Biology subjects

Iwasaki, K.

Publications and source records attributed to Iwasaki, K..

At least 19 recordsLinked to original sources

bioRxiv2026

Distinct senescent β-cell senotypes differentially drive islet aging and dysfunction

Biological aging greatly impacts the bodys ability to handle glucose, and represents a major risk factor the development and progression of type 2 diabetes (T2D). Nonetheless, despite advances in cellular senescence research and the development of new senolytic therapies, the heterogeneity of cellular senescence in the human endocrine pancreas, as well as its roles in normal aging, remains to be elucidated at the single-cell level. Here, we performed single-cell-resolved spatial proteomics and transcriptomics on intact pancreas from 26 donors (ages 20-80) and multiplexed single-cell RNA sequencing and functional assays on dispersed islets from 14 donors (ages 34-69). We identify two discrete SnC subpopulations distinguished by relative expression of CDKN1A and CDKN2A. CDKN1A senescent cells (SnCs) exhibit loss of {beta}-cell identity, impaired insulin secretion, and a proinflammatory SASP associated with increased islet immune infiltration. In contrast, CDKN2A SnCs retain transcriptional identity and functional competence, with lower inflammatory signaling. Together, these findings identify heterogeneous and functionally divergent senotypes in the human pancreas, distinguishing an adaptive (CDKN2A) from a maladaptive (CDKN1A) senescence program, thus providing a mechanism-guided framework for senescence-targeted therapies in T2D.

cell biology↗

bioRxiv2025

Regulatory mechanism of heme-regulated inhibitor through autophosphorylation-driven activation and heme-induced deactivation

Heme-regulated inhibitor (HRI) is a key modulator of hemoglobin synthesis, exerting such effects by sensing intracellular heme levels. Under heme-deficient conditions, HRI dissociates from heme, becomes activated, and phosphorylates the translation initiation factor eIF2. However, the precise regulatory mechanisms governing HRI activation remain incompletely understood. In this study, we delineate part of the regulatory mechanism involving autophosphorylation-dependent activation and heme-mediated deactivation of HRI. HRI formed a dimer in solution through its N-terminal domain, irrespective of its phosphorylation state. However, a mutant with deletion of the N-terminal domain retained autophosphorylation activity, indicating that N-terminal domain-mediated dimerization is not essential for activation. Phosphorylated HRI formed a stable complex with eIF2, whereas the dephosphorylated form failed to bind, indicating that autophosphorylation is required for eIF2 recognition. Biochemical analyses, together with modeling based on the predicted structure, revealed that the phosphate groups of Thr488 and Thr493 interact with adjacent basic residues. These interactions enhance eIF2 recognition and phosphorylation. Additionally, heme-induced deactivation selectively targeted the dephosphorylated kinase domain and suppressed autophosphorylation. These findings provide multiple mechanistic insights into regulation of the activity of HRI, highlighting distinct roles for autophosphorylation, structural elements, and heme responsiveness in controlling its function.

biochemistry↗

bioRxiv2025

Autofluorescence-based Label-free Cell Counting Method in Suspension Culture with Microcarriers

To advance the industrialization of cultured meat and regenerative medicine, scalable and efficient cell culture techniques are essential. Among these, the suspension culture method using microcarriers has emerged as a promising approach for the large-scale cell culture technique. However, monitoring cell growth on the microcarriers remains challenging, particularly in developing cell counting techniques that can be seamlessly integrated into bioprocess workflows without cell detachment, fluorescence labeling and any parameter tuning in the analysis algorithm. In this study, we proposed a versatile image analysis-based cell counting method by using cellular autofluorescence without any parameter tuning. The proposed method estimates the number of cells by applying spatiotemporal averaging to the autofluorescence signals in the microscopic images. Using numerical and cell culture experiments, we demonstrated that the proposed method can estimates the number of cells accurately. This technique, which harnesses the ubiquitous autofluorescence inherent in living cells, offers a cost-effective and practical solution applicable to a broad range of fields requiring high-throughput cell quantification.

systems biology↗

bioRxiv2025

The I-BAR domain protein baiap2l1a is required for protrusion and lateral elongation of epithelial microridge structures

