bims-proteo 2024-10-27 papers

bims-proteo

Biomed News

on Proteostasis

Issue of 2024‒10‒27
thirty papers selected by
Eric Chevet, INSERM

Recruitment of autophagy initiator TAX1BP1 advances aggrephagy from cargo collection to sequestration.
Nuclear proteasomes as a backup for autophagy: interconnected proteostasis pathways.
Intrinsically disordered region amplifies membrane remodeling to augment selective ER-phagy.
EGF-mediated Golgi dynamics and cell migration require CARP2.
MetaDegron: multimodal feature-integrated protein language model for predicting E3 ligase targeted degrons.
Enhanced cell stress response and protein degradation capacity underlie artemisinin resistance in Plasmodium falciparum.
Protocol to identify E3 ligases amenable to biodegraders using a cell-based screening.
Fluorescent Reporters, Imaging, and Artificial Intelligence Toolkits to Monitor and Quantify Autophagy, Heterophagy, and Lysosomal Trafficking Fluxes.
A protein phosphatase 1 specific phosphatase targeting peptide (PhosTAP) to identify the PP1 phosphatome.
Discovery and design of molecular glue enhancers of CDK12-DDB1 interactions for targeted degradation of cyclin K.
Mlf mediates proteotoxic response via formation of cellular foci for protein folding and degradation in Giardia.
Discovery of small molecules for autophagy-lysosome degradation of immune checkpoint proteins.
A structurally informed human protein-protein interactome reveals proteome-wide perturbations caused by disease mutations.
The G1-S transition is promoted by Rb degradation via the E3 ligase UBR5.
Zebrafish reveal new roles for Fam83f in hatching and the DNA damage-mediated autophagic response.
IGF2BP1 phosphorylation in the disordered linkers regulates ribonucleoprotein condensate formation and RNA metabolism.
Induced degradation of SNAP-fusion proteins.
Disruption of tRNA biogenesis enhances proteostatic resilience, improves later-life health, and promotes longevity.
Delineating cysteine-reactive compound modulation of cellular proteostasis processes.
Recruitment to the Proteasome Is Necessary but Not Sufficient for Chemically Induced, Ubiquitin-Independent Degradation of Native Proteins.
Accurate and sensitive interactome profiling using a quantitative protein-fragment complementation assay.
AlphaFold-Multimer accurately captures interactions and dynamics of intrinsically disordered protein regions.
P-stalk ribosomes act as master regulators of cytokine-mediated processes.
Protocol for CRISPR-based endogenous protein tagging in mammalian cells.
Nuclear release of eIF1 restricts start-codon selection during mitosis.
Implications of the Lipidic Ecosystem for the Membrane Binding of ApoE Signal Peptide: Importance of Sphingomyelin.
Cyclophilin D plays a critical role in the survival of senescent cells.
Stepwise phosphorylation and SUMOylation of PIDD1 drive PIDDosome assembly in response to DNA repair failure.
Protein structural context of cancer mutations reveals molecular mechanisms and candidate driver genes.
Transgelin 2 guards T cell lipid metabolism and antitumour function.

EMBO J. 2024 Oct 24.

Recruitment of autophagy initiator TAX1BP1 advances aggrephagy from cargo collection to sequestration.

Bernd Bauer, Jonas Idinger, Martina Schuschnig, Luca Ferrari, Sascha Martens.

  Autophagy mediates the degradation of harmful material within lysosomes. In aggrephagy, the pathway mediating the degradation of aggregated, ubiquitinated proteins, this cargo material is collected in larger condensates prior to its sequestration by autophagosomes. In this process, the autophagic cargo receptors SQSTM1/p62 and NBR1 drive cargo condensation, while TAX1BP1, which binds to NBR1, recruits the autophagy machinery to facilitate autophagosome biogenesis at the condensates. The mechanistic basis for the TAX1BP1-mediated switch from cargo collection to its sequestration is unclear. Here we show that TAX1BP1 is not a constitutive component of the condensates. Its recruitment correlates with the induction of autophagosome biogenesis. TAX1BP1 is sufficient to recruit the TBK1 kinase via the SINTBAD adapter. We define the NBR1-TAX1BP1-binding site, which is adjacent to the GABARAP/LC3 interaction site, and demonstrate that the recruitment of TAX1BP1 to cargo mimetics can be enhanced by an increased ubiquitin load. Our study suggests that autophagosome biogenesis is initiated once sufficient cargo is collected in the condensates.

Keywords:  Aggrephagy p62; NBR1; Quality Control; Selective Autophagy; TAX1BP1

DOI:  https://doi.org/10.1038/s44318-024-00280-5
Autophagy. 2024 Oct 20. 1-2

Nuclear proteasomes as a backup for autophagy: interconnected proteostasis pathways.

Meiyan Jin, Daniel J Klionsky.

  Protein homeostasis (proteostasis) refers to the balance of the cellular protein environment, tightly regulated by pathways governing protein synthesis, folding, trafficking, and degradation. Growing evidence supports the interconnection of these pathways to ensure the robustness of the proteo-stasis network. A recent study by Park et al. showed that, in macroautophagy/autophagy-deficient cells, the loss of proteasome or nuclear pore components causes synthetic lethality, as cytoplasmic proteins that accumulate under impaired autophagy are transported to the nucleus and degraded by nuclear proteasomes. The authors illustrated the mechanistic basis for why cells with conditions such as Huntington disease, where both autophagy and cytoplasm-to-nuclear shuttling are compromised, are more vulnerable to proteostasis perturbation.Abbreviation: UPR: unfolded protein response; UPS: ubiquitin-proteasome system.

Keywords:  Autophagy; nuclear pore complex; nuclear proteasome; proteasome; proteostasis

DOI:  https://doi.org/10.1080/15548627.2024.2416261
Proc Natl Acad Sci U S A. 2024 Oct 29. 121(44): e2408071121

Intrinsically disordered region amplifies membrane remodeling to augment selective ER-phagy.

Sergio Alejandro Poveda-Cuevas, Kateryna Lohachova, Borna Markusic, Ivan Dikic, Gerhard Hummer, Ramachandra M Bhaskara.

