Biology subjects

Tokunaga, K.

Publications and source records attributed to Tokunaga, K..

At least 19 recordsLinked to original sources

bioRxiv2026

Large-scale meta-analysis of over one million individuals reveals the genetic architecture of 127 complex traits in East Asian populations

Genome-wide association studies (GWASs) have disproportionately focused on European (EUR) populations, limiting the characterization of genetic architecture in other ancestries. To address this imbalance, we integrated large-scale biobanks from Japan, Korea, Taiwan, and China to perform the largest phenome-wide meta-analysis to date in East Asian (EAS) populations, encompassing over one million individuals across 127 complex traits. We identified 8,010 previously unreported associations and observed substantial genetic sharing across EAS subpopulations, while also detecting cohort-specific heterogeneity within the broader EAS context. Transethnic analyses revealed moderate genetic correlations between EAS and EUR populations, indicating both shared and ancestry-specific components of disease risk. Pleiotropy analyses highlighted prominent signals within the HLA region, supported by protein-protein interaction connectivity and immune-related pathway enrichment. Decomposition of genome-wide association matrices further uncovered structured cross-trait architectures, revealing a predominantly shared polygenic backbone driven by metabolic, biochemical, and anthropometric traits, together with two discrete latent components enriched for immune-related processes. Together, our findings refine the genetic architecture of complex traits in East Asian populations at unprecedented scale and clarify the balance between shared and population-specific determinants of human diseases.

genetics↗

bioRxiv2026

Selective activation of LH-dependent transcriptional pathways determines ovulatory follicles in the hierarchical ovary of cloudy catshark

The number of offspring produced per reproductive cycle is generally explained by a luteinizing hormone receptor (LHR)-threshold model, in which only follicles expressing sufficient levels of LHR respond to the LH surge and proceed to ovulation. Using the cloudy catshark as a model, we reveal a distinct mechanism that determines the fate of ovulatory (F1) and non-ovulatory (F2) follicles independently of LHR abundance. The cloudy catshark produces only two eggs per reproductive cycle by ovulating a pair of F1 follicles from its single ovary. Both F1 and F2 follicles express comparable levels of lhr, indicating that both retain the capacity to receive the LH signal. Nevertheless, LH stimulation selectively activated ovulation-associated responses in F1 follicles. These included progesterone production via star2 upregulation, as well as transcriptional programs including RUNX transcription factors, prostaglandin signaling (ptgs2, ptger1), and matrix metalloproteinases. Moreover, direct activation of cAMP pathway, bypassing LHR activation, induced progesterone production in F1 but not F2 follicles, indicating that the critical difference in ovulatory responsiveness lies downstream of cAMP. These results demonstrate that follicular ovulatory fate is determined, at least in part, by follicle-specific transcriptional responses to cAMP, rather than by LHR abundance alone, challenging the prevailing LHR-threshold model of ovulatory regulation in vertebrates.

physiology↗

bioRxiv2025

Passive and active demethylation in immune diseases with proliferation driving global hypomethylation

Global DNA hypomethylation is a hallmark of immune-mediated diseases, yet its regulatory significance remains unclear. Replication-associated loss of DNA methylation at solo-WCGW (W = A/T) CpGs has recently been proposed as a consequence of cell division. Here we systematically investigated genome-wide hypomethylation patterns across seven immune-mediated diseases. Most diseases exhibited global hypomethylation, particularly at solo-WCGW CpGs in transcriptionally repressed regions, potentially reflecting increased immune cell proliferation. By contrast, CpG sites whose methylation levels were associated with cytokine exposure or SNP genotypes were predominantly located in transcriptionally active regions. To investigate whether immunological events driving immune cell proliferation may also be imprinted in transcriptionally active regions, we searched for differentially methylated regions (DMRs) correlated with an index reflecting the extent of solo-WCGW hypomethylation. In narcolepsy, we identified a DMR within the T-cell receptor alpha (TRA) locus, and greater hypomethylation was associated with increased clonality of both TRA and TRB repertoires, with a similar pattern in an independent cohort. In multiple sclerosis, a DMR was also detected within the IGH locus encoding the B-cell receptor. Together, these findings suggest that the hypomethylation index captures the impact of disease-specific immune dynamics, while reflecting a shared epigenetic signature of immune cell proliferation across diseases.

