Biology subjects

Williams, J.

Publications and source records attributed to Williams, J..

At least 19 recordsLinked to original sources

bioRxiv2026

Independent evolution of harmala alkaloids in medicinal Syrian rue and ayahuasca plants

Plants from distantly related lineages can produce identical neuroactive molecules, many of which have been independently adopted across human societies for their medicinal and cognition-altering effects. This phenomenon is exemplified by the traditional use of Syrian rue (Peganum harmala) and the ayahuasca vine (Banisteriopsis caapi) for medicine and ritual. Despite diverging over 100 million years ago and inhabiting different continents and climates, both plants synthesize harmala alkaloids (HAs), which inhibit human monoamine oxidase and exhibit diverse neuroactive effects. Here, we combine gene expression analyses, genomics, and enzyme characterization to elucidate HA biosynthesis for both Syrian rue and ayahuasca vine. We reveal multiple layers of biochemical and genetic convergence in these pathways, including independent duplication and neofunctionalization of an ancestral enzyme from serotonin biosynthesis, and lineage-specific repurposing of primary metabolic genes to generate a reactive precursor for alkaloid scaffold formation. Our study resolves biosynthetic routes to HAs in two medicinal plants and demonstrates convergent evolution of specialized metabolism at the levels of enzymes, pathways, and genomes.

biochemistry↗

bioRxiv2026

Sex-Dependent Microbiome-Metabolome Remodeling Associated with Bacteroides acidifaciens Modulation of Acute Respiratory Distress Syndrome

Acute respiratory distress syndrome (ARDS) is a severe lung disorder influenced by the interplay between the intestinal microbiome, host metabolism, and immune function. We previously found that Bacteroides acidifaciens (BA) has opposing effects on staphylococcal enterotoxin B (SEB)-induced ARDS, worsening the condition in female mice while alleviating it in males. This study aimed to examine whether these outcomes are linked to sex-dependent changes in the intestinal microbiome and gut-lung metabolome. C3H/HeJ mice of both sexes were orally colonized with Bacteroides acidifaciens before SEB exposure. Stool and bronchoalveolar lavage fluid (BALF) were collected for analysis using 16S rRNA sequencing and untargeted liquid chromatography mass spectrometry metabolomics. Our findings revealed distinct sex-dependent metabolic and microbial responses. In females, disease exacerbation correlated with elevated lipid mediators (leukotriene B4, 13-oxoODE, and 13S-HODE) alongside shifts in bile acid and redox metabolism. The intestinal microbiome showed changes in Bacteroides, Akkermansia, and Lachnospiraceae. Conversely, males showed a protective phenotype characterized by increased butyrate and alterations in sphingolipid, amino acid, redox, and nucleotide pathways. Integrated analyses identified sex-associated microbial communities and metabolomic profiles, highlighting distinct patterns of lipid and redox associations in females versus sphingolipid and short-chain fatty acid patterns in males. These results suggest that host sex influences how a commensal bacterium affects inflammatory lung injury and point to pathways for future research.

microbiology↗

bioRxiv2026

Folic acid polyglutamylation triggers MAP2K5 signaling to induce adipocyte lipid catabolism

Adipocyte signaling adaptively responds to macronutrients, but how essential micronutrients impact lipid homeostasis remains poorly understood. Here, we demonstrate that folate polyglutamylation, the sequential conjugation of glutamate residues to folate, serves as a dynamic biochemical process regulating metabolic signaling. Using our folate metabolomics platform, we show that polyglutamylated folic acid accumulates in healthy adipose tissue, but is depleted in obesity across mice and humans, independent of circulating folate levels. Genetic ablation of the polyglutamylation enzyme folylpolyglutamate synthase (Fpgs) in adipocytes suppresses lipid catabolism to induce cell-autonomous lipid accumulation, operating independently of canonical adipogenesis or downstream one-carbon flux. Target-engagement proteomics identifies monoglutamylated folic acid as a Map2k5-interacting molecule that inhibits Map2k5 activity, whereas Fpgs-mediated folic acid polyglutamylation acts as a chemical switch that disrupts this interaction to promote lipolysis. In vivo, whole-body inhibition of Map2k5 or adipose-targeted genetic depletion of Fpgs increases body fat mass in the absence of dietary obesogenic triggers. Furthermore, single-cell and single-nuclear transcriptomic analyses establish the Fpgs-Map2k5 pathway as a core transcriptional signature of mouse and human adipose tissues. These findings uncover a non-canonical signaling role for folate that regulates adipocyte lipid homeostasis.

