Nature Communications

Type 2 inflammation is central to chronic rhinosinusitis with nasal polyps (CRSwNP), but it does not fully explain persistent stromal and vascular remodeling. We asked whether complement, adhesion, stromal, and vascular signals converge across tissue, cellular, and spatial scales. We integrated four bulk-transcriptomic cohorts with a single-cell atlas and GeoMx spatial transcriptomics. Network analysis identified eight contrast-associated hub genes and an ABCC9-ACTA2-GPM6A score that separated study-defined groups across cohorts. Single-cell mapping localized these genes mainly to pericytes, fibroblasts, and endothelial cells, while CellChat linked structural and immune compartments through extracellular-matrix and inflammatory signals. In spatial data, C3-C3AR1 showed closer-than-random proximity after multiple-testing correction, whereas ICAM1-ITGAL/ITGB2 did not. Anatomical-site sensitivity analyses retained all eight hub-gene directions. These findings support CASVR as an association framework linking complement-related signals to structural-vascular compartments and identify C3-C3AR1 for independent mechanistic testing.

Neumann et al. introduced CHARM, a compact epigenetic silencer in which a histone H3 tail fused to DNMT3L was proposed to recruit and stimulate endogenous DNMT3A, enabling durable gene repression without a fused DNMT3A catalytic domain. Here, we evaluated the contribution of the H3 tail in independent reporter and endogenous-gene contexts. In an SNRPN reporter system, a KRAB-DNMT3L-dCas9 construct lacking the H3 tail displayed silencing kinetics comparable to CRISPRcharm Kv2, and mutating the critical H3K4 residue to alanine in CRISPRcharm Kv2 did not compromise this silencing. Similarly, after transient delivery of editor mRNAs to HEK293T cells, CRISPRcharm Kv2 did not consistently outperform the corresponding H3-tail-free construct at three endogenous loci, and mutating the critical H3K4 residue to alanine in CRISPRcharm Kv2 did not compromise this activity. These observations suggest that the engineered H3 tail does not confer a general functional advantage within the KRAB-DNMT3L-dCas9 architecture under the conditions tested.

Genomic events and their timing that grant a primary tumour the competence to disseminate remain poorly defined. We performed sequencing of 247 stage I/II primary cutaneous melanomas (CMs) and 60 matched metastases without intervening therapy from a prospectively followed registry cohort with a median followup of 92 months, integrating copy-number, mutational, protein and spatial-transcriptomic analyses. Relapse was not distinguished by oncogenic point mutations, which were largely shared between primaries and metastases, but by somatic copy-number alterations (SCNAs) and global chromosomal instability. We defined OncoCycle, a six-gene copy-number signature (amplification of CDK4, MCL1 and CD276; biallelic loss of CDKN2A, CDKN2B and TP53BP1) that predicted relapse independently of established clinicopathological features in melanoma, and a pan-cancer analysis. In matched pairs, metastatic progression was driven by continued copy-number evolution and reduction in intra-tumoural heterogeneity, rather than by acquired point mutations, and OncoCycle alterations from primary tumours were preserved in metastasis seeding clones. Clonal reconstruction revealed both monoclonal and polyclonal metastasis seeding, and spatial transcriptomics resolved copy-number-defined metastatic subclones occupying and programming distinct immune and stromal niches. Thus, metastatic competence was primed early by focal SCNAs on a background of chromosomal instability, elaborated by continued copy-number evolution during dissemination and spatio-temporal interactions with the tumour-microenvironment.

In the paper by Nguyen et al, published in Nature Communications in 2024, we describe structural polymorphism in fibrils extracted from patients with ATTR amyloidosis. ATTR amyloidosis is characterized by deposition of transthyretin amyloid fibrils across multiple organs, resulting in marked phenotypic variability. Previous studies on ATTR suggest its phenotypic variability may be linked to differences in fibril structure, as demonstrated in amyloid diseases such as AA amyloidosis, tauopathies, or synucleinopathies. Thus far, most ATTR fibrils adopt a strikingly similar fold across patients and affected organs. The most notable exception is described in the study by Nguyen et al., where we report the identification of two co-existing polymorphs in cardiac fibrils from three patients with the ATTRv-I84S variant. These cardiac polymorphs differ in conformation of the segment Gly57 to Gly67, termed the gate, which regulates access to a central polar channel within the fibril core. In one polymorph, the gate adopts an open conformation, exposing the channel, whereas in the other it is closed, shielding it. Whether these polymorphs extend beyond cardiac tissue remains unknown. Here, we address this question by extending our analysis to additional organs from a fourth patient with ATTRv-I84S amyloidosis.

