Nat Cell Biol
· 2026 Jan · PMID 41366519
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Cytosolic phospholipase A2 (cPLA) controls some of the most powerful inflammatory lipids in vertebrates by releasing their metabolic precursor, arachidonic acid, from the inner nuclear membrane (INM). Ca and INM tension...Cytosolic phospholipase A2 (cPLA) controls some of the most powerful inflammatory lipids in vertebrates by releasing their metabolic precursor, arachidonic acid, from the inner nuclear membrane (INM). Ca and INM tension (T) are thought to govern the interactions and activity of cPLA at the INM. However, as compensatory membrane flow from the contiguous endoplasmic reticulum (ER) may prevent T, the conditions permitting nuclear membrane mechanotransduction by cPLA or other mediators remain unclear. To test whether the ER buffers T, we created the genetically encoded, Ca²⁺-insensitive T biosensor amphipathic lipid-packing domain inside the nucleus (ALPIN). Confocal time-lapse imaging of ALPIN- or cPLA-INM interactions, along with ER morphology, nuclear shape/volume and cell lysis revealed a link between T and disrupted ER-nuclear membrane contiguity in osmotically or ferroptotically stressed mammalian cells and at zebrafish wound margins in vivo. By combining ALPIN imaging with Ca-induced ER disruption, we reveal the causality of this correlation, which suggests that compensatory membrane flow from the ER buffers T without preventing it. Besides consolidating the biomechanical basis of cPLA activation by nuclear deformation, our results identify cell stress- and cell death-induced ER disruption as an additional nuclear membrane mechanotransduction trigger.
Zorzan I, Carbognin E, Lauria A
… +18 more, Proserpio V, Benvegnù D, Bertelli F, De Juambelz Urías S, Dalrio C, Panebianco G, Scarfò R, Pensabene E, Arboit M, Paolucci I, Drusin A, Bizzotto D, Sledziowska M, Braghetta P, Ditadi A, Amadei G, Oliviero S, Martello G
Pluripotency, the ability to generate all body cell types, emerges in a disorganized embryonic cell mass. After implantation, these cells form a columnar epithelium and initiate lumenogenesis. During gastrulation, some u...Pluripotency, the ability to generate all body cell types, emerges in a disorganized embryonic cell mass. After implantation, these cells form a columnar epithelium and initiate lumenogenesis. During gastrulation, some undergo epithelial-to-mesenchymal transition to form the primitive streak (PS). The signals controlling these events in humans are largely unknown. Here, to study them, we developed a chemically defined 3D model where conventional pluripotent stem cells self-organize into a columnar epithelium with a lumen, from which PS-like cells emerge. We show that early TGFβ family inhibition prevents epithelial identity, also in murine 3D embryo models and in embryos. ZNF398 acts downstream of TGFβ1, activating the epithelial master regulator ESRP1 while repressing mesenchymal factors CDH2 and ZEB2. After epithelium formation, TGFβ1 stimulation is dispensable for its maintenance. However, treatment via ACTIVIN-a distinct TGFβ family ligand-induces PS efficiently. Thus, signalling of the TGFβ family dynamically governs pluripotent epiblast epithelial identity.
Kozai T, Fernandez-Martinez J, Kapinos LE
… +18 more, Gallardo P, van Eeuwen T, Saladin M, Eliasian R, Mazur A, Zhang W, Tempkin J, Panatala R, Delgado-Izquierdo M, Escribano-Marin R, Feng Q, Lin C, Sali A, Chait BT, Raveh B, Veenhoff LM, Rout MP, Lim RYH
Nat Cell Biol
· 2025 Dec · PMID 41331088
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Nuclear pore complexes (NPCs) mediate selective exchange of macromolecules between the nucleus and cytoplasm, but the organization of their transport barrier has been a matter of debate. Here we used high-speed atomic fo...Nuclear pore complexes (NPCs) mediate selective exchange of macromolecules between the nucleus and cytoplasm, but the organization of their transport barrier has been a matter of debate. Here we used high-speed atomic force microscopy, complemented with orthogonal in vitro and in vivo approaches, to probe the dynamic behaviour of the NPC central channel at millisecond resolution. We found that nuclear transport factors dynamically remodel intrinsically disordered phenylalanine-glycine (FG) domains tethered within the NPC channel, partitioning the barrier into two zones: a rapidly fluctuating annular region and a highly mobile central plug. Increased FG-repeat density in mutant NPCs dampened barrier dynamics and impaired transport. Notably, NPC-like behaviour was recapitulated in DNA origami nanopores bearing transport factors and correctly tethered FG domains but not in in vitro FG hydrogels. Thus, the rotationally symmetric architecture of NPCs supports a nanoscopic barrier organization that contrasts with many of the bulk properties of in vitro FG-domain assemblies.