biochemistry↗

bioRxiv2026

Dysregulated neutrophil extracellular trap formation is a novel driver of clonogenicity and therapeutic vulnerability in CMML

Chronic myelomonocytic leukemia (CMML) is an aggressive hematologic malignancy characterized by excess inflammatory signaling and clonal myeloproliferation. The relative contribution of neutrophils (PMNs) to the inflammatory milieu in CMML is poorly understood. In this study we sought to understand whether neutrophil extracellular trap (NET) formation, a key mediator of neutrophilic inflammation, is dysregulated in CMML and can be therapeutically targeted with a novel peptide inhibitor of NETosis called neonatal NET-inhibitory factor (nNIF). Here, we demonstrate that baseline NET formation is aberrantly increased in primary CMML PMNs transcriptionally primed for NETosis, and that soluble factors produced during CMML NET formation promote clonogenicity in CMML CD34+ hematopoietic cells matched to the same patient. Further, we show that nNIF and clinical agents under investigation in CMML effectively inhibit NETosis, warranting further study of NET inhibitory agents in this rare disease with limited treatment options.

cancer biology↗

bioRxiv2026

Hierarchical neural integration of musical structure during expert performance

Music, which is organized hierarchically (notes, phrases, sections), provides an ideal model for studying fine-grained motor sequence production. However, it is unknown how musicians brains integrate tonal structure over multiple timescales during real-life performance. Here, we scrambled an unfamiliar Tchaikovsky piano suite at four timescales--every 1/2/8 measures, or fully intact--and asked expert pianists to play (sightread) all four versions in the fMRI scanner. Responses in the motor network, default mode network, and hippocampus were strongly impacted by scrambling, indicating that they integrate tonal structure over relatively long timescales. Additionally, the emergence of functionally connected sub-networks between auditory, visual, motor, and default mode network regions across scramble levels supported this hierarchical integration process. Our results cannot be explained by lower-level cues (tempo, timbre, dynamics; local pitch height or rhythmic density) and instead reflect processing of high-level tonal structure. Our study highlights novel mechanisms of complex auditory-motor action planning during live music performance.

neuroscience↗

bioRxiv2026

Multi-site phenotypic characterisation of Kisumu colonies for comparison of insecticide susceptibility between testing centres

Testing of insecticides and insecticide-based vector control tools relies on the use of bioassays whereby insects are exposed and mortality, or another relevant entomological endpoint, is measured. To be able to compare results between products and across time and space standard laboratory strains are used. Insecticide-susceptible strains are often used as comparators in studies involving insecticide-resistant populations. Kisumu is a long-established, widely-used insecticide-susceptible strain of Anopheles gambiae. In this study, 16 facilities across different countries exposed their Kisumu colonies aged 3-5 days to four insecticides (permethrin, alpha-cypermethrin, DDT and pirimiphos-methyl) at a range of concentrations in WHO tube assays to assess susceptibility. Insecticide-treated filter papers were made at the Liverpool School of Tropical Medicine and sent to testing centres, along with a standard protocol and testing conditions requirements, to standardise testing as much as possible. A first observation was that DDT results were affected by the use of Risella oil, the WHO-recommended carrier oil but immiscible in acetone, leading to uneven coating of papers and increased variability in bioassay results (40x variance between colonies). Bioassay data generated by the testing facilities suggest a wide range of susceptibility to different insecticides between colonies of Kisumu: LC50 values between colonies differed, though not significantly, by up to 4x for permethrin, 6x for pirimiphos-methyl, and 15x for alpha-cypermethrin. Some colonies are classified as insecticide resistant based on the WHO definition of <98% mortality on exposure to a discriminating concentration. The assumption that Kisumu is a standard susceptible strain, consistent between facilities is challenged, which has implications when comparing results between studies. We propose genotypic analysis of the colonies to identify signs of resistance markers indicative of contamination from resistant strains or wild mosquitoes, and generation of guidance to support the maintenance and monitoring of susceptible mosquito strains.

