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Masuda, Y.

Publications and source records attributed to Masuda, Y..

16 recordsLinked to original sources
bioRxiv2025

FoxO transcription factors couple the urea cycle and gluconeogenesis by controlling Ass1

Amino acid catabolism during fasting requires coordinated nitrogen disposal and glucose production, but the transcriptional logic linking the urea cycle to gluconeogenesis remains unclear. Forkhead box O (FoxO) transcription factors are key regulators of fasting metabolism, yet their role in controlling urea cycle genes has not been fully defined. Here we identify FoxOs as direct regulators of hepatic argininosuccinate synthase 1 (Ass1). Because FoxOs often act through Kruppel-like factor 15 (Klf15) in amino acid metabolism, we tested whether Ass1 regulation requires Klf15. Acute hepatic FoxO1/3a knockdown in fasted mice selectively reduced Ass1 expression, lowered blood glucose, and shifted urea-cycle amino acids, with arginine decreased and ornithine increased, even in Klf15-deficient livers. Silencing Ass1 phenocopied these metabolic effects, indicating that Ass1 mediates a key FoxO-dependent branch of fasting adaptation. Mechanistically, we mapped a functional FoxO-binding element within an upstream Ass1 enhancer: FoxO1/3a activated the enhancer in reporter assays, EMSA confirmed binding, and in vivo luciferase imaging and liver ChIP demonstrated fasting-inducible enhancer activity and FoxO occupancy. Collectively, these findings establish a FoxO-Ass1 axis that couples ureagenesis to gluconeogenesis and supports metabolic flexibility during fasting. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=158 SRC="FIGDIR/small/695746v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@94ff36org.highwire.dtl.DTLVardef@1ff4f3forg.highwire.dtl.DTLVardef@abd0e5org.highwire.dtl.DTLVardef@1f8d9b_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗
bioRxiv2025

Lateralized vagal oxytocin signaling separately controls feeding and socioemotional functions via hypothalamic oxytocin signaling

Oxytocin neurons in the paraventricular hypothalamus (PVHOxt) regulate feeding, anxiety, and social behaviors. Activation of Oxt receptor (Oxtr) -expressing vagal sensory neurons engages these PVHOxt neurons and improves hyperphagic obesity; however, their roles in anxiety and sociability remain unclear. Here, we activated vagal Oxtr-expressing neurons in male mice using a single intraperitoneal (IP) Oxt injection or chemogenetics. IP Oxt reduced anxiety-like behavior, enhanced social interaction, and suppressed feeding while activating both vagal sensory neurons and PVHOxt neurons. These effects were abolished by chemogenetic inhibition of PVHOxt neurons or central Oxtr blockade. Subdiaphragmatic vagotomy revealed lateralized functions: right-side vagotomy eliminated anxiolytic and prosocial effects, whereas left-side vagotomy blocked feeding suppression. Consistently, chemogenetic activation of left-sided neurons suppressed feeding, while right-sided activation reduced anxiety and increased sociability. These findings identify Oxtr-expressing vagal sensory neurons as a major peripheral pathway in which left- and right-sided inputs differentially control feeding and socioemotional behaviors.

physiology↗
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

Betaproteobacterial clade II nosZ activated under high N2O concentrations in paddy soil microcosms

AimsMicrobial communities in paddy soils act as potential sinks of nitrous oxide (N2O), a notorious greenhouse gas, but their potential to reduce external N2O is unclear. The direct observation of N2O reduction in submerged field soils is technically difficult. Here, we aimed to identify soil microbial clades that underpin the strong N2O mitigation capacity. Methods and ResultsWe constructed paddy soil microcosms with external N2O amendment that enabled the simultaneous evaluation of N2O reductase gene (nosZ) transcripts and N2O consumption. Although the amount of N2O amended was large, it was mostly consumed after 6- 8 days of microcosm incubation. Metatranscriptomic sequencing revealed that betaproteobacterial nosZ, especially those classified as clade II nosZ belonging to the orders Rhodocyclales or Nitrosomonadales, occupied >50% of the nosZ transcripts in three of the five paddy soils used. On the other hand, publicly available shotgun metagenomic sequences of 46 paddy soils were not dominated by betaproteobacterial clade II nosZ sequences, although they were ubiquitous. The same applied to the 16S rRNA sequences of Rhodocyclales or Nitrosomonadales. ConclusionsThe results indicated that betaproteobacterial N2O reducers potentially serve as powerful N2O sinks. Betaproteobacteria holding clade II nosZ can be targets of biostimulation, although further studies are required to understand their ecophysiology. Impact StatementOur results indicate that Rhodocyclales or Nitrosomonadales have the potential to serve as powerful N2O sinks in soils.

