Biology subjects

Kurosaka, H.

Publications and source records attributed to Kurosaka, H..

7 recordsLinked to original sources

bioRxiv2026

Retinoid signaling promotes frontonasal identity while repressing maxillary identity during craniofacial development

Vertebrate facial development depends on the correct specification of embryonic facial prominences, a process known to be governed by region-specific reciprocal signaling pathways between the craniofacial ectoderm, endoderm and mesenchyme. This process is further modulated by transcriptional and epigenetic mechanisms that regulate the expression of essential genes in mesenchymal progenitors derived from cranial neural crest cells. Retinoid signaling plays a critical role in facial development, and both gain-and loss-of-function results in a wide spectrum of facial defects, including orofacial clefts. In this study, we identified retinoid signaling as a critical regulator of cranial neural crest cell specification toward a frontonasal process identity in mice. Rdh10 oxidizes vitamin A (all-trans retinol) to retinal, which is a rate limiting step in the synthesis of retinoic acid. Rdh10 loss-of-function resulted in ectopic formation of whisker pads - a derivative of the maxillary process - within the frontonasal process region. This transformation was evidenced by the mis-expression of maxillary specific transcription factors including Meis2 and Lhx6 in the frontonasal mesenchyme. Furthermore, these transcription factors exhibited increased chromatin accessibility at their consensus binding sites following the loss of retinoid signaling in frontonasal cranial neural crest cells. These results indicate that retinoid signaling acts as a critical regulator specifying frontonasal identity and fate of cranial neural crest cells as they migrate into the frontonasal process, while concomitantly repressing maxillary process fate. These results not only advance our understanding of frontonasal prominence specification and the evolutionary development of craniofacial structures but also offers valuable insights into the etiology and pathogenesis of craniofacial malformations such as orofacial clefts.

developmental biology↗

bioRxiv2026

A Wnt-responsive fibrocartilage progenitor system coordinates postnatal mandibular condylar cartilage growth

Postnatal growth of the mandibular condyle requires coordinated expansion of fibrocartilage and production of chondrocytes, yet the cellular populations that organize this process remain incompletely defined. Here we identify a Wnt-responsive fibrocartilage progenitor population that contributes to postnatal mandibular condylar cartilage growth. Using a direct Wnt activity reporter (R26-WntVis), inducible genetic lineage tracing (Axin2CreERT2), and single-cell transcriptomics, we define a Wnt-enriched progenitor-like cluster localized predominantly within the fibrocartilage zone. Lineage tracing demonstrates that Axin2-lineage cells expand laterally within fibrocartilage and generate vertically aligned chondrocytes in the chondrocartilage compartment, indicating bidirectional growth contribution in vivo. Conditional ablation of {beta}-catenin in Axin2-lineage cells results in depletion of the fibrocartilage compartment and premature activation of chondrogenic differentiation programs, whereas constitutive {beta}-catenin activation disrupts compartmental organization without enhancing proliferation. Mechanistically, we identify Foxm1 as a Wnt-associated proliferative mediator enriched in fibrocartilage, and genetic reduction of Foxm1 cooperates with {beta}-catenin deficiency to impair condylar growth. In parallel, {beta}-catenin loss derepresses TGF-{beta}-Smad signaling and enhances chondrogenic differentiation, indicating that canonical Wnt activity coordinates proliferative maintenance while restraining lineage commitment within the same cellular compartment. Together, these findings identify a Wnt-responsive fibrocartilage progenitor system that regulates postnatal mandibular condylar cartilage growth by coupling Foxm1-associated proliferative maintenance with suppression of TGF-{beta}-dependent chondrogenic differentiation during temporomandibular joint development. Graphical abstractWnt-responsive fibrocartilage progenitors coordinate postnatal mandibular condylar cartilage growth through Foxm1-dependent proliferative maintenance and suppression of TGF-{beta}-driven chondrogenic differentiation.

developmental biology↗

bioRxiv2025

TMEM2 maintains hyaluronan turnover and cartilage homeostasis during early osteoarthritis progression