How perennial, postmitotic multinucleated tissues, such as skeletal myofibers, maintain their identity and transcriptional adaptation to homeostatic perturbations through adult life is an outstanding question. To address this issue, we investigated the consequences of loss of 3D-genome architecture in skeletal muscles by generating myofiber-specific Ctcf-deficient (CtcfmKO) mice. CtcfmKO mice did not exhibit muscular phenotype at birth but spontaneously developed a severe myopathy. Integrated analysis of snRNAseq, ATACseq and promoter-capture Hi-C revealed both common and fiber-type specific patterns of dysregulated gene expression associated with alterations in chromatin accessibility and promoter-based interactions in Ctcf-deficient myonuclei at distinct stages of myopathy development. Decreased chromatin accessibility at promoters and changes in their connectivity with distal elements were observed across all myonuclei as a direct consequence of Ctcf deficiency at early stages and associated with downregulation of genes implicated in myofiber contraction and anabolism, metabolism, adhesion and neuromuscular transmission. Conversely, at later stages, upregulation of genes leading to persistent activation of ER stress/UPR and catabolism resulted from global reconfiguration of chromatin structure and connectivity, partly as indirect consequence of Ctcf deficiency. Notably, type-IIB myonuclei exhibited specific alterations in gene expression that culminated in loss of fiber-type identity and ectopic expression of inflammatory genes. These results reveal a requirement of Ctcf for maintenance of fiber-type identity and transcriptional adaptation in vivo, through multilayered control of 3D genome integrity. They also indicate an unprecedented association between Ctcf deficiency in myofibers and susceptibility to develop myopathies, whereby Ctcf dispensability for developmental myogenesis confers vulnerability to develop myopathic syndromes.

Sequence-to-function (S2F) deep learning models predict genome-wide functional profiles directly from DNA sequences and have significantly advanced our understanding of gene regulation. However, existing S2F models are fundamentally constrained by their initial training datasets, and no single model encompasses the full spectrum of cell types, clinical conditions, and cellular states, while training new models de novo is nontrivial. Here, we develop Kidzoi, a kidney cell-type-specific S2F model obtained by transfer learning from the pretrained model Borzoi using single-nucleus chromatin accessibility data from human kidney tissue. Kidzoi accurately predicts cell-type-specific chromatin accessibility and improves regulatory variant effect prediction by 10-23% over existing models. Furthermore, Kidzoi effectively prioritizes genome-wide association study (GWAS) fine-mapped causal variants associated with kidney function (estimated glomerular filtration rate based on both creatinine and cystatin C; eGFRcr-cys) while resolving their cell-type specificity. Systematic ablation of flanking sequence indicates that a local context of 32-64kbp, representing only 6-12% of the full model input, is sufficient for accurate variant effect prediction. Finally, we show that a multi-task model achieves comparable performance to single-task models at a substantially reduced computational cost.

The global CTX-M pandemic is considered a plasmid-driven phenomenon, yet chromosomal integration of blaCTX-M has been reported in various settings. Screening 451,174 publicly available E. coli genomes and retaining 2,269 CTX-M-E. coli long-read or hybrid assemblies, we found that 33.5% carried at least one chromosomally integrated blaCTX-M. Chromosomal integration was strongly structured by E. coli lineage and CTX-M variant, reaching 68.5% in ST38 and 43.9% in ST131. In the latter, integration rates varied between subclades with modelling, suggesting recent acceleration within the ST131-C2 subclade. Additional resistance genes were frequently co-integrated with blaCTX-M (33%), with architectures largely inherited from plasmid-borne structures. Integrations were mainly ISEcp1- or IS26-mediated (77.3% and 18.9%, respectively) and occurred across the entire E. coli chromosome without spared zones. blaCTX-M chromosomal integration is a widespread, lineage-structured component of the CTX-M pandemic, providing stable vertical inheritance that complements or may supplant plasmid-mediated dissemination in successful ESBL-E. coli clones.