zoology↗

bioRxiv2026

Environment and plant genetics shape barley rhizosphere microbiome structure across contrasting locations

AbstractSoil microorganisms are crucial for plant survival and productivity, but factors governing rhizosphere recruitment across diverse regions remain unclear. This study investigated the rhizosphere microbiome of barley, using elite cultivars across seven location-year trials to evaluate the effects of environmental factors and crop genotype on bacterial and fungal community composition. Three locations were in the US northern Great Plains, and Hawaii was used as a contrasting environment. A greenhouse reciprocal transplant study determined the relative contributions of soil physicochemical factors and soil inoculum to rhizosphere community structure. Using 16S and ITS2 amplicon sequencing, the study characterized bacterial and fungal microbiomes and assessed the contribution of environment, soil chemistry, and barley genetics to microbial community assembly. In locations within the adapted range of barley, Actinobacteriota was the dominant phylum, while Proteobacteria was dominant in Hawaii. Variance partitioning showed that 73% of bacterial and 80% of fungal genera were associated with location-year effects while 53% of bacterial and 36% of fungal genera were responsive to soil factors. Enrichment analysis found 21.6% of bacterial and 51.4% of fungal ASVs were unique to specific barley genetic subpopulations. Results from the reciprocal transplant study validated field observations by demonstrating that 20.7% of the variation in community structure was explained by soil while 18.2% was explained by inoculum source. These findings demonstrate that environmental variation is the dominant constraint on rhizosphere community composition but within these constraints, barley genotype drives recruitment of distinct bacterial and fungal taxa. ImportanceThese findings underscore the complex interplay between plant genotype, environment, and microbial community assembly, providing insights into how barley recruits distinct microbial communities in the rhizosphere across different environments. These insights have the potential to be leveraged for management and plant breeding strategies to optimize plant-microbe interactions for enhancing agricultural sustainability.

microbiology↗

bioRxiv2026

Spatial MALDI-MSI Reveals a Coordinated Vicious Cycle of Oxidative Membrane Damage and Ceramide-Driven Sphingolipid Dysregulation in the Chronically Neuroinflamed Brain

BackgroundChronic neuroinflammation is a major driver of cognitive decline, vascular cognitive impairment, and Alzheimers disease. However, the spatial lipidomic alterations underlying neuroinflammatory brain injury remain poorly defined. Oxidative stress and sphingolipid dysregulation have been implicated, but their regional distribution and interplay in the brain are not well characterized. MethodsWe performed positive-ion mode matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) on coronal brain sections from middle-aged spontaneously hypertensive rats (SHR), a model of chronic neuroinflammation, and normotensive Wistar-Kyoto (WKY) controls. Spatial distributions and relative abundances of multiple lipid classes, including phosphatidylcholines (PCs), sphingomyelins (SMs), hexosylceramides (HexCers), ceramides, phosphatidylserines (PSs), phosphatidylinositols (PIs), phosphatidylethanolamines (PEs), phosphatidic acids (PAs), and sulfatides, were mapped and compared between genotypes. Region-of-interest analysis was used to quantify changes across cortex, hippocampus, and white-matter tracts. ResultsSHR brains exhibited a coordinated lipidomic signature characterized by pronounced oxidative stress and membrane remodeling. Oxidized and short-chain PCs were markedly upregulated (up to 11.6-fold), while major structural diacyl PCs were broadly downregulated. Concurrently, sphingolipids were significantly altered, with robust upregulation of SM(d36:1) (7.5-fold) and multiple HexCer species (1.5-1.9-fold), accompanied by accumulation of ceramides. These changes were accompanied by heterogeneous redistribution of PS, PI, and PE species, particularly within the hippocampus. Sulfatide patterns in white-matter tracts were also altered, suggesting myelin remodeling. Region-of-interest analysis confirmed that the most pronounced lipid alterations were concentrated in the hippocampus and white-matter regions. ConclusionsChronic neuroinflammation induces a spatially organized, multi-class lipid remodeling response in the brain, driven by advanced oxidative membrane damage and a shift toward a pro-apoptotic sphingolipid profile. The convergence of these pathways creates a vicious cycle of membrane injury, mitochondrial dysfunction, and sustained neuroinflammation that is especially prominent in the hippocampus and white matter. These spatially resolved findings provide direct evidence that oxidative stress and sphingolipid dysregulation are central, interrelated mechanisms contributing to neurovascular injury and increased risk of cognitive impairment. The study highlights the power of MALDI-MSI to uncover region-specific lipid pathology and identifies potential lipid-based targets for therapeutic intervention in neuroinflammatory brain disease. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/738110v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@853af9org.highwire.dtl.DTLVardef@e2d079org.highwire.dtl.DTLVardef@e4e065org.highwire.dtl.DTLVardef@480d80_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2026

