Biology subjects

Chen, J.

Publications and source records attributed to Chen, J..

At least 109 records · Page 6Linked to original sources

bioRxiv2026

The Proposed Bone Post-Arterial Type R Capillaries Resolve into Venous and Fatty Acid-Handling Endothelial Cells

Endothelial specialization is increasingly recognized as a fundamental regulator of tissue homeostasis, yet the cellular diversity of the skeletal vasculature remains incompletely resolved. Here, we integrate large-scale single-cell transcriptomics, cross-tissue comparisons, and imaging to comprehensively define endothelial heterogeneity across the skeleton. Our analyses demonstrate that the proposed post- arterial "type R" endothelial population is not a distinct endothelial subtype but instead comprises canonical venous endothelial cells and fatty acid-handling endothelial state. RNA velocity supports a venous continuum, while the proposed type R markers FMO2, and AQP7 lack both endothelial and skeletal specificity. The fatty acid-handling endothelial state, characterized by Lpl and Cd36 is conserved across multiple skeletal sites and non-skeletal tissues, indicating a general endothelial metabolic programme. Within bone, this endothelial state expands following high-fat diet and is suppressed during injury. Together, these findings redefine skeletal endothelial heterogeneity and establish the proposed type R population as part of a venous continuum.

cell biology↗

bioRxiv2026

A surface-intrinsic framework for topology-preserving hippocampal alignment and precision morphometry

Accurate alignment of hippocampal anatomy across individuals remains challenging due to complex and highly variable folding patterns that are not well captured by conventional volumetric approaches. HippUnfold introduced a surface-based representation of the hippocampus, but key components--including coordinate estimation and inter-subject correspondence--were defined in the volumetric domain, making them susceptible to topological errors and interpolation artifacts. Here, we introduce a surface-intrinsic formulation of hippocampal unfolding in which geometry, intrinsic coordinates, and correspondence are defined directly on subject-specific surface manifolds. Intrinsic anterior-posterior and proximal-distal coordinates are computed by solving Laplace equations on the surface, and correspondence is established through surface-based resampling in unfolded space, replacing inverse volumetric warping. Relative to the original HippUnfold approach, this formulation improves test-retest consistency, subject identifiability, and mesh quality, while better preserving subject-specific gyral and sulcal morphology. Surface representations show reduced distortion between folded and unfolded spaces and eliminate misplaced or outlier vertices associated with volumetric warping. These improvements translate to enhanced sensitivity in a clinical application, improving lateralization of temporal lobe epilepsy. These results demonstrate that a surface-intrinsic formulation provides a principled and robust foundation for hippocampal unfolding, enabling topology-preserving alignment and more accurate characterization of inter-individual variability in health and disease.

neuroscience↗

bioRxiv2026

Large-scale functional coupling and computational modelling reveal frontotemporal hotspots in distributed network connectivity during working memory recognition, encoding, and retrieval

Working memory depends on coordinated activity across distributed brain networks, but how these interactions support recognition, encoding, and retrieval remains unclear. Here, we analyzed intracranial local field potentials from 1,652 sites in 36 epilepsy patients performing a naturalistic card-matching task. Phase-amplitude and phase-phase coupling revealed widespread condition-dependent interactions, with frontal and temporal regions emerging as recurrent network hubs. Event-related analyses showed reproducible within-trial coupling dynamics across participants. Directed phase transfer entropy further identified frequency-specific information flow, including frontal regions acting as a theta-band source and an alpha-band sink. Finally, a time-delay-embedded hidden Markov model identified task-locked latent states with condition-dependent occupancy trajectories and distinct coherence profiles. Together, these results suggest that working memory is implemented through dynamic, directionally organized large-scale networks, in which frontotemporal hubs coordinate flexible transitions among distributed neural states rather than operating in isolation.

neuroscience↗

bioRxiv2026

Uni-TINT: Unifying T and Innate Lymphoid Cell Taxonomy with a high resolution, pan-disease and pan-tissue single-cell transcriptomic atlas

