bims-proteo Biomed News
on Proteostasis
Issue of 2025–05–18
35 papers selected by
Eric Chevet, INSERM
  1. BiP, GRP94, Calreticulin and Calnexin Contribute to Development of the Notochord in Medaka Fish.
  2. ATP functions as a pathogen-associated molecular pattern to activate the E3 ubiquitin ligase RNF213.
  3. Structural ubiquitin contributes to K48 linkage specificity of the HECT ligase Tom1.
  4. IRE1α promotes phagosomal calcium flux to enhance macrophage fungicidal activity.
  5. Proteome-wide determinants of co-translational chaperone binding in bacteria.
  6. Ubiquitin-conjugating enzyme UBE2N modulates proteostasis in immunoproteasome-positive acute myeloid leukemia.
  7. Specific branches of the proteostasis network regulate the toxicity associated with mistranslation.
  8. Ribosome association inhibits stress-induced gene mRNA localization to stress granules.
  9. ER stress disrupts insulin release in murine models of type 2 diabetes by impairing retromer action and constitutive secretion.
  10. Protein targeting to the ER membrane: multiple pathways and shared machinery.
  11. The mitophagy receptors BNIP3 and NIX mediate tight attachment and expansion of the isolation membrane to mitochondria.
  12. Site-specific quantification of the in vivo UFMylome reveals myosin modification in ALS.
  13. UFMylation safeguards human hepatocyte differentiation and liver homeostasis by regulating ribosome dissociation.
  14. Cdc48 plays a crucial role in redox homeostasis through dynamic reshaping of its interactome during early stationary phase.
  15. Suppressing proteasome activity enhances sensitivity to actinomycin D in diffuse anaplastic Wilms tumor.
  16. Discovery and optimisation of a covalent ligand for TRIM25 and its application to targeted protein ubiquitination.
  17. Activation mechanism of small heat shock protein HSPB5 revealed by disease-associated mutants.
  18. Visualization of autophagic structures near solid polyQ aggregates reveals how they undermine autophagy.
  19. Quality control of un-imported mitochondrial proteins at a glance.
  20. Mechanical effect of protein glycosylation on BiP-mediated post-translational translocation and folding in the endoplasmic reticulum.
  21. Revealing arginine-cysteine and glycine-cysteine NOS linkages by a systematic re-evaluation of protein structures.
  22. Allosteric regulation of UBIAD1 trafficking from ER to Golgi revealed by chemical genetic screening.
  23. TRIM23 mediates cGAS-induced autophagy in anti-HSV defense.
  24. Small-molecule dissolution of stress granules by redox modulation benefits ALS models.
  25. Interaction networks of SIM-binding groove mutants reveal alternate modes of SUMO binding and profound impact on SUMO conjugation.
  26. RIM and MUNC13 membrane-binding domains are essential for neuropeptide secretion.
  27. EDC4 C-terminal domain scaffolds P-body assembly and links P-body dynamics to p53 mediated tumor suppression.
  28. Hsc70-4: An unanticipated mediator of dsRNA internalization in Drosophila.
  29. Inhibitors of eIF1A-ribosome interaction unveil uORF-dependent regulation of translation initiation and antitumor and antiviral effects.
  30. Prediction of protein subcellular localization in single cells.
  31. Discovery of an LSD1 PROTAC degrader.
  32. Reactive Oxygen Species-Instructed Supramolecular Assemblies Enable Bioorthogonally Activatable Protein Degradation for Pancreatic Cancer.
  33. Beta cells intrinsically sense and limit their secretory activity via mTORC1-RhoA signaling.
  34. Glutamine modulates stress granule formation in cancer cells through core RNA-binding proteins.
  35. The solute carrier superfamily interactome.
  1. Cell Struct Funct. 2025 May 13.
    BiP, GRP94, Calreticulin and Calnexin Contribute to Development of the Notochord in Medaka Fish.
    Serina Kita, Tokiro Ishikawa, Kazutoshi Mori.
      The accumulation of unfolded or misfolded proteins in the endoplasmic reticulum (ER) activates the unfolded protein response (UPR) to maintain the homeostasis of the ER. The UPR consists of the IRE1, PERK and ATF6 pathways in vertebrates. Knockout of the IRE1 and PERK pathways causes defects in liver and pancreatic β cells, respectively, in mice, whereas knockout of the ATF6 pathway causes very early embryonic lethality in mice and medaka fish, a vertebrate model organism. We previously showed that ATF6 knockout in medaka causes a defect in the development of the notochord - the notochord becomes shorter - but that transient overexpression of the ER chaperone BiP via microinjection of BiP mRNA into one-cell stage embryos of these ATF6 knockout rescues this defect. Here, we microinjected mRNA encoding various ER chaperones and found that GRP94, calreticulin and calnexin also partially rescued this defect. Thus, BiP/GRP94 and calreticulin/calnexin greatly contribute to the development of the notochord by controlling the quality of collagens and N-glycosylated proteins (such as laminin and fibrillin), respectively, which have been confirmed necessary to the formation of the notochord in zebrafish genetics.Key words: endoplasmic reticulum, protein folding, molecular chaperone, collagen, glycoprotein.
    Keywords:  collagen; endoplasmic reticulum; glycoprotein; molecular chaperone; protein folding
    DOI:  https://doi.org/10.1247/csf.25009
  2. Nat Commun. 2025 May 13. 16(1): 4414
    ATP functions as a pathogen-associated molecular pattern to activate the E3 ubiquitin ligase RNF213.
    Juraj Ahel, Arda Balci, Victoria Faas, Daniel B Grabarczyk, Roosa Harmo, Daniel R Squair, Jiazhen Zhang, Elisabeth Roitinger, Frederic Lamoliatte, Sunil Mathur, Luiza Deszcz, Lillie E Bell, Anita Lehner, Thomas L Williams, Hanna Sowar, Anton Meinhart, Nicola T Wood, Tim Clausen, Satpal Virdee, Adam J Fletcher.
      The giant E3 ubiquitin ligase RNF213 is a conserved component of mammalian cell-autonomous immunity, limiting the replication of bacteria, viruses and parasites. To understand how RNF213 reacts to these unrelated pathogens, we employ chemical and structural biology to find that ATP binding to its ATPases Associated with diverse cellular Activities (AAA) core activates its E3 function. We develop methodology for proteome-wide E3 activity profiling inside living cells, revealing that RNF213 undergoes a reversible switch in E3 activity in response to cellular ATP abundance. Interferon stimulation of macrophages raises intracellular ATP levels and primes RNF213 E3 activity, while glycolysis inhibition depletes ATP and downregulates E3 activity. These data imply that ATP bears hallmarks of a danger/pathogen associated molecular pattern, coordinating cell-autonomous defence. Furthermore, quantitative labelling of RNF213 with E3-activity probes enabled us to identify the catalytic cysteine required for substrate ubiquitination and obtain a cryo-EM structure of the RNF213-E2-ubiquitin conjugation enzyme transfer intermediate, illuminating an unannotated E2 docking site. Together, our data demonstrate that RNF213 represents a new class of ATP-dependent E3 enzyme, employing distinct catalytic and regulatory mechanisms adapted to its specialised role in the broad defence against intracellular pathogens.
    DOI:  https://doi.org/10.1038/s41467-025-59444-4
  3. Cell Rep. 2025 May 12. pii: S2211-1247(25)00459-0. [Epub ahead of print]44(5): 115688
    Structural ubiquitin contributes to K48 linkage specificity of the HECT ligase Tom1.
    Katrina Warner, Moritz Hunkeler, Kheewoong Baek, Anna Schmoker, Shourya S Roy Burman, Daan Overwijn, Cyrus Jin, Katherine A Donovan, Eric S Fischer.
