Biology subjects

Nishimura, S.

Publications and source records attributed to Nishimura, S..

11 recordsLinked to original sources

bioRxiv2026

Ectopic overproduction of cell wall glucan through membrane perturbation by an antifungal peptide theonellamide A in fission yeast

Ergosterol has multiple functions in filamentous fungi and yeasts, although only a part of the functions seems to be understood. An antifungal peptide, theonellamide A (TNM-A) induces drastic morphological changes in fission yeast cells by targeting plasma membrane ergosterol. TNM-A induces overproduction and ectopic accumulation of cell wall glucan at both growing tips and septum through a yet unknown mechanism. Here we show that TNM-A treatment causes accumulation of 1,3-{beta}-glucan at cell-polarity sites, not by increased activity of 1,3-{beta}-glucan synthase, but by an increased, persistent localization of the glucan synthase enzymes. Screening based on subcellular localization of proteins at periphery or polarity sites suggested the involvement of the Rho family GTPase Cdc42. In agreement, TNM-A induced both activation of Cdc42 and enhancement of membrane trafficking of glucan synthase enzymes. In conclusion, our chemical genetics analyses using TNM-A suggest that membrane ergosterol regulates the activity of Cdc42, which further regulates the localization of glucan synthases and cell wall biosynthesis. Highlights (four sentences)- Thenoellamide A (TNM-A) induces an ectopic overproduction of cell wall glucan. - TNM-A treatment causes increased, persistent localization of glucan synthases at the cell tips and septum. - TNM-A activates Cdc42 and upregulates membrane trafficking of glucan synthases. - Ergosterol is involved in proper activation/inactivation of Cdc42.

cell biology↗

bioRxiv2026

Glutathione acts as an exometabolite that rescues functionally distinct genetic mutations in fission yeast

Microorganisms in nature form communities through diverse interactions, such as mutualism and competition, to adapt to their ecological environments. These interactions seem to be mediated by extracellular metabolites (exometabolites), yet the chemical and biological diversity underlying these processes remains largely unexplored. In this study, we examined the chemical basis of cell-cell communication in the fission yeast Schizosaccharomyces pombe by a genome-wide screen employing 3,420 viable gene deletion mutants. We identified 37 strains that exhibited growth defects in monoculture on a minimal medium but exhibited growth recovery in the vicinity of wild-type colonies (co-culture), suggesting that exometabolites secreted by wild-type cells compensated for the gene deletion. Both lipophilic and water-soluble fractions obtained by solvent partitioning of the wild-type culture supernatant promoted growth recovery. Among the 11 mutants rescued by the water-soluble fraction, six were cysteine auxotrophs, prompting analyses of thiol-containing metabolites by liquid chromatography-mass spectrometry (LC-MS), revealing the presence of glutathione (GSH) in the culture supernatant. GSH restored growth in most strains as a nutrient source. In contrast, GSH rescued cell morphology defects in the hob3{Delta} mutant, lacking the Bin/Amphiphysin/Rvs (BAR) adaptor protein Hob3, through a mechanism independent of nutrition. This research advances understanding of exometabolite-mediated interactions in S. pombe by highlighting the role of GSH as one of the communication molecules that influence cellular processes and shape microbial communities. Author summaryMicroorganisms secrete a wide range of metabolites that control microbial community behavior. These extracellular metabolites (exometabolites) include not only well-studied signaling molecules but also diverse primary and secondary metabolites, suggesting complex interactions among microbes. However, the molecular basis of these interactions remains poorly understood, partly due to challenges in detecting them experimentally. In this study, we surveyed exometabolites involved in cell-cell interactions in the model eukaryotic microorganism Schizosaccharomyces pombe. S. pombe secretes a wide variety of metabolites, including previously reported nitrogen signaling factors (NSFs) and glutathione identified in this work. By analyzing gene deletion mutants that depend on GSH for growth, we provide new insights into how microbes regulate collective behavior by sharing exometabolites.

microbiology↗

bioRxiv2025

A Conserved Stress-Inducible Hub Enhancer Governs Natriuretic Peptide Expression and Undergoes Dynamic and Reversible Activation in Human Heart Failure

