Biology subjects

Inaba, M.

Publications and source records attributed to Inaba, M..

At least 19 recordsLinked to original sources

bioRxiv2026

Fertility Gene Introns Harbor Transposable Elements that Shape Y-Loop Architecture

Transposable elements (TEs) are powerful drivers of genome evolution, yet how they persist under selection and become incorporated into host regulatory networks remains poorly understood. In the Drosophila male germline, TEs are highly expressed during the spermatocyte stage, coinciding with activation of giant fertility genes on the Y chromosome. These genes contain megabase-scale introns enriched for repetitive DNA, and three of these genes form prominent nuclear structures known as Y-loops, providing a unique system to investigate gene regulation. Here, we show that multiple TEs expressed in spermatocytes are transcribed from the introns of Y-linked fertility genes. RNA fluorescence in situ hybridization (FISH) targeting several TEs, including accord2, Juan, and HMS Beagle, illuminates distinct nuclear regions corresponding to kl-2, kl-3, and kl-5, respectively. Genetic perturbation of these fertility gene loci or disruption of RNA-processing factors eliminates these TE transcripts, demonstrating that these TE sequences are embedded within Y chromosome-associated nascent transcripts rather than being independently transcribed. The identity and expression patterns of Y-loop-associated TEs vary extensively among closely related Drosophila species, consistent with the previously documented rapid evolution of Y-linked loci and suggesting that TEs may contribute to the genetic diversification of these giant fertility genes. We propose that continual turnover of repetitive elements within Y-linked introns provides a mechanism by which rapidly evolving repetitive DNA influences germline gene regulation, male fertility, and speciation.

genomics↗

bioRxiv2025

Peristaltic contractions drive gut anisotropic growth through collective cell rearrangements

The gut is the longest organ in the body, and its length is essential for efficient absorption of nutrient in adults. It is known that the gut undergoes massive elongation during prenatal/embryonic stages1,2, in which anisotropic growth takes place. But how this characteristic organ growth is achieved remains unknown. We here demonstrate an unprecedented role of peristaltic movements in the embryonic gut elongation using a caecum as a novel model in chicken embryos. Inhibition of the peristaltic movements by chemical drugs impedes the elongation of caecum. The elongation-halted caecum restores its longitudinal growth by optogenetically introduced peristaltic movements. At the cellular level, we have found that circular smooth muscle cells divide along the circumferential axis in a peristalsis-independent manner, which would otherwise serve as an engine for the radial growth. However, with peristaltic stimulation, the cells expand collectively their distribution along the gut longitudinal axis leading to the elongation of this organ. Thus, peristaltic activity biases circumferentially proliferating smooth muscle cells toward longitudinal rearrangement. This two-step model offers a novel way to identify the peristaltic activity as a critical growth signal for the gut elongation, and also to parsimoniously explain the anisotropic growth of the gut, which undergoes the elongation while retaining its thickness.

developmental biology↗

bioRxiv2025

Brinker regulates reciprocal outcomes of BMP signal between stem cells and differentiating cells

Drosophila male germline stem cells (GSCs) reside at the testis tip, surrounding a cluster of niche cells known as the hub. Bone Morphogenetic Protein (BMP) ligands secreted from the hub exert both contact-dependent and -independent effects. In close proximity to the niche, BMP signaling maintains stem cells by suppressing transcription of the key differentiation factor Bag of Marbles (Bam). In contrast, the diffusible fraction of BMP promotes differentiation of cells by activating bam. How a single signaling pathway produces such opposing outcomes has remained unclear. Here, we show that the diffusible BMP fraction induces bam transcription by repressing the transcriptional repressor Brinker (Brk). We further found that brk mRNA displays a highly heterogeneous expression pattern within interconnected spermatogonia, suggesting that Brk may prime cell fate in a subset of transit-amplifying cells, helping to preserve a population poised for dedifferentiation while maintaining other cells for differentiation. Our findings propose a model in which a single niche-derived factor modulates reciprocal outcomes inside versus outside the niche, which is essential for the tissue homeostasis. Given the broad use of BMP signaling across stem cell niches, this mechanism may represent a general strategy to ensure correct balance between self-renewal and differentiation of stem cells.