Gut microbiome derived folate metabolite suppresses colorectal cancer progression

The gut microbiota influences colorectal cancer (CRC) progression, primarily through the secretion of small molecule metabolites. While numerous microbial products are known to drive CRC, endogenous protective mechanisms remain largely uncharacterized. Utilizing a folate metabolomics platform, we demonstrate that the healthy gut microbiota produces folinic acid (FA), a known chemotherapeutic adjuvant also known as leucovorin. This microbially derived folinic acid is progressively depleted in mouse models of colitis-associated CRC and in human clinical metagenomic cohorts with advancing disease severity. Mechanistically, folinic acid acts as a signaling molecule that directly binds and inhibits the intracellular protease calpain-2. This interaction stabilizes epithelial E-cadherin protein expression and suppresses CRC epithelial-to-mesenchymal transition driving metastasis. Genetically manipulating gut microbial production of FA is sufficient to modulate CRC in vivo, even in the presence of chronic inflammation. This study reframes folinic acid from a chemotherapeutic enhancer to an endogenous microbial metabolite that actively suppresses CRC progression.

cancer biology↗

bioRxiv2026

Hepatic stearoyl-CoA desaturase deficiency ameliorates hyperglycemia through bile acid signaling in an insulin-independent manner

Hyperglycemia in Type 1 Diabetes (T1D) is managed almost exclusively via exogenous insulin therapy, an approach restricted by significant glycemic fluctuations, long-term side effects such as weight gain, and high economic burden. Identifying physiological pathways capable of clearing blood glucose independent of insulin is therefore of paramount clinical importance. Here, we demonstrate that liver-specific stearoyl-CoA desaturase-1 (SCD1) deficiency protects against diabetic hyperglycemia and hepatic steatosis in an insulin-independent manner. SCD1 ablation decreases cellular oleate availability, altering lipid flux and redirecting excess cholesterol into alternative biosynthetic pathways. This redirection drives a 2-fold elevation in hepatic bile acids and a striking 10-fold increase in plasma bile acids, predominantly characterized by the accumulation of taurocholic acid. This shifted bile acid pool stimulates the expression of glucose transporter 1 (Glut1) in the liver via activation of the nuclear hormone receptor FXR, facilitating basal glucose clearance in the absence of insulin. Genetic deletion models show that while the hepatokine FGF21 serves as a partial mediator of this phenotype, the local bile acid-FXR axis remains a sufficient driver of systemic glucose clearance. Finally, we show that dietary oleate supplementation completely reverses this protective phenotype, turning down Glut1 expression and restoring overt diabetes. Together, our findings uncover a novel bile acid-FXR-Glut1 signaling axis triggered by SCD1 inhibition, offering a framework for insulin-independent glycemic control. Article HighlightsHere, we demonstrate that liver-specific stearoyl-CoA desaturase-1 (Scd1) deficiency protects against diabetic hyperglycemia and hepatic steatosis in an insulin-independent manner. Mechanistically, Scd1 ablation redirects excess cholesterol into bile synthesis, predominantly characterized by an increase in liver and plasma taurocholic acid. These shifted bile acids stimulate hepatic glucose transporter 1 (Glut1) expression via the farnesoid X receptor (FXR) activation to facilitate basal glucose clearance. While the hepatokine FGF21 acts as a partial systemic mediator, the local bile acid-Fxr axis remains a sufficient driver of clearance, a protective phenotype completely reversed by dietary oleate supplementation.

biochemistry↗

bioRxiv2026

Conventional dendritic cells type I with an enhanced type-I-IFN signaling underpin anti-tumor immune responses in brain metastases

