Biology subjects

Fukuda, A.

Publications and source records attributed to Fukuda, A..

14 recordsLinked to original sources

bioRxiv2026

Rice Cultivars Carrying the Semi Dwarfing Allele Enables High Yield without Lodging under Hairy Vetch based Green Manure

Green manure is a promising strategy for reducing dependence on chemical fertilizers in crop production. However, vigorous growth due to green manure often leads to high yield accompanied by lodging in rice, hindering its practical use in rice cultivation. Here, we show that rice cultivars carrying a semi-dwarfing sd1/ga20ox2 allele achieve high grain yield without lodging under hairy vetch-based green manure conditions. The leading Japanese cultivar Koshihikari exhibited enhanced vegetative growth, increased panicle number, and consequently higher grain yield and quality under green manure conditions in 2023 and 2024 compared with chemical fertilizer management, although this was accompanied by increased culm length and widespread lodging. Among the four GA20-oxidase genes, green manure significantly upregulated Sd1/GA20ox2 mRNA levels. A temperate japonica cultivar, Nijinokirameki, and an indica cultivar, Hokuriku-193, carrying a non-functional sd1/ga20ox2 allele exhibited no lodging under hairy vetch-based green manure management while achieving improved yield performance. Notably, yields obtained under our hairy vetch-based cultivation system were comparable to or exceeded a recently reported high-yield benchmark observed for Hokuriku-193 under chemical fertilizer management in the same region of Japan. These findings suggest that cultivars harboring non-functional sd1/ga20ox2 alleles enable the practical implementation of annual hairy-vetch-rice rotation for sustainable rice production.

plant biology↗

bioRxiv2026

Epigenetic Control of Spatiotemporal Dynamics of Pancreatic Cancer Cells via Brg1-Rac1 Signaling

Pancreatic ductal adenocarcinoma (PDA) arises from distinct precursor lesions with different clinical outcomes, yet the mechanisms linking epigenetic regulation to invasive cell behavior remain poorly understood. Here, we investigate how the chromatin remodeler Brg1 influences the dynamic properties of cancer cell migration. Using a biomimetic supported membrane system combined with label-free interferometric imaging, we quantitatively analyze the spatiotemporal dynamics of PDA cells derived from pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasms (IPMN). Despite their similar morphology under conventional conditions, PanIN- and IPMN-derived PDA cells exhibit markedly different migration behaviors. PanIN-derived cells migrate faster and display enhanced dynamic remodeling, whereas IPMN-derived cells show persistent elongation with limited displacement. These differences are captured by quantitative analyses of cell trajectories and deformation dynamics. Mechanistically, PanIN-derived PDA cells exhibit elevated Rac1 activity, supporting a model in which a Brg1-Rac1 axis regulates cytoskeletal dynamics and migration behavior. Together, our findings demonstrate that epigenetic regulation is linked to distinct dynamic phenotypes of cancer cells and highlight the importance of quantitative analysis of cell behavior for understanding invasive potential.

biophysics↗

bioRxiv2026

Spatiotemporal Patterns of Active Deformation Reveal Downregulation of Cell-Cell Adhesion in Patient-Derived Colorectal Cancer Organoids with BRAF Mutation

Colorectal cancer (CRC) is the second most common cause of cancer-related mortality. At the molecular level, CRC is associated with genetic mutations and epigenetic modifications that dysregulate various signaling networks. From the biophysical viewpoint, invasive and metastatic cell migration need to be empowered by mechanical forces. In this study, we analyze the dynamic deformation of patient-derived CRC organoids in Fourier space and demonstrate how organoids with protooncogene BRAF mutation exhibit deformation phenotypes at an early stage. The organoids with BRAFmut have significantly lower elasticity and higher viscosity than those with BRAFWT, which mathematically indicated as the weakening of cell-cell adhesion. Immunohistochemical images, qRT-PCR, and TCGA data analysis confirm the downregulation of E-cadherin (CDH1) in BRAFmut organoids as well as in BRAFmut CRC, suggesting that the decrease in cell-cell adhesion in BRAFmut CRC facilitates invasive and metastatic migration. Notably, the recovery of CDH1 expression by pharmacological inhibition of DNA methylation can quantitatively be detected as the change in mechanical properties, suggesting that the complementary combination of dynamic phenotyping, mathematical modelling, and molecular-level analyses has a potential to unravel the mechanistic causality of the critical gene mutation and CRCs prognosis and the response to therapeutic interventions.

biophysics↗

bioRxiv2026

THERM-D Uncovers Distinct Neural Mechanisms Separating Morning and Evening Body Temperature Rhythms in Drosophila