T cells and innate lymphoid cells (ILCs) exhibit extensive phenotypic diversity across tissues and diseases, yet inconsistent annotation limits cross-study comparisons and biological interpretation. We present Uni-TINT, an integrated pan-disease and pan-tissue atlas comprising 3.46 million cells from 1,869 samples spanning 194 studies, 166 disease subtypes, and 66 tissue types. Through systematic, hierarchical manual annotation, Uni-TINT establishes a unified and context-aware taxonomy of T and ILC populations, resolving 207 cell subtypes and states across conventional and unconventional T cells, natural killer (NK) cells, helper ILCs, thymocytes, and hematopoietic progenitors. We identified an immunosuppressive, tumour-associated CD4 T regulatory population which we validated with spatial transcriptomics. Other rare and unconventional populations characterised included CD8 regulatory T cells, invariant NKT cells and memory-like NK cells. Integration of T cell receptor sequencing suggested functional associations between {gamma}{delta} T cell co-receptor expression and TRDV gene usage. Finally, a comparative analysis of healthy and diseased immature cells identified a small population of malignant hematopoietic stem cells carrying chromosomal aberrations and enriched in acute leukaemia of mixed phenotype. Together, Uni-TINT provides a unified reference framework for immune annotation and discovery across health and disease.

immunology↗

bioRxiv2026

Health-associated gut bacteriocins target TLR4 to suppress intestinal inflammation

The human microbiome maintains host immune homeostasis by secreting bioactive metabolites. However, extending beyond well-characterized metabolites, the functions of most microbiome-encoded peptides remain poorly defined. In this study, we developed a multi-cohort metagenomic framework to profile protective class II bacteriocins--unmodified, ribosomally synthesized peptides--that are enriched in healthy individuals but depleted in patients with inflammatory bowel disease (IBD). We have designated these health-associated bacteriocins as gutcins. Two gutcins, which lack canonical antimicrobial activity, potently attenuate intestinal inflammation in murine models. Cryo-electron microscopy (cryo-EM) reveals that one gutcin, named gutcin 03, directly engages the C-terminus of TLR4, blocking its dimerization and downstream inflammatory signaling. Guided by this structural interface, we generated truncated variants with enhanced potency, demonstrating the amenability of these simple peptides to rational optimization. Collectively, our findings reposition class II bacteriocins from antimicrobial agents to endogenous immunomodulatory effectors and establish a structural and mechanistic foundation for their development as next-generation therapeutics for inflammatory intestinal disorders.

microbiology↗

bioRxiv2026

A fast 2C-induction method reveals a barrier role of SP2 for totipotency

The acquisition of totipotency in vitro remains a major challenge, limiting our understanding of early embryogenesis and its clinical and translational applications. Here, we reported a robust and rapid chemical reprogramming system that converts mouse embryonic stem cells (mESCs) into totipotent-like cells (TLCs) within 36 hours, with efficiencies exceeding 70%. The induced cells exhibit transcriptomic, epigenomic, and functional features closely resembling 2-cell (2C) stage embryos, as confirmed by scRNA-seq, ATAC-seq, and chimeric embryo analyses. Single-cell trajectory construction revealed a branching reprogramming process that yields both successful totipotent-like and alternative, non-reprogrammed fates. Mechanistically, we identified the transcription factor SP2 as a barrier to totipotency acquisition, promoting lineage-specific gene expression while repressing totipotency networks. Collectively, our study has established a highly efficient in vitro model for totipotency induction and highlighted molecular barriers shaping cell fate decisions, providing a platform to dissect the mechanisms governing the pluripotency-to-totipotency transition.

developmental biology↗

bioRxiv2026

A Chairside Multimodal Platform for Temporomandibular Joint Biomechanical Assessment: Technical Evaluation and Illustrative Application