Brain metastases are associated with poor prognosis. A better understanding of anti-tumor immune responses in the context of immune specialized microenvironment of the brain is required to develop improved therapeutic strategies for this disease. We demonstrate that the conventional dendritic cells type 1 (cDC1) gene signature positively correlates with a prolonged BrM-dependent survival in melanoma and breast cancer patients. Furthermore, intracranial anti-tumor immune responses in preclinical BrM models consistently rely on cDC1s for tumor growth control, BrM-dependent survival and maintenance of the intra-tumoral CD8+ T cell pool, in contrast to variable, cancer type-dependent cDC1 roles in extracranial tumors. This is underpinned by tumor site-specific cDC1 molecular profiles with distinct Toll like receptor repertoires, upregulation of co-stimulatory molecules and IL-12, and enhanced type-I-IFN signaling in intracranial cDC1s, with the latter driving increased cDC1 activation. cDC1s also promote the conversion of progenitor exhausted CD8+ T cells to transient effectors, which is further enhanced by immune checkpoint blockade therapy. These findings pinpoint cDC1s as a major cell population of interest in the development of future immunotherapies for BrM. Significance statementCharacteristics of immune cells and tumor microenvironment in brain metastases (BrM) are distinct to extracranial tumor sites and characterized by special immunological barriers and suppressive stroma. Conventional dendritic cells type 1 (cDC1s), which are key regulators of anti-tumor immunity, remain largely unexplored in this context. This study deepens our understanding of how anti-tumor immune responses differ between BrM and extracranial tumors by revealing BrM-specific phenotype and function of cDC1s, with the latter being critical for anti-tumor immunity in BrM even in cancer models where extracranial cDC1s demonstrate pro-tumorigenic function, highlighting cDC1s as universal regulators of intracranial anti-tumor immunity. Our study suggests that tumor site-specific augmentation of cDC1s should be pursued to enhance the efficacy of immunotherapies in BrM.

cancer biology↗

bioRxiv2026

Age-Dependent Spatiotemporal Remodeling of Brain Sphingolipids During LPS-Induced Neuroinflammation: MALDI-MSI Reveals Accelerated Sphingomyelin Depletion and Sulfatide Accumulation Linked to Mitochondrial Oxidative Stress

Aging is a major risk factor for exacerbated neuroinflammation and neurodegenerative diseases, yet the underlying lipid metabolic mechanisms remain incompletely understood. Here, we employed high-resolution matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) combined with quantitative peak-area analysis and conceptual kinetic modeling to investigate age-dependent sphingolipid remodeling in the rat brain following intracerebro-ventricular (ICV) LPS challenge. In old rats, MALDI-MSI revealed pronounced and progressive sphingolipid dysregulation compared with young animals. Quantitative analysis showed a dramatic [~]10-fold reduction in SM(d36:1) and [~]4-fold reduction in SM(d42:2), accompanied by significant accumulation of long-chain sulfatides (2.12-fold increase in C24:1-sulfatide and 1.45-fold increase in C24(OH)-sulfatide) at both 24 h and 72 h post-LPS. Spatial imaging demonstrated that these changes were markedly amplified in white matter regions and became more widespread and intense at 72 h. A simplified Michaelis-Menten kinetic model successfully recapitulated the experimental data, identifying increased nSMase2 activity (higher Vmax) as the primary driver of accelerated sphingomyelin hydrolysis and subsequent ceramide rerouting into sulfatide synthesis. This metabolic shift generates excess ceramide that promotes Drp1-mediated mitochondrial fission, elevates mitochondrial ROS production, and disrupts bioenergetics, establishing a feed-forward loop linking sphingolipid remodeling to mitochondrial oxidative stress and white matter vulnerability in the aged brain. These findings provide the first spatially and temporally resolved demonstration of age-dependent sphingolipid metabolic reprogramming during neuroinflammation. By integrating multimodal MALDI-MSI, quantitative lipidomics, and kinetic modeling, this study reveals a previously underappreciated nSMase2-ceramide-mitochondrial axis in neuroinflammaging. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=179 SRC="FIGDIR/small/734865v1_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@be7a9eorg.highwire.dtl.DTLVardef@164499dorg.highwire.dtl.DTLVardef@11cea50org.highwire.dtl.DTLVardef@15b2ac2_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2026

Microbiome Composition Regulates Cathepsin Expression in Vaginal Fluid Across BV Diagnoses and Community State Types