genomics↗

bioRxiv2024

Phased genome assemblies and pangenome graphs of human populations of Japan and Saudi Arabia

The selection of a reference sequence in genome analysis is critical, as it serves as the foundation for all downstream analyses. Recently, the pangenome graph has been proposed as a data model that incorporates haplotypes from multiple individuals. Here we present JaSaPaGe, a pangenome graph reference for Saudi Arabian and Japanese populations, both of which have been significantly underrepresented in previous genomic studies. We constructed JaSaPaGe from high-quality phased diploid assemblies which were made utilizing PacBio high-fidelity long reads, Nanopore long reads, and Hi-C short reads of 9 Saudi and 10 Japanese individuals. Quality evaluation of the pangenome graph by variant calling showed that our pangenome outperformed earlier linear reference genomes (GRCh38 and T2T-CHM13) and showed comparable performance to the pangenome graph provided by the Human Pangenome Reference Consortium (HPRC), with more variants found in Japanese and Saudi samples using their population-specific pangenomes. This pangenome reference will serve as a valuable resource for both the research and clinical communities in Japan and Saudi Arabia.

genomics↗

bioRxiv2024

Multiomics analysis of narcolepsy T cells: global hypomethylation in solo-WCGW motif linked to T cell proliferation

Narcolepsy type 1 (NT1) is a chronic sleep disorder caused by a loss of orexin-producing cells in the brain and involves autoimmune mechanisms, including the presence of autoreactive T cells. We performed a genome-wide DNA methylation analysis using CD4+ and CD8+ T cells of NT1 patients. Analysis of differentially methylated regions as well as multiomics analysis with genomic and transcriptomic data obtained from the same samples indicated that cell chemotaxis pathways are implicated as a cause in the pathogenesis of NT1. Additionally, we found global hypomethylation in both the T cells of NT1 cases (CD4+: P = 1.69E-67; CD8+: P = 4.83E-12). These NT1-associated hypomethylated sites were significantly more abundant in solo-WCGW (sequences without neighboring CpGs, where W is an A or T base; P = 9.87E-194). Solo-WCGW tends to lose DNA methylation over the course of cell divisions, suggesting enhanced T cell proliferation in NT1.

genomics↗

bioRxiv2024

Enhancing Prediction of Human Traits and Behaviors through Ensemble Learning of Traditional and Novel Resting-State fMRI Connectivity Analyses

Recent efforts in cognitive neuroscience have focused on leveraging resting-state functional connectivity (RSFC) data from fMRI to predict human traits and behaviors more accurately. Traditional methods typically analyze RSFC by correlating averaged time-series data between regions of interest (ROIs) or networks, a process that may overlook critical spatial signal patterns. To address this limitation, we introduced a novel linear regression technique that estimates RSFC by predicting spatial brain activity patterns in a target ROI from those in a seed ROI. We applied both traditional and our novel RSFC estimation methods to a large-scale dataset from the Human Connectome Project, analyzing resting-state fMRI data to predict sex, age, personality traits, and psychological task performance. Additionally, we developed an ensemble learner that integrates these methods using a weighted average approach to enhance prediction accuracy. Our findings revealed that hierarchical clustering of RSFC patterns using our novel method displays distinct whole-brain grouping patterns compared to the traditional approach. Importantly, the ensemble model outperformed the traditional RSFC method in predicting human traits and behaviors. Notably, the predictions from the traditional and novel methods showed relatively low similarity, indicating that our novel approach captures unique and previously undetected information about human traits and behaviors through fine-grained local spatial patterns of neural activation. These results highlight the potential of combining traditional and innovative RSFC analysis techniques to enrich our understanding of the neural basis of human traits and behaviors.

