Biology subjects

Su, B.

Publications and source records attributed to Su, B..

At least 19 recordsLinked to original sources

bioRxiv2026

CryoCodex: learning discrete structural representations for cryo-EM map post-processing

Cryo-EM maps frequently suffer from weak structural signals, spatially variable quality, and noise, complicating interpretation and downstream modeling. Existing post-processing approaches treat density enhancement, quality assessment, and masking as separate tasks, often amplifying residual noise or introducing local artifacts. Here, we present CryoCodex, a unified framework that jointly performs density enhancement, local quality estimation, and molecular masking. CryoCodex introduces a multi-head, multi-scale vector-quantization module that projects encoder-derived latent features onto a shared codebook of structural prototypes. By imposing discrete constraints across feature subspaces and spatial scales, the codebook suppresses stochastic noise while preserving recurrent structural patterns. On a non-redundant test set of 126 experimental maps spanning 3-8 [A], CryoCodex outperformed four representative methods, achieving a mean map-model FSC-0.5 of 3.93 [A] versus 4.62 [A] for EMReady2 and 6.87 [A] for the deposited maps. Downstream automated de novo model building with Phenix further confirmed the quality of the CryoCodex-enhanced maps. These gains extended to maps containing nucleic acids, where CryoCodex produced the best map-model agreement and improved model building. Half-map cross-validation indicated that these improvements are anchored to experimental data rather than driven by half-map-specific noise or structural hallucination. CryoCodex also delivered reliable local quality estimates and molecular masks within the same framework. Together, these results establish CryoCodex as a practical tool for cryo-EM map post-processing and structural determination in challenging cases.

bioinformatics↗

bioRxiv2026

An L-threonine-inducible biosensor derived from the tryptophan-repressed trp promoter of Escherichia coli for high-throughput screening of L-threonine overproducers

L-Threonine is an essential amino acid widely used in the feed, food, and pharmaceutical industries, and microbial fermentation is now the dominant route for its production. Further increases in producer titers depend on random mutagenesis coupled with high-throughput screening (HTS), but progress has been limited by the lack of a biosensor with a broad operating range, high sensitivity, and target specificity. Here, we show that L-threonine induces the tryptophan-repressed trp promoter of Escherichia coli, a response that, to our knowledge, has not been described previously. Deletion analysis showed that both the trpO operator and the trpL attenuation element contribute to induction, and that the lowest basal expression occurs only when both elements are present. By optimizing the ribosome-binding site, the promoter -10 region, and TrpR expression, we constructed an L-threonine biosensor on a pACYC backbone. The sensor responded linearly to L-threonine from 0.1 to 30 g/L (R2 = 0.9895), with an induction ratio of up to 51.7-fold. Among the 20 proteinogenic amino acids tested, only L-threonine produced a detectable response. The biosensor distinguished strains with different L-threonine production capacities and guided the directed evolution of homoserine kinase (ThrB) at a conserved motif, yielding variants with brighter reporter fluorescence and increased L-threonine production. This work provides a high-performance biosensor based on native E. coli regulatory elements for screening L-threonine overproducers.

bioengineering↗

bioRxiv2026

Adaptive introgression in Tibetans spans focal and distributed architectures

The Denisovan-derived EPAS1 haplotype is a paradigmatic example of adaptive introgression, but whether archaic ancestry contributed beyond such large-effect loci remains unclear. We analyzed genomes from 1,039 Tibetans, Han Chinese, archaic hominins and ancient Tibetan Plateau individuals. Denisovan-derived adaptive variation was highly localized around EPAS1, whereas Neanderthal-derived variation was distributed across the genome; temporal and elevational patterns reinforced this contrast. Despite individual-level heterogeneity, Neanderthal-introgressed gene repertoires repeatedly converged on immune, metabolic and signal-transduction pathways. A Tibetan-enriched Neanderthal PPARGC1A haplotype was associated with lower fasting glucose and showed concordant support from an independent East Asian type 2 diabetes GWAS. These findings reveal that adaptive introgression can contribute to complex human adaptation through functionally convergent combinations of archaic variants, rather than only through shared large-effect haplotypes.

