Biology subjects

Suda, T.

Publications and source records attributed to Suda, T..

17 recordsLinked to original sources

bioRxiv2026

Highly plastic macrophage niches orchestrate acquired quiescence and reactivation in breast-cancer bone metastasis

Recurrence and metastasis remain major causes of cancer mortality, sustained by therapy-resistant micrometastatic cells. Bone is a frequent site of breast-cancer relapse, yet the cues that reawaken disseminated cells remain poorly defined. We identify a previously unrecognized, highly plastic CXCL16 macrophage population that integrates tumor-associated macrophage programs found in distant metastatic sites such as lung and brain with non-tumor disease-associated traits in bone marrow. These CXCL16 macrophages establish a transient niche that restrains disseminated cancer-cell proliferation. Single-cell transcriptomics delineate functional remodeling of myeloid niches within the bone metastatic microenvironment: a CXCL16 macrophage niche that transiently constrains metastatic growth, and G-CSF macrophage and neutrophil niches that reignite tumor outgrowth. In primary tumors, cancer-associated fibroblasts (CAFs) aberrantly secrete G-CSF in response to cancer-cell signals, expanding G-CSF-receptor-positive subset of cancer cells with high metastatic potential. In advanced human bone metastases, CXCL16 macrophages localize to CAF-rich stroma but are excluded from cancer-cell clusters, indicating immune evasion. Together, these findings uncover CAF-bone-marrow cross-talk as a therapeutic target linking stromal inflammation, immune remodeling, and metastatic progression.

cancer biology↗

bioRxiv2026

Inflammatory restraint and membrane lipid integrity protect hematopoietic stem cells under stress

Life-long production of blood requires the preservation of hematopoietic stem cell (HSCs) regenerative capacity during inflammation. The cytokine, Thrombopoietin (THPO), is essential for HSC maintenance yet its role during inflammatory stress remains incompletely understood. Long-term repopulating potential was rapidly depleted in THPO-deficient HSCs upon poly(I:C) administration through inflammatory pyroptosis. Transcriptomic and chromatin accessibility analyses revealed constitutive interferon (IFN) pathway activation in THPO-deficient HSCs, characterized by enhanced STAT1 signaling, increased accessibility of STAT and IRF motifs, and elevated expression of IFN-stimulated genes. Lipidomic profiling further identified selective shifts in sphingomyelin (SM) species and enrichment of features associated with increased bilayer rigidity. THPO-deficient HSCs displayed elevated membrane SM incorporation, impaired membrane fluidity and altered membrane ultrastructure. Genetic ablation of Stat1 normalized membrane lipid abnormalities and reduced pyroptotic activation and restored HSC survival and regenerative function under inflammatory stress. Together, these findings identify a STAT1 and SM metabolism as critical THPO downstream to protect HSCs from inflammatory pyroptosis. Our results reveal membrane lipid homeostasis as a fundamental mechanism through which cytokine signaling safeguards HSC function during stress.

cell biology↗

bioRxiv2026

Developmental Gfi1 Dynamics Define Hematopoietic Emergenceand Adult Hematopoietic Stem Cell Potency

Gfi1 regulates endothelial-to-hematopoietic transition (EHT) and hematopoietic stem cell (HSC) maintenance; however, its expression dynamics remain unclear. We generated a non-disruptive Gfi1-T2A-mScarlet reporter to track Gfi1 expression. At E10.5, reporter selectively marked EHT in the dorsal aorta, umbilical artery, and vitelline artery. At E9.5, mScarlet-positive endothelial cells were present in the umbilical and vitelline arteries but were scarce in the dorsal aorta, indicating vascular-bed-specific differences in hemogenic activation. Gfi1 was broadly expressed in fetal liver HSCs, while higher expression levels identified HSCs with enhanced multilineage reconstitution and preferential T-cell output. Although Gfi1 expression declined during fetal-to-adult maturation, a subset of adult bone marrow HSCs retained expression and exhibited superior repopulating and self-renewal capacity. Bulk and single-cell transcriptomic analyses linked this subset to dormant and fetal-associated programs, including imprinted genes. Reduced chromatin accessibility at a conserved Gfi1 +26.5kb putative enhancer correlated with developmental Gfi1 downregulation. Thus, Gfi1 dynamics define EHT onset and functionally distinct HSC stemness.