neuroscience↗

bioRxiv2024

Low microbial abundance and community diversity within the egg capsule of the oviparous cloudy catshark (Scyliorhinus torazame) during oviposition

Vertebrate embryos are protected from bacterial infection by various maternally derived immune factors before the embryonic organs are fully developed. However, the defense mechanisms employed by elasmobranch embryos during development remain poorly understood. This study attempted to elucidate the embryonic defense mechanism of elasmobranchs by investigating the intracapsular environment of freshly laid eggs of the oviparous cloudy catshark (Scyliorhinus torazame). The egg capsule of oviparous elasmobranchs is tightly sealed until pre-hatching (early opening of the egg capsule), after which seawater flows into the capsule and the embryos are consequently exposed to the surrounding seawater. We first experimentally examined the resistance of embryos to potential bacterial infections and found that the early embryos were highly vulnerable to environmental pathogens, suggesting that the embryos are protected from the threat of infection before pre-hatching. Indeed, the intracapsular environment of freshly laid eggs exhibited a significantly low bacterial density that was maintained until pre-hatching. Furthermore, the microbiome inside eggs just after oviposition differed markedly from the microbiomes of rearing seawater and adult oviducal gland epithelia; these eggs were predominantly populated by an unidentified genus of Sphingomonadaceae. Overall, this study provides compelling evidence that early embryos of oviparous cloudy catshark are incubated in a clean intracapsular environment that potentially plays a significant role in embryonic development in oviparous elasmobranchs. Our results suggest that maintenance of this clean condition might be attributable to bactericidal or bacteriostatic activities associated with the egg jelly and/or oviducal gland.

microbiology↗

bioRxiv2023

Virological characteristics correlating with SARS-CoV-2 spike protein fusogenicity

The severe acute respiratory syndrome coronavirus (SARS-CoV-2) spike (S) protein is essential in mediating membrane fusion of the virus with the target cells. Several reports demonstrated that SARS-CoV-2 S protein fusogenicity is reportedly closely associated with the intrinsic pathogenicity of the virus determined using hamster models. However, the association between S protein fusogenicity and other virological parameters remains elusive. In this study, we investigated the virological parameters of eleven previous variants of concern (VOCs) and variants of interest (VOIs) correlating with S protein fusogenicity. S protein fusogenicity was found to be strongly correlated with S1/S2 cleavage efficiency and plaque size formed by clinical isolates. However, S protein fusogenicity was less associated with pseudoviral infectivity, pseudovirus entry efficiency, and viral replication kinetics. Taken together, our results suggest that S1/S2 cleavage efficiency and plaque size could be potential indicators to predict the intrinsic pathogenicity of newly emerged SARS-CoV-2 variants.

microbiology↗

bioRxiv2023

Identification of Region-Specific Gene Isoforms in the Human Brain Using Long-Read Transcriptome Sequencing and Their Correlation with DNA Methylation

BackgroundSite specificity is known in neuropsychiatric disorders, and differences in gene expression patterns could potentially explain this mechanism. However, studies using long-read transcriptome sequencing to analyze gene expression in different regions of the human brain have been limited, and none have focused on the hypothalamus, which plays a crucial role in regulating autonomic functions. ResultsWe performed long-read RNA sequencing on 12 samples derived from three different brain regions of the same individuals; the cerebellum, hypothalamus, and temporal cortex. We found that, compared to other regions, many genes with higher expression levels in the cerebellum and temporal cortex were associated with neuronal pathways, whereas those with higher expression levels in the hypothalamus were primarily linked to immune pathways. In addition, we investigated genes with different major isoforms in each brain region, even with similar overall expression levels among regions, and identified several genes, such as GAS7, that express different major isoforms in different regions. Many of these genes are involved in "actin filament-based process" and "cell projection organization" pathways, suggesting that region-dependent isoforms may have distinct roles in dendritic spine and neuronal formation in each region. Furthermore, we investigated the involvement of DNA methylation in these isoforms and found that DNA methylation may be associated with isoforms that have different first exons. ConclusionsOur results provide potentially valuable findings for future research on brain disorders and shed light on the mechanisms underlying isoform diversity in the human brain.

