Biology subjects

Nakanishi, Y.

Publications and source records attributed to Nakanishi, Y..

9 recordsLinked to original sources

bioRxiv2026

Ultrastructurally planar growth cones during neocortical axon navigation

Neurons extend axons toward spatially defined but often distant targets to establish brain circuits. Callosal projection neurons exemplify this organization, linking corresponding cortical territories between the two hemispheres. Growing axons rely on growth cones to regulate direction and speed through densely packed tissues. Although growth cone morphology and dynamics have been extensively characterized in cultured neurons and invertebrate models, how growth cones navigate the three-dimensional environment of the developing mammalian brain remains poorly explored. Here, we combined a genetically encoded electron microscopy (EM)-labeling strategy and zero-shot deep-learning-based tracing with high-resolution volume EM to reconstruct growth cones of identified callosal projection neurons in the developing mouse brain. Mediolaterally extending growth cones exhibited a planar, fan-shaped morphology, preferentially expanded along the dorsoventral axis. Filopodia extended from the edges of this plane and aligned predominantly with the same axis but showed no directional bias toward the growing direction. Longitudinal in vivo imaging revealed that increased growth cone complexity associated with filopodial extension coincided with lower instantaneous velocity but preceded accelerated extension. Together, these findings reveal a dorsoventrally oriented planar growth cone architecture in which filopodial extension precedes accelerated axon growth, suggesting a structural constraint that may restrict axon navigation to the callosal plane.

neuroscience↗

bioRxiv2026

Reprogramming protein interfaces between adenylation and carrier protein domains in nonribosomal peptide synthetases

Nonribosomal peptide synthetases (NRPSs) assemble structurally diverse bioactive natural products through selective communication between adenylation (A) and carrier protein (CP) domains. Although rational rewiring of these protein-protein interactions could enable customized NRPS design, this remains challenging due to the dynamic nature of protein interfaces. Here we show that structure-guided interface reprogramming enables productive non-cognate A- CP pairings across enterobactin, vibriobactin, pyochelin, and vicenistatin biosynthetic systems. Engineered interactions between the A domains EntE, VibE, or PchD and the non-cognate CPs VinL or EntB were validated by biochemical, kinetic, and structural analyses. Reconstituted pathways containing engineered VibE, EntB, and EntF restored enterobactin production and increased yield to 2.2-fold that of the native EntE-EntB-EntF system. Interface-engineered PchD also enhanced production of a non-native salicylic acid-norspermidine conjugate. An X-ray structure and molecular dynamics simulations of an engineered non-cognate A-CP complex reveal recognition principles for programmable NRPS interface design.

biochemistry↗

bioRxiv2026

Integrating spatial and single-cell multi-omics analysis of induced pluripotent stem cell-derived cervical adenocarcinoma model

Human papillomavirus 18 (HPV18) preferentially infects cervical stem cell-like cells and is strongly associated with adenocarcinoma. However, the mechanisms underlying differentiation into cervical adenocarcinoma remain unclear due to the lack of appropriate experimental models. We aimed to establish a model of HPV18-associated cervical adenocarcinoma and elucidate its molecular and cellular differentiation mechanisms. HPV18 E6/E7 were introduced into induced pluripotent stem cell-derived reserve cell-like cells (iRCs) to generate tumor models. Spatial transcriptomics and single-cell multi-omics analyses were performed to integrate histological and molecular data. A distinct component (Gland_A) exhibited morphological and immunohistochemical features of cervical adenocarcinoma and was efficiently induced in iRC-18 tumors. Gland_A showed increased chromatin accessibility and elevated expression of FOXA1, FOXA2, and ALDH1A1. Analysis of clinical samples confirmed enrichment of ALDH1A1 in HPV-associated adenocarcinomas. This model recapitulates key features of HPV18-associated cervical adenocarcinoma and provides insights into its differentiation mechanisms.

cancer biology↗

bioRxiv2025

Epigenomic and transcriptomic germ-free ageing atlas reveals sterile inflammation as an intrinsic ageing feature