      Homologous to E6AP C terminus (HECT) ubiquitin ligases play key roles in essential pathways such as DNA repair, cell cycle control, or protein quality control. Tom1 is one of five HECT ubiquitin E3 ligases in budding yeast S. cerevisiae and is prototypical for a ligase with pleiotropic functions such as ubiquitin chain amplification, orphan quality control, and DNA damage response. Structures of full-length HECT ligases, including the Tom1 ortholog HUWE1, have been reported, but how domains beyond the conserved catalytic module contribute to catalysis remains largely elusive. Here, through cryoelectron microscopy (cryo-EM) snapshots of Tom1 during an active ubiquitination cycle, we demonstrate that the extended domain architecture directly contributes to activity. We identify a Tom1-ubiquitin architecture during ubiquitination involving a non-canonical ubiquitin-binding site in the solenoid shape of Tom1. We demonstrate that this ubiquitin-binding site coordinates a structural ubiquitin contributing to the fidelity of K48 poly-ubiquitin chain assembly.
    Keywords:  CP: Cell biology; CP: Molecular biology; HECT; HUWE1; TOM1; cryo-EM; orphan quality control; protein ubiquitylation; ubiquitin
    DOI:  https://doi.org/10.1016/j.celrep.2025.115688
  4. Cell Rep. 2025 May 09. pii: S2211-1247(25)00465-6. [Epub ahead of print]44(5): 115694
    IRE1α promotes phagosomal calcium flux to enhance macrophage fungicidal activity.
    Michael J McFadden, Mack B Reynolds, Britton C Michmerhuizen, Einar B Ólafsson, Sofia M Marshall, Faith Anderson Davis, Tracey L Schultz, Takao Iwawaki, Jonathan Z Sexton, Mary X D O'Riordan, Teresa R O'Meara.
      The mammalian endoplasmic reticulum (ER) stress sensor inositol-requiring enzyme 1α (IRE1α) is essential for cellular homeostasis and plays key roles in infection responses, including innate immunity and microbicidal activity. While IRE1α functions through the IRE1α-XBP1S axis are known, its XBP1S-independent roles are less well understood, and its functions during fungal infection are still emerging. We demonstrate that Candida albicans activates macrophage IRE1α via C-type lectin receptor signaling independent of protein misfolding, suggesting non-canonical activation. IRE1α enhances macrophage fungicidal activity by promoting phagosome maturation, which is crucial for containing C. albicans hyphae. IRE1α facilitates early phagosomal calcium flux post-phagocytosis, which is required for phagolysosomal fusion. In macrophages lacking the IRE1α endoribonuclease domain, defective calcium flux correlates with fewer ER-early endosome contact sites, suggesting a homeostatic role for IRE1α-promoting membrane contact sites. Overall, our findings illustrate non-canonical IRE1α activation during infection and a function for IRE1α in supporting organelle contact sites to safeguard against rapidly growing microbes.
    Keywords:  CP: Immunology; CP: Microbiology; Candida albicans; IRE1α; calcium; fungal infection; innate immunity; phagosome
    DOI:  https://doi.org/10.1016/j.celrep.2025.115694
  5. Nat Commun. 2025 May 10. 16(1): 4361
    Proteome-wide determinants of co-translational chaperone binding in bacteria.
    Carla Verónica Galmozzi, Frank Tippmann, Florian Wruck, Josef Johannes Auburger, Ilia Kats, Manuel Guennigmann, Katharina Till, Edward P O Brien, Sander J Tans, Günter Kramer, Bernd Bukau.
      Chaperones are essential to the co-translational folding of most proteins. However, the principles of co-translational chaperone interaction throughout the proteome are poorly understood, as current methods are restricted to few substrates and cannot capture nascent protein folding or chaperone binding sites, precluding a comprehensive understanding of productive and erroneous protein biosynthesis. Here, by integrating genome-wide selective ribosome profiling, single-molecule tools, and computational predictions using AlphaFold we show that the binding of the main E. coli chaperones involved in co-translational folding, Trigger Factor (TF) and DnaK correlates with "unsatisfied residues" exposed on nascent partial folds - residues that have begun to form tertiary structure but cannot yet form all native contacts due to ongoing translation. This general principle allows us to predict their co-translational binding across the proteome based on sequence only, which we verify experimentally. The results show that TF and DnaK stably bind partially folded rather than unfolded conformers. They also indicate a synergistic action of TF guiding intra-domain folding and DnaK preventing premature inter-domain contacts, and reveal robustness in the larger chaperone network (TF, DnaK, GroEL). Given the complexity of translation, folding, and chaperone functions, our predictions based on general chaperone binding rules indicate an unexpected underlying simplicity.
    DOI:  https://doi.org/10.1038/s41467-025-59067-9
  6. J Clin Invest. 2025 May 15. pii: e184665. [Epub ahead of print]135(10):
    Ubiquitin-conjugating enzyme UBE2N modulates proteostasis in immunoproteasome-positive acute myeloid leukemia.
    Chiharu Ishikawa, Laura Barreyro, Avery M Sampson, Kathleen M Hueneman, Kwangmin Choi, Sophia Y Philbrook, Issac Choi, Lyndsey C Bolanos, Mark Wunderlich, Andrew G Volk, Stephanie S Watowich, Kenneth D Greis, Daniel T Starczynowski.
      Altered protein homeostasis through proteasomal degradation of ubiquitinated proteins is a hallmark of many cancers. Ubiquitination, coordinated by E1, E2, and E3 enzymes, involves up to 40 E2-conjugating enzymes in humans to specify substrates and ubiquitin linkages. In a screen for E2 dependencies in acute myeloid leukemia (AML), ubiquitin conjugating enzyme E2 N (UBE2N) emerged as the top candidate. To investigate UBE2N's role in AML, we characterized an enzymatically defective mouse model of UBE2N, revealing UBE2N's requirement in AML without an impact on normal hematopoiesis. Unlike other E2s, which mediate lysine-48 (K48) polyubiquitination and degradation of proteins, UBE2N primarily synthesizes K63-linked chains, stabilizing or altering protein function. Proteomic analyses and a whole-genome CRISPR-activation screen in pharmacologically and genetically UBE2N-inhibited AML cells unveiled a network of UBE2N-regulated proteins, many of which are implicated in cancer. UBE2N inhibition reduced their protein levels, leading to increased K48-linked ubiquitination and degradation through the immunoproteasome and revealing UBE2N activity is enriched in immunoproteasome-positive AML. Furthermore, an interactome screen identified tripartite motif-containing protein 21 (TRIM21) as the E3 ligase partnering with activated UBE2N in AML to modulate UBE2N-dependent proteostasis. In conclusion, UBE2N maintains proteostasis in AML by stabilizing target proteins through K63-linked ubiquitination and prevention of K48 ubiquitin-mediated degradation by the immunoproteasome. Thus, inhibition of UBE2N catalytic function suppresses leukemic cells through selective degradation of critical proteins in immunoproteasome-positive AML.
    Keywords:  Hematology; Leukemias; Oncology; Ubiquitin-proteosome system
    DOI:  https://doi.org/10.1172/JCI184665
  7. Nucleic Acids Res. 2025 May 10. pii: gkaf428. [Epub ahead of print]53(9):
    Specific branches of the proteostasis network regulate the toxicity associated with mistranslation.
    Donovan W McDonald, Rebecca N Dib, Christopher De Luca, Ashmi Shah, Martin L Duennwald.
      All cellular functions rely on accurate protein biosynthesis. Yet, many variants of transfer RNA (tRNA) genes that induce amino acid misincorporation are found in human genomes. Mistranslation induces pleiotropic effects on proteostasis, ranging from protein misfolding to impaired protein biosynthesis and degradation. We employ Saccharomyces cerevisiae (budding yeast), a genetically and biochemically tractable model that facilitates quantitative analysis of how specific proteostasis pathways interact with mistranslating tRNAs. We tested two mistranslating tRNASer variants, one inducing proline to serine (P > S), the other arginine to serine (R > S) misincorporation. We found that P > S misincorporation impairs cellular fitness and sensitizes cells to protein misfolding to a greater extent than R > S misincorporation. Of note, we also show that, even though both tRNA variants induce misincorporation of serine, they result in the accumulation of misfolded proteins by distinct mechanisms. Specifically, R > S misincorporation reduces that association of Hsp70 with misfolded proteins, while P > S misincorporation impairs the degradation of nascent polypeptides. Our findings reveal that different mistranslating tRNASer variants impair specific branches of proteostasis and thus compromise cellular fitness by distinct mechanisms.