genomics↗

bioRxiv2023

Determination of the factors responsible for host tropism of SARS-CoV-2-related bat coronaviruses

Differences in host ACE2 genes may affect the host range of SARS-CoV-2-related coronaviruses (SC2r-CoVs) and further determine the tropism of host ACE2 for the infection receptor. However, the factor(s) responsible for determining the host tropism of SC2r-CoVs, which may in part be determined by the tropism of host ACE2 usage, remains unclear. Here, we use the pseudoviruses with the spike proteins of two Laotian SC2r-CoVs, BANAL-20-236 and BANAL-20-52, and the cells expressing ACE2 proteins of eight different Rhinolophus bat species, and show that these two spikes have different tropisms for Rhinolophus bat ACE2. Through structural analysis and cell culture experiments, we demonstrate that this tropism is determined by residue 493 of the spike and residues 31 and 35 of ACE2. Our results suggest that SC2r-CoVs exhibit differential ACE2 tropism, which may be driven by adaptation to different Rhinolophus bat ACE2 proteins.

microbiology↗

bioRxiv2023

Exploring the genetic diversity of the Japanese Population: Insights from a Large-Scale Whole Genome Sequencing Analysis

The Japanese archipelago is a terminal location for human migration, and the contemporary Japanese people represent a unique population whose genomic diversity has been shaped by multiple migrations from Eurasia. Through high-coverage whole-genome sequencing (WGS) analysis of 9,850 samples from the National Center Biobank Network, we analyzed the genomic characteristics that define the genetic makeup of the modern Japanese population from a population genetics perspective. The dataset comprised populations from the Ryukyu Islands and other parts of the Japanese archipelago (Hondo). Low frequency detrimental or pathogenic variants were found in these populations. The Hondo population underwent two episodes of population decline during the Jomon period, corresponding to the Late Neolithic, and the Edo period, corresponding to the Early Modern era, while the Ryukyu population experienced a population decline during the shell midden period of the Late Neolithic in this region. Genes related to alcohol and lipid metabolism were affected by positive natural selection. Two genes related to alcohol metabolism were found to be 12,500 years out of phase with the time when they began to be affected by positive natural selection; this finding indicates that the genomic diversity of Japanese people has been shaped by events closely related to agriculture and food production. Author summaryThe human population in the Japanese archipelago exhibits significant genetic diversity, with the Ryukyu Islands and other parts of the archipelago (Hondo) having undergone distinct evolutionary paths that have contributed to the genetic divergence of the populations in each region. In this study, whole genome sequencing of healthy individuals from national research hospital biobanks was utilized to investigate the genetic diversity of the Japanese population. Haplotypes were inferred from the genomic data, and a thorough population genetic analysis was conducted. The results indicated not only genetic differentiation between Hondo and the Ryukyu Islands, but also marked differences in past population size. In addition, gene genealogies were inferred from the haplotypes, and the patterns were scrutinized for evidence of natural selection. This analysis revealed unique traces of natural selection in East Asian populations, many of which were believed to be linked to dietary changes brought about by agriculture and food production.

genomics↗

bioRxiv2022

Dissecting the cell of origin of aberrant SALL4 expression in myelodysplastic syndrome

Myelodysplastic syndrome (MDS) is a group of heterogeneous diseases characterized by cytologic dysplasia and cytopenias resulting from ineffective hematopoiesis. Oncofetal protein SALL4 is a known oncogene in MDS and its baseline expression level serves as a prognostic biomarker for MDS at the time of diagnosis. In addition, a recent study showed that SALL4 upregulation following hypomethylating agent treatment in MDS patients correlates with poor outcomes. Despite its important mechanistic and diagnostic significance, the cellular identity of bone marrow cells with aberrant SALL4 expression in MDS patients remains unknown. In this study, we analyzed MDS bone marrow cells on single cell level by mass cytometry (CyTOF) and found that SALL4 was mainly aberrantly expressed in the hematopoietic stem and progenitor cells (HSPC) as well as myeloid lineages. Within the HSPC population from MDS patients, SALL4 and p53 were co-expressed, with the highest co-expressing clones harboring pathogenic TP53 mutations. Overall, our study characterizes for the first time the aberrant SALL4 expression in primary MDS patient samples at a single-cell level. Further studies on the SALL4/p53 network for in-depth mechanistic investigation are needed in the future. Key PointsSALL4 expression in various MDS BM cells confirmed by mass cytometry (CyTOF). SALL4 and p53 double positive cells were predominantly found in the hematopoietic stem and progenitor cell (HSPC) population and associated with pathogenic TP53 mutation status.

