bims-proteo Biomed News
on Proteostasis
Issue of 2026–08–23
fifty-one papers selected by
Eric Chevet, INSERM
  1. The UPR drives gametogenesis via an unexpected response, coordinating translation and ER structure.
  2. Human Hsp70 paralogs display selective J-domain interactions tuning proteostasis under stress.
  3. BAG6-RNF115 Couples Protein Quality Control with Ribosome Assembly.
  4. De Novo-Designed Bifunctional Proteins for Targeted Protein Degradation.
  5. Reconstitution of glycan-driven MHC-I recycling reveals calreticulin as mediator between TAPBPR and tapasin.
  6. Selenoprotein S associates with complexes governing membrane protein biogenesis and translation-associated processes.
  7. Astrocytes mobilize a broader repertoire of lysosomal repair mechanisms than neurons.
  8. Endoplasmic reticulum engagement of transcripts triggers VSG mRNA balancing in trypanosomes.
  9. Astrocytes instructively regulate neuronal translation.
  10. FAF1 and FAF2 enhance unfolding by p97-UFD1-NPL4 complex enabling rational design of p97 activators.
  11. ER-liquid condensate contacts sequester FAM134B/C and RhoA to govern cell morphology.
  12. A New Recruitable E3 Ligase UHRF1 Supporting Targeted Protein Degradation: A Minimal Azide as a Recruitment Ligand.
  13. PKA-dependent phosphorylation drives stress granule maturation and controls recovery from prolonged starvation.
  14. A longevity-associated ubiquitin E3 ligase UBE3C governs lamin B1 homeostasis through selective autophagy and delay senescence.
  15. EOLA1 functions in nucleotide salvage through deacetylating free N4-acetylcytidine.
  16. Mitochondrial profiling across macrophage states reveals inhibition of IL-4/IL-13 reprogramming by the integrated stress response.
  17. FAF1 cofactor enhances UFD1/NPL4-p97 unfolding efficiency across ubiquitin chain lengths independent of SUMO2.
  18. Chronic stress disrupts the network among stress granules, P-bodies, and motor proteins.
  19. An ER retention motif controls the heteromeric stoichiometry of hERG1a/1b channels.
  20. Yeast Rai1p and the RAC complex (Ssz1p/Zuo1p) modulate uORF-mediated GCN4 translational control under stress conditions.
  21. Dealing with membrane stress through CASM.
  22. RPL41 stabilizes ribosome dynamics and supports long-protein homeostasis in mammals.
  23. ESCAPE: assigning site-specific activity to covalent ligands in cells by prime editing.
  24. The degrader gets degraded as cysteine fluctuates.
  25. Transmembrane Protein Disulfide Isomerase TMX3 Potentiates Platelet Aggregation and Arterial Thrombosis in Mice.
  26. Human SRP Subunits Coordinate Protein Targeting and mRNA Quality Control.
  27. Analysis of copper toxicity by western blotting of secretory proteins translocated via the Sec61 channel.
  28. Small GTPase RAN-driven PNET2 oligomerization and phase separation at the nuclear lamina promote nuclear envelope integrity in plants.
  29. Structural rearrangements underlying the activation of STIM1 by ER calcium depletion.
  30. Codon optimality modulates cellular stress and innate immune responses triggered by exogenous RNAs.
  31. Disease-specific tau polymorphs are associated with unique protein networks across proteinopathies.
  32. Competing Molecular Interactions Govern the Dynamical Arrest of G3BP1 Condensates.
  33. Loss of BECN1 does not compromise autophagosome formation but alters cargo selection and HLA-restricted antigen presentation of B cells.
  34. The methyltransferase Rrp8 is a positive regulator of autophagy flux in Saccharomyces cerevisiae.
  35. Molecular mechanisms of protein disulfide isomerase antagonism by punicalagin.
  36. Iron-endoplasmic reticulum-extracellular matrix axis regulates cancer cell invasion.
  37. A proximity biotinylation approach for the identification of membrane contact site proteins in Toxoplasma gondii.
  38. PANCS-Inhibitors: A rapid method to directly select for protein-protein interaction inhibitors.
  39. Transcriptome-wide splicing analysis reveals HuR-regulated post-transcriptional programs in osteocytes.
  40. Col4a1 mutation in a mouse model of cerebral small vessel disease causes white matter defects that can be modulated by targeting protein folding.
  41. Deubiquitylase OTUD3 regulates integrated stress response to suppress progression and sorafenib resistance of liver cancer.
  42. A conserved vesicular circuit enables metabolic adaptation and maintains systemic heme homeostasis.
  43. TOLLIP suppresses picornavirus internalization by targeting and degrading VP1 protein through selective autophagy.
  44. Photoreactive crosslinkers capture the ubiquitin-E2-E3 ligases intermediates.
  45. Calcineurin B-mediated Ca 2+ sensing translates stress signal intensity into the assembly of phase-separated condensates at PERK complexes.
  46. Architecture and mechanism of the human p24 cargo receptor.
  47. Phase Separation of TRIM21 Modulates PTPN14 Stability to Drive Flow-Dependent Endothelial Activation and Atherogenesis.
  48. Excess intracellular lactate represses mRNA translation and tumor progression through lactate-tRNA charging.
  49. Surface proteomics of cancer-derived extracellular vesicles reveals universal heat shock proteins and cancer signatures.
  50. Stress-induced secretory pathway disruption causes atypical metalloproteinase transport in PMM2-CDG.
  51. Targeted Degradation of KAT6A: Expanding the Therapeutic Frontier of Epigenetic Drug Discovery.
  1. J Cell Biol. 2026 Oct 05. pii: e202509100. [Epub ahead of print]225(10):
    The UPR drives gametogenesis via an unexpected response, coordinating translation and ER structure.
    Constantine Bartolutti, Alena Lauren Bishop, Yanzhe Ma, Kelsey Van Dalfsen, Elçin Ünal, Marko Jovanovic, Gloria Ann Brar.
      Stress responses, including the unfolded protein response (UPR), are commonly studied via induction with harsh exogenous stressors, leaving endogenous functions of these pathways less well understood. We found that the endogenous UPR that precedes meiosis in budding yeast is required for gamete production but diverges dramatically from previously defined UPR outputs, with only a few characterized UPR targets induced, and mildly. The role of this UPR can be replaced by increasing ER chaperones, reducing bulk translation, or impairing the machinery for protein translocation into the ER. ER integrity appears compromised in premeiotic cells lacking the UPR, as foci of reticulon proteins are seen and correlate strongly with an inability of cells to enter meiosis. These findings indicate that physiological UPR activation supports proteostasis and normal ER structure, preparing cells for meiotic entry by reducing the load of proteins that enter the ER. Overall, our study reveals surprising features of a physiological UPR induction that enables a cell-fate decision.
    DOI:  https://doi.org/10.1083/jcb.202509100
  2. Proc Natl Acad Sci U S A. 2026 Aug 25. 123(34): e2534903123
    Human Hsp70 paralogs display selective J-domain interactions tuning proteostasis under stress.
    Roni Suhler, Lars J W van Beurden, Merav D Shmueli, Ofrah Faust, Rina Rosenzweig.
      Hsp70 chaperones are central regulators of proteostasis, mediating folding, refolding, degradation, and aggregation-prevention. Their activity is tuned by the diverse J-domain proteins (JDPs), which both recruit client proteins and stimulate Hsp70 ATP hydrolysis via interaction with a conserved J-domain. The cytosol contains four Hsp70 paralogs-the stress-inducible HSPA1A/B and HSPA6, and the constitutively expressed HSPA8. Whether these act redundantly or carry distinct cellular functions has remained unresolved. Here, we systematically map the interactions of cytosolic Hsp70s with broad-specificity JDPs to elucidate how paralog identity shapes cellular function. We found that despite the high conservation of the JDP-Hsp70 interaction sites, the affinities of these interactions and their functionality varied greatly. HSPA8 behaves as a generalist, engaging all JDP classes with comparable affinity, consistent with its housekeeping role. By contrast, HSPA1 preferentially binds canonical Class A and B JDPs, while showing only weak binding to Class B'. Therefore, under stress, HSPA1 pairs only with Class A/B JDPs to support robust protein refolding, while freeing Class B' to suppress protein aggregation in an Hsp70-independent manner. Most unexpectedly, HSPA6, the most stress-inducible paralog, binds selectivity to Class B JDPs, losing interactions with both Class A and B'. Thus, under severe stress, HSPA6 works exclusively with Class B JDPs to ensure ATP-dependent protein repair, while freeing Class A and B' JDPs to act independently of Hsp70 to protect damaged/misfolded proteins. Our findings reveal an evolved hierarchy of paralog-specific JDP couplings that dynamically rewires the Hsp70 network from active repair to protection during stress.
    Keywords:  Hsp70 and J-domain proteins; NMR; molecular chaperones; protein folding and aggregation; protein homeostasis
    DOI:  https://doi.org/10.1073/pnas.2534903123
  3. bioRxiv. 2026 Aug 07. pii: 2026.08.06.742952. [Epub ahead of print]
    BAG6-RNF115 Couples Protein Quality Control with Ribosome Assembly.
    Anat Raiff, Colin Zenge, Alban Ordureau, Itay Koren.
      Protein homeostasis relies on protein quality control (PQC) pathways that survey the proteome to eliminate aberrant polypeptides. The BAG6 complex is a central PQC factor that recognizes exposed hydrophobic regions, a feature commonly associated with misfolded, mislocalized, and mistranslated proteins. Whether this surveillance machinery also regulates intact, functional proteins as part of physiological proteostasis has remained unclear. Using unbiased quantitative proteomics, we identify the ribosomal protein RPL22L1 as an endogenous BAG6 substrate whose abundance is controlled by continuous proteasomal degradation. This turnover requires the RNF115 E3 ligase activity but not the canonical BAG6 partner RNF126, defining RPL22L1 as a selective RNF115-dependent substrate. Mechanistically, we map a bipartite hydrophobic degron that distinguishes RPL22L1 from its stable paralog RPL22, and show that BAG6-RNF115-mediated degradation is governed by substrate assembly state. Accordingly, RPL22L1 is protected from degradation upon incorporation into the 60S ribosome, where it substitutes for RPL22. When RPL22 is lost, either genetically or through recurrent inactivating mutations in microsatellite-unstable cancers, the vacant ribosomal binding site permits RPL22L1 incorporation, protecting it from BAG6-mediated degradation. These findings establish unassembly-coupled degradation as a mechanism by which BAG6 regulates the abundance of functional protein components, ensuring that they accumulate only when incorporated into their native macromolecular complexes.
    DOI:  https://doi.org/10.64898/2026.08.06.742952
  4. J Am Chem Soc. 2026 Aug 19. 148(32): 34311-34319
    De Novo-Designed Bifunctional Proteins for Targeted Protein Degradation.
    Bram Mylemans, Boguslawa Korona, Amanda M Acevedo-Jake, Ailsa MacRae, Thomas A Edwards, Danny T Huang, Andrew J Wilson, Laura S Itzhaki, Derek N Woolfson.