    DOI:  https://doi.org/10.1093/nar/gkaf428
  8. Genes Dev. 2025 May 13.
    Ribosome association inhibits stress-induced gene mRNA localization to stress granules.
    Noah S Helton, Benjamin Dodd, Stephanie L Moon.
      The integrated stress response (ISR) is critical for resilience to stress and is implicated in numerous diseases. During the ISR, translation is repressed, stress-induced genes are expressed, and mRNAs condense into stress granules. The relationship between stress granules and stress-induced gene expression is unclear. We measured endogenous stress-induced gene mRNA localization at the single-molecule level in the presence or absence of small molecule translation inhibitors. Reducing ribosome association increases the localization of stress-induced gene mRNAs to stress granules, whereas increasing ribosome association inhibits their localization to stress granules. The presence of upstream open reading frames (uORFs) in mRNA reporters reduces their localization to stress granules in a ribosome-dependent manner. Furthermore, a single initiating ribosome blocks stress granule formation and inhibits mRNA association with preformed stress granules. Thus, uORF-mediated ribosome association inhibits stress-induced gene mRNA localization to stress granules, suggesting a new role for uORFs in limiting RNA condensation.
    Keywords:  ATF4; GADD34; RNA localization; condensate; integrated stress response; ribosome; stress granules; stress-induced genes; translation; upstream open reading frame
    DOI:  https://doi.org/10.1101/gad.352899.125
  9. Cell Rep. 2025 May 13. pii: S2211-1247(25)00462-0. [Epub ahead of print]44(5): 115691
    ER stress disrupts insulin release in murine models of type 2 diabetes by impairing retromer action and constitutive secretion.
    Nancy Sue, Le May Thai, Ebru Boslem, Kwan Yi Chu, Chenxu Yan, Leanne Mackin, William E Hughes, Angela Fontaine-Titley, Deborah Barkauskas, Louise Cottle, Helen E Thomas, Carsten Schmitz-Peiffer, Yan-Chuan Shi, Paul Timpson, David Herrmann, Martin Whitham, Trevor J Biden.
      Using in vitro models of lipotoxicity and islets from the db/db mouse model of type 2 diabetes (T2D), we show that endoplasmic reticulum (ER) stress impairs β cell function. This is unrelated to apoptosis or alterations in insulin content or proinsulin processing, despite expansion of the Golgi compartment. Instead, the constitutive secretory pathway and endocytic recycling are disrupted, as revealed by depletion of glycosylated proteins and syntaxins from the plasma membrane (PM) and accumulation of E-cadherin in the retromer. This involves the PERK arm of the unfolded protein response. Proteomics identified multiple PM proteins mislocalized by ER stress, notably axon-guidance and cell-adhesion proteins, and many with glycosylphosphatidylinositol linkages. A retromer chaperone attenuated defective insulin secretion from islets of both db/db and high-fat-fed mice. By identifying different endpoints and mechanisms, our results redefine the relevance of ER stress to β cell failure. They also implicate retromer chaperones as potential T2D therapeutics.
    Keywords:  CP: Metabolism; CP: Molecular biology; ER stress; Golgi complex; adherens junction; constitutive secretion; endocytosis; insulin secretion; lysosome; pancreatic β cell; plasma membrane; retromer; type 2 diabetes
    DOI:  https://doi.org/10.1016/j.celrep.2025.115691
  10. Crit Rev Biochem Mol Biol. 2025 May 16. 1-47
    Protein targeting to the ER membrane: multiple pathways and shared machinery.
    Wendy N Sánchez, Arnold J M Driessen, Christian A M Wilson.
      The endoplasmic reticulum (ER) serves as a central hub for protein production and sorting in eukaryotic cells, processing approximately one-third of the cellular proteome. Protein targeting to the ER occurs through multiple pathways that operate both during and independent of translation. The classical translation-dependent pathway, mediated by cytosolic factors like signal recognition particle, recognizes signal peptides or transmembrane helices in nascent proteins, while translation-independent mechanisms utilize RNA-based targeting through specific sequence elements and RNA-binding proteins. At the core of these processes lies the Sec61 complex, which undergoes dynamic conformational changes and coordinates with numerous accessory factors to facilitate protein translocation and membrane insertion across and into the endoplasmic reticulum membrane. This review focuses on the molecular mechanisms of protein targeting to the ER, from the initial recognition of targeting signals to the dynamics of the translocation machinery, highlighting recent discoveries that have revealed unprecedented complexity in these cellular trafficking pathways.
    Keywords:  Protein targeting; endoplasmic reticulum; sec61 complex; signal peptide; signal recognition particle; translocon
    DOI:  https://doi.org/10.1080/10409238.2025.2503746
  11. J Cell Biol. 2025 Jul 07. pii: e202408166. [Epub ahead of print]224(7):
    The mitophagy receptors BNIP3 and NIX mediate tight attachment and expansion of the isolation membrane to mitochondria.
    Shun-Ichi Yamashita, Ritsuko Arai, Hiroshi Hada, Benjamin Scott Padman, Michael Lazarou, David C Chan, Tomotake Kanki, Satoshi Waguri.
      BNIP3 and NIX are the main receptors for mitophagy, but their mechanisms of action remain elusive. Here, we used correlative light EM (CLEM) and electron tomography to reveal the tight attachment of isolation membranes (IMs) to mitochondrial protrusions, often connected with ER via thin tubular and/or linear structures. In BNIP3/NIX-double knockout (DKO) HeLa cells, the ULK1 complex and nascent IM formed on mitochondria, but the IM did not expand. Artificial tethering of LC3B to mitochondria induced mitophagy that was equally efficient in DKO cells and WT cells. BNIP3 and NIX accumulated at the segregated mitochondrial protrusions via binding with LC3 through their LIR motifs but did not require dimer formation. Finally, the average distance between the IM and the mitochondrial surface in receptor-mediated mitophagy was significantly smaller than that in ubiquitin-mediated mitophagy. Collectively, these results indicate that BNIP3 and NIX are required for the tight attachment and expansion of the IM along the mitochondrial surface during mitophagy.
    DOI:  https://doi.org/10.1083/jcb.202408166
  12. Cell Rep Methods. 2025 May 02. pii: S2667-2375(25)00084-0. [Epub ahead of print] 101048
    Site-specific quantification of the in vivo UFMylome reveals myosin modification in ALS.
    Ronnie Blazev, Barry M Zee, Hayley Peckham, Yaan-Kit Ng, Christopher T A Lewis, Chengxin Zhang, James W McNamara, Craig A Goodman, Paul Gregorevic, Julien Ochala, Frederik J Steyn, Shyuan T Ngo, Matthew P Stokes, Benjamin L Parker.
      UFMylation is a ubiquitin-like protein modification of Ubiquitin Fold Modifier 1 (UFM1) applied to substrate proteins and regulates several cellular processes such as protein quality control. Here, we describe the development of an antibody-based enrichment approach to immunoprecipitate remnant UFMylated peptides and identification by mass spectrometry. We used this approach to identify >200 UFMylation sites from various mouse tissues, revealing extensive modification in skeletal muscle. In vivo knockdown of the E2 ligase, UFC1, followed by enrichment and analysis of remnant UFMylated peptides quantified concomitant down-regulation and validation of a subset of modification sites, particularly myosin UFMylation. Furthermore, we show that UFMylation is increased in skeletal muscle biopsies from people living with amyotrophic lateral sclerosis (plwALS). Quantification of UFMylation sites in these biopsies with multiplexed isotopic labeling reveal prominent increases in myosin UFMylation. Our data suggest that in vivo UFMylation is more complex than previously thought.