Zhou Y, Jay A, Burget N
… +7 more, Friedrich T, Yoon S, Alsing J, Nir G, Grosschedl R, Vahedi G, Faryabi RB
Nat Cell Biol
· 2026 Jan · PMID 41331087
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Multiple enhancers, often separated by vast genomic distances, regulate key genes. However, how the folding of individual chromatin fibres enables cell-type-restricted multi-enhancer regulation remains unclear. Here, usi...Multiple enhancers, often separated by vast genomic distances, regulate key genes. However, how the folding of individual chromatin fibres enables cell-type-restricted multi-enhancer regulation remains unclear. Here, using acute protein degradation and time-resolved chromatin conformation capture in mantle cell lymphoma, we found that the B cell-lineage-determining factor EBF1 organizes multiple enhancers around sparsely distributed genes essential for B cell identity and oncogenesis. Time-resolved sub-diffraction optical tracing of more than 100,000 chromatin fibres further revealed diverse topological conformations that facilitate multi-enhancer interactions. Mechanistically, we discovered that enhancer positioning at local topological centres is required for promoter engagement, with EBF1 acting as a permeable barrier to loop-extruding cohesin at enhancers. Extending these findings to T cell leukaemia, we show that lineage-determining transcription factors such as EBF1 and TCF1 radially position enhancers within gene loci to enable multi-enhancer regulation of key oncogenes at the single-allele level.
Nat Cell Biol
· 2026 Jan · PMID 41331086
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Targeting oncogenic KRAS holds great promise but is often limited by rapid adaptive resistance. A study now shows that RASH3D19 is regulated by microRNAs and promotes resistance to RAS inhibition by enhancing EGFR dimeri...Targeting oncogenic KRAS holds great promise but is often limited by rapid adaptive resistance. A study now shows that RASH3D19 is regulated by microRNAs and promotes resistance to RAS inhibition by enhancing EGFR dimerization. Targeting RASH3D19 improves sensitivity to RAS inhibitors in preclinical settings.
Treekitkarnmongkol W, Katayama H, Sankaran D
… +17 more, Tai MC, Rauth S, Chen H, Nguyen T, Hara K, Thege FI, Ponnusamy MP, Batra SK, Wang H, Wistuba II, Schmittgen TD, Heymach JV, Kopetz S, Hu T, Yao W, Maitra A, Sen S
Therapeutic targeting of mutant KRAS pathways driving cancers is being actively investigated to identify feedback mechanisms responsible for the development of adaptive resistance to mutant KRAS inhibitors undergoing cli...Therapeutic targeting of mutant KRAS pathways driving cancers is being actively investigated to identify feedback mechanisms responsible for the development of adaptive resistance to mutant KRAS inhibitors undergoing clinical trials. Here we report RASH3D19 as a mediator of RAS pathway activation through a positive feedback loop involving the KRAS-microRNA signalling axis. KRAS-induced miR-222 represses ETS1 expression and downstream transactivation of miR-301a leading to elevation of its target RASH3D19. RASH3D19 facilitates activation of RAS pathways by promoting dimerization and interaction of EGFR with the SOS2, GRB2, SHP2 and GAB1 complex. Genetic deletion of RASH3D19 in mutant KRAS-expressing cancer cells exhibits growth retardation in vitro, in vivo and sensitized pancreatic ductal adenocarcinoma and colorectal cancer cells, organoids and xenografts to mutant KRAS inhibitors, suppressing feedback reactivation of RAS pathways. Therapeutic targeting of RASH3D19 is expected to lead to tumour debulking and alleviating resistance to KRAS inhibitors in mutant KRAS-expressing cancers.
Li R, Li Y, Wu S
… +17 more, Zhou Z, Hou X, Yang P, Zhu J, Xia Y, Wu W, Feng R, Gao Q, Huang K, Liu Y, Zheng J, Zhong X, Huang H, Chang C, Ma W, Songyang Z, Liu F
Nuclear condensates (NCs) are membraneless organelles that enable spatial and functional compartmentalization in the nucleus. Yet, the components and functional co-organization of NCs have been poorly studied. Here, we u...Nuclear condensates (NCs) are membraneless organelles that enable spatial and functional compartmentalization in the nucleus. Yet, the components and functional co-organization of NCs have been poorly studied. Here, we used PhastID to explore the proximal interactome of 18 NCs in HeLa cells. Our data revealed the organizational flow of gene control among these NCs. Crucially, we developed an algorithm to dissect the intricate internal relations of NCs. This algorithm led to key discoveries: the identification of an uncharacterized BUD13 condensate, and the recognition of specific co-organizations between nuclear gems and Cajal bodies for telomerase maturation, and between nuclear gems and histone locus bodies for histone gene pre-mRNA processing. We also created a global reference map to understand NC dynamics under stresses and how disease-related mutations differentially affect NC interactomes. Overall, our work provides a proximal proteome-based atlas for human NCs, substantially advancing our spatiotemporal understanding of nuclear biological events.
