sep 27. e77898
    TROP2-Targeting Chimeras (TRTACs) for Tumor Cell-Selective Membrane Protein Degradation and Enhanced Drug Delivery.
    Luping Chen, Xinying Fu, Wenqian Dong, Xiaobei Deng, Simin Chen, Fujun Wang, Jian Zhao, Shiqun Shao, Liqiang Fan, Jingyu Zhang, Lixin Zhang.
      Extracellular targeted protein degradation (eTPD) systems typically utilize lysosome-targeting receptors (LTRs) to redirect extracellular and membrane proteins for lysosomal degradation. Expanding the repertoire of clinically relevant LTRs could broaden the therapeutic potential of eTPD. Here we report trophoblast cell surface antigen-2 (TROP2), a clinically validated tumor-associated antigen targeted by approved antibody-drug conjugates, as a promising LTR for tumor cell-selective eTPD. We develop TROP2-targeting chimeras (TRTACs) by fusing a TROP2-binding nanobody to target-specific nanobodies. This fully genetically encoded, ∼30 kDa nanobody format enables modular construction, broad target adaptability, and favorable tumor penetration. TRTACs induce degradation of diverse membrane proteins, including epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), and programmed death-ligand 1 (PD-L1), preferentially in tumor cells rather than in nonmalignant cells. We further designed TRTAC-drug conjugates (TRTAC-DCs) by attaching cytotoxic payloads to TRTACs, enabling enhanced drug delivery together with protein degradation. The HER2-targeted TRTAC-DC reduced SKBR3 cell viability by 50% at 0.2 nM and showed potent antitumor activity in vivo, without significant body weight loss or histological abnormalities in major organs. These findings establish TROP2 as a clinically relevant LTR and provide a modular platform that combines tumor cell-selective extracellular protein degradation with enhanced payload delivery.
    Keywords:  EGFR; TROP2‐targeting chimera; degrader‐drug conjugate; extracellular targeted protein degradation; nanobody
    DOI:  https://doi.org/10.1002/advs.77898
  46. Aging Dis. 2026 Sep 09.
    Ubiquitin-Dependent Protein Quality Control in Polyglutamine Diseases.
    Matthew V Prifti, Kristin Richardson, Sokol V Todi.
      Polyglutamine diseases comprise a family of nine age-dependent disorders caused by CAG triplet-repeat expansions that produce misfolded proteins with elongated glutamine tracts. Although these diseases share proteotoxic stress and engagement of protein quality control pathways as responses to their presence and disruptive activities, they differ in protein context, normal function, localization, interactomes and selective vulnerability. Here, we review how expanded polyglutamine proteins engage ubiquitin-dependent protein quality control across Huntington's disease, dentatorubral-pallidoluysian atrophy, spinal and bulbar muscular atrophy, and spinocerebellar ataxias 1, 2, 3, 6, 7 and 17. Rather than treating protein quality control as a uniform response, we emphasize pathway selectivity. A central theme arises across these diseases: mutant polyglutamine proteins are recognized by ubiquitin-related machinery, but recognition does not necessarily produce productive degradation. Instead, disease-specific outcomes are shaped by tissue context, subcellular localization, aggregate state, disease protein fragmentation, normal protein function, compensatory pathway activation and local proteostasis capacity over time. The information synthesized in this review may help the field develop a more comparative and granular understanding of processes and pathways that are shared or divergent among polyglutamine diseases and may also inform more targeted therapeutic strategies for this family of incurable disorders.
    DOI:  https://doi.org/10.14336/AD.2026.0887
  47. Proc Natl Acad Sci U S A. 2026 Oct 06. 123(40): e2528514123
    BAG3 coordinates astrocytic proteostasis of Alzheimer's disease-linked proteins via proteasome, autophagy, and retromer interactions.
    Zachary M Augur, Garrett M Fogo, Courtney R Benoit, Masin A Kearney, Gizem Terzioglu, Zachary R Murphy, Mason R Arbery, Natacha Comandante-Lou, Duc M Duong, Nicholas T Seyfried, Philip L De Jager, Tracy L Young-Pearse.
