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Results for “cell biology”

Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2026

Patterned alginate hydrogel spatially guides collagen fibrillogenesis, viscoelasticity and endothelial cell invasion

Angiogenesis following injury has been shown to be driven by fibrillar proteins of the extracellular matrix (ECM), such as collagen. However, the use of protein-based biomaterials presents some challenges, such as uncontrolled degradation and limited tuneability. We demonstrate how to create patterned interpenetrating networks (IPNs) based on covalently crosslinked alginate and physically crosslinked collagen that provide suitable mechanical properties to support migration of endothelial cells (ECs) in a spatially controlled manner. Low molecular weight alginate is functionalized with norbornene (N) or tetrazine (T), which enables two independent covalent crosslinking methods: UV-mediated and degradable crosslinks with matrix metalloproteinase (MMP) sensitive peptides (Deg) and slower spontaneous N:T non-degradable crosslinks (noDeg). Using photolithography, patterns in degradation, collagen fibrillogenesis, microarchitecture and matrix viscoelasticity are created. The potential of such 3D patterned alginate-collagen (Alg-Col) IPNs to spatially guide EC invasion and proliferation was tested in a microfluidics platform resembling an early healing setting. Only regions combining collagen fibrillogenesis, alginate degradability and viscoelasticity demonstrated EC cell invasion similar to the ones found in vivo following injury. The 3D patterned Alg-Col IPNs are compatible with microfluidics, offer an strategy to widen the applications of protein-based hydrogels and present a versatile platform for tissue engineering and disease modeling.

bioengineering↗

bioRxiv2026

Aberrant neuronal cell cycle re-entry induces late-onset Alzheimer's disease relevant neuropathological and gene expression changes

Aberrant neuronal cell cycle re-entry (NCCR) is an alternative pathogenic mechanism in Alzheimer disease (AD) that has gained substantial support in the literature. The pathogenic role of ectopic NCCR is supported by our past work demonstrating that SV40T-mediated NCCR in adult mice can induce numerous pathologies associated with AD. Since NCCR is chronically induced for an extended period in the mouse model which gives rise to numerous pathologies including neuroinflammation, many of these neuropathological changes could simultaneously participate in driving disease progression. We hypothesized that the NCCR is a primary pathogenic driver and that halting this disease process at a later age could be sufficient for preventing the progression of AD-related pathologies. Here we show that modulation of NCCR at a later age prevents the progression of AD pathologies, including Abeta; and tau pathologies. Furthermore, functional genomics analysis demonstrates the late-onset AD (LOAD)-relevance of NCCR. Our findings suggest that our NCCR mouse model could help identify novel therapeutic targets that could aid in preventing AD progression.

neuroscience↗

bioRxiv2026

Operando Failure Diagnosis and Performance Dynamics in Microbial Fuel Cells Treating Mine Waste

Bench-scale microbial fuel cells (MFCs) treating mining wastewater frequently exhibit operational variability and uncharacterized degradation that obscure true biocatalytic performance. To decouple genuine biological treatment effects from mechanical failures, this paper presents an integrated diagnostic framework validated on two bench-scale systems treating heavy-metal-rich gold mine tailings. The first system evaluates Micractinium inermum algal bio-augmentation (System 1), while the second compares Psychrobacter alimentarius- and Trichococcus patagoniensis-dominated anodic consortia (System 2). To overcome single-reactor constraints, the framework integrates paired time-series statistical modeling, an adaptive percentile-floor change-point detector, equivalent-circuit modeling, and baseline-corrected spectroscopy (XRD/FTIR). Applying the framework to these systems uncovers previously masked dynamics: statistical analysis demonstrates that algal biocatalysis provides no voltage advantage under stable operation (+0.17%) but increases output by 27.54% under diurnal perturbation, while periodicity analysis links these diurnal shifts to the chamber photoperiod. Furthermore, heavy-metal remediation (up to 97.7%) is governed by system-level physicochemical mechanisms rather than algal-specific processes. The change-point detector successfully isolates distinct failure modes, distinguishing a recoverable excursion from terminal structural collapse. Finally, equivalent-circuit modeling reveals that the superior power density of Trichococcus consortia is driven by combined improvements in internal resistance and open-circuit voltage. Ultimately, pairing statistical controls with automated fault detection resolves operational ambiguity, offering a scalable baseline for health monitoring in bio-electrochemical wastewater treatment.

bioengineering↗

bioRxiv2026

THE ROLE OF LIQUID CRYSTAL ORDERING IN THE STRUCTURAL ORGANIZATION OF DNA IN BACTERIA.