  Intrinsically disordered regions (IDRs) play a pivotal role in organellar remodeling. They transduce signals across membranes, scaffold signaling complexes, and mediate vesicular traffic. Their functions are regulated by constraining conformational ensembles through specific intra- and intermolecular interactions, physical tethering, and posttranslational modifications. The endoplasmic reticulum (ER)-phagy receptor FAM134B/RETREG1, known for its reticulon homology domain (RHD), includes a substantial C-terminal IDR housing the LC3 interacting motif. Beyond engaging the autophagic machinery, the function of the FAM134B-IDR is unclear. Here, we investigate the characteristics of the FAM134B-IDR by extensive modeling and molecular dynamics simulations. We present detailed structural models for the IDR, mapping its conformational landscape in solution and membrane-anchored configurations. Our analysis reveals that depending on the membrane anchor, the IDRs collapse onto the membrane and induce positive membrane curvature to varying degrees. The charge patterns underlying this Janus-like behavior are conserved across other ER-phagy receptors. We found that IDRs alone are sufficient to sense curvature. When combined with RHDs, they intensify membrane remodeling and drive efficient protein clustering, leading to faster budding, thereby amplifying RHD remodeling functions. Our simulations provide a perspective on IDRs of FAM134B, their Janus-like membrane interactions, and the resulting modulatory functions during large-scale ER remodeling.

Keywords:  ER remodeling; IDRs; conformational entropy; curvature induction; structural ensemble

DOI:  https://doi.org/10.1073/pnas.2408071121
Cell Rep. 2024 Oct 22. pii: S2211-1247(24)01247-6. [Epub ahead of print]43(11): 114896

EGF-mediated Golgi dynamics and cell migration require CARP2.

Rahul Sharma, Krishanu Dey Das, Srinivasa M Srinivasula.

  In mammalian cells, the Golgi exists in ribbon architecture-individual stacks laterally linked to each other by tubular structures. Golgi architecture changes dynamically to cater to cellular needs. Loss of architecture is linked with pathological conditions like cancer and neurodegeneration. Not much is known about the regulators of Golgi dynamics. Here, we demonstrate that CARP2 (caspase-8- and caspase-10-associated RING-containing protein 2), an endosomal ubiquitin ligase and a known regulator of cell migration, modulates Golgi dynamics. Epidermal growth factor (EGF) treatment modestly increases CARP2 protein and disperses Golgi. An exogenous supply of CARP2 also leads to Golgi dispersal. Conversely, Golgi remains intact in CARP2 knockout (KO) cells upon EGF treatment. CARP2 variants defective in either endosomal association or ligase activity are unable to affect Golgi dispersal. Importantly, CARP2 targets Golgin45 for ubiquitination and degradation in EGF-stimulated cells. Collectively, our findings unravel the existence of crosstalk between endosomal ubiquitin signaling and Golgi dynamics.

Keywords:  CP: Cell biology

DOI:  https://doi.org/10.1016/j.celrep.2024.114896
Brief Bioinform. 2024 Sep 23. pii: bbae519. [Epub ahead of print]25(6):

MetaDegron: multimodal feature-integrated protein language model for predicting E3 ligase targeted degrons.

Mengqiu Zheng, Shaofeng Lin, Kunqi Chen, Ruifeng Hu, Liming Wang, Zhongming Zhao, Haodong Xu.

  Protein degradation through the ubiquitin proteasome system at the spatial and temporal regulation is essential for many cellular processes. E3 ligases and degradation signals (degrons), the sequences they recognize in the target proteins, are key parts of the ubiquitin-mediated proteolysis, and their interactions determine the degradation specificity and maintain cellular homeostasis. To date, only a limited number of targeted degron instances have been identified, and their properties are not yet fully characterized. To tackle on this challenge, here we develop a novel deep-learning framework, namely MetaDegron, for predicting E3 ligase targeted degron by integrating the protein language model and comprehensive featurization strategies. Through extensive evaluations using benchmark datasets and comparison with existing method, such as Degpred, we demonstrate the superior performance of MetaDegron. Among functional features, MetaDegron allows batch prediction of targeted degrons of 21 E3 ligases, and provides functional annotations and visualization of multiple degron-related structural and physicochemical features. MetaDegron is freely available at http://modinfor.com/MetaDegron/. We anticipate that MetaDegron will serve as a useful tool for the clinical and translational community to elucidate the mechanisms of regulation of protein homeostasis, cancer research, and drug development.

Keywords:  E3 ligase; deep-learning; degrons; targeted protein degradation; ubiquitin-proteasome system; web server

DOI:  https://doi.org/10.1093/bib/bbae519
mSphere. 2024 Oct 22. e0037124

Enhanced cell stress response and protein degradation capacity underlie artemisinin resistance in Plasmodium falciparum.

Melissa R Rosenthal, Sukhithasri Vijayrajratnam, Tessa M Firestone, Caroline L Ng.

  Malaria remains a global health burden, killing over half a million people each year. Decreased therapeutic efficacy to artemisinin, the most efficacious antimalarial, has been detected in sub-Saharan Africa, a worrying fact given that over 90% of deaths occur on this continent. Mutations in Kelch13 are the most well-established molecular marker for artemisinin resistance, but these do not explain all artemisinin-resistant isolates. Understanding the biological underpinnings of drug resistance is key to curbing the emergence and spread of artemisinin resistance. Artemisinin-mediated non-specific alkylation leads to the accumulation of misfolded and damaged proteins and activation of the parasite unfolded protein response (UPR). In addition, the parasite proteasome is vital to artemisinin resistance, as we have previously shown that chemical inhibition of the proteasome or mutations in the β2 proteasome subunit increase parasite susceptibility to dihydroartemisinin (DHA), the active metabolite of artemisinins. Here, we investigate parasites with mutations at the Kelch13 and/or 19S and 20S proteasome subunits with regard to UPR regulation and proteasome activity in the context of artemisinin resistance. Our data show that perturbing parasite proteostasis kills parasites, early parasite UPR signaling dictates DHA survival outcomes, and DHA susceptibility correlates with impairment of proteasome-mediated protein degradation. Importantly, we show that functional proteasomes are required for artemisinin resistance in a Kelch13-independent manner, and compound-selective proteasome inhibition demonstrates why artemisinin-resistant Kelch13 mutants remain susceptible to the related antimalarial peroxide OZ439. These data provide further evidence for targeting the parasite proteasome and UPR to overcome existing artemisinin resistance.IMPORTANCEDecreased therapeutic efficacy represents a major barrier to malaria treatment control strategies. The malaria proteasome and accompanying unfolded protein response are crucial to artemisinin resistance, revealing novel antimalarial therapeutic strategies.