      Bcl-2-associated athanogene 3 (BAG3) is a mediator of chaperone-assisted selective autophagy, and in the brain, is most highly expressed in astrocytes. However, its role in astrocytes remains poorly defined. Given the genetic and pathological links of BAG3 to proteostasis and neurodegenerative diseases, we investigated how BAG3 contributes to astrocyte function and Alzheimer's disease (AD). To define its function and relevance, we used single-nucleus RNA sequencing to confirm BAG3 enrichment in astrocytes and employed CRISPR/Cas9 editing of human induced pluripotent stem cells followed by proteomic and transcriptomic profiling, which revealed that BAG3 loss caused greater disruption in astrocytes than in neurons. BAG3-deficient astrocytes displayed reduced autophagy, lysosomal abundance and activity, and proteasome function. Coimmunoprecipitation identified BAG3 known binding partners (e.g., HSPB8, proteasome regulators), as well as an interactor in the retromer complex, VPS35. BAG3 deficiency resulted in altered retromer activity as measured by amyloid precursor protein (APP) localization in endosomes. In addition to validating these binding partners, integrative -omics analyses showed that BAG3 regulates AD-relevant proteins (GFAP, BIN1), as well as HSPB8. Functionally, BAG3 knockout astrocytes exhibited impaired amyloid-β proteostasis when cocultured with APP/PSEN1 mutant neurons, directly linking BAG3 to a disease-relevant astrocyte phenotype. Finally, analysis of postmortem human brain revealed that BAG3 expression marks a stress-responsive astrocyte subtype in aged individuals. Together, these findings demonstrate that BAG3 coordinates astrocyte proteostasis through interactions with regulators of autophagy, proteasome activity, and retromer function, positioning it as a potential therapeutic target and central node of astrocytic protein quality control in neurodegeneration.
    Keywords:  Alzheimer’s disease; BAG3; astrocytes; autophagy; iPSC
    DOI:  https://doi.org/10.1073/pnas.2528514123
  48. bioRxiv. 2026 Sep 23. pii: 2026.09.22.752907. [Epub ahead of print]
    Integrated stress response activation in combination PG3 and cisplatin-treated TP53 -mutated head and neck squamous carcinoma cells induces potent apoptotic response.
    Xiaobing Tian, Praveen R Srinivasan, Wafik S El-Deiry.
      Cisplatin remains as standard chemotherapy for patients with HNSCC, but rapid development of drug resistance has limited patient benefit. The p53 tumor suppressor plays a central role in the cellular response to DNA damage in cancer, triggering apoptosis to prevent propagation of damaged cells in tumor development. TP53 gene mutations occur in 65-86% of HNSCC. Small molecule PG3 induces the integrated stress response (ISR), leading to apoptosis via the HRI-eIF2α-ATF4-PUMA axis. We hypothesized that a combination of PG3 plus cisplatin could increase apoptosis in TP53 -mutated HNSCC cells through enhanced induction of the ISR and ATF4. PG3 synergized with cisplatin to inhibit cell viability, leading to potent apoptosis in TP53 -deficient cells. The effect was regulated through the HRI-ATF4-NOXA pathway. Furthermore, we identified that cisplatin activates HRI and leads to the degradation of CReP (constitutive repressor of eIF2α phosphorylation) via E3 ligase β-TrCP, contributing to the induction of the ISR. We noted decreased ATF4 levels after treatment with cisplatin, CPT, or PG3 and cisplatin. Thus, combined therapy of PG3 plus cisplatin likely results in adaptation and acquired resistance via degradation of ATF4. We targeted the degradation mechanism of ATF4 by inhibiting β-TrCP1, CK1δ, or CK2, respectively. Each approach successfully blocked ATF4 degradation induced by cisplatin or PG3 plus cisplatin and enhanced apoptosis. Our results provide a rational strategy for triple treatments, involving an ISR inducer, a DNA damaging drug, and a β-TrCP inhibitor/CK1δ inhibitor/CK2 inhibitor, to achieve potent and prolonged anti-tumor effects and overcome chemoresistance in HNSCC.
    DOI:  https://doi.org/10.64898/2026.09.22.752907
  49. Exp Mol Med. 2026 Oct 01.
    D-allose reprogrammes the mitophagy-mtDNA-cGAS-STING-ADM2 axis to restore nucleus pulposus cell homeostasis in intervertebral disc degeneration.
    Zhuoyang Zhao, Jiamin Li, Linchuan Lei, Yanting Chen, Zhe Li, Yuhua Zhang, Zhaomin Zheng, Hua Wang, Fan Chen, Jianru Wang.
