Biology subjects

Yoshikawa, M.

Publications and source records attributed to Yoshikawa, M..

10 recordsLinked to original sources

bioRxiv2026

An anti-aggregation region of the SGS3 N-terminal IDR is essential for secondary siRNA biogenesis

Secondary siRNA biogenesis amplifies small RNA signals from target transcripts and plays a pivotal role in plant development and defense responses. The RNA-binding protein SGS3 is essential for this pathway, recruiting RNA-dependent RNA polymerase 6 (RDR6) to Argonaute-small RNA-bound targets. The N-terminal intrinsically disordered region (IDR) of SGS3, which contains a prion-like domain (PrLD), has been reported to drive liquid-liquid phase separation, forming siRNA bodies, and to be required for secondary siRNA production. However, the molecular mechanism by which the N-terminal IDR contributes to secondary siRNA production remains unclear. Here, using in vitro reconstitution and in planta assays, we show that the N-terminal IDR comprises two functional modules: the PrLD and a negatively charged region (NCR). The PrLD is required for phase separation and siRNA body formation but is dispensable for secondary siRNA production. In contrast, mutations in the NCR caused SGS3 to form abnormally large cytoplasmic assemblies and markedly impaired secondary siRNA production. These results suggest that the N-terminal IDR helps maintain SGS3 in a functional, soluble state that supports efficient secondary siRNA biogenesis.

molecular biology↗

bioRxiv2025

Designing Cell Cell Relation Extraction Models: A Systematic Evaluation of Entity Representation and Pre-training Strategies

Extracting cell-cell relations from biomedical literature is essential for understanding intercellular communication in immunity, inflammation, and tissue biology. However, cell-cell relation extraction has not been established as a standalone biomedical relation extraction task, and no benchmark corpus or systematic evaluation framework currently exists. Fully manual corpus construction is costly and difficult to scale, limiting literature-based analyses of cell-cell communication. Here, we define a sentence-level cell-cell relation extraction task and construct complementary manually annotated corpora under realistic annotation constraints. To enable scalable annotation, rule-based literature mining is used solely as an annotation accelerator to identify candidate sentences, while all relation labels are assigned manually. In addition, an independently annotated PubMed corpus without rule-based filtering is constructed to evaluate robustness on natural sentence distributions. Using these resources, we evaluate representative model configurations involving entity indication strategies, classification architectures, and continued pre-training. Our results show that cell-cell relation extraction remains challenging under realistic conditions. Increasing training data size yields consistent performance gains, and specific combinations of entity-aware architectures and continued pre-training provide modest robustness improvements. Nevertheless, performance on unfiltered PubMed sentences remains in the 70% accuracy range, and error analyses indicate that failures cannot be readily explained by simple surface-level factors. Comparisons with general-purpose large language models further suggest that task complexity, rather than model class, is the primary limiting factor. Together, these findings establish a practical foundation for literature-scale cell-cell relation extraction while clarifying its intrinsic limitations.

bioinformatics↗

bioRxiv2025

Distinct dermal fibroblasts direct mechano-chemical signaling to the epidermis during pregnancy

Dermal fibroblasts alter extracellular matrix (ECM) and tissue mechanics dynamically in wound healing and fibrosis, however, are understudied in healthy skin remodeling across life stages. Here, by conducting spatio-temporal transcriptomics, we identified Cdh4+ dermal fibroblast subpopulation that remodels ECM and converts mechano-to-chemical signals to promote epidermal stem cell proliferation in expanding abdominal skin of pregnant mice. Mechanistically, Cdh4+ fibroblasts produce a pregnancy-responsive matrisome, resulting in denser and stiffer dermal fibrils. These fibroblasts then sense the stiffened substrate and convert the mechanical input into a chemical output via the YAP1-TGF{beta}2 axis, leading to the proliferation of epidermal stem cells via TGF{beta} receptor in the expanding skin. Thus, Cdh4+ fibroblasts fine-tune the dermal mechanofield and act as a mechanical-to-chemical signal conversion hub in healthy skin expansion.

developmental biology↗

bioRxiv2025

Novel humanized anti-Nav1.7 antibodies with long-lasting, side-effect-free analgesic effects