Keywords:  Plasmodium falciparum; artemisinin resistance; proteasome; proteasome-mediated protein degradation; proteostasis; ubiquitin; unfolded protein response

DOI:  https://doi.org/10.1128/msphere.00371-24
STAR Protoc. 2024 Oct 24. pii: S2666-1667(24)00578-1. [Epub ahead of print]5(4): 103413

Protocol to identify E3 ligases amenable to biodegraders using a cell-based screening.

Marie Sorbara, Margot Cristol, Anaïs Cornebois, Klervi Desrumeaux, Pierre Cordelier, Nicolas Bery.

  Here, we provide a protocol for the identification of E3 ubiquitin ligases that are functional when implemented as biodegraders using a cell-based screening assay. We describe steps for establishing a stable cell line expressing a GFP-tagged protein of interest (POI), preparing a sub-library of E3 ligases to screen, and performing the cell-based screening. This protocol can be broadly applied to identify any functional E3 ligase in a biodegrader setting. For complete details on the use and execution of this protocol, please refer to Cornebois et al.1.

Keywords:  antibody; cell biology; cell culture; cell-based assays; molecular biology

DOI:  https://doi.org/10.1016/j.xpro.2024.103413
Traffic. 2024 Oct;25(10): e12957

Fluorescent Reporters, Imaging, and Artificial Intelligence Toolkits to Monitor and Quantify Autophagy, Heterophagy, and Lysosomal Trafficking Fluxes.

Mikhail Rudinskiy, Diego Morone, Maurizio Molinari.

  Lysosomal compartments control the clearance of cell-own material (autophagy) or of material that cells endocytose from the external environment (heterophagy) to warrant supply of nutrients, to eliminate macromolecules or parts of organelles present in excess, aged, or containing toxic material. Inherited or sporadic mutations in lysosomal proteins and enzymes may hamper their folding in the endoplasmic reticulum (ER) and their lysosomal transport via the Golgi compartment, resulting in lysosomal dysfunction and storage disorders. Defective cargo delivery to lysosomal compartments is harmful to cells and organs since it causes accumulation of toxic compounds and defective organellar homeostasis. Assessment of resident proteins and cargo fluxes to the lysosomal compartments is crucial for the mechanistic dissection of intracellular transport and catabolic events. It might be combined with high-throughput screenings to identify cellular, chemical, or pharmacological modulators of these events that may find therapeutic use for autophagy-related and lysosomal storage disorders. Here, discuss qualitative, quantitative and chronologic monitoring of autophagic, heterophagic and lysosomal protein trafficking in fixed and live cells, which relies on fluorescent single and tandem reporters used in combination with biochemical, flow cytometry, light and electron microscopy approaches implemented by artificial intelligence-based technology.

Keywords:  ER‐phagy; ER‐to‐lysosome‐associated degradation (ERLAD); artificial intelligence; autophagy; autophagy flux; endolysosomes (EL); heterophagy; lysosomal storage disorders (LSD); lysosomes; tandem fluorescent reporters

DOI:  https://doi.org/10.1111/tra.12957
Proc Natl Acad Sci U S A. 2024 Oct 29. 121(44): e2415383121

A protein phosphatase 1 specific phosphatase targeting peptide (PhosTAP) to identify the PP1 phosphatome.

Meng S Choy, Hieu T Nguyen, Ganesan S Kumar, Wolfgang Peti, Arminja N Kettenbach, Rebecca Page.

  Phosphoprotein phosphatases (PPPs) are the key serine/threonine phosphatases that regulate all essential signaling cascades. In particular, Protein Phosphatase 1 (PP1) dephosphorylates ~80% of all ser/thr phosphorylation sites. Here, we developed a phosphatase targeting peptide (PhosTAP) that binds all PP1 isoforms and does so with a stronger affinity than any other known PP1 regulator. This PhosTAP can be used as a PP1 recruitment tool for Phosphorylation Targeting Chimera (PhosTAC)-type recruitment in in vitro and cellular experiments, as well as in phosphoproteomics experiments to identify PP1-specific substrates and phosphosites. The latter is especially important to further our understanding of cellular signaling, as the identification of substrates and especially phosphosites that are targeted by specific phosphatases lags behind that of their kinase counterparts. Using PhosTAP-based proteomics, we show that, counter to our current understanding, many PP1 regulators are also substrates, that the number of residues between regulator PP1-binding and phosphosites vary significantly, and that PP1 counteracts the activities of mitotic kinases. Finally, we also found that Haspin kinase is a direct substrate of PP1 and that its PP1-dependent dephosphorylation modulates its activity during anaphase. Together, we show that PP1-specific PhosTAPs are a powerful tool for +studying PP1 activity in vitro and in cells.

Keywords:  phosphatase targeting peptide (PhosTAP); phosphoprotein phosphatases (PPP); protein engineering; protein phosphatase 1 (PP1); protein–protein interactions

DOI:  https://doi.org/10.1073/pnas.2415383121
RSC Chem Biol. 2024 Oct 11.

Discovery and design of molecular glue enhancers of CDK12-DDB1 interactions for targeted degradation of cyclin K.

Pompom Ghosh, Maximilian Schmitz, Thiyagamurthy Pandurangan, Solomon Tadesse Zeleke, Sean Chin Chan, John Mosior, Luxin Sun, Vinayak Palve, Dylan Grassie, Kanchan Anand, Sylvia Frydman, William R Roush, Ernst Schönbrunn, Matthias Geyer, Derek Duckett, Andrii Monastyrskyi.

The CDK12 inhibitor SR-4835 promotes the proteasomal degradation of cyclin K, contingent on the presence of CDK12 and the CUL4-RBX1-DDB1 E3 ligase complex. The inhibitor displays molecular glue activity, which correlates with its enhanced ability to inhibit cell growth. This effect is achieved by facilitating the formation of a ternary complex that requires the small molecule SR-4835, CDK12, and the adaptor protein DDB1, leading to the subsequent ubiquitination and degradation of cyclin K. We have successfully solved the structure of the ternary complex, enabling the de novo design of molecular glues that transform four different CDK12 scaffold inhibitors, including the clinical pan-CDK inhibitor dinaciclib, into cyclin K degraders. These results not only deepen our understanding of CDK12's role in cell regulation but also underscore significant progress in designing molecular glues for targeted protein degradation in cancers associated with dysregulated cyclin K activity.

DOI: https://doi.org/10.1039/d4cb00190g
PLoS Pathog. 2024 Oct 21. 20(10): e1012617

Mlf mediates proteotoxic response via formation of cellular foci for protein folding and degradation in Giardia.