cell biology↗

bioRxiv2026

DNA barcoding-based paired daughter cell analysis reveals division preferences of hematopoietic stem cells

Hematopoietic stem cells(HSCs) maintain their pools by stem-stem division and produce mature blood cells through stem-progenitor or progenitor-progenitor division. A paired daughter cell(PDC) assay combined with single cell transplantation is a powerful method to compare the lineage outputs of two HSC daughter cells. However, single-cell transplantation precludes large-scale analysis of daughter-cell pairs, as only one cell can be transplanted per recipient. Here, we developed a DNA barcoding-based PDC assay to overcome this limitation, enabling simultaneous analysis of 476 daughter pairs from individual HSC divisions and revealing that daughter-cell fates are coordinated and that HSC division patterns are biased toward stem-stem and progenitor-progenitor divisions rather than stem-progenitor divisions. These findings indicate that HSC fate outcomes are directed toward symmetric division outcomes. Integration of single-cell RNA sequencing with DNA barcoding revealed a continuum of HSC states--from balanced HSCs to myeloid-biased HSCs and ultimately to a low-output HSC subset--in which progressively reduced production of mature blood cells relative to stem cell expansion, a proxy for stem-stem division bias, exhibits distinct activities of transcription factors and signaling pathways. Overall, our analysis uncovers characteristic patterns of HSC division and links stem maintenance with distinct molecular features.

cell biology↗

bioRxiv2026

BRM preserves sinusoidal niche integrity to prevent microenvironmental senescence and sustain hematopoietic stem cells

During aging and stress, remodeling of the bone marrow (BM) microenvironment compromises hematopoietic stem cell (HSC) maintenance, contributing to an extrinsic HSC aging phenotype. Although niche-derived Notch signaling is essential for hematopoietic regeneration following myelosuppressive injury, the upstream epigenetic mechanisms that regulate this stress-responsive signaling remain poorly understood. Here, we identify the chromatin remodeler BRM (SMARCA2) as a critical regulator of the BM sinusoidal niche that preserves vascular integrity and hematopoietic regeneration. Using reciprocal BM transplantation, we demonstrate that a Brm-deficient microenvironment impairs HSC repopulating capacity and imposes an aging-like myeloid bias characterized by expansion of granulocyte-monocyte progenitors. Following 5-fluorouracil (5-FU)-induced myelosuppression, BrmKO mice exhibit defective sinusoidal regeneration accompanied by endothelial degeneration. Mechanistically, BRM deficiency attenuates endothelial Notch signaling by impairing stress-induced Notch2 expression in sinusoidal endothelial cells (SECs), while simultaneously reducing Jag2 ligand pool through persistent depletion of SECs and impaired stress-induced expansion of Jag2-producing LepR-positive stromal cells. These alterations attenuate endothelial Notch signaling, resulting in defective sinusoidal regeneration, loss of mesenchymal niche support, and progressive displacement of HSCs from the sinusoidal vasculature. Notably, Brm expression is physiologically reduced in aged wild-type SECs and LepR-positive stromal cells. Collectively, our findings identify BRM as a key epigenetic regulator of bone marrow niche integrity and suggest that age-associated BRM decline contributes to niche dysfunction and hematopoietic aging. Key PointsO_LIBRM preserves the bone marrow niche to sustain HSC repopulating capacity and prevent aging-associated myeloid bias. C_LIO_LIBRM preserves stress-responsive Notch signaling in the bone marrow niche, thereby maintaining sinusoidal integrity and HSC localization. C_LI

immunology↗

bioRxiv2025

Plcl1 Regulates Hematopoietic Stem Cell Function During Aging and Stress by Modulating Calcium Dynamics

