Biology subjects

Fujiwara, K.

Publications and source records attributed to Fujiwara, K..

35 records · Page 2Linked to original sources

bioRxiv2024

Tameness selection pressure affects gut virome diversity in mice

The gut microbiome, a complex ecosystem comprising bacteria, viruses, archaea, fungi, and other microorganisms, plays a pivotal role in host health, immunity, and behaviour. Among its components, bacteriophages and viruses that infect bacteria significantly influence microbial community dynamics by affecting bacterial diversity, function, and evolution. Despite extensive research on bacterial components, the viral metagenome (virome) remains relatively unexplored. This study investigated the effect of selective breeding for tameness on the gut virome of wild heterogeneous stock (WHS) mice. WHS mice were selectively bred for active tameness, resulting in four groups: S1 and S2 (selected for tameness), and C1 and C2 (non-selected). In previous study, we observed an increased abundance of Limosilactobacillus reuteri in tame groups compared to non-selected groups, as well as a tameness-promoting effect of L. reuteri with long-term administration. Given the regulatory role of phages in bacterial populations, this study focused on analysing the gut virome using shotgun metagenome sequencing data. From the 84 samples, we generated 6078 non-redundant viral operational taxonomic units (vOTUs) and identified the hosts of 3,065 of these vOTUs. Significant differences in gut virome beta diversity were observed between the selectively bred and control groups, suggesting that tameness selection exerts distinct pressure on the virome. Additionally, phage-host interaction analysis revealed strong correlations between specific phages and their bacterial hosts, indicating a co-occurrence that influences host behaviour. Overall, this study provides novel insights into the role of the gut virome in shaping host behaviour and highlights the broader implications of microbial contributions to domestication and selective breeding outcomes.

microbiology↗

bioRxiv2024

Investigation of host plant contact with Diaphorina citri (Hemipreta: Psyllidae) by detecting D. citri-derived environmental DNA

Citrus greening (Huanglongbing) disease has serious impacts on citrus production. Field monitoring of the Asian citrus psyllid Diaphorina citri (Hemiptera: Psyllidae), a vector of citrus greening disease, is essential to prevent the invasion and spread of this disease. This study reported a new method for determining the presence of D. citri and traces of contact with host plants by collecting leaves of host plants from the survey area and detecting the environmental DNA (eDNA) derived from D. citri. The results showed that the method can determine the presence of D. citri. As D. citri-derived eDNA is contained in DNA solutions of plants with a history of contact with D. citri, we succeeded in detecting not only genes conserved in D. citri mitochondria but also genes of D. citri symbiont organisms consisting of Wolbachia, Wolbachia phage, Candidatus Carsonella sp., and Candidatus Profftella sp. Diaphorina citri-derived eDNA could be detected in host plants even after only 10 min of contact with D. citri and could still be detected 6 months after contact. This technology has the potential to trace D. citri from their traces without individual detection and is expected to greatly contribute to the early detection and invasion warning of citrus greening disease in the future.

molecular biology↗

bioRxiv2024

Decoding the Centromeric Region with a Near Complete Genome Assembly of the Oshima Cherry Cerasus speciosa

The Oshima cherry (Cerasus speciosa), which is endemic to Japan, has significant cultural and horticultural value. In this study, we present a near complete telomere-to-telomere genome assembly for C. speciosa, derived from the old growth "Sakurakkabu" tree on Izu Oshima Island. Using Illumina short-read, PacBio long-read, and Hi-C sequencing, we constructed a 269.3 Mbp genome assembly with a contig N50 of 32.0 Mbp. We examined the distribution of repetitive sequences in the assembled genome and identified regions that appeared to be centromeric. Detailed structural analysis of these putative centromeric regions revealed that the centromeric regions of C. speciosa comprised repetitive sequences with monomer lengths of 166 or 167 bp. Comparative genomic analysis with Prunus sensu lato genome revealed structural variations and conserved syntenic regions. This high-quality reference genome provides a crucial tool for studying the genetic diversity and evolutionary history of Cerasus species, facilitating advancements in horticultural research and the preservation of this iconic species.

