Biology subjects

Inoue, H.

Publications and source records attributed to Inoue, H..

At least 19 recordsLinked to original sources

bioRxiv2026

Adolescent prodromal phenotypes of psychiatric disorders: insights from a novel DISC1-perturbed monkey model

Neurodevelopmental deficits frequently lead to mild but significant adolescent social and cognitive changes, representing prodromal manifestations of adult-onset psychiatric disorders in a cross-disease manner. Although experimental medicine utilizes animal models to address the mechanisms, rodent brains lack critical structures for human higher brain functions such as the dorsolateral prefrontal cortex. To fill this translational gap, we generated monkeys with an impaired DISC1 gene locus, targeting the multifunctional protein essential for neurodevelopment and involved in transdiagnostic mental health risks. Here we analyze the F0 animals by minimizing mosaicism. At ages equivalent to human adolescence, these monkeys exhibited blunted responsiveness to social stimuli and increased internal drives, mirroring altered social behaviors in the prodromal stage in humans. Resting-state functional MRI data indicated disrupted global functional connectivity and reorganization of network topology, marked by a shift in the dominance of brain hubs from prefrontal regions to sensory areas. This primate model provides a unique platform for studying psychiatric disorders, such as schizophrenia, across the developmental trajectory.

neuroscience↗

bioRxiv2026

Rice brown spot resistance gene bsr1 also confers resistance to bacterial blight by suppressing sucrose efflux

Brown spot (BS), caused by the fungal pathogen Bipolaris oryzae, is a major disease threatening global rice production. However, the genetic basis of host BS resistance remains unclear. Here, we identified brown spot resistance 1 (bsr1), a quantitative trait locus conferring BS resistance, by map-based cloning. We show that bsr1 encodes a sucrose transporter and that a near-isogenic line carrying bsr1 (bsr1-NIL) in the susceptible Koshihikari genetic background exhibited resistance to BS by suppressing sucrose efflux into the apoplast after pathogen attack. Furthermore, bsr1-NIL also showed strain-specific resistance to bacterial blight caused by Xanthomonas oryzae pv. oryzae through the same mechanism. These findings demonstrate that bsr1 confers dual resistance to fungal and bacterial diseases by regulating sucrose efflux. Our study identifies a previously unrecognized mechanism underlying resistance to both BS and bacterial blight and highlights bsr1 as a promising target for breeding disease-resistance rice cultivars. Rice (Oryza sativa L.) is a staple food for more than half of the worlds population1. Brown spot (BS), caused by the fungus Bipolaris oryzae, is one of the most prevalent fungal diseases of rice, and its incidence has increased under global warming2. BS infects coleoptiles, leaves, leaf sheaths, panicle branches, glumes, and spikelets, and severe infection can substantially reduce grain yield.

plant 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

Fluorescence in situ hybridization reveals endophytic and epiphytic root colonization of the novel plant growth-promoting bacterium Citrobacter sedlakii CESi7

Plant growth-promoting bacteria are gaining significant attention as promising biofertilizers. However, the inconsistency between in vitro plant growth-promoting traits and actual field performance remains a challenge, driven partly by a limited understanding of in situ colonization. This study characterized the colonization patterns of Citrobacter sedlakii CESi7, a novel plant growth-promoting bacterium, isolated from oil palm waste compost, during Brassica rapa cultivation. The in situ behavior of CESi7 was observed in both sterilized medium and non-sterilized soil using fluorescence in situ hybridization with a strain-targeting probe. The results revealed that CESi7 can establish both epiphytic and endophytic populations that transiently colonize roots. In a sterilized medium, CESi7 was widely distributed throughout the root tissues. Conversely, in non-sterilized soil, the bacterium formed dense aggregates specifically at the root tips. This study provides direct microscopic evidence of the colonization strategy of CESi7, offering crucial insights for its development as an effective biofertilizer.

microbiology↗

bioRxiv2026

Off-target-free chemogenetic platform that decodes physiological roles of target GPCRs

