Biology subjects

Yamagata, K.

Publications and source records attributed to Yamagata, K..

14 recordsLinked to original sources

bioRxiv2026

Co-targeting an AMPK--MAPK axis reprograms CAFs and suppresses PDAC

Pancreatic ductal adenocarcinoma (PDAC) is a fatal cancer characterized by limited therapeutic options and a highly treatment-resistant tumor microenvironment. Beyond tumor-intrinsic genetic alterations, growing evidence indicates that host-microbiome interactions influence cancer progression through microbial metabolites. However, how microbiome-derived metabolites influence oncogenic signaling in PDAC remains unclear. Here, integrated profiling revealed a consistent reduction of the microbial metabolite acetic acid in fecal samples from treatment-naive patients with PDAC and in a genetically defined Drosophila model recapitulating key PDAC driver alterations. Acetic acid activates AMP-activated protein kinase, and pharmacological activation of this pathway together with inhibition of mitogen-activated protein kinase signaling suppressed tumor growth in fly and mouse models. Combined pathway targeting restored AMPK activity and suppressed cancer-associated fibroblast activation. These findings identify a microbiome-associated metabolic vulnerability in PDAC and suggest that coordinated targeting of metabolic and oncogenic signaling may restrain tumor progression and improve therapeutic strategies.

cancer biology↗

bioRxiv2025

Integrin-binding matricellular protein fibulin-5 maintains epidermal stem cell heterogeneity during skin aging

The extracellular matrix (ECM) is crucial in building the extracellular environment and translating extracellular information into biochemical signals that sustain organ functions. Fibulin-5 (Fbln5) is a multifunctional ECM protein essential for forming elastic fibers and regulating cellular functions by binding to integrins. While fibulin-5 expression decreases with age in human skin, the functional implications of this decrease, particularly in epidermal stem cell regulation, have remained largely unexplored. Here, we find that the knocking out of Fbln5 in mice results in early impairments of epidermal stem cell properties, similar to the chronological aging of the skin. The Fbln5 deficiency suppresses the expression of integrins and other cell junction proteins, leading to the inactivation of YAP signaling in epidermal stem cells. This reduction in YAP signaling results in the downregulation of the fast-cycling epidermal stem cell marker SLC1A3 in human skin and primary keratinocytes. These results suggest that, in addition to its known function in elastic fiber formation, fibulin-5 also plays a critical role in regulating the balance of epidermal stem cell populations during skin aging and coordinating the crosstalk between the extracellular environment and intracellular signaling.

developmental biology↗

bioRxiv2025

Differential effects of the D1/S264V mutation in Photosystem II with either PsbA1 or PsbA3 on QB, non-heme Iron, and the associated hydrogen-bond network

The role of the D1/S264 residue and the role of its environment in the proton-coupled electron transfer reaction on the acceptor side of Photosystem II were investigated. To this end, D1/S264V mutants were constructed in the thermophilic cyanobacterium Thermosynechococcus elongatus, with D1 being either PsbA1 or PsbA3. The PSII mutants were investigated using EPR spectroscopy, thermoluminescence, (time-resolved) absorption changes measurements, and oximetry. While the mutation had minor effects in PsbA1-PSII, the S264V mutation in PsbA3-PSII had significant consequences: i) thermoluminescence data show inefficient electron transfer from QA- to QB; ii) re-oxidation of QA- was slowed, by at least a factor of 10; iii) the herbicides inhibit weakly O2 evolution; iv) no Fe2+QB- EPR signal was detected in dark-adapted PSII; instead, v) a large Fe3+ signal was present with vi) modified EPR properties; vii) no QA-Fe2+QB- biradical signal was observed after illumination at 198 K following a flash illumination, confirming the inefficient formation of QB-; viii) either no proton uptake coupled to non-heme iron reduction occurred or with a very slow rate compared to PsbA3-PSII; ix) changes were noted in the electrochromic response associated with QA- formation; and x) increased production of singlet oxygen, both with and without herbicides. The S264V mutation in PsbA3-PSII leads to a significant decrease in the energy gap between the QA-QB and QAQB- states. The effects listed above are discussed regarding the differences between PsbA1-PSII and PsbA3-PSII as those related to the sulfoquinovosyldiacylglycerol, the water molecules and the H-bond network.

biophysics↗

bioRxiv2025

Chronic social defeat causes dysregulation of systemic glucose metabolism via the cerebellar fastigial nucleus