Osteoarthritis (OA) is a degenerative joint disease characterized by progressive disruption of the cartilage extracellular matrix (ECM), yet the molecular mechanisms governing ECM turnover during disease initiation remain incompletely defined. Hyaluronan (HA) is a major structural component of articular cartilage, and its regulated turnover is essential for maintaining tissue integrity. Transmembrane protein 2 (TMEM2) is a cell-surface hyaluronidase capable of degrading high-molecular weight HA under physiological conditions, but its role in joint tissues has remained unclear. Here, we examine the spatiotemporal expression and functional contribution of TMEM2 in articular cartilage using single-cell transcriptomic analysis, histological approaches, and a chondrocyte-specific conditional knockout mouse model. Under physiological conditions, Tmem2 was predominantly expressed in non-calcified articular chondrocytes. Following joint destabilization, Tmem2 expression was transiently increased during early osteoarthritis, coinciding with reduced cartilage HA content, consistent with altered HA turnover. Importantly, genetic ablation of Tmem2 in chondrocytes markedly exacerbated osteoarthritis progression, resulting in accelerated cartilage delamination, increased chondrocyte apoptosis, reduced proliferative activity, and enhanced hypertrophic differentiation. These changes occurred without detectable abnormalities in the synovium, subchondral bone, or osteophyte formation, indicating a cartilage-intrinsic phenotype. Collectively, these findings identify TMEM2 as an important regulator of hyaluronan homeostasis within the cartilage ECM and provide in vivo genetic evidence that TMEM2- dependent HA turnover contributes to the maintenance of articular cartilage integrity during osteoarthritis progression.

molecular biology↗

bioRxiv2024

Compromised actin dynamics underlie the orofacial cleft in Baraitser-Winter Cerebrofrontofacial Syndrome with a variant in ACTB

Craniofacial anomalies encompassing the orofacial cleft are associated with >30% of systemic congenital malformations. Baraitser-Winter Cerebrofrontofacial syndrome (BWCFF) is a rare genetic disorder attributed to variants in the actin beta (ACTB) or actin gamma genes that are correlated with a range of craniofacial abnormalities, including cleft lip and/or palate. The underlying pathological mechanism of BWCFF remains elusive, and it is necessary to investigate the etiology of orofacial clefts in BWCFF. In this study, we identified a missense variant (c.1043C>T: p.S348L) in the ACTB gene from a patient with BWCFF and concomitant cleft lip and palate. Furthermore, we performed functional assessments of this variant using various disease models such as the MDCK cell line and Xenopus laevis. These models revealed a compromised capacity of mutated ACTB to localize at the epithelial junction, consequently affecting the behavior of epithelial cells. Additionally, we made the noteworthy discovery that mutated ACTB exhibited an impaired ability to bind PROFILIN1, a critical factor in actin polymerization. This defective ability may contribute to the molecular etiology of aberrant epithelial cell adhesion and migration, resulting in orofacial cleft formation in BWCFF. Author SummaryAdvances in clinical sequencing have rapidly elucidated the genetic basis of rare diseases affecting multiple organs. In this study, we investigate the role of a variant in the ACTB gene, which is fundamental to cell structure, in causing orofacial clefts in the Baraitser-Winter Cerebrofrontofacial Syndrome (BWCFF), known for its distinctive facial anomalies. Using epithelial cell lines and frog embryo models, we investigated the effects of the ACTB gene alteration on cellular functions. Our results indicate that this genetic change disrupts the normal adhesion and movement of epithelial cells, processes that are critical for facial development. This research underscores the importance of the ACTB gene in shaping the face and suggests new directions for future research, potentially informing the diagnosis and treatment of BWCFF and related conditions.

genetics↗

bioRxiv2023

SLC26A2-mediated sulfate metabolism is essential for the tooth development

The sulfate transporter gene SLC26A2 is responsible for diastrophic dysplasia, which represents skeletal dysplasia in humans. This highlights the importance of sulfate metabolism in skeletal formation. SLC26A2-related chondrodysplasia is also known to exhibit abnormalities in craniofacial and tooth development. Although the function of SLC26A2 in mammals has been investigated using genetic mouse models, the essential role of SLC26A2 during craniofacial and tooth development has not been elucidated. In this study, we demonstrate the pivotal roles of SLC26A2-mediated sulfate metabolism during tooth development. Analysis of Slc26a2 expression reveals that it is predominantly expressed in dental tissues, including odontoblasts and ameloblasts, during tooth development. Slc26a2 knockout mice (Slc26a2-KO-{Delta}exon2) exhibit a retrognathic upper jaw, small upper incisors, and hypoplasia of upper molars. Additionally, upper incisors and molars in Slc26a2-KO-{Delta}exon2 mice display flattened odontoblasts and nuclei that lack intracellular polarity. In contrast, tooth phenotype is not remarkable in lower incisors and molars. Furthermore, the expression of odontoblast differentiation markers, Dspp and Dmp1, is significantly decreased in the upper molars of Slc26a2-deficient mice. Ex vivo organ culture of tooth germs by implantation of Slc26a2-deficient tooth germs under the kidney capsule reveals hypoplasia of the dentin matrix as well as tooth root shortening. In vitro studies using human dental pulp stem cells (hDPSCs) show that the expression levels of Dspp and Dmp1 in shSlc26a2 knockdown cells are significantly decreased compared to control cells. Collectively, our data demonstrate that SLC26A2-mediated sulfate metabolism is essential for tooth development. This study may provide insight into the mechanisms underlying tooth abnormalities in patients with recessively inherited chondrodysplasias caused by mutations in the SLC26A2 gene.