G-protein-coupled receptors (GPCRs) are essential mediators of cellular processes, and their dysfunction is implicated in various diseases. Although pharmacological approaches are powerful for elucidating GPCR functions, careful consideration of potential off-target effects remains crucial. Here, we present an off-target-free chemogenetic platform to dissect the physiological roles of target GPCRs. As a proof-of-concept, we focused on the adenosine A2A receptor (A2AR), a prototypical GPCR involved in neurodegenerative diseases. Leveraging the ligand recognition mechanisms of A2AR, we designed a prominent chemogenetic pair comprising a clinical A2AR antagonist and an engineered A2AR mutant that is insensitive to the antagonist but retains responsiveness to adenosine. Using this pair, we uncovered the critical role of A2AR in neurite outgrowth under hypoxic conditions in neuron-like cells, while avoiding potential off-target effects. Furthermore, we demonstrate the generalizability of this chemogenetic approach to other class A GPCRs, offering a versatile platform for receptor-specific dissection of GPCR signaling.

biochemistry↗

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

A PKD-caveolin axis drives secretory carrier biogenesis at the TGN

Constitutive secretion from the trans-Golgi network (TGN) to the cell surface proceeds via carriers thought to form without a canonical cytoplasmic coat, yet how these carriers are generated remains poorly understood. Here, we identify a distinct population of TGN-to-cell surface carriers transporting influenza hemagglutinin (HA) and uncover a coat-like mechanism underlying their formation. HA carrier biogenesis requires non-vesicular lipid transfer at endoplasmic reticulum (ER)-Golgi membrane contact sites (MCSs) and protein kinase D (PKD) activity. We show that caveolin promotes membrane budding by assembling into cholesterol- and PKD-associated oligomers that act as a membrane-embedded, coat-like scaffold at lipid nanodomain-enriched TGN subdomains. These findings establish caveolin as a structural and regulatory component of TGN export and support a model in which a PKD-caveolin axis couples ER-Golgi lipid transfer to cargo sorting, membrane remodeling and fission during secretory carrier biogenesis.

cell biology↗

bioRxiv2025

Non-linear gene sets for digital biomarkers of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease caused by the loss of motor neurons. Accurate and accessible blood-based diagnosis of neurodegenerative diseases, including ALS, is becoming increasingly critical. It would be of a major clinical advantage to be able to distinguish ALS based on gene expression profiles of blood cells, but this still remains to be established. We analyzed existing data of peripheral blood mononuclear cells (PBMCs) of ALS patients by machine learning, the Maximum Mean Discrepancy (MMD), minimizing the issue of multi-collinearity shown in a multiple regression model, and identified a non-linear combination of gene sets to classify healthy controls and ALS, since non-linear approaches can capture intricate gene-gene interactions and threshold effects, which are often critical for accurately distinguishing disease from healthy states. In the gene expression profiles of PBMCs, a combination of expression levels in 3 genes, PRKAR1A, QPCT, and TMEM71, enabled us to classify ALS with an area under the curve (AUC) accuracy of 0.83 from the public database, which were then confirmed by laboratory blood samples with an AUC accuracy of 0.85. Furthermore, we found the expression levels of PRKAR1A, QPCT, and TMEM71 in motor neurons derived from induced pluripotent stem cells (iPSCs), the cell type at the core of the pathology, classified ALS with an accuracy of AUC 0.79. This approach of discriminating ALS with non-linear gene combinations may be useful for identifying ALS molecular biomarkers for blood-based diagnosis as well as for arriving at a completely new perspective of ALS pathogenesis.

molecular biology↗

bioRxiv2025

A Conserved Stress-Inducible Hub Enhancer Governs Natriuretic Peptide Expression and Undergoes Dynamic and Reversible Activation in Human Heart Failure