Microridges are laterally elongated membrane protrusions from the apical surface of epithelial cells. Microridges are arranged in striking maze-like patterns. They are found on various mucosal epithelia in many animals, including the skin of zebrafish, where they are required to maintain mucus on the skin surface. Recent studies have revealed molecular mechanisms of how microridiges formation involving actin and actin-regulatory proteins. However, the molecular mechanism that deforms epithelial membranes to create microridge protrusions remain unknown. We have found that one of the I-BAR domain proteins, baiap2l1a, which is known to regulate membrane curvature, is required for microridge morphogenesis. CRISPR/Cas9 knockdown showed that baiap2l1a mutant zebrafish had defects in microridge morphogenesis. Baiap2l1a mutant zebrafish had shorter and wider microridges than WT microridges. Baiap2l1a localized to microridges, and its localization proceeded microridge actin formation. Furthermore, the baiap2l1a I-BAR domain, which binds and curves membranes, was sufficient to localize to microridges in zebrafish skin cells. Structure/function experiments revealed that the I-BAR domain alone could partially rescued microridge length in baiap2l1a mutants. A 39 amino acid deletion in the I-BAR domain, which caused the loss of one -helix according to AlphaFold2 simulations, is sufficient to impair microridge localization and failed to rescue microridge elongation in baiap2l1a mutants. These results suggest that the I-BAR domain is required for baiap2l1a microridge localization and function. Eps8like1a, a member of the Eps8 family proteins known as an actin capping and bundling protein genetically interacted with baiap2l1a in microridge elongation. Together, we found that the membrane curvature protein baiap2l1a plays an important role in generating microridges in zebrafish epithelia.

cell biology↗

bioRxiv2025

Drosophila learn about properties of objects through physical interaction

Animals interact with unfamiliar objects to learn about their properties and guide future behavior, but the underlying neurobiological mechanism is not well understood. Here, we developed a behavioral paradigm in which freely walking Drosophila melanogaster are repeatedly guided to spherical objects using a visual cue. Flies exhibited diverse and structured object interaction motifs, including "ball pulling", and "ball walking", that evolved over time. Notably, flies developed a strong preference for immobile over mobile spherical objects, despite their near identical appearance, suggesting they learn about the objects stability through physical interaction. This preference was impaired by silencing specific h{Delta} neurons in the fan-shaped body, previously implicated in spatial navigation but not known to contribute to object interactions. Our results show that h{Delta} neurons also modulate object interaction motifs and fidelity of following visual guidance cues, pointing to a role in balancing goal-directed and exploratory behaviors. These findings establish Drosophila as a model for investigating how internal representations and multimodal feedback contribute to adaptive object interaction.

neuroscience↗

bioRxiv2025

A human and mouse subpopulation of senescent β-cells induces pathologic dysfunction through targetable paracrine signaling

Cellular senescence is a stress response mechanism marked by irreversible growth arrest, upregulation of antiapoptotic pathways, loss of cellular function, and remodelling of the cellular secretory profile. In both humans and mice, pancreatic {beta}-cells undergo senescence with age and insulin resistance. Targeted removal of senescent cells in mouse models of diabetes improves glucose homeostasis, demonstrating the role {beta}-cell senescence in diabetes progression. In contrast, {beta}-cell senescence also promotes immune surveillance, promoting {beta}-cell survival and function. Thus, a better understanding of senescent cells phenotypic and functional heterogeneity is needed to develop effective therapeutic strategies. Herein, we show that subpopulations of senescent {beta}-cells in mice and humans, which were identified through the expression of Cdkn1a (encoding p21Cip1) and Cdkn2a (encoding p16Ink4a) by single-cell RNA sequencing (scRNA-seq), flow cytometry, spatial transcriptomics, and spatial proteomics, exhibit distinct transcriptional and functional identities. The predominant senescent {beta}-cell subpopulation expressed Cdkn1a and was characterized by a lack of glucose responsiveness, high basal insulin secretion, and transcription of canonical SASP factors. The SASP of Cdkn1a-expressing {beta}-cells had non-cell autonomous effects on neighbouring cells. A subset of four SASP factors from Cdkn1a+ cells was sufficient to induce secondary senescence and {beta}-cell dysfunction in vitro. JAK inhibitors (JAK1/2 and JAK1/3) counteracted secondary senescence induction and restored {beta}-cell function in high-fat diet-fed mice and human islets from donors with or without type 2 diabetes. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=196 SRC="FIGDIR/small/648438v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@9b8addorg.highwire.dtl.DTLVardef@1b9c4eborg.highwire.dtl.DTLVardef@12f2964org.highwire.dtl.DTLVardef@1468297_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2024