BackgroundNatriuretic peptides (NPs), encoded by the NPPA and NPPB genes, serve as both diagnostic biomarkers and cardioprotective hormones in heart failure. Their expression is tightly regulated by mechanical load, yet the upstream enhancer mechanisms translating hemodynamic stress into transcriptional activation remain incompletely understood. MethodsWe investigated the Nppa/Nppb super-enhancer (SE), which resides in a well-insulated topologically associating domain (TAD) and regulates stress-inducible expression of Nppa and Nppb. We applied CRISPR-based enhancer perturbation, chromatin accessibility and histone profiling, chromatin conformation assays, genetic mouse models, human iPSC-derived cardiomyocytes, and paired failing human hearts before and after left ventricular assist device (LVAD) unloading. ResultsThrough integrative epigenomic and genetic analyses, we identified a conserved element, CR9, as a dominant hub enhancer within this SE. In neonatal rat cardiomyocytes, CRISPR-based activation and inhibition established that CR9 was both necessary and sufficient for NP induction, whereas neighboring elements (CR7/CR8) displayed modest activity but synergized with CR9, establishing a hierarchical and cooperative SE architecture. In vivo, CR9 deletion in mice markedly suppressed Nppa/Nppb expression and diminished chromatin accessibility and histone acetylation across adjacent enhancers and promoters, underscoring its structural as well as transcriptional role. Reinsertion of CR9, even in reverse orientation, restored transcription, confirming its orientation-independent activity. CR9 function remained confined within TAD boundaries, supporting a locus-specific regulation. RNAscope revealed spatial proximity between CR9 enhancer RNA and Nppa/Nppb nascent transcripts. Developmental profiling revealed a switch in enhancer usage from CR6/CR7 in the fetal heart to CR9 in adulthood, indicating a transition from developmental to stress-inducible regulation. In human induced pluripotent stem cell-derived cardiomyocytes, deletion of CR9 nearly abolished NPPA/NPPB expression, establishing its essential role in a human cellular context. Most notably, in paired human failing hearts before and after LVAD unloading, chromatin accessibility at CR9 was dynamically reversed, providing the first direct evidence that enhancer states are plastic and therapeutically modifiable in the human myocardium. ConclusionsCR9 functions as a stress-inducible hub enhancer that coordinates NP transcription under pathological load. This enhancer exhibits reversible activation in human hearts, underscoring enhancer plasticity as a modifiable regulatory layer mediating stress-responsive transcriptional adaptation in the diseased myocardium. Clinical PerspectiveO_ST_ABSWhat is new?C_ST_ABSO_LIWe identified CR9 as a stress-inducible hub enhancer within the Nppa/Nppb super-enhancer that is both necessary and sufficient for natriuretic peptide induction, defining a hierarchical and cooperative regulatory architecture controlling Nppa and Nppb expression. C_LIO_LIWe uncovered a developmental switch in enhancer usage from CR6 in the fetal heart to CR9 in adulthood, revealing how stress-inducible enhancer activity emerges as the heart transitions from developmental to adaptive regulation. C_LIO_LIUsing paired failing human hearts before and after LVAD unloading, we provide the first direct evidence that enhancer states are dynamically reversible in the human myocardium. C_LI What are the clinical implications?O_LIThe reversibility of CR9 activity in human heart failure provides a molecular basis for how natriuretic peptide levels decrease when cardiac function improves with effective therapy, reflecting the underlying enhancer plasticity of the diseasesd myocardium. C_LIO_LIEnhancer plasticity is an emerging and targetable mechanism in heart failure, raising the possibility that stress-inducible enhancers like CR9 could be engineered to switch on cardioprotective genes when the heart is under stress, offering a new strategy for precision gene therapy. C_LI

molecular biology↗

bioRxiv2025

Nectriatides with no antifungal activity bind to ergosterol and potentiate the antifungal activity of amphotericin B against Candida albicans

Fungal cyclotetrapeptide nectriatide and its synthetic linear derivatives (nectriatide-based compounds, NCTs), having no antifungal activity, potentiated antifungal activity of amphotericin B (AmB) against Candida albicans. The mechanism of action was investigated using fluorescein-conjugated and biotin-tagged probes. Microscopy revealed fluorescent-probe localization at the C. albicans cell membrane. The biotinyl probe binding assay towards membrane lipids showed the highest affinity for ergosterol but no affinity for cholesterol. Ergosterol binding was confirmed by a liposome disruption assay. LC-MS quantification of AmB in C. albicans revealed that NCTs increased AmB binding to C. albicans. These results indicate that NCTs bind to ergosterol on the fungal plasma membrane, subsequently localizing AmB, explaining the observed potentiation of AmB fungicidal activity.