      In targeted protein degradation (TPD), specific subcellular proteins are removed by routing them to the ubiquitin-proteasome, autophagy, or lysosome machinery. For instance, proteolysis-targeting chimeras (PROTACs) are synthetic heterobifunctional small molecules that simultaneously bind the target and an E3 ubiquitin ligase to drive ubiquitination and degradation by the proteasome. Despite considerable success, designing such molecules is challenging, and the number of currently addressable ubiquitin E3 ligases is limited. Here, we design a heterobifunctional de novo protein to trigger the degradation of a common cancer target, resulting in a desired phenotypic output. First, we developed a highly stable and adaptable helix-turn-helix scaffold for presenting multiple binding sites. Next, we use computational protein design to incorporate and embellish hot-spot-binding sites to target the antiapoptotic mediators BCL-xL and MCL-1. We show a 75% success rate for creating submicromolar binders against these targets. Crystal structures of the complexes confirmed the designed binding poses. Then, we designed short linear motifs (SLiMs) into the loop of the scaffold to recruit KLHL20 and the ubiquitin ligase machinery. These designs have low micromolar affinity for KLHL20 comparable to that of the natural SLiMs. Moreover, the bifunctionalized proteins degrade BCL-xL in cells, leading to apoptosis.
    DOI:  https://doi.org/10.1021/jacs.6c07593
  5. EMBO J. 2026 Aug 17.
    Reconstitution of glycan-driven MHC-I recycling reveals calreticulin as mediator between TAPBPR and tapasin.
    Tim J Heinke, Amin Fahim, Niko Popovic, Tobias Rath, Ines K Müller, Nina Morgner, Simon Trowitzsch, Robert Tampé.
      Protein folding in the endoplasmic reticulum (ER) relies on N-linked glycosylation and glycan remodeling to guide quality control. Major histocompatibility complex class I (MHC-I) molecules, essential for adaptive immunity, undergo a specialized maturation pathway involving the peptide-loading complex (PLC), the editor TAPBPR, the UDP-glucose:glycoprotein glucosyltransferase, and the lectin chaperone calreticulin. However, how glycan-dependent mechanisms coordinate MHC-I transfer between these factors has remained unclear. Using a fully reconstituted system, we show that retrograde transfer of peptide-receptive MHC-I from TAPBPR to tapasin requires calreticulin recognition of monoglucosylated MHC-I glycans. While calreticulin's C-terminal acidic helix is dispensable for releasing reglucosylated MHC-I from TAPBPR, it is essential for productive docking of MHC-I onto tapasin. These findings reveal a glycan-surveillance mechanism that enables retrieval of suboptimally loaded MHC-I molecules missed by the initial quality control at the PLC. Our work defines a glycan-dependent chaperone network, finely tuned by a combination of low-micromolar interactions between the constituents, that ensures efficient MHC-I maturation and illustrates fundamental principles of ER protein quality control.
    DOI:  https://doi.org/10.1038/s44318-026-00873-2
  6. Redox Biol. 2026 Aug 07. pii: S2213-2317(26)00335-6. [Epub ahead of print]96 104336
    Selenoprotein S associates with complexes governing membrane protein biogenesis and translation-associated processes.
    Farid Ghelichkhani, Mariia A Kapitonova, Atinuke Odunsi, Erfan Rahmani, Omar G Rosas Bringas, Mohammed Hanzala Kaniyar, John LaCava, Sharon Rozovsky.
      Human selenoprotein S (selenos) is part of the integrated cellular stress response and linked to protein quality control and signaling pathways. Consequently, genetic polymorphisms of selenos are associated with increased risks for diabetes, dyslipidemia, and cardiovascular diseases. Determining the specific roles of selenos in these cellular pathways and diseases has been challenging, as selenos associates with a wide range of protein complexes. Thus, to map the cellular functions of selenos and uncover their interconnections, we used affinity purification and in vivo crosslinking to stabilize transient protein interactions, followed by proteomics to record the resulting selenos interactome. Through mapping of selenos protein partners, we found evidence that selenos associates with complexes responsible for the insertion of membrane proteins into the endoplasmic reticulum (ER) bilayer and their connected quality control components. Furthermore, selenos is also part of metabolic, trafficking, and mitochondrial pathways. Notably, proteins involved in translation preferentially associate with selenos when its C-terminal intrinsically disordered segment containing the redox-active motif is accessible. Together, these results identify the C-terminal redox loop of selenos as a central interaction hub connecting translation with ER membrane protein biogenesis and quality control.
    Keywords:  Derlin; EMC; Selenoprotein S; Selenos; VCP; VIMP
    DOI:  https://doi.org/10.1016/j.redox.2026.104336
  7. iScience. 2026 Aug 21. 29(8): 116866
    Astrocytes mobilize a broader repertoire of lysosomal repair mechanisms than neurons.
    Erin M Smith, Natali L Chanaday, Sandra Maday.
      Lysosomal damage impairs proteostasis and contributes to neurodegenerative diseases, yet cell-type-specific differences in lysosomal repair remain unclear. Using a neuron-astrocyte coculture system, we compared responses to lysosomal injury induced by a lysosomotropic methyl ester. Both neurons and astrocytes showed lysosomal damage, marked by Galectin-3 recruitment to lumenal lysosomal β-galactosides, disrupted lysosomal pH, and engagement of lysophagy receptors TAX1BP1 and p62. However, astrocytes showed a preferential recruitment of ESCRT (endosomal sorting complex required for transport) repair machinery to damaged lysosomes. Additionally, the lysosomal membrane reformation pathway regulated by the RAB7-GTPase-activating protein (GAP), TBC1D15, was more robustly activated in astrocytes. By contrast, the phosphoinositide-initiated membrane tethering and lipid transport (PITT) pathway, mediating lipid transfer between the endoplasmic reticulum (ER) and damaged lysosomes, was engaged in both cell types. Our data reveal a divergence in how neurons and astrocytes mobilize repair pathways to manage lysosomal damage. These data may reflect differences in lysosomal resilience between astrocytes and neurons and inform therapeutic strategies to correct lysosomal dysfunction in neurodegenerative diseases.
    Keywords:  ESCRT; LLOMe; ORP9; PI4K2A; TBC1D15; astrocyte; lysosomal damage; neuron
    DOI:  https://doi.org/10.1016/j.isci.2026.116866
  8. PLoS Pathog. 2026 Aug 18. 22(8): e1014518
    Endoplasmic reticulum engagement of transcripts triggers VSG mRNA balancing in trypanosomes.
    Erick O Aroko, Elisabeth Meyer-Natus, Christopher Batram, Kathrin Weißenberg, Jasmin Henning, Majeed Bakari-Soale, Nicola G Jones, Markus Engstler.
      The cell surface of bloodstream-form African trypanosomes is covered by a dense coat of variant surface glycoproteins (VSGs). By periodically switching the expressed VSG antigen, parasites evade host immune responses. VSG mRNA constitutes ~10% of total cellular mRNA, and depletion of VSG transcripts is lethal. When two VSGs are expressed simultaneously, however, total VSG mRNA levels remain close to wild-type amounts, indicating the presence of a balancing mechanism that limits the overall abundance of these highly expressed transcripts. Using inducible and constitutive expression systems, we found that attenuation of endogenous VSG mRNA requires efficient engagement of ectopic transcripts with the endoplasmic reticulum (ER). This response occurs independently of efficient VSG protein production and does not require a VSG open reading frame. In contrast, abundant transcripts lacking functional ER-engagement signals fail to trigger balancing despite containing the VSG 3' UTR 16-mer stability element. These results indicate that the signal for VSG mRNA regulation is the presence of abundant ER-engaged transcripts (i.e., transcripts undergoing co-translational targeting to the ER), rather than VSG-specific sequence features or productive protein synthesis. Based on our findings together with previous work, we propose an ER-engagement-coupled homeostatic attenuation mechanism in which increased ER-engaged transcript load elicits a regulatory response that reduces endogenous VSG mRNA abundance. This model links the cytoplasmic burden of ER-targeted transcripts to transcriptional control of the nuclear VSG expression site, providing a mechanism that could allow trypanosomes to couple secretory pathway capacity to surface antigen expression. Such a mechanism could enable trypanosomes to maintain secretory pathway homeostasis while supporting rapid surface-coat remodelling during antigenic variation.
    DOI:  https://doi.org/10.1371/journal.ppat.1014518
  9. bioRxiv. 2026 Jul 28. pii: 2026.07.27.741020. [Epub ahead of print]
    Astrocytes instructively regulate neuronal translation.
    Wendy J Liu, Carson C Schultz, Eshaan A Khan, Mauricio M Oliveira, Srinidhi V Kalavai, Charles J Sheehan, Xunzhao Zhang, Richard Sam, Eric Klann.
      Neuronal protein synthesis is essential for synaptic plasticity and long-term memory, yet whether its regulation is shaped by other cell types remains poorly understood. Here, we show that astrocyte-secreted proteins regulate global neuronal translation depending on astrocytic state. Astrocyte-conditioned medium (ACM) increased neuronal translation under basal conditions, an effect enhanced by astrocyte stimulation with the activity-dependent factor BDNF, whereas ACM from neurotoxic reactive astrocytes, a state linked to neuroinflammation and Alzheimer's disease, suppressed neuronal translation. Across these conditions, neuronal mTORC1 activity consistently tracked with translational output, whereas the integrated stress response (ISR) acted through distinct, state-specific mechanisms that did not always track with neuronal translation. Furthermore, we identified astrocyte-secreted apolipoprotein E (APOE) and its associated cargo as a negative regulator of neuronal translation that contributed to the decreased translation induced by neurotoxic reactive astrocytes. We also found that astrocyte-secreted signals required neuronal endocytosis to influence translation and drove synaptic remodeling dependent on glutamatergic signaling and neuronal mTORC1 activity. Together, these findings identify astrocytes as active, instructive regulators of neuronal translation and synaptic structure, with implications for understanding how astrocyte dysfunction may disrupt the translational mechanisms underlying impairments in synaptic plasticity and long-term memory in neurodegenerative disease.
    GRAPHICAL ABSTRACT:
    DOI:  https://doi.org/10.64898/2026.07.27.741020
  10. EMBO J. 2026 Aug 15.
    FAF1 and FAF2 enhance unfolding by p97-UFD1-NPL4 complex enabling rational design of p97 activators.
    Pritha Dasgupta, Ian R Kelsall, Gaurav Anand, Anna Pérez-Ràfols, Thomas A Jowitt, Axel Knebel, Robert Gourlay, Glenn R Masson, Yogesh Kulathu.
      VCP/p97 is an AAA+ ATPase that, together with its cofactors UFD1-NPL4 (p97-UN), unfolds ubiquitylated substrates to maintain cellular homeostasis. The human p97-UN complex associates with additional cofactors, but how these cofactors modulate p97-UN activity is not fully understood. Here, we screen cofactors and identify FAF2 to potently enhance substrate unfolding by p97-UN. Using biochemical and structural approaches, we show how FAF2 engages p97-UN and polyubiquitin to promote unfolding. We define a conserved activation motif in FAF2 that contacts both UFD1 and the ubiquitin proximal to the initiator, thereby stabilizing and supporting the unfolding of the initiator ubiquitin in a UFD1-dependent manner. We leverage the features of the FAF2 activation motif to engineer de novo proteins that potently enhance unfolding, providing a rational strategy to boost p97 activity. Our findings reveal how cofactors can provide additional adaptive control, fine-tuning human p97 activity to unfold challenging substrates and those modified with short ubiquitin chains.
    DOI:  https://doi.org/10.1038/s44318-026-00894-x
  11. Mol Cell. 2026 Aug 17. pii: S1097-2765(26)00513-7. [Epub ahead of print]
    ER-liquid condensate contacts sequester FAM134B/C and RhoA to govern cell morphology.
    Natalia Jimenez-Moreno, Arianna Karageorgiou, Katie Winnington-Ingram, Jimi Wills, Chinmayi Pednekar, Melina Overhoff, Katherine Paine, Matthew Pearson, Lukas Gerasimavicius, Ann Wheeler, Natalia L Kononenko, Joseph A Marsh, Paul Verkade, Alex von Kriegsheim, Jon D Lane, Simon Wilkinson.
