Biology subjects

Kawaguchi, M.

Publications and source records attributed to Kawaguchi, M..

15 recordsLinked to original sources

bioRxiv2026

High CO2 promotes fatty acid-dependent asymbiotic growth of Rhizophagus clarus

Arbuscular mycorrhizal (AM) fungi form symbioses with most terrestrial plants and play an ecologically important role in soil carbon dynamics. However, direct CO2 effects on the fungi remain unclear. We show that CO2-enriched conditions enhance the asymbiotic growth of Rhizophagus clarus in the presence of fatty acids. The response pattern to high CO2 differed between the tested species. These findings provide new insights into the physiological characteristics of AM fungi.

microbiology↗

bioRxiv2026

Lotus japonicus CLV1-Like Receptor HAR1 Promotes Nitrogen Utilization and Growth Under Non-Symbiotic Conditions.

Legumes establish mutualistic symbiosis with nitrogen (N)-fixing bacteria, which allows them to utilize atmospheric N2. Because the maintenance of symbiosis requires abundant carbon (C) sources, legumes regulate the balance between carbon consumption and nitrogen acquisition by systemically controlling the nodule number through CLAVATA1 (CLV1)-like receptors. In Lotus japonicus, the CLV1-like receptor HYPERNODULATION ABERRANT ROOT FORMATION1 (HAR1) acts in shoots to regulate root nodulation and contributes to symbiotic C/N coordination. This raises the possibility that HAR1 may also influence plant growth and nitrogen utilization beyond symbiotic nodulation. In this study, we showed that HAR1 plays a critical role in regulating nitrogen use to enhance growth under conditions of high nitrate availability, even in non-symbiotic environments. Unlike the wild-type, the har1 mutant failed to increase its growth in response to higher nitrate availability. This lack of growth response was associated with a lower rate of net biomass production per unit leaf area and a reduced capacity for biomass production per unit plant nitrogen. We further found that nitrate-responsive TCA cycle-related organic acids were higher in har1 leaves than in wild-type leaves even under low nitrate conditions. Because the HAR1 mutation did not affect photosynthetic traits, we propose that HAR1 promotes growth under non-symbiotic conditions by coordinating nitrogen utilization with primary metabolism. One-sentence summaryLotus japonicus CLV1-like receptor HAR1 improves nitrogen utilization and growth at high nitrate conditions by regulating organic acid metabolism, independent of root nodule symbiosis.

plant biology↗

bioRxiv2026

Early detection of pancreatic ductal adenocarcinoma by single-molecule profiling of pancreatic enzyme activities

Pancreatic ductal adenocarcinoma (PDAC) remains a leading cause of cancer-related mortality, largely due to diagnosis at advanced stages. Early detection through minimally invasive liquid biopsy holds promise for improving patient outcomes. Here, we report a blood-based liquid biopsy platform based on single-molecule enzyme activity profiling (SEAP), which detects proteoform-level alterations in circulating pancreatic enzymes at single-molecule resolution. Using a tissue-centric biomarker discovery strategy, we identified activity signatures of pancreas-specific digestive enzymes associated with PDAC. A combinatorial classifier detected PDAC (stage I-IV) with 95.5% specificity and 75.0% sensitivity (74.2% for stage I-II) across 690 blood samples collected from multiple hospitals and biobanks. Performance was further validated in an independent cohort enriched for early-stage disease, where 54.2% (13/24) of stage IA, 64.3% (9/14) of stage IB, and 21.4% (3/14) of stage 0 lesions were classified as positive. These findings support the clinical potential of SEAP for early detection of PDAC.

biochemistry↗

bioRxiv2025

Cyclophilin A-mediated cis/trans isomerization modulates RIN4 to control intracellular rhizobial infection in legumes