genomics↗

bioRxiv2024

Evolution of the sex-determination gene Doublesex within the termite lineage

The molecular mechanism of sex determination has long been considered conserved in insects. However, recent studies of hemimetabolous insects have challenged this notion. One notable example is termites. In Reticulitermes speratus, a homolog of sex determination gene, Doublesex (RsDsx), exhibits characteristics that are distinct from those of other insects, including sister-group cockroaches. It comprises a single exon, contains only doublesex/mab-3 DNA-binding domain (DM) but lacks a conserved oligomerization domain (OD), and exhibits transcriptional activity only in males. To investigate whether these characteristics are widespread within the termite lineage, we identified Dsx homologs in three different families. The loss of the conserved OD sequences was observed in all termite species examined, whereas the number of exons and expression patterns between sexes varied among families. Particularly, distinctive differences in Dsx were found in species from the Archotermopsidae and Kalotermitidae, both of which have a linear caste developmental pathway. Our findings indicate that diversification of Dsx structure and expression patterns may have contributed to ecological diversification, such as caste developmental pathways, within the termite lineage.

evolutionary biology↗

bioRxiv2024

A role for the S4-domain containing protein YlmH in ribosome-associated quality control in Bacillus subtilis

Ribosomes trapped on mRNAs during protein synthesis need to be rescued for the cell to survive. The most ubiquitous bacterial ribosome rescue pathway is trans-translation mediated by tmRNA and SmpB. Genetic inactivation of trans-translation can be lethal, unless the ribosomes are rescued by ArfA or ArfB alternative rescue factors or the ribosome-associated quality control (RQC) system, which in B. subtilis involves MutS2, RqcH, RqcP and Pth. Using transposon sequencing in a trans-translation-incompetent B. subtilis strain we identify a poorly characterized S4-domain-containing protein YlmH as a novel potential RQC factor. Cryo-EM structures reveal that YlmH binds peptidyl-tRNA-50S complexes in an position analogous to that of S4-domain-containing RqcP, and that, similarly to RqcP, YlmH can co-habit with RqcH. Consistently, we show that YlmH can assume the role of RqcP in RQC in facilitating the addition of polyalanine tails to the truncated nascent polypeptides. While in B. subtilis the function of YlmH is redundant with RqcP, our taxonomic analysis reveals that in multiple bacterial phyla RqcP is absent, while YlmH and RqcH are present, suggesting that in these species the YlmH plays a central role in the RQC.

microbiology↗

bioRxiv2024

Resolution of ribosomal stalling by ABCF ATPases YfmR and YkpA/YbiT

Efficiency of protein synthesis on the ribosome is strongly affected by the amino acid composition of the assembled amino acid chain. Challenging sequences include proline-rich motifs as well as highly positively and negatively charged amino acid stretches. Members of the F subfamily of ABC ATPases (ABCFs) have been long hypothesised to promote translation of such problematic motifs. In this study we have applied genetics and reporter-based assays to characterise the four housekeeping ABCF ATPases of Bacillus subtilis: YdiF, YfmM, YfmR/Uup and YkpA/YbiT. We show that YfmR cooperates with the translation factor EF-P that promotes translation of Pro-rich motifs. Simultaneous loss of both YfmR and EF-P results in a dramatic growth defect. Surprisingly, this growth defect can be largely suppressed though overexpression of an EF-P variant lacking the otherwise crucial 5-amino-pentanolylated residue K32. Using in vivo reporter assays, we show that overexpression of YfmR can alleviate ribosomal stalling on Asp-Pro motifs. Finally, we demonstrate that YkpA/YbiT promotes translation of positively and negatively charged motifs but is inactive in resolving ribosomal stalls on proline-rich stretches. Collectively, our results provide insights into the function of ABCF translation factors in modulating protein synthesis in B. subtilis.

microbiology↗

bioRxiv2023

Beacon-based sleep status and physical activity monitoring in humans.

One out of every five people in Japan is dissatisfied with their sleep and various diseases caused by lack of exercise have been pointed out, but there are few effective remedies for these problems. In this study, we aimed to develop a simple method for measuring behavioral sleep patterns and physical activity using a beacon accelerometer wirelessly connected with a smartphone. A sleep prediction model was created comparing the data obtained from the accelerometer with the sleep status data obtained by a previously validated sleep monitoring system. The Random Forest model was able to classify sleep and wakefulness with a 97.4% and 85.4% precision, respectively, which were comparable to those of conventional acceleration-based sleep monitoring devices. Additionally, the same data acquisition method was used to classify exercise intensity into seven levels and a high correlation (r=0.813, p<0.0001) was found when comparing the classified exercise intensity to metabolic equivalent (MET) values. This suggests that the proposed method can be used for accurate measurement of both behavioral sleep and physical activity classifying over a long period of time.

physiology↗

bioRxiv2023

Tumor- and Nerve-Derived Axon Guidance Molecule Promotes Pancreatic Ductal Adenocarcinoma Progression and Metastasis through Macrophage Reprogramming

