Biology subjects

Hayashi, R.

Publications and source records attributed to Hayashi, R..

At least 19 recordsLinked to original sources

bioRxiv2026

Phylogeny is not taxonomy: lessons on higher classification from turtle and whale barnacles (Cirripedia: Coronuloidea)

Phylogeny and taxonomy are related but not identical: phylogeny estimates common ancestry, whereas taxonomy must also provide diagnosable, stable, informative, and useful classifications. We examine this distinction using turtle and whale barnacles of the superfamily Coronuloidea, a group with an extensive fossil record, marked ecological specialization, a long history of morphology-based classification, and a recent topology-driven reclassification that merged recognized higher taxa to preserve monophyly. These changes reduced diagnosability and biological coherence, weakened historical continuity, and relied on placements insufficiently tested against morphological and phylogenetic evidence. Our reanalysis of published molecular datasets shows that critical relationships are sensitive to marker composition and taxon sampling. Together with morphological considerations on extant and fossil forms, these results indicate that topology alone is insufficient for coherent higher classification. We argue that higher taxa should be evaluated using complementary criteria: diagnosability, stability, utility, and monophyly, rather than by mechanically translating a preferred tree into Linnaean ranks.

zoology↗

bioRxiv2026

Integrated transcriptomics and proteomics define the TRP channel hierarchy in mouse cortex

Transient receptor potential (TRP) channels are evolutionarily conserved polymodal cation channels that mediate diverse sensory functions across the animal kingdom. Although TRP channels play key roles in peripheral sensation, their expression and functional relevance in the cerebral cortex remain poorly defined. Here, we integrate long- and short-read transcriptomics, targeted qPCR and membrane-aware proteomics to quantify TRP family members in adult mouse cortex. Across transcriptomic platforms, cortical TRP expression is dominated by TRPML, TRPC, and TRPM subfamilies, with lower representation of TRPP/TRPV, whereas Trpa1 and Trpv1 lie near empirical detection thresholds. Our proteomic workflow yields reproducible protein-level evidence for a subset of cortical TRPs, including TRPV2, TRPC4, TRPM3, TRPM7 and TRPP2, consistent with transcript rank order, while TRPA1/TRPV1 do not meet replicate-level protein-group detection criteria under 1% FDR control. Together, these multi-platform measurements establish a quantitative reference for cortical TRP biology and a framework for profiling low-abundance ion channels in complex brain tissue.

molecular biology↗

bioRxiv2026

A Naïve RNA Sampling Core Enables Adaptive piRNA Specificity Against Transposable Elements

How piRNA-mediated genome defense achieves specificity against transposons while sampling a complex transcriptome has remained unresolved. Here we show that piRNA biogenesis operates through pervasive, non-specific sampling of cytoplasmic RNAs, with specificity imposed by tissue-specific molecular modules that exploit intrinsic vulnerabilities of transposons. In Drosophila somatic cells, the specificity factor Yb steers basal processing towards uridine-rich RNAs--automatically capturing antisense retrotransposon transcripts due to their intrinsically adenosine-biased genomes. In germline cells lacking Yb, basal sampling generates naive piRNAs loaded into catalytically active Argonaute proteins, which trigger autocatalytic ping-pong amplification upon encountering complementary targets. In both contexts, transposon mobility facilitates the production of antisense RNAs that enable either biased processing or amplification. Thus, piRNA clusters, long associated with pathway specificity, act as sources of transposon antisense sequences, while specificity arises from layering distinct molecular mechanisms onto a shared foundation of indiscriminate transcript sampling, enabling robust and adaptable genome defense without predefined templates.

molecular biology↗

bioRxiv2025

SWARM: A Single-Molecule Workflow for High-Precision Profiling of RNA Modifications

Nanopore direct RNA sequencing promises to decode the epitranscriptome by detecting multiple modifications on individual RNA molecules, but its potential for biological discovery is hampered by high false-positive rates. We present SWARM, an AI-based framework designed to overcome this fundamental limitation. Its key innovation is a crosstalk-aware training strategy that incorporates non-target modifications and orthogonally validated cellular signals, enabling high-precision detection of m6A, pseudouridine ({Psi}), and m5C at single-nucleotide and single-molecule resolution. Using rigorous in vitro and cellular RNA benchmarks, SWARM outperforms existing tools and maintains strong agreement with orthogonal methods. Applying SWARM across mammalian tissues reveals thousands of novel modification sites with confirmed motifs and localisation patterns. Our high-resolution multi-tissue modification map revealed no evidence of widespread m6A-{Psi} interplay in predominant writer contexts, challenging models of a coordinated epitranscriptomic code. We further discovered a previously unrecognised splicing-shaped mode of {Psi} deposition, whereby TRUB1-mediated pseudouridylation preferentially occurs after exon-exon ligation, consistent with local RNA structure stabilisation. SWARM provides a robust, universally applicable tool for epitranscriptome discovery.