Bacterial genome encoded ParMs

ParMs generally exist on low copy number plasmids where they contribute to plasmid segregation and stable inheritance. We carried out bioinformatics analysis, which indicated that ParM genes are not only confined to plasmids but are also occasionally found on genomes. Here we report the discovery and characterization of two chromosome encoded ParMs (cParMs) from the genomes of Desulfitobacterium hafniense (Dh-cParM1) and Clostridium botulinum (Cb-cParM). Both cParMs form filaments, exhibit nucleotide hydrolysis, and possess characteristic ParM subunit structures. Dh-cParM1 forms single and tightly coupled double filaments and is highly conserved on the chromosomes of five of six Desulfitobacterium species. Interestingly, these bacteria have not been reported to harbour plasmids. Cb-cParM possesses unique properties. Its filaments were stable after nucleotide hydrolysis and Pi release, and its ParR, (Cb-cParR) did not affect the initial stage of Cb-cParM polymerization but displayed properties of a depolymerization factor for mature filaments. These results indicate functional, polymerizing ParMs can be encoded on genomes, suggesting that ParM roles may extend to other functions beyond plasmid segregation.

microbiology↗

bioRxiv2024

The fruit fly, Drosophila melanogaster, as a micro-robotics platform.

Engineering small autonomous agents capable of operating in the microscale environment remains a key challenge, with current systems still evolving. Our study explores the fruit fly, Drosophila melanogaster, a classic model system in biology and a species adept at microscale interaction, as a biological platform for micro-robotics. Initially, we focus on remotely directing the walking paths of fruit flies in an experimental arena. We accomplish this through two distinct approaches: harnessing the fruit flies opto-motor response and optogenetic modulation of its olfactory system. These techniques facilitate reliable and repeated guidance of flies between arbitrary spatial locations. We guide flies along predetermined trajectories, enabling them to scribe patterns resembling textual characters through their locomotion. We enhance olfactory-guided navigation through additional optogenetic activation of positive valence mushroom body output neurons. We extend this control to collective behaviors in shared spaces and navigation through constrained maze-like environments. We further use our guidance technique to enable flies to carry a load across designated points in space, establishing the upper bound on their weight carrying capabilities. Additionally, we demonstrate that visual guidance can facilitate novel interactions between flies and objects, showing that flies can consistently relocate a small spherical object over significant distances. Beyond expanding tools available for micro-robotics, these novel behavioral contexts can provide insights into the neurological basis of behavior in fruit flies.

neuroscience↗

bioRxiv2024

5-Deazaflavin (TND1128) and its hybrid analogs are cytoprotective against hydrogen peroxide (H2O2)-induced oxidative stress.

Increased production of reactive oxygen species (ROS) and oxidative stress are implicated in mitochondrial dysfunction, contributing to the pathogenesis of many neurodegenerative diseases. Research is ongoing into a new treatment approach for neurodegeneration, focusing on reactivating dysfunctional mitochondria. Some 5-deazaflavins, such as 10-ethyl-3-methylpyrimido[4,5-b]quinoline-2,4(3H,10H)-dione (TND1128), and four analogs of 5-deazaflavin, including {beta}-nicotinamide mononucleotide ({beta}-NMN), demonstrate efficient self-redox abilities similar to {beta}-NMN, making them potential activators of mitochondrial energy synthesis. This study examines whether TND1128 and its analogs have protective effects against cellular impairment induced by oxidative stress. These compounds exhibit proliferative potential against normal cells. Moreover, TND1128 and its analogs significantly improved cell viability against hydrogen peroxide (H2O2)-induced oxidative stress injury. Our study confirms the cytoprotective effect of these 5-deazaflavins through mitochondrial activation. We anticipate TND1128 and its analogs will serve as mitochondria-stimulating drugs capable of rescuing deteriorating neurons in aging or diseases.