Chronic psychological stress leads to hyperglycemia through the endocrine and sympathetic nervous systems, which contributes to the development of type II diabetes mellitus (T2DM). Higher plasma corticosteroids after stress is one well-established driver of insulin resistance in peripheral tissues. However, previous studies have indicated that only a fraction of patients with depression and post-traumatic disorder (PTSD) who develop T2DM exhibit hypocortisolism, so corticosteroids do not fully explain psychological stress-induced T2DM. Here, we find that chronic social defeat stress (CSDS) in mice enhances gluconeogenesis, which is accompanied by a decrease in plasma insulin, an increase in plasma catecholamines, and a drop in plasma corticosterone levels. We further reveal that these metabolic and endocrinological changes are mediated by the activation of neurons projecting from the cerebellar fastigial nucleus (FN) to the medullary parasolitary nucleus (PSol). These neurons are crucial in shifting the bodys primary energy source from glucose to lipids. Additionally, data from patients with depression reveal correlations between the presence of cerebellar abnormalities and both worsening depressive symptoms and elevated HbA1c levels. These findings highlight a previously unappreciated role of the cerebellum in metabolic regulation and its importance as a potential therapeutic target in depression, PTSD, and similar psychological disorders.

neuroscience↗

bioRxiv2024

Four-dimensional label-free live cell image segmentation for predicting live birth potential of mouse embryos

1Selection of high-quality embryos is critical in assisted reproductive technology (ART), but it relies on visual assessment by experts, and the birth rate remains low. We previously developed a deep learning method to predict the birth of mouse embryos by quantifying the morphological features of cell nuclei. This method involves cell nuclear segmentation on fluorescence microscopy images, but fluorescence labeling of nuclei is not feasible in medical applications. Here, we developed FL2-Net, a nuclear segmentation method for time-series three-dimensional bright-field microscopy images of mouse embryos without fluorescence labeling. FL2-Net outperformed existing state-of-the-art segmentation methods. We predicted the birth potential of mouse embryos from the nuclear features quantified by bright-field microscopy image segmentation. Birth prediction accuracy of FL2-Net (81.63%) exceeded those of other methods and experts (55.32%). We expect that FL2-Net, which can quantify nuclear features of embryos non-invasively and with high accuracy, might be useful in ART.

systems biology↗

bioRxiv2024

Reconstruction of artificial nuclei with nuclear import activity in living mouse oocytes

The cell nucleus is a dynamic structure repeating disassembly and reformation during mitosis. Reformation of the nucleus is essential for cell proliferation, and therefore, the factors required for nuclear reformation are fundamental for eukaryotes. Although various factors have been identified in in vitro systems using frog egg extracts and in vivo imaging of somatic cells, little is known about the factors required for the formation of functional nuclear structures in living mouse eggs. To identify such factors, we used a reconstruction approach to construct an artificial nucleus around DNA in mouse eggs. T4 phage DNA (166 kbp) was microinjected into living mouse oocytes. Amounts of DNA injected and injection timing were examined to determine the conditions appropriate for the formation of functional nuclei. Microinjection of 100-500 ng/{micro}l DNA during metaphase through telophase of the second meiosis, but not the subsequent interphase, was important for the formation of artificial nucleus. This T4 DNA-derived artificial nucleus had the structure of nuclear lamina and nuclear pore complexes, and nuclear transport activity, similar to natural nuclei. These results suggest that exogenous DNA can form a functional nucleus in mouse oocytes, regardless of the sequence or the source of DNA.

cell biology↗

bioRxiv2024

Visualizing histone H4K20me1 in knock-in mice expressing the mCherry-tagged modification-specific intracellular antibody

During development and differentiation, histone modifications dynamically change locally and globally, associated with transcriptional regulation, DNA replication and repair, and chromosome condensation. The level of histone H4 Lys20 monomethylation (H4K20me1) increases during the G2 to M phases of the cell cycle and is enriched in facultative heterochromatin, such as inactive X chromosomes in cycling cells. To track the dynamic changes of H4K20me1 in living cells, we have developed a genetically encoded modification-specific intracellular antibody (mintbody) probe that specifically binds to the modification. Here, we report the generation of knock-in mice in which the coding sequence of the mCherry-tagged version of the H4K20me1-mintbody is inserted into the Rosa26 locus. The knock-in mice, which ubiquitously expressed the H4K20me1-mintbody, developed normally and were fertile, indicating that the expression of the probe does not disturb the cell growth, development, or differentiation. Various tissues isolated from the knock-in mice exhibited nuclear fluorescence without the need for fixation. The H4K20me1-mintbody was enriched in inactive X-chromosomes in developing embryos and in XY bodies during spermatogenesis. The knock-in mice will be useful for the histochemical analysis of H4K20me1 in any cell types.

