Biology subjects

Yamamoto, M.

Publications and source records attributed to Yamamoto, M..

At least 19 recordsLinked to original sources

bioRxiv2026

High-titer convalescent serum from cats recovered from severe fever with thrombocytopenia syndrome protects SFTSV-infected mice

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging, tick-borne zoonosis caused by severe fever with thrombocytopenia syndrome virus (SFTSV) and is associated with high case-fatality rates in humans and cats. No vaccines or antiviral drugs are currently available for the prevention or treatment of feline SFTS. Because cats with SFTS often develop blood viral RNA loads exceeding those in severe human cases, we hypothesized that some naturally recovered cats develop strong neutralizing antibody responses and may serve as a potential source of antibody-rich serum for passive immunotherapy. We compared 80% plaque-reduction neutralization test (PRNT80) titers in convalescent sera from six human SFTS survivors and 14 cats that had recovered from naturally acquired SFTS, then evaluated the therapeutic efficacy of convalescent cat serum in SFTSV-infected, interferon receptor-deficient AG129 mice. Some SFTS-recovered cats showed high PRNT80 titers. Passive administration of high-titer convalescent cat serum early after infection protected SFTSV-infected mice. A single dose given at 2 or 3 days post-infection completely protected mice, whereas treatment at 4 days post-infection resulted in 40% survival. Administration on two consecutive days was associated with lower late-phase viremia in surviving mice. Sera from three independent SFTS-recovered cats improved survival, with greater effects observed for sera with higher neutralizing antibody titers. These findings indicate that some cats recovering from SFTS can develop high neutralizing antibody titers and their sera can exert therapeutic effects in an SFTSV-infected mouse model. These findings support further evaluation of passive immunotherapy as a treatment strategy for feline SFTS.

microbiology↗

bioRxiv2026

IL-17A Mediates Cardiac Hypertrophy in Chronic Kidney Disease through the IL-23/Th17 Axis.

Background: Chronic kidney disease (CKD) is a major risk factor for cardiovascular morbidity, promoting left ventricular hypertrophy (LVH) prior to overt heart failure. The mechanisms underlying early cardiac remodeling in CKD remain incompletely understood. The IL-23/Th17/IL-17A inflammatory axis has emerged as a key mediator of cardiovascular pathology; however, its role in CKD-associated cardiac remodeling remains unclear. This study aimed to elucidate the contribution of this pathway to the early cardiac changes in CKD. Methods: Using a 5/6 nephrectomy (5/6Nx) mouse model of CKD, we evaluated cardiac structure, mitochondrial function, and inflammatory signaling. Cardiomyocyte transcriptomes were analyzed by RamDA-seq to identify molecular alterations. Circulating cytokines were profiled longitudinally, and IL-17A neutralization experiments were performed to assess causality. In addition, serum IL-12p40 concentrations were measured in patients with CKD and correlated with their renal function and cardiac remodeling indices. Results: At 16 weeks post-5/6Nx, mice developed LVH with preserved systolic function, accompanied by increased oxidative stress and mitochondrial structural abnormalities, but without significant myocardial fibrosis. Transcriptomic analysis identified oxidative phosphorylation as the most significantly downregulated pathway in cardiomyocytes. Despite increased mitochondrial DNA copy number and elevated PGC-1 expression, ATP production and mitochondrial respiratory chain proteins were reduced, indicating mitochondrial dysfunction. Circulating IL-12p40 and IL-23 levels increased progressively, concurrent with expansion of splenic Th17 cells and elevated cardiac IL-17A expression. Recombinant IL-17A directly impaired mitochondrial respiration in cultured cardiomyocytes. In vivo, IL-17A neutralization attenuated LVH, reduced oxidative stress, and partially restored mitochondrial protein expression without improving renal function. In patients with CKD, serum IL-12p40 levels correlated inversely with estimated glomerular filtration rate and positively with left ventricular mass index, supporting clinical relevance. Conclusions: Activation of the systemic IL-23/Th17/IL-17A axis contributes to early CKD-associated cardiac remodeling by promoting oxidative stress and mitochondrial dysfunction. Circulating IL-12p40 may serve as a biomarker of cardiorenal remodeling, and the IL-23/IL-17A pathway represents a potential therapeutic target.

