Biology subjects

Kawasaki, K.

Publications and source records attributed to Kawasaki, K..

13 recordsLinked to original sources

bioRxiv2026

Sex Differences in Physiological Hair Cycling and Adhesive Material-Induced Hair Regeneration in Mice

Sex differences in hair growth are clinically evident, but sex-dependent regulation of physiological hair cycling and injury-induced hair regeneration remains incompletely understood. We compared physiological dorsal hair-cycle progression and adhesive material-induced localized hair regeneration in male and female C3H/He mice. Males entered the second and third anagen phases earlier than females, indicating longer telogen phases in females. In contrast, localized hair regrowth after application and removal of a cyanoacrylate adhesive material appeared earlier in females. Ovariectomy induced widespread telogen-to-anagen transition and therefore did not permit isolation of ovarian-hormone effects on the localized response. RNA sequencing of intact dorsal skin identified sex-dependent baseline expression profiles involving inflammation, wound response, and tissue repair. Independent time-course quantitative PCR further demonstrated sex-dependent expression of inflammatory and reparative genes after adhesive material application. Local clodronate liposome administration alone induced delayed perifocal hair growth. When combined with adhesive material application, clodronate treatment markedly delayed wound healing and localized hair regrowth in males, whereas these responses were comparatively preserved in females. These findings show that physiological hair cycling and adhesive material-induced hair regeneration exhibit distinct sex differences and suggest that the localized regenerative response is more macrophage-dependent in males than in females.

developmental biology↗

bioRxiv2026

Polymeric mechanism of enhancer-promoter cooperativity in transcriptional bursting

Emerging evidence suggest that gene expression is controlled though the modulation of transcriptional bursting across species. However, the underlying regulatory mechanisms remain largely uncertain. Recent live-imaging studies have reported that transcription factors (TFs) form a cluster at enhancers just prior to gene activation, thereby locally concentrating the active transcription machinery. This process is thought to be mediated by multivalent interaction between Mediator recruited by TFs. Conversely, transcription itself is also suggested that to influence the assembly of the TF clustering, implicating the presence of a feedback mechanism. To understand the theoretical framework underlying the interplay between TF clustering and transcription, we here develop a polymer micelle model of transcriptional bursting. With this model, multiple RNA polymerase II (Pol II) molecules loaded to the promoter, together with enhancer-bound Mediator, assemble a micelle-like structure due to their connectivity via DNA when the chromatin fiber connecting enhancer and promoter adopts closed conformation, analogous to polysoap micelle. This assembly further recruits freely diffusing Pol II and Mediator in the nucleoplasm even at low concentration up to the optimal size. Our theoretical framework enables quantitative prediction of how dynamic transitions of enhancer-promoter conformation and the stability of the micelle impact the kinetics of transcriptional bursting.

biophysics↗

bioRxiv2025

Praja1 protects cells from DNA damage through direct DNA binding

Praja1 is known as an E3 ubiquitin ligase that regulates multiple functions through protein degradation. It acquired nuclear localization signal after gene duplication and although studies have shown some significant roles of nuclear Praja1, comprehensive analysis still lacks. In this study, we performed comparative proteomics and biochemical analyses to elucidate the functions of Praja1 in the nucleus. First, proteomics analysis applied to nuclear localization deficient Praja1 exhibited signs of DNA damage response fluctuation. Subsequent comet assay revealed Praja1 protecting cells from various DNA damage sources. Similarly, cells lacking Praja1 became more sensitive to DNA damage-induced cell death, while E. coli expressing Praja1 exhibited resistance to DNA damage. To further elucidate the molecular basis of DNA protection, gel shift assay showed direct binding of Praja1 to DNA through electrostatic interactions within its intrinsically disordered region. Further in vitro damaging assay suggested that Praja1 may induce structural changes that enhance DNA repair efficiency upon binding to DNA. Together, these results provide insights into the evolutionarily novel role of nuclear Praja1 in protecting against DNA damage.

molecular biology↗

bioRxiv2024

Isolation of bacteria that catabolize abiotically synthesized tetroses via the formose reaction

Sugars synthesized through abiotic processes, specifically the formose reaction, are promising candidates for next-generation feedstocks for biomanufacturing due to their significantly higher productivity compared to cultivated crops. One of the significant challenges associated with the utilization of abiotically synthesized sugars via the formose reaction is the presence of unusual sugars, which are not metabolized by typical microorganisms. This study aimed to identify microorganisms capable of catabolizing tetroses, which are among the unusual sugars present in the abiotically synthesized sugars. Four model bacteria commonly used in biomanufacturing, including Escherichia coli, were unable to catabolize tetroses and their growth was completely inhibited by 2 to 4 g/L of D-erythrose or 0.5 to 1 g/L of L-erythrulose. We successfully isolated eight phylogenetically diverse bacterial strains from river sediments, capable of utilizing D-erythrose and L-erythrulose as the sole carbon sources. These isolates exhibited higher tolerances to tetroses compared to the model bacteria. The isolates exhibited the ability to grow in high concentrations of the abiotically synthesized sugars, which inhibit the growth of typical microorganisms, and to consume tetroses contained therein. Further investigation into the molecular mechanisms of tetrose metabolisms in the isolates will facilitate the development of biomanufacturing processes utilizing the abiotically synthesized sugars.

