Biology subjects

Sakaguchi, S.

Publications and source records attributed to Sakaguchi, S..

32 records · Page 2Linked to original sources

bioRxiv2022

Transcriptional control of adipogenesis by PATZ1

White adipose tissues (WAT) play a central role in lipid storage and systemic energy, lipid, and glucose homeostasis. Understanding the intricacies of adipocyte formation could inform therapies for obesity and metabolic disorders. We have identified the POZ/BTB and AT Hook Containing Zinc Finger 1 (PATZ1) protein as an adipogenic transcription factor through an unbiased high-throughput cDNA screen for modulators of adipogenesis. PATZ1 is expressed by both human and mouse adipocyte precursor cells (APC) and adipocytes, and its expression negatively correlates with obesity. In cell models, PATZ1 expression promotes adipogenesis through a mechanism dependent on protein-protein interaction and DNA binding. Adipose-specific ablation of PATZ1 in mice leads to decreased lipid storage and fat mass and protection from diet-induced obesity. Genome-wide PATZ1 DNA binding analyses using ChIP-Seq suggest that PATZ1 facilitates adipogenesis through interactions with transcription factor machinery at the promoter regions of key adipogenic factors and histone modifiers. Global PATZ1 protein interaction studies using immunoprecipitation-mass spectrometry (IP-MS) suggest that General Transcription Factor 2I (GTF2I) binding to PATZ1 forms a repressive complex, and knockdown of GTF2I augments PATZ1 adipogenic function. Collectively these findings identify PATZ1 as a regulator of the adipocyte differentiation programs and advance our understanding of the complex transcriptional mechanisms underlying adipose tissue development and homeostasis.

molecular biology↗

bioRxiv2022

24-Nor-ursodeoxycholic acid counteracts TH17/Treg imbalance and ameliorates intestinal inflammation by restricting glutaminolysis in differentiating TH17 cells

Objective24-Nor-ursodeoxycholic acid (NorUDCA) is a novel therapeutic bile acid for treating primary sclerosing cholangitis (PSC), an immune-mediated cholestatic liver disease. Since PSC strongly associates with inflammatory bowel diseases (IBD) driven by TH17/Treg imbalance, we aimed to explore NorUDCAs immunomodulatory potential on intestinal TH17/Treg balance. DesignNorUDCAs impact on TH17/Treg tissue distribution was first assessed in Mdr2-/- mouse model of PSC. We specifically investigated NorUDCAs effect on modulating TH17/Treg balance in a CD4+ T cell driven colitis model induced by adoptive transfer of CD25-CD44lowCD45RBhighCD4+ TNaive cells into Rag2-/- mice, mimicking human IBD. Mechanistic studies were performed using molecular approaches, flow cytometry and metabolic assays in murine TH17 cells in vitro. NorUDCAs signaling effects observed in murine system were further validated in circulating CD4+ T cells from PSC patients with co-existing IBD. ResultsNorUDCA promoted Treg generation in both liver and intestine in the Mdr2-/- model. In the experimental IBD model, NorUDCA attenuated intestinal immunopathology. Mechanistically, NorUDCA demonstrated strong immunomodulatory efficacy in counteracting TH17/Treg imbalance by restricting glutaminolysis in differentiating TH17 cells, thus suppressed -Ketoglutarate-dependent mTORC1 activation, glycolysis and enhanced FOXP3 expression. NorUDCAs impact on mTORC1 signaling was further confirmed in circulating CD4+ T-cells from PSC patients with IBD. ConclusionNorUDCA possesses direct immunometabolic modulatory potency to counteract TH17/Treg imbalance and ameliorate excessive TH17 cell driven intestinal immunopathology. These findings extend future clinical applications of NorUDCA for treatment of TH17 cell-mediated disorders along the gut-liver axis and beyond. Significance of this studyO_ST_ABSWhat is already known on this subject?C_ST_ABSO_LIPSC is an immune-mediated cholestatic liver disease highly associated with IBD where TH17/Treg imbalance drives immunopathogenesis; seeking effective therapeutics covering both liver and intestinal disease in PSC is of high clinical relevance. C_LIO_LIIndependent of anti-cholestatic effects, NorUDCA has recently been shown to possess direct immunomodulatory properties on CD8+ T cell metabolism, lymphoblastogenesis and clonal expansion through targeting mTORC1 signaling. C_LIO_LISince mTORC1 serves as critical metabolic checkpoint orchestrating TH17/Treg axis, inhibiting mTORC1 activity represents a potential treatment avenue counteracting TH17/Treg imbalance under intestinal inflammatory conditions. C_LI What are the new findings?O_LINorUDCA enriches FOXP3+ Treg population in both liver and intestinal tissue in the cholestatic Mdr2-/- mouse model of PSC. C_LIO_LINorUDCA exhibits direct immunomodulatory efficacies in suppressing excess TH17 cell-mediated intestinal immunopathology and promotes FOXP3+ Treg generation in an experimental IBD model. C_LIO_LIMechanistically, NorUDCA counteracts TH17/Treg imbalance by restricting glutaminolysis in differentiating TH17 cells, thus suppresses -Ketoglutarate-dependent mTORC1 activation, glycolysis and enhances FOXP3 expression. C_LIO_LINorUDCAs impact on mTORC1 signaling was further confirmed in circulating CD4+ T cells from patients with PSC and IBD. C_LI How might it impact on clinical practice in the foreseeable future?These findings advance our current understanding of therapeutic potentials of NorUDCA, which might represent a novel therapeutic strategy in the treatment of PSC and concomitant IBD and other TH17-mediated intestinal diseases.