physiology↗

bioRxiv2026

Machine-learning-guided enzyme discovery and redox-system engineering for efficient production of the nylon monomer methyl 12-aminododecanoate in Escherichia coli

Methyl 12-aminododecanoate (ADAME) is a key precursor for Nylon 12 synthesis and an attractive target for sustainable microbial production. However, efficient biosynthesis of ADAME requires the coordinated oxidation and transamination of methyl dodecanoate (DAME), and identifying compatible enzymes for this multistep pathway remains challenging. In this study, we applied a machine learning-based bioinformatic screening approach to identify alternative enzymes for DAME-to-ADAME bioconversion. Among 18 selected distantly related AlkB homologs, AlkBGp01 from Alcanivorax sp. P2S70 showed activity toward DAME, producing 12-hydroxydodecanoic acid methyl ester (HDAME) and 12-oxododecanoic acid methyl ester (ODAME) despite low sequence identity to Pseudomonas putida AlkB. Further screening identified compatible redox partners, AlkG60 from the same species as AlkBGp01 and AlkTp04, whose combination with AlkBGp01 resulted in almost sixfold higher ODAME production than that achieved with P. putida AlkBFGJLT. ODAME production was further increased by more than threefold by conjugating these three newly identified proteins. The optimized single-plasmid system comprising conjugated AlkBGp01, AlkG60, and AlkTp04, together with AlaD and newly identified omega-transaminases EAV41574 produced 0.28 mM gDCW-1 ADAME, representing a 5.6-fold improvement over the previously reported P. putida AlkBFGJLT-based system with AlaD-CV2025. These results demonstrate that machine learning-guided enzyme discovery can identify functional distantly related homologs and compatible enzyme combinations for constructing efficient synthetic pathways. This study also highlights the importance of optimizing pathway architecture, including enzyme conjugation and plasmid configuration, for improving microbial production of bio-based monomer precursors.

bioengineering↗

bioRxiv2026

PEG-Arginase 1: A Novel Therapy for Optic Nerve Injury

Traumatic optic neuropathy (TON) occurs due to direct or indirect injury to the optic nerve and is a significant cause of visual disability. So far, there is no effective treatment. The lack of understanding of the cellular mechanisms by which trauma induces inflammation and damage in retinal neurons is a critical knowledge gap in developing effective therapies. We have studied the role of the arginase 1 (A1) enzyme in this pathology. We have found previously that treatment with a long-acting form of human recombinant A1, pegylated A1 (PEG-A1) after optic nerve crush limits activation of retinal microglia and macrophages (M{Phi}) and reduces inflammation, thereby decreasing injury and protecting visual function. Here we report on studies designed to demonstrate the therapeutic efficacy of PEG-A1 in mouse models of direct and indirect TON and to elucidate the underlying mechanisms. We used ONC to model direct TON and sonication-induced trauma to the supraorbital rim to model indirect TON (SI-TON). At different times after injury, mice were treated with PEG-A1 which was delivered systemically by i.p. injection or locally by intravitreal injection. In order to assess the role of A1-induced activation of the ornithine/polyamine pathway in the protective effects of PEG-A1, some mice were treated with the ornithine decarboxylase (ODC) inhibitor, difluoromethylornithine (DFMO) immediately after the PEG-A1 treatment. Retinal function was determined by OptoMotry and electroretinography. Retinal injury and microglia/M{Phi} activation were assessed by immunofluorescence imaging. Expression of inflammatory cytokines was determined by Western blotting and quantitative RT PCR. Liquid chromatography mass spectrometry was used to analyze changes in arginase/ODC pathway metabolites. Results showed that PEG-A1 treatment improved neuronal survival and visual function whether delivered systemically or intravitreally. This neuroprotection was associated with decreased microglia/M{Phi} activation, decreased inflammatory cytokine expression, and increased formation of L-ornithine and putrescine. Furthermore, DFMO treatment blocked these effects, indicating that PEG-A1 limits retinal injury and preserves vision after ocular injury by activating ODC. ODC processes the arginase product L-ornithine to form polyamines which are known to promote reparative functions. Thus, PEG-A1 therapy offers a new strategy to limit trauma-induced vision loss and promote repair after TON.