Martina Vinopalová, Lenka Arbonová, Zoltán Füssy, Vít Dohnálek, Abdul Samad, Tomáš Bílý, Marie Vancová, Pavel Doležal.

Myeloid leukemia factor 1 (Mlf1) was identified as a proto-oncoprotein that affects hematopoietic differentiation in humans. However, its cellular function remains elusive, spanning roles from cell cycle regulation to modulation of protein aggregate formation and participation in ciliogenesis. Given that structurally conserved homologs of Mlf1 can be found across the eukaryotic tree of life, we decided to characterize its cellular role underlying this phenotypic pleiotropy. Using a model of the unicellular eukaryote Giardia intestinalis, we demonstrate that its Mlf1 homolog (GiMlf) mainly localizes to two types of cytosolic foci: microtubular structures, where it interacts with Hsp40, and ubiquitin-rich, membraneless compartments, found adjacent to mitochondrion-related organelles known as mitosomes, containing the 26S proteasome regulatory subunit 4. Upon cellular stress, GiMlf either relocates to the affected compartment or disperses across the cytoplasm, subsequently accumulating into enlarged foci during the recovery phase. In vitro assays suggest that GiMlf can be recruited to membranes through its affinity for signaling phospholipids. Importantly, cytosolic foci diminish in the gimlf knockout strain, which exhibits extensive proteomic changes indicative of compromised proteostasis. Consistent with data from other cellular systems, we propose that Mlf acts in the response to proteotoxic stress by mediating the formation of function-specific foci for protein folding and degradation.

DOI: https://doi.org/10.1371/journal.ppat.1012617
Eur J Med Chem. 2024 Oct 16. pii: S0223-5234(24)00839-0. [Epub ahead of print]280 116958

Discovery of small molecules for autophagy-lysosome degradation of immune checkpoint proteins.

Kaizhen Wang, Zhihao Qi, Jiazheng Guo, Guoqing Shen, Xiang Ni, Sheng Jiang, Kuojun Zhang, Tianyu Wang, Xiangyu Zhang.

  Targeted protein degradation (TPD) technologies, particularly proteolysis targeting chimeras (PROTACs), have emerged as a promising branch of targeted therapy. Current ubiquitin-proteasome-dependent TPD technologies are limited to targeting intracellular proteins. Although the blockade of immune checkpoints has achieved great clinical success, most immune checkpoints are transmembrane proteins, which are difficult to be ubiquitinated and degraded by PROTACs. Herein, we developed a novel discovery strategy of bifunctional small molecules, which could mediate autophagy-lysosome degradation of immune checkpoints. F-1 was demonstrated to activate the autophagy-lysosome system, and conjugation of F-1 with inhibitors targeting programmed cell death-ligand 1 (PD-L1) or V-domain Ig suppressor of T-cell activation (VISTA) generated a new class of small molecules that effectively induce the degradation of PD-L1 or VISTA in tumor cells. The most promising PD-L1 degrader B3 significantly induced PD-L1 degradation in RKO cells through the autophagy-lysosome system and exhibited good tumor-inhibiting effects in vivo. Our work could expand the development of degraders targeting immune checkpoints and provide a promising discovery strategy for future autophagy-lysosome targeting degradation technology.

Keywords:  Autophagy-lysosome; Cancer immunotherapy; Degrader; PD-1/PD-L1; VISTA

DOI:  https://doi.org/10.1016/j.ejmech.2024.116958
Nat Biotechnol. 2024 Oct 24.

A structurally informed human protein-protein interactome reveals proteome-wide perturbations caused by disease mutations.

Dapeng Xiong, Yunguang Qiu, Junfei Zhao, Yadi Zhou, Dongjin Lee, Shobhita Gupta, Mateo Torres, Weiqiang Lu, Siqi Liang, Jin Joo Kang, Charis Eng, Joseph Loscalzo, Feixiong Cheng, Haiyuan Yu.

To assist the translation of genetic findings to disease pathobiology and therapeutics discovery, we present an ensemble deep learning framework, termed PIONEER (Protein-protein InteractiOn iNtErfacE pRediction), that predicts protein-binding partner-specific interfaces for all known protein interactions in humans and seven other common model organisms to generate comprehensive structurally informed protein interactomes. We demonstrate that PIONEER outperforms existing state-of-the-art methods and experimentally validate its predictions. We show that disease-associated mutations are enriched in PIONEER-predicted protein-protein interfaces and explore their impact on disease prognosis and drug responses. We identify 586 significant protein-protein interactions (PPIs) enriched with PIONEER-predicted interface somatic mutations (termed oncoPPIs) from analysis of approximately 11,000 whole exomes across 33 cancer types and show significant associations of oncoPPIs with patient survival and drug responses. PIONEER, implemented as both a web server platform and a software package, identifies functional consequences of disease-associated alleles and offers a deep learning tool for precision medicine at multiscale interactome network levels.

DOI: https://doi.org/10.1038/s41587-024-02428-4
Sci Adv. 2024 Oct 25. 10(43): eadq6858

The G1-S transition is promoted by Rb degradation via the E3 ligase UBR5.

Shuyuan Zhang, Lucas Fuentes Valenzuela, Evgeny Zatulovskiy, Lise Mangiante, Christina Curtis, Jan M Skotheim.

Mammalian cells make the decision to divide at the G1-S transition in response to diverse signals impinging on the retinoblastoma protein Rb, a cell cycle inhibitor and tumor suppressor. Passage through the G1-S transition is initially driven by Rb inactivation via phosphorylation and by Rb's decreasing concentration in G1. While many studies have identified the mechanisms of Rb phosphorylation, the mechanism underlying Rb's decreasing concentration in G1 was unknown. Here, we found that Rb's concentration decrease in G1 requires the E3 ubiquitin ligase UBR5. UBR5 knockout cells have increased Rb concentration in early G1, exhibited a lower G1-S transition rate, and are more sensitive to inhibition of cyclin-dependent kinase 4/6 (Cdk4/6). This last observation suggests that UBR5 inhibition can strengthen the efficacy of Cdk4/6 inhibitor-based cancer therapies.

DOI: https://doi.org/10.1126/sciadv.adq6858
Open Biol. 2024 Oct;14(10): 240194

Zebrafish reveal new roles for Fam83f in hatching and the DNA damage-mediated autophagic response.