Crista junctions (CJs) are narrow infoldings of the inner mitochondrial membrane, serving as entry points into the cristae microenvironment. The mitochondrial contact site and cristae organizing system (MICOS) complex is a highly conserved protein machinery controlling CJ biogenesis and architecture, but its assembly mechanism driving CJ formation has remained unclear. By determining a crystal structure of a Chaetomium thermophilum Mic60-Mic19 complex and combining it with modelling and mutagenesis studies, we identify an extended helical region (EHR) of the Mic60 mitofilin domain and a further Mic19 helix as crucial elements of complex formation. In vitro cross-linking data and hydrogen-deuterium exchange mass spectrometry resolve domain assembly and conformational dynamics of the Mic60 Mic19 subcomplex and point to the EHR as a molecular switch controlling Mic60 transition from a closed dimer to a Mic19-bound hetero-octamer. Our data provide a structural model for Mic60 activation and therefore MICOS assembly as a prerequisite for crista junction formation.

Newcastle disease virus (NDV) continues to cause major economic losses in poultry, driving many recent virulent outbreaks. Maternally derived antibodies (MDA) can interfere with the development of active immunity following vaccination in young chicks. NDV vaccine strains that are safe for day-old chicks are not safe for administration to embryos. Here, we combined a computationally guided attenuation strategy with an in-ovo vaccination and challenge study to develop and evaluate an embryo-safe NDV vaccine candidate. Using a large NDV sequence compendium, we applied a local RNA folding energy based algorithm to identify folding selected clusters within the NDV fusion protein; we introduced targeted synonymous substitutions within the first ~900 nucleotides to modulate local RNA structure while preserving the encoded protein sequence. The recombinant attenuated variant was propagated in embryonated chicken eggs to generate vaccine stocks for in-ovo vaccination. After 18 days of embryonation, the synthetic variants were well tolerated, achieving more than 97% hatchability, with no pre-challenge morbidity or mortality, while unvaccinated challenge controls showed complete mortality. Together, these data support a model in which structure-aware synonymous modifications can yield an embryo-safe NDV backbone suitable for in-ovo delivery, with protective efficacy exhibiting a clear dose window under high MDA conditions.

{gamma}-Aminobutyric acid (GABA) and glycine mediate fast synaptic inhibition by restraining neuronal excitability and stabilizing neural circuits. The vesicular inhibitory amino acid transporter (VIAAT/VGAT) loads both GABA and glycine into synaptic vesicles, but its substrate-recognition and transport mechanisms remain unclear. Here we report cryo-EM structures of human VIAAT in apo luminal-open and GABA-bound occluded states. VIAAT adopts an 11 transmembrane helix LeuT-like fold, distinct from other vesicular neurotransmitter transporters. In the central pocket, an aromatic cage comprising Phe133, Tyr221, Phe315 and Phe322 mediates recognition of GABA. Structural comparison reveals that Phe315 acts as a luminal gate, rotating away from the central pocket to open the luminal pathway for substrate release. Complemented by functional neuronal rescue experiments and a cytosolic-facing model, our findings elucidate the mechanism for VIAAT-mediated GABA transport and provide a framework for understanding how epilepsy-associated VIAAT variants impair inhibitory neurotransmission.

Cardiac G protein-gated inwardly rectifying potassium-K+ (GIRK) channels, formed by GIRK1/4 heterotetramers or GIRK4 homotetramers, are crucial regulators of heart rate, and their dysfunction is associated with atrial fibrillation. Despite their therapeutic potential as anti-arrhythmic targets, development of selective GIRK modulators has been limited by an incomplete understanding of their structures, gating mechanisms, and pharmacology. Here, we report high-resolution structures of the GIRK4 homotetramer and of GIRK1/4 heterotetramers, revealing that they form in unexpected 1:3 and 2:2 assembly stoichiometries that we corroborate using cell-based single molecule microscopy. Electrophysiological assays and molecular dynamics simulations further show that incorporation of one or two GIRK1 subunits into GIRK4-containing channels progressively opens the helix bundle crossing (HBC) gate and increases basal channel activity. Additional structures of GIRK1/4 complexes with ML297, a small-molecule ligand selective for GIRK1-containing channels, reveal distinct GIRK1-centered intersubunit binding environments. Remarkably, ML297 inhibits GIRK1/4 channels with a 1:3 stoichiometry but potentiates channels with a 2:2 stoichiometry, establishing a direct link between subunit composition, ligand-binding environment, and pharmacological efficacy. Together, these findings provide a structural and mechanistic framework for the rational design of ligands with enhanced selectivity among cardiac and neuronal GIRK channel heterotetramers.