developmental biology↗

bioRxiv2025

Efficacy of natural marine sponges as a passive environmental DNA sampler for freshwater fish diversity monitoring

Environmental DNA (eDNA) analysis is a cost-effective and noninvasive tool for species and biodiversity monitoring in aquatic environments. Passive eDNA sampling is a novel alternative to conventional sampling methods such as water filtration. In this study, we examined the efficacy of the sponge skeleton as a passive eDNA sampler for monitoring freshwater fish. The performance of the passive sampling method was compared with that of standard water filtration in a river environment. Five DNA extraction methods were used in three experiments, and a suitable method for DNA extraction from sponge skeleton was identified. Quantitative fish metabarcoding using MiFish primers revealed no significant differences in species richness between the aqueous and passive sampling methods. Although both the sampling methods showed comparable trends in fish community structure, different clusters were identified for water sampling and passive samplers based on the differences in the DNA concentration of each fish species. The fish diversity in the passive samples was comparable among four DNA extraction methods (except for the filtration method) using the direct capture method. Our results demonstrate the efficacy of passive eDNA sampling for monitoring freshwater fish diversity and the potential use of sponge skeletons as absorption materials for passive eDNA samplers.

ecology↗

bioRxiv2025

The functional organization of chromosome territories in single nuclei during zygotic genome activation

Chromosome territories (CTs) are intricately organized and regulated within the nucleus. Despite remarkable advances in our understanding of genome packaging and gene expression, the interplay among CTs, pairing of parental homologous chromosomes, and genome function during development remains elusive. Here, we employ an Oligopaints-based high-resolution imaging approach to examine variable CT organization in single nuclei during the developmental process of zygotic genome activation. We reveal large-scale chromosome changes with extensive homolog pairing at the whole-chromosome level that decreases locally due to spatial variability in chromosome conformations. In the absence of one homolog copy, the dynamics of CT compaction and RNA polymerase II recruitment are supported by transcriptional changes in haploid embryos. Finally, global inhibition of transcription results in decreased CT opening and no significant impact on CT pairing levels. These findings enhance our understanding of parental genome folding and regulation, which may inform strategies for chromosome-based diseases.

developmental biology↗

bioRxiv2025

Detection of dedifferentiated stem cells in Drosophila testis

Tissue homeostasis relies on the stable maintenance of the stem cell pool throughout an organisms lifespan. Dedifferentiation, a process in which partially or terminally differentiated cells revert to a stem cell state, has been observed in a wide range of stem cell systems, and it has been implicated in the mechanisms for stem cell maintenance. Dedifferentiated stem cells are morphologically indistinguishable from original stem cells, making them challenging to identify. Therefore, whether dedifferentiated stem cells have any distinguishable characteristics compared with original stem cells is poorly understood. The Drosophila testis provides a well-established model to study dedifferentiation. While our previous live imaging analyses have identified dedifferentiation events constantly occurring at steady state, existing genetic marking methods fail to detect most of the dedifferentiated stem cells and thus significantly underestimate the frequency of dedifferentiation events. Here, we established a genetic tool with improved sensitivity and used live imaging and mathematical modeling to evaluate the system. Our findings indicate that the specificity of lineage-specific promoters is critical for successfully identifying dedifferentiated stem cells.

developmental biology↗

bioRxiv2024

Self-organized patterning of peristaltic waves by suppressive actions in a developing gut