BackgroundNatriuretic peptides (NPs), encoded by the NPPA and NPPB genes, serve as both diagnostic biomarkers and cardioprotective hormones in heart failure. Their expression is tightly regulated by mechanical load, yet the upstream enhancer mechanisms translating hemodynamic stress into transcriptional activation remain incompletely understood. MethodsWe investigated the Nppa/Nppb super-enhancer (SE), which resides in a well-insulated topologically associating domain (TAD) and regulates stress-inducible expression of Nppa and Nppb. We applied CRISPR-based enhancer perturbation, chromatin accessibility and histone profiling, chromatin conformation assays, genetic mouse models, human iPSC-derived cardiomyocytes, and paired failing human hearts before and after left ventricular assist device (LVAD) unloading. ResultsThrough integrative epigenomic and genetic analyses, we identified a conserved element, CR9, as a dominant hub enhancer within this SE. In neonatal rat cardiomyocytes, CRISPR-based activation and inhibition established that CR9 was both necessary and sufficient for NP induction, whereas neighboring elements (CR7/CR8) displayed modest activity but synergized with CR9, establishing a hierarchical and cooperative SE architecture. In vivo, CR9 deletion in mice markedly suppressed Nppa/Nppb expression and diminished chromatin accessibility and histone acetylation across adjacent enhancers and promoters, underscoring its structural as well as transcriptional role. Reinsertion of CR9, even in reverse orientation, restored transcription, confirming its orientation-independent activity. CR9 function remained confined within TAD boundaries, supporting a locus-specific regulation. RNAscope revealed spatial proximity between CR9 enhancer RNA and Nppa/Nppb nascent transcripts. Developmental profiling revealed a switch in enhancer usage from CR6/CR7 in the fetal heart to CR9 in adulthood, indicating a transition from developmental to stress-inducible regulation. In human induced pluripotent stem cell-derived cardiomyocytes, deletion of CR9 nearly abolished NPPA/NPPB expression, establishing its essential role in a human cellular context. Most notably, in paired human failing hearts before and after LVAD unloading, chromatin accessibility at CR9 was dynamically reversed, providing the first direct evidence that enhancer states are plastic and therapeutically modifiable in the human myocardium. ConclusionsCR9 functions as a stress-inducible hub enhancer that coordinates NP transcription under pathological load. This enhancer exhibits reversible activation in human hearts, underscoring enhancer plasticity as a modifiable regulatory layer mediating stress-responsive transcriptional adaptation in the diseased myocardium. Clinical PerspectiveO_ST_ABSWhat is new?C_ST_ABSO_LIWe identified CR9 as a stress-inducible hub enhancer within the Nppa/Nppb super-enhancer that is both necessary and sufficient for natriuretic peptide induction, defining a hierarchical and cooperative regulatory architecture controlling Nppa and Nppb expression. C_LIO_LIWe uncovered a developmental switch in enhancer usage from CR6 in the fetal heart to CR9 in adulthood, revealing how stress-inducible enhancer activity emerges as the heart transitions from developmental to adaptive regulation. C_LIO_LIUsing paired failing human hearts before and after LVAD unloading, we provide the first direct evidence that enhancer states are dynamically reversible in the human myocardium. C_LI What are the clinical implications?O_LIThe reversibility of CR9 activity in human heart failure provides a molecular basis for how natriuretic peptide levels decrease when cardiac function improves with effective therapy, reflecting the underlying enhancer plasticity of the diseasesd myocardium. C_LIO_LIEnhancer plasticity is an emerging and targetable mechanism in heart failure, raising the possibility that stress-inducible enhancers like CR9 could be engineered to switch on cardioprotective genes when the heart is under stress, offering a new strategy for precision gene therapy. C_LI

molecular biology↗

bioRxiv2025

Mechanosensitive Stanniocalcin-1 Attenuates Pulmonary Arterial Hypertension by Suppressing Smooth Muscle Cell Proliferation