BackgroundTemporomandibular joint (TMJ) biomechanics can be characterized by mandibular motion, masticatory muscle activity, and bite force generation. When acquired synchronously, these functional variables can serve as model-ready inputs for subject-specific computational analyses of internal joint mechanics. However, existing tools typically measure these signals using separate hardware and software platforms, limiting synchronized acquisition within a clinically practical chairside workflow. MethodsWe developed and technically evaluated a compact multimodal platform for chairside acquisition of TMJ functional data and demonstrated its analytical utility in an illustrative orthognathic surgery application. The platform integrates motion, bite force, muscle activity, acoustic, and event-timing measurements with software for real-time preview, protocol guidance, and synchronized export. We assessed technical performance and chairside feasibility and analyzed representative pre- and postoperative data from an orthognathic surgery patient using kinematic, force-control, and computational modeling workflows. FindingsMotion capture demonstrated submillimeter accuracy, with static and dynamic errors of approximately 0.04 mm and 0.12 mm. Bite force sensors showed excellent linearity (R{superscript 2} = 0.998). Chairside deployment required approximately 15 minutes each for setup and data collection. The illustrative case demonstrated that synchronized chairside data can support preoperative and postoperative kinematic analysis, bite force control capacity assessment, and estimation of TMJ disc stress. InterpretationThe proposed platform enables time-efficient chairside acquisition of synchronized, model-ready multimodal datasets for quantitative TMJ biomechanical assessment. This platform and workflow could support subject-specific biomechanical analysis and future clinical studies of temporomandibular joint function.

bioengineering↗

bioRxiv2026

Multiscale Biomechanical and Electrophysiological Modeling of Nociceptive Activation in Musculoskeletal Joint Disorders: Insights from the Temporomandibular Joint

Background and objectiveMusculoskeletal joint disorders often show inconsistent relationships between structural degeneration and nociceptive pain. Temporomandibular joint (TMJ) disc displacement represents a clinically relevant model for investigating the structure-function-pain relationship. This study aimed to develop a multiscale computational framework integrating biomechanics, three-dimensional (3D) neural morphology, and electrophysiology to quantitatively link TMJ structural alterations, biomechanical loading, and peripheral nociceptive activation. MethodsStrain distributions in the TMJ disc and retrodiscal tissue during mouth opening and clenching were computed in ArtiSynth under varying degrees of displacement. Human TMJ 3D nerve architecture was reconstructed using porcine TMJ nerve mapping data as an anatomical reference, and ion channel dynamics were implemented in NEURON. Model coupling was achieved by mapping biomechanical strain fields onto nociceptor membranes to simulate mechanosensitive currents and action potential propagation to the trigeminal ganglion. ResultsAnterior DDwoR induced a severity dependent strain pattern in the TMJ disc and retrodiscal tissue, including posterior redistribution, increased strain magnitude, prolonged activation, and broader retrodiscal tissue involvement. Displacements of 4, 6, and 8 mm produced larger mechanosensitive currents, broader terminal depolarization, and higher trigeminal firing rates during mouth opening (6, 18, and 28 Hz) and clenching (8, 20, and 28 Hz), whereas 0- and 2-mm displacements produced negligible neural activation. ConclusionsThis study establishes a multiscale biomechanical-electrophysiological framework linking TMJ structural alterations to peripheral nociceptive activation. The framework quantitatively connects macroscale strain patterns with microscale neural activation, suggesting that anterior disc displacement may amplify peripheral nociceptive signaling by increasing the overlap between elevated strain and densely innervated retrodiscal tissue.

bioengineering↗

bioRxiv2026

Deep learning design and in vivo validation of Müller glia-specific cis-regulatory elements

Effective recombinant adeno-associated virus gene therapies require promoters that are compact and cell-type-specific. Ideally, promoters should also exhibit functional conservation when tested in model organisms to ensure that preclinical findings translate reliably to human patients. Here, we introduce a deep learning framework for designing cis-regulatory elements (CREs) meeting these criteria, applied to retinal Muller glia (MG). Using single-cell chromatin accessibility data from human and mouse retinas, we trained species-specific models to predict cell-type accessibility, and designed compact CREs using two complementary strategies. In silico validation predicted that the designed CREs exhibit high MG-specific accessibility (on-target) in both species with minimal off-target accessibility across hundreds of human cell types and tissues. Mechanistic analysis revealed that the predicted MG accessibility is driven by the creation of LHX2 motifs. In vivo validation confirmed that the designed CREs successfully restrict reporter expression to MG in the murine retina. Our deep learning framework is highly generalizable and enables the rapid design of compact, on-target, and species-conserved CREs for precision gene therapy.

