Biology subjects

Ogawa, H.

Publications and source records attributed to Ogawa, H..

17 recordsLinked to original sources

bioRxiv2026

Tumor Microenvironment Modulates Lineage Plasticity in Lung Squamous Cell Carcinoma

Lung squamous cell carcinoma (LUSC) is the second most common type of lung cancer, yet therapeutic options remain limited. A deeper understanding of its biology and molecular pathogenesis is essential for developing new treatment strategies. Here, we investigated the mechanisms of phenotypic plasticity in LUSC by comparing organoid-derived orthotopic lung models (ODOLs) and subcutaneous xenograft models (ODXs). ODXs showed greater tumor growth, squamous differentiation, and extracellular matrix (ECM) organization compared to ODOLs. Transcriptomic analyses revealed upregulation of multiple HIF1 and SOX2 target genes together with enhanced hypoxia signaling in ODXs. CRISPR/Cas9-mediated HIF1-knockout ODXs showed reduced SOX2 expression, tumor growth, and ECM organization, whereas SOX2-knockout ODXs reduced tumor growth without affecting HIF1 and ECM organization. These results indicate that HIF1 regulates squamous lineage maintenance through SOX2 and ECM remodeling. Spatial transcriptomics revealed enrichment of basal cell-like and squamous-differentiated tumor states in ODXs, whereas ODOLs displayed less differentiated phenotypes. These findings identify the tumor microenvironment as a critical determinant of lineage plasticity in LUSC and provide mechanistic insight into how hypoxia shapes tumor differentiation.

Cancer Biology↗

bioRxiv2026

Mirror-image mRNA display uncovers isoform-selective D-peptide macrocycles targeting a cryptic KRAS pocket

Activating KRAS mutations drive millions of cancers diagnosed worldwide,1 yet for decades this oncoprotein was deemed "undruggable", reflecting the challenge of discovering molecules capable of perturbing its complex biological functions, and of translating these discoveries into effective cancer therapeutics.2 Recent advances propelled by innovative screening have identified diverse modalities that bind at or near the switch-II pocket (SII-P) of RAS proteins, including molecular glues,3 macrocyclic peptides,4 fragment-derived small molecules,5 and approved therapies that covalently target KRASG12C.6,7 Unfortunately, resistance to approved therapies has emerged,8,9 highlighting the need for molecules that engage new or underexploited binding sites on RAS oncoproteins with mechanisms complementary to established SII-P inhibitors.10,11 Here we show that mirror-image mRNA display12 enabled the discovery of all-D macrocyclic peptide ligands targeting a cryptic RAS back pocket (CRB-P).13 These ligands engage KRAS(OFF) and KRAS(ON) with equal affinity, exploit a single-residue difference among isoforms to bind KRAS selectively, and successfully inhibit oncogenic signaling in KRAS-mutant cells through a mechanism distinct from SII-P binders. Mirror-image screening directly afforded nanomolar peptide ligands stable toward cellular proteolysis and delivered probes targeting distinct epitopes not accessible by homochiral peptide-display methods. Together, these findings establish the CRB-P as a specifically druggable and mechanistically differentiated site on KRAS with potential for combination with emerging RAS-targeting therapies and substantiate mirror-image mRNA display as a strategy for discovering stable all-D macrocyclic peptides targeting previously inaccessible epitopes on challenging targets.

cancer biology↗

bioRxiv2026

Structural basis for Ca2+-dependent, RyR2-selective stabilization by Ryanozole in CPVT

The cardiac ryanodine receptor (RyR2) is a Ca{superscript 2} release channel essential for excitation-contraction coupling. RyR2 mutations cause severe arrhythmogenic disorders, including catecholaminergic polymorphic ventricular tachycardia (CPVT), through gain-of-function (GOF) effects leading to aberrant Ca{superscript 2} release. We have recently developed Ryanozole, a potential therapeutic compound for CPVT, which selectively stabilizes RyR2 in a Ca2+-dependent manner. Here, we define the mechanism of action of Ryanozole by combining high-resolution cryo-electron microscopy, targeted mutagenesis and functional assays. Ryanozole binds to the interface between the Ca{superscript 2}-binding site and the pore-forming S6 helix, interfering with conformational changes required for Ca{superscript 2}-induced channel opening at low Ca2+. We identified key residues for the binding and isoform-specific modulation of Ryanozole. Notably, Ryanozole-bound RyR2 retains its ability to open at high Ca2+ via unique conformational changes. These findings provide a structural basis for CPVT-targeted therapy and redefine the paradigm of small molecule-based regulation of large ion channels.