This paper presents and critically reviews the results of original and some literature based experimental studies conducted by the authors last years on the structural organization of DNA in dormant (starvation stress), anabiotic dormant (4 HR treatment) E. coli cells, as well as the K12 {Delta}dps strain, which lacks the Dps protein (Dps null E. coli). The experimental data includes small-angle synchrotron radiation diffraction (SAXS) and transmission electron microscopy (TEM) data. Synchrotron radiation diffraction experiments on K12{Delta}dps cells allowed us to conclude that peaks at 44.3, 22.1, and 14.8 angstrom resolutions are associated exclusively with ordered DNA organization. Peaks at 44.3, 22.1, and 14.8 angstrom resolutions are also observed for samples of dormant (starvation stress) cells and anabiotically dormant cells. Therefore, this ordered DNA organization also applies to samples of dormant and anabiotically dormant cells. A model is proposed that considers the ordered DNA organization in the cell as a cholesteric liquid crystal. The powder diffraction pattern calculated based on this model is compared with experimental small angle X ray scattering (SAXS) data obtained on Dps-null cell samples. The model completely reproduces the key features of the experimental diffraction pattern from Dps-null cell samples. Accordingly, the cholesteric liquid crystal model corresponds to DNA packaging in dormant and anabiotically dormant cells. Cholesteric liquid crystal ordering should be further considered in all models of cellular DNA packaging. To address the question of which structural organization of DNA predominates in the cell: the cholesteric liquid crystal or nanocrystalline or whether they coexist and fully manifest themselves under different external conditions, it is necessary to utilize the latest methodological advances in structural analysis.

biophysics↗

bioRxiv2026

HIF1A recruits primate-specific endogenous retroviruses into the human hypoxic and immune responses

Oxygen availability varies profoundly across the human body and changes further during inflammation, infection, tissue injury and disease. Immune cells must therefore continuously adapt their transcriptional and metabolic state based on the oxygen availability to them. Hypoxia-inducible factor 1 (HIF1A) is central to this adaptation and a marker of the cellular response to low oxygen, yet its genomic targets have been assembled from a non-repetitive fraction of the genome, leaving nearly half of the human genome largely unexplored. Here we define the gene and transposable-element (TE) landscape of the human hypoxic response across different human tissues, cell lines, and conditions. This directional TE response was reproduced in transformed cells and in primary immune cells isolated from blood and the physiologically oxygen-restricted tonsil. Single-cell profiling of peripheral blood mononuclear cells (PBMC) under hypoxia, pharmacological HIF stabilization, and interferon stimulation revealed a striking difference between the gene and retrotranscriptome responses. While gene responses were strongly cell-type dependent and in a bidirectional manner, TEs were overwhelmingly activated. This pattern extended to blood and tonsil immune cells, where ~70-90% of tested TE families were induced under hypoxia, with activated tonsil cells showing exclusively induced significant families, including THE1B, alongside increased LTR7 and HERVH. Integrating HIF1A ChIP-seq with transcriptional responses revealed that HIF1A does not engage repetitive DNA indiscriminately. Instead, its binding converged on LTR7, the promoter long terminal repeat of the HERVH endogenous retrovirus. Approximately 80% of HIF1A-bound LTR7 elements contained a canonical hypoxia-response element, and disruption of HIF1A DNA binding dramatically reduced the expression of occupied HERVH loci. CRISPR deletion of individual LTR7/HERVH loci altered the expression of distant and neighboring genes, demonstrating that hypoxia-responsive retroelements can participate directly in host gene regulation and contribute to overall physiology. Our findings reveal the repetitive genome as a previously underappreciated component of oxygen sensing. We propose that HIF1A recruits selected endogenous retroviral elements into the human hypoxic response, extending oxygen-dependent regulation beyond conventional gene promoters and providing an additional regulatory layer through which tissue oxygenation can shape immune-cell state and human physiology.