developmental biology↗

bioRxiv2021

The cell surface hyaluronidase TMEM2 plays an essential role in mouse neural crest cell development and survival

Neural crest cells (NCCs) are a migratory population that gives rise to a diverse cell lineage, including the craniofacial complex, the peripheral nervous system, and a part of the heart. Hyaluronan (HA) is a major component of the extracellular matrix, and its tissue levels are dynamically regulated in during development. Although the synthesis of HA has been shown to exert substantial influence on embryonic morphogenesis, the functional importance of the catabolic side of HA turnover is poorly understood. Here, we demonstrate that the transmembrane hyaluronidase TMEM2 plays an essential role in NCC development and the morphogenesis of their derivatives. Wnt1-Cre-mediated Tmem2 knockout (Tmem2CKO) mice exhibit severe craniofacial and cardiovascular abnormalities. Analysis of Tmem2 expression using Tmem2 knock-in reporter mice reveals that Tmem2 is expressed at the site of NCC delamination in the neural tube and in Sox9-positive emigrating NCCs, suggesting that Tmem2 is critical for NCC development. Consistent with this possibility, linage tracing analysis reveals that the contribution of Wnt1-Cre-labeled cells to NCC derivatives is significantly reduced in a Tmem2-deficient background. Moreover, the emigration of NCCs from the neural tube is greatly reduced in Tmem2CKO mice. In vitro assays demonstrate that Tmem2 expression is essential for the ability of mouse O9-1 NCCs to form focal adhesion on and migrate into HA-containing substrates. Tmem2CKO mice also exhibit increased apoptotic cell death in NCC-derived tissues. Collectively, our data demonstrate that Tmem2 is essential for normal development of NCC-derivatives, including the craniofacial complex, and that TMEM2-mediated HA degradation allows NCCs to generate a tissue environment suitable for efficient focal adhesion assembly and migration. This study reveals the hitherto unrecognized functional importance of the catabolic side of HA metabolism in embryonic development and highlights the pivotal role of Tmem2 in the process. Author SummaryThe functional significance of hyaluronan (HA) in embryonic developmental processes has been demonstrated by studies using genetic manipulation of HA synthesis. However, the expression of HA is regulated not only by its synthesis, but also by its degradation. This issue is of particular importance due to the extremely rapid metabolic turnover of HA. Curiously, mice with mutations/ablations of known hyaluronidase molecules, such as the lysosomal hyaluronidases HYAL1 and HYAL2, exhibit little embryonic phenotypes. This suggests the existence of yet another hyaluronidase molecule that plays a key role in regulating extracellular HA balance in developing tissues. In this context, transmembrane protein 2 (TMEM2) is a novel hyaluronidase that functions on the cell surface. Here, we demonstrate that TMEM2 is expressed at the site of neural crest development and in neural crest cell (NSC)-derived craniofacial tissue, and that NCC-targeted Tmem2 conditional knockout mice develop severe craniofacial defects, which attests to a requirement for TMEM2-mediated extracellular HA degradation in neural crest development. Our in vitro and in vivo analyses on the underlying mechanisms of the phenotype demonstrate that TMEM2 is essential for generating a tissue environment suitable for efficient focal adhesion formation by NCCs. This paper reveals for the first time that the catabolic machinery for HA exerts a specific regulatory role in embryonic morphogenesis, and that its dysregulation of HA degradation leads to severe developmental defects.

developmental biology↗

bioRxiv2021

Intracellular and intercellular gene regulatory networks inference from time-course individual RNA-Seq

Gene regulatory network (GRN) inference is an effective approach to understand the molecular mechanisms underlying biological events. Generally, GRN inference mainly targets intracellular regulatory relationships such as transcription factors and their associated targets. In multicellular organisms, there are both intracellular and intercellular regulatory mechanisms. Thus, we hypothesize that GRNs inferred from time-course individual (whole embryo) RNA-Seq during development can reveal intercellular regulatory relationships (signaling pathways) underlying the development. Here, we conducted time-course bulk RNA-Seq of individual mouse embryos during early development, followed by pseudo-time analysis and GRN inference. The results demonstrated that GRN inference from RNA-Seq with pseudo-time can be applied for individual bulk RNA-Seq similar to scRNA-Seq. Validation using an experimental-source-based database showed that our approach could significantly infer GRN for all transcription factors in the database. Furthermore, the inferred ligand-related and receptor-related downstream genes were significantly overlapped. Thus, the inferred GRN based on whole organism could include intercellular regulatory relationships, which cannot be inferred from scRNA-Seq based only on gene expression data. Overall, inferring GRN from time-course bulk RNA-Seq is an effective approach for understanding the regulatory relationships underlying biological events in multicellular organisms.

bioinformatics↗