microbiology↗

bioRxiv2024

E3 ligase Praja1 mediates ubiquitination and degradation of microtubule-associated protein Tau

RING-H2 type E3 ligase Praja family is composed of Praja1 and Praja2, which promote the degradation of substrates through the ubiquitin-proteasome system. Both paralogs contribute to neuronal maturation and differentiation, indicating a significant role in the nervous system. Aggregation-prone proteins associated with neurodegenerative diseases, including TDP-43 and -synuclein, are degraded and/or suppressed by Praja1. Furthermore, the expression level of the MAPT gene, which is frequently mutated in Alzheimers, is regulated by Praja2. While the Praja family has been shown to recognize various aggregation-prone proteins as substrates, it has not been determined whether Tau, a key protein that aggregates in tauopathies, is also recognized by Praja proteins. In this study, we show that Praja1, but not Praja2, recognizes Tau as a candidate substrate. We observed that Tau expression in human neuroblastoma SH-SY5Y cells decreased depending on the E3 ligase activity of Praja1. Furthermore, Praja1 polyubiquitinated and interacted with Tau, indicating that it is a target substrate. Next, by combining ancestral sequence reconstruction and mutational analysis, we revealed that the Praja1-Tau interaction began via deletion of the N- and C-terminal regions of Praja1, occurring just after the duplication of the Praja family in the common ancestor of placentals. Lastly, to test whether this interaction is disrupted under pathological conditions, P301L Tau was introduced, resulting in a degradation similar to that of wild-type Tau. These results reveal an unidentified mechanism of Tau proteostasis by Praja1 and may provide insight into the pathogenesis of neurodegenerative diseases, including tauopathy.

biochemistry↗

bioRxiv2023

dact1/2 modifies noncanonical Wnt signaling and calpain 8 expression to regulate convergent extension and craniofacial development

Wnt signaling plays crucial roles in embryonic patterning including the regulation of convergent extension during gastrulation, the establishment of the dorsal axis, and later, craniofacial morphogenesis. Further, Wnt signaling is a crucial regulator of craniofacial morphogenesis. The adapter proteins Dact1 and Dact2 modulate the Wnt signaling pathway through binding to Disheveled. However, the distinct relative functions of Dact1 and Dact2 during embryogenesis remain unclear. We found that dact1 and dact2 genes have dynamic spatiotemporal expression domains that are reciprocal to one another suggesting distinct functions during zebrafish embryogenesis. Both dact1 and dact2 contribute to axis extension, with compound mutants exhibiting a similar convergent extension defect and craniofacial phenotype to the wnt11f2 mutant. Utilizing single-cell RNAseq and an established noncanonical Wnt pathway mutant with a shortened axis (gpc4), we identified dact1/2 specific roles during early development. Comparative whole transcriptome analysis between wildtype and gpc4 and wildtype and dact1/2 compound mutants revealed a novel role for dact1/2 in regulating the mRNA expression of the classical calpain capn8. Over-expression of capn8 phenocopies dact1/2 craniofacial dysmorphology. These results identify a previously unappreciated role of capn8 and calcium-dependent proteolysis during embryogenesis. Taken together, our findings highlight the distinct and overlapping roles of dact1 and dact2 in embryonic craniofacial development, providing new insights into the multifaceted regulation of Wnt signaling.

developmental biology↗

bioRxiv2023

Coordinated multiple cellular processes in tongue development

Dysfunction of primary cilia leads to genetic disorder, ciliopathies, which shows various malformations in many vital organs such as brain. Multiple tongue deformities including cleft, hamartoma and ankyloglossia are also seen in ciliopathies, which yield difficulties in fundamental functions such as mastication and vocalization. Here, we found these tongue anomalies in mice with mutation of ciliary protein. Abnormal cranial neural crest-derived cells (CNCC) failed to evoke Hh signal for differentiation of mesoderm-derived cells into myoblasts, which resulted in abnormal differentiation of mesoderm-derived cells into adipocytes. The ectopic adipose subsequently arrested migration of other mesoderm-derived cells and CNCC. Some aberrant CNCC abnormally differentiated into osteoblasts due to the lack of Hh signal, which migrated into tongue to form ectopic bone. Ankyloglossia was caused by aberrant cell migration due to lack of non-canonical Wnt signaling. In addition to ciliopathies, these tongue anomalies are often observed as non-familial condition in human. We found that these tongue deformities could be reproduced in wild-type mice by simple mechanical manipulations in CNCC to disturb cellular processes which were disrupted in mutant mice. Thus, tongue development requires coordinated multiple cellular processes (cell-cell contact, migration and differentiation). Our results provide hints for possible future treatment in ciliopathies.