evolutionary biology↗

bioRxiv2026

Integrated Transcriptomics and Histological Analysis Reveals Gill Adaptation Mechanisms to Hypoxic Stress in Largemouth Bass (Micropterus salmoides)

Largemouth bass (Micropterus salmoides) is a globally significant freshwater aquaculture species known for its rapid growth, high-quality flesh, and excellent nutritional value. Given the increasing incidence of hypoxia in its aquaculture setting, understanding the molecular mechanisms underlying gill adaptation to hypoxic stress is critical for improving resilience in this species. To achieve this, we employed full-length transcript sequencing using PacBio Iso-Seq and high-throughput Illumina RNA-seq to construct a comprehensive transcriptome profile. Histological analyses revealed apparent structural alterations in gill tissues under hypoxic stress (dissolved oxygen: 2.0 +/- 0.5 mg/L, duration: 96 h), including curved and swollen gill filaments, apoptosis, and epithelial cell vacuolation. Comparative transcriptomic analysis revealed pronounced shifts in the expression of genes involved in energy metabolism, angiogenesis, immune responses, cell proliferation, and apoptosis. Notably, the down-regulation of genes associated with cell cycle and proliferation suggests that M. salmoides may conserve energy by suppressing cell division under hypoxic conditions. In contrast, the increased expression of apoptosis-related genes, including Caspase 7, indicates that apoptosis may play a protective role to maintain tissue homeostasis under hypoxia. Furthermore, the suppression of chemokine-mediated immune pathways implies that hypoxia redirects energy toward essential metabolic processes at the cost of local immune responses. GO and KEGG enrichment analyses identified TNF- and chemokine-mediated signaling pathways as key regulators for tissue homeostasis and adaptive responses to hypoxic stress. These findings enhance our understanding of the mechanisms underlying hypoxia adaptation and provide valuable genetic resources for breeding strategies aimed at enhancing hypoxia tolerance in M. salmoides.

genomics↗

bioRxiv2026

The small molecule inhibitor SU056-mediated targeting of YB1 inhibits the Rb pathway in triple-negative breast cancer tumors

Background: Triple-negative breast cancer (TNBC) tumors lack expression of estrogen receptor (ER), progesterone receptor (PR), and HER2, limiting the availability of targeted therapeutic options. As a result, TNBC is characterized by a high propensity for metastasis, rapid recurrence, and poor overall prognosis. Despite significant research efforts, the molecular mechanisms that drive TNBC progression and metastasis remain incompletely understood. Y box binding protein 1 (YB1) is a multifunctional DNA/RNA-binding protein that regulates transcription, translation, and mRNA stability. Aberrant activation of YB1 has been implicated in multiple oncogenic processes, including proliferation, survival, invasion, metastasis, and therapy resistance in several cancer types, including TNBC. Our investigations have revealed a central role for YB1 in regulating key hallmarks of cancer that drive TNBC tumor progression and metastatic dissemination. Methods: Cell viability and drug sensitivity in TNBC cell lines were assessed using the MTT assay to determine IC50 values for pharmacologic inhibitors. Western blotting and quantitative RT PCR were used to quantify protein and mRNA expression levels in TNBC cell lines and tumor samples. Oncogenic phenotypes were evaluated using 2D colony formation assays, 3D tumorsphere growth assays, limiting dilution, and transwell migration assays to assess proliferative, stem-like, and migratory capacities. Cell cycle progression was analyzed using flow cytometry based assays. In vivo therapeutic efficacy was evaluated using preclinical xenograft and patient-derived xenograft (PDX) mouse models of TNBC to assess tumor growth and metastatic progression following treatment. Results: Our studies demonstrate that the small-molecule inhibitor SU056, which specifically targets YB1 for degradation, significantly suppresses the oncogenic behavior of TNBC cell lines and tumors. Pharmacologic inhibition of YB1 reduced TNBC cell proliferation, clonogenic growth, tumorsphere formation, stemness and migratory capacity. Importantly, combined treatment with SU056 and the CDK4/6 inhibitor Palbociclib enhanced the inhibitory effect on TNBC cell growth and tumor progression compared with either agent alone. Mechanistically, both genetic and pharmacologic targeting of YB1 suppressed TNBC tumor growth and metastatic potential through modulation of the Cyclin D/CDK4/6/Rb signaling axis, leading to inhibition of cell cycle progression and induction of G1 arrest. Conclusion: Collectively, these findings identify SU056/CDK4/6i combination as a potential therapeutic option for the treatment of Rb+ TNBC.