    Keywords:  CP: cell biology; UFM1; UFMylation; Ubiquitin Fold Modifier 1; ubiquitin-like modification
    DOI:  https://doi.org/10.1016/j.crmeth.2025.101048
  13. Cell Rep. 2025 May 09. pii: S2211-1247(25)00457-7. [Epub ahead of print]44(5): 115686
    UFMylation safeguards human hepatocyte differentiation and liver homeostasis by regulating ribosome dissociation.
    Shuchun Yang, Li Wang, Ran Gao, Yanchang Li, Duo Zhang, Chenxi Wang, Guang Liu, Jie Na, Ping Xu, Xiaoyue Wang, Yuyan Jia, Yue Huang.
      Ribosomal UFMylation contributes to ribosome heterogeneity and is associated with ribosome-associated quality control at the endoplasmic reticulum. However, the specific pathophysiological functions of ribosomal UFMylation remain unknown. In this study, we systematically demonstrate the significance of UFMylation in the differentiation and maturation of hepatocytes using human embryonic stem cell-derived hepatocyte-like cells and liver bud organoids as experimental platforms. We also develop a strategy to identify UFMylated substrates and confirm that RPL26 is a substrate in the liver. Additionally, we discover that mice with the Rpl26 c.395A>G (p.K132R) mutation are more susceptible to steatosis induced by a high-fat diet. Further investigations reveal a key role of CDK5RAP3 and RPL26 UFMylation in regulating ribosome dissociation. Our findings suggest that ribosome UFMylation serves as an important safeguard for liver development and homeostasis and may represent a potential therapeutic target for nonalcoholic fatty liver disease.
    Keywords:  CP: Metabolism; CP: Molecular biology; UFMylation; liver homeostasis; ribosome
    DOI:  https://doi.org/10.1016/j.celrep.2025.115686
  14. Redox Biol. 2025 May 01. pii: S2213-2317(25)00164-8. [Epub ahead of print]84 103651
    Cdc48 plays a crucial role in redox homeostasis through dynamic reshaping of its interactome during early stationary phase.
    Meytal Radzinski, Tal Oppenheim, Ohad Yogev, Adi Levy, Melamed-Book Naomi, Assaf Kacen, Yifat Merbl, Tommer Ravid, Dana Reichmann.
      Most microbial cells on earth predominantly exist in non-proliferating, dormant conditions, such as the stationary state. The stationary phase is a crucial stage during the cellular lifespan, which requires homeostatic rewiring for long-term viability and rapid responses to environmental changes. Here, we show that entry to the stationary phase in yeast is accompanied by increased cytosolic and mitochondrial oxidation, imposing stress on the proteostasis network. We establish a functional link between redox and protein homeostasis, mediated by a key protein quality control member, Cdc48/p97/VCP. Comparative proteomic analysis of post-mitotic yeast cells reveals that while the global proteome remains largely stable during the first stages of stationary phase, the Cdc48 interactome undergoes significant remodeling, including altered interactions with antioxidants and its cofactors Shp1/Ubx1 and Ubx2. To challenge yeast Cdc48's capacity as a redox-switch protein during the early stages of the stationary phase, we utilized redox proteomics to map changes in reversible oxidation modification on Cdc48's cysteines upon entry to the stationary phase. We revealed the temporal and reversible oxidation of Cdc48-Cys115 as a key regulatory event essential for stationary-phase survival and interactome modulation. Cys115-to-serine mutation significantly reduced longevity and increased oxidative stress sensitivity, correlating with disrupted interactions between Cdc48 and antioxidants, and cofactor Shp1, specifically with the phosphorylated form of Shp1. Taken together, these findings identify a new thiol switch protein in the protein degradation pathway, while further defining novel roles for Cdc48 in reshaping the proteome during the yeast stationary phase.
    DOI:  https://doi.org/10.1016/j.redox.2025.103651
  15. Cell Rep Med. 2025 May 06. pii: S2666-3791(25)00206-X. [Epub ahead of print] 102133
    Suppressing proteasome activity enhances sensitivity to actinomycin D in diffuse anaplastic Wilms tumor.
    Patricia D B Tiburcio, Kenian Chen, Lin Xu, Kenneth S Chen.
      Wilms tumor is the most common pediatric kidney cancer, and diffuse anaplastic Wilms tumor is the most chemoresistant subtype. Here, we explore how Wilms tumor cells evade the chemotherapy actinomycin D, which inhibits ribosomal RNA biogenesis. Using ribosome profiling, protein arrays, and a genome-wide knockout screen, we describe how actinomycin D disrupts protein homeostasis and blocks cell-cycle progression. When ribosomal capacity is limited by actinomycin D treatment, anaplastic Wilms tumor cells preferentially translate proteasome components. Next, we find that the proteasome inhibitor bortezomib sensitizes cells to actinomycin D treatment in vitro and prolongs survival in xenograft models. Lastly, increased levels of proteasome components are associated with anaplastic histology and worse prognosis in Wilms tumor patients. In sum, maintaining protein homeostasis is critical for Wilms tumor proliferation, and it can be therapeutically disrupted by blocking protein synthesis or turnover.
    Keywords:  Wilms tumor; actinomycin D; proteasome; protein homeostasis
    DOI:  https://doi.org/10.1016/j.xcrm.2025.102133
  16. Chem Sci. 2025 May 12.
    Discovery and optimisation of a covalent ligand for TRIM25 and its application to targeted protein ubiquitination.
    Katherine A McPhie, Diego Esposito, Jonathan Pettinger, Daniel Norman, Thilo Werner, Toby Mathieson, Jacob T Bush, Katrin Rittinger.
      The tripartite motif (TRIM) family of RING-type E3 ligases catalyses the formation of many different types of ubiquitin chains, and as such, plays important roles in diverse cellular functions, ranging from immune regulation to cancer signalling pathways. Few ligands have been discovered for TRIM E3 ligases, and these E3s are under-represented in the rapidly expanding field of induced proximity. Here we present the identification of a novel covalent ligand for the PRYSPRY substrate binding domain of TRIM25. We employ covalent fragment screening coupled with high-throughput chemistry direct-to-biology optimisation to efficiently elaborate covalent fragment hits. We demonstrate that our optimised ligand enhances the in vitro auto-ubiquitination activity of TRIM25 and engages TRIM25 in live cells. We also present the X-ray crystal structure of TRIM25 PRYSPRY in complex with this covalent ligand. Finally, we incorporate our optimised ligand into heterobifunctional proximity-inducing compounds and demonstrate the in vitro targeted ubiquitination of a neosubstrate by TRIM25.
    DOI:  https://doi.org/10.1039/d5sc01540e
  17. Proc Natl Acad Sci U S A. 2025 May 20. 122(20): e2425061122
    Activation mechanism of small heat shock protein HSPB5 revealed by disease-associated mutants.
    Christopher N Woods, Maria K Janowska, Lindsey D Ulmer, Jasleen Kaur Sidhu, Natalie L Stone, Ellie I James, Miklos Guttman, Matthew F Bush, Rachel E Klevit.