genomics↗

bioRxiv2026

Compression Sequencing enables ultra-sensitive and scalable scRNA-seq

Current sequencing methods are inefficient and bottlenecked by repeated sampling of highly abundant molecules, which dominate sequencing reads, limit assay throughput and sensitivity for rare targets. For example, single-cell RNA sequencing (scRNA-seq) can profile up to millions of cells, but remains severely constrained by sequencing cost, resulting in shallow gene coverage and high dropout rate. Here we report an information science-inspired method, Compression Sequencing, that tackles this fundamental inefficiency and enables highly improved (>100x) sequencing power. Our method works by performing an accurate and unbiased logarithmic transform on molecular abundances over a wide (5 logs) dynamic range, thus suppressing high-abundance targets and enriching rare ones, while maintaining quantitative accuracy. Applied to scRNA-seq libraries, our method allows ultra-sensitive detection of low-abundance transcripts (2-5x more UMIs), ultra-low sequencing cost (200x reduction), preserves accurate cell types and differential expression analysis over a 500-2,000 gene panel. In AML clinical samples, Compression Sequencing reproduces clinical diagnosis and additionally allows transcriptomic profiling at affordable cost (est. $10 per sample). Our approach thus enables ultra-sensitive and scalable single-cell analysis for large-scale functional genomics studies, drug discovery screens, AI cell model training, as well as affordable single-cell disease diagnostics.

bioengineering↗

bioRxiv2026

Spatial Transcriptomics Reveals Compartment-Specific Immune Activation Signatures in Ileal and Lymph Node Tissue in Treated HIV Infection

People with HIV (PWH) on long-term antiretroviral therapy (ART) continue to experience elevated rates of morbidities and mortality driven by persistent immune activation despite viral suppression. Known contributors include low-level HIV provirus activity, microbial translocation in part from epithelial barrier dysfunction, microbiome dysfunction, and co-infections. However, how these interact and where they predominate across tissue compartments remains incompletely defined. Here, we applied spatial transcriptomics to characterize compartment-specific transcriptional programs in ileum (epithelium, Peyer's patches, lamina propria) and inguinal lymph nodes (B Cell follicles and T cell zone) from ten PWH on long-term ART, stratified by CD4/CD8 ratio into low-ratio and high-ratio groups, with low-ratio as a proxy for immune activation and increased risk for non-AIDS related serious event. Comparison of global expression found significant differences between groups in four of five compartments. Differential expression analysis identified 483 differentially expressed genes across four of five compartments, with the greatest burden in the T-cell zone and none in the lamina propria. Gene set enrichment analysis identified 116 enriched pathways predominantly in the low-ratio group, spanning immune activation, infection-associated, and metabolic programs, with Peyer's patches showing the broadest transcriptional divergence of any compartment. Cross-compartment signals included higher expression of ORMDL3 and ARL17B in the low-ratio group implicating mitochondrial stress and inflammasome activation, lower expression of CCL3L3 and FCMR in the low-ratio group suggesting impaired immune execution, and divergent ribosomal protein programs between B-cell follicles and the T-cell zone. Cell deconvolution identified compartment-specific differences in estimated immune cell proportions, and T-cell zone gene expression showed significant associations with HIV reservoir measures and plasma markers of microbial translocation and immune activation. Together these findings support spatially heterogeneous immune activation as a feature of persistent immune dysregulation in treated HIV infection and provide compartment-resolved, hypothesis-generating evidence for the tissue-specific mechanisms driving inflammation in this population.

bioinformatics↗

bioRxiv2026

An alignment-last approach enables rapid transcriptomic biomarker discovery in large cohorts