cell biology↗

bioRxiv2021

SARS-CoV-2 Lambda variant exhibits higher infectivity and immune resistance

SARS-CoV-2 Lambda, a new variant of interest, is now spreading in some South American countries; however, its virological features and evolutionary trait remain unknown. Here we reveal that the spike protein of the Lambda variant is more infectious and it is attributed to the T76I and L452Q mutations. The RSYLTPGD246-253N mutation, a unique 7-amino-acid deletion mutation in the N-terminal domain of the Lambda spike protein, is responsible for evasion from neutralizing antibodies. Since the Lambda variant has dominantly spread according to the increasing frequency of the isolates harboring the RSYLTPGD246-253N mutation, our data suggest that the insertion of the RSYLTPGD246-253N mutation is closely associated with the massive infection spread of the Lambda variant in South America. HighlightsO_LILambda S is highly infectious and T76I and L452Q are responsible for this property C_LIO_LILambda S is more susceptible to an infection-enhancing antibody C_LIO_LIRSYLTPGD246-253N, L452Q and F490S confer resistance to antiviral immunity C_LI Graphical Abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

microbiology↗

bioRxiv2021

SARS-CoV-2 spike P681R mutation enhances and accelerates viral fusion

During the current SARS-CoV-2 pandemic, a variety of mutations have been accumulated in the viral genome, and currently, four variants of concerns (VOCs) are considered as the hazardous SARS-CoV-2 variants to the human society1. The newly emerging VOC, the B.1.617.2/Delta variant, closely associates with a huge COVID-19 surge in India in Spring 20212. However, its virological property remains unclear. Here, we show that the B.1.617.2/Delta variant is highly fusogenic, and notably, more pathogenic than prototypic SARS-CoV-2 in infected hamsters. The P681R mutation in the spike protein, which is highly conserved in this lineage, facilitates the spike protein cleavage and enhances viral fusogenicity. Moreover, we demonstrate that the P681R-bearing virus exhibits higher pathogenicity than the parental virus. Our data suggest that the P681R mutation is a hallmark that characterizes the virological phenotype of the B.1.617.2/Delta variant and is closely associated with enhanced pathogenicity.

microbiology↗

bioRxiv2021

MARCH8 targets cytoplasmic lysine residues of various viral envelope glycoproteins

The host transmembrane protein MARCH8 is a RING finger E3 ubiquitin ligase that downregulates various host transmembrane proteins, such as MHC-II. We have recently reported that MARCH8 expression in virus-producing cells impairs viral infectivity by reducing virion incorporation of not only HIV-1 envelope glycoproteins but also vesicular stomatitis virus G-glycoprotein through two different pathways. However, the MARCH8 inhibition spectrum remains largely unknown. Here, we investigate the antiviral spectrum of MARCH8 using HIV-1 pseudotyped with a variety of viral envelope glycoproteins. Pseudotyping experiments revealed that viral envelopes derived from the rhabdovirus, arenavirus, coronavirus, and togavirus (alphavirus) families were sensitive to MARCH8-mediated inhibition. Lysine mutations at the cytoplasmic tails of rabies virus-G, lymphocytic choriomeningitis virus glycoproteins, SARS-CoV and SARS-CoV-2 spike proteins, and Chikungunya virus and Ross River virus E2 proteins conferred resistance to MARCH8. Immunofluorescence showed impaired downregulation of the mutants of these viral envelopes by MARCH8, followed by lysosomal degradation, suggesting that MARCH8-mediated ubiquitination leads to intracellular degradation of these envelopes. Indeed, rabies virus-G and Chikungunya virus E2 proteins proved to be clearly ubiquitinated. We conclude that MARCH8 has inhibitory activity on a variety of viral envelope glycoproteins whose cytoplasmic lysine residues are targeted by this antiviral factor.