genomics↗

bioRxiv2026

Pervasive historical introgression across oak phylogeny shapes gene expression

Oaks were once described as the "worst-case scenario" for the biological species concept because of the presumed pervasive hybridization and introgression among species. However, we still lack a full picture of genome-wide introgression across the eight sections of the Quercus phylogeny. Furthermore, we still do not have a complete characterization of what is introgressed and how it functionally influences the recipient species. Firstly, we used 168 whole genomes from 67 Quercus species belonging to the eight sections of the Quercus phylogeny and a super pan-genome to generate a phylogeny and test for ILS and introgression. Introgression was widespread: ancient introgression occurred between ancestors of the eight sections and more recent introgression took place within sections. Secondly, using genome-wide transcriptome, methylome, and chromatin accessibility data from a subset of 19 species sampled from four sections we demonstrated that introgressed genes are more highly and uniformly expressed than non-introgressed ones. This difference in expression level between introgressed and non-introgressed genes mostly results from cis-regulation, and is most likely due to DNA methylation decay in promoter ACRs. In summary, introgression has been a major evolutionary force during the whole evolutionary history of Quercus and had a significant impact on fundamental processes such as gene regulation and expression.

evolutionary biology↗

bioRxiv2026

Multiscale alterations and cortex-wide organizational trends in focal cortical dysplasia

BACKGROUND AND OBJECTIVESFocal cortical dysplasia (FCD) is a leading cause of surgically remediable pharmaco-resistant epilepsy. Previous research has used various MRI sequences to profile FCD lesions, but multi-site studies using personalized features derived from sequences routinely used in the clinic are sparse. This study aimed to quantify changes in cortical morphology, myeloarchitecture, and function within FCD lesions, compare MRI signatures between lesion subtypes and outcomes, and investigate links to macroscale brain organization. METHODSThis cross-sectional study included patients with FCD-related epilepsy aggregated across 3 datasets from Canada and Germany. The 3T MRI acquisitions included T1-weighted and fluid-attenuated inversion recovery (FLAIR) sequences, with the addition of a resting-state functional sequence for two datasets. We derived cortex-wide maps of patient-specific variations in morphology, myeloarchitecture, and function. Variations were quantified in lesions, ipsilateral cortex, and homotopic regions using age- and sex-adjusted normative models. Subgroup analyses explored the role of histological subtype and surgical outcome. Variations were also related to anterior-posterior and sensory-association organizational gradients. RESULTSWe included 159 patients with epilepsy and FCD (47 % female) and 183 healthy control participants (50 % female). Lesions exhibited changes in morphology (cortical thickness; FLAIR blurring) and in specific measures of depth-dependent intracortical myelin (variance and kurtosis of intracortical myelin profiles across depths), but not in local function (regional homogeneity; node strength). Spatial contextualization indicated more pronounced thickness changes in transmodal association cortices, while increased blurring of the gray-white matter interface co-localized with posterior cortical regions. FCD Type IIb lesions contained more marked changes in blurring and depth-dependent intracortical myelin (kurtosis of intracortical myelin profiles across depths) than Type IIa lesions. DISCUSSIONOur findings provide robust evidence that multiscale MRI profiling can identify FCD signatures and contribute to in-vivo subtyping. Furthermore, the novel use of myeloarchitecture profiling and contextualization with macro-scale brain gradients provides new avenues to understand intracortical alterations and the embedding of FCD lesions into broader organizational patterns.

neuroscience↗

bioRxiv2026

A Lipocalin and a Hedgehog-related protein are partners in the C. elegans pre-cuticle apical extracellular matrix

