Biology subjects

Sugawara, T.

Publications and source records attributed to Sugawara, T..

10 recordsLinked to original sources

bioRxiv2025

Retinoic acid-responsive hox genes in hoxba and hoxbb clusters direct pharyngeal pouch formation in zebrafish

The segmented pharyngeal apparatus is crucial for organ development specific to vertebrates, and its formation relies on the proper development of pharyngeal pouches. While retinoic acid (RA) is known to influence pouch formation, the downstream genes involved have been unclear. In this study, we demonstrate that zebrafish mutants lacking both the hoxba and hoxbb clusters--teleost-specific duplicates of the ancestral HoxB cluster--exhibit a significant loss of posterior pharyngeal pouches and related skeletal elements. This phenotype resembles that observed in raldh2 and pax1a;pax1b mutants. We identify hoxb1a and hoxb1b as RA-dependent genes expressed in the pharyngeal region that are essential for pouch formation. Morpholino-mediated knockdown of these genes replicated the pouch defects and decreased pax1a expression, indicating a regulatory pathway linking RA, Hox, and pax1. Our findings uncover a previously unrecognized role of Hox genes in early pouch segmentation and suggest that RA-responsive HoxB clusters were co-opted during vertebrate evolution to initiate pharyngeal regionalization.

developmental biology↗

bioRxiv2024

Claudin-11 regulates immunological barrier formation and spermatogonial proliferation through stem cell factor

Tight junctions (TJs) between adjacent Sertoli cells are believed to form immunological barriers that protect spermatogenic cells expressing autoantigens from autoimmune responses. However, there is no direct evidence that Sertoli cell TJs (SCTJs) do indeed form immunological barriers. Here, we analyzed male mice lacking claudin-11 (Cldn11), which encodes a SCTJ component, and found autoantibodies against antigens of spermatocytes/spermatids in their sera. Defective spermatogenesis was not restored in Cldn11-deficient mice on a genetic background mimicking a severely impaired adaptive immune system. This suggests that defective spermatogenesis is not caused by autoimmune responses against spermatogenic cells. Further analyses showed that Cldn11 knockout impaired Sertoli cell polarization, localization of stem cell factor (SCF) (a key molecule for maintaining differentiating spermatogonia) to the basal compartment of seminiferous tubules, and also proliferation of differentiating spermatogonia. We propose that CLDN11 creates a microenvironment for SCF-mediated spermatogonial proliferation at the basal compartment via Sertoli cell polarization.

developmental biology↗

bioRxiv2023

DNA methylation signature in NSD2 loss-of-function variants appeared similar to that in Wolf-Hirschhorn syndrome

PurposeWolf-Hirschhorn syndrome (WHS), a contiguous gene syndrome caused by the hemizygous deletion of the distal short arm of chromosome 4 where NSD2 is, reportedly exhibits specific DNA methylation signatures in peripheral blood cells. However, responsible genomic loci for signatures are unreported. The objective of the study is to define the loci of WHS-related DNA methylation signatures and to explore the role of NSD2 for the signatures. MethodsWe conducted genome-wide methylation analysis of individuals with WHS or NSD2 variants using array. We studied genome-edited knock in mice or induced pluripotent stem cells to explore the function of NSD2 variants which are observed in congenital anomaly cases. ResultsThree undiagnosed cases with NSD2 variants showed WHS-related DNA methylation signatures. These variants were validated to be NSD2 loss-of-function in induced pluripotent stem cells or genome-edited knock-in mice. p.Pro905Leu variant decreased Nsd2 protein levels, and changed Histone H3-Lysine 36 demethylation levels in similar way in the same genomic regions as Nsd2 knock out mice regulated. Nsd2 knock out mice exhibited common DNA methylation changes. ConclusionThese results revealed that WHS-related DNA methylation signatures are dependent on NSD2 dysfunction and are useful in diagnosing NSD2 variants of unknown significance.