In most legume-rhizobium symbioses, rhizobial colonization occurs through host-derived intracellular infection threads, which enable recruitment of compatible rhizobia while presumably modulating the host immune system to prevent rejection. To investigate how legumes regulate immune responses through post-translational mechanisms during the infection, we focused on Cyclophilin A (CyPA), a peptidyl-prolyl cis/trans isomerase. The model legume Lotus japonicus encodes three canonical CyPA genes. Among them, LjCyPA1 was characterized through CRISPR/Cas9-mediated knockout analysis and shown to be important for normal intracellular infection of compatible rhizobia. A gain-of-function LjCyPA1 variant in a soybean cultivar was able to promote symbiosis with not only compatible but also incompatible rhizobia. Structural modeling followed by genetic analysis demonstrated a functional interaction between LjCyPA1 and the immune hub protein LjRIN4. The cis conformation of LjRIN4 promoted intracellular rhizobial infection, while the trans conformation suppressed it. LjCyPA1 acted with the rhizobial type III secretion system (T3SS) which exhibited a cooperative role between host and symbiont in facilitating infection. Phylogenomic analysis showed that conservation of the CyPA1 orthologue is correlated with the trait of intracellular infection in legumes. Our results contribute to the understanding of how legumes accept symbiotic partners while balancing immune responses.

plant biology↗

bioRxiv2025

Regnase-1 Promotes Tumor-Initiating Activity in Non-Small Cell Lung Cancer

Regnase-1, encoded by the ZC3H12A gene, is a well-known RNase that suppresses inflammation by degrading the mRNAs of inflammatory cytokines. However, its role in cancer pathogenesis, especially in non-small cell lung cancer (NSCLC), remains poorly understood. Through an analysis of public databases, we found that NSCLC patients with higher ZC3H12A expression levels had a worse prognosis than those with lower levels. To explore the function of Regnase-1 in NSCLC, we knocked out the ZC3H12A gene in NSCLC cell lines and compared their transcriptomes with those of parental cells. This analysis identified the SOX2 pathway as a common pathway suppressed by Regnase-1 deficiency. Consistent with the SOX2 contribution to the cancer stemness, Regnase-1 inhibition impaired oncosphere growth and tumor formation of cell lines derived from adenocarcinoma, squamous cell carcinoma and large cell carcinoma. It was also effective for NRF2-activated NSCLC cells, which are highly resistant to most of the therapeutics. Notably, post-tumorigenic suppression of Regnase-1 significantly inhibited tumor growth, suggesting that Regnase-1 could be a promising therapeutic target for post-tumorigenic treatment of NSCLC. Given recent studies describing that Regnase-1 inhibition enhances anti-cancer immunity, we propose that targeting Regnase-1 could be an ideal strategy for controlling intractable cancers by both suppressing cancer cells and activating anti-cancer immunity.

cancer biology↗

bioRxiv2024

Glutathione supersulphide regulates T-cell receptor signalling

Immunometabolism regulates functions and fates of immune cells including T cells. Supersulphides, which are universal metabolites containing catenated sulphur atoms, have various physiological functions based on their unique redox properties. Here we found that activation of T-cell receptor (TCR) signalling was accompanied by supersulphide decrease, which suggests a regulatory contribution of sulphur metabolism to immune function. Consistently, inhibiting supersulphide synthesis facilitated TCR activation and exacerbated allergen-induced type 2 inflammation in mice. Supplementation with glutathione trisulphide (GSSSG), a major endogenous supersulphide, suppressed TCR signalling in naive CD4+ T cells and their differentiation and effectively alleviated the inflammation. Docking simulation revealed interaction of GSSSG with CD3{varepsilon} chain in the TCR/CD3 complex, which was supported by mass spectrometry detection of persulphidated glutathionylation at a functionally important CXXC motif of CD3{varepsilon} chain. This study identified a new post-translational modification with supersulfides and demonstrated a critical contribution of sulphur metabolism to TCR signalling regulation.

biochemistry↗

bioRxiv2024

Effect of Light and Water Agitation on Hatching Processes in Clown Anemonefish Amphiprion ocellaris