Rebecca A Jones, Fay Cooper, Gavin Kelly, David Barry, Matthew J Renshaw, Gopal Sapkota, James C Smith.

  The FAM83 (Family with sequence similarity 83) family is highly conserved in vertebrates, but little is known of the functions of these proteins beyond their association with oncogenesis. Of the family, FAM83F is of particular interest because it is the only membrane-targeted FAM83 protein. When overexpressed, FAM83F activates the canonical Wnt signalling pathway and binds to and stabilizes p53; it therefore interacts with two pathways often dysregulated in disease. Insights into gene function can often be gained by studying the roles they play during development, and here we report the generation of fam83f knock-out (KO) zebrafish, which we have used to study the role of Fam83f in vivo. We show that endogenous fam83f is most strongly expressed in the hatching gland of developing zebrafish embryos, and that fam83f KO embryos hatch earlier than their wild-type (WT) counterparts, despite developing at a comparable rate. We also demonstrate that fam83f KO embryos are more sensitive to ionizing radiation than WT embryos-an unexpected finding, bearing in mind the previously reported ability of FAM83F to stabilize p53. Transcriptomic analysis shows that loss of fam83f leads to downregulation of phosphatidylinositol-3-phosphate (PI(3)P) binding proteins and impairment of cellular degradation pathways, particularly autophagy, a crucial component of the DNA damage response. Finally, we show that Fam83f protein is itself targeted to the lysosome when overexpressed in HEK293T cells, and that this localization is dependent upon a C' terminal signal sequence. The zebrafish lines we have generated suggest that Fam83f plays an important role in autophagic/lysosomal processes, resulting in dysregulated hatching and increased sensitivity to genotoxic stress in vivo.

Keywords:  DNA damage; Fam83f; autophagy; development; hatching; zebrafish

DOI:  https://doi.org/10.1098/rsob.240194
Nat Commun. 2024 Oct 20. 15(1): 9054

IGF2BP1 phosphorylation in the disordered linkers regulates ribonucleoprotein condensate formation and RNA metabolism.

Harald Hornegger, Aleksandra S Anisimova, Adnan Muratovic, Benjamin Bourgeois, Elena Spinetti, Isabell Niedermoser, Roberto Covino, Tobias Madl, G Elif Karagöz.

The insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) is a conserved RNA-binding protein that regulates RNA stability, localization and translation. IGF2BP1 is part of various ribonucleoprotein (RNP) condensates. However, the mechanism that regulates its assembly into condensates remains unknown. By using proteomics, we demonstrate that phosphorylation of IGF2BP1 at S181 in a disordered linker is regulated in a stress-dependent manner. Phosphomimetic mutations in two disordered linkers, S181E and Y396E, modulate RNP condensate formation by IGF2BP1 without impacting its binding affinity for RNA. Intriguingly, the S181E mutant, which lies in linker 1, impairs IGF2BP1 condensate formation in vitro and in cells, whereas a Y396E mutant in the second linker increases condensate size and dynamics. Structural approaches show that the first linker binds RNAs nonspecifically through its RGG/RG motif, an interaction weakened in the S181E mutant. Notably, linker 2 interacts with IGF2BP1's folded domains and these interactions are partially impaired in the Y396E mutant. Importantly, the phosphomimetic mutants impact IGF2BP1's interaction with RNAs and remodel the transcriptome in cells. Our data reveal how phosphorylation modulates low-affinity interaction networks in disordered linkers to regulate RNP condensate formation and RNA metabolism.

DOI: https://doi.org/10.1038/s41467-024-53400-4
RSC Chem Biol. 2024 Oct 21.

Induced degradation of SNAP-fusion proteins.

Savina Abraham Pol, Sara Liljenberg, Jack Barr, Gina Simon, Luis Wong-Dilworth, Danielle L Paterson, Vladimir P Berishvili, Francesca Bottanelli, Farnusch Kaschani, Markus Kaiser, Mariell Pettersson, Doris Hellerschmied.

Self-labeling protein tags are an efficient means to visualize, manipulate, and isolate engineered fusion proteins with suitable chemical probes. The SNAP-tag, which covalently conjugates to benzyl-guanine and -chloropyrimidine derivatives is used extensively in fluorescence microscopy, given the availability of suitable SNAP-ligand-based probes. Here, we extend the applicability of the SNAP-tag to targeted protein degradation. We developed a set of SNAP PROteolysis TArgeting Chimeras (SNAP-PROTACs), which recruit the VHL or CRBN-ubiquitin E3 ligases to induce the degradation of SNAP-fusion proteins. Endogenous tagging enabled the visualization and the selective depletion of a SNAP-clathrin light chain fusion protein using SNAP-PROTACs. The addition of PROTACs to the SNAP-tag reagent toolbox facilitates the comprehensive analysis of protein function with a single gene tagging event.

DOI: https://doi.org/10.1039/d4cb00184b
PLoS Biol. 2024 Oct;22(10): e3002853

Disruption of tRNA biogenesis enhances proteostatic resilience, improves later-life health, and promotes longevity.

Yasir Malik, Yavuz Kulaberoglu, Shajahan Anver, Sara Javidnia, Gillian Borland, Rene Rivera, Stephen Cranwell, Danel Medelbekova, Tatiana Svermova, Jackie Thomson, Susan Broughton, Tobias von der Haar, Colin Selman, Jennifer M A Tullet, Nazif Alic.

tRNAs are evolutionarily ancient molecular decoders essential for protein translation. In eukaryotes, tRNAs and other short, noncoding RNAs are transcribed by RNA polymerase (Pol) III, an enzyme that promotes ageing in yeast, worms, and flies. Here, we show that a partial reduction in Pol III activity specifically disrupts tRNA levels. This effect is conserved across worms, flies, and mice, where computational models indicate that it impacts mRNA decoding. In all 3 species, reduced Pol III activity increases proteostatic resilience. In worms, it activates the unfolded protein response (UPR) and direct disruption of tRNA metabolism is sufficient to recapitulate this. In flies, decreasing Pol III's transcriptional initiation on tRNA genes by a loss-of-function in the TFIIIC transcription factor robustly extends lifespan, improves proteostatic resilience and recapitulates the broad-spectrum benefits to late-life health seen following partial Pol III inhibition. We provide evidence that a partial reduction in Pol III activity impacts translation, quantitatively or qualitatively, in both worms and flies, indicating a potential mode of action. Our work demonstrates a conserved and previously unappreciated role of tRNAs in animal ageing.