Tumour-associated macrophages (TAMs) play a pivotal role in shaping the immune microenvironment of hepatocellular carcinoma (HCC), influencing tumour progression and immunotherapy response. WDR4, a tRNA-binding cofactor...Tumour-associated macrophages (TAMs) play a pivotal role in shaping the immune microenvironment of hepatocellular carcinoma (HCC), influencing tumour progression and immunotherapy response. WDR4, a tRNA-binding cofactor of the N-methylguanosine (mG) methyltransferase complex, remains poorly understood in its independent functions. Here we show that WDR4 is significantly upregulated in HCC-associated TAMs and correlates with poor prognosis. Loss of WDR4 in monocyte-derived macrophages, but not in resident Kupffer cells, reprogrammes TAMs towards an antitumoral phenotype and suppresses HCC progression. Mechanistically, cytoplasmic WDR4 acts independently of mG modification by directly interacting with eIF4E2 to enhance eIF4E-mediated selective translation of ABCA1, thereby promoting membrane cholesterol efflux and maintaining pro-tumoral polarization. Targeted silencing of WDR4 in TAMs using a CpG-small interfering RNA delivery system enhances antitumour immunity, inhibits tumour progression and improves the efficacy of anti-PD-1 therapy. Our findings identify WDR4 as a key regulator of TAM polarization and a promising therapeutic target to enhance immunotherapeutic outcomes.
Rai A, Huynh K, Cross J
… +9 more, Poh QH, Fang H, Claridge B, Duong T, Duarte C, Shaw JE, Marwick TH, Meikle P, Greening DW
Nat Cell Biol
· 2025 Dec · PMID 41315767
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Extracellular vesicles (EVs) are an essential signalling entity in human plasma implicated in health and disease. Still, their core protein and lipid componentry, which lie at the centre of EV form and function, remain p...Extracellular vesicles (EVs) are an essential signalling entity in human plasma implicated in health and disease. Still, their core protein and lipid componentry, which lie at the centre of EV form and function, remain poorly defined. Here we performed high-resolution density gradient fractionation of over 140 human plasma samples to isolate circulating EVs, and systematically constructed their quantitative proteome (4,500 proteins) and lipidome (829 lipids) landscapes using mass spectrometry. We identified a highly conserved panel of 182 proteins (including ADAM10, STEAP23 and STX7) and 52 lipids (including PS, PIPs, Hex2Cer and PAs), providing a deep survey of hallmark molecular features and biological pathways characteristic to circulating EVs. We also mapped the surfaceome diversity, identifying 151 proteins on the EV surface. We further established a set of 42 proteins and 114 lipids features that served as hallmark features of non-EV particles in plasma. We submit ADAM10 and PS(36:1) as conserved EV biological markers that precisely differentiate between EV and non-EV particles. Our findings, which can be explored via an open-source Shiny web tool ( evmap.shinyapps.io/evmap/ ), will serve as a valuable repository to the research community for a clearer understanding of circulating EV biology.
Yuan M, Jin K, Yan H
… +21 more, Schroeder A, Luo C, Yao S, Dumoulin B, Levinsohn J, Luo T, Clemenceau JR, Jang I, Kim M, Liu Y, Deng M, Furth EE, Wilson P, Nayak A, Lubo I, Solis Soto LM, Wang L, Park JH, Susztak K, Hwang TH, Li M
Nat Cell Biol
· 2025 Dec · PMID 41298871
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Spatial omics technologies have transformed biomedical research by enabling high-resolution molecular profiling while preserving the native tissue architecture. These advances provide unprecedented insights into tissue s...Spatial omics technologies have transformed biomedical research by enabling high-resolution molecular profiling while preserving the native tissue architecture. These advances provide unprecedented insights into tissue structure and function. However, the high cost and time-intensive nature of spatial omics experiments necessitate careful experimental design, particularly in selecting regions of interest (ROIs) from large tissue sections. Currently, ROI selection is performed manually, which introduces subjectivity, inconsistency and a lack of reproducibility. Previous studies have shown strong correlations between spatial molecular patterns and histological features, suggesting that readily available and cost-effective histology images can be leveraged to guide spatial omics experiments. Here we present Smart Spatial omics (S2-omics), an end-to-end workflow that automatically selects ROIs from histology images with the goal of maximizing molecular information content in the ROIs. Through comprehensive evaluations across multiple spatial omics platforms and tissue types, we demonstrate that S2-omics enables systematic and reproducible ROI selection and enhances the robustness and impact of downstream biological discovery.