cancer biology↗

bioRxiv2026

Sex specificity of inhibitory gating deficits in individuals with high autistic traits

Empirical evidence from traditional inhibitory control tasks regarding inhibitory deficits in high autistic traits has been mixed, indicating that this issue remains controversial. Although sex differences are widely documented in autistic cognitive profiles, their role in inhibitory gating mechanisms remains underexplored. Given that the expression of inhibitory gating deficits may be modulated by the social versus non-social nature of stimuli, and no prior study has investigated this topic by integrating both sex differences and stimulus domain, we addressed these two questions with the attribute amnesia paradigm. We manipulated stimulus type (non-social vs. social). In Experiment 1, participants performed a location task with animal drawings as targets and were unexpectedly asked to report animal identity on a surprise trial. High autistic trait females showed significantly higher accuracy on the surprise trial than all other groups, reflecting a failure to actively filter out task-irrelevant non-social information, that is, a reduced inhibitory gating efficiency. In Experiment 2, using face stimuli and a self-vs. other-face design, this gating deficit was no longer expressed: all groups performed at chance levels on the identity judgment, regardless of autistic trait level, sex, or face type. This dissociation aligns with a dual-mechanism framework: the inhibitory gating deficit in high autistic trait females is specific to non-social stimuli and masked by camouflaging for social ones. This study demonstrates that the inhibitory gating deficit in high autistic trait females is not a global impairment but rather a stimulus-dependent one, highlighting the need to consider sex and stimulus type.

neuroscience↗

bioRxiv2026

Multidimensional variation and population stratification across 8000 complete human centromeres

Human centromeres are indispensable for the faithful segregation of chromosomes during cell division, yet their highly repetitive nature has historically precluded comprehensive characterization, leaving fundamental questions about their sequence diversity, evolution trajectories and function dynamics unresolved. Here, we generated 6,312 complete human centromere sequences from 320 phased genome assemblies in Asian Pan-Genome project phase 1. By integrating the assemblies from the Human Pangenome Reference Consortium (HPRC) and Human Genome Structural Variation Consortium (HGSVC), we constructed a multidimensional genetic variation map encompassing over 8,000 gapless centromeres. Centromeric satellite arrays account for 4.19% to 6.01% of the whole genome, with substantial variations in size and architecture across chromosomes. Using a refined alpha satellite clustering approach that captures global diversity, we identified 195 higher-order repeat (HOR) arrays, 56.4% of which are absent from the T2T-CHM13 reference genome. Extensive structural variations across multiple dimensions exhibit population stratification, including centromeric haplotypes (CenHaps), ultra-large pericentric inversions spanning up to 36.6 Mbp, and inter-chromosomal HOR sharing that reflects sequence exchange among chromosomes. Integrating CENP-A CUT&Tag experiments and long-read-based DNA methylation profiles, we demonstrate that 16.8% of centromeres harbor multiple potential kinetochore assembly sites, and CenHap-specific local HOR homogenization is associated with kinetochore positioning. Despite global suppression of recombination at centromeres, we observed asymmetric linkage disequilibrium flanking centromeres and an ancient recombination event within the centromere of chromosome 19. Furthermore, contrary to the prevailing assumption of high mutation rates in centromeres, our estimates based on stringent orthology reveal no significantly higher single-base substitution rates for centromeres relative to flanking pericentromeric regions with substantial variations across chromosomes, despite extraordinary structural plasticity. Collectively, these multi-scale centromeric variations provide a global view of human centromere diversity and population stratification, fundamentally redefine centromere evolution through a dual-track model balancing structural innovation with mutational constraint, and establish an essential resource for investigating centromere biology and a baseline reference for diagnosing centromere-associated disorders.