Gut peristalsis is a wave-like movement of a local contraction along the gut, and plays important roles in nutrient digestion and absorption. When peristaltic waves emerge in embryonic guts, randomly distributed origins of peristaltic waves (OPWs) become progressively confined to specific sites. We have investigated how this random-to-organized positioning is achieved using the caecum as a model in chicken embryos. While prominent OPWs, recognized as active (spontaneous) contractions, are located at endpoints of the intact caecum, other regions are also found to possess latent rhythm unveiled by fragmentation of a caecum into pieces, showing that the latent rhythm is normally suppressed in the intact gut. Analyses with caecum fragments demonstrate that the latent rhythm is spatially patterned in an early gut, to which negative impact by primitive passing waves contributes; the more passing waves a region experiences to undergo forced/passive contractions, the slower latent rhythm this region acquires. This patterned latent rhythm underlies the final positioning of OPWs at later stages, where a site with faster latent rhythm dominates neighboring slower rhythm, surviving as a "winner" by macroscopic lateral inhibition. Thus, the random-to-organized patterning of OPWs proceeds by self-organization within the caecum, in which two distinct mechanisms, at least, are employed; suppressive actions by primitive waves followed by macroscopic lateral inhibition.

developmental biology↗

bioRxiv2024

Draft genome of Palmaria palmata and intraspecific genetic variations in the North-East Atlantic

The rhodophyte Palmaria palmata (L.) Weber & Mohr is one of the target species of a growing European seaweed industry due to its high content of protein and essential amino acids which makes it suitable for human food, dietary supplements, and as salmon feed. However, the lack of a published nuclear genome limits phylogenetics analyses and gene function investigations which could help the development of a breeding programme. We present the first draft genome of P. palmata that was obtained with PacBio HiFi long read sequencing with average coverage of 10x, consisting of an assembly of 1.05 Gb, N50=2.75Mb and BUSCO completeness of 72.1%. Additionally, a population study on the whole genome of 33 P. palmata individuals from across the Northern East Atlantic area found three main clusters consistent with their geographic distribution: (1) Denmark and Norway, (2) France and western Ireland, (3) Faroe Islands. All individuals from Northern Ireland share ancestry with western Ireland and Denmark, and some individuals from the Faroe Islands show admixture from Faroe, western Ireland and Northern Ireland. These results represent a fundamental step towards breeding and genetic studies to further explore the vastly unexploited economic potential of Palmaria palmata. Highlights- We report the first draft genome of Palmaria palmata, from PacBio HiFi long reads. - The size of the genome, 1.05 Gb, is among the largest so far among the Rhodophyta. - Busco completeness of 72.1% and contig N50 of 2.75 Mb indicate good quality. - The genomes of 33 more individuals from North Atlantic Europe have been sequenced. - Phylogenetic analysis found three clusters consistent with geographic distribution.

genomics↗

bioRxiv2024

Asymmetric stem cell division maintains the genetic heterogeneity of tissue cells

Within a given tissue, the stem cell niche provides the microenvironment for stem cells suitable for their self-renewal. Conceptually, the niche space constrains the size of a stem-cell pool, as the cells sharing the niche compete for its space. It has been suggested that either neutral-or non-neutral-competition of stem cells changes the clone dynamics of stem cells. Theoretically, if the rate of asymmetric division is high, the stem cell competition is limited, thus suppressing clonal expansion. However, the effects of asymmetric division on clone dynamics have never been experimentally tested. Here, using the Drosophila germline stem cell (GSC) system, as a simple model of the in-vivo niche, we examine the effect of division modes (asymmetric or symmetric) on clonal dynamics by combining experimental approaches with mathematical modeling. Our results suggest that the rate of asymmetric division positively correlates with the time a stem cell clone takes to expand. Taken together, our data suggests that asymmetric division is essential for maintaining the genetic variation of stem cells and thus serves as a critical mechanism for safeguarding fertility over the animal age or preventing multiple disorders caused by the clonal expansion of stem cells.

developmental biology↗

bioRxiv2024

The gut contractile organoid: a novel model for studying the gut motility regulated by coordinating signals between interstitial cells of Cajal and smooth muscles