Clown anemonefish (Amphiprion ocellaris) exhibit an environmentally cued hatching strategy, wherein parents generate water flow to encourage embryos to hatch after sunset on the eighth day post spawning. Although previous studies have shown that hatching requires complete darkness and water agitation, the regulatory mechanisms underlying these environmental cues remain elusive. This study investigated the expression patterns of hatching enzymes and how darkness and water agitation affect their secretion and the digestion of the egg envelope (chorion) in clown anemonefish. We revealed that hatching enzymes were expressed from the early developmental stages and decreased towards hatching day, similar to other fish species. The hatching gland cells were located on the tail, which may be associated with egg shape and chorion breakage. On the day of hatching, chorion digestion occurred exclusively in complete darkness, regardless of water agitation. In addition, ionomycin, a Ca2+ ionophore and an inducer of hatching enzyme secretion, triggered chorion digestion even under light-on conditions. Furthermore, water agitation expedited chorion digestion in the dark. In conclusion, our findings indicate that darkness triggers the secretion of hatching enzymes mediated by changes in Ca2+ levels, whereas water agitation assists chorion digestion activity and/or hatching enzyme secretion. These responses to environmental cues contribute to the rapid and synchronised hatching in clown anemonefish. Summary statementLight conditions and water agitation regulate the secretion of hatching enzymes and the chorion digestion activity, thereby facilitating a rapid hatching response to environmental changes in clown anemonefish.

physiology↗

bioRxiv2020

Asymbiotic mass production of the arbuscular mycorrhizal fungus Rhizophagus clarus

Arbuscular mycorrhizal (AM) symbiosis is a mutually beneficial interaction between fungi and land plants and promotes global phosphate cycling in terrestrial ecosystems. AM fungi are recognised as obligate symbionts that require root colonisation to complete a life cycle involving the production of propagules, asexual spores. Recently it has been shown that Rhizophagus irregularis can produce infection-competent secondary spores asymbiotically by adding a fatty acid, palmitoleic acid. Further, asymbiotic growth can be supported using myristate as a carbon and energy source for their asymbiotic growth to increase fungal biomass. However, spore production and the ability of these spores to colonise host roots were still limited compared to co-culture of the fungus with plant roots. Here we show that a combination of two plant hormones, strigolactone and methyl jasmonate, induces production of a large number of infection-competent spores in asymbiotic cultures of Rhizophagus clarus HR1 in the presence of myristate and organic nitrogen. Inoculation of asymbiotically-generated spores promoted the growth of Welsh onions, as observed for spores produced by symbiotic culture system. Our findings provide a foundation for elucidation of hormonal control of the fungal life cycle and development of new inoculum production schemes.

microbiology↗

bioRxiv2020

Conservation and host-specific expression of non-tandemly repeated heterogenous ribosome RNA gene in arbuscular mycorrhizal fungi

The ribosomal RNA-encoding gene (rDNA) has a characteristic genomic nature: tens to thousands of copies in a genome, tandemly repeated structure, and intragenomic sequence homogeneity. These features contribute to ribosome productivity via physiological and evolutionary processes. We reported previously the exceptional absence of these features in the model arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis. Here we examine the phylogenetic distribution of the exceptional rDNA features in the genus Rhizophagus via improving the genome sequence of R. clarus. Cross-species comparison indicated similarity of their rDNAs not only in the genomic features but also in the distribution of intragenomic polymorphic sites on the paralogs. Ribosomal RNA comprises multiple domains with different functions. The two Rhizophagus species commonly exhibited a variation enrichment site, ES27L, which is related to translational fidelity and antibiotic sensitivity. Variation enrichment on ES27L has not been observed in other organisms lacking the three rDNA features such as malaria parasites and Cyanidioschyzon merolae. Expression profiling of rDNAs in R. irregularis revealed that rDNA paralogs are expressed differently in association with host plant species. Our results suggest a broad distribution of the disarranged rDNA across AM fungi and its involvement in the successful association with the broad range of host species.

genomics↗

bioRxiv2020

RADSex: a computational workflow to study sex determination using Restriction Site-Associated DNA Sequencing data