cell biology↗

bioRxiv2023

ISWI chromatin remodeling complexes recruit NSD2 and H3K36me2 in pericentromeric heterochromatin

Histone H3 lysine36 dimethylation (H3K36me2) is generally distributed in the gene body and euchromatic intergenic regions. However, we found that H3K36me2 is enriched in pericentromeric heterochromatin in some mouse cell lines. We here revealed the mechanism of heterochromatin targeting of H3K36me2. Among several H3K36 methyltransferases, NSD2 was responsible for inducing heterochromatic H3K36me2. Depletion and overexpression analyses of NSD2-associating proteins revealed that NSD2 recruitment to heterochromatin was mediated through the imitation switch (ISWI) chromatin remodeling complexes, such as BAZ1B-SMARCA5 (WICH), which directly binds to AT-rich DNA via a BAZ1B domain containing AT-hook-like motifs. The abundance and stoichiometry of NSD2, SMARCA5, and BAZ1B or BAZ2A could determine the localization of H3K36me2 in different cell types. To explore the physiological role of heterochromatic H3K36me2, we analyzed mouse tissues and embryos. As a result, H3K36me2 was found in heterochromatin at the 2- to 4-cell stages of mouse preimplantation embryos, suggesting its involvement in developmental regulation. SummaryThe authors discovered histone H3K36me2, which is believed to be enriched in potentially active genomic regions, is also located in transcriptionally inactive regions called heterochromatin in some cell types. The detailed molecular mechanism of its heterochromatin targeting is now revealed.

cell biology↗

bioRxiv2022

AVP neurons act as the primary circadian pacesetter cells in vivo

The central circadian clock of the suprachiasmatic nucleus (SCN) is a network consisting of various neurons and glia. Individual cells have the autonomous molecular machinery of a cellular clock, but their intrinsic periods are considerably variable. Here, we show that arginine vasopressin (AVP) neurons set the ensemble period of the SCN network to control circadian behavior rhythm. Artificial lengthening of cellular periods by deleting casein kinase 1 delta (CK1{delta}) in the whole SCN lengthened the free-running period of behavior rhythm to an extent similar to CK1{delta} deletion specific to AVP neurons. In SCN slices, PER2::LUC reporter rhythms of these mice did not recapitulate the period lengthening. However, in vivo calcium rhythms of both AVP and vasoactive intestinal peptide (VIP) neurons demonstrated lengthened periods similar to the behavioral rhythm upon AVP neuron-specific CK1{delta} deletion. These results indicate that AVP neurons act as the primary determinant of the SCN ensemble period.

neuroscience↗

bioRxiv2022

ROR1 plays a critical role in pancreatic tumor-initiating cells with a partial EMT signature.

Tumor-initiating cells are the major drivers of chemoresistance and relapse, making them attractive targets for cancer therapy. However, the identity of tumor- initiating cells in human pancreatic ductal adenocarcinoma (PDAC) and the key molecules underlying their traits remain poorly understood. Here, we show that a partial epithelial-mesenchymal transition (EMT)-like subpopulation marked by high expression of receptor tyrosine kinase-like orphan receptor 1 (ROR1) is the origin of heterogeneous tumor cells in PDAC. We demonstrate that ROR1 depletion suppresses tumor growth, recurrence after chemotherapy, and metastasis. Mechanistically, ROR1 induces the expression of AURKB by activating E2F to enhance PDAC proliferation. Furthermore, epigenomic analyses reveal that ROR1 is transcriptionally dependent on YAP/BRD4 binding at the enhancer region, and targeting this pathway reduces ROR1 expression and prevents PDAC growth. Collectively, our findings reveal a critical role of ROR1high cells as tumor-initiating cells and the functional importance of ROR1 in PDAC progression, thereby highlighting its therapeutic targetability. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=192 HEIGHT=200 SRC="FIGDIR/small/499868v2_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@167bf72org.highwire.dtl.DTLVardef@49df52org.highwire.dtl.DTLVardef@1e480a3org.highwire.dtl.DTLVardef@b987bf_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2022

Hypoxia causes pancreatic β-cell dysfunction by activating a transcriptional repressor BHLHE40