microbiology↗
bioRxiv2025

Nanopore sequencing-based measurement of paramyxovirus RNA editing reveals virus-specific differences in editing efficiency of mRNA, antigenome and genome

Paramyxovirus polymerase recognizes an RNA editing signal on the viral genome and transcribes mRNA in which guanine nucleotides are inserted in a template-independent manner. This enables the synthesis of multiple proteins from a single gene, which is important for viral growth. We developed a method to quantify RNA editing efficiency using Oxford Nanopore Technologies MinION platform. We performed sequence analysis of reverse transcription (RT)-PCR amplicons with the RNA editing sites in cells infected with Sendai virus (SeV) and canine distemper virus (CDV). By modifying RT primers, we simultaneously assessed RNA editing efficiency in mRNA, antigenome and genome. We observed distinct differences in mRNA editing efficiency between SeV and CDV. Notably, while RNA editing in SeV is confined to mRNA, in CDV it is also observed in antigenome/genome. (Anti)genomes harboring extra nucleotides may deviate from a multiple-of-six sequence, suggesting that RNAs not following the "Rule of Six" are produced in CDV-infected cells. HighlightsO_LIA method was established to quantify RNA editing efficiency in RNAs from paramyxovirus-infected cells using the MinION platform. C_LIO_LIRNA editing efficiency in mRNA differs between Sendai virus and canine distemper virus. C_LIO_LIAlthough RNA editing is confined to mRNA during Sendai virus infection, it is also observed in the antigenome and genome during canine distemper virus infection. C_LI

microbiology↗
bioRxiv2024

Comprehensive evaluation of pipelines for diagnostic biomarkers of major depressive disorder using multi-site resting-state fMRI datasets

The objective diagnostic and stratification biomarkers developed with resting-state functional magnetic resonance imaging (rs-fMRI) data are expected to contribute to more effective treatment for mental disorders. Unfortunately, there are currently no widely accepted biomarkers, partially due to the large variety of analysis pipelines for developing them. In this study we comprehensively evaluated analysis pipelines using a large-scale, multi-site fMRI dataset for major depressive disorder (MDD) (1162 participants from eight imaging sites). We explored the combinations of options in four subprocesses of analysis pipelines: six types of brain parcellation, four types of estimations of functional connectivity (FC), three types of site difference harmonization, and five types of machine learning methods. 360 different MDD diagnostic biomarkers were constructed using the SRPBS dataset acquired with unified protocols (713 participants from four imaging sites) as a discovery dataset and evaluated with datasets from other projects acquired with heterogeneous protocols (449 participants from four imaging sites) for independent validation. To identify the optimal options regardless of the discovery dataset, we repeated the same procedure after swapping the roles of the two datasets. We found pipelines that included Glassers parcellation, tangent-covariance, no harmonization, and non-sparse machine learning methods tended to result in high classification performance. The diagnosis results of the top 10 biomarkers showed high similarity, and weight similarity was also observed between eight of the biomarkers, except two that used both data-driven parcellation and FC computation. We applied the top 10 pipelines to the datasets of other mental disorders (autism spectral disorder: ASD and schizophrenia: SCZ) and eight of the ten biomarkers showed sufficient classification performances for both disorders, except two pipelines that included Pearson correlation, ComBat harmonization and random forest classifier combination. HighlightsO_LIWe evaluated the analysis pipelines of rsFC biomarker development. C_LIO_LIFour subprocesses in them were investigated with two multi-site datasets. C_LIO_LIGlassers parcellation, tangent covariance, and non-sparse methods were preferred. C_LIO_LIThe weight patterns of eight of the top 10 biomarkers showed high commonality. C_LIO_LIEight of the top 10 pipelines were successful for developing SCZ/ASD biomarkers. C_LI

neuroscience↗
bioRxiv2023

Inference of gene regulatory networks using time-series single-cell RNA-seq data with CRISPR perturbations