BackgroundIdiopathic pulmonary arterial hypertension (IPAH) is driven by progressive pulmonary vascular remodeling, particularly pulmonary arterial smooth muscle cell (PASMC) proliferation. Current combination vasodilator therapies have markedly improved outcomes; however, prognosis remains poor in subgroups such as patients with respiratory comorbidities, highlighting the need for novel therapies. Elevation of intravascular hydrostatic pressure is a hallmark of IPAH, yet its direct role in PASMC pathobiology remains largely unexplored. We developed a cyclic hydrostatic pressurization culture system to model hypertensive hemodynamic stress in vitro and identify pressure-responsive mediators. MethodsPASMCs from 4 patients with IPAH were exposed to high hydrostatic pressure (70/40 mmHg, 60 bpm). Transcriptomic profiling identified differentially expressed genes, validated by qPCR. Functional studies included piezo-type mechanosensitive ion channel component 1 (PIEZO1) modulation, recombinant human stanniocalcin-1 (rhSTC1) treatment, bromodeoxyuridine (BrdU) incorporation, and western blotting for cell-cycle regulators. In vivo, chronic hypoxia-induced pulmonary hypertension was assessed in wild-type and Stc1-/- mice by hemodynamic and histological analyses, with or without intratracheal rhSTC1 administration. ResultsRNA sequencing revealed STC1 as a robustly pressure-induced gene in IPAH PASMCs. PIEZO1 activation upregulated STC1, whereas knockdown blunted this response. Elevated STC1 expression was observed in PASMCs of IPAH lung tissues, and rhSTC1 reduced PASMC proliferation and increased p-p53, p21, and p27 expression. In the chronic hypoxia model, Stc1-/- mice exhibited higher right ventricular systolic pressure (RVSP) (43.7 {+/-} 1.3 vs. 30.6 {+/-} 0.9 mmHg) and greater pulmonary arterial medial thickness (39.1 {+/-} 2.5% vs. 26.5 {+/-} 1.3%) than wild-type mice. CD68-positive macrophages were increased in Stc1-/- mice under normoxia and further elevated with hypoxia. In wild-type and Stc1-/- PAH models, intratracheal administration of rhSTC1 markedly reduced medial thickening, CD68-positive macrophage accumulation, and RVSP in both wild-type and Stc1-/- mice. ConclusionsWe demonstrate that elevated hydrostatic pressure drives STC1 expression via PIEZO1, conferring potent anti-remodeling effects in IPAH. STC1 supplementation represents a potential therapeutic strategy that addresses an urgent medical need not fulfilled by conventional therapies. Clinical PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LIWe established a novel hydrostatic pressurization system to recapitulate idiopathic pulmonary arterial hypertension (IPAH) hemodynamic conditions in vitro. C_LIO_LIStanniocalcin-1 (STC1) is a hydrostatic pressure-responsive gene in pulmonary arterial smooth muscle cells (PASMCs) from IPAH patients, induced via the mechanosensitive receptor piezo type mechanosensitive ion channel component 1 (PIEZO1). C_LIO_LIExogenous STC1 suppresses PASMC proliferation and attenuates pulmonary vascular remodeling in chronic hypoxia-induced PAH models. C_LI What Are the Clinical Implications?O_LISTC1 supplementation represents a potential therapeutic strategy for IPAH, acting through a non-vasodilatory mechanism. C_LIO_LISTC1 supplementation may offer benefit in patients with limited response to current vasodilator therapies or with comorbid respiratory disease. C_LIO_LITargeting mechanotransduction pathways could expand treatment options for pulmonary hypertension. C_LI

molecular biology↗

bioRxiv2025

HiFAA: a high-confidence framework for transcription factor footprinting with ATAC-seq

Accurate inference of transcription factor (TF) binding from chromatin accessibility remains a major challenge, as conventional methods often miss true binding events, yield unstable motif rankings, or fail to resolve dynamic changes. Here we present high-confidence transcription factor footprint analysis using ATAC-seq (HiFAA), a framework that integrates topologically associating domain-restricted analysis, single-base-resolution footprint construction, and footprint ranking based on footprint bottom height. HiFAA markedly improves sensitivity while maintaining high specificity, enabling detection of binding events overlooked by existing analyses. It also ranks truly bound TFs with higher motif scores, enhancing predictive accuracy, and maintains consistently high geometric mean (G-mean) values, providing a stable balance between sensitivity and specificity. Furthermore, its high-resolution footprints capture subtle structural variations, allowing precise detection of developmental stage-specific TF-binding. By uniting sensitivity, specificity, and dynamic resolution, HiFAA establishes a robust paradigm for reliable TF footprinting from ATAC-seq data.