Hypoxia can occur in pancreatic {beta}-cells in type 2 diabetes. Although hypoxia exerts deleterious effects on {beta}-cell function, the associated mechanisms are largely unknown. Here, we show that the transcriptional repressor basic helix-loop-helix family member e40 (BHLHE40) is highly induced in hypoxic mouse and human {beta}-cells and suppresses insulin secretion. Conversely, BHLHE40 deficiency in hypoxic MIN6 cells or in the {beta}-cells of ob/ob mice reversed the insulin secretion. Mechanistically, BHLHE40 represses expression of Mafa, which encodes the transcription factor musculoaponeurotic fibrosarcoma oncogene family A (MAFA), by attenuating binding of pancreas/duodenum homeobox protein 1 (PDX1) to its enhancer region. Impaired insulin secretion in hypoxic {beta}-cells was recovered by MAFA expression. Collectively, this work identifies BHLHE40 as a key hypoxia-induced transcriptional repressor in {beta}-cells and its implication in the {beta}-cell dysfunction in type 2 diabetes.

biochemistry↗

bioRxiv2021

Deep learning-based algorithm for predicting the live birth potential of mouse embryos

In assisted reproductive technology (ART), embryos produced by in vitro fertilization (IVF) are graded according to their live birth potential, and high-grade embryos are preferentially transplanted. However, the rate of live birth following clinical ART remains low worldwide, suggesting that grading is inaccurate. One explanation is that grading is classically based on the characteristic shape of embryos at a limited number of developmental stages and does not consider the shape of embryos and intracellular structures, e.g., nuclei, at various stages important for normal embryogenesis. Therefore, here we developed a Normalized Multi-View Attention Network (NVAN) that directly predicts live birth potential from nuclear structural features in live-cell fluorescence images taken of mouse embryos across a wide range of stages. The classification accuracy of our method was 83.87%, which greatly exceeded that of existing machine-learning methods and that of visual inspection by embryo culture specialists. By visualizing the features that contributed most to the prediction of live birth potential, we found that the size and shape of the cell nucleus at the morula stage and at the time of cell division were important for live birth prediction. We anticipate that our method will help ART and developmental engineering as a new basic technology for IVF embryo selection.

developmental biology↗

bioRxiv2021

Chromosome counting in the mouse and human zygote using low-invasive super-resolution live-cell imaging

In preimplantation embryos, an abnormal chromosome number causes developmental failure and a reduction in the pregnancy rate. Conventional chromosome testing methods requiring biopsy reduce the risk of associated genetic diseases; nevertheless, the reduction in cell number also reduces the pregnancy rate. Therefore, we attempted to count the chromosomes in mouse (Slc:ICR) embryos using super-resolution live-cell imaging as a new method of chromosome counting that does not reduce the cell number or viability. We counted the forty chromosomes at the first mitosis by injecting embryos with histone H2B-mCherry mRNA under conditions by which pups could be obtained; however, the results were often an underestimation of chromosome number and varied by embryo and time point. Therefore, we developed a method to count the chromosomes via CRISPR/dCas-mediated live-cell fluorescence in situ hybridization targeting the sequence of the centromere region, enabling us to count the chromosomes more accurately in mouse embryos. The methodology presented here may provide useful information for assisted reproductive technologies, such as those used in livestock animals/humans, as a technique for assessing the chromosomal integrity of embryos prior to transfer. Impact StatementLow-toxic super-resolution observation enables chromosome counting in preimplantation embryos without cell collection.

developmental biology↗

bioRxiv2018

Development of Convolutional Neural Network Based Instance Segmentation Algorithm to Acquire Quantitative Criteria of Mouse Development

In embryology, analytical approaches such as segmentation are applied to acquiring quantitative criteria from time-series three-dimensional microscopic images. When used to segment cells or intracellular organelles, several current machine learning techniques outperform traditional image processing algorithms. However segmentation algorithms still have unsolved problems, especially in bioimage processing. The most critical issue is that the existing algorithms can perform only semantic segmentation, which distin-guishes whether a pixel is within an object (for example, nucleus) or not. In this study, we implemented a novel segmentation algorithm, based on deep learning, which segments each nucleus and adds different labels to the detected objects. This segmentation algorithm is called instance segmentation. Our instance segmentation algorithm, implemented as a neural network, which we named QCA Net, substantially out-performed 3D U-Net, which is the best semantic segmentation algorithm that uses deep learning. Using QCA Net, we quantified the nuclear number, volume, surface area, and center of gravity coordinates during the development of mouse embryos. In particular, QCA Net distinguished nuclei of embryonic cells from those of polar bodies formed in meiosis. We consider that QCA Net can greatly contribute to bioimage segmentation in embryology by generating quantitative criteria from segmented images.

systems biology↗