      Endoplasmic reticulum (ER)-phagy receptors have elusive physiological functions beyond ER remodeling. To gain insight into these, we screen for cytoplasmic interactors of well-known receptors in mouse pancreatic ductal adenocarcinoma cells. Our data subsequently reveal a non-canonical action of PRKAR1A, a subunit of protein kinase A (PKA), in binding and activating FAM134B/C. At a molecular level, an amphipathic helix in the otherwise-disordered C terminus of FAM134B directly binds PRKAR1A dimers. Multimodal imaging reveals that this interaction occurs at interorganellar contact sites between the ER and liquid-like condensates of PRKAR1A. Mechanistically, ER-condensate contacts promote clustering of FAM134B/C with LC3B and recruit lysosomes, forming degradation hubs. Unexpectedly, these structures are found to control cytoplasmic signaling through FAM134B/C-mediated lysosomal sequestration of RhoA. Underscoring the physiologic relevance of this, these cytoplasmic-facing actions of FAM134B/C result in RhoA-dependent cell morphologic and migratory control in response to cyclic AMP (cAMP) stimuli. Thus, interorganellar contacts expand the roles of FAM134B/C ER-phagy receptors beyond ER degradation.
    Keywords:  ER-phagy; LLPS; RHO GTPase; autophagy; cAMP; cancer; cell invasion; cytoskeleton; liquid-like condensate; lysosome
    DOI:  https://doi.org/10.1016/j.molcel.2026.07.027
  12. Adv Sci (Weinh). 2026 Aug 19. e77201
    A New Recruitable E3 Ligase UHRF1 Supporting Targeted Protein Degradation: A Minimal Azide as a Recruitment Ligand.
    Zehong Lin, Kang Duan, Rui Wan, Tao Jiang, Jinzhe Wei, Miao Sun, Mariia A Skryl'nikova, Maxim A Gureev, Zhi-Min Zhang, Zhang Zhang, Ke Ding, Nan Ma, Tongzheng Liu, Yi Tan, Zhengqiu Li.
      Targeted protein degradation represents a promising therapeutic strategy, yet its broader application is often limited by the scarcity of usable E3 ligases. Glutathione peroxidase 4 (GPX4) is a key target for inducing ferroptosis, but achieving sustained and potent inhibition remains challenging with conventional enzymatic inhibitors. Herein, we report the first small-molecule GPX4 degraders that incorporate either electrophilic warheads or a minimal azide group as an E3 recruitment ligand. The azide-based degrader DK-5070 effectively drives potent GPX4 degradation, achieving a DC50 of 17.4 nM and a Dmax of 84%, thereby outperforming larger PROTAC-based degraders. Notably, DK‑5070 exhibits potent antitumor activity both in vitro (IC50 = 47.21 nM) and in vivo (TGI = 41.8%), demonstrating significant efficacy as a GPX4 degrader. Mechanistic studies reveal that degradation is mediated through recruitment of the oncogenic E3 ligase UHRF1, which is frequently overexpressed in tumors, underscoring the potential for tumor-specific protein degradation. This demonstrated small-molecule degraders that recruit UHRF1 to facilitate targeted degradation of GPX4. In this system, the azide group functions as a minimal recruitment ligand, thereby expanding the E3 ligase toolbox and offering a promising strategy for targeted cancer therapy.
    Keywords:  azide; cell biology; chemistry; gpx4; in vivo; ligand; protein degradation; targeted therapy; therapeutic strategy; ubiquitin ligase
    DOI:  https://doi.org/10.1002/advs.77201
  13. Cell Rep. 2026 Aug 18. pii: S2211-1247(26)00813-2. [Epub ahead of print]45(8): 117735
    PKA-dependent phosphorylation drives stress granule maturation and controls recovery from prolonged starvation.
    Sonja Kroschwald, Caroline Wilson-Zbinden, Federico Uliana, Anastasia Timofiiva, Jiangtao Zhou, Sung Sik Lee, Ludovic Gillet, Martina Zanella, Alaa Othman, Raffaele Mezzenga, Matthias Peter.
      During environmental stress, cells form dynamic biomolecular condensates called stress granules (SGs) that can undergo a maturation process towards more solid-like material states. Deposition of SG proteins in insoluble aggregates is a hallmark of neurodegenerative pathologies, provoking inquiry into the pathological link and mechanisms underlying SG maturation. Here we show that yeast SGs mature into a solid-like state during long-term stationary phase driven by protein kinase A (PKA)-dependent phosphorylation of the SG proteome. Catalytic PKA subunits condense in SGs upon stationary phase where SG-localized PKA activity is maintained. PKA phosphorylates key SG components, including the pyruvate kinase Cdc19, which is necessary and sufficient for Cdc19 maturation into amyloid-like structures. Inhibiting PKA during long-term stationary phase prevents SG maturation, which alters metabolism and delays ordered re-start of cell growth after re-feeding. These results describe a SG maturation mechanism selectively activated during chronic stress that preserves SG integrity and promotes cell survival.
    Keywords:  CP: cell biology; PKA; chronic stress; phase separation; phase transition; phosphorylation; protein kinase A; reversible amyloids; starvation; stress granule maturation; stress recovery; stress response
    DOI:  https://doi.org/10.1016/j.celrep.2026.117735
  14. bioRxiv. 2026 Aug 06. pii: 2026.08.05.741138. [Epub ahead of print]
    A longevity-associated ubiquitin E3 ligase UBE3C governs lamin B1 homeostasis through selective autophagy and delay senescence.
    Di Guan, Seungsoo Kim, Karim Omar, Ying Hao, Gordon Huang, HyeRim Han, Jiping Yang, Naftali Horwitz, Phuong-Anh Dinh, Jihyun Hwang, Haiyuan Yu, Yousin Suh.
      Nuclear lamina integrity is fundamental to cellular homeostasis across the lifespan 1 , and its progressive deterioration is closely linked to human aging 2 . Yet, the regulatory mechanism that govern this decline and how they might be counteracted in long-lived individuals remain poorly defined. Here, by combining whole-exome sequencing of Ashkenazi Jewish centenarians with GTEx transcriptomes, we identify ubiquitin E3 ligase UBE3C strongly associated with exceptional longevity and progressively declines with age across human tissues. UBE3C knockdown triggers premature senescence and destabilizes key nuclear lamina components Lamin B1 (LMNB1) and Lamin B receptor (LBR), while the longevity-associated UBE3C variant delays senescence and preserves LMNB1/LBR expression. Mechanistically, UBE3C interacts directly with LMNB1/LBR and modulates their abundance via selective autophagy. Notably, we uncover the ER- resident autophagy trigger CKAP4 3 bridges UBE3C and LMNB1. UBE3C loss enhances LMNB1-CKAP4 binding, linking nuclear lamina turnover to autophagy. Together, our findings establish UBE3C as a central guardian of nuclear lamina maintenance during senescence and offering novel insights into interventions against age-related nuclear lamina deterioration.
    DOI:  https://doi.org/10.64898/2026.08.05.741138
  15. bioRxiv. 2026 Aug 04. pii: 2026.08.01.741249. [Epub ahead of print]
    EOLA1 functions in nucleotide salvage through deacetylating free N4-acetylcytidine.
    Sebastien Relier, Sarah Schiffers, Hamid Beiki, Maria Prigge, Nishu Tyagi, Cyrinne Achour, Ayush Raman, A Maxwell Burroughs, L Aravind, Shalini Oberdoerffer.
      RNA-based medicines rely on modified nucleotides to promote immune evasion and in vivo efficacy. Nucleotides generated from RNA degradation are either exported or recycled through metabolically favorable salvage pathways, though whether modified nucleotides are efficiently recycled remains unclear. N 4 -acetylcytidine (ac⁴C) is a naturally occurring modification in rRNA and tRNA that has shown promise in therapeutic mRNA applications. However, N 4 -acetylation impairs cytidine deamination, the first step in cytidine salvage. Here, we investigate the endogenous mechanisms that enable ac⁴C metabolism. Through sensitive sequence and structural analyses, we identify the uncharacterized human ASCH domain protein EOLA1 as a key ac⁴C deacetylase in nucleotide salvage. EOLA1 inactivation leads to free intracellular ac⁴C accumulation and increased cytotoxicity upon nucleotide export inhibition. While steady-state ac⁴C levels in cellular RNAs remain unchanged, EOLA1-dependent regulation of free ac⁴C is evident basally and is exacerbated by exogenous mRNA delivery. Proteomic analyses place EOLA1 in proximity to ribosomal proteins, adjacent to endogenous ac⁴C sources. In vitro assays confirm EOLA1 specificity for ac⁴C, and structural analysis reveals a narrow nucleotide-binding pocket consistent with mononucleotide selectivity. These findings identify EOLA1 as a bona fide ac⁴C eraser and uncover a previously unrecognized pathway for recycling modified nucleotides with relevance to therapeutic RNA design.
    DOI:  https://doi.org/10.64898/2026.08.01.741249
  16. Sci Adv. 2026 Aug 21. 12(34): eaed6318
    Mitochondrial profiling across macrophage states reveals inhibition of IL-4/IL-13 reprogramming by the integrated stress response.
    Joan Blanco-Fernandez, Miriam Lisci, Mads M Foged, Chloé Chapuis, Tim Pflästerer, Tatjana Kleele, Alexis A Jourdain.
      Mitochondria drive cellular reprogramming by integrating metabolism and signaling. In macrophages, mitochondria are central to immunometabolic responses to external cues, but the extent to which they are remodeled and participate in macrophage reprogramming remains unclear. Here, we integrate transcriptomics with whole-cell and purified mitochondrial proteomics to profile lipopolysaccharide (LPS)/interferon-γ (IFN-γ)- and interleukin-4 (IL-4)/IL-13-stimulated macrophages. We reveal a notable disconnect between mitochondrial transcript and protein levels following either stimulus and a signal transducer and activator of transcription 6 (STAT6)-dependent increase in mitochondrial DNA (mtDNA) expression and intramitochondrial translation in IL-4/IL-13 macrophages. We demonstrate that pharmacological inhibition of mitochondrial translation or individual respiratory chain complexes variably impairs reprogramming, whereas ATP synthase inhibition uniquely triggers a heme-regulated inhibitor (HRI)-dependent integrated stress response (ISR) through mitochondrial hyperpolarization, thereby preventing IL-4/IL-13 reprogramming. Mechanistically, we show that restoring mitochondrial membrane potential or inhibiting the ISR rescues IL-4/IL-13-mediated reprogramming. Together, we identify mtDNA expression, intramitochondrial translation, and mitochondrial membrane potential as critical, drug-sensitive determinants of the IL-4/IL-13 response.
    DOI:  https://doi.org/10.1126/sciadv.aed6318
  17. bioRxiv. 2026 Aug 03. pii: 2026.07.31.742138. [Epub ahead of print]
    FAF1 cofactor enhances UFD1/NPL4-p97 unfolding efficiency across ubiquitin chain lengths independent of SUMO2.
    Abigail A Lemmon, Christopher D Lima.