Canonical transcriptomic analysis requires committing from the outset to a reference genome or transcriptome, which imposes a predefined feature set, usually annotated genes or isoforms. Alignment and annotation dilute the signal through feature-level aggregation, discard any sequence absent from the reference, and require reprocessing the entire dataset for each new question (mutations, fusions, transposable elements). Here, we introduce the alignment-last paradigm, in which the read becomes the unit of comparison across samples, and alignment is deferred to annotate only the relevant sequences. Querying the merome, a reference-free cohort k-mer index, with just a handful of reads (about 0.01% of a sample's) reveals the cohort's transcriptomic structure in bulk and single-cell data. At single-cell resolution, these reads outperform genes for cell classification and rediscover, without supervision, a transposable-element signature (VL30) of exhausted T cells. Finally, unsupervised read-level differential analysis recovers established lncRNA biomarkers; uncovers new prognostic transposable-element reads in adrenocortical carcinoma and sarcomas; and extracts signals even from reads that fail to align.

bioinformatics↗

bioRxiv2026

De novo design of CR2 binder as vaccine scaffold

Efficient B cell activation during vaccine-induced humoral immunity relies on both B cell receptor (BCR) antigen recognition and synergistic signaling from co-receptors. Complement receptor 2 (CR2), the primary BCR co-receptor on B cells, lowers the activation threshold and amplifies downstream kinase signaling by orders of magnitude when engaged by complement fragment C3d decorated antigens. Targeting CR2 therefore represents a rational vaccine enhancement strategy, yet native C3d suffers from low affinity, poor stability, and manufacturing challenges. Here, we report the de novo design of a highly stable, high-affinity CR2 binder using deep learning driving protein design methods. Biophysical characterization, high-resolution cryoEM structural determination, and functional assays in vitro and in vivo confirm that the designed binder matches computational design models and specifically engages CR2 to boost B cell activation. When fused to antigen as a vaccine scaffold, the trimeric CR2 binder elicits robust humoral immune responses comparable to nanoparticle vaccines, while retaining the simplicity of single-chain protein production. Our work establishes a modular CR2 targeting vaccine scaffold platform with broad translational potential for next-generation protein vaccines.

immunology↗

bioRxiv2026

Bacterial Peptidoglycan Extends Lifespan by Activating Lysosomal Activity through V-ATPase Binding

Lysosomal dysfunction is a hallmark of aging, yet whether microbial components actively regulate this organelle to influence longevity remains unknown. Here, we identify bacterial peptidoglycan (PGN), a major cell wall component degraded by host lysozyme, as an evolutionarily conserved activator of lysosomal function that extends lifespan in both C. elegans and mice. We show that aging leads to an intestinal decline in lysozyme expression, which impairs bacterial cell-wall digestion and results in systemic PGN deficiency. Late-life PGN supplementation (starting at 18 months of age) significantly prolongs mouse lifespan and improves healthspan. Mechanistically, PGN localizes to lysosomes and directly binds V-ATPase subunits, enhancing ATP hydrolysis activity and promoting lysosomal acidification. This effect is abolished by V-ATPase inhibition (bafilomycin A1) or genetic disruption of lysosomal components (cup-5 and vha-12 mutants), confirming that functional V-ATPase is strictly required for lysosomal function and the longevity benefit. Importantly, PGN restores lysosomal acidification in aged cells, alleviates cellular senescence markers, and improves multiple hallmarks of aging including locomotion and muscle integrity. Collectively, these findings reveal an evolutionarily conserved mechanism whereby hosts exploit bacterial cell wall components to maintain cellular homeostasis, establishing a gut microbiome-lysosome-longevity axis with implications for microbiome-based anti-aging interventions.

physiology↗

bioRxiv2026

Entorhinal grid coding as a functional link between tau accumulation and episodic memory in human aging

Episodic memory decline is a common feature of cognitively normal aging, but its extent varies markedly across individuals. Although entorhinal tau pathology is thought to be a key contributor to episodic memory impairment, the neural mechanisms linking early tau accumulation to memory differences remain unclear. Grid-cell computations in the entorhinal cortex, which provide scaffolds for organizing experiences into episodic memories, offer one candidate mechanism. Here, we combined virtual-reality functional MRI, multivariate analysis, tau PET, and delayed word-list recall in cognitively normal older adults to test whether tau-related alterations in entorhinal coding are associated with worse episodic memory. Weaker left entorhinal grid-cell-like signal was associated with poorer memory performance, and individuals with higher left entorhinal tau burden showed weaker grid-cell-like signal. This association was specific to the canonical six-fold signal and was not explained by entorhinal volume, mean diffusivity, or intracortical myelination. A cross-sectional Bayesian mediation analysis further demonstrated that bilateral medial temporal tau burden is related to memory indirectly through left entorhinal grid-cell-like signal. Together, these findings provide evidence that entorhinal grid codes may constitute a functional pathway linking tau accumulation to memory variability in normal aging.