biochemistry↗

bioRxiv2026

A macrocyclic peptide-based fusion inhibitor targeting SARS-CoV-2 Spike S2 subunit

Continuous emergence of SARS-CoV-2 variants carrying mutations in Spike presents a significant challenge for durable antiviral agents. Here we screen for random 13-amino acid non-mimetic macrocyclic peptides that bind to Spike and identify PA-001 that inhibits SARS-CoV-2 infection with high potency at 0.23-2.9 nM as 50% inhibitory concentration (IC50). PA-001 bound to Spike S2 subunit and inhibited the membrane fusion during virus entry. Through drug-resistant selection, we revealed that PA-001 targeted the fusion peptide proximal region (FPPR) in S2, which has not been recognized as a drug target to date. Consistent with its highly conserved amino acid sequences beyond strains, PA-001 exhibited broad antiviral activity against all tested SARS-CoV-2 variants, in contrast to clinically-approved S1-targeting antibodies that lost activity to Omicron variants. PA-001 suppressed SARS-CoV-2 propagation and disease progression in mouse- and hamster-infection models, both by administration prophylactically and therapeutically. Combination therapy with remdesivir further enhanced antiviral profiles. In clinical phase-I trial, PA-001 was well-tolerated and showed high systemic exposure, with 4,300-10,300-fold concentration of IC50 as maximum plasma concentration by single administration to healthy volunteers. These evidence propose FPPR as an unexpected antiviral drug target accessible by macrocyclic peptides and identify PA-001 as a potent anti-SARS-CoV-2 fusion inhibitor.

microbiology↗

bioRxiv2025

Developmental changes in the morphology and three-dimensional arrangement of antennal hair plates in crickets

Antennae of insects are essential mechanosensory organs that facilitate active tactile exploration and spatial navigation. Hair-plate sensilla at the base of the antenna flagellum provide proprioceptive inputs to detect their position. In hemimetabolous insects, such as crickets, the first instar immediately after hatching also possesses antennae, but the developmental dynamics and spatial organization of antennal hair plates remain poorly understood. Here, we present a comprehensive three-dimensional analysis of the antennal hair plates in crickets across developmental stages, from the first instar to adulthood. Using scanning electron microscopy and confocal laser scanning microscopy, we demonstrated that hair plate sensilla were present from the first instars and maintained a highly stereotyped spatial arrangement throughout development. Three-dimensional quantification revealed that new sensilla added during molting were formed at specific sites within the hair plate clusters that had existed at the previous stage, maintaining the spatial pattern despite the substantial growth of the antenna. Multidimensional analyses indicated that the spatial arrangement of sensilla was consistent across individuals, suggesting that organization was genetically determined. Retrograde labeling of sensory afferents showed that sensory neurons in the hair plates converged their axons, extended axon collaterals into the ipsilateral region of the antennal mechanosensory and motor center, and ultimately projected to the subesophageal ganglion. There was no apparent difference in their projection site among hair plates, suggesting no evidence of topographic organization. Our findings highlight the conserved spatial organization of hair-plate sensilla in crickets, suggesting a robust proprioceptive system that provides reliable feedback of antennal position throughout development.

neuroscience↗

bioRxiv2024

Biochemical evidence for the diversity of LHCI proteins in PSI-LHCI from the red alga Galdieria sulphuraria NIES-3638

Red algae are photosynthetic eukaryotes whose light-harvesting complexes (LHCs) associate with photosystem I (PSI). In this study, we examined characteristics of PSI-LHCI, PSI, and LHCI isolated from the red alga Galdieria sulphuraria NIES-3638. The PSI-LHCI supercomplexes were purified using anion-exchange chromatography followed by hydrophobic interaction chromatography, and finally by trehalose density gradient centrifugation. PSI and LHCI were similarly prepared following the dissociation of PSI-LHCI with Anzergent 3-16. Polypeptide analysis of PSI-LHCI revealed the presence of PSI and LHC proteins, along with a red-lineage chlorophyll a/b-binding-like protein (RedCAP), which is distinct from LHC proteins within the LHC protein superfamily. RedCAP, rather than LHC proteins, exhibited tight binding to PSI. Carotenoid analysis of LHCI identified zeaxanthin, {beta}-cryptoxanthin, and {beta}-carotene, with zeaxanthin particularly enriched, which is consistent with other red algal LHCIs. A Qy peak of chlorophyll a in the LHCI absorption spectrum was blue-shifted compared with those of PSI-LHCI and PSI, and a fluorescence emission peak was similarly shifted to shorter wavelengths. Based on these results, we discuss the diversity of LHC proteins, including RedCAP, in red algal PSI-LHCI supercomplexes.