Apical extracellular matrices (aECMs) line exposed body surfaces to shape tissues and protect them from the environment. These aECMs often organize into complex patterns and structures, but how such matrices assemble remains poorly understood. Caenorhabditis elegans cuticle patterns initiate within the transient pre-cuticle, which then helps direct the placement of cuticle collagens. Pre-cuticle patterns arise through post-secretory sorting, which must involve specific molecular interactions among them. Consistent with such a model, Alphafold3 predicts a high confidence physical interaction between two pre-cuticle proteins, the lipocalin LPR-3 and the Hedgehog-related protein WRT-10, with a conserved N-terminal region of LPR-3 forming a {beta}-strand that incorporates into the {beta}-barrel-like structure of the WRT-10 WRT domain. Genetic studies showed that WRT-10 requires this LPR-3 region in order to become properly patterned in the pre-cuticle matrix. Furthermore, WRT-10 and the LPR-3 {beta}-strand region are required to pattern a specific cuticle substructure, the lateral alae ridges, but not for other LPR-3-dependent matrix roles. These data indicate that LPR-3 and WRT-10 are functional partners and support a "landing pad" model whereby physical interactions between them allow LPR-3 to recruit WRT-10 to specific aECM regions. Similar mechanisms may explain how other members of the C. elegans Hh-r family associate with the aECM. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/739337v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@1fdaf57org.highwire.dtl.DTLVardef@2b752org.highwire.dtl.DTLVardef@51a91forg.highwire.dtl.DTLVardef@13665ad_HPS_FORMAT_FIGEXP M_FIG C_FIG Article SummaryAll animal skin is covered by a set of proteins, sugars and lipids that comprise the apical extracellular matrix (aECM). These matrix components can be organized into patterned ridges and other distinctive structures. This study addresses how such patterns form in the developing cuticle of the nematode C. elegans. The study provides evidence for a regulatory mechanism that enables one matrix protein to establish a pattern and then recruit a second protein into the same pattern.

developmental biology↗

bioRxiv2026

Insulin-degrading enzyme activity is modulated by interaction with pyrroline-5-carboxylate reductase 1

Insulin-degrading enzyme (IDE, insulysin, insulinase) is a peptidase that hydrolyzes a number of bioactive peptides including insulin and the amyloid beta peptide, making it a promising therapeutic target for diabetes and Alzheimers disease. Aspects of its physiological role are still controversial, however. In an attempt to further define IDEs role in cells, we used co-immunoprecipitation experiments to identify potential IDE interacting proteins. The enzyme pyrroline-5-carboxylate reductase 1 (PYCR1) was found associated with IDE in three different cell lines, and the two proteins colocalize in HeLa cells. Purified PYCR1 activates IDE toward small peptide substrates, suggesting a modulatory role for the interaction in vivo. Modeling suggests that the unstructured N-terminal region of PYCR1 inserts into allosteric sites of IDE, contributing to the observed activation. Deleting this sequence alters, but does eliminate, the interaction between PYCR1 and IDE. Since pyrroline-5-carboxylate reductase 1 is a mitochondrial protein, we posit that their interaction could regulate a previously described mitochondrial pool of IDE, which may serve to degrade mitochondrial targeting sequences or amyloid beta peptide that localizes to that organelle.

biochemistry↗

bioRxiv2026

A novel AR molecular glue degrader via bivalent engagement of the N-terminal domain and RNF213-UBE2J2 complex

Targeted protein degradation enables the elimination of disease-relevant proteins through induced proximity between E3 ligases and substrates. Current androgen receptor (AR) therapies act via the ligand-binding domain (LBD) but fail against constitutively active LBD mutants or splice variants such as AR-V7. Here, we identify GZL626, a small-molecule degrader that induces proteasome-dependent degradation of both full-length AR and AR-V7. GZL626 suppresses AR transcriptional activity, inhibits proliferation of AR-positive prostate cancer cells, and blocks SARS-CoV-2 infection. Mechanistically, GZL626 binds the intrinsically disordered AR N-terminal domain (NTD) and promotes formation of a noncanonical RNF213-UBE2J2-AR ternary complex that drives AR ubiquitination and degradation. The AR DNA-binding domain (DBD) acts as a structural hub within this assembly. Distinct from PROTACs or classical molecular glues, GZL626 engages both AR and RNF213 bivalently, remodeling protein interactions to stabilize a functional E3-E2-substrate complex. These findings establish GZL626 as a first-in-class NTD degrader with potential to overcome resistance in advanced prostate cancer.