Neuropathic pain remains difficult to treat effectively because of the limitations of current pain medications. The Nav1.7 sodium channel plays a crucial role in pain sensation. The development of selective inhibitors has been challenging because of the high similarity among Nav channel subtypes. To address this issue, we developed monoclonal antibodies that selectively target Nav1.7 and humanized them for clinical use for the first time. When administered systemically to neuropathic pain model rats, a potent analgesic effect was observed that lasted for at least 96 hours. Electrophysiological studies revealed that the antibody reduced mechanically-evoked and spontaneous neuronal activity in the model. Importantly, the antibodies did not impact physiological pain or motor function. Collectively, our findings suggest that these novel Nav1.7-targeting antibodies are potentially effective analgesic drugs for chronic pain, including neuropathic pain. Our novel humanized anti-Nav1.7 antibodies have the potential to be used for the development of new analgesic, and one such antibody, S-151128 is currently in clinical trial.

pharmacology and toxicology↗

bioRxiv2024

RTL4, a retrovirus-derived gene implicated in autism spectrum disorder, is a microglial gene that responds to noradrenaline in the postnatal brain

RTL4, a gene acquired from a retrovirus, is a causative gene in autism spectrum disorder. Its KO mice exhibit increased impulsivity, impaired short-term spatial memory, failure to adapt to novel environments, and delayed noradrenaline (NA) recovery in the frontal cortex. However, due to its very low expression in the brain, it remains unknown which brain cells express RTL4 and its dynamics in relation to NA. Here, using knock-in mice carrying endogenous Rtl4 fused to Venus, we demonstrated that RTL4 is a microglial gene with several important properties. The RTL4-Venus fusion protein was detected as a secreted protein in the midbrain, hypothalamus, hippocampus and amygdala in the postnatal brain. Its signal intensity was high during critical periods of neonatal adaptation to novel environments and was upregulated by various stimuli, such as handling the mice, mild environmental changes and administration of isoproterenol, an agonist of adrenergic beta receptors. It was decreased by anesthesia but was maintained by the administration of milnacipran, an NA reuptake inhibitor. These results suggest that RTL4 intensity depends on the arousal state via NA and is somehow related to the NA reuptake process. In vitro mixed glial culture experiments demonstrated that Rtl4 is a microglial gene and suggested that RTL4 secretion responds rapidly to isoproterenol. Taken together with previous results from Rtl4 KO mice, microglial RTL4 plays an important role in the NA response and it is likely involved in the development of the NAergic neuronal network in the brain.

molecular biology↗

bioRxiv2023

ORP9-PH domain-based fluorescent reporters for visualizing phosphatidylinositol 4-phosphate dynamics in living cells

Fluorescent reporters that visualize phosphatidylinositol 4-phosphate (PI4P) in living cells are indispensable to elucidate the roles of this fundamental lipid in cell physiology. However, currently available PI4P reporters have limitations, such as Golgi-biased localization and low detection sensitivity. Here, we present a series of fluorescent PI4P reporters based on the pleckstrin homology (PH) domain of oxysterol-binding protein-related protein 9 (ORP9). We show that the green fluorescent protein AcGFP1-tagged ORP9-PH domain can be used as a fluorescent PI4P reporter to detect cellular PI4P across its wide distribution at multiple cellular locations, including the plasma membrane (PM), Golgi, endosomes, and lysosomes with high specificity and contrast. We also developed blue, red, and near-infrared fluorescent PI4P reporters suitable for multicolor fluorescence imaging experiments. Finally, we demonstrate the utility of the ORP9-PH domain-based reporter to visualize dynamic changes in the PI4P distribution and level in living cells upon synthetic ER-PM membrane contact manipulation and GPCR stimulation. This work offers a new set of genetically encoded fluorescent PI4P reporters that are practically useful for the study of PI4P biology.

cell biology↗

bioRxiv2022

The bHLH transcription factor MpHYPNOS is a critical regulator of gemma dormancy in the liverwort Marchantia polymorpha.