bioinformatics↗

bioRxiv2025

A vagal sensory--hypothalamic oxytocin--brown adipose tissue pathway mediates cholecystokinin-induced thermogenesis

Thermoregulation is essential for survival in homeothermic animals. Vagal sensory nerves are well known to detect visceral signals and regulate physiological functions, including feeding, metabolism and immunity. However, their role in thermoregulation remains poorly understood. Cholecystokinin (CCK), a gut hormone released postprandially, activates vagal sensory nerves via CCK-A receptors (CCK-AR). Exogenous CCK has been reported to induce thermogenesis in the intrascapular brown adipose tissue (iBAT), but the involvement of vagal sensory nerves and the central neural mechanisms that mediate this effect are not fully understood. In this study, we assessed the thermogenic effect of intraperitoneally (I.P.) administered CCK-8 and investigated the underlying autonomic reflex pathways. I.P. CCK-8 transiently and dose-dependently increased rectal temperature. This response was significantly attenuated by pharmacological blockade of CCK-AR, subdiaphragmatic vagotomy, or knockdown of CCK-AR primarily targeting in vagal sensory neurons. In addition, CCK-8 activated sympathetic nerve activity via vagal afferents. CCK-8-induced thermogenesis was blunted by iBAT sympathectomy or {beta}3-adrenergic receptor blockade. Furthermore, I.P. CCK-8 activated oxytocin neurons in the paraventricular nucleus of the hypothalamus (PVHOXT). Chemogenetic inhibition of PVHOXT neurons or intracerebroventricular administration of an oxytocin receptor (OXTR) antagonist attenuated the thermogenic response. These findings demonstrate, for the first time, the full neural circuitry underlying CCK-induced thermogenesis by delineating its afferent input (CCK-AR-expressing vagal sensory neurons), central integrative hub (PVHOXT neurons and OXTR signaling), and efferent output (iBAT sympathetic nerves). This study further suggests that CCK-AR -expressing vagal sensory neurons may contribute to thermoregulation under physiological conditions in which CCK is endogenously released. Key points summaryO_LIVagal sensory nerves, which connect the gut and the brain, play a key role in regulating meal-related physiology, however their role in thermoregulation remains incompletely understood. C_LIO_LIThis study reveals for the first time the full autonomic reflex pathways underlying thermogenic effect of the gut hormone cholecystokinin (CCK), comprising afferent input (CCK-A receptor-expressing vagal afferents), a central integrative hub (oxytocin neurons in the hypothalamic paraventricular nucleus; PVHOXT neurons), and efferent output (intrascapular brown adipose tissue via sympathetic nerves). C_LIO_LIBoth CCK-A receptors-expressing vagal afferents and sympathetic nerves innervating brown adipose tissue are required for thermogenesis induced by exogenous CCK-8. C_LIO_LIActivation of PVHOXT neurons by CCK-8 is critically involved in mediating this thermogenic effect. C_LIO_LIThis newly identified gut-brain-fat axis may contribute to part of diet-induced thermogenesis, and its impairment could be involved in the development of metabolic disorders such as obesity. C_LI

physiology↗

bioRxiv2025

Evaluation of real-time PCR performance for detecting Diaporthe destruens in sweet potatoes

Sweet potato foot rot disease caused by Diaporthe destruens is a major threat to sweet potato production. Rapid and accurate detection in advance is essential to secure healthy plants or to implement effective control. In this study, we evaluated the detection performance of a real-time PCR method previously developed by our research group. The evaluation was conducted using sweet potato samples collected in the field, including those naturally infected with D. destruens. Based on the real-time PCR results and the actual occurrence of the disease, a receiver operating characteristic (ROC) curve was generated, and the area under the curve (AUC) was calculated. The AUC values ranged from 0.7041571 to 0.9204286, confirming that this real-time PCR method is a well-balanced detection method for both tubers and stems. Furthermore, to minimize the occurrence of false negatives, the cycle threshold (Ct) cutoff was set at 35, and detection performance (sensitivity, specificity, accuracy, etc.) was analyzed. Then, by considering abnormal melting temperature (Tm) values as negative, high accuracy was achieved, with values of 0.871212 for tubers and 0.715447 for stems. This study not only evaluated the detection performance of the real-time PCR for foot rot pathogens but also contributes to providing information on the methodology and significance of performance evaluation in various real-time PCR genetic detection methods.