bioinformatics↗

bioRxiv2025

Whole-genome sequencing of CRFK and PG-4 cells to infer the phenotype of the original donors

BackgroundCrandell-Rees Feline Kidney (CRFK; kidney-derived) cells and PG-4 cells (astrocyte-derived) have been in use and have been passaged for decades in laboratories worldwide; however, no detailed information on the genetic background of the donor individuals is available, particularly regarding phenotype characteristics such as coat and iris color. ResultsWe performed whole-genome sequencing of CRFK and PG-4 cells. We analyzed the resulting data to infer the phenotype of the individual from which the cells were derived, specifically for the coat color, coat length, coat pattern, and iris color. Our data suggested that CRFK cells originated from a cat with long black fur lacking stripes and with non-blue irises; PG-4 cells originated from a cat with long bicolored white and black fur without stripes, and with non-blue irises. Analysis of publicly available RNA-seq data confirmed that genes associated with coat phenotype and iris color are expressed in the skin and eyes, as well as in various other organs. ConclusionsVariants of the genes affecting coat phenotype and iris color may influence physiological functions throughout the body. These results shed light on the previously unknown genetic background of commonly used feline cultured cells and the phenotype of the donor individuals. These findings may facilitate more accurate interpretation of data obtained from cultured feline cells and provide guidelines for developing cell lines with domestic cat genotypes exhibiting diverse phenotypes through genome editing. This will help to provide clarity regarding deafness in cats with white fur and blue irises and elucidate the effects of melanocyte destruction caused by KIT gene mutations on the nervous system and the influence of other coat-related factors in organs other than the skin or functions independent of coat formation.

genomics↗

bioRxiv2025

A fish finder for birds: route-dependent foraging behavior of the brown booby (Sula leucogaster) on a passenger ferry

This manuscript reports a novel case of route-dependent foraging behavior in the brown booby (Sula leucogaster), based on visual observations from a passenger ferry between Chichijima and Hahajima Islands, Ogasawara, Japan. On four dates in 2017, the number of accompanying boobies and their plunge-diving behavior were recorded using only handheld counters. More birds and dives were observed on routes departing from the nesting-rich Hahajima, with peak activity mid-route. These findings suggest that the ferry acts as a "fish finder," flushing prey such as flying fish for the birds to exploit. The study highlights how low-tech observations from public transport can provide valuable behavioral insights and underscores the potential of citizen science in seabird ecology.

ecology↗

bioRxiv2025

Integrative taxonomic revision of turtle barnacle genus Platylepas (Cirripedia: Coronuloidea): an ancient host-range expansion to sea snakes and sirenians

The barnacle genus Platylepas contains obligate epibionts of marine vertebrates, but the taxonomic status and evolutionary history of the sea-snake specialist, P. ophiophila, have remained unclear. Here, we reassess species boundaries and host associations in Platylepas using shell-morphological comparisons and sequence data from mitochondrial COI and nuclear H3 genes from specimens collected from sea turtles, sea snakes, and a dugong. Our results show that P. ophiophila is a valid species, phylogenetically and morphologically distinct from the widespread turtle barnacles P. decorata and P. hexastylos. Furthermore, we report the first record of P. ophiophila from a sirenian host (Dugong dugon), a discovery that necessitates the re-examination of historical records of barnacles on sirenians worldwide. Our phylogeny places P. decorata, a turtle epibiont, as the earliest diverging lineage within the genus. This is consistent with the hypothesis that the ancestor of Platylepas was associated with sea turtles, a conclusion supported by the ichnofossil record. The subsequent divergence of the P. ophiophila clade may represent an important host-use transition: an ancient host shift from sea turtles to the taxonomically disparate but ecologically linked inhabitants--sea snakes and sirenians--of seagrass ecosystems. This study clarifies species limits within Platylepas and provides a framework for discussing host associations and potential host shifts in light of existing molecular and fossil evidence.

zoology↗

bioRxiv2025

Conserved but mechanistically diverse piRNA defence against gypsy LTR retrotransposons in insect ovaries