neuroscience↗

bioRxiv2026

A mouse-adapted Staphylococcus aureus strain enables lifelong neonatal colonization and elicits a Th17-dominated immune response

The opportunistic pathogen Staphylococcus aureus persistently colonizes the anterior nares of up to 20% of the human population, yet there were no persistent mouse colonization models to study host-pathogen interaction. Using the mouse-adapted S. aureus strain JSNZ (CC88-MSSA), we established a neonatal S. aureus colonization model in C57BL/6N mice. Natural neonatal colonization was achieved by vertical transmission in a JSNZ-positive breeding colony. Offspring were followed for up to 69 weeks and found persistently colonized in the nose and cecum with high bacterial loads. Adult mice were colonized by intranasal inoculation of JSNZ; controls received PBS. The colonization patterns and the S. aureus-specific T cell responses were then monitored over a period of 28 days and compared between age-matched mice colonized as neonates or adults. The neonatal group remained persistently colonized in nose and gut with high bacterial densities. In contrast, mice colonized as adults had lower and declining bacterial loads in the nose. Some eliminated S. aureus from the nares, while all remained colonized in the gut. Neonatally colonized mice exhibited reduced nasal chemokine levels, which may have favored the prolonged S. aureus persistence. Ex vivo re-stimulation of cervical lymph node cells with an S. aureus antigen cocktail revealed a Th17-dominated antigen-specific T cell response in both colonized groups. The lymph node cells secreted large amounts of IL-17, but Th1-, Th2-associated and regulatory cytokines were also detected. The cytokine patterns were similar in both colonized groups except for IL-5, which was more abundant upon neonatal colonization. In conclusion, vertical transmission of the mouse-adapted S. aureus strain JSNZ reliably establishes persistent high-density neonatal colonization, providing a physiologically relevant model for the study of S. aureus host interactions. Route and timing of colonization do not fundamentally affect the T cell response to S. aureus.

immunology↗

bioRxiv2026

Profiling and modulating astrocyte borders at injected biomaterials in mice

Astrocyte border formation is a conserved neuroprotective response to neural tissue disruption, yet astrocyte border states at implanted biomaterials remain less well characterized than injury responses. Here, we developed the Astrocyte Border Characterization (ABC) Tool, which leverages a shear-thinning, injectable biomaterial to locally deliver astrocyte-specific RiboTag AAVs and small molecule regulators in the mouse striatum, enabling molecular profiling and phenotypic modulation of astrocyte border (AB) cells. Spatially precise delivery of AAV using the ABC Tool yielded enhanced specificity and robust RiboTag expression in AB cells from 7-70 days post injection. Temporal transcriptomic profiling of AB cells revealed predominantly acute, transient changes in genes governing dedifferentiation, proliferation, metabolic reprogramming, and inflammation regulation. Persistent changes accounted for only 14% of regulated genes but involved critical gain of functions in immune regulation and host defense that mirrored astrocyte border responses at chronic CNS injuries. Local delivery of indiscriminate or astrocyte-selective ablation molecules delayed, rather than prevented, border formation, ultimately yielding thicker astrocytes borders with increased inflammation and fibrosis at the biomaterial-tissue interface. Conversely, local delivery of {beta}-hydroxybutyrate (BHB) from the ABC Tool altered key aspects of the transcriptional reprogramming to attenuate chronic astrocyte reactivity and prevent biomaterial contraction without exacerbating inflammation or fibrosis. Our findings establish the ABC Tool as a bioassay for studying and manipulating astrocyte borders at implanted biomaterials and identify focal metabolic regulation as a strategy to modulate AB cell phenotypes and enhance the CNS biocompatibility of biomaterials.

neuroscience↗

bioRxiv2026

Sphingolipid metabolism-related genes as key regulatory hubs in white smoke inhalation induced lung injury

