Biology subjects

Johnson, K.

Publications and source records attributed to Johnson, K..

At least 19 recordsLinked to original sources

bioRxiv2026

Gadusol in epidermal mucus protects larval zebrafish from UVB-induced DNA damage

Ultraviolet B (UVB) radiation causes DNA damage and poses a threat to organisms exposed to sunlight. Zebrafish embryos are protected from UVB by maternally deposited gadusol, a small-molecule sunscreen, but it has been unclear how protection is maintained as maternal gadusol is depleted during larval development. Here, we show that gadusol synthesis in the developing zebrafish skin is required for UVB defense throughout larval development. Maternal gadusol persists through early larval stages, but juvenile fish depend solely on zygotic gadusol for protection against DNA damage and death after UVB exposure. Melanin provides a more modest contribution to survival. Gadusol is present in the epidermal mucus layer at the body's outermost surface, positioning it to absorb UVB before it reaches underlying cells. Depletion of epidermal mucocytes increases UVB-induced DNA damage and activates a DNA damage response. Together, these findings identify a maternal-to-zygotic handoff in gadusol-based photoprotection and support a model in which gadusol in surface mucus intercepts UVB before it reaches living tissues.

developmental biology↗

bioRxiv2026

Loss of SUR1 and Kir6.2 ATP-sensitive potassium channel subunits produces comparable neuronal excitability and morphine antinociception

ATP-sensitive potassium (KATP) channels are integral membrane proteins that play a crucial role in linking cellular metabolism to neuronal excitability. KATP channels in peripheral nociception and pain emphasize metabolic gating as a determinant of dorsal root ganglia and spinal cord excitability, particularly in pathophysiological states. Activation of KATP channels reduces excitability and and also contributes to opioid induced analgesia. KATP channels are composed of multiple subunits, that are assembled in several subtypes, including Kir6.2/SUR1 which is found predominantly in neurons in the nervous system. It remains unclear whether both Kir6.2 and SUR1 subunits are required to regulate neuronal excitability under naive conditions and to mediate morphine-induced antinociception. These experiments aim to determine whether neuron-specific loss of either Kir6.2 or SUR1 increases dorsal root ganglion (DRG) neuronal excitability, and attenuates morphine antinociception in mice. We found that loss of SUR1 or Kir6.2 activity or expression both increased DRG excitability in whole cell patch clamp studies. Using DRG cultured from SUR1 and Kir6.2 floxed animals, we also found that loss of either subunit decreased potassium flux induced by diazoxide. Morphine induced antinociception was also attenuated after Cre-mediated recombination in SUR1 and Kir6.2 floxed animals. Chronic morphine administration also decreased Kir6.2 (gene: Kcnj11) RNA signal in mouse DRG, but not spinal cords. Together, these findings suggest that, despite evidence from single-cell sequencing and other datasets indicating potentially distinct expression patterns of SUR1 and Kir6.2, these subunits most likely function cooperatively to form functional KATP channels under normal physiological conditions.

neuroscience↗

bioRxiv2026

Standardized benthic imaging with small, modular ROVs: an open-source framework developed in temperate kelp forests