Long-term hematopoietic stem cells (HSCs) can generate all blood lineages but typically remain quiescent, becoming activated only in response to acute stress. We previously demonstrated that quiescent HSCs exhibit heterogeneity in intracellular calcium levels. However, the mechanisms underlying this heterogeneity and its physiological relevance remain unclear. Herein, we identify phospholipase C-like 1 (Plcl1), a noncatalytic protein that binds inositol 1,4,5-trisphosphate (IP3), as being selectively enriched in the most quiescent HSC subset. Loss-of-function studies revealed that Plcl1 deficiency at steady state reduced basal intracellular calcium levels and skewed the HSC compartment toward CD41 subsets while preserving overall HSC numbers and long-term reconstitution capacity. Under acute hematopoietic stress, Plcl1 loss accelerated and amplified platelet rebound and the expansion of non-canonical megakaryocyte progenitors (ncMkPs), indicating activation of the thrombopoietic bypass pathway. In aged HSCs, Plcl1 deficiency exacerbated aging-related features, including expansion of the HSC pool, accumulation of CD41 HSCs and ncMkPs, and myeloid-skewed differentiation with impaired competitive reconstitution. These changes were accompanied by diminished induction of calcium-responsive immediate-early genes. Collectively, we identified Plcl1 as an intrinsic regulator that stabilizes calcium dynamics in HSCs, thereby restraining stress- and aging-associated megakaryocytic priming and preserving stem cell function.

cell biology↗

bioRxiv2025

Molecular basis of the autoregulatory mechanism of motor neuron-related splicing factor 30

Motor neuron-related splicing factor 30 (SPF30, also known as SMNDC1) is a paralog of the survival motor neuron protein that regulates the expression of various genes by affecting mRNA splicing. SPF30 has an autoregulatory mechanism that controls its expression. However, the detailed molecular mechanisms determining cellular levels of SPF30 remain unclear. Here, we demonstrated that SPF30 expression was controlled via the negative autoregulatory feedback, whereby increased SPF30 expression caused the inclusion of cassette exon within intron 2 and/or the generation of a newly spliced variant with exon 4a (produced by splicing 17 bp upstream of the canonical intron 3 and exon 4 junctions). Altered transcripts with cassette exon or exon 4a were subjected to nonsense-mediated mRNA decay, leading to reduced SPF30 mRNA levels. Conversely, the loss of SPF30 protein resulted in a drastic reduction in exon 4a inclusion compared to cassette exon inclusion, suggesting that exon 4a inclusion contributes more to adjusting SPF30 expression levels. An in vivo splicing assay designed to reflect exon 4a inclusion levels demonstrated that a short stretch of sequence within exon 4 of SPF30 mRNA was required for exon 4a inclusion. Additionally, the C-terminal region of SPF30 was crucial for the autoregulatory mechanism. Specifically, the C-terminal region of SPF30, including the latter part of -helix and a kink-like structure, was required for binding to RNA containing exon 4a. Collectively, these results reveal the molecular basis of the autoregulatory mechanism underlying SPF30 gene expression.

molecular biology↗

bioRxiv2024

Quantitative phase imaging with temporal kinetics predicts hematopoietic stem cell diversity

Innovative identification technologies for hematopoietic stem cells (HSCs) have advanced the frontiers of stem cell biology. However, most analytical techniques capture only a single snapshot, disregarding the temporal context. A comprehensive understanding of the temporal heterogeneity of HSCs necessitates live-cell, real-time and non-invasive analysis. Here, we developed a prediction system for HSC diversity by integrating single-HSC ex vivo expansion technology with quantitative phase imaging (QPI)-driven machine learning. By analyzing single-cell kinetics with QPI, we discovered previously undetectable diversity among HSCs that snapshot analysis fails to capture. Our QPI-driven algorithm quantitatively evaluates the stemness of individual HSCs and incorporates temporal information to significantly improve prediction accuracy. This platform marks a paradigm shift from "identification" to "prediction", enabling us to forecast HSC status by analyzing their past temporal kinetics.

cell biology↗

bioRxiv2024

Genotype dependence of virulence-associated genes in ESBL-producing Escherichia coli isolated from the blood of patients with urinary tract infections