      Intervertebral disc degeneration (IVDD) is driven by progressive loss of nucleus pulposus cell (NPC) homeostasis, yet the stress-responsive mechanisms governing this process remain incompletely defined. Here, we identify a mitochondrial stress-innate immune signalling axis in which impaired mitophagy promotes mitochondrial DNA (mtDNA) leakage, aberrant activation of the cGAS-STING pathway, suppression of the cytoprotective peptide adrenomedullin 2 (ADM2) and consequent NPC senescence and apoptosis. Using integrated transcriptomic analyses, genetic models and human disc specimens, we establish ADM2 as a previously unrecognized downstream effector linking innate immune activation to disc cell fate control. Importantly, we demonstrate that D-allose, a naturally occurring rare sugar with an excellent safety profile, reprogrammes this pathogenic axis by restoring mitophagy, limiting cytosolic mtDNA accumulation, suppressing cGAS-STING signalling and preserving ADM2-dependent cytoprotection. In vivo, D-allose markedly attenuates IVDD, with genetic ablation of STING providing complementary evidence that suppression of this pathway confers robust protection against degeneration. Together, these findings define a mitophagy-mtDNA-cGAS-STING-ADM2 axis as a central regulator of IVDD and reveal D-allose as a physiologically compatible strategy for rebalancing mitochondrial stress signalling and restoring NPC homeostasis.
    DOI:  https://doi.org/10.1038/s12276-026-01844-7
  50. Cell Stress Chaperones. 2026 Sep 30. pii: S1355-8145(26)00079-9. [Epub ahead of print] 100221
    The barcoded binding assay: Multiplexed profiling of chaperone binding selectivity for hydrophobic substrates.
    Shawn P Allen, Matthew L Wohlever.
      Membrane proteins contain hydrophobic sequences that must be chaperoned during protein folding and transport to the lipid bilayer. Failures in this essential process result in the formation of potentially toxic aggregates. Our mechanistic understanding of how chaperones interact with hydrophobic sequences is limited due to the technical challenges of measuring interactions between aggregation prone proteins in a detergent-free environment. As hydrophobic substrates are prone to non-specific interactions, traditional binding assays that utilize one substrate and one chaperone per assay struggle to demonstrate that an interaction is specific and physiological. To address these technical challenges, we developed the barcoded binding assay, which allows for the measurement of chaperone binding selectivity to a complex pool of substrates in a detergent-free environment. In this assay, a pool of 35S-labeled barcoded substrates is produced by in vitro translation in the presence of the chaperone of interest and chaperone selectivity is quantified by immunoprecipitation, SDS PAGE, and autoradiography. We previously used this assay to identify novel substrate binding preferences for the Ubiquilin family of chaperones. Here, we provide an overview of the assay, demonstrate the benefits of the multiplexed barcoded binding assay over a single substrate approach, and provide a detailed protocol for how this assay can be adapted to work with a broad array of chaperones.
    Keywords:  Ubiquilin; chaperone-substrate binding; in vitro translation; membrane protein biogenesis; proteostasis
    DOI:  https://doi.org/10.1016/j.cstres.2026.100221
  51. Proc Natl Acad Sci U S A. 2026 Oct 06. 123(40): e2620180123
    Desulfinylation of the deubiquitinase USP7 by SRXN1 attenuates oxidative acute liver injury.
    Mengyun Ke, Jong-Won Kim, Meishu Xu, Jingyuan Wang, Lingyi Liu, Syamprasad N P, Bin Yang, Xiaofei Wang, Huatian Li, Songrong Ren, Kate S Carroll, Song Li, Wen Xie.