microbiology↗

bioRxiv2024

Latent-TGF-β has a domain swapped architecture

The multifunctional cytokine TGF-{beta} is produced in a latent form (L-TGF-{beta}) where a RGD containing homodimeric prodomain forms a "ring" encircling mature TGF-{beta} shielding it from its receptors. Thus L-TGF-{beta} must be activated to function, a process driven by dynamic allostery resulting from integrin binding the L-TGF-{beta} RGD motif. Here we provide critical evidence that defines a domain-swapped architecture of L-TGF-{beta}, an essential component in the dynamic allostery mechanism of L-TGF-{beta} activation.

biophysics↗

bioRxiv2024

Formation of giant ER sheets by pentadecanoic acid causes lipotoxicity in fission yeast

Excess amounts of saturated fatty acids are toxic to organisms, a condition termed lipotoxicity, which is often accompanied by pleiotropic cellular and tissue dysfunction. Here we show that pentadecanoic acid (C15:0) exerts toxicity on the fission yeast Schizosaccharomyces pombe by generating an aberrantly planar endoplasmic reticulum (ER) structure, which we named a "giant ER sheet." Untargeted lipidomics revealed that C15:0 is incorporated into complex lipids depending on an acyl-CoA ligase Lcf1 and an acyl-CoA transferase Slc1, thereby increasing the saturation level of the acyl chains. The toxicity and giant ER sheet formation were abolished by deleting Lcf1 or Slc1, indicating that the incorporation of C15:0 into glycerophospholipids causes giant ER sheet formation. The giant ER sheets disrupted the correct migration of Mid1, a protein determining the cell division site, and physically blocked septum formation, hindering correct cell separation. Our results suggest that the ER is the primary site targeted by saturated fatty acids, leading to lipotoxicity.

cell biology↗

bioRxiv2022

Excess glutamate release triggers subunit-specific homeostatic receptor scaling

Ionotropic glutamate receptors (GluRs) are targets for modulation in Hebbian and homeostatic synaptic plasticity and are remodeled by development, experience, and disease. Although much is known about activity-dependent mechanisms that regulate GluR composition and abundance, the role of glutamate itself in these processes is unclear. To determine how glutamate sculpts GluR receptive fields, we have manipulated synaptically released glutamate and generated precise CRISPR mutations in the two postsynaptic GluR subtypes at the Drosophila neuromuscular junction, GluRA and GluRB. We first demonstrate that GluRA and GluRB compete to establish postsynaptic receptive fields, and that proper GluR abundance and localization can be orchestrated in the absence of any synaptic glutamate release. However, excess glutamate release adaptively tunes postsynaptic GluR abundance, echoing GluR receptor scaling observed in mammalian systems. Unexpectedly, when GluRA vs GluRB competition is eliminated, excess glutamate homeostatically regulates GluRA abundance, while GluRB abundance is now insensitive to glutamate modulation. Finally, Ca2+ impermeable GluRA receptors are no longer sensitive to homeostatic regulation by glutamate. Thus, excess glutamate, GluR competition, and Ca2+ signaling collaborate to selectively target GluR subtypes for homeostatic regulation at postsynaptic compartments.

neuroscience↗

bioRxiv2021

Desiccation-induced fibrous condensation of CAHS protein from an anhydrobiotic tardigrade

Anhydrobiosis is one of the most extensively studied forms of cryptobiosis that is induced in certain organisms as a response to desiccation. Anhydrobiotic species has been hypothesized to produce substances that can protect their biological components and/or cell membranes without water. In extremotolerant tardigrades, highly hydrophilic and heat-soluble protein families, cytosolic abundant heat-soluble (CAHS) proteins, have been identified, which are postulated to be integral parts of the tardigrades response to desiccation. However, the molecular mechanisms underlying these protein functions remain to be fully elucidated. In this study, we perfomed in vitro and in vivo characterizations of the self-assembling property of CAHS1 protein, a major isoform of CAHS proteins from Ramazzottius varieornatus, using a series of spectroscopic and microscopic techniques. Our in vitro observations showed that CAHS1 proteins homo-oligomerized via the C-terminal -helical region and formed a hydrogel as their concentration increased, and that these molecular assembling processes were reversible. Furthermore, our in vivo observations demonstrated that the overexpressed CAHS1 proteins formed condensates under desiccation-mimicking conditions. These data strongly suggested that, upon drying, the CAHS1 proteins form oligomers and eventually underwent sol-gel transition in tardigrade cytosols. Thus, it is proposed that the CAHS1 proteins form the cytosolic fibrous condensates, which presumably have variable mechanisms for the desiccation tolerance of tardigrades. These findings provide insights into the protective mechanisms involved in the anhydrobiosis of tardigrades.

