bims-proteo 2025-08-24 papers

bims-proteo

Biomed News

on Proteostasis

Issue of 2025–08–24
twelve papers selected by
Eric Chevet, INSERM

Proteostasis and Unfolded Protein Response Dynamics in Human Neuron and Mouse Glia Co-culture Reveal Cell-Specific Aging Responses.
FICD (FIC Domain Protein Adenylyl Transferase) Deficiency Protects Mice From Hypertrophy-Induced Heart Failure and Promotes BiP (Binding Immunoglobulin Protein) -Mediated Activation of the Unfolded Protein Response and Endoplasmic Reticulum-Selective Autophagy in Cardiomyocytes.
Degradation of Abl Utilizing an Immunoproteasome N-Degron Prodrug.
Edge curvature drives endoplasmic reticulum reorganization and dictates epithelial migration mode.
Transcriptomic Profiling of Small Molecule Atf6 Modulators in the Retina.
ER-resident CCDC134 safeguards TLR4 maturation by maintaining gp96 stability.
The cysteine-rich domain of SEP15, a selenoprotein co-chaperone of the ER chaperone, UDP-glucose:glycoprotein glucosyltransferase, adopts a novel fold.
Investigational eIF2B activator DNL343 modulates the integrated stress response in preclinical models of TDP-43 pathology and individuals with ALS in a randomized clinical trial.
Site-Specific Nanobody Inhibitors of the Proteasomal Deubiquitinase UCH37.
Multiple golgins are required to support extracellular matrix secretion, modification, and assembly.
ESCRT-III is recruited by human herpesvirus 6A nuclear egress complex to promote nuclear egress of the nucleocapsid.
SEC61B regulates calcium flux and platelet hyperreactivity in diabetes.

bioRxiv. 2025 Aug 13. pii: 2025.08.11.669714. [Epub ahead of print]

Proteostasis and Unfolded Protein Response Dynamics in Human Neuron and Mouse Glia Co-culture Reveal Cell-Specific Aging Responses.

Lea A Barny, Sarah K Garcia, Aiden J Houcek, Burak Uzay, Minsoo Kim, Ege T Kavalali, Lars Plate.

Proteostasis, or protein homeostasis, is a tightly regulated network of cellular pathways essential for maintaining proper protein folding, trafficking, and degradation. Neurons are particularly vulnerable to proteostasis collapse due to their post-mitotic and long-lived nature and thus represent a unique cell type to understand the dynamics of proteostasis throughout development, maturation, and aging. Here, we utilized a dual-species co-culture model of human excitatory neurons and mouse glia to investigate cell type- specific, age-related changes in the proteostasis network using data-independent acquisition (DIA) LC-MS/MS proteomics. We quantified branch-specific unfolded protein response (UPR) activation by monitoring curated effector proteins downstream of the ATF6, IRE1/XBP1s, and PERK pathways, enabling a comprehensive, unbiased evaluation of UPR dynamics during neuronal aging. Species-specific analysis revealed that aging neurons largely preserved proteostasis, although they showed some signs of collapse, primarily in ER-to-Golgi transport mechanisms. However, these changes were accompanied by upregulation of proteostasis-related machinery and activation of the ATF6 branch, as well as maintenance of the XBP1s and PERK branches of the UPR with age. In contrast, glia exhibited broad downregulation of proteostasis factors and UPR components, independent of neuronal presence. Furthermore, we quantified stimulus-specific modulation of select UPR branches in aged neurons exposed to pharmacologic ER stressors. These findings highlight distinct, cell-type-specific stress adaptations during aging and provide a valuable proteomic resource for dissecting proteostasis and UPR regulation in the aging brain.
Significance: Understanding how the unfolded protein response (UPR) and proteostasis network change with age is often studied in model organisms, where pathways are assessed across mixed cell types. Such systems can obscure cell-type-specific regulation. Here, we evaluate age-associated remodeling of the UPR and proteostasis network in a dual-species co-culture of human neurons and mouse glia using DIA proteomics. This approach enables species-specific proteomic profiling without physical separation, supported by a customizable data analysis pipeline. We show that neurons and glia exhibit divergent age-related responses, with neurons maintaining adaptive proteostasis and glia showing broader declines. The analytical framework presented here supports future studies to uncover additional cell-type-specific aging phenotypes or to probe the effects of pharmacologic or physical manipulation of biological systems.