Inflammageing is a hallmark of ageing. Commensal microbiota plays crucial roles in maintaining tissue homeostasis, yet its impact on cellular ageing and inflammageing remains poorly understood. Here we present a comprehensive single-cell epigenomic and transcriptomic atlas of tissues from mice aged under specific pathogen-free (SPF) or germ-free (GF) conditions. Microbiota conferred beneficial effects in young mice but accelerated various ageing features in old, such as age-related AP-1 pathway upregulation, senescence and transcriptomic alterations, likely due to age-associated dysbiosis. Strikingly, inflammatory signatures persisted across cell types in aged GF mouse tissues, establishing sterile inflammation as an intrinsic feature of ageing. Age-associated B cells expanded equally under GF and SPF conditions, raising the possibility that they function as intrinsic, microbiota-independent drivers of inflammageing and potential therapeutic targets. The atlas provides a resource for distinguishing intrinsic ageing features from those modulated by the microbiota, illuminating mechanisms of cellular ageing and potential anti-ageing interventions. HighlightsSterile inflammation and age-associated B cell expansion are prominent intrinsic ageing features The upregulation of age-associated AP-1 pathways and senescence of alveolar macrophages are attenuated in germ-free condition Germ-free condition induces premature ageing-like features in young mouse tissues but delays ageing features in old mice across multiple cell types

genomics↗

bioRxiv2021

The cell surface hyaluronidase TMEM2 plays an essential role in mouse neural crest cell development and survival

Neural crest cells (NCCs) are a migratory population that gives rise to a diverse cell lineage, including the craniofacial complex, the peripheral nervous system, and a part of the heart. Hyaluronan (HA) is a major component of the extracellular matrix, and its tissue levels are dynamically regulated in during development. Although the synthesis of HA has been shown to exert substantial influence on embryonic morphogenesis, the functional importance of the catabolic side of HA turnover is poorly understood. Here, we demonstrate that the transmembrane hyaluronidase TMEM2 plays an essential role in NCC development and the morphogenesis of their derivatives. Wnt1-Cre-mediated Tmem2 knockout (Tmem2CKO) mice exhibit severe craniofacial and cardiovascular abnormalities. Analysis of Tmem2 expression using Tmem2 knock-in reporter mice reveals that Tmem2 is expressed at the site of NCC delamination in the neural tube and in Sox9-positive emigrating NCCs, suggesting that Tmem2 is critical for NCC development. Consistent with this possibility, linage tracing analysis reveals that the contribution of Wnt1-Cre-labeled cells to NCC derivatives is significantly reduced in a Tmem2-deficient background. Moreover, the emigration of NCCs from the neural tube is greatly reduced in Tmem2CKO mice. In vitro assays demonstrate that Tmem2 expression is essential for the ability of mouse O9-1 NCCs to form focal adhesion on and migrate into HA-containing substrates. Tmem2CKO mice also exhibit increased apoptotic cell death in NCC-derived tissues. Collectively, our data demonstrate that Tmem2 is essential for normal development of NCC-derivatives, including the craniofacial complex, and that TMEM2-mediated HA degradation allows NCCs to generate a tissue environment suitable for efficient focal adhesion assembly and migration. This study reveals the hitherto unrecognized functional importance of the catabolic side of HA metabolism in embryonic development and highlights the pivotal role of Tmem2 in the process. Author SummaryThe functional significance of hyaluronan (HA) in embryonic developmental processes has been demonstrated by studies using genetic manipulation of HA synthesis. However, the expression of HA is regulated not only by its synthesis, but also by its degradation. This issue is of particular importance due to the extremely rapid metabolic turnover of HA. Curiously, mice with mutations/ablations of known hyaluronidase molecules, such as the lysosomal hyaluronidases HYAL1 and HYAL2, exhibit little embryonic phenotypes. This suggests the existence of yet another hyaluronidase molecule that plays a key role in regulating extracellular HA balance in developing tissues. In this context, transmembrane protein 2 (TMEM2) is a novel hyaluronidase that functions on the cell surface. Here, we demonstrate that TMEM2 is expressed at the site of neural crest development and in neural crest cell (NSC)-derived craniofacial tissue, and that NCC-targeted Tmem2 conditional knockout mice develop severe craniofacial defects, which attests to a requirement for TMEM2-mediated extracellular HA degradation in neural crest development. Our in vitro and in vivo analyses on the underlying mechanisms of the phenotype demonstrate that TMEM2 is essential for generating a tissue environment suitable for efficient focal adhesion formation by NCCs. This paper reveals for the first time that the catabolic machinery for HA exerts a specific regulatory role in embryonic morphogenesis, and that its dysregulation of HA degradation leads to severe developmental defects.

developmental biology↗

bioRxiv2020

Master Transcription Factors Regulate the DNA Methylation Landscape During Hepatocyte Differentiation

BackgroundHepatocytes are the dominant cell type of the human liver, with functions in metabolism, detoxification, and in producing secreted proteins. During the process of hepatocyte differentiation, gene regulation and master transcription factors have been extensively investigated, whereas little is known about how the epigenome is regulated, particularly the dynamics of DNA methylation, and the upstream factors that have critical roles. ResultsBy examining changes in the transcriptome and the methylome during in vitro hepatocyte differentiation, we identified putative DNA methylation-regulating transcription factors, which are likely involved in DNA demethylation and maintenance of hypo-methylation in a differentiation stage-specific manner. Of these factors, we further reveal that GATA6 induces DNA demethylation together with chromatin activation at a binding-site-specific manner during endoderm differentiation. ConclusionsThese results provide an insight into the spatiotemporal regulatory mechanisms exerted on the DNA methylation landscape by transcription factors, and uncover a new role for transcription factors in early liver development.