(1) Kelp forests support ecologically, commercially, and culturally important species, yet their benthic communities remain difficult to monitor consistently across space and time. Scientific SCUBA surveys provide valuable observations but can be constrained by safety, bottom time, depth, and the spatial extent that divers can efficiently cover, motivating complementary approaches for collecting standardized benthic imagery. (2) We developed and field-tested a standardized, open-source framework for benthic visual-imaging surveys using a small, modular remotely operated vehicle (ROV). Across four cost-scalable hardware configurations, we integrated navigation sensors, custom BlueOS Extensions, and our surftrak contribution to ArduSub, which enables fixed-altitude flight above the seafloor, and developed two open-source desktop applications that streamline field operations (rov_flight_ops) and imagery processing (rov_imagery_processing). We also evaluated power supplies, benthic lighting, camera settings, field protocols, and telemetry-processing workflows over 303 transects conducted on 64 survey days from 2022 to 2026. (3) The ROV operated across soft-sediment and rocky habitats, currents of 0-3 kt, and underwater visibility of 0-10 m. surftrak held the ROV within 0.2 m of its target altitude for 96% of 21 h of survey transects, producing an approximately 1 m^2 standardized image footprint. Sufficient illumination and optimized camera settings yielded high-resolution imagery in visibility below 2 m, while topside power increased endurance and operational flexibility. Combining Doppler velocity log and underwater acoustic positioning data through the vehicle's extended Kalman filter improved navigation and post hoc transect reconstruction. During 2026 surveys, rov_flight_ops and rov_imagery_processing automated flight logging, log retrieval, fault diagnosis, and the sorting, development, and telemetry annotation of survey imagery. Dense kelp canopies nevertheless constrained vehicle operation, and downward-facing imagery provided limited observations of cryptic or obscured organisms. (4) We provide configuration-specific bills of materials, hardware and wiring documentation, software and firmware inventories, camera settings, open-source field and imagery-processing software, step-by-step field protocols, and pre- and post-dive checklists to support replication and adaptation of the framework. Together, these resources provide a field-tested starting point for expanding standardized benthic imaging while complementing diver surveys across kelp forests and other shallow coastal ecosystems.

ecology↗

bioRxiv2026

Disentangling shape and size variation to compare canopy architecture in maize

{middle dot} Comparing maize canopies is confounded by two problems: plants differ in the number of phytomers they produce, and phytomer size scales with that number. Position along the shoot is therefore an unreliable basis for comparison, confounding shape with size. {middle dot} We first position the floral transition (tassel initiation, T.I.) within shoot development, revealing it as the landmark that organizes a conserved canopy shape. This provides the foundation for aligning canopies that differ in phytomer number. {middle dot} Focusing on the leaf blade, we used functional data analysis on a maize panel across controlled and field environments to decompose variation in blade-length profiles, which was dominated (77%) by overall amplitude, identifying scaling as the major source of size variation. Because blades enlarge with each phytomer until the transition, later T.I. yields a larger maximum; this T.I.-linked component explains up to one-third of scaling variation, reinforced by a shared genetic association between T.I. and blade size with the flowering-time gene ZmCCT10. {middle dot} Regressing out this component leaves a residual that defines a new, heritable axis of intrinsic scaling, independent of flowering time. It distinguishes hyper- and hypo-scaling genotypes consistently across environments, offering a selection target decoupled from phenology.

plant biology↗

bioRxiv2026

A platform for automated training of mammalian cell physiology

Controlling cell physiology is difficult, not only because of cells complexity, but also their capacity for real-time adaptation to interventions, leading to challenges such as drug resistance and transgene silencing. Accumulating evidence suggests that this adaptivity resembles classical forms of learning defined in behavioral science. However, a lack of appropriate platforms has led to gaps in our understanding of cells capacity for adaptive problem-solving in physiological and transcriptional space. Here, we present a device, the Cell Trainer, capable of performing a wide variety of automated training experiments on non-neural mammalian cells, using timed drug pulses as the stimulus, and a mobile fluorescence microscope to capture images of responses, across replicate cultures. The Cell Trainer can operate in either an open-loop (feedforward) or closed-loop (feedback-controlled) mode, and our image analysis pipeline can report the behaviors of individual cells throughout each experiment and quantify population heterogeneity. We showcase the ability of the Cell Trainer to execute experimental protocols and perform single-cell analyses in both modes. We first demonstrate with a feedforward experiment in which myoblasts are repeatedly pulsed with dimethyl sulfoxide (DMSO) and their discrete calcium responses are analyzed, revealing sensitization-like dynamics. Next, we demonstrate a feedback control scheme wherein the fluorescence of a pH/voltage reporter in kidney cells is maintained below a threshold level with controlled pulses of acid. To accelerate research in the field of cell training, learning, and memory, we are openly sharing the Cell Trainer schematics and software with the research community. This platform provides a flexible tool for studying how cellular physiological states can be shaped by patterned stimulation and feedback control through approaches that work with the native adaptive competencies of cells.

bioengineering↗

bioRxiv2026

UNRAVELLING A HIDDEN SUBTYPE: MULTIOMICS REVEAL PSORIASIS-LIKE SIGNATURE WITH SURGICAL RELEVANCE IN A SUBSET OF PERIANAL FISTULIZING CROHN'S DISEASE