Approximately 80% of urinary tract infections (UTIs) are caused by uropathogenic Escherichia coli (UPEC). The prevalence of extended-spectrum {beta}-lactamase (ESBL)- producing E. coli in UPEC isolates belonging to specific multilocus sequence-type clones is a global concern. Relations between the pathogenicity of UPEC virulence factors and genotype have been discussed. However, the specific virulence factors and genotypes associated with a higher likelihood of causing UTIs remain unclear. This study analyzed the genotypes and virulence-associated genes of 46 ESBL-producing strains isolated from the blood of patients with UTIs who visited the Department of Emergency Medicine from 2017 to 2022 to elucidate the characteristics of UPEC strains associated with severe infections. Most phylogroups of clinical isolates were B2, except for D, found in three strains. The dominant multilocus sequence typing (MLST) was ST131, followed by ST73, ST95, and ST38, frequently observed in UPEC strains. ST131 strains were more resistant to levofloxacin (87.5%) than non-ST131 strains (50.0%) and caused fewer sepsis cases than non-ST131 strains. Based on in silico analysis, of 23 clinical isolates, the genes detected in all clinical strains could have important role in invasive UTIs. Clustering analysis highlighted the genotype MLST dependence of UPEC-specific virulence-associated genes. The absence of several UPEC-specific gene loci or the presence of ST38-specific indicated atypical repertories of virulence-associated genes in ST38 strains. Genes encoding secretion systems, which were found in enteropathogenic E. coli, were less detected in ST131 strains. These results suggested that the correlation of MLST and repertories should be considered to understand UPEC virulence. IMPORTANCE STATEMENTSeveral virulence genes, their diverse repertories depending on the strains, and various pathological indications complicate the pathogenicity of uropathogenic Escherichia coli (UPEC). The genes detected in all clinical isolates in this study would play a crucial role in UPEC pathogenicity. In contrast, depending on the genotype, the distribution of UPEC- specific genes and other pathogenic E. coli virulence genes in some genotype strains varied. Secretion system genes related to the pathogenicity of enteropathogenic E. coli (EPEC) were less in the strains of the most prevalent genotype, ST131; this was relevant to the number of sepsis cases. These results clearly indicated genotype dependencies on virulence-associated gene repertories and ambiguity in UPEC and EPEC. This study proposed that analyzing genotype-specific virulence factors and sharing virulence genes with EPEC will bring new insights into UPEC pathogenicity.

microbiology↗

bioRxiv2024

JMJD3 mediated senescence is required to overcome stress induced hematopoietic defects.

Cellular senescence in stem cells compromises regenerative capacity, promotes chronic inflammation, and is implicated in aging. Hematopoietic stem and progenitor cells (HSPCs) are responsible for producing mature blood cells, however, how cellular senescence influences their function is largely unknown. Here, we show that JMJD3, a histone demethylase, activates cellular senescence via p16Ink4a upregulation in competition with Polycomb group proteins, and reprograms HSPC integrity to overcome hematopoietic defects induced by replicative and oncogenic stresses. JMJD3 deficiency impaired stem cell potential, proper cell cycle regulation, and WNT pathway activation in HSPCs under stress conditions. These impaired phenotypes were rescued through exogenous and retroviral introduction of p16Ink4a. This JMJD3-p16INK4a axis in hematopoiesis is age-dependent and is distinct from cellular senescence. Treatment with a JMJD3 selective inhibitor attenuated leukemic potential during cellular senescence. Taken together, these results demonstrate that JMJD3-p16INK4a mediates cellular senescence and plays critical roles in the functional integrity of HSPCs under stress conditions, suggesting a new link for aging and anti-cancer therapies.

cell biology↗

bioRxiv2023

Remodeling of the secretory pathway is coordinated with de novo membrane formation in budding yeast gametogenesis

Gametogenesis in budding yeast involves a large-scale rearrangement of membrane traffic to allow de novo formation of a membrane, called the prospore membrane (PSM). However, the mechanism underlying this event is not fully elucidated. Here, we show that the number of endoplasmic reticulum exit sites (ERES) per cell fluctuates and switches from decreasing to increasing upon the onset of PSM formation. Reduction in ERES number, presumably accompanying a transient stall in membrane traffic, resulting in the loss of preexisting Golgi apparatus from the cell, was followed by local ERES regeneration, leading to Golgi reassembly in nascent spores. We have revealed that protein phosphatase-1 (PP-1) and its development-specific subunit, Gip1, promote ERES regeneration through Sec16 foci formation. Furthermore, sed4{Delta}, a mutant with impaired ERES formation, showed defects in PSM growth and spore formation. Thus, ERES regeneration in nascent spores facilitates the segregation of membrane traffic organelles, leading to PSM growth.

cell biology↗

bioRxiv2023

Metabolic regulation in erythroid differentiation by systemic ketogenesis in fasted mice