cell biology↗

bioRxiv2026

Multidimensional characterization of the physiological and behavioral effects of TCB-2 in mice

Background and PurposeSerotonergic psychedelics affect behavior and physiology, but the relationships among these effects remain poorly understood. In rodents, the head-twitch response is used as a measure of psychedelic-like activity, yet it does not capture changes in physiological state or the performance of learned behaviors. Here, we investigated the acute effects of the 5-HT2A receptor agonist TCB-2 across several behavioral and physiological measures and examined how these effects were modified by pretreatment with the 5-HT2A receptor antagonist volinanserin. Experimental ApproachMice were tested in head-fixed and freely moving conditions. During a learned auditory trace-conditioning task, we measured licking, pupil area, eye position, and blinking. We measured locomotor activity in an open field and quantified head-twitch responses using a DeepLabCut-based method. To examine the contribution of 5-HT2A receptors, mice were pretreated with the 5-HT2A receptor antagonist volinanserin. Key ResultsTCB-2 caused pupil constriction without detectable changes in eye position or blinking when administered alone. TCB-2 also reduced licking at the highest dose, but the cue-locked temporal pattern of licking remained evident. In freely moving mice, TCB-2 reduced locomotor activity and produced a dose-dependent increase in head-twitch responses. Volinanserin partially attenuated TCB-2-induced pupil constriction and reduced head-twitch responses under some conditions, but it did not consistently prevent the other effects of TCB-2. Conclusions and ImplicationsTCB-2 produced distinct effects across physiological and behavioral measures rather than a uniform disruption of behavioral function. Pronounced pupil constriction and head-twitch responses occurred without detectable changes in eye position or blinking, while the temporal organization of conditioned licking was retained despite a reduction in its magnitude. The incomplete and variable effects of volinanserin preclude definitive conclusions about the receptor mechanisms underlying each response. Combining automated head-twitch detection with physiological and task-related measurements provides a broader framework for comparing the pharmacological profiles of serotonergic compounds. What is already knownO_LIClassical psychedelics produce characteristic effects primarily through serotonin 5-HT2A receptor activation. C_LIO_LIHead-twitch responses capture only one dimension of psychedelic-like drug action. C_LI What this study addsO_LITCB-2 reduced locomotion and licking while preserving the cue-locked pattern of conditioned licking. C_LIO_LIPupil constriction occurred without detectable changes in eye position or blinking. C_LI Clinical significanceO_LIMultidimensional phenotyping can distinguish the physiological and behavioral profiles of serotonergic compounds. C_LIO_LIComplementary measures may improve preclinical evaluation of emerging serotonergic therapeutics. C_LI

pharmacology and toxicology↗

bioRxiv2026

Developmental expression of the skeletal muscle determination gene, MyoD, is regulated by novel enhancer elements that interact with the core enhancer and distal regulatory region

MyoD plays a central role in determining the skeletal muscle lineage in vertebrate embryos. The core enhancer (CE) and distal regulatory region (DRR) are the only known MyoD enhancers, and together, they recapitulate all major aspects of MyoD expression in the embryo. However, knocking out each enhancer individually has only modest effects on MyoD expression. Here, we show that embryos lacking both enhancers maintain muscle-specific MyoD expression, indicating the existence of unknown MyoD regulatory elements. Precision run-on sequencing together with available ChIP-seq and DNase I hypersensitivity datasets identified three new candidate enhancer regions within 96 kb of MyoD 5 flanking sequences. Transgenic analysis revealed that DNA elements at -36 and -60 kb are active in all muscle-forming regions, each recapitulating aspects of endogenous MyoD expression. Enhancer activities in muscle regulatory factor-deficient mice suggest that they are components of the auto- and cross-regulatory circuitry that maintains MyoD expression. Analysis of Hi-C data showed that the entire -96 kb region constitutes a loop domain, within which multiple interactions between elements and with the MyoD gene were detected. A larger loop domain delimited by CTCF sites was also identified from -96 kb to +220 kb relative to the MyoD transcriptional start site. These data indicate that the newly identified enhancers are key components of a cis regulatory network that controls the activation and maintenance of MyoD expression in the embryo. One sentence summaryCis regulation of MyoD transcription during development

developmental biology↗

bioRxiv2026

Minimally invasive measurement of maternal transcripts enables predicting the developmental potential of mammalian zygotes