biophysics↗

bioRxiv2026

Predicting the protein interaction landscape of a mycobacterial pathogen

High-dimensional phenotypic screens of bacterial loss-of-function mutant libraries have determined gene-gene connections and specific phenotypes for thousands of bacterial genes in many species, but deciphering the underlying mechanisms remains decidedly low-throughput. Here, we demonstrate the utility of proteome-wide AI-based protein-protein interaction (PPI) predictions for overcoming this gap by using pooled-AlphaFold3 to assess all [~]1.3 million possible pairwise interactions in the proteome of Mycobacterium leprae. We identify [~]2,000 strong and intermediate PPIs that underlie a significant fraction of phenotypes and gene-gene connections observed in large-scale chemical genomics screens from Mycobacterium tuberculosis, Mycobacterium smegmatis, and Corynebacterium glutamicum. This combined approach predicts specific functions for dozens of previously uncharacterized core, conserved, and essential mycobacterial proteins. We highlight new information derived from the study, including insights into mycobacterial envelope assembly, peptidoglycan remodeling, and new modulators of the central dogma enzymes RNA polymerase and DNA gyrase. These data establish combined pooled-AlphaFold3 PPI prediction and high-throughput genomics approach as the gold standard for large-scale characterization of protein function.

systems biology↗

bioRxiv2026

Gut Virome Diversity and Health Status in Rescued Pangolins: Metaviromics Reveals Parvovirus as a Key Virus Associated with Disease

BackgroundPangolins are critically endangered mammals that suffer from high rates of gastrointestinal disease during captivity, yet the role of the gut virome in their health remains unexplored. This study presents the first comprehensive characterization of the gut DNA virome in Malayan (Manis javanica) and Chinese (M. pentadactyla) pangolins across different health states. ResultsMetaviromic sequencing of 16 fecal samples from healthy, diarrheal, pneumonic, free-ranging, and deceased pangolins generated 7.2-11.8 Gb clean data per sample. A total of 12 viral phyla, 26 families, 219 genera, and 1,132 species were identified. Caudovirales phages (Siphoviridae, Myoviridae, and Podoviridae) dominated the gut virome of healthy individuals, with phage content exceeding 90% in most healthy samples. However, diseased and deceased individuals exhibited a significant reduction in phage proportion (94.5% vs. 53.3%, P = 0.02), accompanied by a dramatic increase in eukaryotic viruses--particularly Parvoviridae, which accounted for 66% of the virome in deceased Malayan pangolins. Iridoviridae (38%) and Polydnaviridae (34%) dominated in deceased Chinese pangolins. SIMPER analysis identified Parvoviridae as the primary contributor to virome dissimilarity between healthy and diseased Malayan pangolins, whereas Iridoviridae and Polydnaviridae were the key contributors for Chinese pangolins. LEfSe analysis revealed 12 biomarker viruses in free-ranging pangolins (predominantly Staphylococcus and Escherichia phages), six in healthy Chinese pangolins (predominantly Streptococcus phages and Lactobacillus viruses), and six in deceased Malayan pangolins (predominantly vertebrate viruses). KEGG functional annotation revealed that genes related to DNA replication, repair, and recombination were the most abundant, suggesting frequent genomic recombination within the pangolin gut virome. ConclusionsThe gut virome of pangolins is closely associated with health status. Disease induces a shift from a phage-dominated to a eukaryotic virus-dominated virome, with Parvoviridae and Iridoviridae emerging as candidate pathogenic viruses in Malayan and Chinese pangolins, respectively. These findings provide molecular evidence for virome monitoring in pangolin conservation and highlight the need for targeted surveillance of ImportancePangolins are the most trafficked mammals in the world, and all eight species are critically endangered. During rescue and captive care, these animals suffer from high rates of gastrointestinal disease, yet the role of viruses in their gut health has never been studied. This research provides the first comprehensive map of the gut virus community in pangolins, revealing that healthy individuals carry a virus community dominated by beneficial bacterial viruses called phages, while sick and dying animals show a dramatic shift toward disease-causing viruses. Notably, parvoviruses emerge as potential pathogens in Malayan pangolins, while iridoviruses dominate in deceased Chinese pangolins. These findings offer a new tool for monitoring pangolin health in rescue centers: by tracking the balance between beneficial phages and harmful viruses, veterinarians may be able to detect illness earlier and improve survival rates. The study also highlights the importance of environmental exposure for maintaining a healthy gut virus community, supporting the creation of semi-natural enclosures for rescued pangolins. Beyond conservation, this work contributes to our understanding of how viruses move between wildlife and humans, which is critical for preventing future disease outbreaks.