Viral integration is common in human papillomavirus-positive oropharyngeal squamous cell carcinoma (HPV+ OPSCC), but its structural and clinical significance remains unclear. We integrated targeted capture sequencing of 197 HPV+ OPSCCs with long-read nanopore sequencing of 17 tumors and models to characterize HPV-host rearrangements. We developed a novel quantitative informatics framework that classifies integration structures as Type1 (simple, single/low-copy) or Type2 (clustered, high-copy, rearranged). HPV integration occurred in 83% of tumors, with recurrent hotspots including TP63, MYC, CD274, TRAF2, and KMT2C. Type2 events were enriched for HPV E2 disruption, focal human amplifications, preferential transcription, and noncanonical HPV-host RNA fusion isoforms. Type2 tumors showed increased HPV oncogene expression and activation of MYC, NF-{kappa}B/TNF, PI3K-AKT-mTOR, Hippo/YAP-TAZ, and epithelial stemness programs. Multi-region and longitudinal analyses revealed intratumoral heterogeneity and clonal selection of Type2 architectures during progression, supporting integration structure as a potential marker of aggressive behavior and risk stratification.

Transcriptional states can provide functional information beyond genomic alterations, but whether states derived from nonmalignant physiological perturbations remain informative in cancer is unclear. We separated global RNA-output suppression from relative transcriptional remodeling induced by dihydrotestosterone in mouse adrenal zona fasciculata cells and derived a cross-sex transcriptional signature. Projection of this signature to human orthologs defined an androgen-suppressed transcriptional state (ASTS) across cancer cell lines. Integration with pharmacogenomic and CRISPR dependency datasets showed that higher ASTS was associated with resistance to EGFR-targeted drugs and reduced EGFR dependency, together with increased dependency on mitochondrial pathways, including mitochondrial translation and respiratory electron transport. These associations were not attributable to a single major cancer lineage and remained evident after adjustment for canonical oncogenic drivers and cell-state covariates. ASTS provides a proof of concept that an endocrine-derived transcriptional state can reveal candidate functional vulnerabilities in an unrelated cancer context.

The human brain transforms a brief glance into a neural representation flexible enough to serve many behavioural goals. This flexibility may reflect multidimensional visual representations read out selectively across tasks, rather than goal-specific instantiation. Combining a large-scale EEG dataset with a behavioural battery of 12 tasks spanning 200 objects, including perceptual, categorical and social decisions, we tested how behaviours are supported by stimulus-evoked representations. Information relevant to 11 of 12 tasks was accessible in the neural signal, emerging early and overlapping in time. Orthogonal behavioural dimensions showed distinct temporal profiles, indicating a genuinely multidimensional representation. The neural code converged on a human-relatedness axis, while feature-level information remained accessible. Behavioural accessibility was thus broad but structured. Object familiarity alone was reliable in behaviour yet absent from the shared neural signal, revealing a limit on what a glance makes accessible. We propose that sustained, recurrent processing serves this broad behavioural accessibility.

Mechanistic understanding of viral protein-protein interactions enables global health security and pandemic preparedness, yet experimental characterisation remains difficult, costly, and often restricted to specialised laboratories. Here, we describe an in silico campaign using AlphaFold2 and AlphaFold-Multimer to predict monomers and dimer structures within 2,812 viral proteomes from 23 viral families relevant to human health. We report high-confidence predicted structures of 5,279 hetero-, and 2,749 homo-dimers. Structural clustering of interfaces reduces the set fivefold to 1,598 consolidated by form and function, of which 471 (29.5%) lacked detectable similarity to experimentally determined interfaces in the PDB. These clusters yielded two notable findings: new data on vaccinology-relevant glycoproteins, and accurate prediction of the specificity and cleavage-site recognition of viral proteases, which are targets for anti-viral development. Viral monomers and high-confidence dimers are available for interactive browsing in the Pandemic Preparedness Portal of the AlphaFold Database (https://alphafold.ebi.ac.uk/), and the full dataset is available for bulk download.