DOI: https://doi.org/10.1371/journal.pbio.3002853
Nat Chem Biol. 2024 Oct 24.

Delineating cysteine-reactive compound modulation of cellular proteostasis processes.

Ashley R Julio, Flowreen Shikwana, Cindy Truong, Nikolas R Burton, Emil R Dominguez, Alexandra C Turmon, Jian Cao, Keriann M Backus.

Covalent modulators and covalent degrader molecules have emerged as drug modalities with tremendous therapeutic potential. Toward realizing this potential, mass spectrometry-based chemoproteomic screens have generated proteome-wide maps of potential druggable cysteine residues. However, beyond these direct cysteine-target maps, the full scope of direct and indirect activities of these molecules on cellular processes and how such activities contribute to reported modes of action, such as degrader activity, remains to be fully understood. Using chemoproteomics, we identified a cysteine-reactive small molecule degrader of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nonstructural protein 14 (nsp14), which effects degradation through direct modification of cysteines in both nsp14 and in host protein disulfide isomerases. This degrader activity was further potentiated by generalized electrophile-induced global protein ubiquitylation, proteasome activation and widespread aggregation and depletion of host proteins, including the formation of stress granules. Collectively, we delineate the wide-ranging impacts of cysteine-reactive electrophilic compounds on cellular proteostasis processes.

DOI: https://doi.org/10.1038/s41589-024-01760-9
ACS Chem Biol. 2024 Oct 22.

Recruitment to the Proteasome Is Necessary but Not Sufficient for Chemically Induced, Ubiquitin-Independent Degradation of Native Proteins.

Madeline Balzarini, Joel Tong, Weijun Gui, Isuru M Jayalath, Bin-Bin Schell, Thomas Kodadek.

Targeted protein degradation (TPD) is a promising strategy for drug development. Most degraders function by forcing the association of the target protein (TP) with an E3 Ubiquitin (Ub) ligase, which, in favorable cases, results in the polyubiquitylation of the TP and its subsequent degradation by the 26S proteasome. An alternative strategy would be to create chemical dimerizers that bypass the requirement for polyubiquitylation by recruiting the target protein directly to the proteasome. Direct-to-proteasome degraders (DPDs) may exhibit different characteristics than ubiquitin-dependent degraders, but few studies of this type of TPD have been published, largely due to the dearth of suitable proteasome ligands. To facilitate studies of DPDs, we report here a mammalian cell line in which the HaloTag protein is fused to the proteasome via Rpn13, one of the ubiquitin receptors. In these cells, a chloroalkane serves as a covalent proteasome ligand surrogate. We show that chimeric molecules comprised of a chloroalkane linked to a ligand for the BET family of proteins or the Cdk2/7/9 family of kinases result in ubiquitin-independent degradation of some of these target proteins. We use this system, the first that allows facile degradation of native proteins in a ubiquitin-independent fashion, to probe two issues: the effect of varying the length of the linker connecting the chloroalkane and the target ligand and the selectivity of degradation within the protein families engaged by the target ligand.

DOI: https://doi.org/10.1021/acschembio.4c00422
Cell Rep Methods. 2024 Oct 21. pii: S2667-2375(24)00263-7. [Epub ahead of print]4(10): 100880

Accurate and sensitive interactome profiling using a quantitative protein-fragment complementation assay.

Natalia Lazarewicz, Gaëlle Le Dez, Romina Cerjani, Lunelys Runeshaw, Matthias Meurer, Michael Knop, Robert Wysocki, Gwenaël Rabut.

  An accurate description of protein-protein interaction (PPI) networks is key to understanding the molecular mechanisms underlying cellular systems. Here, we constructed genome-wide libraries of yeast strains to systematically probe protein-protein interactions using NanoLuc Binary Technology (NanoBiT), a quantitative protein-fragment complementation assay (PCA) based on the NanoLuc luciferase. By investigating an array of well-documented PPIs as well as the interactome of four proteins with varying levels of characterization-including the well-studied nonsense-mediated mRNA decay (NMD) regulator Upf1 and the SCF complex subunits Cdc53 and Met30-we demonstrate that ratiometric NanoBiT measurements enable highly precise and sensitive mapping of PPIs. This work provides a foundation for employing NanoBiT in the assembly of more comprehensive and accurate protein interaction maps as well as in their functional investigation.

Keywords:  CP: Systems biology; Cdc53; Irc20; Met30; Nam7; NanoBiT; Saccharomyces cerevisiae; Upf1; budding yeast; interactome; protein-protein interaction

DOI:  https://doi.org/10.1016/j.crmeth.2024.100880
Proc Natl Acad Sci U S A. 2024 Oct 29. 121(44): e2406407121

AlphaFold-Multimer accurately captures interactions and dynamics of intrinsically disordered protein regions.

Alireza Omidi, Mads Harder Møller, Nawar Malhis, Jennifer M Bui, Jörg Gsponer.

  Interactions mediated by intrinsically disordered protein regions (IDRs) pose formidable challenges in structural characterization. IDRs are highly versatile, capable of adopting diverse structures and engagement modes. Motivated by recent strides in protein structure prediction, we embarked on exploring the extent to which AlphaFold-Multimer can faithfully reproduce the intricacies of interactions involving IDRs. To this end, we gathered multiple datasets covering the versatile spectrum of IDR binding modes and used them to probe AlphaFold-Multimer's prediction of IDR interactions and their dynamics. Our analyses revealed that AlphaFold-Multimer is not only capable of predicting various types of bound IDR structures with high success rate, but that distinguishing true interactions from decoys, and unreliable predictions from accurate ones is achievable by appropriate use of AlphaFold-Multimer's intrinsic scores. We found that the quality of predictions drops for more heterogeneous, fuzzy interaction types, most likely due to lower interface hydrophobicity and higher coil content. Notably though, certain AlphaFold-Multimer scores, such as the Predicted Aligned Error and residue-ipTM, are highly correlated with structural heterogeneity of the bound IDR, enabling clear distinctions between predictions of fuzzy and more homogeneous binding modes. Finally, our benchmarking revealed that predictions of IDR interactions can also be successful when using full-length proteins, but not as accurate as with cognate IDRs. To facilitate identification of the cognate IDR of a given partner, we established "minD," which pinpoints potential interaction sites in a full-length protein. Our study demonstrates that AlphaFold-Multimer can correctly identify interacting IDRs and predict their mode of engagement with a given partner.