plant biology↗

bioRxiv2024

Structural basis for molecular assembly of fucoxanthin chlorophyll a/c-binding proteins in a diatom photosystem I supercomplex

Photosynthetic organisms exhibit remarkable diversity in their light-harvesting complexes (LHCs). LHCs are associated with photosystem I (PSI), forming a PSI-LHCI supercomplex. The number of LHCI subunits, along with their protein sequences and pigment compositions, has been found to differ greatly among the PSI-LHCI structures. However, the mechanisms by which LHCIs recognize their specific binding sites within the PSI core remain unclear. In this study, we determined the cryo-electron microscopy structure of a PSI supercomplex incorporating fucoxanthin chlorophyll a/c-binding proteins (FCPs), designated as PSI-FCPI, isolated from the diatom Thalassiosira pseudonana CCMP1335. Structural analysis of PSI-FCPI revealed five FCPI subunits associated with a PSI monomer; these subunits were identified as RedCAP, Lhcr3, Lhcq10, Lhcf10, and Lhcq8. Through structural and sequence analyses, we identified specific protein-protein interactions at the interfaces between FCPI and PSI subunits, as well as among FCPI subunits themselves. Comparative structural analyses of PSI-FCPI supercomplexes, combined with phylogenetic analysis of FCPs from T. pseudonana and the diatom Chaetoceros gracilis, underscore the evolutionary conservation of protein motifs crucial for the selective binding of individual FCPI subunits. These findings provide significant insights into the molecular mechanisms underlying the assembly and selective binding of FCPIs in diatoms.

plant biology↗

bioRxiv2023

SHARPIN enhances ferroptosis in synovial sarcoma cells via NF-κB- and PRMT5-mediated PGC1α reduction

Sarcoma is a rare type of cancer for which new therapeutic agents are required. Ferroptosis is a nonapoptotic cell death triggered by iron-mediated lipid peroxidation. We found that transferrin receptor 1 (TFRC), an iron uptake protein, was expressed at higher levels in sarcoma cell lines than in noncancer and carcinoma cell lines. Glutathione peroxidase 4 (GPX4) protects cells against ferroptosis, and its inhibition using RAS-selective lethal 3 (RSL3) had an antitumor effect that was more pronounced in sarcoma cell lines, particularly synovial sarcoma cells, than in non-sarcoma cells. Because NF-{kappa}B can provoke ferroptosis, we examined the role of SHARPIN, an activator of NF-{kappa}B, in sarcoma. We found that SHARPIN expression is significantly associated with reduced survival in cohorts of patients with cancer, including sarcoma. In addition, SHARPIN promotes the sensitivity of sarcoma cells to ferroptosis. Further analyses revealed that the PGC1/NRF2/SLC7A11 axis and BNIP3L/NIX-mediated mitophagy are regulated through NF-{kappa}B and PRMT5 downstream of SHARPIN. Our findings suggest that ferroptosis could have a therapeutic effect in sarcoma, particularly in subpopulations with high TFRC and SHARPIN expression.

cancer biology↗

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↗

bioRxiv2021

Microtubule inhibitors enhance DNA transfection efficiency through autophagy receptor p62/SQSTM1

Ectopic gene expression is an indispensable tool in biology and medicine, but is often limited by the low efficiency of DNA transfection. We previously reported that depletion of the autophagy receptor p62/SQSTM1 enhances DNA transfection efficiency by preventing the degradation of transfected DNA. Therefore, p62 is a potential target for drugs to increase transfection efficiency. To identify such drugs, a non-biased high-throughput screening was applied to over 4,000 compounds from the Osaka University compound library, and their p62-dependency was evaluated. The top-scoring drugs were mostly microtubule inhibitors, such as colchicine and vinblastine, and all of them showed positive effects only in the presence of p62. To understand the p62-dependent mechanisms, the time required for p62-dependent ubiquitination, which is required for autophagosome formation, was examined using polystyrene beads that were introduced into cells as materials that mimicked transfected DNA. Microtubule inhibitors caused a delay in ubiquitination. Furthermore, the level of phosphorylated p62 at S405 was markedly decreased in the drug-treated cells. These results suggest that microtubule inhibitors inhibit p62-dependent autophagosome formation. Our findings demonstrate for the first time that microtubule inhibitors suppress p62 activation as a mechanism for increasing DNA transfection efficiency and provide solutions to increase efficiency.