Maternal transcripts are stored in the oocyte cytoplasm during oogenesis and play a pivotal role in early embryonic development after fertilization. However, specific maternal transcripts that reflect the developmental potential of embryos have not been systematically identified, and the use of maternal transcript levels as an indicator of successful development has not been explored. Here, we link the maternal transcriptome to the zygotes developmental potential by examining transcripts in a single polar body. The transcriptome of a zygote or an oocyte was highly similar to that of its accompanying polar body in mouse, cow, and human. We have identified a set of maternal transcripts whose expression levels fluctuate between poor- and good-quality zygotes. Specifically, Sipa1and Zmym6 were identified as marker transcripts that accurately reflect the developmental potential of zygotes. Using these marker genes, combined with machine learning, the development of zygotes to the blastocyst stage was successfully predicted with more than 80% specificity as early as 12 hours after fertilization. Furthermore, our prediction platform significantly improved implantation rates and live births to term. Thus, we have demonstrated a minimally invasive method for identifying maternal transcripts associated with zygote developmental potential. Our developed prediction system provides a generalizable conceptual framework for human infertility treatment to reduce the risk of implantation failure by excluding embryos with low developmental potential, especially when early embryos are transferred, and for livestock propagation to assess selected expressed maternal trait-associated variants before embryo transfer.

developmental biology↗

bioRxiv2026

A single PLAT domain protein couples reproductive arrest and carotenoid pigmentation during diapause in the two-spotted spider mite, Tetranychus urticae Koch

Adult females of the two-spotted spider mite, Tetranychus urticae Koch, enter a photoperiodically induced diapause to overwinter. Diapause in T. urticae is accompanied by reproductive arrest and the orange body coloration that arises from the accumulation of astaxanthin esters. How these two traits are coordinated at the molecular level remains poorly understood. Here, we compared the proteomes of adult females reared under diapause-inducing (long-night) and non-diapause-inducing (short-night) photoperiods using liquid chromatography-tandem mass spectrometry, followed by RNA interference (RNAi) of candidate genes. The carotenoid biosynthesis enzymes phytoene desaturase (TuPDS) and lycopene cyclase/phytoene synthase (TuLCPS), both encoded by genes horizontally transferred from fungi, were more abundant in diapausing females than in non-diapausing females. RNAi of the genes encoding TuPDS and TuLCPS markedly reduced orange pigmentation as well as {beta}-carotene and astaxanthin contents, demonstrating that these enzymes are required for diapause-associated pigmentation. Our proteomic analysis further identified a single PLAT (Polycystin-1, Lipoxygenase, Alpha-toxin) domain protein, TuPLAT10, as one of the most strongly upregulated proteins in diapausing females. The PLAT domain is a lipid-binding module, suggesting a role for TuPLAT10 in lipid metabolism. In addition to the suppression of orange pigmentation, RNAi of the TuPLAT10 gene restored reproduction even under diapause-inducing conditions and selectively reduced TuPDS and TuLCPS protein levels, despite the absence of sequence similarity to their genes. We propose that TuPLAT10 acts as a lipid-allocation switch that, in response to photoperiodic information, partitions fatty acids between astaxanthin esterification and yolk lipid supply, thereby coupling reproductive arrest and carotenoid pigmentation during diapause in T. urticae.

physiology↗

bioRxiv2026

Systemic injection of metabotropic glutamate 2/3 receptor antagonist LY341495 disrupts reward-related behaviors in mice.