Keywords:  AlphaFold2; fuzziness; interactions; intrinsically disordered protein

DOI:  https://doi.org/10.1073/pnas.2406407121
Cell. 2024 Oct 14. pii: S0092-8674(24)01139-5. [Epub ahead of print]

P-stalk ribosomes act as master regulators of cytokine-mediated processes.

Anna Dopler, Ferhat Alkan, Yuval Malka, Rob van der Kammen, Kelly Hoefakker, Daniel Taranto, Naz Kocabay, Iris Mimpen, Christel Ramirez, Elke Malzer, Olga I Isaeva, Mandy Kerkhoff, Anastasia Gangaev, Joana Silva, Sofia Ramalho, Liesbeth Hoekman, Maarten Altelaar, Roderick Beijersbergen, Leila Akkari, Jonathan Wilson Yewdell, Pia Kvistborg, William James Faller.

  Inflammatory cytokines are pivotal to immune responses. Upon cytokine exposure, cells enter an "alert state" that enhances their visibility to the immune system. Here, we identified an alert-state subpopulation of ribosomes defined by the presence of the P-stalk. We show that P-stalk ribosomes (PSRs) are formed in response to cytokines linked to tumor immunity, and this is at least partially mediated by P-stalk phosphorylation. PSRs are involved in the preferential translation of mRNAs vital for the cytokine response via the more efficient translation of transmembrane domains of receptor molecules involved in cytokine-mediated processes. Importantly, loss of the PSR inhibits CD8+ T cell recognition and killing, and inhibitory cytokines like transforming growth factor β (TGF-β) hinder PSR formation, suggesting that the PSR is a central regulatory hub upon which multiple signals converge. Thus, the PSR is an essential mediator of the cellular rewiring that occurs following cytokine exposure via the translational regulation of this process.

Keywords:  P-stalk; T cells; cytokines; melanoma; ribosome

DOI:  https://doi.org/10.1016/j.cell.2024.09.039
STAR Protoc. 2024 Oct 19. pii: S2666-1667(24)00569-0. [Epub ahead of print]5(4): 103404

Protocol for CRISPR-based endogenous protein tagging in mammalian cells.

Yu-Xi Xiao, Jiarun Wei, Jason Moffat.

  Tracking the localization and proximal interaction partners of endogenous proteins provides valuable functional insight. Here, we present a protocol for CRISPR-based endogenous protein tagging in mammalian cells. We describe steps for endogenously tagging human TSC22D2 and MAP4, including designing Cas9 and Cas12a guides for knockin, modularized repair template design and cloning, and procedures for lipid transfection and electroporation. This protocol accommodates Cas nucleases in plasmid expression or ribonucleoprotein complex (RNP) formats. This "endo-tagging" approach offers flexibility and broad applicability. For complete details on the use and execution of this protocol, please refer to Xiao et al.1.

Keywords:  CRISPR; Cell Biology; Genetics; High-Throughput Screening; Microscopy; Molecular Biology

DOI:  https://doi.org/10.1016/j.xpro.2024.103404
Nature. 2024 Oct 23.

Nuclear release of eIF1 restricts start-codon selection during mitosis.

Jimmy Ly, Kehui Xiang, Kuan-Chung Su, Gunter B Sissoko, David P Bartel, Iain M Cheeseman.

Regulated start-codon selection has the potential to reshape the proteome through the differential production of upstream open reading frames, canonical proteins, and alternative translational isoforms1-3. However, conditions under which start codon selection is altered remain poorly defined. Here, using transcriptome-wide translation-initiation-site profiling4, we reveal a global increase in the stringency of start-codon selection during mammalian mitosis. Low-efficiency initiation sites are preferentially repressed in mitosis, resulting in pervasive changes in the translation of thousands of start sites and their corresponding protein products. This enhanced stringency of start-codon selection during mitosis results from increased association between the 40S ribosome and the key regulator of start-codon selection, eIF1. We find that increased eIF1-40S ribosome interaction during mitosis is mediated by the release of a nuclear pool of eIF1 upon nuclear envelope breakdown. Selectively depleting the nuclear pool of eIF1 eliminates the change to translational stringency during mitosis, resulting in altered synthesis of thousands of protein isoforms. In addition, preventing mitotic translational rewiring results in substantially increased cell death and decreased mitotic slippage in cells that experience a mitotic delay induced by anti-mitotic chemotherapies. Thus, cells globally control stringency of translation initiation, which has critical roles during the mammalian cell cycle in preserving mitotic cell physiology.

DOI: https://doi.org/10.1038/s41586-024-08088-3
Chembiochem. 2024 Oct 23. e202400469

Implications of the Lipidic Ecosystem for the Membrane Binding of ApoE Signal Peptide: Importance of Sphingomyelin.

Sasmita Pradhan, Lipika Mirdha, Tanusree Sengupta, Hirak Chakraborty.

  The unidirectional movement of nascent secretory proteins in the cell is primarily assisted by the signal recognition particles (SRP). However, this does not completely justify the importance of the signal peptide (SP) which gets eliminated after the protein translocation. We have earlier demonstrated that a negatively charged lipid such as POPG plays an important role in the higher binding affinity and cholesterol-discriminating ability of the apolipoprotein E (ApoE) SP. In this present work, we aimed to understand the role of sphingomyelin, an important constituent of ER, on the membrane binding of ApoE SP. Our results demonstrate that sphingomyelin promotes membrane binding but cannot discriminate cholesterol. However, sphingomyelin shows a synergistic effect with POPG toward the membrane binding of the ApoE SP. We have further shown that the membrane domains do not have any impact on the binding of ApoE SP. Based on our results we propose that the lipid composition of the endoplasmic reticulum (ER) where ApoE translocates, enhances the binding of the ApoE signal peptide to the ER membrane.

Keywords:  Binding affinity; Lipid composition; Lipidic ecosystem; Signal peptide; Synergistic effect

DOI:  https://doi.org/10.1002/cbic.202400469
EMBO J. 2024 Oct 24.

Cyclophilin D plays a critical role in the survival of senescent cells.

Margherita Protasoni, Vanessa López-Polo, Camille Stephan-Otto Attolini, Julian Brandariz, Nicolas Herranz, Joaquin Mateo, Sergio Ruiz, Oscar Fernandez-Capetillo, Marta Kovatcheva, Manuel Serrano.