      The AAA+ protein p97/VCP and its cofactor UFD1/NPL4 interact with and unfold ubiquitinated proteins to promote disaggregation and unfolding for recycling or to prepare substrates for proteasomal degradation. The cofactor Fas-associated factor 1 (FAF1) is suggested to reduce the length of ubiquitin chain required for substrate unfolding by UFD1/NPL4-p97 and to interact with SUMO. Here, we employ in vitro reconstitution of UFD1/NPL4-p97 and FAF1/UFD1/NPL4-p97 complexes and fluorescent substrates modified with SUMO2-polyubiquitin hybrid or polyubiquitin-only chains of varying lengths to assess initial rates of unfolding. These assays reveal that FAF1 enhances initial rates of unfolding relative to UFD1/NPL4-p97 in a manner that is independent of SUMO2 and semi-dependent on ubiquitin chain length. Unlike preferences observed for yeast Ufd1/Npl4-Cdc48, these data suggest that the FAF1 cofactor does not contribute to preferential unfolding of the SUMO2-polyubiquitin substrates tested. Further dissection of FAF1 reveals that it significantly increases the rate of unfolding for all ubiquitin chain lengths tested with its greatest differential impact observed when unfolding chains with four to ten ubiquitin molecules that are considered physiologically relevant. Using cryoEM we resolve a series of reconstructions that reveal FAF1/UFD1/NPL4-p97 bound to substrate in non-translocating and translocating states. Observed interactions between a helix of FAF1 and UFD1 throughout the unfolding process are consistent with AlphaFold models and recent reports suggesting that FAF1 may stabilize interactions between UFD1, NPL4, and p97 to promote substrate engagement and unfolding.
    DOI:  https://doi.org/10.64898/2026.07.31.742138
  18. J Cell Sci. 2026 Aug 21. pii: jcs.264801. [Epub ahead of print]
    Chronic stress disrupts the network among stress granules, P-bodies, and motor proteins.
    Yuichiro Adachi, Masashi Masuda, Yutaka Taketani, Paul J Anderson, Pavel V Ivanov.
      Stress granules (SGs) are dynamic RNA condensates that assemble rapidly in response to cellular stress following translational arrest, thereby promoting adaptation and influencing disease pathogenesis. Although SG assembly and disassembly during acute stress have been extensively characterized, their regulation under chronic stress remains poorly understood. We previously showed that chronic stress preconditioning suppresses the earliest stages of SG assembly through translation-dependent mechanisms. Whether chronic stress also impairs subsequent SG maturation has remained unknown. Here, we demonstrate that chronic stress limits SG maturation by disrupting the MYH9-dependent interaction network centered on the core SG nucleator G3BP1. Loss of this interaction reduces SG size and impairs docking between SGs and processing bodies (PBs), thereby restricting the maturation of nascent SGs. In parallel, chronic stress decreases expression of the SG nucleator UBAP2L, an essential regulator of SG-PB docking, further exacerbating these defects. Together, our findings identify chronic stress as a regulator of the MYH9-G3BP1-UBAP2L axis and reveal that chronic stress inhibits SG maturation through translation-independent mechanisms.
    Keywords:  G3BPs; MYH9; P-bodies; Stress granules; UBAP2L
    DOI:  https://doi.org/10.1242/jcs.264801
  19. Proc Natl Acad Sci U S A. 2026 Aug 25. 123(34): e2605305123
    An ER retention motif controls the heteromeric stoichiometry of hERG1a/1b channels.
    Sudharsan Kannan, Liliana R Ernandez, Gail A Robertson.
      The human ether-à-go-go related gene (hERG) encodes a potassium channel essential for cardiac repolarization and neuronal excitability. In the heart, heteromeric assemblies of hERG1a and hERG1b subunits produce cardiac IKr, and mutations in either subunit are associated with long QT syndrome. Although hERG1a and 1b contain identical transmembrane and C-terminal cytosolic domains, they differ in N-terminal cytosolic domains, with hERG1b harboring an arginine-based endoplasmic reticulum (ER) retention/retrieval motif that limits its surface expression in the absence of hERG1a. While the association of hERG1a and 1b subunits is known to influence channel function, the stoichiometry of heteromeric hERG channels and mechanisms that regulate it have remained unresolved. Here, using single molecule photobleaching step analysis in HeLa cells, we show that heteromeric hERG1a/1b channels assemble predominantly with a fixed 2:2 stoichiometry. Mutation of the hERG1b ER retention motif disrupts this bias, resulting in a broader, near-random distribution of subunit compositions. Independent functional assays using dominant-negative pore mutants in Xenopus oocytes yielded quantitative current suppression consistent with a 2:2 assembly and similarly revealed loss of stoichiometric bias upon RXR mutation. Together, these results establish the oligomeric composition of hERG1a/1b channels and identify ER retention as a previously unrecognized determinant of heteromeric stoichiometry.
    Keywords:  arrhythmia; endoplasmic reticulum; hERG; protein trafficking; stoichiometry
    DOI:  https://doi.org/10.1073/pnas.2605305123
  20. RNA. 2026 Aug 17. pii: rna.081039.126. [Epub ahead of print]
    Yeast Rai1p and the RAC complex (Ssz1p/Zuo1p) modulate uORF-mediated GCN4 translational control under stress conditions.
    Kristina Jendruchova, Adriana Subrtova, Leos Shivaya Valasek.
      Translation reinitiation (REI) is one of the most important gene-specific regulatory mechanisms by which eukaryotic cells influence expression of main translons, for example during highly conserved integrated stress response (ISR). In S. cerevisiae, expression of the key stress response gene, GCN4, is controlled by an intricate interplay among four short upstream translons (uTranslons, formerly uORFs), resulting in high or low levels of REI at GCN4 depending on the growth conditions. Under nutrient rich conditions, GCN4 expression is repressed, but upon amino acid starvation, it is derepressed, despite of a general translational shut down. Capitalizing on our screening reporter system, we identified three new factors influencing efficiency of REI after translation of GCN4 uTranslons: Rai1p (an RNA quality control and processing factor), and Ssz1p and Zuo1p (members of the Ribosome Associated Complex [RAC]). Importantly, we showed that depletion of these factors deregulated derepression of Gcn4p synthesis under starvation. Furthermore, we found that similar to RAC, Rai1p associates with 40S subunits and actively translating ribosomes. We also explored interactomes of these proteins. Collectively, we present three previously unknown factors that co-regulate stress response to amino acid starvation in the budding yeast by unique mechanisms.
    Keywords:  GCN4; RAI1; SSZ1 and ZUO1 (RAC complex); ribosome recycling; translation reinitiation
    DOI:  https://doi.org/10.1261/rna.081039.126
  21. Autophagy. 2026 Aug 21. 1-10
    Dealing with membrane stress through CASM.
    Håvard Styrkestad Haukaas, Sven R Carlsson, Alf Håkon Lystad.
      Compartments of the endolysosomal and secretory pathways encounter diverse insults - from loss of ion gradients and osmotic imbalance to physical membrane disruption - creating a need for rapid, localized surveillance and response systems. Recent work demonstrates that conjugation of ATG8 to single membranes (CASM) provides such specificity via stress-responsive targeting mechanisms that recruit the ATG8 conjugation machinery through pathways distinct from macroautophagy/autophagy. Here, we review recent advances in how membrane stress is sensed, coupled to CASM initiation, and converted into downstream cellular responses.
    Keywords:  ATG16L1; ATG16L2; ATG8; STIL; TECPR1; VAIL
    DOI:  https://doi.org/10.1080/15548627.2026.2717033
  22. Nucleic Acids Res. 2026 Aug 10. pii: gkag797. [Epub ahead of print]54(15):
    RPL41 stabilizes ribosome dynamics and supports long-protein homeostasis in mammals.
    Mina Hirata, Maho Fujino, Kazuya Ichihara, Ronghao Tang, Momoko Narita, Wakana Iwasaki, Chisa Shiraishi, Taichi Shiraishi, Atsushi Hatano, Toru Suzuki, Tokichiro Abe, Tomoyo Takami, Emi Mishiro-Sato, Atsushi Toyoda, Masaki Matsumoto, Yoshikazu Tanaka, Takuhiro Ito, Toshifumi Inada, Keiichi I Nakayama, Takeshi Yokoyama, Akinobu Matsumoto.
      Ribosomal protein L41 (RPL41 or eL41) is the smallest ribosomal protein and forms the eukaryote-specific bridge, eB14, near the decoding center; however, its role in mammalian translation remains unclear. In this study, we established RPL41-deficient models of human HEK293T cells and mice to define its function. Cryo-electron microscopy revealed that RPL41 constrains intersubunit conformational dynamics without inducing major local static rearrangements. Loss of RPL41 altered A-site dynamics, slowed elongation, modestly increased amino acid misincorporation, and modestly enhanced readthrough of collision-inducing reporter sequences. Quantitative proteomic analysis suggested that these translational defects compromise long-protein homeostasis, as evidenced by increased insolubility and reduced abundance of long proteins. In vivo, Rpl41-/- mice were viable but exhibited growth retardation and decreased abundance of long proteins in tissues. Our findings reveal a conserved role for RPL41 in maintaining ribosome dynamics and translational fidelity, indicating that RPL41 supports ribosome function and long-protein homeostasis in mammals.
    DOI:  https://doi.org/10.1093/nar/gkag797
  23. bioRxiv. 2026 Jul 29. pii: 2026.07.28.741261. [Epub ahead of print]
    ESCAPE: assigning site-specific activity to covalent ligands in cells by prime editing.
    Jason E Tse, William R Brothers, Rachel E Hayward, Sabrina Barbas, Kai Sheng, Kathryn R Spencer, Kristen E DeMeester, Evert Njomen, James R Williamson, David R Liu, Bruno Melillo, Gene W Yeo, Benjamin F Cravatt, Haoxin Li.
      Chemical proteomics has identified covalent ligands targeting cysteine residues across many hundreds of human proteins. The functional effects of these liganding events, however, remain challenging to assign at scale. Here we describe ESCAPE (Endogenous Site-specific Competition Assays using Prime Editors), a platform for the site-resolved functional analysis of covalent ligands in cells. In this method, cysteine-to-serine substitutions are generated by prime editing to abrogate covalent ligand-protein interactions, and the impact of these edits on ligand-induced cellular phenotypes is quantified through allele frequency-based resistance scores. Applied to ligandable cysteines mapped by activity-based protein profiling in 50+ proteins, ESCAPE identified multiple covalent ligand-protein interactions that impair cancer cell growth, including azetidine butynamides that target a non-orthosteric cysteine in the RNA helicase DDX49 to disrupt 18S rRNA processing, 40S ribosome assembly, and protein synthesis. ESCAPE thus provides a scalable framework for the functional characterization of covalent ligands targeting structurally and mechanistically diverse proteins.
    DOI:  https://doi.org/10.64898/2026.07.28.741261
  24. Cell Chem Biol. 2026 Aug 20. pii: S2451-9456(26)00284-9. [Epub ahead of print]33(8): 1074-1076
    The degrader gets degraded as cysteine fluctuates.
    Junyi Chen, Haopeng Xiao.
      In a recent issue of Molecular Cell, Ramage et al. identify a Cullin-RING E3 ubiquitin ligase complex defined by the substrate adaptor LRRC58 that inversely regulates the abundance of LRRC58 and its cognate substrate, CDO1, in response to cysteine levels.1.
    DOI:  https://doi.org/10.1016/j.chembiol.2026.07.010
  25. Arterioscler Thromb Vasc Biol. 2026 Aug 20.
    Transmembrane Protein Disulfide Isomerase TMX3 Potentiates Platelet Aggregation and Arterial Thrombosis in Mice.
    Fengwu Chen, Xiaofeng Yan, Yuxin Zhang, Zhenzhen Zhao, Aizhen Yang, Yue Lu, Miao Jiang, Yue Han, Depei Wu, Yi Wu.
       BACKGROUND: Several members of the PDI (protein disulfide isomerase) family have been shown to regulate platelet function and thrombosis. TMX3 (thioredoxin-related transmembrane protein 3), the sole transmembrane member of the PDI family containing catalytically inactive thioredoxin-like domains, has been implicated in thrombotic diseases, but its role in hemostasis and thrombosis remains uninvestigated.