Animal body temperature rises throughout the day and peaks in the evening, a pattern conserved across diurnal endotherms and ectotherms. However, the mechanisms driving the robust body temperature rhythms (BTR) remain largely unclear. Here, we developed a machine learning-based platform, temperature homeostasis evaluation of rhythmicity in model Drosophila (THERM-D), enabling continuous, high-throughput analyses of BTR. Using THERM-D, we identified robust BTR patterns reflecting flies morning and evening behaviors and revealed the function of CRYPTOCHROME (CRY)-negative clock neurons. About half of all clock neurons lack CRY, yet their function was unclear. Newly developed Gal4 drivers targeting CRY-negative neurons demonstrated that these neurons control the morning temperature rise without affecting evening BTR. The data suggests that separate clock circuits regulate morning and evening BTR. Thus, THERM-D elucidated the role of CRY-negative clock neurons, which are specialized for BTR regulation and distinct from the circuits controlling sleep-wake cycles.

neuroscience↗

bioRxiv2026

Spatial proteomics reveals prefrontal circuit diversity in socioemotional behaviour

Understanding how molecular diversity across long-range neural circuits governs brain function remains a central challenge in neuroscience. Here, we developed a projection-specific spatial proteomics approach and revealed robust presynaptic molecular divergence across six projection-defined pathways of the medial prefrontal cortex (mPFC), including efferent projections to the basolateral amygdala (BLA), nucleus accumbens (NAc), thalamus (Thal), hypothalamus (HT) and cortex (CTX), as well as the afferent projection from the BLA to the mPFC. Among these pathways, we identify BLTP2 (KIAA0100), a previously uncharacterized transmembrane protein, as highly enriched in the projection from the mPFC to the BLA. BLTP2 localizes to excitatory presynaptic terminals and is enriched at synapses that are activated during memory formation. Loss of BLTP2 impairs synaptic structure and transmission, and reduces activity-dependent remodelling, resulting in selective deficits in contextual fear memory, anxiety-related behavior, and social behavior. Mechanistically, BLTP2 promotes presynaptic assembly by recruiting Neurexin 1. These findings reveal projection-specific presynaptic molecular diversity and provide mechanistic insights into circuit-level vulnerabilities in neuropsychiatric disorders.

neuroscience↗

bioRxiv2025

Dissecting Agronomically Favorable Genotypes in Temperate Japonica Rice via Haplotype Analysis of a Japan-MAGIC Population

O_LICrop breeding assembles genomic variants into cultivars via crossing and selection. Phenotypic selection has improved yield and lodging tolerance but has limited genetic insights. We show how specific genomic variants and their combinations underpin advances in modern rice breeding in Japan. C_LIO_LIThrough genome-wide association study using a multi-parent advanced-generation intercross population derived from four temperate japonica cultivars, we identified 11 quantitative trait loci (QTLs) for key agronomic traits, including days to heading, shoot biomass, panicle length, and culm length under field conditions. GA20ox1, GA20ox2, and Hd1 were among the QTLs, and their natural variants were well conserved in temperate japonica cultivars bred in Japan, underscoring distinct selection pressures at these loci. C_LIO_LIBy integrating genotype data with 5-year yield-performance-evaluation trials of elite cultivars, we found that cultivars carrying multiple-copy GA20ox1 and functional Hd1, together with non-functional ga20ox2, tended to have shorter culms and higher grain yield than cultivars with multiple-copy GA20ox1, functional Hd1, and GA20ox2. This yield advantage was consistent across latitudes in Japan. C_LIO_LIThese results reveal favorable genotype combinations underlying modern japonica improvement and provide a genomic framework for breeding semi-dwarf, high-yielding cultivars adapted to temperate rice-growing regions in Asia. C_LI

genomics↗

bioRxiv2025

Upregulation of the lncRNA XACT sustains pluripotency, blocks lineage specification, and drives germ cell tumor-like transcriptional programs in human pluripotent stem cells

Long noncoding RNAs (lncRNAs) represent a vast class of regulatory transcripts and are spatiotemporally controlled, yet only a few have been functionally implicated in human development. Here, we identify the X-linked lncRNA XACT, abundantly but transiently expressed during early human embryogenesis, as a critical regulator of pluripotency, lineage specification, and cancer-like states. In human pluripotent stem cells (hPSCs), XACT overexpression--but not depletion--sustains self-renewal without exogenous factors and prevents lineage commitment. Mechanistically, XACT upregulation drives hyper-elevation of the core pluripotency factors OCT4 and NANOG at the protein level by repressing their 3' untranslated regions (UTRs). XACT overexpression confers context-dependent states: in standard hPSC medium it promotes a naive-like program, whereas in the absence of exogenous factors it drives transcriptomic states resembling testicular germ cell tumors, linking misregulation of a developmentally restricted lncRNA to tumorigenic potential. In hPSC-based models of post-implantation development, XACT expression normally declines, whereas its sustained expression disrupts embryonic progression, while depletion has little effect. Finally, transcriptomic analysis of post-implantation human embryos showed that XACT levels correlate positively with pluripotency-associated gene networks. Together, these findings establish XACT as a potent, human-specific modulator of pluripotency and early embryogenesis, and suggest that its aberrant upregulation may underlie both developmental failure and germ cell tumorigenesis.