  Senescent cells play a causative role in many diseases, and their elimination is a promising therapeutic strategy. Here, through a genome-wide CRISPR/Cas9 screen, we identify the gene PPIF, encoding the mitochondrial protein cyclophilin D (CypD), as a novel senolytic target. Cyclophilin D promotes the transient opening of the mitochondrial permeability transition pore (mPTP), which serves as a failsafe mechanism for calcium efflux. We show that senescent cells exhibit a high frequency of transient CypD/mPTP opening events, known as 'flickering'. Inhibition of CypD using genetic or pharmacologic tools, including cyclosporin A, leads to the toxic accumulation of mitochondrial Ca2+ and the death of senescent cells. Genetic or pharmacological inhibition of NCLX, another mitochondrial calcium efflux channel, also leads to senolysis, while inhibition of the main Ca2+ influx channel, MCU, prevents senolysis induced by CypD inhibition. We conclude that senescent cells are highly vulnerable to elevated mitochondrial Ca2+ ions, and that transient CypD/mPTP opening is a critical adaptation mechanism for the survival of senescent cells.

Keywords:  Cellular Senescence; Cyclophilin D; Mitochondria; Senolytic Therapy; mPTP Flickering

DOI:  https://doi.org/10.1038/s44318-024-00259-2
Nat Commun. 2024 Oct 24. 15(1): 9195

Stepwise phosphorylation and SUMOylation of PIDD1 drive PIDDosome assembly in response to DNA repair failure.

Richa B Shah, Yuanyuan Li, Honglin Yu, Ela Kini, Samuel Sidi.

SUMOylation regulates numerous cellular stress responses, yet targets in the apoptotic machinery remain elusive. We show that a single, DNA damage-induced monoSUMOylation event controls PIDDosome (PIDD1/RAIDD/caspase-2) formation and apoptotic death in response to unresolved DNA interstrand crosslinks (ICLs). SUMO-1 conjugation occurs on conserved K879 in the PIDD1 death domain (DD); is catalyzed by PIAS1 and countered by SENP3; and is triggered by ATR phosphorylation of neighboring T788 in the PIDD1 DD, which enables PIAS1 docking. Phospho/SUMO-PIDD1 proteins are captured by nucleolar RAIDD monomers via a SUMO-interacting motif (SIM) in the RAIDD DD, thus compartmentalizing nascent PIDDosomes for caspase-2 recruitment. Denying SUMOylation or the SUMO-SIM interaction spares the onset of PIDDosome assembly but blocks its completion, thus eliminating the apoptotic response to ICL repair failure. Conversely, removal of SENP3 forces apoptosis, even in cells with tolerable ICL levels. SUMO-mediated PIDDosome control is also seen in response to DNA breaks but not supernumerary centrosomes. These results illuminate PIDDosome formation in space and time and identify a direct role for SUMOylation in the assembly of a major pro-apoptotic device.

DOI: https://doi.org/10.1038/s41467-024-53412-0
Cell Rep. 2024 Oct 22. pii: S2211-1247(24)01256-7. [Epub ahead of print]43(11): 114905

Protein structural context of cancer mutations reveals molecular mechanisms and candidate driver genes.

Diego Chillón-Pino, Mihaly Badonyi, Colin A Semple, Joseph A Marsh.

  Advances in protein structure determination and modeling allow us to study the structural context of human genetic variants on an unprecedented scale. Here, we analyze millions of cancer-associated missense mutations based on their structural locations and predicted perturbative effects. By considering the collective properties of mutations at the level of individual proteins, we identify distinct patterns associated with tumor suppressors and oncogenes. Tumor suppressors are enriched in structurally damaging mutations, consistent with loss-of-function mechanisms, while oncogene mutations tend to be structurally mild, reflecting selection for gain-of-function driver mutations and against loss-of-function mutations. Although oncogenes are difficult to distinguish from genes with no role in cancer using only structural damage, we find that the three-dimensional clustering of mutations is highly predictive. These observations allow us to identify candidate driver genes and speculate about their molecular roles, which we expect will have general utility in the analysis of cancer sequencing data.

Keywords:  CP: Cancer; CP: Genomics; Missense mutations; gain-of-function; loss-of-function; oncogenes; protein stability; protein structure; structural bioinformatics; tumor suppressors

DOI:  https://doi.org/10.1016/j.celrep.2024.114905
Nature. 2024 Oct 23.

Transgelin 2 guards T cell lipid metabolism and antitumour function.

Sung-Min Hwang, Deepika Awasthi, Jieun Jeong, Tito A Sandoval, Chang-Suk Chae, Yusibeska Ramos, Chen Tan, Matías Marin Falco, Camilla Salvagno, Alexander Emmanuelli, Ian T McBain, Bikash Mishra, Lionel B Ivashkiv, Dmitriy Zamarin, Evelyn Cantillo, Eloise Chapman-Davis, Kevin Holcomb, Diana K Morales, Xiaoqing Yu, Paulo C Rodriguez, Jose R Conejo-Garcia, Martin Kaczocha, Anna Vähärautio, Minkyung Song, Juan R Cubillos-Ruiz.

Mounting effective immunity against pathogens and tumours relies on the successful metabolic programming of T cells by extracellular fatty acids1-3. Fatty-acid-binding protein 5 (FABP5) has a key role in this process by coordinating the efficient import and trafficking of lipids that fuel mitochondrial respiration to sustain the bioenergetic requirements of protective CD8+ T cells4,5. However, the mechanisms that govern this immunometabolic axis remain unexplored. Here we report that the cytoskeletal organizer transgelin 2 (TAGLN2) is necessary for optimal fatty acid uptake, mitochondrial respiration and anticancer function in CD8+ T cells. TAGLN2 interacts with FABP5 to facilitate its cell surface localization and function in activated CD8+ T cells. Analyses of ovarian cancer specimens revealed that endoplasmic reticulum (ER) stress responses induced by the tumour microenvironment repress TAGLN2 in infiltrating CD8+ T cells, thereby enforcing their dysfunctional state. Restoring TAGLN2 expression in ER-stressed CD8+ T cells increased their lipid uptake, mitochondrial respiration and cytotoxic capacity. Accordingly, chimeric antigen receptor T cells overexpressing TAGLN2 bypassed the detrimental effects of tumour-induced ER stress and demonstrated therapeutic efficacy in mice with metastatic ovarian cancer. Our study establishes the role of cytoskeletal TAGLN2 in T cell lipid metabolism and highlights the potential to enhance cellular immunotherapy in solid malignancies by preserving the TAGLN2-FABP5 axis.

DOI: https://doi.org/10.1038/s41586-024-08071-y