The study of sex determination and sex chromosome organisation in non-model species has long been technically challenging, but new sequencing methodologies are now enabling precise and high-throughput identification of sex-specific genomic sequences. In particular, Restriction Site-Associated DNA Sequencing (RAD-Seq) is being extensively applied to explore sex determination systems in many plant and animal species. However, software designed to specifically search for sex-biased markers using RAD-Seq data is lacking. Here, we present RADSex, a computational analysis workflow designed to study the genetic basis of sex determination using RAD-Seq data. RADSex is simple to use, requires few computational resources, makes no prior assumptions about type of sex-determination system or structure of the sex locus, and offers convenient visualization through a dedicated R package. To demonstrate the functionality of RADSex, we re-analyzed a published dataset of Japanese medaka, Oryzias latipes, where we uncovered a previously unknown Y chromosome polymorphism. We then used RADSex to analyze new RAD-Seq datasets from 15 fish species spanning multiple systematic orders. We identified the sex determination system and sex-specific markers in six of these species, five of which had no known sex-markers prior to this study. We show that RADSex greatly facilitates the study of sex determination systems in non-model species and outperforms the commonly used RAD-Seq analysis software STACKS. RADSex in speed, resource usage, ease of application, and visualization options. Furthermore, our analysis of new datasets from 15 species provides new insights on sex determination in fish.

bioinformatics↗

bioRxiv2020

Virtual-freezing fluorescence imaging flow cytometry

By virtue of the combined merits of flow cytometry and fluorescence microscopy, imaging flow cytometry (IFC) has become an established tool for cell analysis in diverse biomedical fields such as cancer biology, microbiology, immunology, hematology, and stem cell biology. However, the performance and utility of IFC are severely limited by the fundamental trade-off between throughput, sensitivity, and spatial resolution. For example, at high flow speed (i.e., high throughput), the integration time of the image sensor becomes short, resulting in reduced sensitivity or pixel resolution. Here we present an optomechanical imaging method that overcomes the trade-off by virtually "freezing" the motion of flowing cells on the image sensor to effectively achieve 1,000 times longer exposure time for microscopy-grade fluorescence image acquisition. Consequently, it enables high-throughput IFC of single cells at >10,000 cells/s without sacrificing sensitivity and spatial resolution. The availability of numerous information-rich fluorescence cell images allows high-dimensional statistical analysis and accurate classification with deep learning, as evidenced by our demonstration of unique applications in hematology and microbiology.

cell biology↗

bioRxiv2019

Myristate as a carbon and energy source for the asymbiotic growth of the arbuscular mycorrhizal fungus Rhizophagus irregularis

Arbuscular mycorrhizal (AM) fungi, forming symbiotic associations with land plants, are obligate symbionts that cannot complete their natural life cycle without a host. Recently, fatty acid auxotrophy of AM fungi is supported by studies showing that lipids synthesized by the host plants are transferred to the fungi and that the latter lack genes encoding cytosolic fatty acid synthases (1-7). Therefore, to establish an asymbiotic cultivation system for AM fungi, we tried to identify the fatty acids that could promote biomass production. To determine whether AM fungi can grow on medium supplied with fatty acids or lipids under asymbiotic conditions, we tested eight saturated or unsaturated fatty acids (C12-C18) and two {beta}-monoacylglycerols. Only myristate (C14:0) led to an increase in biomass of Rhizophagus irregularis, inducing extensive hyphal growth and formation of infection-competent secondary spores. However, such spores were smaller than those generated symbiotically. Furthermore, we demonstrated that R. irregularis can take up fatty acids in its branched hyphae and use myristate as a carbon and energy source. Myristate also promoted the growth of Rhizophagus clarus and Gigaspora margarita. Finally, mixtures of myristate and palmitate accelerated fungal growth and induced a substantial change in fatty acid composition of triacylglycerol compared with single myristate application, although palmitate was not used as a carbon source for cell wall biosynthesis in this culture system. In conclusion, here we demonstrate that myristate boosts asymbiotic growth of AM fungi and can also serve as a carbon and energy source. Significance statementThe origins of arbuscular mycorrhizal (AM) fungi, which form symbiotic associations with land plants, date back over 460 million years ago. During evolution, these fungi acquired an obligate symbiotic lifestyle, and thus depend on their host for essential nutrients. In particular, fatty acids are regarded as crucial nutrients for the survival of AM fungi owing to the absence of genes involved in de novo fatty acid biosynthesis in the AM fungal genomes that have been sequenced so far. Here, we show that myristate initiates AM fungal growth under asymbiotic conditions. These findings will advance pure culture of AM fungi.

microbiology↗

bioRxiv2019

Genome sequence of the euryhaline Javafish medaka, Oryzias javanicus: a small aquarium fish model for studies on adaptation to salinity.