cell biology↗

bioRxiv2021

Action selection based on multiple-stimulus aspects in the wind-elicited escape behavior of crickets

Animals detect approaching predators via sensory inputs through various modalities and immediately show an appropriate behavioral response to survive. Escape behavior is essential to avoid the predators attack and is more frequently observed than other defensive behaviors. In some species, multiple escape responses are exhibited with different movements. It has been reported that the approaching speed of a predator is important in choosing which escape action to take among the multiple responses. However, it is unknown whether other aspects of sensory stimuli, that indicate the predators approach, affect the selection of escape responses. We focused on two distinct escape responses (running and jumping) to a stimulus (short airflow) in crickets and examined the effects of multiple stimulus aspects (including the angle, velocity, and duration) on the choice between these escape responses. We found that the faster and longer the airflow, the more frequently the crickets jumped, meaning that they could choose their escape response depending on both velocity and duration of the stimulus. This result suggests that the neural basis for choosing escape responses includes the integration process of multiple stimulus parameters. It was also found that the moving speed and distance changed depending on the stimulus velocity and duration during running but not during jumping, suggesting higher adaptability of the running escape. In contrast, the movement direction was accurately controlled regardless of the stimulus parameters in both responses. The escape direction depended only on stimulus orientation, but not on velocity and duration. Summary statementWhen air currents triggering escape are faster and longer, crickets more frequently jump than run. Running speed and distance depend on stimulus velocity and duration, but direction control is independent.

animal behavior and cognition↗

bioRxiv2021

Sense of absence: Spatial perception through active sensing by insect antennal mechanosensory system

Animals perceive their surroundings by using various modalities of sensory inputs to navigate their locomotion. Nocturnal insects such as crickets use mechanosensory inputs mediated by their antennae to navigate under dark conditions. Active sensing with voluntary antennal movements improves spatial information, but it remains unclear how accurately the insects can perceive the surrounding space by using their antennal system. Crickets exhibit escape behavior in response to a short air-puff, which is detected by the abdominal mechanosensory organ called cerci and is perceived as a "predator approach" signal. We placed objects of different shapes at different locations with which the cricket actively made contact using its antenna. We then examined the effects on wind-elicited escape. The crickets changed their movement trajectory depending on the shape and location of the objects so that they could avoid collision with these obstacles even when the escape behavior was triggered by another modality of stimulus. For instance, when a wall was placed in front of the crickets so that it was detected by one side of their antenna, the escape trajectory in response to a stimulus from behind was significantly biased toward the side opposite the wall. However, if the antenna on the free side without the wall was ablated, this modulation to avoid collision diminished, suggesting that the antenna on the free side provided information of "absence" of obstacles. This study demonstrated that crickets were able to perceive spatial information, including the presence or absence of objects by active sensing with their antennal system. Summary StatementCrickets can acquire spatial information such as shape, location and orientation of objects through active sensing by antennal mechanosensory system, which also provides information about the absence of objects.

animal behavior and cognition↗

bioRxiv2020

Gating mechanism of cardiac ryanodine receptor 2 upon calcium ion binding

Cardiac ryanodine receptor (RyR2) is a large Ca2+ release channel in the sarcoplasmic reticulum and indispensable for excitation-contraction coupling in the heart. RyR2 is activated by Ca2+ and RyR2 mutations are implicated in severe arrhythmogenic diseases. Yet, the structural basis underlying channel opening and how mutations affect the channel remain unknown. Here, we addressed gating mechanism of RyR2 by combining high-resolution structures determined by cryo-electron microscopy with quantitative functional analysis of channels carrying various mutations in specific residues. We demonstrated two fundamental mechanisms for channel gating: interactions close to the channel pore stabilize the channel to prevent hyperactivity and a series of interactions in the surrounding regions is necessary for channel opening upon Ca2+ binding. Mutations at the residues involved in the former and the latter mechanisms cause gain-of-function and loss-of-function, respectively. Our results reveal gating mechanisms of the RyR2 channel and alterations by pathogenic mutations at the atomic level.

biochemistry↗

bioRxiv2020

Origin and evolution of the Haustoriidae (Amphipoda): A eulogy for the Haustoriidira