pharmacology and toxicology↗

bioRxiv2023

Location analysis of presynaptically active and silent synapses in single-cultured hippocampal neurons

A morphologically present but non-functioning synapse is termed a silent synapse. Silent synapses are categorized into "postsynaptically silent synapses," where AMPA receptors are either absent or non-functional, and "presynaptically silent synapses," where neurotransmitters cannot be released from nerve terminals. The presence of presynaptically silent synapses remains enigmatic, and their physiological significance is highly intriguing. In this study, we examined the distribution and developmental changes of presynaptically active and silent synapses in individual neurons. Our findings show a gradual increase in the number of excitatory synapses, along with a corresponding decrease in the percentage of presynaptically silent synapses during neuronal development. To pinpoint the distribution of presynaptically active and silent synapses, i.e., their positional information, we enhanced the traditional Sholl analysis and introduced a novel method termed "donut analysis." Our results indicate that the distribution of presynaptically silent synapses within a single neuron does not exhibit a distinct pattern during synapse development in different areas. However, irrespective of neuronal development, the proportion of presynaptically silent synapses tends to rise as the projection site moves farther from the cell body, suggesting that synapses near the cell body may exhibit higher synaptic transmission efficiency. This study represents the first observation of changes in the distribution of presynaptically active and silent synapses within a single neuron. Additionally, we propose that donut analysis can serve as a valuable analytical tool for evaluating synaptic positional information. Scope statementA morphologically present but non-functioning synapse is termed a silent synapse. The presence of presynaptically silent synapses remains enigmatic, and their physiological significance is highly intriguing. This study focused on the distribution and developmental changes of presynaptically active and silent synapses in individual neurons. To pinpoint the distribution of presynaptically active and silent synapses, we enhanced the traditional Sholl analysis and introduced a novel method termed "donut analysis." We found that the distribution of presynaptically silent synapses within a single neuron does not exhibit a distinct pattern during synapse development in different areas. However, irrespective of neuronal development, the proportion of presynaptically silent synapses tends to rise as the projection site moves farther from the cell body. This is a new paper that applies "Sholl analysis," a method invented 70 years ago that is now the gold standard of morphological analysis of the single neuron. This study represents the first observation of changes in the distribution of presynaptically active and silent synapses within a single neuron. Additionally, we propose that donut analysis can serve as a valuable analytical tool for evaluating synaptic positional information for the design of "synaptic maps" in neural circuits.

neuroscience↗

bioRxiv2022

Three-dimensional motions of GroEL during substrate protein recognition

GroEL is a bacterial chaperonin responsible for the assisted folding of non-native and misfolded polypeptides into biologically active proteins. The adaptive nature of the recognition mechanism of chaperonins toward client polypeptides inherently lends itself to structural heterogeneity, which hampers detailed analyses of intermolecular recognition and binding. In this report, we used single-particle cryo-EM and multiple rounds of focused mask three-dimensional classification to reveal a landscape of distinct snapshots of endogenous GroEL complexed with an unfolded substrate, the water-soluble domain of human UDP glucuronosyltransferase 1A (UGT1A), at 2.7-3.5 [A] resolution. We demonstrate that UGT1A occupies the GroEL ring asymmetrically, engaging with 2-3 contiguous subunits and that a subunit bound to UGT1A exhibits a wider range of conformational dynamics, consistent with AlphaFold models. These data reveal molecular motions during initial substrate capture at near-atomic detail.

biophysics↗

bioRxiv2022

The role of geology in creating stream climate-change refugia along climate gradients