developmental biology↗

bioRxiv2022

Extension of bacterial rDNA sequencing to concurrent epigenetic analysis and its application to 16S meta-epigenetics

Although polymerase chain reaction (PCR) amplification and sequencing of the 16S rDNA region has been used in a wide range of scientific fields, it does not provide DNA methylation information. We describe a simple add-on method to investigate 5-methylcytosine residues in the bacterial 16S rDNA region from clinical samples or flora. Single-stranded bacterial DNA after bisulfite conversion was preferentially amplified with multiple displacement amplification (MDA) at pH neutral, and the 16S rDNA region was analyzed using nested bisulfite PCR and sequencing. 16S rDNA bisulfite sequencing can provide clinically important bacterial DNA methylation status concurrently with intact 16S rDNA sequence information. We used this approach to identify novel methylation sites and a methyltransferase (M. MmnI) in Morganella morganii. Next, we analyzed bacterial flora from clinical specimens of small amount and identified different methylation motifs among Enterococcus faecalis strains. The method developed here, referred to as "add-on" to the conventional 16S rDNA analysis, is the most clinically used bacterial identification genetic test, which provides additional information that could not be obtained with the conventional method. Since the relationship between drug resistance in bacteria and DNA methylation status has been reported, bacterial epigenetic information would be useful in clinical testing as well. Our analysis suggests that M. MmnI has a promotive effect on erythromycin resistance. 16S rDNA bisulfite PCR and sequencing coupled with MDA at pH neutral is a useful add-on tool for analyzing 16S meta-epigenetics.

microbiology↗

bioRxiv2021

Over 400 million years of cooperation: Untangling the chondrocranium-dermatocranium connection

The cranial endo- and dermal skeletons, which comprise the vertebrate skull, evolved independently over 470 million years ago and form separately during embryogenesis. In mammals, much of the cartilaginous chondrocranium is transient, undergoing endochondral ossification or disappearing, so its role in skull morphogenesis is not well studied and it remains an enigmatic structure. We provide complete three-dimensional (3D) reconstructions of the laboratory mouse chondrocranium from embryonic day 13.5 through 17.5 using a novel methodology of uncertainty-guided segmentation of phosphotungstic enhanced 3D microcomputed tomography images with sparse annotation. We evaluate the embryonic mouse chondrocranium and dermatocranium in 3D and delineate the effects of a Fgfr2 variant on embryonic chondrocranial cartilages and on the association with forming dermal bones using the Fgfr2cC342Y/+ Crouzon syndrome mouse. We show that the dermatocranium develops outside of and in shapes that conform to the chondrocranium. Results reveal direct effects of the Fgfr2 variant on embryonic cartilage, on chondrocranium morphology, and on the association between chondrocranium and dermatocranium development. Histologically we observe a trend of relatively more chondrocytes, larger chondrocytes, and/or more matrix in the Fgfr2cC342Y/+ embryos at all timepoints before the chondrocranium begins to disintegrate at E16.5. The chondrocrania and forming dermatocrania of Fgfr2cC342Y/+ embryos are relatively large, but a contrasting trend begins at E16.5 and continues into early postnatal (P0 and P2) timepoints, with the skulls of older Fgfr2cC342Y/+ mice reduced in most dimensions compared to Fgfr2c+/+ littermates. Our findings have implications for the study and treatment of human craniofacial disease, for understanding the impact of chondrocranial morphology on skull growth, and potentially on the evolution of skull morphology.

developmental biology↗

bioRxiv2020

Requirement of Irf6 and Esrp1/2 in frontonasal and palatal epithelium to regulate craniofacial and palate morphogenesis in mouse and zebrafish