      Protein S-sulfinylation is a reversible oxidative cysteine modification, but its contribution to acute liver injury such as those induced by acetaminophen (APAP) overdose has not been reported. Sulfiredoxin-1 (SRXN1) is the only oxidoreductase known to reduce the sulfinylated proteins, yet the substrates and mechanisms through which it protects against acute liver injury are unclear. Here, we show that hepatic protein S-sulfinylation was markedly upregulated in APAP-overdose patients and mice. Hepatocyte-specific knockout or pharmacological inhibition of Srxn1 sensitized mice to APAP-induced acute liver injury. In contrast, overexpression of SRXN1, but not its oxidoreductase-dead C99S mutant, protected mice from APAP-induced liver injury. Mechanistically, we identified the deubiquitinase ubiquitin-specific protease 7 (USP7) as an SRXN1 substrate required for hepatoprotection. USP7 inhibition worsened APAP-induced liver injury, whereas its overexpression alleviated injury by stabilizing its deubiquitinase substrate heme oxygenase-1 (HO-1). Loss of SRXN1 enhanced S-sulfinylation of USP7 at Cys315, promoting its ubiquitination and degradation. SRXN1 expression correlated with USP7 levels in human liver samples. Delayed lipid nanoparticle (LNP) delivery of SRXN1 messenger RNA (mRNA) mitigated established APAP-induced liver injury beyond the therapeutic window of N-acetylcysteine. These findings define an SRXN1-USP7-HO-1 axis as a promising therapeutic target for oxidative liver injury and highlight LNP-mediated SRXN1 mRNA delivery as a potential treatment approach.
    Keywords:  S-sulfinylation; SRXN1; mRNA-LNP therapy; oxidative acute liver injury; ubiquitin-specific protease 7
    DOI:  https://doi.org/10.1073/pnas.2620180123
  52. J Biol Chem. 2026 Sep 29. pii: S0021-9258(26)02484-1. [Epub ahead of print] 113612
    CAPZ regulates endosomal-exosomal trafficking of plasma membrane PD-L1 independently of canonical autophagy.
    Peng Xu, Zuodong Ye, Yanni Zhu, Chuen-Fuk Chan, Yanhua Li, Huiru Huang, Chengyi Zhu, Xiaoyin Zhang, Yijing Wang, Yilin Wang, Naixin Lin, Wei Sun, Meiyu Peng, Yingying Lu, Jianbo Yue.
      Programmed death-ligand 1 (PD-L1) undergoes continuous endocytosis and post-endocytic sorting that determine its recycling to the plasma membrane, lysosomal degradation, and exosomal secretion. Although PD-L1 internalization depends on RAB5-mediated endocytosis, whether canonical autophagy contributes to the subsequent sorting of internalized PD-L1 remains unclear. Here, we show that canonical autophagy is dispensable for the post-endocytic fate of cell-surface PD-L1. Genetic disruption of core autophagy components, including LC3B, ATG4B, ATG5, and ATG7, did not impair delivery of internalized PD-L1 to early endosomes, multivesicular bodies (MVBs), late endosomes, or extracellular vesicles. Pharmacologic inhibition of autophagosome-lysosome fusion increased PD-L1 accumulation in RAB5- and CD63-positive compartments, but this effect persisted in cells lacking LC3B, ATG5, or ATG7, further indicating that canonical autophagy is not required for PD-L1 endosomal sorting or exosomal secretion. Instead, we identify CAPZ, a CAPZα-CAPZβ heterodimer best known for actin filament capping, as a regulator of PD-L1 post-endocytic sorting. Loss of CAPZ reduced PD-L1 delivery to CD63-positive MVBs and incorporation into exosomes while increasing its accumulation in RAB11-positive recycling endosomes, resulting in elevated cell-surface PD-L1. Functionally, CAPZ-deficient tumor cells were less sensitive to peripheral blood mononuclear cell-mediated killing, consistent with increased surface PD-L1 and enhanced immune-evasive capacity. Together, these findings indicate that CAPZ-dependent endosomal maturation controls the balance between PD-L1 recycling and MVB/exosomal sorting independently of canonical autophagy, thereby influencing PD-L1 surface abundance and tumor immune evasion.
    DOI:  https://doi.org/10.1016/j.jbc.2026.113612
  53. Philos Trans R Soc Lond B Biol Sci. 2026 Oct 01. pii: 20250266. [Epub ahead of print]381(1960):
    Proteostasis and ageing dissidence.
    Neethu Babu, Katie Whalen, Brian Freeman.
      Proteostasis, the process governing the dynamic regulation of protein synthesis, folding and degradation, is critical for maintaining cell function and organismal health. Ageing disrupts this intricate system, leading to inactive, misfolded and/or aggregated proteins that contribute to age-associated pathologies. Here, we explore current findings on proteostasis and its deterioration during ageing with an attention to three major pathways: molecular chaperone (MC), ubiquitin-proteasome system (UPS) and autophagy-lysosomal pathway. Components in all three paths undergo age-dependent decline in both expression and function, with each impairment having select initial outcomes on the health of a proteome. For example, loss in the MC path may cause nascent chain defects, whereas a decline in the UPS may result in the accumulation of toxic aggregates. Notably, the interconnectedness of these pathways results in reciprocal reactions in all three. Understanding how each path connects to the others and how these connections are regulated offers promising strategies to restore proteome integrity and extend a healthy lifespan. This article is part of the Theo Murphy meeting issue 'ProteostaSys: a systems view of proteostasis'.