The gut undergoes peristaltic movements regulated by intricate cellular interactions. How these interactions emerge in the developing gut remains poorly explored due to a lack of model system. We here developed a novel contractile organoid that is derived from the muscle layer of chicken embryonic hindgut. The organoid contained smooth muscle cells (SMCs) and interstitial cells of Cajal (ICCs; pacemaker) with few enteric neurons, and underwent periodic contractions. The organoid was formed by self-organization with morphological arrangements of ICCs (internal) and SMCs (peripheral), allowing identification of these cells in live. GCaMP-Ca2+ imaging analyses revealed that Ca2+ transients between ICC-ICC, SMC-SMC or SMC-ICC were markedly coordinated. Pharmacological studies further suggested a role of gap junctions in ICC-to-SMC signaling, and also possible mechanical feedback from SMCs contraction to ICCs pace-making activities. In addition, two organoids with different rhythm became synchronized when mediated by SMCs, unveiling a novel contribution of SMCs to ICCs pace-making. The gut contractile organoid developed in this study offers a useful model to understand how the rhythm coordination between/among ICCs and SMCs is regulated and maintained during gut development.

developmental biology↗

bioRxiv2024

Soma-to-germline BMP signal is essential for Drosophila spermiogenesis

In the Drosophila testis, developing germ cells are encapsulated by somatic support cells throughout development. Soma-germline interactions are essential for successful spermiogenesis. However, it is still not fully understood what signaling events take place between the soma and the germline. In this study, we found that a Bone Morphogenetic Protein (BMP) ligand, Glass bottom boat (Gbb), secreted from somatic cyst cells (CCs), signals to differentiating germ cells to maintain proper spermiogenesis. Knockdown of Gbb in CCs or the type I BMP receptor Saxophone (Sax) in germ cells leads to a defect in sperm head bundling and decreased fertility. Our Transmission Electron Microscopy (TEM) analyses revealed that the mutant germ cells have aberrant morphology of mitochondria throughout the stages of spermiogenesis and exhibit a defect in nebenkern formation. Elongating spermatids show uncoupled nuclei and elongating mitochondrial derivatives, suggesting that improper mitochondrial development may cause the sperm bundling defect. Taken together we propose a new role of soma-derived BMP signaling, which is essential for spermiogenesis.

developmental biology↗

bioRxiv2023

Dedifferentiating germ cells regain stem-cell specific polarity checkpoint prior to niche reentry

In the Drosophila germline stem cell system, maintenance of the stem cell pool requires "dedifferentiation", in which differentiating cells reattach to the niche and reacquire stem cell properties. However, the mechanism of dedifferentiation remains poorly understood. Here, using long-term live imaging, we show that dedifferentiated cells immediately re-enter mitosis with correct spindle orientation after reattachment to the niche. Analysis of cell cycle markers revealed that these dedifferentiating cells are all in G2 phase. In addition, we found that the observed G2 block during dedifferentiation likely corresponds to a centrosome orientation checkpoint (COC), a previously reported polarity checkpoint. We show that re-activation of a COC is likely required for the dedifferentiation thus ensuring asymmetric division even in dedifferentiated stem cells. Taken together, our study demonstrates the remarkable ability of dedifferentiating cells to reacquire the ability to divide asymmetrically.

developmental biology↗

bioRxiv2022

Homophilic interaction of E-Cadherin prevents cell-cell fusion between developing germline and surrounding epithelia in Drosophila ovary

In the Drosophila ovary, developing germline cysts are encapsulated by somatic follicle cell epithelia and E-Cadherin localizes to the interface of these tissues. E-Cadherin mutants have been shown to have multiple defects in oogenesis. Therefore, it is difficult to determine E-Cadherin function on germline-soma interaction. In this study, we characterize E-Cadherin function, specifically focusing on germline-soma interaction. Unexpectedly, knockdown of E-Cadherin either in the germline or follicle cells results in excess formation of membrane protrusions at the interface of these cells, which leads to a cell-cell fusion and indicates that homophilic interaction of E-Cadherin is required for maintenance of the tissue boundary between these two adjacent tissues. The fate of follicle cells fused to the germline becomes compromised, leading to a defective individualization of germline cysts. We propose that homophilic interaction of E-Cadherin facilitates a barrier between adjacent tissues, demonstrating a unique model of cell-fate disturbance caused by cell-cell fusion.

cell biology↗

bioRxiv2022

Decapentaplegic ligand ensures niche space restriction inside and outside of Drosophila testicular niche