molecular biology↗

bioRxiv2022

NeoRdRp: A comprehensive dataset for identifying RNA-dependent RNA polymerase of various RNA viruses from metatranscriptomic data

RNA viruses are distributed throughout various environments, and most RNA viruses have recently been identified by metatranscriptome sequencing. However, due to the high nucleotide diversity of RNA viruses, it is still challenging to identify novel RNA viruses from metatranscriptome data. To overcome this issue, we created a dataset of RNA-dependent RNA polymerase (RdRp) domains that are essential for all RNA viruses belonging to Orthornavirae. Genes with RdRp domains from various RNA viruses were clustered based on their amino acid sequence similarity. For each cluster, a multiple sequence alignment was generated, and a hidden Markov model (HMM) profile was created if the number of sequences was greater than three. We further refined the 426 HMM profiles by detecting the RefSeq RNA virus sequences and subsequently combined the hit sequences with the RdRp domains. As a result, a total of 1,182 HMM profiles were generated from 12,502 RdRp domain sequences, and the dataset was named NeoRdRp. Almost all NeoRdRp HMM profiles successfully detected RdRp domains, specifically in the UniProt dataset. Furthermore, we compared the NeoRdRp dataset with two previously reported methods for RNA virus detection using metatranscriptome sequencing data. Our methods successfully identified most of the RNA viruses in the datasets; however, some RNA viruses were not detected, as in the cases of the other two methods. The NeoRdRp can be repeatedly improved by adding new RdRp sequences and is applicable as a system for detecting various RNA viruses from diverse metatranscriptome data.

microbiology↗

bioRxiv2021

Single-cell transcriptome atlas of Drosophila gastrula 2.0

During development, positional information directs cells to specific fates, leading them to differentiate with their own transcriptomes and express specific behaviors and functions. However, the mechanisms underlying these processes in a genome-wide view remain ambiguous, partly because the single-cell transcriptomic data of early developing embryos containing both accurate spatial and lineage information is still lacking. Here, we report a new single-cell transcriptome atlas of Drosophila gastrulae, divided into 65 transcriptomically distinct clusters. We found that the expression profiles of plasma-membrane-related genes, but not those of transcription factor genes, represented each germ layer, supporting the nonequivalent contribution of each transcription factor mRNA level to effector gene expression profiles at the transcriptome level. We also reconstructed the spatial expression patterns of all genes at the single-cell stripe level as the smallest unit. This atlas is an important resource for the genome-wide understanding of the mechanisms by which genes cooperatively orchestrate Drosophila gastrulation.