Identifying climate-change refugia is a key adaptation strategy for reducing global warming impacts. Knowledge of the effects of underlying geology on thermal regime along climate gradients and the ecological responses to the geology-controlled thermal regime is essential to plan appropriate climate adaptation strategies. The dominance of volcanic rocks in the watershed is used as a landscape-scale surrogate for cold groundwater inputs to clarify the importance of underlying geology. Using statistical models, we explored the relationship between watershed geology and the mean summer water temperature of mountain streams along climate gradients in the Japanese archipelago. Summer water temperature was explained by the interaction between the watershed geology and climate in addition to independent effects. The cooling effect associated with volcanic rocks was more pronounced in streams with less summer precipitation or lower air temperatures. We also examined the function of volcanic streams as cold refugia under contemporary and future climatic conditions. Community composition analyses revealed that volcanic streams hosted distinct stream communities composed of more cold-water species compared with non-volcanic streams. Scenario analyses revealed a geology-related pattern of thermal habitat loss for cold-water species. Non-volcanic streams rapidly declined in thermally suitable habitats for lotic sculpins even under the lowest emission scenario (RCP 2.6). In contrast, most volcanic streams will be sustained below the thermal threshold, especially for low and mid-level emission scenarios (RCP 2.6, 4.5). However, the distinct stream community in volcanic streams and geology-dependent habitat loss for lotic sculpins was not uniform and was more pronounced in areas with less summer precipitation or lower air temperatures. Although further studies are needed to understand underlying mechanisms of the interplay of watershed geology and climate, findings highlight that watershed geology, climate variability, and their interaction should be considered simultaneously for effective management of climate-change refugia in mountain streams.

ecology↗

bioRxiv2022

LabGym: A versatile computational tool for automatic quantification of user-defined animal behaviors

Quantifying animal behavior is important for many branches of biological research. Current computational tools for behavioral quantification typically rely on a few pre-defined, simplified features to identify a behavior. However, such an approach restricts the information used and the tools applicability to a limited range of behavior types or species. Here we report a new tool, LabGym, for quantifying animal behaviors without such limitations. Combining a novel approach for effective evaluation of animal motion with customizable convolutional recurrent networks for capturing spatiotemporal details, LabGym provides holistic behavioral assessment and accurately identify user-defined animal behaviors without restrictions on behavior types or animal species. It then provides quantitative measurements of each behavior, which quantify the behavior intensity and the body kinematics during the behavior. LabGym requires neither any intermediate step for processing features that causes information loss nor programming knowledge from users for post-hoc analysis. It tracks multiple animals simultaneously in various experimental settings for high-throughput and versatile analysis. It also provides users a way to generate visualizable behavioral datasets that are valuable resources for the research community. We demonstrate its efficacy in capturing subtle behavioral changes in animals ranging from soft-bodied invertebrates to mammals.

animal behavior and cognition↗

bioRxiv2022

Mechanistic insights into intramembrane proteolysis by E. coli site-2 protease homolog RseP

Site-2 proteases are a conserved family of intramembrane proteases that cleave transmembrane substrates to regulate signal transduction and maintain proteostasis. Here, we elucidated crystal structures of inhibitor-bound forms of bacterial site-2 proteases including E. coli RseP. Our observations are consistent with a rearrangement of the RseP domains surrounding the active center to expose the substrate-binding site where a conserved electrostatic linkage between the transmembrane and membrane-associated domains mediates the conformational changes, suggesting that RseP has a gating mechanism to regulate substrate entry. Mutational analysis also supports that the substrate transmembrane helix is unwound by strand addition to the intramembrane {beta} sheet and is clamped at the active center for efficient cleavage. Furthermore, this substrate accommodation mechanism appears to be common across distinct intramembrane proteases.

biochemistry↗

bioRxiv2021

Identification of bacterial drug-resistant cells by the convolutional neural network in transmission electron microscope images