Axon guidance molecules were found to be the gene family most frequently altered in pancreatic ductal adenocarcinoma (PDA) through mutations and copy number changes. However, the exact molecular mechanism regarding PDA development remained unclear. Using genetically engineered mouse models to examine one of the axon guidance molecules, semaphorin 3D (SEMA3D), we found a dual role for tumor-derived SEMA3D in malignant transformation of pancreatic epithelial cells and a role for nerve-derived SEMA3D in PDA development. This was demonstrated by the pancreatic-specific knockout of the SEMA3D gene from the KRASG12D and TP53R172H mutation knock-in, PDX1-Cre (KPC) mouse model which demonstrated a delayed tumor initiation and growth comparing to the original KPC mouse model. Our results showed that SEMA3D knockout skews the macrophages in the pancreas away from M2 polarization, providing a potential mechanistic role of tumor-derived SEMA3D in PDA development. The KPC mice with the SEMA3D knockout remained metastasis-free, however, died from primary tumor growth. We then tested the hypothesis that a potential compensation mechanism could result from SEMA3D which is naturally expressed by the intratumoral nerves. Our study further revealed that nerve-derived SEMA3D does not reprogram macrophages directly, but reprograms macrophages indirectly through ARF6 signaling and lactate production in PDA tumor cells. SEMA3D increases tumor-secreted lactate which is sensed by GPCR132 on macrophages and subsequently stimulates pro-tumorigenic M2 polarization in vivo. Tumor intrinsic- and extrinsic-SEMA3D induced ARF6 signaling through its receptor Plexin D1 in a mutant KRAS-dependent manner. Consistently, RNA sequencing database analysis revealed an association of higher KRASMUT expression with an increase in SEMA3D and ARF6 expression in human PDAs. Moreover, multiplex immunohistochemistry analysis showed an increased number of M2-polarized macrophages proximal to nerves in human PDA tissue expressing SEMA3D. Thus, this study suggests altered expression of SEMA3D in tumor cells lead to acquisition of cancer-promoting functions and the axon guidance signaling originating from nerves is "hijacked" by tumor cells to support their growth. Other axon guidance and neuronal development molecules may play a similar dual role which is worth further investigation. One sentence summaryTumor- and nerve-derived SEMA3D promotes tumor progression and metastasis through macrophage reprogramming in the tumor microenvironment. STATEMENT OF SIGNIFICANCEThis study established the dual role of axon guidance molecule, SEMA3D, in the malignant transformation of pancreatic epithelial cells and of nerve-derived SEMA3D in PDA progression and metastasis. It revealed macrophage reprogramming as the mechanism underlying bothroles. Together, this research elucidated how inflammatory responses promote invasive PDA progression and metastasis through an oncogenic process.

cancer biology↗

bioRxiv2023

Patchy and widespread distribution of bacterial translation arrest peptides associated with the protein localization machinery

Regulatory arrest peptides exert cellular functions via mechanisms involving regulated translational arrest. Monitoring substrates, a class of arrest peptides, feedback-regulate the expression of the Sec or YidC protein localization machinery. Previously, only a limited number of monitoring substrates were identified. In this study, we performed a bacterial domain-wide search, followed by in vivo and in vitro analyses, leading to a comprehensive identification of many novel Sec/YidC-related arrest peptides that showed patchy, but widespread, phylogenetic distribution throughout the bacterial domain. Identification of five novel arrest-inducing sequences suggests that bacteria have evolved various arrest-inducing mechanisms. We also identified many arrest peptides that share an R-A-P-P like sequence, suggesting that this sequence could serve as a common evolutionary seed that could overcome the species-specific structures of ribosomes, to evolve arrest peptides. Our comprehensive phylogenetic study revealed that arrest peptide is a prevalent mechanism for the gene regulation of the protein localization machinery.

molecular biology↗

bioRxiv2023

Inference of Selective Force on House Mice Genomes during Secondary Contact in East Asia

The house mouse (Mus musculus), commensal to humans, has spread globally via human activities, leading to secondary contact between genetically divergent subspecies. This pattern of genetic admixture can provide insights into the selective forces at play in this well-studied model organism. Our analysis of 163 house mouse genomes, with a particular focus on East Asia, revealed substantial admixture between the subspecies castaneus and musculus, particularly in Japan and southern China. We revealed, despite the admixture, all Y chromosomes in the East Asian samples belonged to the musculus-type haplogroup, potentially explained by genomic conflict under sex ratio distortion due to varying copy numbers of ampliconic genes on sex chromosomes. We also investigated the influence of selection on the post-hybridization of the subspecies castaneus and musculus in Japan. Even though the genetic background of most Japanese samples closely resembles the subspecies musculus, certain genomic regions overrepresented the castaneus-like genetic components, particularly in immune-related genes. Furthermore, a large genomic block containing a vomeronasal/olfactory receptor gene cluster predominantly harbored castaneus-type haplotypes in the Japanese samples, highlighting the crucial role of olfaction-based recognition in shaping hybrid genomes.