    METHODS: We generated hematopoietic and endothelial cell-specific (Tie2-Cre) and platelet-specific (Pf4-Cre) TMX3 conditional knockout mice to assess the contribution of TMX3 to hemostasis and thrombosis. The functions of TMX3 in platelet aggregation, granule secretion, spreading, clot retraction, and calcium mobilization were examined. Wild-type and catalytically inactive TMX3 proteins were produced to determine the catalytic capacity of TMX3 toward αIIbβ3. Calcium ionophore, eptifibatide, ADP, and apyrase were used to investigate the regulatory mechanisms of TMX3 in platelet granule secretion.
    RESULTS: Expression of TMX3 on the platelet surface increased with convulxin stimulation. TMX3 knockout mice exhibited prolonged tail-bleeding times and decreased platelet accumulation in both FeCl3- and laser-induced thrombosis models. TMX3-deficient platelets had decreased P-selectin expression, αIIbβ3 activation, platelet aggregation, and ATP release, while maintaining normal platelet spreading, clot retraction, granule biogenesis, and Ca2+ mobilization. Mechanistically, TMX3 bound tightly to αIIbβ3 and generated thiols in αIIb and β3, particularly in αIIb. Meanwhile, TMX3 regulated calcium mobilization-triggered platelet dense granule secretion, independent of αIIbβ3-mediated outside-in signaling. The impaired aggregation of TMX3-deficient platelets resulted from defective ADP secretion. Recombinant catalytically inactivated TMX3 protein inhibited platelet aggregation and ATP release and significantly reduced platelet accumulation and fibrin deposition in the laser-induced thrombosis model, whereas recombinant wild-type TMX3 protein neither enhanced human platelet aggregation and ATP release nor rescued the impaired aggregation and ATP release of TMX3-deficient platelets.
    CONCLUSIONS: Our data highlight a critical role for TMX3 in platelet function and thrombus formation by interacting with αIIbβ3 integrin and directly regulating platelet granule secretion.
    Keywords:  apyrase; endothelial cells; fibrin; integrins; thrombosis
    DOI:  https://doi.org/10.1161/ATVBAHA.125.323282
  26. bioRxiv. 2026 Aug 03. pii: 2026.07.31.742061. [Epub ahead of print]
    Human SRP Subunits Coordinate Protein Targeting and mRNA Quality Control.
    Morgana K Kellogg, Elena B Tikhonova, Paul F D'Cunha, Andrey L Karamyshev.
      The Signal Recognition Particle (SRP) targets secretory proteins to the endoplasmic reticulum (ER) for their subsequent transport and protects their mRNAs from degradation by the RAPP pathway. The SRP is an RNA-protein complex consisting of six protein subunits and one noncoding RNA assembled into S- and Alu-domains. However, the distinct roles of individual SRP subunits remain undetermined. Using a molecular dissection of the human SRP through the depletion of specific subunits, we demonstrate that S-domain subunits have a primary role in mRNA protection and protein targeting, while Alu-domain subunits are dispensable. The SRP54 subunit is essential for both processes and ribosome association, while other S-domain subunits (SRP19, SRP68, and SRP72) serve supportive roles. The expression of SRP subunits is dependent on each other within the same domains, suggesting independent domain assembly. These data provide new insights into the multidomain organization and distinct functions of human SRP subunits and establish a framework for understanding the molecular mechanisms of SRP-related human diseases.
    DOI:  https://doi.org/10.64898/2026.07.31.742061
  27. Methods Enzymol. 2026 ;pii: S0076-6879(26)00156-4. [Epub ahead of print]733 253-278
    Analysis of copper toxicity by western blotting of secretory proteins translocated via the Sec61 channel.
    Smriti Anand, Prateeksha Thakur, Vandana Anjana, Santoshi Acharjee, Rajshree Pal, Sugandh Sharma, Raghuvir Singh Tomar.
      As a trace element, copper plays a vital role in regulating cellular homeostasis by serving as a cofactor for various enzymes. Therefore, maintaining an adequate copper concentration inside the cell is crucial. Even moderate dysregulation of homeostasis can lead to cytotoxicity, protein aggregation, resulting in cell death. One of the significant cellular consequences in Saccharomyces cerevisiae is the inhibition of Sec61-mediated protein translocation in the secretory pathway, resulting in the accumulation of the cytosolic form of precursor secretory proteins, such as Gas1 and CPY, in the cytosol. The accumulation of unprocessed precursor proteins in the cytosol provides a method to read out translocon dysfunction. This chapter describes experimental approaches for analysing copper-induced translocation defects in yeast, with a primary focus on Western blot-based detection of precursor and mature secretory protein species. Given the high conservation of copper homeostasis and the secretory pathway across eukaryotes, these methods offer a robust framework for investigating the molecular mechanisms underlying copper toxicity and its impact on protein folding and maturation. This biochemical assay further enables the identification and characterization of chemical or genetic modulators that mitigate copper-induced translocation defects. Compounds that restore Sec61 function or improve ER targeting and maturation reduce the accumulation of precursor forms and promote the recovery of mature protein species. Thus, this approach provides a robust platform for assessing copper toxicity and for screening chemical agents that rescue ER translocation and protein maturation under metal stress conditions.
    Keywords:  CPY; Copper homeostasis; ER; Gas1; Protein aggregation; Sec61 protein translocation; Secretory pathway protein; Western blot
    DOI:  https://doi.org/10.1016/bs.mie.2026.05.028
  28. Mol Plant. 2026 Aug 18. pii: S1674-2052(26)00264-9. [Epub ahead of print]
    Small GTPase RAN-driven PNET2 oligomerization and phase separation at the nuclear lamina promote nuclear envelope integrity in plants.
    Xiao Liu, Dingrong Jin, Chunqian Wei, Yang Xu, Yu Tang.
      The nuclear envelope is a fundamental organizer of eukaryotic cells, yet how its architecture and integrity are regulated in plants remains poorly understood. Here we identify that plant inner nuclear membrane protein PNET2 as a central scaffold that maintains nuclear envelope integrity and genome stability. Loss of PNET2 compromises nuclear membrane structure and sensitizes cells to DNA damage, whereas overexpression drives aberrant expansion of the nuclear membrane. Mechanistically, PNET2 cooperates with the nuclear lamin protein KAKU4 and CRWN1 within the nuclear lamina to promote nuclear membrane remodeling, a process driven by biomolecular condensate formation via their intrinsically disordered regions. We further uncover a direct interaction between PNET2 and the small GTPase RAN. Structural modelling and biochemical analyses reveal its active GTP-bound form stimulates PNET2 oligomerization, potentially promoting its phase separation to drive membrane expansion. Genetic analyses confirm that PNET2 and RAN function in a shared pathway essential for nuclear membrane integrity. Together, our findings define a regulatory module that orchestrates GTPase signaling to control nuclear membrane homeostasis in plants, positioning PNET2 as a key nexus linking membrane dynamics, nuclear lamina organization, and genome protection.
    Keywords:  PNET2; nuclear envelope; nuclear integrity; nuclear lamina; phase separation; small GTPase RAN
    DOI:  https://doi.org/10.1016/j.molp.2026.08.006
  29. Proc Natl Acad Sci U S A. 2026 Aug 25. 123(34): e2602410123
    Structural rearrangements underlying the activation of STIM1 by ER calcium depletion.
    Ruoyi Qiu, Richard S Lewis.
      In resting cells, STIM1, the dimeric endoplasmic reticulum (ER) Ca2+ sensor that controls store-operated Ca2+ entry (SOCE), is held in a Ca2+-bound inactive state by multiple intramolecular restraints, or brakes. Receptor-evoked release of Ca2+ from the ER causes a large conformational change in STIM1 that releases the brakes and exposes the CRAC activation domain (CAD), enabling it to bind and open store-operated Orai1 channels in the plasma membrane. We performed single-molecule Förster resonance energy transfer (smFRET) measurements with purified STIM1 to better understand how Ca2+ release from the luminal domain of STIM1 drives the conformational changes in the cytosolic domain that underlie CAD release. We find that Ca2+ removal releases the CAD from CC1α1 (the "CC1 clamp") without obligatory formation of the CC1 coiled-coil that has been associated with CAD release in cells. Surprisingly, the CAD rearranges dramatically during release, as the two hairpin protomers that create its characteristic V-shaped structure are spread apart. Locking the two protomers together by cysteine crosslinking prevents CAD release, suggesting that the CAD must rearrange to escape the CC1 clamp. Our data support a model in which ER depletion-induced dimerization of the luminal SAM domains drives the cytosolic domain into multiple intermediate states including a 3-helix bundle of CC1α1/2/3, releasing the CC1 clamp and allowing the CAD to escape through a "fold-out" mechanism. Subsequent formation of the CC1 coiled-coil enables the CAD to revert to its original shape and extends it toward the plasma membrane to activate Orai1.
    Keywords:  STIM1; calcium signaling; single-molecule FRET; store-operated calcium entry
    DOI:  https://doi.org/10.1073/pnas.2602410123
  30. J Mol Biol. 2026 Aug 19. pii: S0022-2836(26)00368-2. [Epub ahead of print] 169995
    Codon optimality modulates cellular stress and innate immune responses triggered by exogenous RNAs.
    Chotiwat Seephetdee, Nada Bejar, Kathrina D Castillo, Eric Chau, Thy Y Nguyen, Biana Godin, Daniel L Kiss.
      Despite advancements in RNA sequence design, evidence regarding the preferential use of synonymous codons on cellular stress and innate immune responses is lacking. To this end, we developed a new codon optimality formula to re-engineer the coding sequences of three luciferase reporters. We demonstrate that mRNAs enriched in optimal codons elicited dramatic increases in luciferase activities compared to less optimal sequences both in vitro and in vivo. Notably, transfecting low optimality test RNAs suppress the translation of co-transfected control mRNAs in dual reporter assays. Transcriptomic profiling revealed temporally distinct waves of cellular stress, comprising an early ribotoxic stress response followed by a sustained integrated stress response and interferon-driven innate immune program. Cells responded to both dsRNA contaminants, which was mitigated by cellulose purification, and also rare codon-mediated ribosome stalling signal activating GCN2 independently of dsRNA. Translation suppression by low optimality mRNAs was driven by eIF2α phosphorylation, which was suppressed by a GCN2 inhibitor, but not by other eIF2α kinase inhibitors, and both cellulose purification and GCN2 inhibition each produced partial translational rescue. Using nucleoside-modified or circular RNAs also fully or partially abrogated these responses. Finally, only optimal, circular RNAs have enhanced RNA lifespan and duration of protein expression. Our results show that RNA sequence, composition, and structure all govern RNA translatability. Further, RNA sequences with poor codon optimality are immunogenic and induce cellular stress. Together, we show that RNA coding sequence design is a key consideration for both mRNA and circular RNA therapeutics.
    Keywords:  RNA therapeutics; circRNA; codon optimality; innate immune evasion; reporter assays; stress response
    DOI:  https://doi.org/10.1016/j.jmb.2026.169995
  31. EMBO J. 2026 Aug 18.
    Disease-specific tau polymorphs are associated with unique protein networks across proteinopathies.
    Nicha Puangmalai, Nemil Bhatt, Nopparat Suthprasertporn, Ernesto G Miranda-Morales, Nikita Shchankin, Madison Samples, Ahmet E Balci, Jia Yi Liew, Mauro Montalbano, Yingxin Zhao, Rakez Kayed.