genetics↗

bioRxiv2021

Chemophoresis Engine: Universal Principle of ATPase-driven Cargo Transport

Cell polarity regulates the orientation of the cytoskeleton members that directs intracellular transport for cargo-like organelles, using chemical gradients sustained by ATP or GTP hydrolysis. However, how cargo transports are directly mediated by chemical gradients remains unknown. We previously proposed a physical mechanism that enables directed movement of cargos, referred to as chemophoresis. According to the mechanism, a cargo with reaction sites is subjected to a chemophoresis force in the direction of the increased concentration. Based on this, we introduce an extended model, the chemophoresis engine, as a general mechanism of cargo motion, which transforms chemical free energy into directed motion through the catalytic ATP hydrolysis. We applied the engine to plasmid motion in a parABS system to demonstrate the the self-organization system for directed plasmid movement and pattern dynamics of ParA-ATP concentration, thereby explaining plasmid equi-positioning and pole-to-pole oscillation observed in bacterial cells and in vitro experiments. We mathematically show the existence and stability of the plasmid-surfing pattern, which allows the cargo-directed motion through the symmetry-breaking transition of the ParA-ATP spatiotemporal pattern. Finally, based on its generality, we discuss the chemophoresis engine as a universal principle of hydrolysis-driven intracellular transport. SIGNIFICANCEThe formation of organelle/macromolecule patterns depending on chemical concentration under non-equilibrium conditions, first observed during macroscopic morphogenesis (1), has recently been observed at the intracellular level as well, and its relevance as intracellular morphogen has been demonstrated in the case of bacterial cell division. These studies have discussed how cargos maintain positional information provided by chemical concentration gradients/localization. However, how cargo transports are directly mediated by chemical gradients remains unknown. Based on the previously proposed mechanism of chemotaxis-like behavior of cargos (referred to as chemophoresis, (2, 3)), we introduce a chemophoresis engine as a physicochemical mechanism of cargo motion, which transforms chemical free energy to directed motion. The engine is based on the chemophoresis force to make cargoes move in the direction of the increasing ATPase(-ATP) concentration and an enhanced catalytic ATPase hydrolysis at the positions of the cargoes. Applying the engine to ATPase-driven movement of plasmid-DNAs in bacterial cells, we constructed a mathematical model to demonstrate the self-organization for directed plasmid motion and pattern dynamics of ATPase concentration, as is consistent with in vitro and in vivo experiments. We propose that this chemophoresis engine works as a universal principle of hydrolysis-driven intracellular transport.

biophysics↗

bioRxiv2021

Early Dynamics of Chronic Myeloid Leukemia on Nilotinib Predicts Deep Molecular Response

Chronic myeloid leukemia (CML) is a myeloproliferative disorder caused by the BCR-ABL1 tyrosine kinase1,2. ABL1-selective tyrosine kinase inhibitors (TKIs) including nilotinib have dramatically improved the prognosis of patients with CML3-7. The ultimate goal of CML treatment is likely to be TKI-free maintenance of deep molecular response (DMR), which is defined by quantitative measurement of BCR-ABL1 transcripts on the international scale (IS)8, and durable DMR is a prerequisite to reach this goal9. Thus, an algorithm to enable the early prediction of DMR achievement on TKI therapy is quite valuable. Here, we show that our mathematical framework based on a clinical trial dataset10 can accurately predict the response to frontline nilotinib. We found that our simple dynamical model can predict nilotinib response by using two common laboratory findings (observation values): IS11,12 and white blood cell (WBC) count. Furthermore, our proposed method identified patients who failed to achieve DMR with high fidelity according to the data collected only at three initial time points during nilotinib therapy. Since our model relies on the general properties of TKI response, our framework would be applicable to CML patients who receive frontline nilotinib or other TKIs in clinical practice. Significance StatementChronic myeloid leukemia (CML) is a myeloproliferative disorder caused by the BCR-ABL1 tyrosine kinase. The goal of this treatment is the sequential achievement of deep molecular response (DMR). Tyrosine kinase inhibitors (TKIs) are effective in the reduction because they inhibit CML cell proliferation. However, because of individual differences in the TKI efficacy, some patients are unable to achieve DMR, so that early prediction of DMR reachability is necessary for personalized medicine. By combining time series analysis and mathematical modeling, we developed a mathematical method that accurately predicts patients who do not achieve DMR in the early stages of TKI administration. Our prediction method gives a basis of effective personalized treatments for CML patients.

systems biology↗

bioRxiv2020

Polygamous breeding system identified in the distylous genus Psychotria: P. manillensis in the Ryukyu archipelago, Japan

Distyly is a genetic polymorphism composed of long- and short-styled flowers in a population. The evolutionary breakdown of distyly has been reported in many taxa, and mainly involves a shift toward monomorphism or dioecism. However, a shift toward monoecism has not been reported in distylous species. Psychotria (Rubiaceae), one of the world largest genera, consists of distylous species and their derivatives. In our preliminary study, however, we identified some monoecious individuals in a population of Psychotria manillensis. To understand the breeding system and reproductive biology of P. manillensis, we investigated floral traits, open fruit set, and flower visitors, and performed hand pollination and bagging experiments in five populations of Okinawa and Iriomote islands, Ryukyu Islands, Japan. The populations of P. manillensis were composed mainly of monoecious individuals (54%), followed by female (30%), male (14%), and hermaphroditic (2%) individuals at the time of flower collection. Of the collected flowers, 93% were functionally unisexual (male or female), whereas only 6.5% were perfect (hermaphroditic). However, some individuals changed sex mainly towards increasing femaleness during the flowering period. Moreover, 35% of the studied plants changed their sexual expression over the years. P. manillensis showed self-compatibility and no agamospermy. The fruit set under open pollination varied among populations and years (1.8-21.9%), but it was significantly higher than that of auto-selfing (0.68-1.56%). Wasps and flies were the main flower visitors and probably the main pollinators of the species. In conclusion, P. manillensis was revealed to be polygamous, involving monoecious, female, male, and hermaphroditic individuals. This is the first report of the polygamous breeding system not only in the genus Psychotria, but also in all heterostylous taxa.