plant biology↗

bioRxiv2025

Global loss of responsiveness in key regulator metabolites and elevated enzyme proteins as metabolic dysregulation in skeletal muscle and liver of obese mice during starvation

Starvation induces complex metabolic adaptations in skeletal muscle, a key tissue for maintaining energy homeostasis; however, these adaptations are largely impaired in obesity. How obesity alters global metabolic adaptations to starvation in skeletal muscle remains unclear. Here, we analyzed the metabolic adaptations on a trans-omics scale during starvation in skeletal muscle from wild-type (WT) and leptin-deficient obese (ob/ob) mice. We measured multi-omics data during starvation and constructed global trans-omics networks in WT and ob/ob mice. We found that starvation induces "responsiveness" in WT mice, characterized by increases or decreases in key regulator metabolites, including ATP and AMP, as well as enzyme proteins, leading to global regulation of metabolic pathways, which was lost in ob/ob mice. In contrast, during starvation, ob/ob mice exhibit "difference" in comparison to WT mice, manifested by the persistently elevated expression of metabolic enzymes. These features were similarly found in liver, another key metabolic organ. Thus, global loss of responsiveness and elevated enzyme proteins are systemic features of metabolic dysregulation in ob/ob mice.

systems biology↗

bioRxiv2024

A novel RyR2-selective stabilizer prevents and ameliorates severe arrhythmias in stress-induced ventricular tachycardia

Background and PurposeAberrant activation of the type 2 ryanodine receptor (RyR2) causes lethal arrhythmias, such as catecholaminergic polymorphic ventricular tachycardia (CPVT). Developing drugs that suppress RyR2 hyperactivation may be key to novel arrhythmia treatments. This study evaluated the antiarrhythmic potential of Ryanozole, a recently developed novel RyR2 modulator with high affinity and selectivity, using CPVT mouse models harboring mutant RyR2s. Experimental approachIn vitro effects of Ryanozole were evaluated by ER Ca2+-based assay and [3H]Ryanodine binding assay using RyR2-expressing HEK293 cells. Two lines of mice with different arrhythmia severity, RyR2-R420W and -K4750Q, were employed for in vivo assessments. Intracellular Ca2+ signals were analyzed in isolated cardiomyocytes using Cal520. Antiarrhythmic effects were evaluated by ECG under catecholaminergic challenge in anesthetized mice and during spontaneous arrhythmias in conscious mice. ECG and echocardiographic parameters were evaluated before and after drug administration. Key resultsRyanozole inhibited both wild type and mutant RyR2s with similar IC50 of 15-40 nM. The inhibition was more potent at lower cytosolic Ca2+ concentrations. It suppressed Ca2+ waves and Ca2+ sparks without affecting action potential-evoked Ca2+ transients. Ryanozole effectively prevented adrenaline-induced arrhythmias and rapidly terminated spontaneous arrhythmias during daily activity. Importantly, Ryanozole did not impair cardiac conduction or contractility, unlike conventional antiarrhythmic drugs. Conclusions and implicationsRyanozole strongly suppresses RyR2 under diastolic Ca2+ conditions, thereby preventing the arrhythmogenic trigger of aberrant Ca2+ release. This mechanism likely provides potent antiarrhythmic effects while preserving cardiac function. Ryanozole is a promising therapeutic candidate treating RyR2-mediated arrhythmias, such as CPVT.