Folate is co-administered with fluoropyrimidines in colorectal cancer to potentiate thymidylate synthase inhibition, yet its therapeutic effects are paradoxically inconsistent across disease and treatment settings. Here we show that one-carbon metabolism remodels epithelial identity before treatment and thereby alters fluoropyrimidine response. Folate restoration shifted colorectal cancer cells along a polarized epithelial axis from a stem-like toward a differentiated state, whereas folate deprivation favored the opposite configuration. These pretreatment states differed markedly in 5-fluorouracil sensitivity. Folate restoration increased sensitivity across colorectal cancer cell lines and patient-derived organoids, whereas folate deprivation or methotrexate increased therapy-induced senescence. State-dependent sensitivity was not explained by proliferation alone, as stem-like cells were least responsive under the same conditions in which their S-phase occupancy was highest. Mechanistically, folate restoration increased methylation potential, supported EZH2-dependent H3K27me3 remodeling, reorganized chromatin accessibility and transcription-factor engagement, and reduced canonical WNT activity. In human colorectal tissue, stem-like program activity was higher and differentiation program activity lower in carcinoma than in normal mucosa, with adenomas intermediate, a pattern opposite to folate-induced remodeling. Folate is therefore not merely a cofactor for thymidylate synthase inhibition but an instructive metabolic regulator of the epithelial state that stratifies fluoropyrimidine response.

Respiratory complex I, a crucial metabolic enzyme, uses the reducing potential of NADH to pump protons across energy-transducing membranes, but its mechanism of catalysis remains unknown. Here, guided by detailed structural information, we combine targeted mutagenesis and biophysical analyses to define the proton-pumping pathways in the three antiporter-like (proton-pumping) subunits of complex I from Paracoccus denitrificans and Escherichia coli. In Pd-CI, a four-proton pump, one of the three proton-uptake pathways is inactive, reconciling the four-proton/three-subunit mismatch. In Ec-CI two pathways are inactive - and Ec-CI pumps only two protons. We define the structural determinants for redox-coupled proton uptake, establish a mechanistic framework for proton pumping, and reveal how the proton-pumping stoichiometry has been adapted to deliver efficient energy conservation in diverse bioenergetic environments.

The gut microbiome develops from birth and is associated with cognition and mental health, but its relationship with early brain maturation remains unclear. We studied 223 typically developing infants aged 0-3 years using multimodal MRI and paired fecal shotgun metagenomics. Microbiome-wide association analysis linked microbial features to cortical morphology, white-matter microstructure, and functional networks. Regularized canonical correlation analysis identified structural-metabolic and functional-taxonomic covariance patterns. Sparse partial least-squares analysis recapitulated both full-sample CCA patterns and their defining structural and functional phenotypes, providing cross-method corroboration. Longitudinal canonical-score slopes did not significantly covary, whereas earlier neuroimaging CCA1 scores predicted microbiome CCA1 scores 1-6 months later in bidirectional autoregressive cross-lag models. These observational results identify coordinated developmental patterns and temporal ordering without establishing causality.

Analysis of the Allen Institute Visual Coding Neuropixels dataset revealed a ~5 Hz synchronization of visual cortical activity in awake, immobile mice. During bouts of this rhythm, synchronized cortical bursts coincided with changes in the occurrence of sharp-wave ripples (SWRs) and dentate spikes and in SWR properties. Cortical synchronization, SWRs, and dentate spikes were also coordinated with slower pupil-size fluctuation, revealing that awake SWRs are embedded within nested brainwide rhythms.

Multidrug-resistant pathogens demand novel resistance-breaking strategies. The oligoarylamide albicidin and pyrrolobenzodiazepines (PBDs) are potent antibacterials with unrelated scaffolds, yet both are neutralized by albA gene products in many Gram-negatives: AlbA, an albicidin-responsive self-upregulating MerR-family factor, and the smaller AlbAS, its unique ligand binding domain (LBD) alone. How AlbA couples ligand sensing to transcriptional activation has remained elusive. Here, using an integrative multidisciplinary approach, we define this mechanism and show that albicidin and PBDs elicit different responses. Crystal structures reveal C8-linked PBDs bound to AlbAS at the N- and C-terminal subdomains (NTD, CTD) of the central tunnel in poses dictated by C8-tail chemistry. This plasticity sequesters diverse PBDs with nanomolar affinities. Cryo-EM shows AlbA as an autoinhibited dimer in which a reciprocal arm closes the NTD end of the partner's tunnel, restricting PBD binding to the CTD while the DNA binding domains (DBDs) are mobile. Albicidin binding is incompatible with this arrangement, whereas PBDs, as CTD plugs, do not perturb the native equilibrium between autoinhibited and promoter-competent states. We visualized the latter by cryo-EM in the RNAP-DNA-AlbA complex. By blocking albicidin-mediated enhancement in vitro, PBDs act as resistance-breaking partners for albicidin and a route to overcoming albA-dependent resistance to oligoarylamide antibiotics.