      Tau protein aggregates adopt distinct conformations across tauopathies, yet the protein interactions engaged by disease-specific polymorphs remain poorly characterized. Here, we demonstrate that conformationally distinct tau polymorphs associate with disease-specific interaction networks across Alzheimer's disease (AD), progressive supranuclear palsy (PSP), and dementia with Lewy bodies (DLB). Interactome profiling of tau aggregates from PBS- and sarkosyl-soluble brain fractions identified 493 high-confidence interactors exhibiting remarkable disease specificity. As an exploratory feature discovery machine learning classification discriminated against diseases using as few as four to six protein features. AD tau selectively engaged glycolytic enzymes, TCA cycle components, and glutamate/GABA cycling machinery, with the astrocytic transporter SLC1A2 showing 27-fold enrichment. PSP tau exhibited extensive interactor depletion alongside selective proteasome enrichment, whereas DLB tau associated with neurogenesis modulators while depleting neuroinflammatory mediators. Interaction patterns were corroborated by parallel reaction monitoring mass spectrometry and proximity ligation assays and corresponded to disease-specific post-translational modification profiles. These findings show that tau polymorph conformations are associated with disease-specific interaction networks, providing molecular insight into tauopathy heterogeneity.
    DOI:  https://doi.org/10.1038/s44318-026-00897-8
  32. bioRxiv. 2026 Jul 31. pii: 2026.07.31.741769. [Epub ahead of print]
    Competing Molecular Interactions Govern the Dynamical Arrest of G3BP1 Condensates.
    Anurag Singh, Vicky Liu, Tharun Selvam Mahendran, Aoon Rizvi, Bhargavi Gindra, Brian Freibaum, Hong Joo Kim, J Paul Taylor, Rohit V Pappu, Priya R Banerjee.
      G3BP1 is a central scaffold of stress granules (SGs). Upon cellular stress, G3BP1 forms complex coacervates with translationally repressed mRNAs and recruits multiple RNA-binding proteins to form reversible biomolecular condensates. Persistent SGs are linked to age-dependent dynamical arrest and impaired disassembly. Here, we employ active and passive nanoscale rheology with optical tweezers to show that G3BP1 condensates evolve from being dominantly viscous fluids to dynamically arrested network glasses characterized by nanoscale caging and elastic memory. Integrating atomistic and coarse-grained simulations with experiments, we find that electrostatic interactions between the oppositely charged intrinsically disordered regions drive condensate ageing. RNA modulates these interactions in a length-and structure-dependent manner, delaying dynamic arrest, whereas Caprin-1 binding to the NTF2L domain has little effect. Together, these findings reveal how competing inter-IDR and IDR-RNA interactions govern condensate ageing and material-state transitions. The findings have broader implications for the regulation of SG dynamics in cells.
    DOI:  https://doi.org/10.64898/2026.07.31.741769
  33. Autophagy. 2026 Aug 21.
    Loss of BECN1 does not compromise autophagosome formation but alters cargo selection and HLA-restricted antigen presentation of B cells.
    Svenja L Walter, Felix Baur Lermer, Sophie Buchser, Sibilla Sander, Kennith Brian Castelino, Marcel Wacker, Eanna Fennell, Lisa Rieble, Saskia von Boxberg, Sandra Schmid, Christopher A Hess, Laure-Anne Ligeon, Juliane Walz, Muriel Mari, Fulvio Reggiori, Jörn Dengjel, Christian Münz.
      BECN1 (beclin 1) is a member of the nucleation complex and considered crucial for induction of macroautophagy/autophagy, leading to the formation and ultimate degradation of autophagosomes. We found that in human B lymphoblastoid cell lines (LCLs) deficient of BECN1 (BECN1-KO), autophagosome formation was intact and autophagic flux could be induced upon nutrient starvation or MTOR inhibition. Remarkably, autophagosomal cargo differed significantly between BECN1-KO and control (CTRL) LCLs, revealing a preferred formation of autophagosomes at the endoplasmic reticulum (ER) and not at endosomes/lysosomes in BECN1-KO LCLs. Endosomal TLR3 (toll like receptor 3) was less frequently incorporated within autophagosomes in BECN1-KO LCLs. Additionally, several proteins of the ER-resident peptide loading complex for MHC class I antigen presentation were found enriched in autophagosomes from BECN1-KO LCLs, resulting in a diminished detection of BECN1-KO LCLs by T cells. Hence, BECN1 seems to be dispensable for autophagosome formation but rather contributes to cargo selection of phagophores and immunosurveillance.
    Keywords:  B cells; Beclin-1; CD8+ T cells; MHC class I; peptide loading complex
    DOI:  https://doi.org/10.1080/15548627.2026.2719432
  34. MicroPubl Biol. 2026 ;2026
    The methyltransferase Rrp8 is a positive regulator of autophagy flux in Saccharomyces cerevisiae.
    Swaroopa Badenahalli Narasimhaiah, Elizabeth Delorme-Axford.
      Macroautophagy (hereafter referred to as autophagy) is a dynamic pathway of cellular degradation and recycling that is conserved from yeast to humans. The products of autophagic degradation may be used for anabolic reactions during nutrient-limited conditions. Thus, autophagy serves both metabolic and quality control functions. However, the role of nucleolar proteins in autophagy remains largely unexplored. Here we identify Ribosomal RNA processing 8 (Rrp8) as a positive regulator of autophagy flux in the yeast Saccharomyces cerevisiae. Our work provides insight into the role of the conserved nucleolar protein Rrp8 in regulating cellular responses to starvation.
    DOI:  https://doi.org/10.17912/micropub.biology.002252
  35. Biochem Biophys Res Commun. 2026 Aug 19. pii: S0006-291X(26)01226-X. [Epub ahead of print]834 154462
    Molecular mechanisms of protein disulfide isomerase antagonism by punicalagin.
    Osamede C Owegie, Rong-Guang Xu, Quinn P Kennedy, Tanvi Penubothu, Ivan Hancco Zirena, Oren Levy, Quoc Quang Luu, Yiliang Chen, Pavel Davizon-Castillo, Yunfeng Chen, Moua Yang.
      Punicalagin, an ellagic acid polyphenol from pomegranate, has been proposed as an antagonist of protein disulfide isomerase (PDI) and endoplasmic reticulum resident protein 57 (ERp57)-thiol isomerases that regulate protein folding and extracellular thrombotic signaling. Here, biochemical oxidase and reductase assays on PDI showed that punicalagin inhibits both activities with micromolar potency, thereby extending earlier work that described disulfide reductase inhibition. Using purified domains to determine potential interaction sites indicated that punicalagin antagonizes both the N-terminal a and C-terminal a' domains similarly. Broader profiling of the PDI family demonstrated that punicalagin selectively inhibits the oxidase activity of multiple thiol isomerases while leaving ERp72 unaffected. In parallel, jump-dilution experiments revealed a reversible mechanism of inhibition. Thiol labeling of PDI's catalytic cysteines detected no change in the redox state, supporting a noncovalent, allosteric mechanism. Complementary spectrophotometric and fluorometric assays showed that punicalagin coordinates with zinc to further inhibit PDI, suggesting metal complexation as an additional feature that may shape its interaction with thiol isomerases. Extensive molecular docking and molecular dynamics simulations showed that punicalagin binds stably and preferentially to defined sites on both the N- and C-terminal domains through extensive hydrogen bonding and van der Waals contacts. Finally, artificial intelligence-driven network analysis identified PDI as a high-confidence target of punicalagin and related galloylated polyphenols, alongside additional signaling proteins. Together, these findings provide further mechanistic framework for punicalagin-mediated antagonism of PDI and highlight galloylated polyphenols as promising scaffolds for PDI-targeted therapeutics.
    Keywords:  Galloylated polyphenol; Oxidase; Protein disulfide isomerase; Punicalagin; Reductase; Thiol isomerase
    DOI:  https://doi.org/10.1016/j.bbrc.2026.154462
  36. bioRxiv. 2026 Aug 03. pii: 2026.06.29.735430. [Epub ahead of print]
    Iron-endoplasmic reticulum-extracellular matrix axis regulates cancer cell invasion.
    Arun Asif, Kavya Panjwani, Kavya Nair, Percy Smith, Oruko Dancan, Isaiah Crosbourne, Joseph DeLuca, Taylor Humphrey, Ramon Bossardi Ramos, David T Corr, Teresita Padilla-Benavides, Margarida Barroso.
      Intracellular iron homeostasis is increasingly recognized as a regulator of cancer cell behavior, but how iron distribution influences extracellular matrix (ECM) organization and invasion remains poorly understood. Here, we show that loss of divalent metal transporter 1 (DMT1/SLC11A2) disrupts intracellular iron homeostasis and promotes cancer cell invasion through an iron-ER-ECM axis. In MDA-MB-231 cells, DMT1 knockout (KO) reduced total iron content but increased the labile iron pool (LIP) in both 2D and 3D culture models, indicating altered intracellular iron distribution. Across transcriptomic and phenotypic readouts, DMT1-dependent effects were more evident in 3D than in 2D models, with DMT1 KO inducing endoplasmic reticulum (ER) stress and impaired collagen/ECM organization. Functionally, the DMT1-loss phenotype was marked by reduced 2D motility, whereas in 3D spheroid models DMT1 KO cells displayed enhanced invasive outgrowth in both Matrigel and collagen matrices. Iron chelation further modulated this phenotype in a DMT1-dependent manner. Pharmacologic induction of ER stress phenocopied the loose spheroid architecture and invasive behavior, supporting ER stress as a mechanistic link between altered iron handling and ECM destabilization. Together, these findings identify intracellular iron distribution, rather than total iron abundance alone, as a determinant of ECM integrity and context-dependent cancer cell invasion. One Sentence Summary: Iron misrouting, not iron excess, drives ER stress and collagen failure that unleash invasion in triple-negative breast cancer.
    DOI:  https://doi.org/10.64898/2026.06.29.735430
  37. bioRxiv. 2026 Aug 05. pii: 2026.08.05.743015. [Epub ahead of print]
    A proximity biotinylation approach for the identification of membrane contact site proteins in Toxoplasma gondii.
    Kaelynn V Parker, Diego Huet.
      Inter-organellar communication is crucial for cellular function. Inside the cell, organelles interact with each other via membrane contact sites (MCSs). These structures mediate the close apposition of two organellar membranes to allow for the exchange of metabolites, ions and lipids. Most of what is known about MCSs comes only from a handful of well-studied metazoans, particularly yeast and mammals. Apicomplexans are parasites that drive human disease throughout the world. Yet, little is known about the makeup or function of their MCSs, leaving a gap in our understanding of how organelles communicate beyond conventional model eukaryotes. Here, we used a proximity biotinylation approach to map the surface proteome of three organelles in the model apicomplexan Toxoplasma gondii : the apicoplast-a non-photosynthetic plastid found only in apicomplexans-its single mitochondrion and the endoplasmic reticulum. By subtracting a cytosolic spatial reference, our high-stringency proteomic analysis uncovered candidate proteins localized simultaneously to multiple organellar surfaces suggesting their role as MCS components. We then validate our approach by characterizing a candidate involved in the association between the apicoplast and the mitochondrion. Overall, our findings provide a valuable approach to identify MCSs in apicomplexans and set the stage to apply our approach to other organelles in these pathogens.
    Highlights: Generation of surface proteomes for the apicoplast, mitochondrion, and ER in Toxoplasma gondii Mapping of the first endoplasmic reticulum and mitochondrial surface proteomes in T. gondii Identified novel membrane contact site candidate proteinsValidated a membrane contact site candidate mediating mitochondrion-apicoplast interactions.