      Found from bacteria to humans, small heat shock proteins (sHSPs) are the least understood protein chaperones. HSPB5 (or αB-crystallin) is among the most widely expressed of the 10 human sHSPs, including in muscle, brain, and eye lens where it is constitutively present at high levels. A high content of disorder in HSPB5 has stymied efforts to uncover how its structure gives rise to function. To uncover its mechanisms of action, we compared human HSPB5 and two disease-associated mutants, R120G and D109H. Expecting to learn how the mutations lead to loss of function, we found instead that the mutants are constitutively activated chaperones while wild-type HSPB5 can transition reversibly between nonactivated (low activity) and activated (high activity) states in response to changing conditions. Techniques that provide information regarding interactions and accessibility of disordered regions revealed that the disordered N-terminal regions (NTR) that are required for chaperone activity exist in a complicated interaction network within HSPB5 oligomers and are sequestered from solvent in nonactivated states. Either mutation or an activating pH change causes rearrangements in the network that expose parts of the NTR, making them more available to bind an aggregating client. Although beneficial in the short-term, failure of the mutants to adopt a state with lower activity and lower NTR accessibility leads to increased coaggregation propensity and, presumably, early cataract. The results support a model where chaperone activity and solubility are modulated through the quasi-ordered NTR and its multiple competing interactions.
    Keywords:  protein aggregation; protein chaperones; small heat shock proteins
    DOI:  https://doi.org/10.1073/pnas.2425061122
  18. Autophagy. 2025 May 14. 1-3
    Visualization of autophagic structures near solid polyQ aggregates reveals how they undermine autophagy.
    Hana Popelka, Daniel J Klionsky.
      Aggregates of polyglutamine (polyQ) repeat extensions are known markers of several, predominantly inherited, neurodegenerative diseases. Removal of polyQ is essential for cellular proteostasis and macroautophagy/autophagy has been proposed to be an important tool in the clearance of polyQ aggregates. The mechanism of recognition and encapsulation of these aggregates within autophagosomes is largely unknown. A study described in this article employed in situ correlative cryo-electron tomography to visualize polyQ aggregates interacting with autophagic compartments. The tomograms revealed that only amorphous polyQ, but not fibrils, are engulfed by double-membrane structures and that SQSTM1/p62 is the receptor involved in recognition of polyQ during autophagy. Solidified amorphous polyQ and subsequent fibrils arrest the normal formation of autophagosomes and impair autophagy. Findings of the study described here have implications for therapies that rely on autophagy in targeting polyQ neurodegeneration.Abbreviation: cryo-CLEM, cryo-correlative light and electron microscopy; cryo-ET, cryo-electron tomography; ER, endoplasmic reticulum; HD, Huntington disease; HTT, huntingtin; polyQ, polyglutamine repeats.
    Keywords:  Amorphous phase; SQSTM1/p62; cryo-electron tomography, fibrillar phase; huntingtin; neurodegeneration
    DOI:  https://doi.org/10.1080/15548627.2025.2503578
  19. J Cell Sci. 2025 May 01. pii: jcs263757. [Epub ahead of print]138(9):
    Quality control of un-imported mitochondrial proteins at a glance.
    Megan Balzarini, John Kim, Hilla Weidberg.
      Mitochondria are metabolic hubs that are essential for cellular homeostasis. Most mitochondrial proteins are translated in the cytosol and imported into the organelle. However, import machineries can become overwhelmed or disrupted by physiological demands, mitochondrial damage or diseases, such as metabolic and neurodegenerative disorders. Impaired import affects mitochondrial function and causes un-imported pre-proteins to accumulate not only in the cytosol but also in other compartments, including the endoplasmic reticulum and nucleus. Quality control pathways have evolved to mitigate the accumulation of these mistargeted proteins and prevent proteotoxicity. In this Cell Science at a Glance article and the accompanying poster, we summarize the fate of un-imported mitochondrial proteins and the compartment-specific quality control pathways that regulate them.
    Keywords:  Mitochondrial protein import; Mitochondrial stress; Protein quality control
    DOI:  https://doi.org/10.1242/jcs.263757
  20. Biophys Rev. 2025 Apr;17(2): 435-447
    Mechanical effect of protein glycosylation on BiP-mediated post-translational translocation and folding in the endoplasmic reticulum.
    Christian A M Wilson, Hilda M Alfaro-Valdés, Merve Kaplan, Cecilia D'Alessio.
      About one-third of the proteins synthesized in eukaryotic cells are directed to the secretory pathway, where close to 70% are being N-glycosylated. N-glycosylation is a crucial modification for various cellular processes, including endoplasmic reticulum (ER) glycoprotein folding quality control, lysosome delivery, and cell signaling. The defects in N-glycosylation can lead to severe developmental diseases. For the proteins to be glycosylated, they must be translocated to the ER through the Sec61 translocon channel, either via co-translationally or post-translationally. N-glycosylation not only could accelerate post-translational translocation but may also enhance protein stability, while protein folding can assist in their movement into the ER. However, the precise mechanisms by which N-glycosylation and folding influence translocation remain poorly understood. The chaperone BiP is essential for post-translational translocation, using a "ratchet" mechanism to facilitate protein entry into the ER. Although research has explored how BiP interacts with protein substrates, there has been less focus on its binding to glycosylated substrates. Here, we review the effect of N-glycosylation on protein translocation, employing single-molecule studies and ensembles approaches to clarify the roles of BiP and N-glycosylation in these processes. Our review explores the possibility of a direct relationship between translocation and a ratchet effect of glycosylation and the importance of BiP in binding glycosylated proteins for the ER quality control system.
    Supplementary Information: The online version contains supplementary material available at 10.1007/s12551-025-01313-x.
    Keywords:  BiP Chaperone; ER translocation; Protein N-glycosylation; Ratchet mechanism
    DOI:  https://doi.org/10.1007/s12551-025-01313-x
  21. Commun Chem. 2025 May 13. 8(1): 146
    Revealing arginine-cysteine and glycine-cysteine NOS linkages by a systematic re-evaluation of protein structures.
    Sophia Bazzi, Sharareh Sayyad.
      Nitrogen-oxygen-sulfur (NOS) linkages act as allosteric redox switches, modulating enzymatic activity in response to redox fluctuations. While NOS linkages in proteins were once assumed to occur only between lysine and cysteine, our investigation shows that these bonds extend beyond the well-studied lysine-NOS-cysteine examples. By systematically analyzing over 86,000 high-resolution X-ray protein structures, we uncovered 69 additional NOS bonds, including arginine-NOS-cysteine and glycine-NOS-cysteine. Our pipeline integrates machine learning, quantum-mechanical calculations, and high-resolution X-ray crystallographic data to systematically detect these subtle covalent interactions and identify key predictive descriptors for their formation. The discovery of these previously unrecognized linkages broadens the scope of protein chemistry and may enable targeted modulation in drug design and protein engineering. Although our study focuses on NOS linkages, the flexibility of this methodology allows for the investigation of a wide range of chemical bonds and covalent modifications, including structurally resolvable posttranslational modifications (PTMs). By revisiting and re-examining well-established protein models, this work underscores how systematic data-driven approaches can uncover hidden aspects of protein chemistry and inspire deeper insights into protein function and stability.
    DOI:  https://doi.org/10.1038/s42004-025-01535-w
  22. Proc Natl Acad Sci U S A. 2025 May 20. 122(20): e2426532122
    Allosteric regulation of UBIAD1 trafficking from ER to Golgi revealed by chemical genetic screening.
    Dong-Jae Jun, Marc M Schumacher, Youngah Jo, Rebecca A Faulkner, Yangyang Yang, Jet Tsien, Tian Qin, Russell A DeBose-Boyd.