DOI: https://doi.org/10.1101/2025.08.11.669714
J Am Heart Assoc. 2025 Aug 19. 14(16): e040192

FICD (FIC Domain Protein Adenylyl Transferase) Deficiency Protects Mice From Hypertrophy-Induced Heart Failure and Promotes BiP (Binding Immunoglobulin Protein) -Mediated Activation of the Unfolded Protein Response and Endoplasmic Reticulum-Selective Autophagy in Cardiomyocytes.

Shannon M Lacy, Rebecca J Taubitz, Nicholas D Urban, Samantha N Turowski, Eric N Primack, Eric D Smith, Adam S Helms, Daniel E Michele, Matthias C Truttmann.

   BACKGROUND: Cardiomyocytes require the HSP70 (heat shock protein 70) chaperone BiP (binding immunoglobulin protein) to maintain proteostasis in the endoplasmic reticulum (ER) following cardiac stress. The adenylyl transferase FICD (FIC domain protein adenylyl transferase) is increasingly recognized to regulate BiP activity through the posttranslational addition of an adenosine monophosphate moiety to BiP surface residues. However, the physiological impact of FICD-mediated BiP regulation in the context of cardiovascular health is unknown.
METHODS: We assessed 6-month and 12-month-old wild-type and FICD knockout mice in a transverse aortic constriction hypertrophy paradigm. We determined cardiac function and injury using echocardiography, histological stainings, and biochemical approaches. In complementary assays, we used isolated neonatal wild-type and FICD knockout cardiomyocytes and cardiac fibroblasts to quantitatively assess cell-type specific adaptations in proteostasis and ER stress responses.
RESULTS: We find that FICD deficiency prevents pressure overload-associated heart failure, hypertrophy, and fibrosis. At a cellular level, we observe that FICD-mediated BiP AMPylation blunts the induction of the unfolded protein response and impairs BiP interaction with FAM134B, an ER-selective autophagy receptor, thus limiting ER-selective autophagy induction under stress. In contrast, FICD loss significantly increases BiP-dependent unfolded protein response induction and ER-selective autophagy in stressed cardiomyocytes. We also uncover cell type-specific consequences of FICD activity in response to ER stress, positioning FICD as a critical proteostasis regulator in cardiac tissue.
CONCLUSIONS: Our results highlight a novel regulatory paradigm controlling stress resilience in cardiomyocytes and offer a rationale to consider FICD as a therapeutic target to treat cardiac hypertrophy.

Keywords:  BiP; ER; FICD; hypertrophy; proteostasis

DOI:  https://doi.org/10.1161/JAHA.124.040192
J Med Chem. 2025 Aug 19.

Degradation of Abl Utilizing an Immunoproteasome N-Degron Prodrug.

Cody A Loy, Timothy J Harris, Shawn E Vinogradsky, Darci J Trader.

Targeted protein degradation is an emerging therapeutic strategy that leverages the cell's natural protein clearance pathways to eliminate proteins of interest (POIs). One common approach involves bifunctional molecules that link an E3 ligase ligand to a POI ligand. However, these large molecules often suffer from poor cell permeability, and expanding ligands to new E3 ligases remains a challenge. As an alternative, we explored the use of N-degrons, single amino acid signals attached to small molecules, to recruit E3 ligases and induce POI ubiquitination and degradation. To enhance tissue selectivity and stability, we developed a caged N-degron degrader incorporating a short peptide sequence recognized by the immunoproteasome (iCP). Using this method, we targeted the Abelson proto-oncogene (Abl). Abl was effectively degraded in cells expressing the iCP, and prolonged degradation at lower concentrations was observed. These findings demonstrate that caging N-degrons offers a promising strategy for improving targeted protein degradation approaches through this mechanism.

DOI: https://doi.org/10.1021/acs.jmedchem.5c01168
Nat Cell Biol. 2025 Aug 18.

Edge curvature drives endoplasmic reticulum reorganization and dictates epithelial migration mode.

Simran Rawal, Pradeep Keshavanarayana, Diya Manoj, Purnati Khuntia, Sanak Banerjee, Basil Thurakkal, Rituraj Marwaha, Fabian Spill, Tamal Das.