Organisms have evolved highly adaptable defence systems to silence mobile genetic elements such as transposons. While individual transposons may emerge and vanish over time, the long-term evolutionary stability of the pathways that suppress them remains poorly understood. In the ovarian somatic cells of Drosophila, envelope-carrying gypsy long terminal repeat (LTR) retrotransposons are silenced by the Piwi-interacting RNA (piRNA) pathway, which depends exclusively on Piwi - an evolutionary derivative of an Aubergine/Piwi gene duplication unique to flies. Recent studies, however, have revealed that envelope-carrying gypsy elements are widespread across metazoans and particularly abundant in insect genomes, raising questions about the origin of ovarian somatic piRNA defence. Here we demonstrate that Aedes and Anopheles mosquitoes, as well as stingless bees and crickets, all produce piRNAs targeting gypsy elements in ovarian somatic cells, indicating a continuous arms race in this cellular niche for more than 400 million years of insect evolution. Notably, in Aedes aegypti, ovarian somatic cells express the same piRNA clusters as other somatic tissues, where they are known to target RNA viruses - suggesting a shared origin of anti-viral and anti-retrotransposon defences. Furthermore, we reveal striking lineage-specific differences in ovarian somatic piRNA biogenesis: slicing-independent phasing dominates in dipterans, ping-pong amplification in bees, and slicing-dependent phasing in crickets. Together, these findings indicate that distinct piRNA pathways have independently evolved at different timepoints to silence the same class of retrotransposons in insect evolution.

evolutionary biology↗

bioRxiv2025

Single-molecule multimodal timing of in vivo mRNA synthesis

mRNA synthesis requires extensive pre-mRNA maturation, the organisation of which remains unclear. Here, we directly sequence pre-mRNA without metabolic labelling or amplification to resolve transcription and multimodal pre-mRNA processing at single-molecule resolution. Using poly(A) tail measurement, we distinguish transcriptionally engaged pre-mRNA from polyadenylated transcripts, revealing that splicing is substantially delayed compared to previous estimates. Splicing is rare for at least 10 kb behind elongating RNA polymerase II such that thousands of genes remain largely unspliced during transcription, revising the notion that splicing is predominantly co-transcriptional. Progressive splicing becomes apparent only on polyadenylated transcripts, suggesting that splicing is commonly activated after 3' end formation. Unexpectedly, we find abundant m6A on unspliced pre-mRNA, indicating that a substantial portion of RNA methylation precedes splicing. This m6A methylation is suppressed around exon boundaries, indicating that m6A topology is established prior to exon junction complex deposition. Finally, we demonstrate that 3' end cleavage occurs multiple kilobases behind transcription in coordination with efficient transcription termination, and further highlight recursive 3' end formation across hundreds of genes. Conserved from human cells to mouse tissues, we illustrate a revised timeline for mRNA synthesis wherein cleavage, termination, and m6A deposition occur earlier than, or at least partially decoupled from, the bulk of splicing, reframing the sequence of early mammalian gene expression.

genomics↗

bioRxiv2024

Non-gonadal PIWI protein, Aubergine, regulates regenerative stem cell proliferation and tumourigenesis in the Drosophila adult intestine.

The PIWI-interacting RNA (piRNA) biosynthesis pathway is best studied for its role suppressing Drosophila germline transposable elements. Piwi, the founding member of the pathway, is involved in adult intestinal stem cell (ISC) homeostasis. Whether a broader role of the PIWI pathway exists in the intestine, remains unknown. Here, we characterise a role of the PIWI family protein Aubergine (Aub) in ISCs. While dispensable for basal ISC self-renewal, upregulation of Aub by damage-induced reactive oxygen species drives regenerative ISC proliferation through increased protein synthesis, including translation of ISC factors Myc and Sox21a. Unexpectedly, such roles of Aub in ISCs appear uncoupled from its piRNA regulatory function. Additionally, Aub and mammalian PIWIL1, mediate tumorigenic intestinal growth in Drosophila and human organoids, respectively. Our results reveal regulated protein translation as a fundamental aspect of regenerative ISC function and discover a central role of Aub in such process.