Single-cell RNA-seq analysis coupled with CRISPR-based perturbation (scCRISPR) has enabled the inference of gene regulatory networks (GRNs) with causal relationships. However, a snapshot of scCRISPR data may not lead to an accurate inference, since a gene knockout can influence multi-layered downstream over time. Here, we developed RENGE, a new computational method that infers GRNs using a time-series scCRISPR dataset. RENGE models the propagation process of the effects elicited by a gene knockout on its GRN. It can distinguish between direct and indirect regulations, which allows for the inference of regulations by genes that are not knocked out. RENGE therefore outperforms current methods in the accuracy of inferring GRNs. When used on a dataset we derived from human-induced pluripotent stem cells, RENGE yielded a GRN consistent with multiple databases and literature. Accurate inference of GRNs by RENGE would enable the identification of key factors for various biological systems.

bioinformatics↗
bioRxiv2023

Analysis of histone H3K4me3 modifications in bovine placenta derived from different calf-production methods

In ruminants, overgrowth of offspring produced by in vitro fertilization (IVF) is a common problem. Abnormal epigenetic modifications caused by environmental factors during the early embryonic period are suspected as an etiology of overgrowth. In this study, we investigated the genome-wide histone H3K4me3 profiles of bovine placentae that play a pivotal role in fetal development and compared their characteristics between artificial insemination (AI)- and IVF-derived samples. Cotyledons were harvested from the placentae obtained at parturition of 5 AI- and 13 IVF-derived calves, and chromatin immunoprecipitation sequencing was performed for H3K4me3. We confirmed no significant maternal tissue contamination in the samples we used. The revealed H3K4me3 profiles reflected the general characteristics of the H3K4me3 modification, which is abundantly distributed in the promoter region of active genes. By extracting common modifications from multiple samples, the genes involved in placenta-specific biological processes could be enriched. Comparison with the H3K4me3 modifications of blastocyst samples was also effective for enriching the placenta-specific features. Principal component analysis suggested the presence of differential H3K4me3 modifications in AI- and IVF-derived samples. The genes contributing to the difference were related to the developmental biological processes. Imprinted genes such as BEGAIN, ZNF215, and DLX5 were among the extracted genes. Principal component and discriminant analyses using only male samples categorized the samples into three groups based on fetal weight and calf-production methods. To our knowledge, this is the first study to profile the genome-wide histone modifications of bovine fetal placentae and reveal their differential characteristics between different calf-production methods.

genomics↗
bioRxiv2023

Genome-scale CRISPR-Cas9 screen identifies novel host factors as potential therapeutic targets for SARS-CoV-2 infection.

Although many host factors important for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection have been reported, the mechanisms by which the virus interacts with host cells remain elusive. Here, we identified tripartite motif containing (TRIM) 28, TRIM33, euchromatic histone lysine methyltransferase (EHMT) 1, and EHMT2 as novel proviral factors involved in SARS-CoV-2 infection by CRISPR-Cas9 screening. We demonstrated that TRIM28 plays a role(s) in viral particle formation and that TRIM33, EHMT1, and EHMT2 are involved in viral transcription and replication using cells with suppressed gene expression. UNC0642, a compound that specifically inhibits the methyltransferase activity of EHMT1/2, strikingly suppressed SARS-CoV-2 growth in cultured cells and reduced disease severity in a hamster infection model. This study suggests that EHMT1/2 may be a novel therapeutic target for SARS-CoV-2 infection.

microbiology↗
bioRxiv2022

Global soil metagenomics reveals ubiquitous yet previously-hidden predominance of Deltaproteobacteria in nitrogen-fixing microbiome

Biological nitrogen fixation is a fundamental process sustaining all life on earth. In the soil, Alphaproteobacteria, Betaproteobacteria, and Cyanobacteria are traditionally considered the major groups of nitrogen fixation population. This conventional knowledge has been accumulated by numerous PCR amplicon surveys for nitrogenase genes. However, this prevalent view appears to be inconsistent with results of recent PCR-free meta-omics investigations. Here, we show the global distribution and diversity of terrestrial diazotrophic microbiome, updated by fully leveraging the planetary collections of soil metagenomes along with recently expanded culture collections. After extensive analysis of 1,445 soil metagenomic samples, we proved that the previously-undervalued Anaeromyxobacteraceae and Geobacteraceae within Deltaproteobacteria are ubiquitous and predominant groups of diazotrophic microbiome in the soils, spanning a broad spectrum of environments, with different geographic origins and land usage types, including aerobic (croplands, grasslands) and anaerobic ones (paddy soils, sediments). Our results delineate a revised scheme of soil nitrogen fixation, which is inclusive of both conventional diazotrophs and the so far undervalued Deltaproteobacteria.

microbiology↗
bioRxiv2022

Is single-step genomic REML with the algorithm for proven and young more computationally efficient when less generations of data are present?