Metabotropic glutamate 2/3 receptors (mGluR2/3) have been implicated in depression, anxiety, learning, and memory, yet their role in reward-related behavior remains poorly understood. Here, we examined the effects of the selective mGluR2/3 antagonist LY341495 on reward-related behaviors in mice using a combination of head-fixed and freely moving behavioral paradigms. In a head-fixed temporal conditioning task, mice developed anticipatory licking and pupil dilation preceding the delivery of a 10% sucrose solution, indicating successful temporal anticipation of reward. Systemic administration of LY341495 dose-dependently reduced both anticipatory and consummatory licking. Normalization analyses further suggested that LY341495 reduced the overall expression of licking behavior while largely preserving its temporal organization. LY341495 also induced pupil dilation and attenuated reward-proximity-related pupillary modulation. To determine whether these effects reflected non-specific motor impairments, we assessed locomotor activity in an open-field task and measured ultrasonic vocalizations (USVs) during courtship interaction. LY341495 did not significantly alter locomotion, excretion, or USV production, suggesting preserved general motor and orofacial motor function. In contrast, LY341495 dose-dependently reduced food intake and decreased social preference, indicating that the effects of mGluR2/3 antagonism extend beyond ingestive behavior to social reward-related processes. These findings demonstrate that mGluR2/3 signaling contributes to the regulation of reward-related behaviors across multiple reward domains independently of general motor dysfunction. These results provide new insight into glutamatergic mechanisms underlying motivation and reward processing and may have implications for neuropsychiatric disorders characterized by anhedonia and motivational deficits.

neuroscience↗

bioRxiv2026

Lipopolysaccharide-induced systemic inflammation alters courtship ultrasonic vocalizations in male mice.

BackgroundSickness behavior comprises a coordinated constellation of motivational, cognitive, and social alterations that emerge during systemic inflammation. Although reductions in locomotion, feeding, and social engagement have been extensively characterized, how inflammation affects ultrasonic vocal communication--an ethologically relevant index of social motivation in rodents--remains insufficiently understood. Here, we investigated how systemic immune activation alters male-female social communication in mice by jointly assessing ultrasonic vocalizations (USVs) and approach behavior. MethodsSexually experienced male mice received an intraperitoneal injection of lipopolysaccharide (LPS), and their interactions with a novel estrous female were evaluated 24 h later by quantifying USVs and approach behavior. ResultsLPS administration robustly suppressed both the total number of USVs and the duration of male approach behavior, indicating a pronounced reduction in social motivation. Beyond this quantitative suppression, LPS also induced qualitative changes in vocal output, including shifts in the proportional use of specific USV subtypes and alterations in acoustic features such as sound pressure. ConclusionsThese findings demonstrate that USVs capture multiple dimensions of inflammation-induced disruption of social communication, reflecting not only diminished motivation to engage socially but also changes in the structure of communicative signals themselves. By revealing that systemic immune activation reshapes both social approach behavior and vocal communication patterns, this study establishes USV analysis as a sensitive and translationally relevant behavioral readout for probing neuroimmune mechanisms underlying the social and communicative disturbances characteristic of sickness behavior. More broadly, our results highlight the utility of vocal communication analyses for elucidating how inflammatory processes perturb social circuits and communicative function in health and disease.

animal behavior and cognition↗

bioRxiv2026

Phenotypic diversity of yeasts curated in the 5th edition of The Yeasts: A data-driven visualization approach

Yeasts have long served as key experimental systems in genetics, cell biology, and fermentation research; however, these studies have largely focused on a limited number of model species. In contrast, yeasts are a fungal group with remarkable physiological and ecological diversity. To provide a global overview of yeast diversity, we compiled and visualized phenotypic information for approximately 1,300 yeast species documented in The Yeasts (5th edition), including carbon utilization profiles, fermentation capacity, growth temperature ranges, and reported isolation sources. Taxonomic reconciliation revealed extensive reannotation, with approximately 44% of the species undergoing name changes and recognized genera increasing from 143 to 233. Integrative analyses revealed pronounced phylogenetic structuring of metabolic breadth. Many basidiomycetous yeasts, particularly Agaricomycotina, exhibited broader, generalist-like carbon utilization, whereas ascomycetous yeasts, especially Saccharomycotina, more frequently displayed sugar-centered, specialist-like narrower profiles; model yeasts such as Saccharomyces cerevisiae and Schizosaccharomyces pombe fell at the narrow end of this spectrum. Fewer than half of all species fermented glucose, a trait largely confined to Saccharomycotina. In addition, nearly one-fifth of species failed to grow at 30 {degrees}C, the standard laboratory temperature. By reconstructing and visualizing decades of dispersed taxonomic knowledge accumulated in The Yeasts, this study reframes yeasts not merely as laboratory model organisms but as metabolically diverse fungi whose phenotypic diversity reflects diverse ecological contexts. The analytical framework presented here provides a foundation for integrating standardized quantitative phenotypes and newly described species and offers a starting point for exploring the latent ecological and metabolic potential of yeast diversity.