genomics↗

bioRxiv2026

Intranucleolar Invasion of Cajal Body Remnants Suppresses Ribosomal Biogenesis in cis and Telomerase Functions in trans

Cellular processes are compartmentalized within immiscible heterotypic condensates, yet the functional consequences of losing their physical segregation remain unclear. Here, we show that genetic inactivation of the RNA chaperone SMN forces the aberrant intermixing of the two most prominent nuclear condensates: nucleolus and Cajal Body (CB). Upon SMN depletion, CB components invade the nucleolus and undergo reduced mobility and solubility consistent with a liquid-to-gel-like hardening transition. The CB-scaffold coilin aberrantly enriches at the nucleolar FC/DFC boundary and occupies rDNA chromatin, thereby locally suppressing rRNA production. Concurrently, this sequestration globally impairs coilin targeting to snRNA/snoRNA loci and limits telomerase access to telomeres, reducing telomeric synthesis. Crucially, genetic coilin depletion alone alleviates this mistargeting and rescues these functional impairments across condensates. Our findings reveal an inter-condensate rheostat model in which the loss of CB-nucleolar immiscibility is directly sensed, communicated, and executed by CB remnants, thereby proportionally coupling the functional outputs of otherwise distinct RNPs essential for splicing, translation, and genomic integrity.

molecular biology↗

bioRxiv2026

SCD-dependent lipid metabolism licenses alternative macrophage activation and macrophage plasticity

Lipid metabolic reprogramming accompanies macrophage activation, yet our understanding of why macrophages profoundly reshape their lipid composition remains unclear. Here, we identify stearoyl-CoA desaturase (SCD) as a lipid-metabolic checkpoint required for the acquisition of the alternatively activated macrophage (AAM) cell state. We show that SCD maintains lipid desaturation balance by ensuring the conversion of newly synthesized saturated long-fatty acids (SFAs) into monounsaturated fatty acids (MUFAs). Critically, disruption of SCD resulted in the rapid accumulation of saturated phosphatidic acid (PA) and hyperactivation of mTORC1, which perturbed the acquisition of stable AAM epigenetic and transcriptional programs. Notably, inhibition of upstream lipogenic enzymes (e.g., ACC and FASN) did not impair AAM polarization, underscoring that an imbalance between newly synthesized saturated and monounsaturated fatty acids, not the loss of de novo lipogenesis, determines AAM identity. Unexpectedly, the loss of SCD redirected IL-4-activated macrophages into an aberrant cell state poised for pro-inflammatory responses, indicating that disruption of this lipid metabolic checkpoint leads to misinterpretation of anti-inflammatory instructional cues. Rescue of SCD-deficient macrophages via mTORC1 inhibition confirmed a causal role for the SCD-PA-mTORC1 axis in controlling macrophage plasticity. Finally, we find that SCD deficiency and resultant PA accumulation attenuated Toxoplasma gondii-driven AAM polarization and reduced pathogen burden. Together, these data establish the SCD-PA lipid signaling axis as a critical metabolic regulator of macrophage plasticity with direct consequences for host defense.

immunology↗

bioRxiv2026

Decoding Immune Reconstitution Failure in People Living with HIV through Single-cell Genomics