From single-cell extrusion to centimetre-sized wounds, epithelial gaps of various sizes and geometries appear across organisms. Their closure involves two orthogonal modes: lamellipodial crawling at convex edges and purse string-like movements at concave edges. The mechanisms driving this curvature-dependent migration remain unclear. Here we perform an intracellular cartography to reveal that in both micropatterned and naturally arising gaps, the endoplasmic reticulum (ER) undergoes edge curvature-dependent morphological reorganizations, forming tubules at convex edges and sheets at concave edges. This reorganization depends on cytoskeleton-generated protrusive and contractile forces. Mathematical modelling reveals that these morphologies minimize strain energy under their respective geometric regime. Functionally, ER tubules at the convex edge favour perpendicularly oriented focal adhesions, supporting lamellipodial crawling, while ER sheets at the concave edge favour parallelly oriented focal adhesions, supporting purse string-like movements. Altogether, ER emerges as a central mechanotransducer, integrating signals from cytoskeletal networks to orchestrate two orthogonal modes of cell migration.

DOI: https://doi.org/10.1038/s41556-025-01729-3
FASEB J. 2025 Aug 31. 39(16): e70940

Transcriptomic Profiling of Small Molecule Atf6 Modulators in the Retina.

Monica Sophia Diaz-Aguilar, Angela Galdamez, Leon Chea, Masako Le, Soyoung Park, Hyejung Min, Eun-Jin Lee, Jonathan H Lin.

  The Unfolded Protein Response (UPR) maintains endoplasmic reticulum (ER) homeostasis and is essential for retinal health. Activating Transcription Factor 6 (ATF6) controls a key UPR branch and upregulates genes that mitigate ER stress. Small molecule modulators of ATF6 have been characterized in cell culture models that increase or decrease the amount of the cleaved, transcriptional activator domain of ATF6 generated from the full-length precursor. However, the effects of these small molecule ATF6 modulators remain unclear in vivo, in part because of the lack of antibodies that robustly detect the cleaved, activated form of ATF6 in model organisms like mice. Here, we used targeted RNA sequencing (RNA-seq) to assess the transcriptional response to intraocular delivery of Ceapin-A7 (an ATF6 inhibitor) and AA147 (an ATF6 activator) in the mouse retina. Using this strategy, we demonstrate that Ceapin-A7 significantly suppressed ATF6 transcriptional targets, whereas AA147 induced ATF6-regulated genes in retinal tissue of the eye. We also show that neither small molecule ATF6 modulator caused retinal cell death, compromised vision, or triggered ER stress by histology, functional testing, and transcriptional analysis. These results identify a transcriptional strategy to sensitively detect and quantify Ceapin-A7 and AA147 modulation of ATF6 in vivo. These findings also identify nontoxic conditions for further in vivo evaluation of small molecule ATF6 modulators in mouse vision loss disease models linked to ER stress.

Keywords:  RNA‐seq; activating transcription factor 6; endoplasmic reticulum; retina; transcriptome; unfolded protein response

DOI:  https://doi.org/10.1096/fj.202501459R
Proc Natl Acad Sci U S A. 2025 Aug 26. 122(34): e2512154122

ER-resident CCDC134 safeguards TLR4 maturation by maintaining gp96 stability.

Yang Bai, Chao Zhang, Hao Liu, Fan Deng, Zeyu Wu, Wanyan Deng, Zengzhang Zheng, Rui Min, Shenglin Mei, He Kang, Huiqing Yu, Youdong Pan, Judy Lieberman, Jingxia Zhao, Xing Liu.

  Toll-like receptor 4 (TLR4), a pattern-recognition receptor located on the plasma membrane, senses extracellular danger signals to initiate inflammatory immune responses. It is initially synthesized in the endoplasmic reticulum (ER), undergoes N-linked glycosylation, and is subsequently transported to the Golgi before ultimately reaching the plasma membrane. However, the mechanisms underlying the processing and maturation of TLR4 in the ER remain elusive. Through whole genome-wide CRISPR screening, CCDC134 was identified as a critical and essential factor for TLR4-dependent inflammatory response. Localization of CCDC134 in the ER lumen rather than its exosome-mediated secretion is required for its role in TLR4 signaling. Loss of CCDC134 results in the retention of TLR4 in the ER for subsequent ER-associated degradation, and thus blockade of TLR4 maturation and plasma membrane trafficking. Defects in TLR4 processing and maturation in the ER in CCDC134-depleted cells are caused by aberrant hyperglycosylation and destabilization of glycoprotein 96 (gp96), a key chaperone of TLR4. These results suggest that CCDC134 controls gp96 glycosylation to facilitate TLR4 maturation in the ER.