biochemistry↗

bioRxiv2020

Distinct target-specific mechanisms homeostatically stabilize transmission at pre-and post-synaptic compartments

Neurons must establish and stabilize connections made with diverse targets, each with distinct demands and functional characteristics. At Drosophila neuromuscular junctions, synaptic strength remains stable in a manipulation that simultaneously induces hypo-innervation on one target and hyper-innervation on the other. However, the expression mechanisms that achieve this exquisite target-specific homeostatic control remain enigmatic. Here, we identify the distinct target-specific homeostatic expression mechanisms. On the hypo-innervated target, an increase in postsynaptic glutamate receptor (GluR) abundance is sufficient to compensate for reduced innervation, without any apparent presynaptic adaptations. In contrast, a target-specific reduction in presynaptic neurotransmitter release probability is reflected by a decrease in active zone components restricted to terminals of hyper-innervated targets. Finally, loss of postsynaptic GluRs on one target induces a compartmentalized, homeostatic enhancement of presynaptic neurotransmitter release called presynaptic homeostatic potentiation that can be precisely balanced with the adaptations required for both hypo- and hyper-innervation to maintain stable synaptic strength. Thus, distinct anterograde and retrograde signaling systems operate at pre- and post-synaptic compartments to enable target-specific, homeostatic control of neurotransmission.

neuroscience↗

bioRxiv2020

Intron retention as a new marker of the pre-disease state and its recovery to the normal state by a traditional Japanese multi-herbal medicine

Intron retention (IR) is an important regulatory mechanism that affects gene expression and protein functions. Using klotho mice at the pre-symptomatic state, we discovered that retained-introns accumulated in several organs including the liver and that among these retained introns in the liver a subset was recovered to the normal state by a Japanese traditional herbal medicine. This is the first report of IR recovery by a medicine. IR-recovered genes fell into two categories: those involved in liver-specific metabolism and in splicing. Metabolome analysis of the liver showed that the klotho mice were under starvation stress. In addition, our differentially expressed gene analysis showed that liver metabolism was actually recovered by the herbal medicine at the transcriptional level. By analogy with the widespread accumulation of intron-retained pre-mRNAs induced by heat shock stress, we propose a model in which retained-introns in klotho mice were induced by an aging stress and in which this medicine-related IR recovery is indicative of the actual recovery of liver-specific metabolic function to the healthy state. Accumulation of retained-introns was also observed at the pre-symptomatic state of aging in wild-type mice and may be an excellent marker for this state in general.

molecular biology↗

bioRxiv2019

Comprehensive epigenome characterization reveals diverse transcriptional regulation across human vascular endothelial cells

BackgroundEndothelial cells (ECs) make up the innermost layer throughout the entire vasculature. Their phenotypes and physiological functions are initially regulated by developmental signals and extracellular stimuli. The underlying molecular mechanisms responsible for the diverse phenotypes of ECs from different organs are not well understood.\n\nResultsTo characterize the transcriptomic and epigenomic landscape in the vascular system, we cataloged gene expression and active histone marks in nine types of human ECs (generating 148 genome-wide datasets) and carried out a comprehensive analysis with chromatin interaction data. We identified 3,765 EC-specific enhancers, some of which were associated with disease-associated genetic variations. We also identified various candidate marker genes for each EC type. Notably, reflecting the developmental origins of ECs and their roles in angiogenesis, vasculogenesis and wound healing.\n\nConclusionsWhile the importance of several HOX genes for early vascular development and adult angiogenesis in pathological conditions has been reported, a systematic analysis of the regulation and roles of HOX genes in mature tissue cells has been lacking. These datasets provide a valuable resource for understanding the vascular system and associated diseases.

genomics↗