Biology subjects

Taguchi, S.

Publications and source records attributed to Taguchi, S..

8 recordsLinked to original sources

bioRxiv2026

Oncogenic NPM-ALK reprograms the TGM1 1 interactome toward oncogenic signaling and transcriptional states

Oncogenic NPM-ALK drives aberrant signaling networks that promote malignant phenotypes; however, the molecular mechanisms linking oncogenic signaling to downstream cellular programs remain incompletely understood. Among candidate regulatory factors, transglutaminase 1 (TGM1) has not been functionally characterized in this context. Here, we investigated the role of TGM1 in NPM-ALK-expressing cells by combining proximity-dependent proteomics with functional analyses. Using a TurboID- based approach, we mapped the TGM1-associated protein network and identified extensive remodeling of this network upon NPM-ALK expression. Proteomic analyses revealed that NPM-ALK reduced TGM1-associated proteins involved in genome maintenance and DNA repair, while enhancing associations with proteins linked to cytoplasmic translation and PI3K-AKT signaling pathways. Consistent with these findings, TGM1 deficiency impaired cell proliferation without significantly affecting cell viability, indicating a specific role in maximal proliferative capacity. Furthermore, proteomic and functional analyses suggested a link between TGM1 and AKT signaling pathways. Together, these findings suggest that oncogenic NPM-ALK reprograms the TGM1 interactome toward oncogenic signaling and transcriptional states, positioning TGM1 within signaling networks associated with proliferative cellular phenotypes.

molecular biology↗

bioRxiv2025

Deep Learning Pipeline for Segmentation and Quantification of Overlapping Membranes in 2D Image

Quantitative morphological analysis is crucial for understanding cellular processes. While 3D Z-stack imaging offers high-resolution data, the complexity of 3D structures makes direct interpretation and manual annotation challenging and time-consuming, especially for large datasets. Maximum Intensity Projection (MIP) is a common strategy to create more interpretable 2D representations, but this inevitably leads to artificial overlaps between structures, significantly hindering accurate automated segmentation of individual instances by conventional methods or standard deep learning tools. To address this critical challenge in 2D projection analysis, we developed DeMemSeg, a deep learning pipeline based on Mask R-CNN, specifically designed to segment overlapping membrane structures, called prospore membranes (PSMs) during yeast sporulation. DeMemSeg was trained on a custom-annotated dataset, leveraging a systematic image processing workflow. Our optimized model accurately identifies and delineates individual, overlapping PSMs, achieving segmentation performance and derived morphological measurements that are statistically indistinguishable from expert manual annotation. Notably, DeMemSeg successfully generalized to segment PSMs from unseen data acquired from gip1{Delta} mutant cells, capturing the distinct morphological defects in PSMs. DeMemSeg thus provides a robust, automated solution for objective quantitative analysis of complex, overlapping membrane morphologies directly from widely used 2D MIP images, offering a practical tool and adaptable workflow to advance cell biology research.

cell biology↗

bioRxiv2022

Automation of yeast spot assays using an affordable liquid handling robot

The spot assay of the budding yeast Saccharomyces cerevisiae is an experimental method that is used to evaluate the effect of genotypes, medium conditions, and environmental stresses on cell growth and survival. Automation of the spot assay experiments from preparing a dilution series to spotting to observing spots continuously has been implemented based on large laboratory automation devices and robots, especially for high-throughput functional screening assays. However, there has yet to be an affordable solution for the automated spot assays suited to researchers in average laboratories and with high customizability for end-users. To make reproducible spot assay experiments widely available, we have automated the plate-based yeast spot assay of budding yeast using Opentrons OT-2 (OT-2), an affordable liquid-handling robot, and a flatbed scanner. We prepared a 3D-printed mount for the Petri dish to allow for precise placement of the Petri dish inside the OT-2. To account for the uneven height of the agar plates, which were made by human hands, we devised a method to adjust the z-position of the pipette tips which is based on the weight of each agar plate. During the incubation of the agar plates, a flatbed scanner was used to automatically take images of the agar plates over time, allowing researchers to quantify and compare the cell density within the spots at optimal time points a posteriori. Furthermore, the accuracy of the newly developed automated spot assay was verified by performing spot assays with human experimenters and the OT-2 and quantifying the yeast-grown area of the spots. This study will contribute to the introduction of automated spot assays and the automated acquisition of growth processes in conventional laboratories that are not adapted for high-throughput laboratory automation.

bioengineering↗

bioRxiv2022

Structural model of microtubule dynamics inhibition by Kinesin-4 from the crystal structure of KLP-12 - tubulin complex

Kinesin superfamily proteins are microtubule-based molecular motors driven by the energy of ATP hydrolysis. Among them, the kinesin-4 family is a unique motor that inhibits microtubule dynamics. Although mutations of kinesin-4 cause several diseases, its molecular mechanism is unclear because of the difficulty of visualizing the high-resolution structure of kinesin-4 working at the microtubule plus-end. Here, we report that KLP-12, a C. elegans kinesin-4 ortholog of KIF21A and KIF21B, is essential for proper length control of C. elegans axons, and its motor domain represses microtubule polymerization in vitro. The crystal structure of the KLP-12 motor domain complexed with tubulin, which represents the high-resolution structural snapshot of inhibition state of microtubule-end dynamics, revealed the bending effect of KLP-12 for tubulin. Comparison with the KIF5B-tubulin and KIF2C-tubulin complexes, which represent the elongation and shrinking forms of microtubule ends, respectively, showed the curvature of tubulin introduced by KLP-12 is in between them. Taken together, KLP-12 controls the proper length of axons by modulating the curvature of the microtubule ends to inhibit the microtubule dynamics.