developmental biology↗

bioRxiv2021

Deeply hidden genome organization directly mediated by SATB1

Mammalian genomes are organized by multi-level folding, yet how this organization contributes to cell type-specific transcription remain unclear. A nuclear protein SATB1 forms a SATB1-rich subnuclear structure that resists high-salt extraction. SATB1 binds in the minor groove of double-stranded base-unpairing regions (BURs), genomic elements with high unwinding propensities. We uncovered that SATB1 establishes a two-tiered chromatin organization, one through indirect binding and another by direct binding of BURs. Published ChIP-seq datasets show SATB1 binding to highly accessible chromatin at enhancers and CTCF sites, but not to BURs. By employing urea ChIP-seq, which retains only directly bound protein:DNA complexes, we found that BURs, but not CTCF sites, are direct SATB1 binding targets genome-wide. BURs bound to the SATB1 nuclear substructure interact with accessible chromatin crossing multiple topologically associated domains (TADs). SATB1 is required for these megabase-scale interactions linked to cell type-specific gene expression. BURs are highly enriched within lamina associated domains (LADs), but some ([~]10%) are found in gene-rich accessible chromatin outside LADs as well. Only a subset of BURs is bound to SATB1 depending on cell type. Notably, despite the mutually exclusive SATB1-binding profiles uncovered by the two ChIP-seq methods, we found most peaks in both profiles are valid and require SATB1. Based on these and previous data, we propose that the SATB1 protein network forms a chromatin scaffold, providing an interface that connects accessible chromatin to a subnuclear architectural structure, thereby facilitating the three-dimensional organization linked to cell type-specific gene expression.

genomics↗

bioRxiv2021

Myasthenia gravis-specific aberrant neuromuscular gene expression by medullary thymic epithelial cells in thymoma

Myasthenia gravis (MG) is a neurological disease caused by autoantibodies against neuromuscular-associated proteins. While MG is frequently developed in thymoma patients, the etiologic factors for MG are not well understood. Here, by constructing a comprehensive atlas of thymoma using bulk and single-cell RNA-seq, we identified ectopic expression of neuromuscular molecules in MG-associated thymoma (MG-thymoma). These molecules were originated from a distinct subpopulation of medullary thymic epithelial cells (mTECs), which we named neuromuscular mTECs (nmTECs). MG-thymoma also exhibited microenvironments dedicated to autoantibody production, including ectopic germinal center formation, T follicular helper cell accumulation, and type 2 conventional dendritic cell migration. Cell-cell interaction analysis also predicted the interaction between nmTECs and T/B cells via CXCL12-CXCR4. The enrichment of nmTECs presenting neuromuscular molecules within MG-thymoma was further confirmed by immunohistochemically and by cellular composition estimation from MG-thymoma transcriptome. Altogether, this study suggests that nmTECs play a significant role in MG pathogenesis via ectopic expression of neuromuscular molecules.

immunology↗

bioRxiv2021

24-Nor-Ursodeoxycholic acid reshapes immunometabolism in CD8+ T cells and alleviates hepatic inflammation

Background & Aims24-NorUrsodeoxycholic acid (NorUDCA) is novel therapy for immune-mediated liver diseases such as primary sclerosing cholangitis (PSC) where dysregulated T cells including CD8+ T cells cause liver immunopathology. We hypothesized that NorUDCA may directly modulate CD8+ T cell effector function thus contributing to its therapeutic efficacy independent of anti-cholestatic effects. MethodsNorUDCA effects on CD8+ T cell function in vivo were investigated in a hepatic injury model system induced by excessive CD8+ T cell immune response upon non-cytolytic lymphocytic choriomeningitis virus (LCMV) infection. Mechanistic studies included molecular and biochemical approaches, flow cytometry and metabolic assays in mouse CD8+ T cells in vitro. Mass spectrometry (MS) was used to identify potential targets modulated by NorUDCA in CD8+ T cells. NorUDCA signaling effects observed in murine systems were validated in peripheral T cells from healthy volunteers and PSC patients. ResultsIn vivo NorUDCA ameliorated hepatic injury and systemic inflammation upon LCMV infection. Mechanistically, NorUDCA demonstrated a strong immunomodulatory efficacy in CD8+ T cells affecting lymphoblastogenesis, mTORC1 signaling and glycolysis of CD8+ T cells. With MS, we identified that NorUDCA regulates CD8+ T cells via targeting mTORC1. NorUDCAs impact on mTORC1 signaling was further confirmed in circulating human CD8+ T cells. ConclusionsNorUDCA possesses a yet-unrecognized direct modulatory potency on CD8+ T cells and attenuates excessive CD8+ T cell hepatic immunopathology. These findings may be relevant for treatment of immune-mediated liver diseases such as PSC and beyond.

molecular biology↗

bioRxiv2020

Scalable, multimodal profiling of chromatin accessibility and protein levels in single cells