evolutionary biology↗

bioRxiv2022

Formation of multiple flagella caused by a mutation of the flagellar rotor protein FliM in Vibrio alginolyticus

The marine bacterium Vibrio alginolyticus forms only a single flagellum at the cell pole. In Vibrio, two proteins (GTPase FlhF and ATPase FlhG) regulate flagellar number at the cell pole. We previously isolated a mutant strain characterized as NMB155 that forms multiple flagella despite the absence of mutations in flhF and flhG. NMB155 also exhibited straight swimming without a directional change in flagellar rotation. Whole-genome sequencing of NMB155 identified an E9K mutation in FliM that is a component of the C-ring in the flagellar rotor. Mutations in FliM result in defects in flagellar formation (fla) and flagellar rotation (che or mot); however, there are few reports indicating that FliM mutations increase the number of flagella. Here, we determined that the E9K mutation confers the multi-flagellar phenotype and also the che phenotype. The co-expression of wild-type FliM and FliM-E9K indicated that they were competitive in regard to determining the flagellar number. It had been shown that the ATPase activity of FlhG corresponds to the flagellar number. We observed that the ATPase activity of FlhG was increased by the addition of FliM but not by the addition of FliM-E9K. This indicates that the N-terminal region of FliM that includes the E9 residue interacts with FlhG to increase its ATPase activity, and the E9K mutation may inhibit this interaction. We concluded that FliM downregulate FlhG activity to inhibit the formation of additional flagella. ImportanceThe flagellar rotor generates a driving force to rotate the flagellum and is not involved in controlling the number of flagella in Vibrio. However, we observed that the E9K mutation in the rotor protein FliM confers multiple flagella. Our findings reveal a novel regulatory mechanism controlling flagellar number.

microbiology↗

bioRxiv2022

Mode selection mechanism in traveling and standing waves revealed by Min wave reconstituted in artificial cells

Reaction-diffusion coupling (RDc) generates spatiotemporal patterns, including two dynamic wave modes: traveling and standing waves. Although mode selection plays a significant role in the spatiotemporal organization of living cell molecules, the mechanism for selecting each wave mode remains elusive. Here, we investigated a wave mode selection mechanism using Min waves reconstituted in artificial cells, emerged by the RDc of MinD and MinE. Our experiments and theoretical analysis revealed that the balance of membrane binding and dissociation from the membrane of MinD determines the mode selection of the Min wave. We successfully demonstrated that the transition of the wave modes can be regulated by controlling this balance and found hysteresis characteristics in the wave mode transition. These findings highlight a novel role of the balance between activators and inhibitors as a determinant of the mode selection of waves by RDc and depict a novel mechanism in intracellular spatiotemporal pattern formations. TeaserActivator-inhibitor balance determines whether a nonlinear wave in live cells becomes a traveling wave or standing wave.

biophysics↗

bioRxiv2021

Whole-genome sequencing analysis reveals the population history of Mus musculus in Madagascar

In Madagascar, the house mouse (Mus musculus) is thought to have colonized with human activities and is now one of the most abundant rodents on the island. In this study, we determined the whole-genome sequences of five Madagascar house mice captured from the wild. We examined the evolutionary history of the population by analyzing the mitochondrial and autosomal genomes. We confirmed that the mitochondrial genome lineages of Madagascar house mice formed a monophyletic clade placed at one of the most basal positions in the species. An analysis of autosomal genomic sequences indicates that the Madagascar house mice are genetically members of M. m. castaneus (CAS), but also contain genetic elements of M. m. domesticus (DOM) resulting from hybridization between subspecies. The signature of a strong population bottleneck 1000-3000 years ago was observed in both mitochondrial and autosomal genomic data. All samples showed strong genetic affinity to many CAS samples across a wide range of Indian Ocean coastal and island regions, with divergence time estimated around 4000 years ago. These findings support that the Madagascar house mice started to colonize the island with human agricultural activity, and experienced complex history for the establishment.