    Keywords:  ageing; autophagy–lysosome pathway; molecular chaperone; proteostasis; ubiquitin–proteasome system
    DOI:  https://doi.org/10.1098/rstb.2025.0266
  54. bioRxiv. 2026 Sep 13. pii: 2026.09.12.751112. [Epub ahead of print]
    Toxoplasma gondii GRA66 prevents premature egress driven by the host phospholipase RARRES3, independently of RNF213.
    Caroline de Moraes de Siqueira, Stephanie Martinez-Beltran, Emma A Johnston, Jörn Coers, Jeroen P J Saeij.
      Toxoplasma gondii replicates inside a host-derived parasitophorous vacuole (PV), and interferon-gamma (IFNγ) induces host restriction factors that target this compartment. Previous CRISPR screens identified the dense granule protein GRA66 and the GRA57/GRA70/GRA71 complex as required for parasite fitness in IFNγ-stimulated human cells, but the host pathways they oppose were unknown. Here, we show that GRA66, a PV membrane (PVM)-associated protein predicted to be an N-acylphosphatidylethanolamine (NAPE)-hydrolyzing phospholipase D, is required to prevent premature egress driven by the host phospholipase and acyltransferase RARRES3. Loss of GRA66 caused premature parasite egress, host cell death, and impaired replication in human cells. These phenotypes persisted in cells lacking RNF213, the E3 ubiquitin ligase that dominates IFNγ-dependent Toxoplasma restriction in human cells, and Δgra66 vacuoles recruited less RNF213 and ubiquitin than wild type, indicating an RNF213-independent mechanism rather than an exaggerated RNF213 response. Complementation with a catalytic-site mutant of the GRA66 zinc-binding motif failed to restore any of these phenotypes, indicating a requirement for its predicted enzymatic activity. Deleting RARRES3 rescued the premature egress of Δ gra66 and catalytic-mutant parasites but not of Δgra70 parasites, whereas the replication defect persisted, revealing a second, RARRES3-independent consequence of GRA66 loss. RARRES3 was recruited to the PVM and intravacuolar network after IFNγ stimulation, and structural modeling supported assignment of GRA66 to the NAPE-phospholipase D family with an intact di-zinc active site. These findings define a lipid-centered, RNF213-independent arm of human cell-autonomous immunity and identify the parasite effector required to withstand it.
    Importance: Toxoplasma gondii infects a large fraction of the world's population and persists for life inside host cells, sheltered within a compartment called a vacuole. The immune signal interferon gamma normally holds the parasite in check, but how human cells attack this vacuole, and how the parasite resists, is only partly understood because humans lack the main anti-vacuolar defenses mice use. Here, we show that human cells recruit a lipid-remodeling enzyme, RARRES3, to this vacuole, and that RARRES3 drives the parasite to exit prematurely, triggering host cell death. The parasite counters with a lipid-modifying enzyme of its own, GRA66, which is required to prevent that premature exit. This host defense works independently of the pathway thought to dominate human control of Toxoplasma . Our findings identify lipid remodeling of the pathogen-containing vacuole as a distinct arm of human cell-intrinsic immunity, and the parasite enzyme that counteracts it as a potential drug target.
    DOI:  https://doi.org/10.64898/2026.09.12.751112
  55. Proc Natl Acad Sci U S A. 2026 Oct 06. 123(40): e2605220123
    Tandem RNA-binding domain architecture drives PKR activation through an intramolecular interface.
    Chong Han, Chen Li, Yi-Feng Xu, Ming Rao, Shao-Qing Zhang, Ling-Ling Chen, Jiaquan Liu.