      Schnyder corneal dystrophy (SCD) is a rare autosomal dominant condition characterized by the opacification of the cornea owing to the abnormal deposition of cholesterol. SCD-associated mutations have been identified in the gene encoding UbiA prenyltransferase domain-containing protein-1 (UBIAD1), which uses geranylgeranyl pyrophosphate (GGpp) to synthesize the vitamin K2 subtype menaquinone-4 (MK-4). Beyond its enzymatic role, UBIAD1 serves as a key regulator of the endoplasmic reticulum (ER)-localized enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), the rate-limiting enzyme in the mevalonate pathway that produces cholesterol and nonsterol isoprenoids such as GGpp and MK-4. Sterol-induced binding to UBIAD1 inhibits the sterol-accelerated ER-associated degradation (ERAD) of HMGCR to maintain the synthesis of nonsterol isoprenoids under conditions of cholesterol repletion. GGpp dissociates the HMGCR-UBIAD1 complex, triggering maximal ERAD of HMGCR and ER-to-Golgi translocation of UBIAD1. However, SCD-associated UBIAD1 resists this GGpp-induced dissociation and remains sequestered in the ER. ER retention of UBIAD1 leads to inhibition of HMGCR ERAD, promoting increased synthesis and accumulation of cholesterol. Here, chemical genetic screening was utilized to identify molecules that restored Golgi localization of SCD-associated UBIAD1 (N102S) and thereby relieve inhibition of HMGCR ERAD. We found that the chemotherapeutic tyrosine kinase inhibitor Apatinib stimulated ER-to-Golgi transport of both N102S and wild type UBIAD1. This effect required GGpp but was independent of Apatinib's tyrosine kinase inhibition. Apatinib-mediated Golgi transport of UBIAD1 enhanced the ERAD of HMGCR. Photoaffinity labeling studies indicated that Apatinib binds directly to UBIAD1, suggesting that the drug allosterically activates GGpp-induced transport of UBIAD1 from the ER to the Golgi.
    Keywords:  ER-to-Golgi transport; ERAD; cholesterol; endoplasmic reticulum; nonsterol isoprenoids
    DOI:  https://doi.org/10.1073/pnas.2426532122
  23. Nat Commun. 2025 May 13. 16(1): 4418
    TRIM23 mediates cGAS-induced autophagy in anti-HSV defense.
    Dhiraj Acharya, Zuberwasim Sayyad, Helene Hoenigsperger, Maximilian Hirschenberger, Matthew Zurenski, Kannan Balakrishnan, Junji Zhu, Sebastian Gableske, Jiro Kato, Shen-Ying Zhang, Jean-Laurent Casanova, Joel Moss, Konstantin M J Sparrer, Michaela U Gack.
      The cGAS-STING pathway, well-known to elicit interferon (IFN) responses, is also a key inducer of autophagy upon virus infection or other stimuli. Whereas the mediators for cGAS-induced IFN responses are well characterized, much less is known about how cGAS elicits autophagy. Here, we report that TRIM23, a unique TRIM protein harboring both ubiquitin E3 ligase and GTPase activity, is crucial for cGAS-STING-dependent antiviral autophagy. Genetic ablation of TRIM23 impairs autophagic control of HSV-1 infection. HSV-1 infection or cGAS-STING stimulation induces TBK1-mediated TRIM23 phosphorylation at S39, which triggers TRIM23 autoubiquitination and GTPase activity and ultimately elicits autophagy. Fibroblasts from a patient with herpes simplex encephalitis heterozygous for a dominant-negative, kinase-inactivating TBK1 mutation fail to activate autophagy by TRIM23 and cGAS-STING. Our results thus identify the cGAS-STING-TBK1-TRIM23 axis as a key autophagy defense pathway and may stimulate new therapeutic interventions for viral or inflammatory diseases.
    DOI:  https://doi.org/10.1038/s41467-025-59338-5
  24. Nat Chem Biol. 2025 May 14.
    Small-molecule dissolution of stress granules by redox modulation benefits ALS models.
    Hiroyuki Uechi, Sindhuja Sridharan, Jik Nijssen, Jessica Bilstein, Juan M Iglesias-Artola, Satoshi Kishigami, Virginia Casablancas-Antras, Ina Poser, Eduardo J Martinez, Edgar Boczek, Michael Wagner, Nadine Tomschke, António M de Jesus Domingues, Arun Pal, Thom Doeleman, Sukhleen Kour, Eric Nathaniel Anderson, Frank Stein, Hyun O Lee, Xiaojie Zhang, Anatol W Fritsch, Marcus Jahnel, Julius Fürsch, Anastasia C Murthy, Simon Alberti, Marc Bickle, Nicolas L Fawzi, André Nadler, Della C David, Udai B Pandey, Andreas Hermann, Florian Stengel, Benjamin G Davis, Andrew J Baldwin, Mikhail M Savitski, Anthony A Hyman, Richard J Wheeler.
      Neurodegenerative diseases, such as amyotrophic lateral sclerosis, are often associated with mutations in stress granule proteins. Aberrant stress granule condensate formation is associated with disease, making it a potential target for pharmacological intervention. Here, we identified lipoamide, a small molecule that specifically prevents cytoplasmic condensation of stress granule proteins. Thermal proteome profiling showed that lipoamide stabilizes intrinsically disordered domain-containing proteins, including SRSF1 and SFPQ, which are stress granule proteins necessary for lipoamide activity. SFPQ has redox-state-specific condensate dissolving behavior, which is modulated by the redox-active lipoamide dithiolane ring. In animals, lipoamide ameliorates aging-associated aggregation of a stress granule reporter protein, improves neuronal morphology and recovers motor defects caused by amyotrophic lateral sclerosis-associated FUS and TDP-43 mutants. Thus, lipoamide is a well-tolerated small-molecule modulator of stress granule condensation, and dissection of its molecular mechanism identified a cellular pathway for redox regulation of stress granule formation.
    DOI:  https://doi.org/10.1038/s41589-025-01893-5
  25. Sci Adv. 2025 May 16. 11(20): eadp2643
    Interaction networks of SIM-binding groove mutants reveal alternate modes of SUMO binding and profound impact on SUMO conjugation.
    Laura A Claessens, Matty Verlaan-de Vries, Nienke Broekman, Martijn S Luijsterburg, Alfred C O Vertegaal.
      The best-characterized mode of noncovalent SUMO interaction involves binding of a SUMO-interaction motif (SIM) to a conserved binding groove in SUMO. Our knowledge on other types of SUMO interactions is still limited. Using SIM-binding groove SUMO2/3 mutants and mass spectrometry, we identified proteins that bind to SUMO in an alternate manner. Domain-enrichment analysis characterized a group of WD40 repeat domain-containing proteins as SIM-independent SUMO interactors, and we validated direct binding of SEC13 and SEH1L to SUMO in vitro. Using AlphaFold-3 modeling and in vitro mutational analysis, we identified residues in the WD40 domain of SEC13 and SUMO2/3's C terminus involved in the interaction. Furthermore, SIM-binding groove mutants failed to interact with SUMO E3 ligases belonging to the PIAS family, RANBP2, ZNF451, and TOPORS, leading to loss of covalent conjugation to most of SUMO target proteins. Together, our dataset serves as a unique resource and offers valuable insights on the intricacies of the SUMO interactome and SUMO targets.
    DOI:  https://doi.org/10.1126/sciadv.adp2643
  26. J Cell Biol. 2025 Jul 07. pii: e202409196. [Epub ahead of print]224(7):
    RIM and MUNC13 membrane-binding domains are essential for neuropeptide secretion.
    Fiona H Murphy, Adlin Abramian, Remco V Klaassen, Frank Koopmans, Claudia M Persoon, August B Smit, Ruud F Toonen, Matthijs Verhage.
      Neurons release neurotransmitters from synaptic vesicles (SVs) and neuropeptides from dense-core vesicles (DCVs). The presynaptic proteins RIM and MUNC13 play key roles in both pathways. It remains unclear how DCVs are targeted to release sites and whether RIM and MUNC13 are involved in this process. Here, we show that three membrane-binding domains in RIM and MUNC13 regulate DCV exocytosis differently from SV exocytosis. Using neuropeptide secretion assays with single-vesicle resolution and peptidomics analysis of endogenous neuropeptide release in MUNC13/RIM null neurons, we demonstrate that MUNC13 is essential for DCV exocytosis. The RIM N terminus prevents MUNC13 degradation via the proteasome, and inhibiting proteasomal degradation partially rescues DCV exocytosis in RIM's absence. Unlike SV exocytosis, the PIP2-binding RIM C2B domain and MUNC13 C1-C2B polybasic face are redundant for DCV exocytosis, while the lipid-binding MUNC13 C2C domain is crucial. These results show that RIM and MUNC13 synergistically regulate DCV exocytosis through membrane interactions and reveal new mechanistic differences between SV and DCV exocytosis.