developmental biology↗

bioRxiv2024

A germline PAF1 paralog complex ensures cell type-specific gene expression

Animal germline development and fertility rely on paralogs of general transcription factors that recruit RNA polymerase II to ensure cell type-specific gene expression. It remains unclear whether gene expression processes downstream of such paralog-based transcription is distinct from that of canonical RNA polymerase II genes. In Drosophila, the testis-specific TBP-associated factors (tTAFs) activate over a thousand spermatocyte-specific gene promoters to enable meiosis and germ cell differentiation. Here, we show that efficient termination of tTAF-activated transcription relies on testis-specific paralogs of canonical Polymerase Associated Factor 1 Complex (PAF1C) proteins, which form a testis-specific PAF1C (tPAF). Consequently, tPAF mutants cause aberrant expression of hundreds of downstream genes due to read-in transcription. Furthermore, tPAF facilitates expression of Y-linked male fertility factor genes, and thus broadly maintains spermatocyte-specific gene expression. Consistently, tPAF is required for the segregation of meiotic chromosomes and male fertility. Supported by comparative in vivo protein interaction assays, we provide a mechanistic model for the functional divergence of tPAF and PAF1C and for transcription termination as a developmentally regulated process required for cell type-specific gene expression.

molecular biology↗

bioRxiv2023

Ancient origins of endogenous retroviruses in Metazoans

Retrotransposons or endogenous retroviruses (ERVs) essentially carry open reading frames of gag and pol, which are utilized to selfishly replicate themselves in the host germline genome. One rare example of ERVs that additionally carry envelope genes is Ty3/gypsy errantiviruses in Drosophila. Though they are structurally analogous to retroviruses, it remained unclear whether envelope-containing Ty3/gypsy elements represent recent, lineage-specific acquisitions of viral fusogens or an ancient association between retrotransposons and envelope-like genes. We systematically searched for intact envelope-containing ERVs that are homologous to Ty3/gypsy in invertebrate metazoan genomes and found that they are widespread across taxa including ancient animals such as cnidarians, ctenophores and tunicates. Many elements occur as multiple highly similar copies in their respective genomes, consistent with recent genomic expansion in some host lineages. Envelope genes are classified into those that resemble glycoprotein F from paramyxoviruses and glycoprotein B from herpesviruses, and both types are equally abundant and widespread. Phylogenetic and structural analyses revealed that envelope genes have largely diverged with pol genes as well as with the host organisms throughout their evolutionary history and recombined infrequently, suggesting that the envelope acquisition to ERVs is ancient and likely dates to before the split of bilaterian and non-bilaterian animals in Pre-Cambrian era.

evolutionary biology↗

bioRxiv2022

Mechanistic divergence of piRNA biogenesis in Drosophila

Organisms require mechanisms to distinguish self and non-self RNA. This distinction is crucial to initiate the biogenesis of piRNAs. In Drosophila ovaries, PIWI-guided slicing and the recognition of piRNA precursor transcripts by the DEAD-box RNA helicase Yb are the two known mechanisms to licence an RNA for piRNA biogenesis in the germline and the soma, respectively. Both, the PIWI proteins and Yb are highly conserved across most Drosophila species and are thought to be essential to the piRNA pathway and for silencing transposons. However, we find that species closely related to D. melanogaster have lost the yb gene, as well as the PIWI gene Ago3. We show that the precursor RNA is still selected in the absence of Yb to abundantly generate transposon antisense piRNAs in the soma. We further demonstrate that D. eugracilis, which lacks Ago3, is completely devoid of ping-pong piRNAs and exclusively produces phased piRNAs in the absence of slicing. Thus, there are more possible routes through which the piRNA pathway can achieve specificity than previously suggested.

molecular biology↗

bioRxiv2022

Mechanistic insights into RNA surveillance by the canonical poly(A) polymerase Pla1 of the MTREC complex.

The S. pombe orthologue of the human PAXT complex, Mtl1-Red1 Core (MTREC), is an eleven-subunit complex which targets cryptic unstable transcripts (CUTs) to the nuclear RNA exosome for degradation. It encompasses the canonical poly(A) polymerase Pla1, responsible for polyadenylation of nascent RNA transcripts as part of the cleavage and polyadenylation factor (CPF/CPSF). In this study we identified and characterised the interaction between Pla1 and the MTREC complex core component Red1 and analysed the functional relevance of this interaction in vivo. Our crystal structure of the Pla1-Red1 complex showed that a 58-residue fragment in Red1 binds to the RNA recognition motif domain of Pla1 and tethers it to the MTREC complex. Structure-based Pla1-Red1 interaction mutations showed that Pla1, as part of MTREC complex, hyper-adenylates CUTs for their efficient degradation. Interestingly, the Red1-Pla1 interaction was also required for the efficient assembly of the fission yeast facultative heterochromatic islands. Together, our data suggest a complex interplay between the RNA surveillance and 3-end processing machineries.

molecular biology↗

bioRxiv2022

Identification of m6A and m5C RNA modifications at single-molecule resolution from Nanopore sequencing