Objective White smoke inhalation injury (WSI) causes severe acute lung damage with no specific therapy currently available. Sphingolipid metabolism is implicated in pulmonary inflammation, but its transcriptional regulatory landscape in WSI remains unexplored. This study aimed to identify key sphingolipid metabolism related genes and evaluate their regulatory roles and therapeutic potential in WSI. Methods We established a rat model of WSI and performed integrated bulk RNA sequencing, weighted gene coexpression network analysis (WGCNA), and single-cell RNA sequencing (scRNAseq) to screen for differentially expressed sphingolipid metabolism-related genes (DESRGs). Protein-protein interaction (PPI) network with four centrality algorithms was used to prioritize hub genes. In silico gene knockout and molecular docking were conducted to assess regulatory functions and identify potential drug candidates. Results We identified 22 DESRGs that were predominantly enriched in DNA replication and cell cycle pathways rather than canonical sphingolipid metabolic processes. PPI consensus prioritized three hub genes--Top2a, Ttk, and Ccna2--with Top2a exhibiting the highest expression in epithelial cells and significant downregulation after smoke exposure. ScRNAseq revealed immune cell infiltration and epithelial differentiation trajectories. Virtual knockout showed that Top2a depletion affected the largest transcriptomic fraction (~0.4%) and was enriched in lysosome biogenesis, innate immunity, phagocytosis, and lipid catabolism. Molecular docking identified thalidomide as a high affinity ligand for Top2a (Vina score: -8.5 kcal/mol). Conclusion Our multiomics integrative framework identifies Top2a as a central regulatory hub linking sphingolipid associated inflammation to epithelial responses in WSI, and nominates thalidomide as a potential drug repurposing candidate. These findings provide prioritized targets for future translational investigation.

bioinformatics↗

bioRxiv2026

X-ray crystallographic fragment screening reveals novel and conformationally dynamic ligand-binding sites in Mycobacterium tuberculosis FtsZ

Tuberculosis is a leading cause of death globally due to an infectious agent. There is ongoing need for novel mechanisms to inhibit M. tuberculosis (Mtb) growth and infection to improve patient outcomes. FtsZ, a GTPase that assembles into protofilaments at the division site of a replicating cell to produce two individual cells, is an attractive target as an essential protein in bacterial cell division. Here we describe a crystallographic fragment screening campaign of MtbFtsZ. 1,070 crystals were soaked with fragments and 714 datasets were used for downstream PanDDA analysis. 149 datasets exhibited PanDDA-generated event map density to support modeling of fragment binding. 15 novel sites are described. Both the ON and the OFF conformations of FtsZ are found in the asymmetric unit. Asymmetric binding of fragments to each chain in the model is observed. These crystallographic fragment screening results additionally provide opportunities for fragment growing and merging to develop FtsZ binders into drug-like molecules or conformation specific chemical probes.

biophysics↗

bioRxiv2026

From Bile Acids to a Gas-Producing Microbiome Phenotype: A Novel Mechanism of Host-Microbiome Communication

Background Microbiome-derived metabolites regulate host physiology, yet bacterial gaseous metabolites remain largely overlooked. Traditionally regarded as fermentation end-products, bacterial gases may act as biologically active mediators of host-microbiome communication. We hypothesized that bile acids regulate bacterial gaseous metabolism and influence host epithelial responses. Methods A high gas-producing clinical Escherichia coli isolate from a patient with moderately severe acute pancreatitis was cultured with selected primary and secondary bile acids. Gas production was assessed by pressure measurements, GC-TCD and GC-MS. Biological activity was evaluated by indirect exposure of Caco-2 and PANC-1 epithelial cells, followed by apoptosis/necrosis assays and whole-transcriptome RNA sequencing. Results Bile acids markedly reshaped bacterial gaseous metabolism. Cholic acid and deoxycholic acid promoted intense gas production, whereas chenodeoxycholic acid almost completely abolished it. Despite minimal apoptosis and necrosis, bacterial gaseous metabolites induced extensive transcriptional remodeling. Caco-2 cells showed stronger responses than PANC-1 cells, particularly to deoxycholic acid-derived gases, involving inflammatory signaling, extracellular matrix remodeling, epithelial plasticity, stress responses, and cancer-associated genes including PTGS2, MMP1, PLAUR, NR4A2, and SERPINE1. PANC-1 cells exhibited a more restricted response involving oxidative stress, proteostasis, and autophagy-associated pathways. Conclusions Our findings indicate that bacterial gases are a previously underrecognized class of microbiome-derived signaling molecules capable of modulating host gene expression independently of direct bacterial contact. We identify a gas-producing microbiome phenotype regulated by bile acid composition, linking microbial metabolism with epithelial signaling. These findings expand the concept of host-microbiome communication and provide a framework for investigating bacterial gaseous metabolites in intestinal and pancreatic diseases.