Recent technological advances have enabled massively parallel chromatin profiling with single-cell Assay for Transposase Accessible Chromatin by sequencing (scATAC-seq) in thousands of individual cells. Here, we extend these approaches and present ATAC with Select Antigen Profiling by sequencing, ASAP-seq, a tool to simultaneously profile accessible chromatin and protein levels in thousands of single cells. Our approach pairs sparse scATAC-seq data with robust detection of hundreds of cell surface and intracellular protein markers and optional capture of mitochondrial DNA (mtDNA) for clonal tracking, thus concomitantly capturing three distinct modalities in single cells. Importantly, ASAP-seq uses a novel bridging approach that repurposes antibody:oligo conjugates designed for existing technologies that pair protein measurements with single cell RNA-seq. We demonstrate the utility of ASAP-seq by revealing coordinated and distinct changes in chromatin, RNA, and surface proteins during native hematopoietic differentiation, peripheral blood mononuclear cell stimulation, and as a combinatorial decoder and reporter of multiplexed perturbations in primary T cells.

genomics↗

bioRxiv2020

Model-based prediction of spatial gene expression via generative linear mapping

Decoding spatial transcriptomes from single-cell RNA sequencing (scRNA-seq) data has become a fundamental technique for understanding multicellular systems; however, existing computational methods lack both accuracy and biological interpretability due to their model-free frameworks. Here, we introduced Perler, a model-based method to integrate scRNA-seq data with reference in situ hybridization (ISH) data. To calibrate differences between these datasets, we developed a biologically interpretable model that uses generative linear mapping based on a Gaussian-mixture model using the Expectation-Maximization algorithm. Perler accurately predicted the spatial gene expression of Drosophila embryos, zebrafish embryos, mammalian liver, and mouse visual cortex from scRNA-seq data. Furthermore, the reconstructed transcriptomes did not over-fit the ISH data and preserved the timing information of the scRNA-seq data. These results demonstrated the generalizability of Perler for dataset integration, thereby providing a biologically interpretable framework for accurate reconstruction of spatial transcriptomes in any multicellular system.

genomics↗

bioRxiv2020

VIRTUS: a pipeline for comprehensive virus analysis from conventional RNA-seq data

The possibility that RNA transcripts from clinical samples contain plenty of virus RNAs has not been pursued actively so far. We here developed a new tool for analyzing virus-transcribed mRNAs, not virus copy numbers, in the data of conventional and single-cell RNA-sequencing of human cells. Our pipeline, named VIRTUS (VIRal Transcript Usage Sensor), was able to detect 763 viruses including herpesviruses, retroviruses, and even SARS-CoV-2 (COVID-19), and quantify their transcripts in the sequence data. This tool thus enabled simultaneously detecting infected cells, the composition of multiple viruses within the cell, and the endogenous host gene expression profile of the cell. This bioinformatics method would be instrumental in addressing the possible effects of covertly infecting viruses on certain diseases and developing new treatments to target such viruses. Availability and implementationVIRTUS is implemented using Common Workflow Language and Docker under a CC-NC license. VIRTUS is freely available at https://github.com/yyoshiaki/VIRTUS. Supplementary informationSupplementary data are available at Bioinformatics online.

bioinformatics↗

bioRxiv2020

Geographic and subsequent biotic isolations led to a diversity anomaly of Heterotropa (Aristolochiaceae) in insular versus continental regions of the Sino-Japanese Floristic Region

The Sino-Japanese Floristic Region is highly diverse with respect to temperate plants. However, the reasons for this diversity are poorly understood because most studies have only considered geographic isolation caused by climatic oscillations. Heterotropa (genus Asarum; Aristolochiaceae) diverges here and shows high species diversity in insular systems (63 species) compared to continental areas (25 species). Heterotropa shows low dispersal ability with small distribution ranges, implying diversification by geographic events, and high floral diversity, implying pollinator-mediated diversification. To reveal how abiotic and biotic factors have shaped the diversity anomaly of Heterotropa, we conducted phylogenetic analysis using ddRAD-seq and chloroplast genome datasets including 79 species, estimation of floral trait evolution, and comparison of isolation factors within clades based on distribution range and floral trait analysis. Phylogenetic analysis indicates that Heterotropa originated in mainland China and expanded to the Japanese Archipelago in the Miocene, and the major clades almost correspond to geographic distributions. Floral traits evolved repeatedly in the tip nodes within the clades. Although the major clades include a high proportion of species pairs showing isolation by floral traits, there are no conditional relationships between two isolation factors, indicating that most species pairs with floral trait isolation are distributed allopatrically. The repeated exposure and submergence of land-bridges caused by climatic oscillations would have led to significant population fragmentation in insular systems. Thus, the diversity anomaly of Heterotropa would have resulted from geographic and climatic events during the Miocene, while subsequent repeated floral trait evolution would have followed geographic isolation during the Pleistocene.