biophysics↗

bioRxiv2021

CAMSAP2 organizes a γ-tubulin-independent microtubule nucleation centre

Microtubules are dynamic polymers consisting of {beta}-tubulin heterodimers. The initial polymerization process, called microtubule nucleation, occurs spontaneously via {beta}-tubulin. Since a large energy barrier prevents microtubule nucleation in cells, the {gamma}-tubulin ring complex is recruited to the centrosome to overcome the nucleation barrier. However, detachment of a considerable number of microtubules from the centrosome is known to contribute to fundamental processes in cells. Here, we present evidence that minus-end-binding calmodulin-regulated spectrin-associated protein 2 (CAMSAP2) serves as a strong nucleator for microtubule formation from soluble {beta}-tubulin independent of {gamma}-tubulin. CAMSAP2 significantly reduces the nucleation barrier close to the critical concentration for microtubule polymerization by stabilizing the longitudinal contacts among {beta}-tubulins. CAMSAP2 clusters together with {beta}-tubulin to generate nucleation intermediates, from which numerous microtubules radiate, forming aster-like structures. Our findings suggest that CAMSAP2 supports microtubule growth by organizing a nucleation centre as well as by stabilizing microtubule nucleation intermediates.

cell biology↗

bioRxiv2017

A randomized comparison of different hormone replacement protocols for thawed blastocyst transfer

BackgroundThere are no randomized controlled trials evaluating the pregnancy rates after thawed blastocyst transfers in patients treated with various hormone replacement regimens.\n\nMethodsA prospective randomized controlled trial was conducted to evaluate the outcomes in three different hormone replacement protocols for thawed blastocyst transfer. A total of 330 women (median age 38.2 years) who were undergoing IVF at our clinic were enrolled.\n\nResultsSerum estradiol (E2) levels were 267.71 pg/ml in Premarin group, 391.22 pg/ml in Estrogel group and 495.12 pg/ml in Estrana tape group. Therefore, serum E2 levels in Estrana tape group were higher than those of the other two groups (P<0.01). The pregnancy rate in the Estrogel group was higher than that in the Premarin group (30.0% versus 17.3%, P=0.026, odds ratio 2.05, 95% confidence interval: 1.09-3.87). Furthermore, the pregnancy rate in the Estrana tape group was higher than that in the Estrogel group (43.6% versus 30.0%, P=0.036, odds ratio 1.81, 95% confidence interval: 1.04-3.14).\n\nConclusionThe serum E2 levels contributed to differences observed in the pregnancy rate among the three different protocols. Thus, Estrana tape has an advantage as a hormone replacement protocol for thawed blastocyst transfer.

clinical trials↗

bioRxiv2017

Abnormally high levels of serum α-klotho result in a poor outcome for clinical pregnancy-A prospective cohort study-

BackgroundThe klotho protein has been extensively studied. However, there are no studies examining the association between serum alpha klotho levels and the clinical outcome of post-clinical pregnancy.\n\nMethodsWe conducted a prospective cohort study in 42 patients (median age 37.4 years) to evaluate the association between serum alpha klotho levels during the follicular phase of preimplantation and the clinical outcome data of post-clinical pregnancy. The patients provided informed consent at our clinic. The serum alpha klotho levels were evaluated using a human soluble alpha klotho assay kit. The fetal chromosomal abnormalities were investigated at our clinic. We also assessed the clinical outcomes of post-clinical pregnancies.\n\nResultsThe serum alpha klotho level during the follicular phase of preimplantation for non-pregnant women was 544.31 pg/ml (mean). The clinical pregnancy rate was 38.1%. There were chromosomal abnormalities observed in four unborn children (9.5%; Down syndrome, etc). The serum alpha klotho levels during the follicular phase of preimplantation in the chromosomal abnormalities group were higher than in the group without chromosomal abnormalities (P=0.029, abnormalities group 659.26 pg/ml [mean] versus control 530.23 pg/ml [mean]). A multiple logistic regression analysis showed the chromosomal abnormalities rates in unborn children were positively influenced by serum alpha klotho levels during the follicular phase of preimplantation (p=0.0008) and the patients age (p=0.008).\n\nConclusionPrevious studies have demonstrated that increased alpha klotho levels in human serum are positively correlated with health. However, abnormally high levels of serum alpha klotho during the follicular phase of preimplantation may predict a poor outcome for clinical pregnancy.

genetics↗

bioRxiv2017

Can the location of a trophectoderm biopsy contribute to human blastocyst development ?

The influence of the location of a trophectoderm biopsy in human blastocysts on the development of those blastocysts has not yet been investigated. In our prospective study (n=92), our multivariate logistic regression analysis indicated that blastocoel development was influenced by the location of the trophectoderm biopsy (p=0.049) and by the type of human blastocyst used (fresh or thawed) (p=0.037), regardless of the patients age (p=0.507) and the number of days for the human blastocyst in the pretrophectoderm biopsy (p=0.239). Therefore, when a trophectoderm biopsy is close to the inner cell mass (ICM) in human blastocysts, it improves the progress of blastocoel development.\n\nClinical evidence suggests that the progress of blastocoel development is a predictor of clinical outcomes after single blastocyst transfer. Therefore, when the trophectoderm biopsy is done from near the ICM, improvement of clinical outcomes after single blastocyst transfer may be expected.

developmental biology↗