    DOI:  https://doi.org/10.64898/2026.08.05.743015
  38. bioRxiv. 2026 Aug 06. pii: 2026.08.05.743105. [Epub ahead of print]
    PANCS-Inhibitors: A rapid method to directly select for protein-protein interaction inhibitors.
    Matthew J Styles, Victoria Cochran Xie, Sandrine Legault, Joshua A Pixley, Bryan C Dickinson.
      Aberrant protein-protein interactions (PPIs) drive myriad diseases. Inhibiting these PPIs often relies on discovering molecules that bind to one of the proteins and hoping that this binding inhibits the PPI. Molecular binder discovery often takes months, but a discovery process that ensures that the resulting molecule not only binds a target protein, but selectively inhibits a target PPI, could dramatically accelerate these endeavors. Here, we develop Phage-Assisted Non-Continuous Selection of PPI Inhibitors (PANCS-Inhibitors): a rapid screening platform that directly selects for molecules capable of disrupting a pre-formed PPI. We demonstrate this new platform using three clinically relevant oncogenic PPIs: KRas-Raf, Mdm2-p53, and Myc-Max. PANCS-Inhibitors can be used to both improve known PPI inhibitors and for de novo discovery of mini-protein PPI inhibitors that function in mammalian cells. This platform has the potential to rapidly generate inhibitors for many clinically relevant PPIs, which can be used as starting points for therapeutic development.
    DOI:  https://doi.org/10.64898/2026.08.05.743105
  39. iScience. 2026 Aug 21. 29(8): 116529
    Transcriptome-wide splicing analysis reveals HuR-regulated post-transcriptional programs in osteocytes.
    Ziqiu Fan, Aseel Marahleh, Hideki Kitaura, Jiayi Ren, Abdulrahman Mousa, Fumitoshi Ohori, Kuniyasu Niizuma, Sherif Rashad, Hiroyasu Kanetaka.
      Post-transcriptional gene regulation is central to maintaining cellular homeostasis. Among its mechanisms, alternative splicing (AS) fine-tunes cellular adaptation to stress. This study employed an approach combining RNA splicing analysis with RNA-binding protein (RBP) motif enrichment in primary osteocytes cultured in high-glucose conditions. Our analysis identified the RBP human antigen R (HuR) as a top candidate associated with AS regulation. Loss of HuR reshaped the transcriptome through gene expression and splicing changes, converging on two major pathways: stress response and translational control. Functional validation revealed that HuR depletion heightened oxidative stress, impaired mitochondrial function, and rewired key translational signals, while preserving global protein output. Mechanistically, we identified TXNIP mRNA-protein uncoupling following HuR knockdown (KD), characterized by elevated mRNA but reduced protein expression. Collectively, these findings support HuR's role as a key post-transcriptional regulator of osteocyte metabolic adaptation under high-glucose stress, with potential implications for hyperglycemic bone fragility.
    Keywords:  HuR/Elavl1; RNA-binding proteins; TXNIP; alternative splicing; hyperglycemia; mTOR; osteocytes; translation
    DOI:  https://doi.org/10.1016/j.isci.2026.116529
  40. Matrix Biol. 2026 Aug 17. pii: S0945-053X(26)00086-7. [Epub ahead of print] 102044
    Col4a1 mutation in a mouse model of cerebral small vessel disease causes white matter defects that can be modulated by targeting protein folding.
    Gaia Brezzo, Alexandra Pokhilko, Jon Moss, Juraj Koudelka, Maurits A Jansen, Ross Lennen, Gerry Thompson, Nela Fialova, Sofia Marina Konstantinidou, Joyce Yau, Erin Boland, Anna Williams, Stuart M Allan, Alessandra Granata, Sanjay Sinha, Tao Wang, Hugh S Markus, Roman Fischer, M Zameel Cader, Tom Van Agtmael, Karen Horsburgh.
      Collagen IV, encoded by genes COL4A1/COL4A2, is a major component of the basement membrane, a specialised extracellular matrix (ECM) structure. Mutations in these genes cause a genetic form of cerebral small vessel disease (cSVD), a leading cause of stroke and dementia. White matter abnormalities are a hallmark of cSVD and are closely linked to cognitive decline and dementia. While white matter defects occur in patients with COL4A1/2 mutations, they remain understudied and their mechanisms are unclear. To address these knowledge gaps, we combined magnetic resonance diffusion tensor imaging, pathology, ultrastructural investigations, behaviour and proteomic analysis of white matter in an established mouse model of cSVD due to a Col4a1 mutation (Col4a1+/Svc). The studies revealed that Col4a1+/Svc mice have reduced myelinating oligodendrocyte pools, axonal myelination defects, and altered white matter structural integrity as well as cognitive impairments. Proteomic analysis of isolated white matter from Col4a1+/Svc mice identified extensive changes to ECM and basement membrane composition. Furthermore, this provided evidence for altered endoplasmic reticulum (ER) biology including ER stress. To determine if white matter defects can be attenuated by targeting protein folding in the ER by promoting collagen IV secretion, we treated mice with the FDA-approved chemical chaperone 4-phenylbutyric acid. This revealed increased myelinating oligodendrocytes and improved axon-glial integrity in Col4a1+/Svc mice. These data provide novel insight into the pathomolecular mechanisms of collagen IV mutations in white matter abnormalities in cSVD and identify a modifiable pathway as a putative therapeutic target.
    Keywords:  Cerebral small vessel disease; Col4a1; ER stress; basement membrane; proteomics; white matter
    DOI:  https://doi.org/10.1016/j.matbio.2026.102044
  41. Cell Rep. 2026 Aug 17. pii: S2211-1247(26)00862-4. [Epub ahead of print]45(8): 117784
    Deubiquitylase OTUD3 regulates integrated stress response to suppress progression and sorafenib resistance of liver cancer.
    Hongmiao Dai, Bo Wu, Yingwei Ge, Yang Hao, Lijie Zhou, Ruolin Hong, Jinhao Zhang, Wenli Jiang, Yuting Zhang, Hongchang Li, Lingqiang Zhang.
      
    DOI:  https://doi.org/10.1016/j.celrep.2026.117784
  42. bioRxiv. 2026 Aug 03. pii: 2026.07.31.742071. [Epub ahead of print]
    A conserved vesicular circuit enables metabolic adaptation and maintains systemic heme homeostasis.
    Sohini Dutt, Simon Beardsley, Xiaojing Yuan, Ran Yu, Audrey Belot, Blake Carrington, Kevin Bishop, Sandeepan Ghosh, Jiayu Li, Xuedi Zhang, Jean-Martin Harder, Rutwick Surya Ulhas, Shilpa Dilip Kumar, Vijaya Pandey, Raman Sood, Harold Smith, Michael Krause, James A Wohlschlegel, Ute A Hellmich, Caiyong Chen, Paul Liu, Iqbal Hamza.
      Metabolic homeostasis depends on adaptive control of intracellular metabolite flux, yet how such control is reconfigured when canonical transport pathways fail is unknown. Here we define a conserved vesicular circuit that preserves systemic heme balance by rerouting intracellular heme flux. We show that loss of the intestinal heme exporter MRP-5 in Caenorhabditis elegans causes lethal heme sequestration within endolysosomal compartments. This defect is bypassed by disabling the vesicular adaptor AP-3, which stabilizes and reroutes the heme importer HRG-1, restoring heme export without increasing cytosolic heme. Unbiased genetics and transcriptomics identify two previously uncharacterized SLC49A family members, HRG-13 and HRG-14, as heme exporters with distinct affinities that engage in a vesicular importer-exporter handoff. Live imaging reveals heme-enhanced contacts between HRG-1 and SLC49A-containing vesicles, consistent with direct vesicular transfer. This circuitry extends to vertebrates as disruption of the SLC49A3 homolog impairs erythropoiesis in zebrafish and causes intracellular heme overload, premature hemoglobinization and apoptosis in differentiating human erythroid cells. Together, these findings establish SLC49A3 proteins as conserved heme exporters and uncover a general principle of metabolic adaptation in which reprogramming intracellular compartmentalization, rather than increasing nutrient supply, restores systemic homeostasis.
    DOI:  https://doi.org/10.64898/2026.07.31.742071
  43. Autophagy. 2026 Aug 16. 1-20
    TOLLIP suppresses picornavirus internalization by targeting and degrading VP1 protein through selective autophagy.
    Wei Zhang, Yang Yang, Wenhua Shao, Xingyan Dong, Tingting Zhou, Huifang Yan, Hong Tian, Weijun Cao, Zixiang Zhu, Fan Yang, Haixue Zheng.
      Foot-and-mouth disease virus (FMDV) represents a major threat to global livestock production. The capsid protein VP1 is crucial for infection; however, the host factors and mechanisms responsible for VP1 restriction remain poorly understood. We previously identified the host chaperone DNAJA3 as a host restriction factor that inhibits FMDV infection by promoting VP1 degradation through the autophagy-lysosomal pathway. Here, we elucidate the molecular mechanism by which DNAJA3 mediates degradation of VP1. We demonstrate that DNAJA3 recruits the autophagy cargo receptor TOLLIP to facilitate selective autophagic degradation of VP1. Mechanistically, TOLLIP directly interacts with the 1-37 amino acid (aa) region of VP1 through its N-terminal and C-terminal domains, and full-length TOLLIP is required for efficient VP1 degradation. Furthermore, DNAJA3 recruits the E3 ubiquitin ligase TRIM21 to promote VP1 polyubiquitination through K27-, K48-, and K63-linked ubiquitination. TOLLIP additionally restricts FMDV internalization by modulating early endosomal trafficking in an autophagy-dependent manner. In vivo, tollip-deficient suckling mice exhibit increased susceptibility to FMDV infection. Moreover, TOLLIP exerts antiviral activity against multiple picornaviruses, including Senecavirus A and Enterovirus 71. Notably, FMDV downregulates endogenous TOLLIP expression through the protease activity of the viral 3C protein, enabling the virus to evade host autophagic surveillance. Collectively, our study identifies a novel DNAJA3-TRIM21-TOLLIP axis that restricts FMDV infection through selective autophagy, establishes TOLLIP as a host restriction factor against picornaviruses, and reveals potential molecular targets for antiviral intervention.
    Keywords:  Autophagy; FMDV; TRIM21; early endosomal trafficking; ubiquitination
    DOI:  https://doi.org/10.1080/15548627.2026.2715275
  44. Biochem J. 2026 Sep 02. 483(9): 1709-1711
    Photoreactive crosslinkers capture the ubiquitin-E2-E3 ligases intermediates.
    Tingting Yao, Tatiana G Kutateladze.
      E3 ligases catalyze the final step in transferring ubiquitin to protein substrates, but resulting intermediates are characterized by conformational flexibility. Recent work by Chandler and colleagues ( Biochem J. (2026) 483, 1115-1130) demonstrates that photocross-linking activity-based probes can trap transient conformations of the intermediates, providing a valuable tool to map interfaces during the ubiquitin transfer process.
    Keywords:  crosslinker; ubiquitin ligases; ubiquitin signalling
    DOI:  https://doi.org/10.1042/BCJ20260338
  45. bioRxiv. 2026 Aug 04. pii: 2026.08.03.742535. [Epub ahead of print]
    Calcineurin B-mediated Ca 2+ sensing translates stress signal intensity into the assembly of phase-separated condensates at PERK complexes.
    Sebastian M Bairo, Macarena Fernandez, Gonzalo Quasollo, Andrea Pellegrini, Juan C de Batista, Santiago Asis, Mauricio Martin, Deborah Holstein, James D Lechleiter, Gabriela E Gomez, Mariano Bisbal, Mariana Bollo.