genomics↗

bioRxiv2019

Dear Editorial manager PLoS ONEA survey of vaccine-induced measles IgG antibody titer and the verification of changes in temporal differences of measles vaccination in young adults

In Japan, sporadic measles cases increased rapidly in 2019 compared to that in past 6 years. To clarify the persistence of immunity against measles in young adult, this study explored the prevalence of IgG antibody titer against measles virus in 18-to 24-year-old young adult participants. Measles-specific IgG antibody titers determined by enzyme immunoassay in serum samples collected from 506 participants between 18 to 24 years were assessed through statistical analyses. Multiple regression analysis revealed that the distribution of measles IgG antibody titers was significantly correlated with medical history (P < 0.05), while there was no significant difference among the number of vaccinations related to measles IgG titers. Further, measles IgG titers were significantly different, which was determined by the temporal change that elapsed period after last vaccination (P < 0.05). These results indicate that periodic vaccination against measles is required for young and older adults to prevent even sporadic measles infection.

microbiology↗

bioRxiv2017

Neutrophil elastase inhibitor sivelestat ameliorates gefitinib-naphthalene-induced acute pneumonitis in mice

Background and objectiveGefitinib, an epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI), is an effective therapeutic agent for non-small cell lung cancer with EGFR mutations. It can cause severe acute pneumonitis in some patients. We previously demonstrated that mice with naphthalene-induced airway epithelial injury developed severe gefitinib-induced pneumonitis and that neutrophils played important roles in the development of the disease. This study aimed to investigate the effects of the neutrophil elastase inhibitor sivelestat on gefitinib-induced pneumonitis in mice.\n\nMethodsC57BL/6J mice received naphthalene (200 mg/kg) intraperitoneally on day 0. Gefitinib (250 or 300 mg/kg) was orally administered to mice from day -1 until day 13. Sivelestat (150 mg/kg) was administered intraperitoneally from day 1 until day 13. Bronchoalveolar lavage fluid (BALF) and lung tissues were sampled on day 14.\n\nResultsSivelestat treatment significantly reduced the protein level, neutrophil count, neutrophil elastase activity in BALF, and severity of histopathologic findings on day 14 for mice administered with 250 mg/kg of gefitinib. Moreover, sivelestat treatment significantly improved the survival of mice administered with 300 mg/kg of gefitinib. Conclusions: These results indicate that sivelestat is a promising therapeutic agent for severe acute pneumonitis caused by gefitinib.\n\nSummary statementNeutrophil elastase inhibitor sivelestat is a promising therapeutic agent for severe acute pneumonitis caused by gefitinib.

pharmacology and toxicology↗

bioRxiv2016

Liberation from equations: An equation-free method reveals the ecological interaction networks within complex microbial ecosystems

O_LIMapping the network of ecological interactions is key to understanding the composition, stability, function and dynamics of microbial communities. In recent years various approaches have been used to reveal microbial interaction networks from metagenomic sequencing data, such as time-series analysis, machine learning and statistical techniques. Despite these efforts it is still not possible to capture details of the ecological interactions behind complex microbial dynamics.\nC_LIO_LIWe developed the sparse S-map method (SSM), which generates a sparse interaction network from a multivariate ecological time-series without presuming any mathematical formulation for the underlying microbial processes. The advantage of the SSM over alternative methodologies is that it fully utilizes the observed data using a framework of empirical dynamic modelling. This makes the SSM robust to non-equilibrium dynamics and underlying complexity (nonlinearity) in microbial processes.\nC_LIO_LIWe showed that an increase in dataset size or a decrease in observational error improved the accuracy of SSM whereas, the accuracy of a comparative equation-based method was almost unchanged for both cases and equivalent to the SSM at best. Hence, the SSM outperformed a comparative equation-based method when datasets were large and the magnitude of observational errors were small. The results were robust to the magnitude of process noise and the functional forms of inter-specific interactions that we tested. We applied the method to a microbiome data of six mice and found that there were different microbial interaction regimes between young to middle age (4-40 week-old) and middle to old age (36-72 week-old) mice.\nC_LIO_LIThe complexity of microbial relationships impedes detailed equation-based modeling. Our method provides a powerful alternative framework to infer ecological interaction networks of microbial communities in various environments and will be improved by further developments in metagenomics sequencing technologies leading to increased dataset size and improved accuracy and precision.\nC_LI

ecology↗