Immune non-responders (INRs), a subset of people living with HIV (PLWH), fail to achieve full immune reconstitution and remain at increased risk of morbidity, mortality, and non-AIDS-related illnesses. As the mechanisms underlying this impaired immune recovery remain poorly understood, we performed single-cell multi-omics profiling on peripheral blood mononuclear cells from 43 INRs, 47 immune responders and 53 healthy donors. Our dataset comprises 2,744,009 transcriptomes and 1,226,658 chromatin-accessibility profiles across 58 identified immune cell types. INRs exhibited markedly elevated inflammatory signaling, increased apoptotic activity, and dysregulation of immune-activating ligand-receptor interactions. Additionally, we identified 2,996 interaction cis-eGenes and 5,938 cis-caPeaks, and validated key cell type-specific cis-xQTL effects associated with immune reconstitution failure. Furthermore, we identified 3,551 sc-eQTLs and 872 cell-state interaction eQTLs (ieQTLs) in CD4+ T cells. A genotype-specific upregulation of OAS3 in INRs, likely regulated by STAT1, was identified based on these results. We developed scPRISM (single cell Predictive Reconstitution Immune Status Model), a multi-modal framework that integrates scRNA-seq and scATAC-seq data to perform the dual tasks of differentiating disease states and predicting gene expression from chromatin accessibility, thereby deciphering the cell type-specific cis-regulatory basis of the disease. Our study delineates the transcriptional and epigenomic landscape of immune dysregulation in INRs and provides new genetic insights into HIV-associated immune reconstitution failure. These findings offer a foundation for developing biomarkers and precision therapies to restore immune function in PLWH.

genetics↗

bioRxiv2026

Selective encoding failure of self-face identity in subthreshold depression

Subthreshold depression is associated with significant functional impairment and elevated risk of major depressive disorder. A negative self-concept may disrupt the implicit positive association evoked by ones own face, impairing incidental encoding of self-relevant information. Whether subthreshold depression involves a selective deficit in encoding self-face identity remains unclear. The attribute amnesia paradigm is well suited to address this question because it can dissociate attentional selection from working memory encoding. Using this paradigm, we examined the issue across two experiments. Experiment 1 employed nonsocial stimuli (animal drawings) and confirmed an intact attribute amnesia effect in subthreshold depression (n = 30) comparable to healthy controls (n = 30), ruling out a generalized encoding deficit. Experiment 2 replaced targets with faces (self or other) and revealed a selective enhancement of the attribute amnesia effect for self-face identity in subthreshold depression. Specifically, on the surprise trial, accuracy for self-face identity dropped to near-chance levels in the subthreshold depression group, whereas no such deficit emerged for other-faces or in controls. Encoding recovered rapidly once explicit memory expectations were introduced, indicating intact basic encoding capacity. These findings suggest that subthreshold depression is associated with a specific impairment in incidentally encoding self-face identity. This impairment likely stems from a negative self-concept that weakens self-face salience under incidental encoding conditions. By capturing this selective encoding failure, the present study reveals that the self-processing deficit in subthreshold depression can arise at the gating stage between attention and working memory consolidation.

neuroscience↗

bioRxiv2026

Presenilin 1 (PS1) located at mitochondrial inner membrane regulates mitochondrial cristae junction proteins arrangement and cristae formation in HEK293 cells

Presenilin 1 (PS1), a key pathogenic factor in familial Alzheimers disease, is implicated in regulation of mitochondrial functions, yet its precise sub-mitochondrial localization and underlying mechanisms remain poorly understood. In this study, we generated PS1 knockout (PS1 KO) cell lines to investigate the role of PS1 in mitochondrial structure and function. Our results demonstrated that PS1 is directly localized to the mitochondrial inner membrane. PS1 deficiency led to reduced ATP production, impaired mitochondrial respiration capacity, decreased mitochondrial membrane potential, disrupted Ca2+ homeostasis, and elevated reactive oxygen species (ROS) accumulation. Moreover, loss of PS1 caused abnormal mitochondrial cristae structure. Further analysis revealed that PS1 interacts with mitochondrial inner membrane proteins. Its absence promotes ATAD3A oligomerization and disrupts its arrangement at mitochondrial cristae junctions, leading to expansion of the mitochondria-associated membrane (MAM) and instability of mitochondrial DNA (mtDNA). Our findings demonstrate that PS1 acts as a central regulator of mitochondrial cristae morphogenesis by modulating protein interaction networks at cristae junctions, thereby illuminating fundamental molecular mechanisms contributing to mitochondrial dysfunctions in Alzheimers disease.