Keywords:  CCDC134; TLR4; gp96; inflammation

DOI:  https://doi.org/10.1073/pnas.2512154122
bioRxiv. 2025 Aug 13. pii: 2025.08.11.669745. [Epub ahead of print]

The cysteine-rich domain of SEP15, a selenoprotein co-chaperone of the ER chaperone, UDP-glucose:glycoprotein glucosyltransferase, adopts a novel fold.

Robert V Williams, Kevin P Guay, Owen Hurlbut Lesk, Daniel N Hebert, Lila M Gierasch.

Proteins targeted to the secretory pathway are involved in a myriad of biological processes but can only do so when properly folded. Within the endoplasmic reticulum, glycoprotein folding is regulated by the enzyme UDP-glucose:glycoprotein glucosyltransferase (UGGT) and its oxidoreductase partner, the 15-kDa selenoprotein (SEP15 aka SELENOF). The interaction between these two chaperones is poorly understood, limiting understanding of their function. SEP15 is comprised of two domains, a C-terminal thioredoxin-like domain, the structure of which has been reported (PDB 2A4H), and an approximately 50-residue long N-terminal cysteine-rich domain (CRD), of unknown structure. Here, we use a combination of AlphaFold structural predictions and NMR spectroscopy to elucidate the structure of the SEP15 CRD, which mediates the interaction with UGGT. These data reveal that this domain forms a previously undescribed helical fold stabilized by three disulfide bonds between residues C10-C42, C21-C43, and C24-C39. Furthermore, our results validate our reported model of the UGGT/SEP15 complex and lay the foundation for future studies of its interaction with glycoprotein substrates.

DOI: https://doi.org/10.1101/2025.08.11.669745
Nat Commun. 2025 Aug 18. 16(1): 7690

Investigational eIF2B activator DNL343 modulates the integrated stress response in preclinical models of TDP-43 pathology and individuals with ALS in a randomized clinical trial.

Brittany N Flores, Seungyoon B Yu, Isaac V Cohen, Melania H Fanok, Wei Luan, Romeo D Maciuca, Linus D Sun, Richard M Tsai, Maurits Vissers, Lars Smits, Tommy M Bunte, Anna Bakardjiev, Srijana Balasundar, Meredith E K Calvert, Marcus Y Chin, Sarah K Dobbins, William E Dowdle, Meng Fang, Jules A A C Heuberger, Connie L Ha, Fen Huang, Takashi Miyamoto, Maksim Osipov, Lidia Madrid San Martin, Katie Saund, David Tatarakis, Anthony Q Vu, Chenling Xiong, Gene W Yeo, Geert Jan Groeneveld, Leonard H van den Berg, Shyeilla Dhuria, Anthony A Estrada, Danna Jennings, Thomas Sandmann, Carole Ho, Kimberly Scearce-Levie, Ernie Yulyaningsih, Adam K Walker, Gilbert Di Paolo, Lesley A Kane, Matthew D Troyer, Joseph W Lewcock.

Neuronal TDP-43 aggregates are a hallmark ALS pathology. The integrated stress response (ISR) occurs downstream of TDP-43 pathology and may promote neurodegeneration. Here we demonstrate that a CNS penetrant small molecule eIF2B activator inhibits the ISR in cellular models of ALS and the brain of an inducible mouse model of TDP-43 pathology, where it transiently slowed progression of locomotor deficits and neurodegeneration. ISR activation was observed in ALS patient spinal cord and CSF. The investigational drug DNL343 was advanced into Phase 1 and Phase 1b randomized, double-blind, placebo-controlled trials in healthy and ALS participants, respectively (NCT04268784/NCT05006352); the primary objective in both studies was to investigate the safety and tolerability DNL343. DNL343 demonstrated a half-life supporting once-daily dosing and showed extensive CSF distribution. DNL343 was generally well tolerated and reduced ISR biomarkers in peripheral blood mononuclear cells and CSF of ALS participants. Therefore, DNL343 is a useful investigational drug to explore the effects of ISR inhibition in ALS models and individuals with neurological diseases.