pharmacology and toxicology↗

bioRxiv2024

Isotope analysis of bird eye lens provides early-life information

Early-life environment has a long-lasting effect on later life, though its estimation is often prevented in the wild because of a lack of available methods. Recently, isotope analysis of eye lenses has attracted considerable interest as a means to reconstruct the environmental conditions experienced by animals during the developmental period. This analysis has mostly been confined to fish for practical reasons and remains to be resolved for application to other animals. In this study, we broadened its applicability by developing a novel approach and verifying its usability for the reconstruction of early-life environments. We performed a feeding experiment using Japanese quail (Coturnix japonica), in which we administered two diets: one composed mainly of C3 plants (low{delta} 13C and high{delta} 15N) and the other of C4 plants (high{delta} 13C and low{delta} 15N). Quails in the control group were continuously fed a C3-based diet from hatching until 200 days old, whereas those in the treatment groups (T10, T15, T20, and T40) were switched from the C3 to the C4-based diet at 10, 15, 20, and 40 days after hatching, respectively. We found that the{delta} 13C in the eye lenses of the treatment groups decreased from the center layer to the middle layer of the lens and then increased toward the outer layer, thus reflecting the diet change. In contrast, those of the control group exhibited a decreasing trend and equilibrated at the middle layer of the eye lens, with no increase thereafter. This novel approach revealed the postnatal feeding histories of the diet-shift experiment. The high{delta} 13C values observed in the center of the eye lenses would reflect the prenatal feeding environment, i.e., the C4-based diet consumed by their mothers, which is further reinforced by higher{delta} 15N values at this position due to the consumption of egg yolk-derived nutrition. These results indicate that the avian eye lens can be used as an "isotopic chronicle," which is a useful tool for reconstructing chronological isotopic information about their early-life history.

ecology↗

bioRxiv2024

Investigation of host plant contact with Diaphorina citri (Hemipreta: Psyllidae) by detecting D. citri-derived environmental DNA

Citrus greening (Huanglongbing) disease has serious impacts on citrus production. Field monitoring of the Asian citrus psyllid Diaphorina citri (Hemiptera: Psyllidae), a vector of citrus greening disease, is essential to prevent the invasion and spread of this disease. This study reported a new method for determining the presence of D. citri and traces of contact with host plants by collecting leaves of host plants from the survey area and detecting the environmental DNA (eDNA) derived from D. citri. The results showed that the method can determine the presence of D. citri. As D. citri-derived eDNA is contained in DNA solutions of plants with a history of contact with D. citri, we succeeded in detecting not only genes conserved in D. citri mitochondria but also genes of D. citri symbiont organisms consisting of Wolbachia, Wolbachia phage, Candidatus Carsonella sp., and Candidatus Profftella sp. Diaphorina citri-derived eDNA could be detected in host plants even after only 10 min of contact with D. citri and could still be detected 6 months after contact. This technology has the potential to trace D. citri from their traces without individual detection and is expected to greatly contribute to the early detection and invasion warning of citrus greening disease in the future.

molecular biology↗

bioRxiv2024

Structural robustness and temporal vulnerability of the starvation-responsive metabolic network in liver of healthy and obese mice

Adaptation to starvation is a multi-molecular and temporally ordered process, that could be impaired in obesity. To elucidate how the healthy liver regulates various molecules in a temporally ordered manner during starvation and how obesity disrupts this process, we measured time course multiomic data in the liver of wild-type (WT) and leptin-deficient obese (ob/ob) mice during starvation. Using the measured data, we constructed a starvation-responsive metabolic network, that is a transomic network including responsive molecules and their regulatory relationships during starvation, and analyzed the structure of the network. In WT mice, ATP and AMP, the energy indicators, regulated various metabolic reactions in the network as the hub molecules, both of which were not responsive in ob/ob mice. However, the structural properties of the network were maintained in ob/ob mice. In WT mice, the molecules in the network were temporally ordered through metabolic process coordinated by the hub molecules including ATP and AMP and were positively or negatively co-regulated. By contrast, both temporal order and co-regulation were disrupted in ob/ob mice. Taken together, the starvation-responsive metabolic network is structurally robust, but temporally vulnerable by the loss of responsiveness of the hub molecules in obesity. In addition, we proposed a potential therapeutic target to treat the negative effects of obesity on intermittent fasting to extend lifespan. One Sentence SummaryHub molecules activate or inhibit various molecules in a temporally ordered manner in healthy liver, and the regulatory network is structurally robust but temporally vulnerable to obesity.

systems biology↗