microbiology↗

bioRxiv2026

Breakdown of Rheological Universality in the Vegetal Hemisphere of Ascidian Embryos Mapped by Atomic Force Microscopy

Mechanical regulation plays an essential role in the organization of early embryogenesis. In ascidian embryos, cells in the animal hemisphere exhibit periodic stiffening-softening cycles, and their cell rheological properties follow a common master curve, indicating a form of universality. In contrast, cells in the vegetal hemisphere show cell-to-cell differences in stiffness even within the same endodermal lineage, but their rheological behavior has not been characterized. Here, using atomic force microscopy (AFM), we investigated the spatiotemporal dynamics of single-cell power-law rheological states in the vegetal hemisphere during early cleavage. We found that both the elastic modulus (stiffness) and the fluidity (power-law exponent) differ among endodermal cells, and that these differences change in a stage-dependent manner. This result indicates that vegetal cells do not exhibit a single common rheological behavior, in contrast to animal hemisphere cells, suggesting that mechanical properties in the vegetal hemisphere are not uniformly regulated but are patterned in space and time during cleavage. Our findings indicate that this mechanical diversification is linked to the progression of early morphogenesis and may contribute to the emergence of distinct cell behaviors during development.

biophysics↗

bioRxiv2026

Cell-autonomous thermogenesis of macrophage alters its antibacterial function

Dynamic temperature gradients exist across the bodies of endothermic animals, from the core to peripheral organ, resulting in the physiological cold environment in superficial regions. Consequently, macrophages distributed throughout the body must be able to adapt not only to thermoneutral conditions but also to colder environments. In fact, it is known that environmental temperature influences macrophage immune responses. However, the thermo-responsive mechanisms of macrophage have been largely unexplored. Here we show that macrophage themselves maintains intracellular temperature under physiological cold condition by increasing proton leak index (defined as mitochondrial proton leak per spare respiratory capacity). We further identified a contribution of ADP/ATP carrier (AAC) to this increase in proton leak index. This cell-autonomous thermogenesis pathway, which does not depend on neural or hormonal inputs, highlights the potential for local and organ-specific temperature regulation. Moreover, cold stress reduced mitochondrial membrane potential, which in turn suppressed the expression of the antimicrobial peptide Resistin-like molecule alpha (RETNLA) and diminished antibacterial properties. Together, these findings suggest that macrophages generate heat whereas compromising antibacterial properties, thereby increasing susceptibility to bacterial infection in physiological cold environment. This adaptation mechanism may underscore the important role of temperature homeostasis in non-adipocyte cells.

physiology↗

bioRxiv2025

Adaptation of an herbivorous arthropod to green tea plants by overcoming catechin defenses

Green tea catechins are known antioxidants that benefit human health and protect tea plants from biotic stressors. However, some herbivores can counteract catechin defenses and can use tea plants as a host. Among herbivorous mites, an extreme generalist Tetranychus urticae has not been reported as a tea pest. Instead, T. kanzawai, another generalist, has some populations that thrive on tea plants. Here, we investigated the mechanism of the adaptation of these mites to tea plants. Comparative study of the intra- and inter-specific variations in mite performances uncovered differences in their behavioral and xenobiotic responsiveness to green tea catechins. We showed that green tea catechins exert complex defensive roles. They were repellent and toxic to T. urticae and tea non-adapted T. kanzawai mites. In addition, they had an antifeedant effect on tea non-adapted T. kanzawai mites. Matching the catechin structure, we identified an intradiol ring-cleavage dioxygenase DOG15, a gene horizontally transferred from fungi, as one required for the adaptation of T. kanzawai mites to tea plants. The DOG15 gene has an enhanced inducible expression in tea-adapted T. kanzawai mites, with mRNA and protein levels up to 31.6 and 12.1 times higher than in T. urticae mites fed on tea plants. Furthermore, we identified two amino acid substitutions in DOG15 between Tetranychus species leading to the increased efficacy of the T. kanzawai encoded enzyme toward cleavage of green tea catechins. Thus, we showed that mite adaptation to tea plants occurred in a two-step process. The amino acid substitutions in DOG15 predispose T. kanzawai but not T. urticae for the adaptation to tea plants. Further increased expression of modified DOG15 enables T. kanzawai mites to efficiently detoxify green tea catechins, leading to intra- and inter-specific differences in mites ability to use tea plants as a host. Our findings reveal how a horizontally transferred gene can be co-opted through structural and regulatory changes to overcome plant chemical defenses, with implications for herbivore host adaptation and tea pest management.