ABSTRACTOrofacial clefts are among the most common human congenital malformations. Irf6 and Esrp1 are two key genes important for palate development, conserved across vertebrates. In the zebrafish, we found that irf6 regulates the expression of esrp1. Using RNAscope, we detailed overlapping Irf6 and Esrp1/2 gene expression in the mouse frontonasal prominence ectoderm, lambda joint periderm, palate and lip epithelium. In the zebrafish, irf6 and esrp1/2 share expression in the pre-gastrulation periderm and the embryonic frontonasal ectoderm, oral epithelium ventral to the anterior neurocranium (ANC), and the developing stomodeum. Genetic disruption of irf6 and esrp1/2 in the zebrafish resulted in cleft of the ANC. In the esrp1/2 zebrafish mutant, cleft of the mouth opening formed and appeared to tether into the ANC cleft. Lineage tracing of the anterior cranial neural crest cells revealed that cleft of the ANC resulted not from migration defect, but from impaired chondrogenesis. Molecular analysis of the aberrant cells localized within the ANC cleft revealed that this cell population espresses sox10, col1a1 and irf6 and is adjacent to cells expressing epithelial krt4. Detailed morphogenetic analysis of mouse Irf6 mutant revealed mesenchymal defects not observed in the Esrp1/2 mutant. Analysis of breeding compound Irf6;Esrp1;Esrp2 mutant suggests that these genes interact where the triple mutant is not observed. Taken together, these studies highlight the complementary analysis of Irf6 and Esrp1/2 in mouse and zebrafish models and captured an unique aberrant embryonic cell population that contributes to cleft pathogenesis. Future work characterizing this unqiue sox10+, col1a1+, irf6+ cell population will yield additional insight into cleft pathogenesis.Competing Interest StatementThe authors have declared no competing interest.View Full Text

developmental biology↗

bioRxiv2020

ALX1-related Frontonasal Dysplasia Results From Defective Neural Crest Cell Development and Migration

A pedigree of subjects with frontonasal dysplasia (FND) presented with bilateral oblique facial clefts and ocular phenotypes. Genome sequencing and analysis identified a L165F missense variant in the homeodomain of the transcription factor ALX1 which was imputed to be pathogenic. Induced pluripotent stem cells (iPSC) were derived from the subjects and differentiated to neural crest cells (NCC). NCC derived from ALX1L165F/L165F iPSC were more sensitive to apoptosis, showed an elevated expression of several neural crest progenitor state markers, and exhibited impaired migration compared to wild type controls. NCC migration was also evaluated in vivo using lineage tracing in a zebrafish model, which revealed defective migration of the anterior NCC stream that contributes to the median portion of the anterior neurocranium, phenocopying the clinical presentation. Analysis of human NCC culture media revealed a change in the level of bone morphogenic proteins (BMP), with a low-level of BMP2 and a high level of BMP9. Soluble BMP2 and BMP9 antagonist treatments were able to rescue the defective migration phenotype. Taken together, these results demonstrate a mechanistic requirement of ALX1 in NCC development and migration.

cell biology↗

bioRxiv2020

Neural oscillatory activities in separate frequencies encode hierarchically distinct visual features

Although most previous studies in cognitive neuroscience have focused on the change of the neuronal firing rate under various conditions, there has been increasing evidence that indicates the importance of neuronal oscillatory activities in cognition. In the visual cortex, specific time-frequency bands are thought to have selectivity to visual stimuli. Furthermore, several recent studies have shown that several time-frequency bands are related to frequency-specific feedforward or feedback processing in inter-areal communication. However, few studies have investigated detailed visual selectivity of each time-frequency band, especially in the primate inferior temporal cortex (ITC). In this work, we analyze frequency-specific electrocorticography (ECoG) activities in the primate ITC by training encoding models that predict frequency-specific amplitude from hierarchical visual features extracted from a deep convolutional neural network (CNNs). We find that ECoG activities in two specific time-frequency bands, theta (around 5 Hz) and gamma (around 20-25 Hz) bands, are better predicted from CNN features than the other bands. Furthermore, theta- and gamma-band activities are better predicted from higher and lower layers in CNNs, respectively. Our visualization analysis using CNN-based encoding models qualitatively show that theta- and gamma-band encoding models have selectivity to higher- and lower-level visual features, respectively. Our results suggest that neuronal oscillatory activities in theta and gamma bands carry distinct information in the hierarchy of visual features, and that distinct levels of visual information are multiplexed in frequency-specific brain signals.

neuroscience↗

bioRxiv2018

Quantification of gene expression patterns to reveal the origins of abnormal morphogenesis

The earliest developmental origins of dysmorphologies are poorly understood in many congenital diseases. They often remain elusive because the first signs of genetic misregulation may initiate as subtle changes in gene expression, which can be obscured later in development due to secondary phenotypic effects. We here develop a method to trace back the origins of phenotypic abnormalities by accurately quantifying the 3D spatial distribution of gene expression domains in developing organs. By applying geometric morphometrics to 3D gene expression data obtained by Optical Projection Tomography, our approach is sensitive enough to find regulatory abnormalities never previously detected. We identified subtle but significant differences in gene expression of a downstream target of the Fgfr2 mutation associated with Apert syndrome. Challenging previous reports, we demonstrate that Apert syndrome mouse models can further our understanding of limb defects in the human condition. Our method can be applied to other organ systems and models to investigate the etiology of malformations.

developmental biology↗