Haustoriid amphipods, despite their ubiquity in coastal sand or mud, have received little recent attention and their systematics and phylogenetics are largely unresolved. Some efforts have been made at classifying the family within the broader Amphipoda, but there is persistent incongruence in its placement among different authors and techniques. Furthermore, there exists no phylogenetic hypothesis of intrafamilial relationships despite the potential for rich biogeographic information to be gained given the specific habitat requirements of haustoriids and their limited dispersal abilities. In this work, we evaluate the competing hypotheses on the phylogenetic position of the Haustoriidae within Amphipoda by examining new and previously published sequences of nearly 100 species across 38 families. We find strong support for the Haustoriidae as basal gammarids, and that other families placed within the parvorder "Haustoriidira" are spread across Amphipoda. The radiation began during the Eocene and may have been driven in North America by the rapid filling of a coastal niche opened by the Chesapeake Bay impact crater. Unlike previous work, we find that the Pacific-endemic genus Eohaustorius is the most basal haustoriid, and that it separated from the rest of the family ~31 Mya. Finally, based on ancestral reconstructions, we provide taxonomic recommendations for relationships within Haustoriidae, including the elevation of a new genus, Cryptohaustorius. We conclude by recommending that the "Haustoriidira" be abandoned.

zoology↗

bioRxiv2020

Novel inhibitors of E. coli lipoprotein diacylglyceryl transferase are insensitive to resistance caused by lpp deletion

Lipoprotein diacylglyceryl transferase (Lgt) catalyzes the first step in the biogenesis of Gram-negative bacterial lipoproteins which play crucial roles in bacterial growth and pathogenesis. We demonstrate that Lgt depletion in a clinical uropathogenic Escherichia coli strain leads to permeabilization of the outer membrane and increased sensitivity to serum killing and antibiotics. Importantly, we identify the first ever described Lgt inhibitors that potently inhibit Lgt biochemical activity in vitro and are bactericidal against wild-type Acinetobacter baumannii and E. coli strains. Unlike inhibition of other steps in lipoprotein biosynthesis, deletion of the major outer membrane lipoprotein, lpp, is not sufficient to rescue growth after Lgt depletion or provide resistance to Lgt inhibitors. Our data validate Lgt as a novel druggable antibacterial target and suggest that inhibition of Lgt may not be sensitive to one of the most common resistance mechanisms that invalidate inhibitors of downstream steps of bacterial lipoprotein biosynthesis and transport.

microbiology↗

bioRxiv2020

Individual identification of Japanese macaques (Macaca fuscata) using a face recognition system and a limited number of learning images

Individual identification is an important technique in animal research that requires researcher training and specialized skillsets. Face recognition systems using artificial intelligence (AI) deep learning have been put into practical use to identify in humans and animals, but a large number of annotated learning images are required for system construction. In wildlife research cases, it is difficult to prepare a large amount of learning images, which may be why systems using AI have not been widely used in field research. To investigate the development of a system that identifies individuals using a small number of learning images, we constructed a system to identify individual Japanese macaques (Macaca fuscata yakui) with a low error rate from an average of 20 images per individual. The characteristics of this system were augmentation of data, simultaneous determination by four individual identification models and identification from a majority of five frames to ensure reliability. This technology has a high degree of utility for various stakeholders and it is expected that it will advance the development of individual identification systems by AI that can be widely used in field research.

zoology↗

bioRxiv2017

Rapid degradation and 3D CLEM of condensin reveal chromatin compaction uncoupled from chromosome architecture in mitosis

The requirement for condensin in chromosome formation in somatic cells remains unclear as imperfectly condensed chromosomes do form in conventional condensindepleted cells. Here we have dissected the role of condensin at different stages of vertebrate mitosis by combining auxin-mediated rapid depletion of condensin subunit SMC2 with chemical genetics to obtain near-synchronous mitotic entry of chicken DT40 cells. We analysed the outcomes by live and fixed-cell microscopy methods, including 3D correlative light and serial block face scanning electron microscopy. Following rapid depletion of condensin, chromosomal defects were obvious. The chromatin was compacted normally, but formed a single mass of mitotic chromosomes clustered at one side of a bent mitotic spindle. Cultures arrest at prometaphase, eventually exiting mitosis without segregating chromosomes. Experiments titrating the auxin concentration suggest a previously unsuspected dual role of condensin, as different condensin levels are required for anaphase chromosome segregation and formation of a normal chromosome architecture.

cell biology↗