Drosophila male germline stem cells (GSCs) reside at the tip of the testis and surround a cluster of niche cells. It has been believed that the niche-derived Decapentaplegic (Dpp) has a role in maintaining stem cells in close proximity but has no role in the differentiating cells spaced one-cell layer away. However, the range of Dpp diffusion has never been tested. Here, using genetically encoded nanobodies called Morphotrap, we physically block Dpp diffusion without interfering with niche-stem cell signaling and find that diffusible fraction of Dpp is required to ensure differentiation of GSC daughter cells, opposite of its role in maintenance of GSC in the niche. Our work provides an example in which a soluble niche ligand induces opposed cellular responses in stem cells and in differentiating descendants so that the niche can tightly restrict its space. This may be a common mechanism to regulate tissue homeostasis. One sentence summaryBMP ligand diffuses from the niche and has dual, and opposite roles on stem cells and differentiating daughter cells.

developmental biology↗

bioRxiv2022

Newly raised anti-c-Kit antibody detects interstitial cells of Cajal in the gut of chicken embryos

The gut peristaltic movement, a wave-like propagation of a local contraction, is important for the transportation and digestion of ingested materials. Among three types of cells, the enteric nervous system (ENS), smooth muscle cells, and interstitial cells of Cajal (ICCs), the ICCs have been thought to act as a pacemaker, and therefore it is important to decipher the cellular functions of ICCs for the understanding of gut peristalsis. c-Kit, a tyrosine kinase receptor, has widely been used as a marker for ICCs. Most studies with ICCs have been conducted in mammals using commercially available anti-c-Kit antibody. Recently, the chicken embryonic gut has emerged as a powerful model to study the gut peristalsis. However, since the anti-c-Kit antibody for mammals does not work for chickens, cellular mechanisms by which ICCs are regulated have largely been unexplored. Here, we report a newly raised polyclonal antibody against the chicken c-Kit protein. The specificity of the antibody was validated by both Western blotting analyses and immunocytochemistry. Co-immunostaining with the new antibody and anti- smooth muscle actin (SMA) antibody successfully visualized ICCs in the chicken developing hindgut in the circular muscle- and longitudinal muscle layers: as previously shown in mice, common progenitors of ICCs and smooth muscle cells at early stages were double positive for SMA and c-Kit, and at later stages, differentiated ICCs and smooth muscle cells exhibited only c-Kit and SMA, respectively. A novel ICC population was also found that radially extended from the submucosal layer to circular muscle layer. Furthermore, the new antibody delineated individual ICCs in a cleared hindgut. The antibody newly developed in this study will facilitate the study of peristaltic movement in chicken embryos.

developmental biology↗

bioRxiv2021

Distribution map of peristaltic waves in the chicken embryonic gut reveals importance of ENS and inter-region cross talks along the gut axis