evolutionary biology↗

bioRxiv2025

Structural and Thermodynamic Determinants of Anti-PGE2 mAb Specificity

We investigated the molecular determinants underlying the highly specific recognition of prostaglandin (PG) E2 and its structural analogues by an anti-PGE2 monoclonal antibody (mAb) and its Fab fragment using high-resolution X-ray crystallography coupled with detailed thermodynamic analyses, including conditions that mimic macromolecular crowding. Crystal structures of the Fab fragment in complex with PGE2, PGE1, and ONO5 were determined at 1.9 [A], 1.7 [A], and 1.7 [A] resolution, respectively, and the antigen-free Fab structure was solved at 2.0 [A]. The Fab accommodates the L-shaped PG scaffold within a deep antigen-binding crevasse formed between the {beta}-sheet frameworks of the VH and VL domains, where ligand recognition is mediated by multiple water-bridged hydrogen bonds and extensive hydrophobic contacts, including putative CH-O and CH-{pi} interactions. Comparison of antigen-free and ligand-bound structures revealed compensatory conformational rearrangements accompanied by reorganization of bound water networks. Upon PG binding, a short 310 helix forms at the tip of CDR-H1, stabilized by a cluster of hydrogen bonds and capping water molecules. This helix formation is associated with rotation of the VL domain and an increased Fab elbow angle. The CDR-H1 loop undergoes a flip-in transition in which Phe29H inserts into a hydrophobic cavity while Glu31H is extruded toward solvent, accompanied by {pi}-{pi} stacking between Tyr27H and Tyr32H. These changes collectively reduce hydrophobic surface area and promote enthalpy-driven ligand binding. The flexible -chain of PGE1, lacking the C5-C6 double bond, adopts altered conformations that disrupt C1-carboxyl interactions with three aromatic residues and lead to closure of a secondary water channel, consistent with subtle but meaningful differences in binding thermodynamics and crystallographic temperature factors relative to PGE2 and ONO5. Isothermal titration calorimetry (ITC) revealed high-affinity, enthalpy-driven PG binding to the Fab, in agreement with differential scanning calorimetry (DSC), which showed increased thermal stability upon complex formation. Under macromolecular crowding conditions induced by high bovine serum albumin (BSA) concentrations, PG binding to the intact mAb--but not to the Fab--exhibited improved discrimination between PGE2 and PGE1, suggesting that reduced entropy-enthalpy compensation under crowding amplifies subtle structural mismatches in ligand binding.

molecular biology↗

bioRxiv2025

Gelsolin Counteracts ER Stress-Driven Inflammatory Circuits in Psoriasis-like Dermatitis

Psoriasis is a chronic inflammatory skin disorder driven by amplified communication between immune cells and keratinocytes. Here, we show that imiquimod (IMQ) triggers organelle stress responses that directly contribute to this pathogenic circuit. In dendritic cells (DCs), IMQ promotes formation of ER-mitochondria contact sites (MAMs), inducing ER stress and activation of the unfolded protein response (UPR). These pathways act independently of, yet converge with, TLR7/MyD88 signaling to enhance IL-23 expression. IMQ also increases cytosolic Ca{superscript 2}+, facilitating NLRP3 inflammasome activation and release of mitochondrial DNA (mtDNA). In parallel, keratinocytes exposed to IMQ activate UPR-dependent genes, including Defb14 (mBD14), a psoriasis-associated antimicrobial peptide. Extracellular mtDNA and mBD14 then cooperatively stimulate plasmacytoid DCs through TLR9, establishing a feed-forward inflammatory loop. We further identify Gelsolin as a direct IMQ-binding protein that mitigates IMQ-induced ER stress; its loss amplifies ER stress, UPR activation, and oxidative stress, and its expression is reduced in human psoriatic lesions. Thus, MAM-UPR signaling links intracellular organelle stress to the intercellular networks that drive psoriatic inflammation, with Gelsolin acting as a critical intrinsic safeguard.

immunology↗

bioRxiv2025

The electrophilic metabolite of kynurenine, kynurenine-CKA, targets C151 in Keap1 to derepress Nrf2.