Background and aimsPerianal fistulizing Crohns disease (pCD) affects 20% of patients with Crohns disease (CD) and severely impacts quality of life. Current therapies fail to provide sustained relief, and the molecular underpinnings of pCD remain poorly understood. This study aimed to elucidate the molecular landscape of perianal fistulas through multiomic profiling. MethodsPaired fistula-tract and adjacent rectal mucosal biopsies were collected from 63 patients (48 with pCD, 15 with cryptoglandular fistula [CPTGL]). Longitudinal sampling generated 101 unique molecular profiles, comprising of both RNA sequencing (RNA-seq) and 16S rRNA sequencing from both tissue sites, followed by integrative multiomics and gene network analyses. ResultsUnsupervised clustering revealed three patient clusters primarily defined by host gene expression, with minimal contribution from microbial profiles. Fistulae in clusters 1 and 2 showed strong immune activation and epithelial-mesenchymal transition (EMT). In contrast, cluster 3 fistulae displayed keratinization and metabolic reprogramming resembling psoriatic skin, together with reduced JAK-STAT signalling. Cluster 3 was enriched for patients classified as TOpClass:2a, who are more suitable for surgical repair (p = 0.02). Conversely, cluster 2, characterized by rectal keratinization and EMT in both fistula and rectum, showed the highest MRI inflammatory-mass score (p = 0.02) and a greater risk of subsequent ileostomy (Kaplan-Meier; p = 0.008). An independent RNAseq dataset validated the keratinization signature in a subset of pCD fistulae. ConclusionIntegrated multiomic analysis identified distinct molecular subtypes of pCD with surgical and therapeutic relevance. These findings refine the molecular understanding of pCD and support a precision medicine approach. What You Need to Know?O_ST_ABSBACKGROUND AND CONTEXTC_ST_ABSO_LIPerianal fistulas affect 1 in 5 Crohns disease patients and have a severe negative impact on the quality of life of the patients. C_LIO_LIMost advanced IBD therapies are not efficacious for perianal Crohns disease. C_LIO_LISurgical repair is not suitable for all patients. C_LI NEW FINDINGSO_LIThis study is the first to delineate molecularly defined patient subtypes in perianal fistulizing Crohns disease. It further characterizes a psoriasis-like keratinization program in a subset of fistulas. Finally, it identifies molecular features and histological indicators that may explain-and potentially help predict-favorable surgical outcomes in selected patients C_LI LIMITATIONSO_LIFunctional and mechanistic studies will be required to further dissect the drivers and dynamics of the epithelial remodeling identified. C_LI CLINICAL RESEARCH RELEVANCEO_LIRaises the possibility of refining of existing clinical stratification using molecular markers for improved therapeutic management and outcome. C_LI BASIC RESEARCH RELEVANCEO_LIProposes keratinization as an opposing molecular process to EMT within the perianal fistula tract. C_LIO_LIIdentifies robust gene signatures associated with EMT and keratinization in the fistula for further experimental and clinical studies. C_LI

immunology↗

bioRxiv2026

Harmonized nucleoside mass spectrometry enables reproducible cross-platform RNA modification quantification

RNA modification analysis by LC-MS/MS is central to epitranscriptomics, yet quantitative comparison across laboratories and instrument platforms remains poorly standardized. Here, we performed a community-driven benchmarking study during the first Human RNome Project workshop to systematically evaluate cross-platform reproducibility of ribonucleoside mass spectrometry workflows. Using the same analytical column and gradient, standardized RNA samples, and shared reagents, we compared nucleoside quantification across quadrupole, time-of-flight, and orbitrap-based LC-MS platforms employing distinct acquisition strategies. While chromatographic separation was highly reproducible across systems, nucleoside-specific MS response behavior differed substantially between platforms and limited direct comparability of relative signal intensities. These response differences varied across analytes and concentration ranges, demonstrating that harmonized chromatography alone is insufficient for transferable quantitative analysis. Stable isotope-labeled internal standard (SILIS) normalization substantially reduced platform-and method-dependent response and improved agreement for most evaluated modifications. External calibration improved agreement between qTOF and Orbitrap workflows for a subset of modifications but did not fully resolve residual intersystem differences. Based on these findings, we establish benchmark-derived recommendations for harmonized relative and absolute RNA modification quantification, including guidance for calibration design, quality control, and data reporting. Together, this work provides a methodological framework for reproducible nucleoside LC-MS/MS workflows and establishes a foundation for large-scale comparative epitranscriptomic studies. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=107 SRC="FIGDIR/small/739095v1_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@6e4e98org.highwire.dtl.DTLVardef@1ea13beorg.highwire.dtl.DTLVardef@b65858org.highwire.dtl.DTLVardef@1623337_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2026