      Endoplasmic reticulum (ER) stress activates protein kinase RNA-like ER kinase (PERK), which initially promotes adaptive responses but remains the only active UPR branch during prolonged stress, mediating both early cytoprotective and chronic pro-apoptotic signaling. Recently, we identified translocon-generated Ca 2+ microdomains that promote PERK phosphorylation during early UPR, revealing a mechanism by which local Ca 2+ signals regulate UPR activation. However, the molecular mechanism linking these Ca 2+ microdomains to PERK activation remains elusive. Previously, we showed that calcineurin (CN), a Ca 2+ -dependent heterodimer composed of catalytic (CNA) and regulatory (CNB) subunits, exerts a non-canonical pro-survival function by promoting PERK autophosphorylation. Here, using super-resolution microscopy, CRISPR-Cas9 editing, in silico analyses, and optogenetic droplet assays, we identify CNB as a local Ca 2+ sensor that couples translocon-generated Ca 2+ signals to liquid condensate assembly, thereby promoting adaptive PERK phosphorylation. These findings establish CNB-mediated condensate assembly as a mechanism that translates local Ca 2+ signals into spatially organized early adaptive PERK signaling.
    DOI:  https://doi.org/10.64898/2026.08.03.742535
  46. Proc Natl Acad Sci U S A. 2026 Aug 25. 123(34): e2610235123
    Architecture and mechanism of the human p24 cargo receptor.
    Zhengkang Hua, Di Zhang, Min Zhang, Xinlin Hu, Chengtao Kang, Huanhuan Sun, Ping Yang, Chu Qi, Xinyuan Lin, Jiameng Li, Hongjun Yu.
      The p24 transmembrane emp24 domain family of cargo receptors is central to secretory trafficking, yet its molecular organization and regulatory principles have remained elusive. Here, we present cryo-EM structures of human p24 complex in three pH states, integrated with systematic functional analyses focused on glycosylphosphatidylinositol-anchored protein (GPI-AP) cargo. We identify a tightly interlocked p24α2-p24β1-p24δ1-p24γ2 heterotetramer as the minimal unit for GPI-AP engagement, resolving long-standing uncertainties regarding p24 stoichiometry and higher-order assembly. We reveal functional redundancy between p24α2 and p24α3, alongside selective incorporation of p24γ2 required for cargo binding. Systematic screening identifies two stable subassemblies: p24β1-p24δ1 and p24α2-p24β1-p24δ1, suggesting a stepwise assembly pathway. Comparative structural analysis across multiple pH states uncovers a conserved proton-sensing network centered on Coiled-coil layers 0 and +1 that drives coordinated transmembrane rearrangements, underlying pH-dependent cargo release. Our findings establish an integrated framework linking p24 architecture, isoform diversity, and pH regulation, offering a basis for understanding p24 trafficking complexity.
    Keywords:  GPI-anchored protein; TMED; cargo transport; p24; secretory pathway
    DOI:  https://doi.org/10.1073/pnas.2610235123
  47. Adv Sci (Weinh). 2026 Aug 18. e77262
    Phase Separation of TRIM21 Modulates PTPN14 Stability to Drive Flow-Dependent Endothelial Activation and Atherogenesis.
    Xue He, Qiannan Ma, Xue Wang, Jiaxu Mang, Chufeng Li, Bingjun Wu, Liu Yao, Yequn Chen, Qiankun Bao, Yi Zhu, Jinlong He.
      Local flow patterns determine the uneven distribution of atherosclerotic lesions, yet the underlying mechanism remains poorly defined. We previously reported that protein tyrosine phosphatase nonreceptor type 14 (PTPN14) is implicated in atherosclerosis. Its role in flow-dependent endothelial dysfunction and the upstream regulatory mechanisms remain incompletely understood. Here, we show that PTPN14 functions as a flow-sensitive regulator in endothelial cells, with its protein stability reduced under disturbed flow. Endothelial-specific deletion of PTPN14 led to exacerbated endothelial inflammation and atherosclerosis in vivo. We further identify TRIM21 as a critical E3 ubiquitin ligase that binds PTPN14 via its SPRY domain and promotes K48-linked polyubiquitination at lysine 956, resulting in proteasomal degradation of PTPN14. Notably, disturbed flow induced TRIM21 LLPS in a cytoskeleton-dependent manner, facilitating the recruitment of PTPN14 into co-condensates and enhancing its ubiquitination. Endothelial-specific TRIM21 overexpression aggravated disturbed flow-induced endothelial inflammation and atherosclerosis. In contrast, endothelial-specific TRIM21 deficiency attenuated induced endothelial activation and atherosclerosis; importantly, this atheroprotective effect was abolished by concomitant PTPN14 deletion, establishing a functional TRIM21-PTPN14 regulatory axis in vivo. Together, these findings uncover phase separation-dependent ubiquitin signaling as a mechanistic link between hemodynamic forces and endothelial dysfunction and suggest the TRIM21-PTPN14 pathway as a potential therapeutic target in atherosclerosis.
    Keywords:  PTPN14; TRIM21; atherosclerosis; endothelial mechanotransduction; shear stress
    DOI:  https://doi.org/10.1002/advs.77262
  48. Mol Cell. 2026 Aug 17. pii: S1097-2765(26)00512-5. [Epub ahead of print]
    Excess intracellular lactate represses mRNA translation and tumor progression through lactate-tRNA charging.
    Xinyu Wang, Chi Lin, Yixin Zhang, Huan Wang, Biru Shi, Zhiwen Bu, Pengtao Xu, Huimin Zheng, Yanhong Wang, Xiaojing Lan, Qi Hu, Min Huang, Ying Wang, Huiru Tang, Yichen Xu.
      Tumor cells rely on sustained protein synthesis despite fluctuating metabolic stress. To examine how metabolic state directly influences translational output, we investigated lactate utilization. Intracellular accumulation of lactate, a central glycolytic product, acutely represses mRNA translation. Mechanistically, alanyl-tRNA synthetase 1 (AARS1) charges tRNAs with lactate instead of amino acids. Unlike the rapid and selective transfer of alanine to cognate tRNAAla, slower lactyl transfer permits lactate modification of non-cognate tRNAs, broadly compromising elongation fidelity. Functionally, this direct metabolic control over a fundamental process of the central dogma reshapes the translatome, operating as an intrinsic metabolic brake that aligns biosynthetic capacity with energy state. Notably, aggressive tumors elevate lactate transporters, limiting intracellular lactate accumulation and evading translational repression. Pharmacological blockade of monocarboxylate transporters restores intracellular lactate accumulation, re-establishes translational repression, and impairs tumor progression in mice. These findings uncover a metabolite-tRNA charging event directly rewiring translational output and reveal a metabolic vulnerability with therapeutic potential.
    Keywords:  cancer; lactate; lactyl-tRNA; metabolism-translation coupling; tRNA charging; translation control
    DOI:  https://doi.org/10.1016/j.molcel.2026.07.026
  49. Mol Cell Proteomics. 2026 Aug 21. pii: S1535-9476(26)00145-3. [Epub ahead of print] 101649
    Surface proteomics of cancer-derived extracellular vesicles reveals universal heat shock proteins and cancer signatures.
    Leyla Ayse Erozenci, Fenna Feenstra, Sander R Piersma, Thang V Pham, Jianang Liu, Joke Wortel, Jan R T van Weering, Natasja F J Dits, Guido Jenster, R Jeroen A van Moorselaar, Irene V Bijnsdorp, Connie R Jimenez.
      Extracellular vesicles (EVs) secreted by cancer cells are a valuable source of non-invasive biomarkers, as they are released into biofluids and reflect the molecular makeup of their cells of origin. To enable direct capture and detection of cancer-derived EVs, detailed knowledge of their surface proteome is essential. Here, we combined mild enzymatic treatment with mass spectrometry to analyze outward-facing proteins of EVs across a panel of 18 cell lines. This approach identified 1094 EV-associated surface proteins, including 144 transmembrane proteins involved in cellular communication and transport. Peptide-level analyses revealed that nearly 50% of multi-pass transmembrane proteins, such as ATP1A1, displayed their intracellular domains on the external EV surface. Out of a core set of 112 EV surface proteins shared across all cancer types, we report a top list of 15 highly abundant EV-surface proteins including ATP1A1, heat shock proteins HSPA8 and HSP90AB1 and nuclear proteins linked to EV biogenesis and cancer progression. This comprehensive profiling of EV surface proteomes from diverse cancer types reveals a robust set of pan-cancer EV surface markers with potential for diagnostic analysis. Significance: Over the past decade, EVs have emerged as critical mediators of intercellular communication in both physiological and pathological contexts. While the EV proteome is well-known, detailed knowledge is missing on which proteins are outward facing on the EV surface that would enable to develop tools to isolate cancer-specific EVs. We report a comprehensive EV surface proteome across multiple cancer types, revealing both universally present surface proteins and tumor type-specific markers. A top list of 15 pan-cancer EV surface proteins included heat shock proteins HSPA8 and HSP90AB1 as well as ATP1A1. These findings not only advance our understanding of the molecular architecture of the EV surface but also lay the foundation for developing targeted EV capture strategies for diagnostic and therapeutic applications across diverse cancers.
    Keywords:  Cancer; LC-MS/MS proteomics; extracellular vesicle; pan-cancer; surface proteins
    DOI:  https://doi.org/10.1016/j.mcpro.2026.101649
  50. iScience. 2026 Aug 21. 29(8): 117152
    Stress-induced secretory pathway disruption causes atypical metalloproteinase transport in PMM2-CDG.
    Chia-Lun Wu, Kali Wiggins, Lynn Dukes-Rimsky, Kishore Garapati, Akhilesh Pandey, Richard Steet, Heather Flanagan-Steet.
      Matrix metalloproteinase (Mmp) dysfunction has been implicated as a driver of cartilage and neuromuscular pathologies in a common congenital disorder of glycosylation, PMM2-CDG. Since Mmp activity and interactions can be regulated by their glycans, these abnormalities were thought to likely arise from direct effects on enzyme glycosylation. However, here we show that disruptions in secretory pathway morphology alter the trafficking of Mmp2 and its activator, membrane-type Mmp MT1-Mmp. Biochemical and visual studies indicate enhanced processing by a furin proconvertase causes MT1-Mmp to directly exit the ER, bypassing the Golgi. This unconventional route of cell surface transport prevents MT1-Mmp and Mmp2 from interacting inside chondrocytes, causing pro-Mmp2 to accumulate in the Golgi. Importantly, defects in Mmp trafficking do not appear to correspond to direct defects in the N-glycosylation of either enzyme, suggesting that stress-induced alterations in secretory pathway organization may instead underlie the atypical Mmp trafficking in PMM2-CDG.
    Keywords:  CDG; MMP; glycosylation; secretory pathway; unconventional trafficking
    DOI:  https://doi.org/10.1016/j.isci.2026.117152
  51. ACS Med Chem Lett. 2026 Aug 13. 17(8): 1718-1720
    Targeted Degradation of KAT6A: Expanding the Therapeutic Frontier of Epigenetic Drug Discovery.
    Anna C Renner, Robert B Kargbo.
      Targeted protein degradation continues to reshape modern drug discovery by extending therapeutic intervention beyond conventional inhibition. A recent patent application discloses KAT6 degraders that selectively degrade KAT6A, an oncogenic histone acetyltransferase implicated in multiple malignancies. These degraders demonstrate potent target removal, durable pathway suppression, and enhanced antiproliferative activity, highlighting protein degradation as a promising strategy for epigenetic cancer therapy.
    DOI:  https://doi.org/10.1021/acsmedchemlett.6c00361
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