cell biology↗

bioRxiv2024

Highly efficient XIST reactivation in female hPSC by transient dual inhibition of TP53 and DNA methylation during Cas9 mediated genome editing

Erosion of X-chromosome inactivation (XCI) complicates disease and developmental modeling in female human pluripotent stem cells (hPSCs). Previous studies demonstrated that Cas9-mediated editing of the XIST promoter via non-homologous end joining (NHEJ) or homology-directed repair (HDR) with a selection cassette upstream of XIST can trigger DNA demethylation and XIST reactivation, restoring XCI. Here, we show that NHEJ-mediated XCI reacquisition is more stable during differentiation than HDR. We further developed a novel, efficient XIST reactivation method by combining TP53 inhibition with DNA methylation maintenance suppression during Cas9-mediated NHEJ, offering a robust approach to achieving stable XCI in female hPSCs for diverse applications.

cell biology↗

bioRxiv2023

Phylogenomics reveals the demography of reverse colonization in a rail species

AimIslands have recently been recognized as potential sources of biodiversity, challenging the traditional view that their small population sizes and low genetic diversity limit such roles. This raises the question of how insular genetic variation becomes incorporated into continental populations, contrary to expectations of unidirectional colonization. Here, we investigate whether and how island-derived genetic variation has influenced a continental population through population establishment and gene flow in a bird species where frequent trans-ocean dispersal is expected. LocationContinental East Asia (Russian Far East), Japanese Archipelago TaxonSwinhoes Rail (Coturnicops exquisitus) Materials and MethodsWe apply integrative phylogenomics to reconstruct the spatiotemporal history of the species. Colonization sequences and gene flow are inferred by comparing four different phylogenetic reconstruction methods, using mitochondrial sequences obtained by Sanger sequencing and genome-wide data obtained by genotyping by sequencing (MIG-seq). We assess a history of colonization and gene flow based on summary statistics, demographic trajectory inference by Stairway Plot2, demographic modeling by fastsimcoal2, and species distribution modeling. ResultsAnalyses collectively supported asymmetric gene flow from the island to the continental population, following divergence around the Middle Pleistocene. Post-divergence, the island maintained a large and stable population, while the continental population underwent a severe bottleneck, suggesting a significant evolutionary role of the island for the continental population. Additionally, evidence of recent re-establishment of the island by continental individuals indicates dynamic exchange and persistence within a continent-island metapopulation. Main conclusionsThe maintenance of insular genetic variation within a dynamic continent-island metapopulation may have enabled the island to act as a genetic and demographic reservoir for the continental population. Thus, continent-island metapopulation dynamics may be a key evolutionary pathway through which island populations contribute to continental genetic diversity.

evolutionary biology↗

bioRxiv2023

Synaptic and peptidergic connectomes of the Drosophila circadian clock

The circadian clock and its output pathways play a pivotal role in optimizing daily processes. To obtain novel insights into how diverse rhythmic physiology and behaviors are orchestrated, we have generated the first comprehensive connectivity map of an animal circadian clock using the Drosophila FlyWire brain connectome. Intriguingly, we identified additional dorsal clock neurons, thus showing that the Drosophila circadian network contains [~]240 instead of 150 neurons. We also revealed extensive contralateral synaptic connectivity within the network and discovered novel indirect light input pathways to the clock neurons. Interestingly, we observed sparse monosynaptic connectivity between clock neurons and down-stream higher-order brain centers and neurosecretory cells known to regulate behavior and physiology. Therefore, we integrated single-cell transcriptomics and receptor mapping to decipher putative paracrine peptidergic signaling by clock neurons. Our analyses identified additional novel neuropeptides expressed in clock neurons and suggest that peptidergic signaling significantly enriches interconnectivity within the clock network.

neuroscience↗

bioRxiv2023

Direct photoreception of a pituitary endocrine cell, melanotroph, induces a hormone release

In addition to canonical photoreception by the eye, many other organs express non-visual photoreceptors although their biological significance is mostly unknown. Here, we discovered a novel phenomenon in which the pituitary of medaka directly receives light, which induces hormone release. Ca2+ imaging analysis revealed that a melanotroph, a pituitary endocrine cell secreting melanocyte-stimulating hormone (MSH), robustly increases [Ca2+]i during short-wavelength light irradiation. Moreover, we identified Opn5m as the key molecule of this mechanism. The significance of this phenomenon was suggested to be involved in UV protection because knockout of opn5m significantly reduced the expression of tyrosinase, the rate-limiting enzyme for melanogenesis, in the skin. These results suggest a novel mechanism in which direct reception of short-wavelength light by pituitary endocrine cells triggers the pathway to enhance UV protection. One-Sentence SummaryAn endocrine cell of the pituitary was proven to be a photoreceptive cell that enables autonomous hormone release.