    DOI:  https://doi.org/10.1083/jcb.202409196
  27. RNA. 2025 May 13. pii: rna.080561.125. [Epub ahead of print]
    EDC4 C-terminal domain scaffolds P-body assembly and links P-body dynamics to p53 mediated tumor suppression.
    Yu-Hsuan Cheng, Ting-Wen Chen, Wei-Chung Chiang, Jean-Cheng Kuo, Yi-Sheng Ho, Michelle Noble, Chung-Te Chang.
      Processing bodies (P-bodies) are membrane-less organelles in eukaryotic cells that play a central role in mRNA metabolism, including mRNA decay, storage, and translational repression. However, the molecular mechanisms governing their assembly remain incompletely understood. Here, we identify the C-terminal domain of EDC4 as the minimal region required for P-body formation, with residues 1266-1401 driving phase separation and EDC4 condensation. To investigate the functional relevance of P-body integrity, we employed the microprotein Nobody (NBDY) as a selective perturbation tool. Our results revealed that the NBDY 22-41 peptide directly binds the EDC4 C-terminal domain and inhibits its self-association, thereby selectively dissolving P-bodies without affecting the canonical mRNA decay pathway. Using this tool, we further examined the impact of P-body disruption on gene expression. Transcriptome profiling combined with quantitative validation revealed that P-body loss activates the p53 pathway and enhances the stability of associated transcripts. Consistent with these findings, clinical data show that NBDY overexpression is associated with p53 pathway activation in various cancers, and the NBDY 22-41 fragment reduces tumor cell proliferation and invasion, suggesting a potentially complex role of P-body dynamics in cancer biology. Together, our study defines the EDC4 C-terminal domain as a core scaffold for P-body assembly and uncovers a regulatory role of P-body dynamics in p53-mediated gene expression, with potential implications for cancer biology.
    Keywords:  EDC4; NBDY; P-body; mRNA decapping complex; p53 pathway
    DOI:  https://doi.org/10.1261/rna.080561.125
  28. Sci Adv. 2025 May 16. 11(20): eadv1286
    Hsc70-4: An unanticipated mediator of dsRNA internalization in Drosophila.
    Sabrina J Fletcher, Eugenia S Bardossy, Lorena Tomé-Poderti, Thomas Moss, Vanesa Mongelli, Lionel Frangeul, Hervé Blanc, Yann Verdier, Joelle Vinh, Shaeri Mukherjee, Maria-Carla Saleh.
      The small interfering RNA pathway is the primary antiviral defense mechanism in invertebrates and plants. This systemic mechanism relies on the recognition, transport, and internalization of double-stranded RNA (dsRNA). Our aim was to identify cell surface proteins that bind extracellular dsRNA and mediate its internalization in Drosophila cells. We used coimmunoprecipitation coupled with proteomics analysis and found that silencing heat shock cognate protein 70-4 (Hsc70-4), a constitutively expressed heat shock protein, impairs dsRNA internalization. Unexpectedly, despite lacking a predicted transmembrane domain, Hsc70-4 localizes to the cell membrane via lipid interactions. Antibody blocking experiments revealed an extracellular domain on Hsc70-4 that is essential for dsRNA internalization. Intriguingly, this dsRNA-specific binding capacity of Hsc70-4 functions independently of its chaperone activity. These findings not only highlight Hsc70-4 as a previously uncharacterized and essential component in the dsRNA internalization process but also offer promising insights for advancing RNA interference-based technologies to combat pests and vector-borne diseases.
    DOI:  https://doi.org/10.1126/sciadv.adv1286
  29. EMBO J. 2025 May 12.
    Inhibitors of eIF1A-ribosome interaction unveil uORF-dependent regulation of translation initiation and antitumor and antiviral effects.
    Daniel Hayat, Ariel Ogran, Shaked Ashkenazi, Alexander Plotnikov, Roni Oren, Mirie Zerbib, Amir Ben-Shmuel, Rivka Dikstein.
      During translation initiation, eIF1A binds the ribosome through its N- and C-terminal tails, but the functional importance of this temporal interaction in mammalian cells is lacking. Using a high-throughput drug screen targeting eIF1A-RPS10 interaction, we identified inhibitors (1Ais) for eIF1A, RPS10, or both. Applying 1Ais in biochemical assays along specific and global translation experiments, we confirmed known functions of eIF1A and uncovered new roles for both eIF1A and RPS10. Specifically, the eIF1A N-terminal tail (NTT) binding inhibitors revealed the requirement of eIF1A for translation re-initiation. Moreover, a cytosine at position +5 relative to the start codon AUG, located near eIF1A-NTT in the 48S structure, enhances sensitivity to 1Ais, suggesting that the initiating ribosome recognizes a broader AUG context than the conventional Kozak. Additionally, eIF1A-specific 1Ais predominately affect cancer-related pathways. In xenograft models of ovarian cancer, these 1Ais reduced tumor growth without apparent toxicity. Furthermore, inhibition of RPS10, but not eIF1A, modulates a context-dependent regulatory translation initiation at CUG codon of SARS-CoV-2 and impedes infection. Our study underscores 1Ais as effective means to study the role of eIF1A and RPS10 in translation and suggests their targeted inhibition as potential therapies for cancer and viral infections.
    Keywords:  Ovarian Cancer; Rps10; SARS-CoV-2; Translation Reinitiation; eIF1A
    DOI:  https://doi.org/10.1038/s44318-025-00449-6
  30. Nat Methods. 2025 May 13.
    Prediction of protein subcellular localization in single cells.
    Xinyi Zhang, Yitong Tseo, Yunhao Bai, Fei Chen, Caroline Uhler.
      The subcellular localization of a protein is important for its function, and its mislocalization is linked to numerous diseases. Existing datasets capture limited pairs of proteins and cell lines, and existing protein localization prediction models either miss cell-type specificity or cannot generalize to unseen proteins. Here we present a method for Prediction of Unseen Proteins' Subcellular localization (PUPS). PUPS combines a protein language model and an image inpainting model to utilize both protein sequence and cellular images. We demonstrate that the protein sequence input enables generalization to unseen proteins, and the cellular image input captures single-cell variability, enabling cell-type-specific predictions. Experimental validation shows that PUPS can predict protein localization in newly performed experiments outside the Human Protein Atlas used for training. Collectively, PUPS provides a framework for predicting differential protein localization across cell lines and single cells within a cell line, including changes in protein localization driven by mutations.
    DOI:  https://doi.org/10.1038/s41592-025-02696-1
  31. Proc Natl Acad Sci U S A. 2025 May 20. 122(20): e2425812122
    Discovery of an LSD1 PROTAC degrader.
    Amir Hosseini, Xing Qiu, Yan Xiong, Ki Him Nicholas Chiang, Jerrel Catlett, Ines Kaltheuner, Zhijie Deng, Sudipta Ghosh, Yang Shi, Jian Jin.
      Aberrant expression of lysine-specific demethylase 1 (LSD1) has been implicated in various cancers, including acute myeloid leukemia (AML). Recent studies have revealed both catalytic and noncatalytic oncogenic functions of LSD1, which cannot be effectively addressed by traditional small-molecule inhibitors. Therefore, to remove LSD1 and mitigate its oncogenic activity, we utilized the proteolysis-targeting chimera (PROTAC) approach and developed an LSD1 PROTAC degrader MS9117, which recruits the E3 ligase cereblon (CRBN). MS9117 induces LSD1 degradation in a concentration-, time-, CRBN-, and proteasome-dependent manner. Importantly, MS9117 effectively degrades LSD1 and demonstrates superior antiproliferative effects in AML cells, compared to the existing pharmacological LSD1 inhibitors. Furthermore, MS9117 also sensitized nonacute promyelocytic leukemia AML cells to all-trans retinoic acid treatment. Moreover, we developed two negative controls of MS9117, MS9117N1 and MS9117N2, which do not degrade LSD1 or inhibit leukemia cell growth, further confirming the mechanism of action of MS9117. Overall, MS9117 serves as a valuable chemical tool and a potential therapeutic to target both the catalytic and scaffolding functions of LSD1. With several LSD1 inhibitors already in clinical development, the LSD1 degraders such as MS9117 offer an additional option for future clinical studies.