ecology↗

bioRxiv2020

Angulin-1 seals tricellular contacts independently of tricellulin and claudins

Tricellular tight junctions (tTJs) are specialized tight junctions (TJs) that seal the intercellular space at tricellular contacts (TCs), where the vertices of three epithelial cells meet. Tricellulin and angulin family membrane proteins are known constituents of tTJs, but the molecular mechanism of tTJ formation remains elusive. Here, we investigated the roles of angulin-1 and tricellulin in tTJ formation in MDCK II cells by genome editing. Angulin-1-deficient cells lost the plasma membrane contact at TCs with impaired epithelial barrier function. The COOH-terminus of angulin-1 bound to the TJ scaffold protein ZO-1 and disruption of their interaction influenced the localization of claudins at TCs, but not the tricellular sealing. Strikingly, the plasma membrane contact at TCs was formed in tricellulin- or claudin-deficient cells. These findings demonstrate that angulin-1 is responsible for the plasma membrane seal at TCs independently of tricellulin and claudins.

cell biology↗

bioRxiv2019

Microrheology for Hi-C Data Reveals the Spectrum of the Dynamic 3D Genome Organization

The 1-dimensional information of genomic DNA is hierarchically packed inside the eukaryotic cell nucleus and organized in 3-dimensional (3D) space. Genome-wide chromosome conformation capture (Hi-C) methods have uncovered the 3D genome organization and revealed multiscale chromatin domains of compartments and topologically associating domains (TADs). Moreover, single-nucleosome live-cell imaging experiments have revealed the dynamic organization of chromatin domains caused by stochastic thermal fluctuations. However, the mechanism underlying the dynamic regulation of such hierarchical and structural chromatin units within the micro-scale thermal medium remains unclear. Microrheology is a way to measure dynamic viscoelastic properties coupling between thermal microenvironment and mechanical response. Here, we propose, to our knowledge, a new microrheology for Hi-C data to analyze the compliance property as a barometer of rigidness and flexibility of genomic regions along with the time evolution. Our method allows conversion of a Hi-C matrix into the spectrum of the rheological property along the genomic coordinate of a single chromosome. To demonstrate the technique, we analyzed Hi-C data during the neural differentiation of mouse embryonic stem cells. We found that TAD boundaries behave as more rigid nodes than the intra-TAD region. The spectrum clearly shows the rheological property of the dynamic chromatin domain formations at an individual time scale. Furthermore, we characterized the appearance of synchronous and liquid-like inter-compartment interactions in differentiated cells. Together, our microrheology provides physical insights revealing the dynamic 3D genome organization from Hi-C data.\n\nSIGNIFICANCEGenomic DNA is hierarchically packed inside the eukaryotic cell nucleus, and the genome organization in 3D contributes to proper genome functions at the multiscale chromatin domains. Although thermal fluctuations inevitably drive movements of the genome molecules in the micro-scale cell environment, there is no method, as yet, to quantify such dynamic 3D genome organization of hierarchical and structural chromatin units. Here, we describe a method to calculate rheological properties as barometers of flexibility and liquid-like behavior of genomic regions. We show that biologically relevant boundaries between chromatin domains are more rigid than the inside at a particular time scale. Our method allows interpretation of static and population-averaged genome conformation data as dynamic and hierarchical 3D genome picture.

biophysics↗

bioRxiv2019

PHi-C: deciphering Hi-C data into polymer dynamics

Computational modelling methods for Hi-C data have revealed averaged and static features of the 3D genome in cell nuclei. Here, we describe a 4D simulation method, PHi-C (Polymer dynamics deciphered from Hi-C data), that depicts dynamic 3D genome features through polymer modelling. This method allows for demonstrations of dynamic characteristics of genomic loci and chromosomes, as observed in live-cell imaging experiments, and provides physical insights into Hi-C data.

bioinformatics↗

bioRxiv2018

Epigenetic-scale comparison of human iPSCs generated by retrovirus, Sendai virus or episomal vectors

Human induced pluripotent stem cells (iPSCs) are established by introducing several reprogramming factors, such as OCT3/4, SOX2, KLF4, c-MYC. Because of their pluripotency and immortality, iPSCs are considered to be a powerful tool for regenerative medicine. To date, iPSCs have been established all over the world by various gene delivery methods. All methods induced high-quality iPSCs, but epigenetic analysis of abnormalities derived from differences in the gene delivery methods has not yet been performed. Here, we generated genetically matched human iPSCs from menstrual blood cells by using three kinds of vectors, i.e., retrovirus, Sendai virus, and episomal vectors, and compared genome-wide DNA methylation profiles among them. Although comparison of aberrant methylation revealed that iPSCs generated by Sendai virus vector have lowest number of aberrant methylation sites among the three vectors, the iPSCs generated by non-integrating methods did not show vector-specific aberrant methylation. However, the differences between the iPSC lines were determined to be the number of random aberrant hyper-methylated regions compared with embryonic stem cells. These random aberrant hyper-methylations might be a cause of the differences in the properties of each of the iPSC lines.

cell biology↗