O_LIDormancy is a key process employed by land plants to adapt to harsh terrestrial environments. The liverwort Marchantia polymorpha produces dormant propagules called gemmae for asexual reproduction. The plant hormone abscisic acid (ABA) plays important roles in regulating dormancy in both the seeds of flowering plants and the gemmae of M. polymorpha. C_LIO_LIBased on previous transcriptome analysis, we identified the basic helix-loop-helix transcription factor MpHYPNOS (MpHYP) as a key regulator of gemma dormancy. C_LIO_LIKnock-out mutants of MpHYP showed much higher germination rates of gemmae in gemma cups than ABA-related mutants, while the growth and development of these mutants resembled that of the wild type. Transient induction of MpHYP caused irreversible growth arrest of gemmae and thalli. Transcriptome and RT-qPCR analyses revealed that MpHYP represses the expression of cell cycle-related genes and induces ABA biosynthesis and ABA-responsive genes. Indeed, ABA levels increased in MpHYP overexpression lines and decreased in Mphyp knock-out lines. However, the growth arrest caused by MpHYP overexpression was not suppressed by a mutation in an ABA receptor gene. C_LIO_LIThese findings suggest that MpHYP regulates gemma dormancy and thallus growth mainly through the ABA-independent pathway, providing clues about ABA-dependent and independent regulation of dormancy in land plants. C_LI

plant biology↗

bioRxiv2020

Synthetic protein condensates that recruit and release protein activity in living cells

Compartmentation of proteins into biomolecular condensates or membraneless organelles formed by phase separation is an emerging principle for the regulation of cellular processes. Creating synthetic condensates that accommodate specific intracellular proteins on demand would have various applications in chemical biology, cell engineering and synthetic biology. Here, we report the construction of synthetic protein condensates capable of recruiting and/or releasing proteins of interest in living mammalian cells in response to a small molecule or light. We first present chemogenetic protein-recruiting and -releasing condensates, which rapidly inhibited and activated signaling proteins, respectively. An optogenetic condensate system was successfully constructed that enables reversible release and sequestration of protein activity using light. This proof-of-principle work provides a new platform for chemogenetic and optogenetic control of protein activity in mammalian cells and represents a step towards tailor-made engineering of synthetic protein condensates with various functionalities.

synthetic biology↗

bioRxiv2020

Ribosome stalling caused by the Argonaute-miRNA-SGS3 complex regulates production of secondary siRNA biogenesis in plants

The path of ribosomes on mRNAs can be impeded by various obstacles. One such example is halting of ribosome movement by microRNAs, though the exact mechanism and physiological role remain unclear. Here, we find that ribosome stalling caused by the Argonaute-microRNA-SGS3 complex regulates the production of secondary small interfering RNAs (siRNAs) in plants. We show that the double-stranded RNA-binding protein SGS3 directly interacts with the 3' end of the microRNA in an Argonaute protein, resulting in ribosome stalling. Importantly, microRNA-mediated ribosome stalling positively correlates with efficient production of secondary siRNAs from target mRNAs. Our results illustrate a role for paused ribosomes in regulation of small RNA function that may have broad biological implications across the plant kingdom.

molecular biology↗

bioRxiv2019

Controlling gene activation by enhancers through a drug-inducible topological insulator

While regulation of gene-enhancer interaction is better understood, its application remains limited. Here, we reconstituted arrays of CTCF binding sites and devised a synthetic topological insulator with tetO for chromatin-engineering (STITCH). By coupling STITCH with tetR linked to the KRAB domain to induce heterochromatin and disable the insulation, we developed a drug-inducible system to control gene activation by enhancers. We applied this to dissect MYC regulation in human pluripotent stem cells. Insertion of STITCH between MYC and the enhancer down-regulated MYC and affected its target transcriptome. Progressive mutagenesis of STITCH led to preferential escalation of the gene-enhancer interaction, corroborating the strong insulation ability of STITCH. The STITCH insertion altered epigenetic states around MYC. Time-course analysis by drug induction uncovered deposition and removal of H3K27me3 repressive marks follows and reflects, but does not precede and determine, the expression change. Thus the tool provided important insights in gene regulation, demonstrating its potency.

genetics↗