microbiology↗

bioRxiv2020

Naturally mutated spike proteins of SARS-CoV-2 variants show differential levels of cell entry

The causative agent of the coronavirus disease 2019 (COVID-19) pandemic, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is steadily mutating during continuous transmission among humans. Such mutations can occur in the spike (S) protein that binds to the angiotensin-converting enzyme-2 (ACE2) receptor and is cleaved by transmembrane protease serine 2 (TMPRSS2). However, whether S mutations affect SARS-CoV-2 infectivity remains unknown. Here, we show that naturally occurring S mutations can reduce or enhance cell entry via ACE2 and TMPRSS2. A SARS-CoV-2 S-pseudotyped lentivirus exhibits substantially lower entry than SARS-CoV S. Among S variants, the D614G mutant shows the highest cell entry, as supported by structural observations. Nevertheless, the D614G mutant remains susceptible to neutralization by antisera against prototypic viruses. Taken together, these data indicate that the D614G mutation enhances viral infectivity while maintaining neutralization susceptibility.

microbiology↗

bioRxiv2020

Super-rapid quantitation of the production of HIV-1 harboring a luminescent peptide tag

In virological studies using HIV-1 proviral clones, virus production is normally monitored by either an RT assay or a p24 antigen capture ELISA. However, these assays are costly and time-consuming for routine handling of a large number of HIV-1 samples. In addition, sample dilution is always required in ELISA to determine p24 protein levels because of the very narrow range of detectable concentrations in this assay. Here, we establish a novel HIV-1 production assay system to solve the aforementioned problems by using a recently developed small peptide tag called HiBiT. First, we constructed a novel full-length proviral HIV-1 DNA clone and a lentiviral packaging vector in which the HiBiT tag was added to the C terminus of the integrase. Tagging the integrase with the HiBiT sequence did not impede the production or infectivity of the resultant viruses. Electron microscopy revealed normal morphology of the virus particles. Most importantly, by comparing between ELISA and the HiBiT luciferase assay, we successfully obtained an excellent linear correlation between p24 concentrations and HiBiT-based luciferase activity. Overall, we conclude that HiBiT-tagged viruses can replace the parental HIV-1 and lentiviral vectors, which enables us to perform a super-rapid, inexpensive, convenient, simple, and highly accurate quantitative assay for HIV-1 production.

microbiology↗

bioRxiv2020

MARCH8 inhibits viral infection by two different mechanisms

Membrane-associated RING-CH 8 (MARCH8) inhibits infection with both HIV-1 and vesicular stomatitis virus G-glycoprotein (VSV-G)-pseudotyped viruses by reducing virion incorporation of envelope glycoproteins. The molecular mechanisms by which MARCH8 targets envelope glycoproteins remain unknown. Here, we show two different mechanisms by which MARCH8 inhibits viral infection. Viruses pseudotyped with the VSV-G mutant, in which cytoplasmic lysine residues were mutated, were insensitive to the inhibitory effect of MARCH8, whereas those with a similar lysine mutant of HIV-1 Env remained sensitive to it. Indeed, the wild-type VSV-G, but not its lysine mutant, was ubiquitinated by MARCH8. Furthermore, the MARCH8 mutant, which had a disrupted cytoplasmic tyrosine motif that is critical for intracellular protein sorting, did not inhibit HIV-1 Env-mediated infection, while it still impaired infection by VSV-G-pseudotyped viruses. Overall, we conclude that MARCH8 reduces viral infectivity by downregulating envelope glycoproteins through two different mechanisms mediated by a ubiquitination-dependent or tyrosine motif-dependent pathway.

microbiology↗