Suspension-Based Human Esophagoids Recapitulate WNT2B-Dependent Regulation of Esophageal Basal Progenitors

Background & AimsThe human esophagus undergoes a tightly regulated developmental program, transitioning from a simple columnar epithelium in early development to a mature stratified squamous tissue essential for adult barrier function. Here, we constructed a developmental cell atlas spanning early development to adulthood and leveraged it to generate physiologically relevant in vitro models. MethodsWe utilized single-cell RNA sequencing and spatial multiplex proteomics of human esophageal tissue from early development through adulthood. We established a feeder-supported 2D culture system and a Matrigel-free, suspension-based 3D esophagoid model in a 96-well format. To interrogate WNT2B function, we analyzed patient tissue harboring WNT2B loss-of-function mutations and performed WNT inhibition in esophagoids. ResultsSequencing profiling identified stage-specific epithelial populations: multiciliated and GPC3 basal cells were unique to early development; KRT14 basal and CRNN luminal cells were adult-specific; and COL17A1, LY6D, and KRT4 populations were shared across stages. Spatially organized WNT2B, KIT, and VWC2 mesenchymal subtypes were identified. The 2D system preserved both epithelial and mesenchymal compartments with transcriptional fidelity. Esophagoids exhibited basal-to-luminal stratification, mesenchymal compartmentalization, and required stromal interactions for formation. WNT2B repressed self-renewal of TP63 basal progenitors and inhibited proliferation, confirmed by pharmacologic inhibition of WNT in the in vitro esophagoids. ConclusionsWe present a stage-resolved atlas of human esophageal development and a scalable esophagoid platform recapitulating esophageal architecture. WNT2B regulates progenitor dynamics by restraining basal cell self-renewal. Esophagoids provide a physiologically relevant system for modeling esophageal development and disease. Visual Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=139 SRC="FIGDIR/small/733451v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1af5743org.highwire.dtl.DTLVardef@8a061dorg.highwire.dtl.DTLVardef@1975c3forg.highwire.dtl.DTLVardef@292ce9_HPS_FORMAT_FIGEXP M_FIG C_FIG Key Findings and ImplicationsO_LIDevelopmental Atlas: The study presents a comprehensive transcriptional and structural atlas of the human esophageal epithelium, identifying conserved and stage-specific epithelial populations from early development to adulthood. Notably, stage-specific gene expression of multiciliated and GPC3 basal cells were unique to early development, while KRT14 basal and CRNN luminal cells were adult specific, with COL17A1+ (basal), LY6D+ (epibasal), and KRT4+ (middle), shared at all stages. C_LIO_LIMesenchymal Diversity: Spatial and transcriptional profiling revealed distinct mesenchymal subtypes, including WNT2B, KIT, and VWC2 populations, which are spatially organized and contribute to epithelial-mesenchymal signaling. These findings reinforce the role of stromal-epithelial interactions in esophageal development. C_LIO_LI2D Esophagus Cell Culture System: A feeder-supported 2D cell culture system was developed that retains both epithelial and mesenchymal populations, preserving transcriptional fidelity and enabling long-term expansion for mechanistic studies. C_LIO_LI3D Esophagoid Model: A suspension-based 3D organoid system was optimized using a 96-well format, enabling high-throughput generation of esophagoids with robust epithelial stratification and mesenchymal compartmentalization. These organoids recapitulate key features of the human esophagus, including basal-to-luminal organization, and require stromal interactions for formation. C_LIO_LIFunctional Role of WNT2B in esophagus development: Both in vivo and in vitro analyses demonstrated that WNT2B regulates epithelial progenitor dynamics and tissue architecture by repressing self-renewal of basally localized TP63+ cells and inhibiting proliferation. Loss-of-function models and WNT pathway modulation confirmed its role in epithelial-mesenchymal crosstalk and organoid integrity. C_LI

developmental biology↗

bioRxiv2026

Phylogenetically estimated neutral rates and fitness effects of mutations to influenza proteins