The Kelch-like ECH-associated protein 1 / Nuclear factor-erythroid 2 p45-related factor 2 (Keap1/Nrf2) system responds to a wide array of structurally diverse small molecules, of both exogenous and endogenous origin, by inducing a robust cytoprotective program that allows adaptation during oxidative, metabolic and inflammatory stress. Here, we report that exposure to the tryptophan metabolite kynurenine and its electrophilic derivative kynurenine-carboxyketoalkene (Kyn-CKA) leads to an increase in the abundance of transcription factor Nrf2 and induction of Nrf2-target genes, including NAD(P)H:quinone oxidoreductase 1 (NQO1), in murine and human cells. Additionally, both kynurenine and Kyn-CKA activate the aryl hydrocarbon receptor (AhR). Using cellular thermal shift assays, we found that Kyn-CKA increases the thermal stability of Keap1-mCherry fusion protein, but not free mCherry, indicating target engagement of Keap1, the principal repressor of Nrf2. The use of purified recombinant BTB domain of Keap1 and its C151S mutant counterpart revealed that Kyn-CKA reacts with wild-type, but not C151S mutant, Keap1-BTB, and at a much faster rate than with the small molecule thiol N-acetyl cysteine, demonstrating Kyn-CKA is targeted to react with C151 by the surrounding protein environment. In close agreement, Kyn-CKA increased the abundance of Nrf2 and expression of NQO1 in mouse embryonic fibroblast (MEF) cells expressing wild-type Keap1, but its inducer potency was greatly diminished in C151S-Keap1 mutant MEFs. Experiments in WT, AhR-knockout, and Nrf2-knockout primary murine bone marrow-derived macrophages showed that Nrf2 is required for the acute anti-inflammatory activity of Kyn-CKA, whereas AhR is dispensable. Together, these findings demonstrate that Kyn-CKA targets C151 in Keap1 to derepress Nrf2 and reveal that Nrf2, but not AhR, is a main contributor to the anti-inflammatory activity of Kyn-CKA in macrophages.

biochemistry↗

bioRxiv2025

Novel Role of AcylCoA:cholesterol acyltransferase 1 (ACAT1/SOAT1) in Diabetic Retinopathy

Hypercholesterolemia and excessive cholesterol ester (CE) production have been linked to chronic inflammation and vascular dysfunction during cardiovascular disease. Upregulation of AcylCoA:cholesterol acyltransferase 1 (ACAT1/SOAT1), the enzyme responsible for retinal CE formation, has been implicated in pathological retinal neovascularization. Here we determine the role of this process in diabetic retinopathy (DR). Ins2Akita diabetic mice were treated with the specific ACAT1/SOAT1 inhibitor K604 (10 mg/Kg, i.p.) beginning at 10 weeks for 2 weeks or 8 months for 2 months. ACAT1/SOAT1 expression and CE formation were assayed along with oxidative stress, inflammation, vascular pathology, and neuronal function. ACAT1/SOAT1 expression was also assayed in human retinas and vitrectomy specimens. Retinas from early-stage Ins2Akita mice exhibited increases in CE deposition, superoxide production, and expression of ACAT1/SOAT1, LDLR, TREM1, MCSF, and VEGF along with leukostasis, vascular leakage, acellular capillary formation, retinal ganglion cell loss, and impaired visual function. Late-stage increases in CE, ACAT1/SOAT, oxidative stress, inflammation, and impaired visual function were also observed. These changes were significantly inhibited by K604 treatment. The protective effects were independent of changes in systemic glucose or body weight. Human retina and vitrectomy samples also showed increases in ACAT1/SOAT1 and CE, respectively. Specific inhibition of ACAT1/SOAT1 with K604 normalizes ACAT1/SOAT1 expression and CE formation and prevents increases in oxidative stress and inflammation and preserves retinal structure and function in both early and late stages of DR. These findings identify ACAT1/SOAT1 as a promising therapeutic target for both early intervention and later stage treatment of DR. One sentence summaryInhibiting cholesterol esterification limits retinal neurovascular injury in diabetes.

cell biology↗