      Tandem repeats of RNA-binding domains (RBDs), often linked by intrinsically disordered sequences, are prevalent among RNA-binding proteins (RBPs). The inherent flexibility of these arrangements, however, raises the question of whether such domains can adopt defined configurations critical for function. Here, we reveal that in Protein Kinase R (PKR)-an RBP that triggers the innate immune response upon sensing double-stranded RNAs (dsRNAs)-its two dsRNA-binding domains (dsRBDs) assemble into a unique architecture through intramolecular interactions. This dsRBD1:dsRBD2 interface allows PKR to convert diffusive, transient contacts with RNA into a stabilized complex on the duplex, establishing a scaffold required for downstream protein dimerization and phosphorylation. Disrupting this dsRBD1:dsRBD2 interface dismantles the tandem dsRBD architecture and severely impairs PKR activity both in vitro and in cells. Together, these findings uncover an exceptional advantage of the tandem dsRBD arrangement and underscore intramolecular RBD coupling as a mechanism for regulating RBP activity.
    Keywords:  Protein Kinase R; dsRNA-binding domain; immunogenic dsRNA; single-molecule imaging
    DOI:  https://doi.org/10.1073/pnas.2605220123
  56. Cell Mol Life Sci. 2026 Sep 30. pii: 350. [Epub ahead of print]83(1):
    Biphasic SMN condensates safeguard stress-induced cytoplasmic mislocalization of splicing RNPs.
    Yannick L Riedel, Jessica Dressler, Maximilian T Schilling, Oliver J Gruss.
      Biogenesis of UsnRNPs occurs in distinct steps in the nucleus and the cytoplasm. Sequential cytoplasmic actions of CLNS1A in the PRMT5 complex and of the SMN complex assemble the Sm core structure consisting of RNA and proteins. Nuclear SMN, condensed in Cajal Bodies, promotes late maturation steps. Whether cytoplasmic SMN undergoes condensation, and how this contributes to UsnRNP biogenesis or homeostasis, is poorly defined. Here, we show that molecular crowding stress induces rapid, reversible condensation of cytoplasmic SMN into droplets and filamentous assemblies, S-bodies, that sequester mislocalized cytoplasmic UsnRNPs along microtubules. During stress recovery, S-bodies undergo microtubule-dependent reorganization into split SMN-CLNS1A condensates, Janus bodies, that promote clearance of mislocalized UsnRNPs. Strikingly, cellular models of the SMN-associated disease spinal muscular atrophy fail to assemble S-bodies and to clear cytoplasmic UsnRNPs. Our findings identify stress-induced SMN condensation as a mechanism to buffer and resolve cytoplasmic UsnRNP mislocalization and reveal impaired tolerance to molecular crowding as a hallmark of spinal muscular atrophy.
    Keywords:  Microtubules; Spinal muscular atrophy (SMA); Stress; Survival motor neurons (SMN); UsnRNP
    DOI:  https://doi.org/10.1007/s00018-026-06459-9
  57. bioRxiv. 2026 Sep 26. pii: 2026.09.25.754437. [Epub ahead of print]
    Variant-Specific Rewiring of Lysosomal Positioning by STARD9 Produces Opposite Cholesterol Trafficking Outcomes.
    Mehdi Dianatpour, Dipanjan Banerjee, HyunKyung Kim, Jiyoung Moon, Rui Zhu, Hang Zhou, Arpita Neogi, Trojan Rugira, Shrikant Mane, Adebowale Adeniran, Brisa Paliuqi, Ho-Joon Lee, Lucila Ohno-Machado, Arya Mani.
      Residual cardiovascular risk persists despite lipid-lowering therapy, independent of LDL and Lp(a), reflecting cholesterol dysregulation. We identify STARD9, a lysosomal cholesterol-sensing kinesin, as a causal gene for divergent familial dyslipidemias and premature atherosclerosis. Its START domain binds cholesterol to govern lysosomal positioning. Rare variants segregate with autosomal dominant hypercholesterolemia and premature coronary disease, while common variants associate with reduced HDL and elevated triglycerides. Both converge on lysosomal cholesterol sequestration, ER depletion, mTORC1 SREBP2 activation, impaired autophagy, and NF-kB inflammation, yet diverge through opposite lysosomal positioning: the rare variant disperses lysosomes peripherally, phenocopying START domain loss across cholesterol sequestration, lysosomal positioning, and TFEB activation, triggering LDLR degradation and CASM/TFEB-dependent efflux that preserves HDL, whereas the common variant causes perinuclear retention that suppresses TFEB dependent lipid-handling transcripts, yielding hypertriglyceridemia and low HDL. In Stard9-knockout mice, enterocyte cholesterol sequestration drives SREBP2 dependent NPC1L1 upregulation and apical localization, increasing intestinal cholesterol absorption. These findings position lysosomal cholesterol trafficking as a targetable node in statin-refractory cardiovascular disease.