genetics↗

bioRxiv2021

Evolutionary transition of doublesex regulation in termites and cockroaches: from sex-specific splicing to male-specific transcription

The sex determination gene doublesex (dsx), which encodes a transcription factor with two domains: oligomerisation domain 1 (OD1) and OD2, is conserved among insects. The sex-specific Dsx splicing isoforms regulate the transcription of target genes and trigger sex differentiation in all holometabolous insects examined to date. However, in some hemimetabolous insects, dsx is less conserved and not spliced sexually. Here, to elucidate evolutionary changes in dsx in the gene structure and its regulatory manner in termites, we searched genome and/or transcriptome databases for the OD1 and OD2 of dsx in six termite species and their sister group (woodroach). Molecular phylogenetic analysis identified OD1 sequences of termites and a woodroach clustered with dsx of holometabolous insects and regarded them as dsx orthologues. In the woodroach, a dsx orthologue containing OD2 was spliced sexually, as previously shown in other insects. However, OD2 were not found in all termite dsx orthologues. These orthologues were encoded by only a single exon in three termites with genome information; they were not alternatively spliced, but transcribed in a male-specific manner in two species examined. Evolution of dsx regulation from sex-specific splicing to male-specific transcription might be occurred at the early stage of social evolution in termites.

evolutionary biology↗

bioRxiv2021

Insights into Mus musculus subspecies population structure across Eurasia revealed by whole-genome sequence analysis

For more than 100 years, house mice (Mus musculus) have been used as a key animal model in biomedical research. House mice are genetically diverse, yet their genetic background at the global level has not been fully understood. Previous studies suggested that they originated in South Asia and diverged into three major subspecies almost simultaneously, approximately 350,000-500,000 years ago; however, they have spread across the world with the migration of modern humans in prehistoric and historic times ([~]10,000 years ago to present), and undergone secondary contact, which have complicated the genetic landscape of wild house mice. In this study, we sequenced the whole genomes of 98 wild house mice collected from Eurasia, particularly East Asia, Southeast Asia, and South Asia. We found that although wild house mice consist of three major genetic groups corresponding to the three major subspecies, individuals representing admixture between subspecies are much more ubiquitous than previously recognized. Furthermore, several samples showed an incongruent pattern of genealogies between mitochondrial and autosomal genomes. Using samples likely retaining the original genetic components of subspecies with least admixture, we estimated the pattern and timing of divergence among the subspecies. The results are important for understanding the genetic diversity of wild mice on a global level and the information will be particularly useful in future biomedical and evolutionary studies using laboratory mice established from these wild mice.

genomics↗

bioRxiv2019

ResQ, a release factor-dependent ribosome rescue factor in the Gram-positive bacterium Bacillus subtilis

Rescue of the ribosomes from dead-end translation complexes, such as those on truncated (non-stop) mRNA, is essential for the cell. Whereas bacteria use trans-translation for ribosome rescue, some Gram-negative species possess alternative and release factor (RF)-dependent rescue factors, which enable an RF to catalyze stop codon-independent polypeptide release. We now discover that the Gram-positive Bacillus subtilis has an evolutionarily distinct ribosome rescue factor named ResQ. Genetic analysis shows that B. subtilis requires the function of either trans-translation or ResQ for growth, even in the absence of proteotoxic stresses. Biochemical and cryo-EM characterization demonstrates that ResQ binds to non-stop stalled ribosomes, recruits homologous RF2, but not RF1, and induces its transition into an open active conformation. Although ResQ is distinct from E. coli ArfA, they use convergent strategies in terms of mode of action and expression regulation, indicating that many bacteria may have evolved as yet unidentified ribosome rescue systems.

molecular biology↗

bioRxiv2019

Cell-sized confinement controls generation and stability of a protein wave for spatiotemporal regulation in cells

Min system, which determines the cell division plane of bacteria, uses the localization change of protein (Min wave) emerged by a reaction-diffusion coupling. Although previous studies have shown that cell-sized space and boundaries modulate shape and speed of Min waves, its effects on Min wave emergence was still elusive. Here, by using a fully confined microsized space as a mimic of live cells, we revealed that confinement changes conditions for Min wave emergence. In the microsized space, an increase of surface-to-volume ratio changed the localization efficiency of proteins on membranes, and therefore, suppression of the localization change was necessary to produce stable Min wave generations. Furthermore, we showed that the cell-sized space more strictly limits parameters for wave emergence because confinement inhibits instability and excitability of the system. These results illuminate that confinement of reaction-diffusion systems works as a controller of spatiotemporal patterns in live cells.

biophysics↗