Influenza viruss rapid evolution is shaped by both neutral mutation and selection. Phylogenetics can be used to study these processes, but this approach has typically only been applied to a few thousand influenza genome sequences at once. Here, we built phylogenetic trees with >100,000 influenza sequences, and then used these trees to estimate neutral rates of mutations to the viruss genome. Neutral rates varied by up to ~100-fold among the 12 nucleotide mutation types (A[->]C,A[->]G, etc.). These rates were highly correlated among influenza, SARS-CoV-2, and HIV, though more nuanced context-dependent patterns showed marked differences between influenza and SARS-CoV-2. We also estimated fitness effects of mutations by comparing the number of times a mutation was observed to occur along the branches of a tree to the number of times we expect it to have occurred under neutrality. We estimated effects for ~33,000 nonsynonymous and ~8,000 synonymous mutations spanning all influenza proteins. This compendium of estimated effects helps map the relationship between sequence and fitness in a natural setting, including regions where synonymous mutations are under functional constraint, and for proteins with limited experimentally measured effects. We built interactive heatmaps of the estimated fitness effects to help readers explore these data (see https://matsen.group/flu-mut-rates). Altogether, this work places influenzas mutation rates in a broader cross-viral context and deepens our understanding of how mutation and selection shape influenza evolution in nature at a site-specific level.

bioinformatics↗

bioRxiv2026

Extracellular vesicles from wild-type Epstein-Barr virus-transformed B-cells export host DNA and the viral lncRNA EBER1

Epstein-Barr virus (EBV) infection is nearly ubiquitous and strongly linked to multiple sclerosis (MS), but how EBV-infected B cells communicate with distal tissues remains unclear. We performed an integrated multiomic characterization of small extracellular vesicles (sEVs) released from spontaneous lymphoblastoid cell lines (SLCLs) derived from healthy donors and patients with MS, transformed ex vivo by endogenous wild-type EBV. Proteomics identified over 6,000 shared proteins enriched in nucleic acid-binding and chromatin-associated factors. EV-associated DNA resolved into two structurally distinct compartments: DNase-sensitive, high-molecular weight DNA associated with the vesicle corona and DNase-resistant, nucleosome-sized ([~]130-150 bp) DNA. Both compartments were overwhelmingly host-derived and broadly genomically distributed, whereas EBV DNA was minimal. In contrast, viral RNA cargo was dominated by the EBV noncoding RNA EBER1, which was strikingly enriched across all lines and confirmed within individual vesicles by ddPCR and super-resolution microscopy. EBER1 has previously been detected in MS brain tissue, yet its route to the CNS has remained unexplained. Our findings identify sEVs as a plausible vehicle for disseminating this immunostimulatory viral ncRNA beyond sites of latency, pointing to EV-mediated export of EBER1 as a candidate mechanism linking peripheral EBV infection to distal tissue signaling in MS and beyond.

immunology↗

bioRxiv2025

Variable Schwann cell merlin inactivation is targetable with TEAD1 inhibition in schwannomas.

Schwann cell tumors occur frequently in association with the vestibular nerves, leading to sensorineural hearing loss, and brainstem compression. In humans, unilateral vestibular schwannomas (VS) occur sporadically (VSspo)1, or bilaterally with neurofibromatosis type 2 syndrome (NF2) - VSnf2.2 VS formation is driven by sub-haploid NF2 gene dosage3, typically by biallelic loss.4,5 Loss of merlin promotes hippo/TEAD dependent transcriptional reprogramming, proliferation, and paracrine signaling that varies across time, and tumor volume.4,6 These variations lead to a clinically unpredictable course, and incomplete response to treatment. We hypothesized that Schwann cell merlin inactivation state determines cell-wise hippo/TEAD dependency and drives schwannoma pathogenesis. We analyzed clinical samples from VSspo and VSnf2 with a multi-omics approach and detected variation in merlin activity within tumor Schwann cell population. We found that tumor-driving merlin-depleted Schwann cells (Schwannmer-) exhibited elevated hippo activity that was predominantly driven by TEAD1. In-silico TEAD1 perturbation led to a reversal to merlin intact Schwann phenotype. These findings, and tumor cell growth suppression were confirmed in NF2fl/fl;Peri-Cre mouse model7, and in human derived schwannoma cells treated with a pan-TEAD auto palmitoylation inhibitor VT3989.8 Our computational and experimental results confirm that TEAD1 inhibition could be a potent, targeted strategy for schwannomas.