    DOI:  https://doi.org/10.64898/2026.09.25.754437
  58. Natl Sci Rev. 2026 Sep;13(18): nwag446
    Stepwise-coordinated polymeric nanoplatform enables traceless cytosolic protein delivery and autophagy blockade for enhanced tumor therapy.
    Peng Zhang, Yaowei Zhu, Hongyu Zhang, Shuai Tang, Zhihui Guo, Xuan Yi, Chunsheng Xiao, Xuesi Chen.
      Autophagic degradation of proteins following cellular internalization and endosomal escape presents a significant, yet underexplored, challenge in intracellular protein delivery. This highlights the need for delivery carriers capable of concurrently achieving cytosolic delivery and autophagy inhibition. Here, we report a stepwise-coordinated polymeric nanoplatform based on a tailor-made amphiphilic triblock copolymer (mPEG-b-PGBA-b-PGCQ) that enables traceless cytosolic delivery of functional proteins while concurrently suppressing autophagy. This system incorporates three modular blocks: an mPEG stealth shell, a PGBA block for reversible protein conjugation via pH-sensitive iminoboronates, and a PGCQ block grafted with autophagy inhibitor hydroxychloroquine (HCQ) via self-immolative disulfide linkages. Using ribonuclease A (RNase A) as a model therapeutic, we demonstrated efficient protein release in acidic endolysosomes, simultaneous HCQ-promoted endosomal escape, and glutathione-triggered HCQ liberation in cytosol. The released HCQ significantly enhanced RNase A-induced apoptosis by inhibiting both the autophagic degradation of the protein and RNase A-induced mitophagy. Furthermore, by integrating cytosolic delivery with autophagy inhibition, this platform produced notable functional enhancement across multiple proteins, including green fluorescent protein, β-galactosidase, and horseradish peroxidase, demonstrating its broad utility. Our work establishes a potent traceless delivery and autophagy-blockade strategy for boosting intracellular protein efficacy, with considerable potential in both basic research and biomedical applications.
    Keywords:  autophagy blockade; polymer nanocarrier; protein delivery; protein drug; tumor therapy
    DOI:  https://doi.org/10.1093/nsr/nwag446
  59. Cancer Immunol Res. 2026 Sep 28.
    ULK1-dependent autophagy initiation promotes MHC-I degradation and immune evasion in HPV-positive head and neck cancer.
    Lexi Vu, Nicholas S Giacobbi, Mohamed I Khalil, Canchai Yang, William J Eckerman, Evelyn D Gomez Recinos, Johnathon D Garber, Abigail Z Bennett, Bryttan Adams, Hyomin Son, Prerna Chahal, Dara M Villa, Tanisha Srivastava, Katie R Martin, Craig C Welbon, Caitlin S Williamson, William C Spanos, Jeffrey P MacKeigan, Andrew J Olive, Dohun Pyeon.
      Antigen presentation by major histocompatibility complex class I (MHC-I) is critical for tumor cell killing by CD8+ T cells. In human papillomavirus-positive head and neck cancer (HPV+ HNC), where MHC-I downregulation is frequent despite favorable immune cell infiltration, lower MHC-I levels are associated with poor responses to immune checkpoint inhibitor therapy. However, the mechanism of MHC-I degradation remains elusive. Genome-wide CRISPR screens in HPV+ HNC identified components of the ULK1 and PIK3C3 autophagy initiation complexes among top negative regulators of cell-surface MHC-I. In contrast, inhibiting post-initiation stages of autophagy did not restore cell-surface MHC-I, highlighting a critical role for autophagy initiation in regulation of MHC-I. Mechanistically, we showed that MHC-I is recruited from the ER to autophagosomes by the cargo receptor NDP52. Finally, inhibition of autophagy initiation suppressed HPV+ HNC tumor growth in vivo and enhanced the CD8+ T cell-mediated antitumor response. Our findings suggest that autophagic degradation of MARCHF8-ubiquitinated MHC-I is a key immune evasion mechanism in HPV+ HNC.
    DOI:  https://doi.org/10.1158/2326-6066.CIR-26-0421
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