Gut peristaltic movements recognized as the wave-like propagation of a local contraction are crucial for effective transportation and digestion/absorption of ingested materials. Although the physiology of gut peristalsis has been well studied in adults, it remains largely unexplored how the cellular functions underlying these coordinated tissue movements are established along the rostral-caudal gut axis during development. The chicken embryonic gut serves as an excellent experimental model for elucidating the endogenous potential and regulation of these cells since peristalsis occurs even though no ingested material is present in the moving gut. By combining video-recordings and kymography, we provide a spatial map of peristaltic movements along the entire gut posterior to the duodenum: midgut (jejunum and ileum), hindgut, caecum, and cloaca. Since the majority of waves propagate bidirectionally at least until embryonic day 12 (E12), the sites of origin of peristaltic waves (OPWs) can unambiguously be detected in the kymograph. The spatial distribution map of OPWs has revealed that OPWs become progressively confined to specific regions/zones along the gut axis during development by E12, and that such specific zones are largely conserved between different individuals implying genetic regulation for OPW determination. We have also found that the enteric nervous system (ENS) is essential for the OPW patterning since an ablation of ENS or blocking neural activity by tetrodotoxin disrupts the confined pattern of OPWs, resulting in a failure of transportation of inter-luminally injected ink. Finally, we have discovered a functional coupling of the endpoint of hindgut with the cloaca. When surgically separated, the cloaca ceases its acute contractions that would normally occur concomitantly with the peristaltic rhythm of the hindgut. Our findings shed light on the intrinsic regulations of gut peristalsis, including unprecedented ENS contribution and inter-region cross talk along the gut axis. Contribution to the field statementIt has been well accepted that the gut peristalsis is important in adults, where a luminal content (ingested material) mechanically influences peristalsis. However, the endogenously regulated cellular mechanisms that initiate and sustain peristalsis have poorly been explored, and this might be a reason why few therapeutic treatments have been available for gut diseases associated with peristaltic dysfunction. Recent studies have shown that peristaltic movements occur in the embryonic gut, suggestive of genetic involvement. However, how the peristaltic movement is coordinately patterned along the gut axis remains unknown, because most studies have used isolated fragments of the gut. In our study, we examined the entire gut posterior to the duodenum of chicken embryos to produce a spatial map of peristaltic movements during gut development. Using the map of origins of peristaltic waves (OPWs), we found previously unappreciated roles of enteric nervous system for the OPW patterning and for the physiological functions of embryonic gut tissues. Furthermore, we propose that peristaltic movements might mediate inter-region crosstalk along the gut axis. Our findings provide novel insights into the mechanisms by which the gut peristalsis is established during development at the cellular basis.

developmental biology↗

bioRxiv2021

Interchromosomal interaction of homologous Stat92E alleles regulates transcriptional switch during stem-cell differentiation

The strength of pairing of homologous chromosomes differs in a locus-specific manner and is correlated to gene expression states. However, the functional impact of homolog pairing on local transcriptional activity is still unclear. Drosophila male germline stem cells (GSCs) constantly divide asymmetrically to produce one GSC and one differentiating gonialblast (GB). The GB then enters the differentiation program in which stem cell specific genes are quickly downregulated. Here we demonstrate that a change in local pairing state of the Stat92E locus is required for the downregulation of the Stat92E gene during differentiation. Using OligoPaint fluorescent in situ hybridization (FISH), we show that the interaction between homologous regions of Stat92E is always tight in GSCs and immediately loosened in GBs. When one of the Stat92E locus was absent or relocated to another chromosome, Stat92E did not pair and failed to downregulate, suggesting that the pairing is required for switching of transcriptional activity. The defect in downregulation of Stat92E was also observed upon knockdown of global pairing or anti-pairing factors. Moreover, the Stat92E enhancer element, but not cis-transcription, is required for the change in pairing state, indicating that it is not a consequence of transcriptional changes. GSCs are known to inherit pre-existing histones H3 and H4, while newly synthesized histones are distributed in GBs. When this histone inheritance was compromised, the change in Stat92E pairing did not occur, suggesting that it is an intrinsically programmed process during asymmetric stem cell division. We propose that the change of local pairing state may be a common process to reprogram gene activity during cell-differentiation.

developmental biology↗

bioRxiv2021

Acetylated microtubules are required for maintenance of the barrier between two adjacent tissues

Microtubule acetylation is found in populations of stable, long-lived microtubules, occurring on the conserved lysine 40 (K40) residue of -tubulin, catalyzed by -tubulin acetyltransferases (TATs). K40 acetylation has been shown to stabilize microtubules via enhancing microtubule resilience against mechanical stress. Here we show that Drosophila CG17003/leaky (Lky), an TAT, is required for proper oogenesis. We found that loss of lky disrupted the cell junction between germline cyst and follicle epithelial cells, adjacent cells that form an egg chamber. This resulted in leakage of germline contents into somatic follicle cells. The follicle cells that received germline-derived nanos gene product failed to maintain their cell fate, leading to an egg chamber fusion. The same phenotype was observed upon replacement of major -tubulin84BK40 with -tubulin84BK40A (non-acetylable tubulin), suggesting -tubulinK40 acetylation is required for the boundary integrity of these two adjacent tissues. Taken together, this study provides the first in vivo function of tubulin acetylation in maintaining the integrity of a tissue barrier.

developmental biology↗