Systemic ketogenesis affects murine erythroid differentiation under fasting condition, while less ketone body {beta} OHB boosts fatty acid synthesis and mevalonate pathway along with decreased levels of histone acetylation, which are beneficial for erythroid differentiation and maturation undergoing stressed erythropoiesis. AbstractErythroid terminal differentiation and maturation depends on enormous energy supply. During periods of fasting, ketone bodies from the liver are transported into circulation and utilized as crucial fuel for peripheral tissues. However, the effects of fasting or ketogenesis on erythroid behavior remain unknown. Here, we generated a mouse model with insufficient ketogenesis by conditionally knocking out the gene encoding the hepatocyte-specific ketogenic enzyme hydroxymethylglutary-CoA synthase 2 (Hmgcs2 KO). Intriguingly, erythroid maturation was enhanced with boosted fatty acid synthesis in bone marrow of hepatic Hmgcs2 KO mouse under fasting condition, suggesting that systemic ketogenesis has a profound effect on erythropoiesis. Moreover, we observed significantly activated fatty acids synthesis and mevalonate pathway along with reduced histone acetylation in immature erythrocytes under less systemic ketogenesis condition. Our findings revealed an innovative insight to erythroid differentiation, in which metabolic homeostasis and histone acetylation mediated by ketone bodies are essential factors in adaptation towards nutrient deprivation and stressed erythropoiesis.

cell biology↗

bioRxiv2023

Context-Dependent Modification of PFKFB3 in Hematopoietic Stem Cells Promotes Anaerobic Glycolysis and Ensures Stress Hematopoiesis

Metabolic pathways are plastic and rapidly change in response to stress or perturbation. Current metabolic profiling techniques require lysis of many cells, complicating the tracking of metabolic changes over time after stress in rare cells such as hematopoietic stem cells (HSCs). Here, we aimed to identify the key metabolic enzymes that define differences in glycolytic metabolism between steady-state and stress conditions in HSCs and elucidate their regulatory mechanisms. Through quantitative 13C metabolic flux analysis of glucose metabolism using high-sensitivity glucose tracing and mathematical modeling, we found that HSCs activate the glycolytic rate-limiting enzyme phosphofructokinase (PFK) during proliferation and oxidative phosphorylation (OXPHOS) inhibition. Real-time measurement of adenosine triphosphate (ATP) levels in single HSCs demonstrated that proliferative stress or OXPHOS inhibition led to accelerated glycolysis via increased activity of PFKFB3, the enzyme regulating an allosteric PFK activator, within seconds to meet ATP requirements. Furthermore, varying stresses differentially activated PFKFB3 via PRMT1-dependent methylation during proliferative stress and via AMPK-dependent phosphorylation during OXPHOS inhibition. Overexpression of Pfkfb3 induced HSC proliferation and promoted differentiated cell production, whereas inhibition or loss of Pfkfb3 suppressed them. This study reveals the flexible and multilayered regulation of HSC glycolytic metabolism to sustain hematopoiesis under stress and provides techniques to better understand the physiological metabolism of rare hematopoietic cells. Key PointsO_LICombined isotope tracing, mathematical modeling, and single cell ATP analysis enable high-resolution evaluation of blood cell metabolism. C_LIO_LIUnder stress, HSCs quickly accelerate glycolysis to meet ATP demands and maintain hematopoiesis via context-dependent PFKFB3 activation. C_LI

cell biology↗

bioRxiv2023

Mitochondrial translation regulates terminal erythroid differentiation by maintaining iron homeostasis

A lack of the mitochondrial tRNA taurine modifications mediated by mitochondrial tRNA translation optimization 1 (Mto1) was recently shown to induce proteostress in embryonic stem cells. Since erythroid precursors actively synthesize the hemoglobin protein, we hypothesized that Mto1 dysfunctions may result in defective erythropoiesis. Hematopoietic-specific Mto1 conditional knockout (cKO) mice were embryonic lethal due to niche-independent defective terminal erythroid differentiation. Mechanistically, mitochondrial oxidative phosphorylation complex-I was severely defective in the Mto1 cKO fetal liver and this was followed by cytoplasmic iron accumulation. Overloaded cytoplasmic iron promoted heme biosynthesis and enhanced the expression of embryonic hemoglobin proteins, which induced an unfolded protein response via the IRE1-Xbp1 signaling pathway in Mto1 cKO erythroblasts. An iron chelator rescued erythroid terminal differentiation in the Mto1 cKO fetal liver in vitro. The new point of view provided by this novel non-energy-related molecular mechanism may lead to a breakthrough in mitochondrial research.