plant biology↗

bioRxiv2019

Seasonal and annual fluctuations of deer populations estimated by a Bayesian state-space model

Deer overabundance is a contributing factor in the degradation of plant communities and ecosystems worldwide. The management and conservation of the deer-affected ecosystems requires us to urgently grasp deer population trends and to identify the factors that affect them. In this study, we developed a Bayesian state-space model to estimate the population dynamics of sika deer (Cervus nippon) in a cool-temperate forest in Japan, where wolves (Canis lupus hodophilax) are extinct. The model was based on field data collected from block count surveys, road count surveys by vehicles, mortality surveys during the winter, and nuisance control for 12 years (2007-2018). We clarified the seasonal and annual fluctuation of the deer population. We found two peaks of deer abundance (2007 and 2010) over 12 years. In 2011 the estimated deer abundance decreased drastically and has remained at a low level then. The deer population increased from spring to autumn and decreased from autumn to winter in most years. The seasonal fluctuation we detected could reflect the seasonal migration pattern of deer and the population recruitment through fawn births in early summer. In our model, snowfall accumulation, which can be a lethal factor for deer, may have slightly affected their mortality during the winter. Although we could not detect a direct effect of snow on population dynamics, snowfall decrease due to global warming may decelerate the winter migration of deer; subsequently, deer staying on-site may intensively forage evergreen perennial plants during the winter season. The nuisance control affected population dynamics. Even in wildlife protection areas and national parks where hunting is regulated, nuisance control could be effective in buffering the effect of deer browsing on forest ecosystems.

ecology↗

bioRxiv2019

A novel defective recombinant porcine enterovirus G virus carrying a porcine torovirus papain-like cysteine protease gene and a putative anti-apoptosis gene in place of viral structural protein genes

Enterovirus G (EV-G) belongs to the family of Picornaviridae. Two types of recombinant porcine EV-Gs carrying papain-like cysteine protease (PLCP) gene of porcine torovirus, a virus in Coronaviridae, are reported. Type 1 recombinant EV-Gs are detected in pig feces in Japan, USA, and Belgium and carry the PLPC gene at the junction site of 2C/3A genes, while PLPC gene replaces the viral structural genes in type 2 recombinant EV-G detected in pig feces in a Chinese farm. We identified a novel type 2 recombinant EV-G carrying the PLCP gene with flanking sequences in place of the viral structural genes in pig feces in Japan. The ~0.3 kb-long upstream flanking sequence had no sequence homology with any proteins deposited in GenBank, while the downstream ~0.9 kb-long flanking sequence included a domain having high amino acid sequence homology with a baculoviral inhibitor of apoptosis repeat superfamily. The pig feces, where the novel type 2 recombinant EV-G was detected, also carried type 1 recombinant EV-G. Although the phylogenetic analysis suggested that these two recombinant EV-Gs have independently evolved, type 1 recombinant EV-G might have served as a helper virus by providing viral structural proteins for dissemination of the type 2 recombinant EV-G.

microbiology↗

bioRxiv2018

Genetic basis of transgressive segregation in rice heading phenotypes

Transgressive segregation produces hybrid progeny phenotypes that exceed parental phenotypes. Unlike heterosis, extreme phenotypes caused by transgressive segregation are heritably stable. We examined transgressive phenotypes of flowering time in rice. Our previous study examined days to flowering (heading; DTH) in six F2 populations for which the parents had distal DTH, and found very few transgressive phenotypes. Here, we demonstrate that transgressive segregation in F2 populations occurred between parents with proximal DTH. DTH phenotypes of the A58 x Kitaake F2 progenies frequently exceeded those of both parents. Both A58 and Kitaake are japonica rice cultivars adapted to Hokkaido, Japan, which is a high-latitude region, and have short DTH. Among the four known loci required for short DTH, three loci had common alleles in A58 and Kitaake, and only the one locus had different alleles. This result indicates that there is a similar genetic basis for DTH between the two varieties. We identified five new quantitative trait loci (QTLs) associated with transgressive DTH phenotypes by genome-wide single nucleotide polymorphism (SNP) analysis. Each of these QTLs showed different degrees of additive effects on DTH, and two QTLs had epistatic effect on each other. These results demonstrated that genome-wide SNP analysis facilitated detection of genetic loci associated with the extreme phenotypes and revealed that the transgressive phenotypes were produced by exchanging complementary alleles of a few minor QTLs in the similar parental genotypes.

genetics↗