cell biology↗

bioRxiv2026

Structural basis for CTCF-mediated chromatin organization

Eukaryotic DNA is organized across multiple scales to support genome compaction, appropriate gene expression, and DNA recombination. A central player in these processes is the CCCTC binding factor (CTCF), which defines specific chromatin loop structures and insulates enhancer elements from promoters. Chromatin is organized in a distinct pattern around CTCF-bound sites; however, the role of this patterning remains unclear. Here, we report cryo-electron microscopy structures of reconstituted CTCF-nucleosome complexes, revealing that CTCF dimerization promotes the oligomerization of nucleosomes into defined higher-order assemblies involving specific histone-histone and CTCF-CTCF interactions. Notably, CTCF does not oligomerize efficiently on non-chromatinized DNA substrates. Disruption of identified CTCF interaction interfaces in cells results in a marked decrease in chromatin looping and functionally impairs cellular differentiation. These results indicate that chromatin structure at CTCF sites plays an important role in supporting higher-order interactions between distal regions of the genome and that these interactions are important for supporting cell-type-specific gene expression.

molecular biology↗

bioRxiv2026

PathDiffusion: modeling protein folding pathway using evolution-guided diffusion

Despite remarkable advances in protein structure prediction, a fundamental question remains unresolved: how do proteins fold from unfolded conformations into their native states? Here, we introduce PathDiffusion, a novel generative framework that simulates protein folding pathways using evolution-guided diffusion models. PathDiffusion first extracts structure-aware evolutionary information from 52 million predicted structures the AlphaFold database. Then an evolution-guided diffusion model with a dual-score fusion strategy is trained to generate high-fidelity folding pathways. Unlike existing deep learning methods, which primarily sample equilibrium ensembles, PathDiffusion explicitly models the temporal evolution of folding. On a benchmark of 52 proteins with experimentally validated folding pathways, PathDiffusion accurately reconstructs the order of folding events. We further demonstrate its versatility across four challenging applications: (1) recapitulating Antons molecular dynamics trajectory for 12 fast-folding proteins, (2) reproducing functionally important local folding-unfolding transitions in 20 proteins, (3) characterizing conformational ensembles of 50 intrinsically disordered proteins, and (4) resolving distinct folding mechanisms among 3 TIM-barrel proteins. We anticipate that PathDiffusion will be a valuable tool for probing protein folding mechanisms and dynamics at scale.

bioinformatics↗

bioRxiv2025

Tibetan near-complete pangenome reveals complex variants underlying high-altitude adaptation

Genomic diversity in indigenous populations offers critical insights into human evolution and population-specific adaptation. Here, we generated 70 near-complete, fully phased genome assemblies from 35 high-altitude Tibetan trios and identified 4.55 million small variants and 63,031 structural variants (SVs) absent from the current global pangenome reference. Comparative analyses with a 595 globally diverse long-read genome dataset revealed 207 previously uncharacterized Tibetan-enriched SVs spanning 275 genes implicated in high-altitude physiological adaptation. Genotyping these SVs in 1,164 Tibetan and Han individuals with 88 physiological traits reveals five variants significantly associated with cardiopulmonary and hematological phenotypes central to hypoxia adaptation. We further map 2.45 Gb of archaic hominin sequences in Tibetans and uncover 18 previously unrecognized introgressed regions enriched in Tibetans with genes related to hematopoiesis and lung function. These regions harbor five Tibetan-enriched SVs, indicating that archaic introgression contributed SVs relevant to human adaptation. Together, these findings reveal substantial uncharacterized genomic diversity in Tibetans and highlight a previously underappreciated role of SVs and archaic SVs in shaping human evolution and high-altitude adaptation.