cell biology↗

bioRxiv2021

Depth-targeted intracortical microstroke by two-photon photothrombosis in rodent brain

SignificancePhotothrombosis is a widely used model of ischemic stroke in rodent experiments. In the photothromboris model, the photosensitizer Rose Bengal is systemically introduced to the blood stream and activated by green light to induce aggregation of platelets that eventually cause vessel occlusion. Since the activation of Rose Bengal is a one-photon phenomenon and the molecules in the illuminated area (light path) are subject to excitation, targeting of thrombosis is unspecific especially in the depth dimension. We have developed a photothrombosis protocol that can target a single vessel in the cortical parenchyma by two-photon excitation. AimWe aim to induce a thrombotic stroke in the cortical parenchyma by two-photon activation of Rose Bengal so that we confine photothrombosis within a vessel of a target depth. ApproachFITC-dextran is injected into the blood stream to visualize the cerebral blood flow in anesthetized adult mice with a cranial window. After a target vessel is chosen by two-photon imaging (950 nm), Rose Bengal is injected into the blood stream. The scanning wavelength is changed to 720 nm and photothrombosis was induced by scanning the target vessel. ResultsTwo-photon depth-targeted single vessel photothrombosis was achieved with a success rate of 84.9{+/-}1.7% within 80 s. Attempts without Rose Bengal (i.e., only with FITC) did not result in photothrombosis at the excitation wavelength of 720 nm. ConclusionsWe described a protocol that achieves depth-targeted single vessel photothrombosis by two-photon excitation. Simultaneous imaging of blood flow in the targeted vessel using FITC dextran enabled the confirmation of vessel occlusion and prevention of excess irradiation that possibly induces unintended photodamage.

neuroscience↗

bioRxiv2021

m6A demethylase ALKBH5 promotes tumor cell proliferation by destabilizing IGF2BPs target genes and worsens the prognosis of patients with non-small cell lung cancer

The modification of N6-methyladenosine (m6A) in RNA and its eraser ALKBH5, an m6A demethylase, play important roles across various steps of human carcinogenesis. However, the involvement of ALKBH5 in non-small cell lung cancer (NSCLC) development remains to be completely elucidated. The current study revealed that the expression of ALKBH5 were increased in NSCLC and increased expression of ALKBH5 worsened the prognosis of patients with NSCLC. In vitro study revealed that ALKBH5 knockdown suppressed cell proliferation ability of PC9 and A549 cells as well as promoted G1 arrest and increased the number of apoptotic cells. Furthermore, ALKBH5 overexpression increased the cell proliferation ability of the immortalized cell lines. Microarray analysis and western blotting revealed that the expression of CDKN1A or TIMP3 were increased by ALKBH5 knockdown. These alterations were offset by a double knockdown of both ALKBH5 and one of the IGF2BPs. The decline of mRNAs was, at least partly, owing to the destabilization of these mRNAs by one of the IGF2BPs. In conclusions, the ALKBH5-IGF2BPs axis promotes cell proliferation and tumorigenicity, which in turn causes the unfavorable prognosis of NSCLC.

cancer biology↗

bioRxiv2018

Cell division history determines hematopoietic stem cell potency

Loss of stem cell self-renewal may underpin aging. Here, we combined single cell profiling, deep-learning, mathematical modelling and in vivo functional studies to explore how hematopoietic stem cell (HSC) division patterns evolve with age. We trained an artificial neural network (ANN) to accurately identify cell types in the hematopoietic hierarchy and predict their age from their gene expression patterns. We then used this ANN to compare daughter cell identities immediately after HSC divisions, and found that HSC self-renewal declines sharply with age. Furthermore, while HSC cell divisions are deterministic and intrinsically-regulated in young and old age, they become stochastic and niche-sensitive in mid-life. Analysis of evolving division patterns indicated that the self-renewal ability an individual HSC depends upon number times it has divided previously. We propose a model of HSC proliferation that accurately predicts the accumulation of HSCs with age, and reconciles the stochastic and instructive views of fate commitment.

cell biology↗