The emergence of bacteria that are resistant to antibiotics is common in areas where antibiotics are used widely. The current standard procedure for detecting bacterial drug resistance is based on bacterial growth under antibiotic treatments. Here we describe the morphological changes in enoxacin-resistant Escherichia coli cells and the computational method used to identify these resistant cells in transmission electron microscopy (TEM) images without using antibiotics. Our approach was to create patches from TEM images of enoxacin-sensitive and enoxacin-resistant E. coli strains, use a convolutional neural network for patch classification, and identify the strains on the basis of the classification results. The proposed method was highly accurate in classifying cells, achieving an accuracy rate of 0.94. Using a gradient-weighted class activation mapping to visualize the region of interest, enoxacin-resistant and enoxacin-sensitive cells were characterized by comparing differences in the envelope. Moreover, Pearsons correlation coefficients suggested that four genes, including lpp, the gene encoding the major outer membrane lipoprotein, were strongly associated with the image features of enoxacin-resistant cells.

systems biology↗

bioRxiv2021

Astrocyte Ca2+ Signaling is Facilitated in an Scn1a+/- Mouse Model of Dravet Syndrome

Dravet syndrome (DS) is an infantile-onset epileptic encephalopathy. More than 80% of DS patients have a heterozygous mutation in SCN1A, which encodes a subunit of the voltage-gated sodium channel, Nav1.1, in neurons. The roles played by astrocytes, the most abundant glial cell type in the brain, have been investigated in the pathogenesis of epilepsy; however, the specific involvement of astrocytes in DS has not been clarified. In this study, we evaluated Ca2+ signaling in astrocytes using genetically modified mice that have a loss-of-function mutation in Scn1a. We found that the slope of spontaneous Ca2+ spiking was increased without a change in amplitude in Scn1a+/- astrocytes. In addition, ATP-induced transient Ca2+ influx and the slope of Ca2+ spiking were also increased in Scn1a+/- astrocytes. These data indicate that perturbed Ca2+ dynamics in astrocytes may be involved in the pathogenesis of DS.

neuroscience↗

bioRxiv2021

SIPA1L1/SPAR1 is a non-PSD protein involved in GPCR signaling

SIPA1L1 (also known as SPAR1) has been proposed to regulate synaptic functions that are important in maintaining normal neuronal activities, such as regulating spine growth and synaptic scaling, as a component of the postsynaptic density (PSD)-95/N-methyl-D-aspartate receptor (NMDA-R)-complex. However, contrary to this view, our super-resolution and immunoelectron microscopic analyses demonstrate that SIPA1L1 is mainly localized to general submembranous and cytoplasmic regions in neurons, but scarcely to PSD. Our screening for native interactors of SIPA1L1 identified spinophilin and neurabin-1, regulators of G protein-coupled receptor (GPCR) signaling, but rejected PSD-95/NMDA-R-complex components. Furthermore, Sipa1l1-/- mice showed normal spine size distribution and NMDA-R-dependent synaptic plasticity. Nevertheless, Sipa1l1-/- mice showed aberrant responses to 2-adrenergic receptor (a spinophilin target) or adenosine A1 receptor (a neurabin-1 target) agonist stimulation and striking behavioral anomalies, such as hyperactivity, enhanced anxiety, learning impairments, social interaction deficits, and enhanced epileptic seizure susceptibility. Our findings revealed unexpected properties of SIPA1L1, suggesting a possible association of SIPA1L1 deficiency with neuropsychiatric disorders related to dysregulated GPCR signaling, such as epilepsy, attention deficit hyperactivity disorder (ADHD), autism, or fragile X syndrome.

neuroscience↗

bioRxiv2021

Atomic structure of human sapovirus capsid by single particle cryo-electron microscopy

Sapovirus is a cause of acute gastroenteritis in humans and animals. Infants and younger children have the greatest disease burden. Although it shares many similarities with norovirus, the lack of detailed structural information has hampered the development of vaccines and therapeutics. Here, we investigated the human sapovirus VLP by single particle cryo-electron microscopy and are the first to report the atomic structure of the capsid at 2.9 [A] resolution. The atomic model revealed the domain interactions of the capsid protein and functionally important amino acid residues. The extended loop from the P1 subdomain was involved in interactions in the P2 domain, forming unique arch-like dimeric protrusions of capsid proteins. All hypervariable regions that are important candidates for immune response or receptor binding, formed a large cluster at the top of the P domain. These results pave the way for developing vaccines, antiviral drugs, and diagnostic systems for this infectious disease.

microbiology↗