DOI: https://doi.org/10.1038/s41467-025-63031-y
bioRxiv. 2025 Aug 13. pii: 2025.08.11.669711. [Epub ahead of print]

Site-Specific Nanobody Inhibitors of the Proteasomal Deubiquitinase UCH37.

Yanfeng Li, Lin Hui Chang, Rishi Patel, Elizaveta I Shestoperova, Jiale Du, Chittaranjan Das, Eric R Strieter.

Dysregulation of the ubiquitin (Ub) proteasome system (UPS) is linked to numerous human diseases, making its components, particularly deubiquitinases (DUBs), attractive therapeutic targets. UCH37 (also known as UCHL5), a proteasomal DUB, plays roles in protein degradation, DNA repair, and transcription and is overexpressed in several cancers. Despite its relevance, the precise functions of UCH37 within the proteasome and the INO80 chromatin remodeling complex remain unclear. Current small-molecule inhibitors exhibit broad, off-target effects, limiting their utility both therapeutically and as probes for UCH37 function. Here, we report the development of highly potent, selective nanobody-based inhibitors for UCH37. Using yeast surface display, we generated nanobodies targeting distinct Ub-binding sites on UCH37, with binding specificity confirmed by X-ray crystallography and hydrogen-deuterium exchange mass spectrometry. We find that the nanobody targeting the canonical Ub binding site serves as a pan-UCH37 inhibitor, while the one binding to the K48 chain-specific site selectively inhibits Ub chain debranching. Expression of this Ub chain debranching-specific nanobody in cells enabled identification of proteins whose degradation depends on UCH37. This work provides novel tools for investigating the biological functions of UCH37 and lays the groundwork for evaluating its potential as a therapeutic target.

DOI: https://doi.org/10.1101/2025.08.11.669711
J Cell Biol. 2025 Oct 06. pii: e202411167. [Epub ahead of print]224(10):

Multiple golgins are required to support extracellular matrix secretion, modification, and assembly.

George Thompson, Anna Hoyle, Philip A Lewis, M Esther Prada-Sanchez, Joe Swift, Kate Heesom, Martin Lowe, David Stephens, Nicola L Stevenson.

The secretion of extracellular matrix (ECM) proteins is vital to the maintenance of tissue health. One major control point of this process is the Golgi apparatus, whose dysfunction causes numerous connective tissue disorders. We therefore sought to investigate the role of two Golgi organizing proteins, GMAP210 and Golgin-160, in ECM secretion. CRISPR knockout of either protein had distinct impacts on Golgi organization, with Golgin-160 knockout causing Golgi fragmentation and vesicle accumulation, and GMAP210 loss leading to cisternal fragmentation, dilation, and the accumulation of tubulovesicular structures. Both golgins were required for fibrillar collagen organization and glycosaminoglycan synthesis suggesting nonredundant functions in these processes. Furthermore, proteomics analysis revealed both shared and golgin-specific changes in the secretion of ECM proteins. We therefore propose that golgins are collectively required to create the correct physical-chemical space to support efficient ECM protein secretion, modification, and assembly. This is the first time that Golgin-160 has been shown to be required for ECM secretion.

DOI: https://doi.org/10.1083/jcb.202411167
J Virol. 2025 Aug 19. e0084425

ESCRT-III is recruited by human herpesvirus 6A nuclear egress complex to promote nuclear egress of the nucleocapsid.

Aila Gulijiahani, Jun Arii, Vuk Isakovic, Jing Rin Huang, Yasushi Kawaguchi, Yasuko Mori.