BackgroundThe genus Oryzias is constituted of 35 medaka-fish species each exhibiting various ecological, morphological and physiological peculiarities and adaptations. Beyond of being a comprehensive phylogenetic group for studying intra-genus evolution of several traits like sex determination, behaviour, morphology or adaptation through comparative genomic approaches, all medaka species share many advantages of experimental model organisms including small size and short generation time, transparent embryos and genome editing tools for reverse and forward genetic studies. The Java medaka, Oryzias javanicus, is one of the two species of medaka perfectly adapted for living in brackish/sea-waters. Being an important component of the mangrove ecosystem, O. javanicus is also used as a valuable marine test-fish for ecotoxicology studies. Here, we sequenced and assembled the whole genome of O. javanicus, and anticipate this resource will be catalytic for a wide range of comparative genomic, phylogenetic and functional studies.\n\nFindingsComplementary sequencing approaches including long-read technology and data integration with a genetic map allowed the final assembly of 908 Mbp of the O. javanicus genome. Further analyses estimate that the O. javanicus genome contains 33% of repeat sequences and has a heterozygosity of 0.96%. The achieved draft assembly contains 525 scaffolds with a total length of 809.7 Mbp, a N50 of 6.3 Mbp and a L50 of 37 scaffolds. We identified 21454 expressed transcripts for a total transcriptome size of 57, 146, 583 bps.\n\nConclusionsWe provide here a high-quality draft genome assembly of the euryhaline Javafish medaka, and give emphasis on the evolutionary adaptation to salinity.

genomics↗

bioRxiv2018

Cell type-specific regulation of nutrient absorption, metabolism and transfer in arbuscular mycorrhizal fungi

Arbuscular mycorrhizal fungi (AMF) establish symbiotic relationships with most land plants, mainly for the purpose of nutrient exchange. Many studies have revealed the regulation of absorption, metabolism, and transfer of nutrients in AMF and the genes involved in these processes. However, the spatial regulation of the genes among the structures comprising each developmental stage are not well understood. Here, we demonstrate the structure-specific transcriptome of the model AMF species, Rhizophagus irregularis. We performed an ultra-low input RNA-seq analysis, SMART-seq2, comparing five extraradical structures, germ tubes, runner hyphae, branched absorbing structures, immature spores, and mature spores. In addition, we reanalyzed the recently reported RNA-seq data comparing intraradical hyphae and arbuscules. Our analyses captured the distinct features of each structure and revealed the structure-specific expression patterns of genes related to absorption, metabolism, and transfer of nutrients. Of note, the transcriptional profiles suggest the distinct functions of branched absorbing structures in nutrient absorption. These findings provide a comprehensive dataset to advance our understanding of the transcriptional dynamics of fungal nutrition in this symbiotic system.

plant biology↗

bioRxiv2017

Evidence of non-tandemly repeated rDNAs and their intragenomic heterogeneity in Rhizophagus irregularis

Arbuscular mycorrhizal fungus (AMF) species are one of the most widespread symbionts of land plants. Our substantially improved reference genome assembly of a model AMF, Rhizophagus irregularis DAOM-181602 (total contigs = 210), facilitated discovery of repetitive elements with unusual characteristics. R. irregularis has only ten or eleven copies of complete 45S rDNAs, whereas the general eukaryotic genome has tens to thousands of rDNA copies. R. irregularis rDNAs are highly heterogeneous and lack a tandem repeat structure. These findings provide evidence for the hypothesis that rDNA heterogeneity depends on the lack of tandem repeat structures. RNA-Seq analysis confirmed that all rDNA variants are actively transcribed. Observed rDNA/rRNA polymorphisms may modulate translation by using different ribosomes depending on biotic and abiotic interactions. The non-tandem repeat structure and intragenomic heterogeneity of AMF rDNA/rRNA may facilitate adaptation to a various environmental condition including the broad host range.

genomics↗