genomics↗

bioRxiv2025

A near-complete lamprey genome illuminates ancestral vertebrate innovations

The origin of vertebrates is characterized by a suite of phenotypic innovations, yet the underlying genetic mechanisms remain poorly understood. As a living representative of jawless vertebrates, the lamprey serves as a pivotal model for investigating the genomic basis of early vertebrate evolutioin. Here, we report a near telomere-to-telomere (T2T) genome assembly of the reissner lamprey (Lethenteron reissneri), generated from sperm DNA that is not subject to programmed genome rearrangement. The 1.25 Gb assembly resolves over 97% of chromosome sequences into single contigs, enabling reconstruction of previously inaccessible genomic regions. Centromeres display highly diversified, lamprey-specific repeat units and structural architectures, whereas telomeres contain both conserved and lineage-specific satellite repeats. Deep transcriptomic profiling across representative organs and developmental stages substantially improved genome annotation. Comparative genomic analyses revealed a major expansion of 1,562 gene families at the base of the vertebrate lineage, and they are significantly enriched for functions related to neural development, skeletal formation, and cardiac function, echoing the known phenotypic innovations in vertebrates. Notably, the CDH2 gene family underwent vertebrate-specific copy number expansion and acquired conserved cis-regulatory elements (CREs), highlighting its potential role in shaping the early vertebrate circulatory system. Histological analyses and functional knockout experiments demonstrated that one CDH2 paralog (CDH2_H) is essential for lamprey heart development and chamber formation, playing a critical role in the emergence of the closed circulatory system in early vertebrates. This high-quality lamprey genome provides a foundational resource for dissecting the genetic basis of key innovations that shaped vertebrate evolution.

genomics↗

bioRxiv2025

Transcriptome profiles of telocytes along the intestinal crypt-villus axis

Background & aimsIntestinal epithelial cells rely on a complex array of signals from the stromal microenvironment to determine their fate and function along the crypt-villus axis, but the precise cellular sources and combinations of signals regulating each position remain poorly defined. MethodsHere, we present a high-resolution atlas of a unique mesenchymal cell type previously identified as telocytes, mapped along the crypt-villus axis using single-cell RNA sequencing integrated with in-situ approaches and reporter mouse models. ResultsWe identify four spatially distinct telocyte subtypes: crypt, villus base, villus mid and villus tip, each with a unique gene expression profile. Crypt telocytes are enriched for canonical Wnt signaling components that support stem and progenitor cell maintenance, while villus base telocytes are a source of Bmps (bone morphogenic proteins) and express contractility-associated genes, suggesting a role in tissue architecture. Villus mid telocytes display signatures of immune regulation and inflammation, while villus tip telocytes are specialized for the regulation of ribonucleoprotein complexes and nutrient sensing. ConclusionsThis atlas provides a foundational resource for understanding how diverse telocyte populations coordinate signaling environments and shape intestinal homeostasis through both epithelial interactions and extensive crosstalk among themselves.

cell biology↗

bioRxiv2025

Cryo-FIB Lift-out and Electron Tomography Workflow for Bacteria-Nanopillar Interface Imaging Under Native Conditions: Investigating Dragonfly Inspired Bactericidal Titanium Surfaces

Dragonfly- and cicada-wing-inspired titanium nanostructured surfaces exhibit promising bactericidal properties. However, direct visualisation of the bacteria-material interface under hydrated conditions remains limited, restricting experimental interrogation how bacteria interact with nanostructured surfaces. This limitation reflects a long-standing methodological gap, as visualisation of bacteria-nanostructure interfaces has largely relied upon indirect or dehydrated imaging approaches. Cryo-electron tomography (cryo-ET) enables three-dimensional visualisation of cellular ultrastructure in a native hydrated environment, but imaging bacteria attached to metallic nanostructures by cryo-ET requires preparation of thin electron-transparent lamellae. Obtaining such lamellae from vitrified bacteria on titanium substrates is technically challenging because cryo-focused ion beam (cryo-FIB) milling must simultaneously section soft biological material and hard metal whilst bacteria embedded in vitreous ice are not directly visible. Here, we establish a correlative cryogenic workflow enabling targeted extraction and cryo-ET imaging of defined bacterium-nanopillar interfaces. Individual bacteria interacting with titanium nanopillars are identified by correlative cryo-fluorescence microscopy, followed by targeted cryo-FIB lift-out, transfer to a receiver grid, and thinning into electron-transparent lamellae. The data presented demonstrate that bacteria-nanostructure interfaces can be targeted, extracted, and structurally analysed in situ under fully hydrated conditions. This workflow provides a methodological framework for future cryo-ET studies of bacteria-nanotopography interactions.

microbiology↗