  Herpesviruses replicate their genomes and package them into capsids within the host cell nucleus. These capsids must then translocate from the nucleus to the cytoplasm through a process designated nuclear egress. The virus-encoded nuclear egress complex (NEC), consisting of a nuclear matrix protein and a nuclear membrane protein, plays a crucial role in this process. Although the role of NEC for nucleo-cytoplasmic transport of capsids is conserved in Herpesviridae, some of the binding partners of the NEC components are specific for individual viruses. The NECs of alpha- and gammaherpesviruses recruit the Endosomal Sorting Complex Required for Transport III (ESCRT-III) to the inner nuclear membrane for efficient nuclear egress of capsids. In contrast, the role of ESCRT-III for nuclear egress of betaherpesviruses, including human cytomegalovirus (HCMV) and human herpesvirus 6A (HHV-6A), has not been elucidated. Here, we show that ESCRT-III is recruited to the nuclear rim in cells expressing the NEC of HCMV or of HHV-6A. Inhibition of ESCRT-III impaired HHV-6A replication and nuclear egress of the capsids. Mechanistically, ESCRT-III adaptor ALIX interacts with HHV-6A NEC membrane protein U34 and thus contributes to HHV-6A replication. From these observations, we conclude that, like at least some alpha- and gammaherpesviruses, HHV-6A NEC recruits ESCRT-III through ALIX to promote viral capsid nuclear egress.IMPORTANCEESCRT-III performs reverse-topology scission involved in many diverse cellular processes, including cytokinesis, endosome maturation, autophagy, membrane repair, and viral budding. Nucleo-cytoplasmic transport of herpesvirus capsids requires scission at the inner nuclear membrane. In alpha- and gammaherpesviruses, this process requires ESCRT-III, but it is not known whether this is also the case for betaherpesviruses. Here, we show that ESCRT-III is also important for nuclear egress of capsids of the betaherpesvirus human herpesvirus 6A. These results imply that ESCRT-III-mediated inner nuclear membrane scission is a conserved feature in the virion maturation process of Herpesviridae. Our findings thus suggest that ESCRT-III is a potential therapeutic target also for betaherpesvirus infections.

Keywords:  ESCRT-III; HHV-6; herpesviruses; nuclear egress

DOI:  https://doi.org/10.1128/jvi.00844-25
J Clin Invest. 2025 Aug 15. pii: e184597. [Epub ahead of print]135(16):

SEC61B regulates calcium flux and platelet hyperreactivity in diabetes.

Yvonne X Kong, Rajan Rehan, Cesar L Moreno, Søren Madsen, Yunwei Zhang, Huiwen Zhao, Miao Qi, Callum B Houlahan, Siân P Cartland, Declan Robertshaw, Vincent Trang, Frederick Jun Liang Ong, Michael Liu, Edward Cheng, Imala Alwis, Alexander Dupuy, Michelle Cielesh, Kristen C Cooke, Meg Potter, Jacqueline Stöckli, Grant Morahan, Maggie L Kalev-Zylinska, Matthew T Rondina, Sol Schulman, Jean Y H Yang, G Gregory Neely, Simone M Schoenwaelder, Shaun P Jackson, David E James, Mary M Kavurma, Samantha L Hocking, Stephen M Twigg, James C Weaver, Mark Larance, Freda H Passam.

  Platelet hyperreactivity increases the risk of cardiovascular thrombosis in diabetes and failure of antiplatelet drug therapies. Elevated basal and agonist-induced calcium flux is a fundamental cause of platelet hyperreactivity in diabetes; however, the mechanisms responsible for this remain largely unknown. Using a high-sensitivity, unbiased proteomic platform, we consistently detected over 2,400 intracellular proteins and identified proteins that were differentially released by platelets in type 2 diabetes. We identified that SEC61 translocon subunit β (SEC61B) was increased in platelets from humans and mice with hyperglycemia and in megakaryocytes from mice with hyperglycemia. SEC61 is known to act as an endoplasmic reticulum (ER) calcium leak channel in nucleated cells. Using HEK293 cells, we showed that SEC61B overexpression increased calcium flux into the cytosol and decreased protein synthesis. Concordantly, platelets in hyperglycemic mice mobilized more calcium and had decreased protein synthesis. Platelets in both humans and mice with hyperglycemia had increased ER stress. ER stress induced the expression of platelet SEC61B and increased cytosolic calcium. Inhibition of SEC61 with anisomycin decreased platelet calcium flux and inhibited platelet aggregation in vitro and in vivo. These studies demonstrate the existence of a mechanism whereby ER stress-induced upregulation of platelet SEC61B leads to increased cytosolic calcium, potentially contributing to platelet hyperreactivity in diabetes.

Keywords:  Calcium channels; Cardiology; Cell biology; Hematology; Platelets; Proteomics

DOI:  https://doi.org/10.1172/JCI184597