cancer biology↗

bioRxiv2025

Dynamic proteome profiling uncovers age-related impairments in proteostasis and the protective effects of resistance exercise in human skeletal muscle

A loss of proteostasis is a primary hallmark of ageing that has emerged from mechanistic studies in model organisms, but little is currently known about changes to proteostasis in the muscle of older humans. We used stable isotope labelling (deuterium oxide; D2O) in vivo, and peptide mass spectrometry of muscle samples to investigate differences in proteome dynamics between the muscle of younger (28 {+/-} 5 y; n=4) and older (69 {+/-} 3 y; n=4) men during either habitual activity or resistance exercise training. We quantified the abundance of 1787 proteins and the turnover rate of 1046 proteins in bi-lateral samples of vastus lateralis (n=32 samples total) taken before and after a 15-day program including 5 sessions of unilateral leg-press exercise (3 sets of 10 repetitions at 90% of 10 RM). Our protein abundance profiling revealed a stoichiometric imbalance within the proteostasis network in aged skeletal muscle, including subunits of eIF3, subunits of 40S and 60S ribosomal proteins. The rate of bulk, mixed-protein synthesis was not different between younger and older men, but most ribosomal proteins were less abundant in the muscle of older participants, suggesting ribosomes in older muscle may exhibit increased translational efficiency to maintain similar levels of protein turnover compared to ribosomes in younger muscle. Resistance exercise partially restored age-related disruptions to the proteostasis network. In older skeletal muscle, resistance exercise specifically increased the absolute turnover rate (ATR) of mixed mitochondrial proteins, with increased fractional turnover rate (FTR) of prohibitin 1 (PHB1) and profilin-1 (PROF1), and increased abundance of prohibitin 2 (PHB2). These adaptations may suggest resistance exercise promotes mitochondrial proteostasis by facilitating the synthesis and maintenance of key mitochondrial proteins. Thus, our Dynamic Proteome Profiling data provide an impetus for further exploration of the role of proteostasis in maintaining skeletal muscle quality and supports resistance exercise as a potential therapeutic strategy to promote healthy skeletal muscle ageing in humans. In BriefNishimura et al. used Dynamic Proteome Profiling to uncover whether the distorted proteomic landscape of ageing skeletal muscle is associated with altered turnover of specific proteins. Basal muscle from older men exhibits a divergence in protein abundance between subunits of eIF3 and subunits of 40S and 60S ribosomal proteins, whereas resistance exercise partially restored age-related disruptions in the muscle proteome. In older muscle, protein-specific turnover generally increases after resistance exercise, independent of changes in protein abundance, suggesting improved protein quality and renewal. Created in BioRender. Nishimura, Y. (2025) https://BioRender.com/p2a1aio HighlightsO_LIDynamic Proteome Profiling in human skeletal muscle ageing C_LIO_LIAgeing alters muscle proteostasis C_LIO_LIMixed-muscle protein synthesis does not differ between younger and older men C_LIO_LIResistance exercise increased mitochondrial protein turnover specifically in older muscle C_LIO_LIProtein-specific responses to resistance exercise differed between age groups C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/684531v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@11d0c8corg.highwire.dtl.DTLVardef@22f451org.highwire.dtl.DTLVardef@d62118org.highwire.dtl.DTLVardef@16eda9_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2025

Versatile NTP recognition and domain fusions expand the functional repertoire of the ParB-CTPase fold beyond chromosome segregation