    Keywords:  AML; ATRA; LSD1; PROTAC; degrader
    DOI:  https://doi.org/10.1073/pnas.2425812122
  32. J Am Chem Soc. 2025 May 15.
    Reactive Oxygen Species-Instructed Supramolecular Assemblies Enable Bioorthogonally Activatable Protein Degradation for Pancreatic Cancer.
    Qingxin Yao, Ziyang Wu, Jiaan Li, Xiaoqian Hu, Hanlin Xu, Xingyu Jiang, Yuan Gao.
      Proteolysis-Targeting Chimeras (PROTACs) represent a transformative therapeutic platform for targeted protein degradation across diverse disease indications. However, their potent catalytic activity in normal tissues raises significant concerns regarding off-target toxicity. Here, we present a novel supramolecular self-assembly platform for the bioorthogonal control of PROTAC prodrug activation, enabling tumor-specific protein degradation with minimized systemic toxicity. By exploiting the overproduction of reactive oxygen species (ROS) in pancreatic cancer cells, the supramolecular self-assembly approach selectively accumulates bioorthogonal reaction triggers within the targeted malignant cells, which subsequently facilitates the spatiotemporally controlled activation of the bioorthogonally caged PROTAC. This tumor-selective activation mechanism demonstrates enhanced degradation efficiency in pancreatic cancer cells compared to normal cells. In vivo studies reveal potent tumor growth inhibition with complete preservation of major organ histology, confirming the therapeutic index enhancement achieved through a controllable activation strategy. This biomimetic activation platform establishes a generalizable framework for safer PROTAC-based therapies by integrating tumor-specific microenvironmental cues with bioorthogonal reaction engineering.
    DOI:  https://doi.org/10.1021/jacs.5c04857
  33. Cell Rep. 2025 May 09. pii: S2211-1247(25)00418-8. [Epub ahead of print]44(5): 115647
    Beta cells intrinsically sense and limit their secretory activity via mTORC1-RhoA signaling.
    Saar Krell, Amit Hamburg, Ofer Gover, Kfir Molakandov, Gil Leibowitz, Kfir Sharabi, Michael D Walker, Aharon Helman.
      Precise regulation of insulin secretion by pancreatic β cells is essential to prevent excessive insulin release. Here, we show that the nutrient sensor mechanistic Target of Rapamycin Complex 1 (mTORC1) is rapidly activated by glucose in β cells via the insulin secretion machinery, positioning mTORC1 as a sensor of β cell activity. Acute pharmacological inhibition of mTORC1 during glucose stimulation enhances insulin release, suggesting that mTORC1 acts as an intrinsic feedback regulator that restrains insulin secretion. Phosphoproteomic profiling reveals that mTORC1 modulates the phosphorylation of proteins involved in actin remodeling and vesicle trafficking, with a prominent role in the RhoA-GTPase pathway. Mechanistically, mTORC1 promotes RhoA activation and F-actin polymerization, limiting vesicle movement and dampening the second phase of insulin secretion. These findings identify a glucose-mTORC1-RhoA signaling axis that forms an autonomous feedback loop to constrain insulin exocytosis, providing insight into how β cells prevent excessive insulin release and maintain metabolic balance.
    Keywords:  CP: Metabolism; CP: Molecular biology; RhoA-GTPase; Torin-1; actin remodeling; activity sensor; autonomous regulation; insulin secretion; mTORC1; negative feedback loop; pancreatic β cell; rapamycin
    DOI:  https://doi.org/10.1016/j.celrep.2025.115647
  34. J Cell Sci. 2025 May 16. pii: jcs.263679. [Epub ahead of print]
    Glutamine modulates stress granule formation in cancer cells through core RNA-binding proteins.
    Gabriel P Faber, Gilad Gross, Oz Mualem, Matan Y Avivi, Hiba Waldman Ben-Asher, Orly Yaron, Orit Shefi, Rakefet Ben-Yishay, Dana Ishay-Ronen, Yaron Shav-Tal.
      Cytoplasmic stress granules (SGs) induced by various stresses have been linked to cancer and other disorders. Which active energy pathways are required for SG formation remains unclear. We used nutrient deprivation to show that glutamine is the sole amino acid source governing whether cancer cells form SGs. Metabolic profiling revealed the essential functions of glutamine and glucose in SG formation under limiting metabolic conditions. Providing glutamine during metabolic stress restored ATP levels in cancer cells and revived many essential gene expression patterns. Myc, a known regulator of the shift between glucose and glutamine metabolism, showed increased expression as cells moved to glutamine uptake. Inhibition of MYC prevented SG formation even with glutamine present and increased cell death after arsenite exposure. The RNA-binding proteins G3BP1/2 were required for glutamine utilization, with G3BP1/2 knockout cells displaying a heavier reliance on glucose, yielding reduced cell survival and inability to properly utilize glutamine. Altogether, we show that cancer cells require glutamine for SG formation under nutrient deprivation, and its absence reduces cell survival, lowering ATP levels below an energy threshold required for SG formation.
    Keywords:  Glutamine; Myc; RNA FISH; Stress granules
    DOI:  https://doi.org/10.1242/jcs.263679
  35. Mol Syst Biol. 2025 May 12.
    The solute carrier superfamily interactome.
    Fabian Frommelt, Rene Ladurner, Ulrich Goldmann, Gernot Wolf, Alvaro Ingles-Prieto, Eva Lineiro-Retes, Zuzana Gelová, Ann-Katrin Hopp, Eirini Christodoulaki, Shao Thing Teoh, Philipp Leippe, Brianda L Santini, Manuele Rebsamen, Sabrina Lindinger, Iciar Serrano, Svenja Onstein, Christoph Klimek, Barbara Barbosa, Anastasiia Pantielieieva, Vojtech Dvorak, Thomas J Hannich, Julian Schoenbett, Gilles Sansig, Tamara A M Mocking, Jasper F Ooms, Adriaan P IJzerman, Laura H Heitman, Peter Sykacek, Juergen Reinhardt, André C Müller, Tabea Wiedmer, Giulio Superti-Furga.
      Solute carrier (SLC) transporters form a protein superfamily that enables transmembrane transport of diverse substrates including nutrients, ions and drugs. There are about 450 different SLCs, residing in a variety of subcellular membranes. Loss-of-function of an unusually high proportion of SLC transporters is genetically associated with a plethora of human diseases, making SLCs a rapidly emerging but challenging drug target class. Knowledge of their protein environment may elucidate the molecular basis for their functional integration with metabolic and cellular pathways and help conceive pharmacological interventions based on modulating proteostatic regulation. We aimed at obtaining a global survey of the SLC-protein interaction landscape and mapped the protein-protein interactions of 396 SLCs by interaction proteomics. We employed a functional assessment based on RNA interference of interactors in combination with measurement of protein stability and localization. As an example, we detail the role of a SLC16A6 phospho-degron and the contributions of PDZ-domain proteins LIN7C and MPP1 to the trafficking of SLC43A2. Overall, our work offers a resource for SLC-protein interactions for the scientific community.
    Keywords:  AP-MS; Protein–protein Interactions; Proteostasis; SLC Superfamily; Trafficking
    DOI:  https://doi.org/10.1038/s44320-025-00109-1
This page is being maintained by Thomas Krichel. It was last updated on 2025‒05‒18 at 7:48.