PurposeBacterial vaginosis (BV) is associated with disruption of the vaginal microbiome and extracellular matrix (ECM) remodeling, yet the contribution of host proteases to this process remains unclear. This study investigated whether expression and activity of cathepsins K, L, S, and V differ by BV diagnosis and community state type (CST). We hypothesized that BV and BV associated CSTs would exhibit increased expression and activity of collagen and elastin-degrading cathepsins. MethodsVaginal fluid samples were collected and classified by BV diagnosis and CST. Cathepsin expression was evaluated by Western blotting to distinguish inactive and active enzyme forms. Proteolytic activity was assessed using multiplex cathepsin zymography. Statistical analyses compared cathepsin expression and activity across diagnoses and CSTs. Principal component analysis and linear regression were performed to assess associations between cathepsin activity, microbial diversity, and CST. ResultsProcathepsin K expression was significantly increased in BV-positive and CST IV samples, while total cathepsin L expression was significantly elevated in samples with Nugent-intermediate scores. Cathepsins S and V showed variation in inactive and active forms in Nugent-intermediate and CST III samples. In contrast, total cathepsin activity, including cathepsins K and V, did not significantly differ across BV diagnoses or CSTs. Overall, cathepsin activity varied between individuals rather than by clinical classification. ConclusionsCathepsin expression and maturation state differ by microbiome composition, suggesting that the vaginal microbiome may regulate post-translational processing of cathepsins. As a result, cathepsin activity appears to be regulated at the individual level rather than strictly by BV diagnosis or CST. These findings link vaginal microbiome composition to ECM remodeling and potential adverse reproductive outcomes.

bioengineering↗

bioRxiv2026

New Human IPSC Models of Late-onset Alzheimer's Disease Polygenic Risk Identify Multiple Impairments of Microglial Function

Genetic discoveries implicate microglia in late-onset Alzheimers disease (AD). We modelled AD in a powerful study of 51 human induced pluripotent stem cell (iPSC) microglia derived from high-polygenic risk AD or low-risk cognitively well individuals, sampled from a large cohort. We explored mitochondrial function, cytokine secretion, endocytosis, phagocytosis, lipid accumulation, calcium store release, and chemotaxis under basal conditions and immune challenge. High polygenic risk was independently associated with significant functional deficits in iPSC microglia under immune challenge, in mitochondrial ATP production (p=0.005, -13%), and inflammatory cytokine release (IL-6: p=0.018, -42%; TNF p=0.026, -38.5%). Furthermore, a selective deficit in amyloid-{beta} uptake was identified (p=0.00477, -5.9%). Deficits in inflammatory cytokine release were driven by APOE {varepsilon}4. These findings reflect primary changes in AD pathogenesis predating plaque formation and validate a human in vitro platform for late-onset AD to further understand disease mechanisms and screen drug or genetic therapies.

neuroscience↗

bioRxiv2026

Profiling and Targeting of Regulatory RNAs to Upregulate Gene Expression

Transcription of long noncoding RNAs (lncRNAs), including enhancer RNAs (eRNAs) and promoter-associated RNAs (paRNAs), collectively termed regulatory RNAs (regRNAs), is a hallmark of active gene expression, yet it remains unknown whether regRNAs can be targeted to selectively enhance transcription in cis. We developed regRNA Capture-seq, a high-throughput method to profile regRNAs, and applied it to primary human hepatocytes, annotating thousands of regRNAs at [~]2,000 enhancers and promoters. Using this approach, we interrogated a genetically validated enhancer of the ornithine transcarbamylase (OTC) gene, mutations of which cause OTC deficiency (OTCD), the most common urea cycle disorder. Antisense oligonucleotides (ASOs) targeting enhancer-derived regRNAs led to dose-dependent upregulation of OTC in hepatocytes. Mechanistically, ASOs altered regRNA structure, elevated regRNA levels, displaced transcriptional repressors, and increased H3K27 acetylation at the targeted enhancer. This work establishes a potential therapeutic strategy for addressing haploinsufficiency and highlights regRNAs as actionable targets for ASO-mediated upregulation of gene expression.

genomics↗

bioRxiv2026

Microbial metabolism of methotrexate produces a STAT3 signaling molecule that alleviates gut inflammation