Nucleotide triphosphate (NTP)-dependent molecular switches regulate essential cellular processes by cycling between active and inactive states through nucleotide binding and hydrolysis. These mechanisms were long thought to rely exclusively on ATPase or GTPase proteins, until the discovery of CTPase activity in the bacterial chromosome segregation protein ParB. In the ParABS system, CTP binding enables ParB accumulation around the centromere-like parS DNA sites to activate the ATPase ParA, thereby facilitating chromosome partitioning to daughter cells. CTP hydrolysis then releases ParB from DNA for recycling. This discovery uncovered a new regulatory principle, but the broader diversity of proteins employing a CTPase mechanism remains unclear. Here, we conduct a large-scale survey of proteins harboring the ParB-CTPase fold across bacteria, archaea, bacteriophages, and eukaryotes. While many ParB-like proteins follow the canonical ParABS organization with ParA partners, we also identify numerous orphan homologs encoded outside of the parAB operon, frequently linked to mobile genetic elements that may have driven their rapid diversification. The ParB-CTPase folds in these divergent proteins are often fused to lineage-specific domains with diverse predicted biological activities. We further demonstrate that while many homologs retain CTP-binding, others instead bind ATP or GTP, revealing a broader spectrum of nucleotide specificities than previously appreciated. Our findings establish the ParB-CTPase fold as a widely distributed and evolutionarily versatile NTP-binding module, repeatedly co-opted through domain fusion and shifts in nucleotide specificity to enable functions far beyond the classical ParABS-mediated DNA segregation.

microbiology↗

bioRxiv2025

Phosphoproteomic dysregulation drives tumor proliferation in Cushing's disease

Pituitary adenomas constitute up to 20% of primary brain tumors, yet somatic mutations are only found in 15% of pituitary adenomas. Epigenomic dysregulation has been proposed as a tumorigenic mechanism in pituitary adenomas causing Cushings disease (CD). We created paired datasets of human CD adenomas and en-route margin adult human pituitary glands and assayed their chromatin accessibility, DNA methylation, transcriptomic, proteomic and phospho-proteomic landscapes. In CD adenomas, we found epigenetic reactivation of a neurodevelopmental phosphoprotein program typically lost in the post-natal pituitary gland. CD cells overexpressed PPP1R17, a potent endogenous inhibitor of the ubiquitous protein phosphatase PP2A. Mechanistically, PPP1R17 overexpression in normal murine pituitary cells recapitulated the adenoma phenotype, and PPP1R17-mediated tumorigenesis was reversible using an FDA-approved small molecule PP2A agonist both in-vitro and in-vivo. Our findings highlight aberrant peptide phosphorylation as a targetable mechanism in CD. Significance statementCushings disease (CD) causes significant morbidity and mortality despite best medical and surgical treatment. Surgery is the mainstay of treatment, but carries perioperative risks and is frequently followed by remission. There is a paucity of effective medical treatments, due in part to a limited understanding of tumor mechanisms. The majority of CD adenomas are wild-type, with no known causal mutations. Our study identifies phosphoproteomic dysregulation as a mechanism of CD tumorigenesis common to wild-type and mutant CD adenomas. We target this pathway in-vivo and in-vitro using an FDA-approved small molecule. Our study proposes a novel therapeutic strategy for patients with CD.

cancer biology↗

bioRxiv2025

Human-mouse cross-species comparison identifies common and unique aspects of intestinal mesenchyme development

Using single-cell RNA sequencing (scRNA-seq) and histological approaches, we examine cross-species cellular identity, diversity and organization of mesenchymal (fibroblast) populations in the developing mouse and human intestines. In both species, we defined 7 fibroblast populations. Using cross-species integration and label transfer approaches we find that each mesenchymal cell subtype in the murine intestine is highly concordant to a cell type/state in the human intestine, and vice-versa, suggesting a strong conservation of mesenchymal cell types/states across species. Despite this conservation, we also observe that individual lineage-defining genes are not always shared and can be found in different mesenchymal populations. High resolution spatial analysis via fluorescent in situ hybridization (FISH) and immunofluorescence (IF) confirmed these findings and revealed that transcriptionally-defined sub-types of intestinal mesenchymal cells in mice and humans are organized within similar spatial domains.