microbiology↗

bioRxiv2026

FUNCTIONAL FRAGMENTATION AND STRUCTURAL DRIVERS OF THALAMO-CORTICAL CIRCUITS IN TEMPORAL LOBE EPILEPSY

AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSObjectiveC_ST_ABSIn temporal lobe epilepsy (TLE), the thalamus acts as a nexus in a pathophysiological network that implicates mesiotemporal, subcortical, and neocortical regions. Studying a large multimodal and multicentre dataset, we profiled thalamic, hippocampal, and neocortical functional connectivity (FC), assessed structural mediators, and examined clinical associations. MethodsWe studied resting-state FC alongside structural and diffusion MRI data in 250 unilateral TLE patients and 259 healthy controls, with measures aggregated across four independent datasets. Data were processed using open-access neuroinformatics workflows and analyzed at a subregional level to maximize anatomical precision. Statistical analysis and mediation models assessed between-group FC changes, structural contributors, and clinical correlations. ResultsCompared to controls, TLE patients presented with reduced thalamo-cortical FC, which was most marked in mesiotemporal, fronto-central, and occipital regions. Thalamo-hippocampal FC was also reduced, with effects seen in all CA subfields. In the thalamus, FC reductions peaked in the ventral posterior nucleus when considering neocortical target regions and in the mediodorsal nucleus when considering hippocampal target regions. While ipsilateral hippocampal volume and diffusion changes mediated thalamo-hippocampal FC, thalamo-cortical FC appeared decoupled from structural alterations. Findings were consistent in left and right TLE patients, in patients with short and long disease duration, and across imaging sites, suggesting that thalamo-cortical FC imbalances are a consistent signature of TLE. Conversely, thalamo-hippocampal FC was elevated in patients with focal-to-bilateral-tonic-clonic seizures and FC alterations were more marked in the subgroup of operated patients that became seizure-free after surgery. ConclusionOur multi-site findings demonstrate marked thalamic circuit fragmentation in TLE. Ipsilateral findings robustly showed subdivision-specific effects, which point to both mesiotemporal co-lateralization as well as broader system-level involvement. Mediation analyses furthermore confirmed a key role of hippocampal pathology in disrupted thalamo-hippocampal connectivity in TLE, while broader thalamo-cortical fragmentation becomes increasingly independent of mesiotemporal compromise. Critically, thalamic FC represents a network substrate for seizure generalization and can serve as a prognostic indicator for surgical outcome. These results underscore the contribution of the thalamus as a hub in macroscale dysfunction in TLE.

neuroscience↗

bioRxiv2026

MKMC enables reference-free transcriptomic analysis using k-mer representations

Traditional RNA-seq analysis depends heavily on genome alignment and gene annotation, limiting its utility in non-model organisms and introducing biases that can obscure regulatory complexity. We present MKMC (Multi-sample Kmer Counter), a scalable, reference-free toolkit for RNA-seq analysis that leverages k-mer-based statistics to detect biological variation without requiring alignment. MKMC integrates fast k-mer counting, abundance matrix generation, normalization, dimensionality reduction, and differential analysis into a unified workflow. Across diverse datasets, MKMC recapitulates key biological signals--including sex differences in killifish liver--and matches alignment-based pipelines in differential expression analysis and transcriptomic age prediction. Notably, MKMC detects isoform-specific events missed by traditional methods, one of which we validated using in situ hybridization. These results reveal previously hidden isoform-level regulatory events that contribute to sex-and age-associated transcriptional programs. MKMC offers a robust, extensible alternative to alignment-based approaches, enabling transcriptomic discovery across both model and non-model systems. While we focus here on RNA-seq as a primary application, MKMC is broadly applicable to any k-mer-based analysis of next-generation sequencing data.

bioinformatics↗