Methotrexate (MTX) therapy in inflammatory bowel disease (IBD) is often limited by inter-individual variability in clinical response and adverse effects. Gut microbiota contribute to MTX therapeutic response and toxicity by metabolizing MTX and altering its bioavailability. However, how inflammation alters microbial MTX metabolism and how its metabolites influence the host remain poorly understood. Here, we identify Clostridium asparagiforme as a potent and efficient metabolizer of methotrexate, producing deoxyaminopteroic acid (DAMPA) in the distal gastrointestinal tract. We demonstrate that DAMPA preserves mitochondrial integrity by promoting mitophagy in intestinal epithelial cells through mitochondrial STAT3 signaling. DAMPA administration attenuates intestinal inflammation in vivo, and improves metabolic dysfunction associated with IBD. Together, these findings reveal an unappreciated role for a gut microbial MTX metabolite in mediating epithelial homeostasis during intestinal inflammation, thus reframing microbial MTX metabolism from passive drug detoxification to active regulation of host mitochondrial and inflammatory homeostasis.

microbiology↗

bioRxiv2025

Tracking the zombie ant graveyards' dynamics at Mulu National Park, Borneo, Malaysia

Ophiocordyceps unilateralis sensu lato is a parasitoid fungus that predominantly infects ants from the Camponotini tribe, inducing behavioural changes to enhance its survival and reproduction. The most notable change occurs during the final infection phase, where the infected ant leaves its nest to perform the death grip, hanging upside down and biting onto a substrate to anchor itself. In the tropics, these infected ants congregate at specific locations known as zombie graveyards, which have optimal conditions for spore dispersal. This study examines the spatial dynamics of known graveyards at a national park in Borneo, Malaysia, by assessing existing sites and identifying new ones. We located old graveyards at Night Walk and Paku Trail and discovered a new one at Botany Loop. The density of ant cadavers per quadrat increased slightly, from 1-3 ants/m2 to 2-4 ants/m2. While the graveyard size at Night Walk remained similar, Paku Trail experienced a nearly two-fold reduction, decreasing from 197 m2 to 97 m2. At Night Walk, three ant hosts were identified, with Colobopsis leonardi being the dominant ant host across all graveyards. Without genetic tests, we could not confirm whether the same O. unilateralis infects all ants in the graveyards. Our findings suggest a general site fidelity of zombie graveyards, with hotspot densities varying spatially, potentially influenced by microclimatic changes. Long-term monitoring of graveyard dynamics is essential for understanding natural infections and the effects of environmental conditions on parasitoid-host ecology.

ecology↗

bioRxiv2025

4-1BB expression and signaling regulates MAIT cell activation and effector functions

Mucosal associated invariant T (MAIT) cells are abundant unconventional T cells that recognize microbe-derived riboflavin metabolites presented by MR1. Upon recognition, they become activated, produce proinflammatory cytokines, chemokines and cytotoxic molecules. MAIT cells are also activated by cytokines independently of T cell receptor (TCR) engagement, however these two signals can also act in concert to finetune MAIT cell functions. Additionally, multiple other co-stimulatory signals have also been reported that can boost MAIT cell effector responses to TCR or cytokine stimulation. However, a comprehensive study exploring the role of surface bound TNF receptor superfamily (TNFRSF) molecule 4-1BB, well known for its co-stimulatory function during conventional T cell activation, is lacking in the context of MAIT cells. In this study, we show that 4-1BB is the earliest and most highly expressed TNFRSF co-stimulatory molecule on MAIT cells upon activation by Escherichia coli, with expression seen as early as six hours where it was predominantly MR1 mediated. 4-1BB expression on MAIT cells following late activation was due to both TCR signaling and cytokine signaling. We found marked differences in MAIT cell activation and cytokine expression between 4-1BB+ and 4-1BB- MAIT cells suggesting 4-1BB expression on MAIT cells is associated with functional superiority. By blocking 4-1BB signaling or co-culturing with a 4-1BBL overexpressing cell line we demonstrated an important role of co-stimulation via 4-1BB in MAIT cells during activation. Expression and signaling via 4-1BB also enhanced T-bet and Blimp1 expression. In summary, our study confirms a role for 4-1BB signaling during MAIT cell activation.

immunology↗