developmental biology↗

bioRxiv2025

Developing future resilience from signatures of adaptation across the sorghum pangenome

While the green revolution adapted a handful of crops to homogenous and high-input industrialized agriculture, much of the global population still relies on local food production from low-input smallholder farms that grow highly variable crop cultivars. The high diversity of the grain and bioenergy crop sorghum 1-4, and many other crops that were not homogenized during the green revolution 5, not only provides the raw materials for breeders to make substantial gains in cultivar improvement, but also constrains breeding efforts due to highly specialized locally adapted plant phenotypes 6. Here, we construct a 33-member pangenome and identify trait-associated variants in 1,988 cultivars and landraces. We then apply these resources to explore the complex interplay between historical contingency, ongoing adaptation, and the potential for future gains through climate-aware genome-enabled breeding. Specifically, our analyses conclusively demonstrate that multiple nested, deeply diverged, and previously uncharacterized structural variants in the domestication gene SHATTERING1 distinguish the previously established multicentric origin of sorghum. We then apply landscape genomics tests to reveal how gene flow, adaptation, and secondary contact created the complex genetic mosaic in current global breeding networks. Further analysis of climate-gene associations highlights candidate loci underlying adaptation, including the biosynthetic gene cluster for the cyanogenic glucoside dhurrin. Combined, the pangenome-informed variants developed here will enable both trait discovery and subsequent marker assays to accelerate breeding and provide a framework for similar applications in other diverse and non-model crops.

genomics↗

bioRxiv2025

Root vulnerability to embolism and lack of physiological recovery limit the competitive ability of an invasive palm

O_LIClimate change and invasive species both threaten forest ecosystems. While exacerbated droughts drive tree mortality, species invasion alters forest composition. Invasive plants may be more vulnerable to drought due to acquisitive traits and low embolism resistance, but their potentially superior recovery capacity raises key uncertainties in predicting future species distributions. C_LIO_LIWe compared the drought resistance and recovery of an invasive palm (Trachycarpus fortunei) with two native species (Ilex aquifolium & Tilia cordata) from the Southern Alps. Saplings were exposed to increasing intensities of drought-induced xylem embolism, then rewatered for 45 days. Among others, we tracked net assimilation (Anet), stomatal conductance (gs), and leaf water potential ({Psi}leaf) before and after the drought, and modeled the time to hydraulic failure in leaves, stems, and roots. C_LIO_LIWhile the invasive palm had exceptionally drought-tolerant leaves, its roots were the most drought-sensitive among all species. The palm maintained positive gas exchange until {Psi}leaf of -4 MPa, as some native species; however, it failed to recover from the most severe drought, with simulations showing root embolism preceding leaf failure, unlike native species. C_LIO_LIInvasive plants may match or exceed natives drought resistance, but their inability to recover from extreme droughts could limit their future spread. C_LI

ecology↗

bioRxiv2025

YeiE regulates YeiH to implement sulfite stress resistance in Salmonella enterica serotype Typhimurium

Sulfites are toxic reactive sulfur species that are ubiquitous in nature. Bacteria are exposed to sulfites during the metabolism of sulfur-containing amino acids, during respiration using sulfur-containing electron acceptors, in host environments, and as food preservatives. Despite their broad distribution, the mechanisms bacteria use to resist the toxic effects of sulfites are poorly understood. Recent work showed that the LysR-type transcriptional regulator YeiE binds to sulfite to activate sulfite reduction gene expression. Using the model enteric pathogen, Salmonella enterica serotype Typhimurium, we show that YeiE is an autoregulator whose expression is enhanced by sulfite. We demonstrate here that the mechanism of sulfite stress resistance is distinct from pathways of sulfur assimilation for cysteine biogenesis. YeiE is necessary for survival in sulfite stress due to its regulation of the poorly characterized adjacent gene, yeiH. A yeiH deletion mutant is more sensitive to the growth-inhibiting effects of sulfite than a yeiE deletion mutant, demonstrating YeiH is the primary driver of sulfite stress resistance in vitro. This work provides a fundamental advance in understanding the sulfite stress response in bacteria with broad implications for food safety and host-pathogen interactions.

microbiology↗