Efficient computing techniques allow the estimation of variance components for virtually any traditional dataset. When genomic information is available, variance components can be estimated using genomic REML (GREML). If only a portion of the animals have genotypes, single-step GREML (ssGREML) is the method of choice. The genomic relationship matrix (G) used in both cases is dense, limiting computations depending on the number of genotyped animals. The algorithm for proven and young (APY) can be used to create a sparse inverse of G[Formula] with close to linear memory and computing requirements. In ssGREML, the inverse of the realized relationship matrix (H-1) also includes the inverse of the pedigree relationship matrix, which can be dense with long pedigree, but sparser with short. The main purpose of this study was to investigate whether costs of ssGREML can be reduced using APY with truncated pedigree and phenotypes. We also investigated the impact of truncation on variance components estimation when different numbers of core animals are used in APY. Simulations included 150K animals from 10 generations, with selection. Phenotypes (h2 = 0.3) were available for all animals in generations 1-9. A total of 30K animals in generations 8 and 9, and 15K validation animals in generation 10 were genotyped for 52,890 SNP. Average information REML and ssGREML with G-1 and [Formula] using 1K, 5K, 9K, and 14K core animals were compared. Variance components are impacted when the core group in APY represents the number of eigenvalues explaining a small fraction of the total variation in G. The most time-consuming operation was the inversion, with more than 50% of the total time. Next, numerical factorization consumed nearly 30% of the total computing time. On average, a 7% decrease in the computing time for ordering was observed by removing each generation of data. APY can be successfully applied to create the inverse of the genomic relationship matrix used in ssGREML for estimating variance components. To ensure reliable variance component estimation, it is important to use a core size that corresponds to the number of largest eigenvalues explaining around 98% of total variation in G. When APY is used, pedigrees can be truncated to increase the sparsity of H and slightly reduce computing time for ordering and symbolic factorization, with no impact on the estimates. Lay SummaryThe estimation of variance components is computationally expensive under large-scale genetic evaluations due to several inversions of the coefficient matrix. Variance components are used as parameters for estimating breeding values in mixed model equations (MME). However, resulting breeding values are not Best Linear Unbiased Predictions (BLUP) unless the variance components approach the true parameters. The increasing availability of genomic data requires the development of new methods for improving the efficiency of variance component estimations. Therefore, this study aimed to reduce the costs of single-step genomic REML (ssGREML) with the Algorithm for Proven and Young (APY) for estimating variance components with truncated pedigree and phenotypes. In addition, we investigated the influence of truncation on variance components and genetic parameter estimates. Under APY, the size of the core group influences the similarity of breeding values and their reliability compared to the full genomic matrix. In this study, we found that to ensure reliable variance component estimation it is required to consider a core size that corresponds to the number of largest eigenvalues explaining around 98% of the total variation in G to avoid biased parameters. In terms of costs, the use of APY slightly decreased the time for ordering and symbolic factorization with no impact on estimations. Teaser TextEstimation of variance components is becoming computationally challenging due to the increasing size of genomic information. We investigated the impacts of using the algorithm for proven and young (APY) in genetic evaluations. The use of APY has no impact on variance components and genetic parameters estimation.

genomics↗
bioRxiv2020

Iodine staining as a useful probe for distinguishing amyloid fibril polymorphs

ABSTRACTIt is recently suggested that amyloid polymorphism, i.e., structural diversity of amyloid fibrils, has a deep relationship with pathology. However, its prompt recognition is almost halted due to insufficiency of analytical methods for detecting polymorphism of amyloid fibrils sensitively and quickly. Here, we propose that iodine staining, a historically known reaction that was firstly found by Virchow, can be used as a method for distinguishing amyloid polymorphs. When insulin fibrils were prepared and iodine-stained, they exhibited different colors depending on polymorphs. Each of the colors was inherited to daughter fibrils by seeding reactions. The colors were fundamentally represented as a sum of three absorption bands in visible region between 400-750 nm, and the bands showed different titration curves against iodine, suggesting that there are three specific iodine binding sites. The analysis of resonance Raman spectra and polarization microscope suggested that several polyiodide ions composed of I3− and/or I5− were formed on the grooves or the edges of β-sheets. It was concluded that the polyiodide species and conformations formed are sensitive to surface structure of amyloid fibrils, and the resultant differences in color will be useful for detecting polymorphism in a wide range of diagnostic samples.Competing Interest StatementThe authors have declared no competing interest.View Full Text

biochemistry↗
bioRxiv2019

Recombinant human soluble thrombomodulin is associated with attenuation of sepsis-induced renal impairment by inhibition of extracellular histone release