physiology↗

bioRxiv2022

Fe(III)-dependent anaerobic methane-oxidizing activity in a deep underground borehole demonstrated by in-situ pressure groundwater incubation

The family Methanoperedenaceae archaea mediate anaerobic oxidation of methane (AOM) in various terrestrial environments. In this study, we newly developed a high-pressure laboratory incubation system by controlling hydraulic pressure ranging from ambient to 5 MPa. Using the system, we investigated groundwater from 214- and 249-m deep boreholes at Horonobe Underground Research Laboratory, Japan, where the high and low abundances of Methanoperedenaceae archaea have been revealed by genome-resolved metagenomics, respectively. We incubated the groundwater samples amended with or without amorphous Fe(III) as an electron acceptor and 13C-labelled methane at an in-situ pressure of 1.6 MPa. After three to seven-day incubation, AOM activities were not detected from the 249-m deep groundwater but from the 214-m deep groundwater. The AOM rates were 93.7 {+/-} 40.6 and 27.7 {+/-} 37.5 nM/day with and without Fe(III) amendment. To clarify the differences in AOM activity between the 214- and 249-m deep groundwater samples, we characterized Fe(III) contents in suspended particulates collected by filtration. The particulates were not visible in the 249-m deep groundwater on the filter, while they were abundant and contained Fe(III)-bearing phyllosilicates in the 214-m deep groundwater. These results support the in-situ activity of Fe(III)-dependent AOM in the deep subsurface borehole.

microbiology↗

bioRxiv2021

Loss of TBK1 activity leads to TDP-43 proteinopathy through lysosomal dysfunction in human motor neurons

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by motor neuron loss accompanied by cytoplasmic localization of TDP-43 proteins and their insoluble accumulations. Haploinsufficiency of TBK1 has been found to associate with or cause ALS. However, the cell-autonomous mechanisms by which reduced TBK1 activity contributes to human motor neuron pathology remain elusive. Here, we generated a human cellular model harboring loss-of-function mutations of TBK1 by gene editing and found that TBK1 deficiency was sufficient to cause TDP-43 pathology in human motor neurons. In addition to its functions in autophagy, we found that TBK1 interacted with endosomes and was required for normal endosomal maturation and subsequent lysosomal acidification. Surprisingly, TDP-43 pathology resulted more from the dysfunctional endo-lysosomal pathway than the previously recognized autophagy inhibition mechanism. Restoring TBK1 levels ameliorated lysosomal dysfunction and TDP-43 pathology and maintained normal motor neuron homeostasis. Notably, using patient-derived motor neurons, we found that haploinsufficiency of TBK1 sensitized neurons to lysosomal stress, and chemical regulators of endosomal maturation rescued the neurodegenerative process. Together, our results revealed the mechanism of TBK1 in maintaining TDP-43 and motor neuron homeostasis and suggested that modulating endosomal maturation was able to rescue neurodegenerative disease phenotypes caused by TBK1 deficiency.

neuroscience↗

bioRxiv2020

Full-length 16S rRNA gene amplicon analysis of human gut microbiota using MinION™ nanopore sequencing confers species-level resolution

BackgroundSpecies-level genetic characterization of complex bacterial communities has important clinical applications in both diagnosis and treatment. Amplicon sequencing of the 16S ribosomal RNA (rRNA) gene has proven to be a powerful strategy for the taxonomic classification of bacteria. This study aims to improve the method for full-length 16S rRNA gene analysis using the nanopore long-read sequencer MinION. We compared it to the conventional short-read sequencing method in both a mock bacterial community and human fecal samples. ResultsWe modified our existing protocol for full-length 16S amplicon sequencing by MinION. A new strategy for library construction with an optimized primer set overcame PCR-associated bias and enabled taxonomic classification across a broad range of bacterial species. We compared the performance of full-length and short-read 16S amplicon sequencing for the characterization of human gut microbiota with a complex bacterial composition. The relative abundance of dominant bacterial genera was highly similar between full-length and short-read sequencing. At the species level, MinION long-read sequencing had better resolution for discriminating between members of particular taxa such as Bifidobacterium, allowing an accurate representation of the sample bacterial composition. ConclusionsOur present microbiome study, comparing the discriminatory power of full-length and short-read sequencing, clearly illustrated the analytical advantage of sequencing the full-length 16S rRNA gene, which provided the requisite species-level resolution and accuracy in clinical settings.

microbiology↗