microbiology↗

bioRxiv2026

Integrated Transcriptomic and CRISPR Dependency Analysis Prioritizes a CDK1-AURKB Mitotic Vulnerability Axis in Diffuse Intrinsic Pontine Glioma

Diffuse intrinsic pontine glioma (DIPG), now classified within diffuse midline glioma, H3K27-altered, remains a lethal pediatric brainstem tumor with limited therapeutic options. Here, we integrated public DIPG transcriptomic datasets, protein-protein interaction modeling, functional enrichment, immune deconvolution, survival analysis, and DepMap CRISPR dependency data to nominate candidate mitotic vulnerabilities. Differential expression analysis comparing 27 DIPG tumors with 6 brainstem low-grade glioma comparator samples identified a proliferative transcriptional program enriched for chromosome segregation, nuclear division, and cell-cycle pathways. Network analysis prioritized a compact mitotic hub module containing CDK1, AURKB, TOP2A, CDC20, CDCA8, and related G2/M regulators. CIBERSORT analysis of an independent DIPG cohort inferred low cytotoxic T-cell signal, consistent with an immune-cold phenotype, although immune-cell fractions require orthogonal validation. Survival analysis showed that neither inferred immune scores nor a composite mitotic hub score significantly stratified overall survival. DepMap CRISPR gene-effect data nominated CDK1, AURKB, TOP2A, and BIRC5 as candidate dependencies across brain tumor models. These findings provide a computational framework for prioritizing mitotic vulnerabilities in DIPG and support experimental validation in disease-relevant models.

bioinformatics↗

bioRxiv2026

Lipid-ASO therapeutics exhibit differential tissue targeted delivery upon systemic or local CNS administration

Antisense oligonucleotides (ASOs) are a powerful therapeutic modality, but their full potential is hindered by pharmacokinetic properties that affect tissue and cellular delivery. Lipid conjugation is increasingly used to modulate ASO's biodistribution and promote extrahepatic activity, yet lipid dependent effects on in vivo functional delivery, particularly in the central nervous system (CNS), remain less explored. Here, we performed a side by side in vivo comparison of cholesterol, palmitic acid (C16:0), docosanoic acid (C22:0), and eicosapentaenoic acid (C20:5) conjugated to a fully phosphorothioated 3 10 3 LNA gapmer ASO targeting the Malat1 long non coding RNA. Lipid-ASO conjugates were administered systemically or locally in the brain of mice and evaluated for tissue level and cellular level distribution by imaging, qPCR and single-cell RNA sequencing, simultaneously annotating cell origin and global transcriptional changes within the cell. Following systemic administration in mice, lipid conjugation improved overall multi organ efficacy compared to unconjugated ASO, but with pronounced tissue specific differences. Single cell sequencing of liver and heart transcriptomes revealed lipid dependent cellular uptake patterns and transcriptional responses distinct from administration of unconjugated ASO. After intracerebroventricular administration, selected fatty acid conjugates enhanced silencing in deep brain regions such as the striatum, whereas cholesterol conjugation impaired functional delivery despite increased CNS retention. Light-sheet microscopy showed restricted parenchymal penetration of cholesterol ASOs compared with broader but heterogeneous distribution of palmitic acid conjugate. Together, these findings demonstrate that lipid identity critically determines ASO efficacy, productive cellular uptake, and regional CNS engagement, emphasizing the need for context specific lipid design in ASO therapeutic development.

pharmacology and toxicology↗