Multiple organ dysfunction induced by sepsis often involves kidney injury. Extracellular histones released in response to damage-associated molecular patterns are known to facilitate sepsis-induced organ dysfunction. Recombinant human soluble thrombomodulin (rhTM) and its lectin-like domain (D1) exert anti-inflammatory effects and neutralize damage-associated molecular patterns. However, the effects of rhTM and D1 on extracellular histone H3 levels and kidney injury remain poorly understood. Our purpose was to investigate the association between extracellular histone H3 levels and kidney injury, and to clarify the effects of rhTM and D1 on extracellular histone H3 levels, kidney injury, and survival in sepsis-induced rats. Rats in whom sepsis was induced via cecal ligation and puncture were used in this study. Histone H3 levels, histopathology of the kidneys, and the survival rate of rats at 24 h after cecal ligation and puncture were investigated. Histone H3 levels increased over time following cecal ligation and puncture. Histopathological analyses indicated that the distribution of degeneration foci among tubular epithelial cells of the kidney and levels of histone H3 increased simultaneously. Administration of rhTM and D1 significantly reduced histone H3 levels compared with that in the vehicle-treated group and improved kidney injury. The survival rates of rats in rhTM- and D1-treated groups were significantly higher than that in the vehicle-treated group. The results of this study indicated that rhTM and its D1 similarly reduce elevated histone H3 levels, thereby reducing acute kidney injury. Our findings also proposed that rhTM and D1 show potential as new treatment strategies for sepsis combined with acute kidney injury.

pharmacology and toxicology↗
bioRxiv2019

Exact p-values for large-scale single step genome-wide association, with an application for birth weight in American Angus

BACKGROUNDSingle Step GBLUP (SSGBLUP) is the most comprehensive method for genomic prediction. Point estimates of marker effects from SSGBLUP are often used for Genome Wide Association Studies (GWAS) without a formal framework of hypothesis testing. Our objective was to implement p-values for GWAS studies in the ssGBLUP framework, showing algorithms, computational procedures, and an application to a large beef cattle population. METHODSP-values were obtained based on the prediction error (co)variance for SNP, which uses the inverse of the coefficient matrix and formulas to compute SNP effects. RESULTSComputation of p-values took a negligible time for a dataset with almost 2 million animals in the pedigree and 1424 genotyped sires, and no inflation was observed. The SNP passing the Bonferroni threshold of 5.9 in the -log10 scale were the same as those that explained the highest proportion of additive genetic variance, but the latter was penalized (as GWAS signal) by low allele frequency. CONCLUSIONThe exact p-value for SSGWAS is a very general and efficient strategy for QTL detection and testing. It can be used in complex data sets such as used in animal breeding, where only a proportion of pedigreed animals are genotyped.

genomics↗
bioRxiv2018

Age-related macular degeneration affects the optic radiation white matter projecting to locations of retinal damage

PurposeWe investigated the impact of age-related macular degeneration (AMD) on visual acuity and the visual white matter.\n\nMethodsWe combined an adaptive cortical atlas and diffusion-weighted magnetic resonance imaging (dMRI) and tractography to separate optic radiation (OR) projections to different retinal eccentricities in human primary visual cortex. We exploited the known anatomical organization of the OR and clinically relevant data to segment the OR into three primary components projecting to fovea, mid- and far-periphery. We measured white matter tissue properties - (fractional anisotropy, linearity, planarity, sphericity) along the aforementioned three components of the optic radiation to compare AMD patients and controls.\n\nResultsWe found differences in white matter properties specific to OR white matter fascicles projecting to primary visual cortex locations corresponding to the location of retinal damage (fovea). Additionally, we show that the magnitude of white matter properties in AMD patients correlates with visual acuity. In sum, we demonstrate a specific relation between visual loss, anatomical location of retinal damage and white matter damage in AMD patients. Importantly, we demonstrate that these changes are so profound that can be detected using magnetic resonance imaging data with clinical resolution. The conserved mapping between retinal and white matter damage suggests that retinal neurodegeneration might be a primary cause of white matter degeneration in AMD patients.\n\nConclusionsThe results highlight the impact of eye disease on brain tissue, a process that may become an important target to monitor during the course of treatment.

neuroscience↗