The epitranscriptome embodies many new and largely unexplored functions of RNA. A significant roadblock hindering progress in epitranscriptomics is the identification of more than one modification in individual transcript molecules. We address this with CHEUI (CH3 (methylation) Estimation Using Ionic current). CHEUI predicts N6-methyladenosine (m6A) and 5-methylcytidine (m5C) in individual molecules from the same sample, the stoichiometry at transcript reference sites, and differential methylation between any two conditions. CHEUI processes observed and expected nanopore direct RNA sequencing signals to achieve high single-molecule, transcript-site, and stoichiometry accuracies in multiple tests using synthetic RNA standards and cell line data. CHEUIs capability to identify two modification types in the same sample reveals a co-occurrence of m6A and m5C in individual mRNAs in cell line and tissue transcriptomes. CHEUI provides new avenues to discover and study the function of the epitranscriptome.

bioinformatics↗

bioRxiv2021

The Mechanism of Neurite Outgrowth Induction by Novel Synthetic Retinobenzoic Acids

Retinoids are a family of vitamin A-derived molecules and include the biologically active metabolite, retinoic acid (RA). RA acts as a specific modulator of neuronal differentiation and proliferation. However, teratogenicity and a large excess of RA have been found in animal studies. Thus, development of effective and stable retinoids is desirable. In this study, we showed that treatment with novel synthetic retinobenzoic acids promotes neurite outgrowth in a selected subpopulation of the human neuroblastoma cell line SK-N-SH. Furthermore, we found that, although acting via a different mechanism, retinobenzoic acids have the same neurite outgrowth-inducing effect as RA. Retinoids, including RA, bind to nuclear retinoic acid receptors (RARs). Therefore, we examined the expression of RARs in retinobenzoic acid-treated cells. Similar to already known retinoids, novel synthetic retinobenzoic acids promote the upregulation of RAR{beta} and have no effect on RAR or {gamma}. These results suggest that retinobenzoic acids act via RAR{beta} during neurite outgrowth. Moreover, stimulation with RA or retinobenzoic acids significantly increased the phosphorylation levels of both ERK1/2 and mTOR. ERK1/2 and mTOR inhibition blocked the retinobenzoic acid-induced increase in neurite outgrowth, suggesting that retinobenzoic acids promoted neurite outgrowth by activating the ERK1/2 and mTOR signaling pathways. Notably, the RA-induced increase in neurite outgrowth was blocked by the ERK1/2 inhibitor U0126, but not by the mTOR inhibitor rapamycin. In addition, ERK1/2 inhibition blocked the upregulation of RAR{beta} promoted by RA and retinobenzoic acids. In contrast, mTOR inhibition had no effect on upregulation of RAR{beta}. Our results show that novel synthetic retinobenzoic acids induce neurite outgrowth by a different mechanism than RA. These findings suggest that activation of both ERK1/2, which results in downstream regulation of RAR{beta}, and mTOR, are responsible for the novel synthetic retinobenzoic acid-induced neurite outgrowth in human neuroblastoma cells.

cell biology↗

bioRxiv2020

Reduced intra- and inter-individual diversity of semantic representations in the brains of schizophrenia patients

Schizophrenia patients often manifest semantic processing deficits. It has been proposed that these deficits stem from disorganized semantic representations in the brain. However, no study has yet examined the neural correlates of semantic disorganization by directly evaluating semantic representations in the brain. We used voxelwise modeling on functional magnetic resonance imaging signals to evaluate the semantic representations associated with several thousand words in individual patient brains. We then compared the structural properties of semantic representations to those in healthy controls. The variability of semantic representations was smaller both within individual patients and across patients compared to controls. Surrogate data analysis suggests that the observed reduction in representational variability is associated with disorganization of categorical information. To our knowledge, these findings provide the first evidence for sematic disorganization in schizophrenia at the level of brain representations.

neuroscience↗

bioRxiv2020

Lab-on-a-graphene-FET detection of key molecular events underpinning influenza virus infection and effect of antiviral drugs

Small solid-state devices are candidates for accelerating biomedical assays/drug discovery, however their potential remains unfulfilled. Here, we demonstrate that graphene-field effect transistors (FET) can be used to successfully detect the key molecular events underlying viral infections and the effect of antiviral drugs. Our device success is achieved by bio-mimicking the host-cell surface during an influenza infection at the graphene channel. In-situ AFM confirms the biological interactions at the sialic acid-functionalized graphene: viral hemagglutinin (HA) binds to sialic acid, and neuraminidase (NA) reacts with the sialic acid-HA complex. The graphene-FET detects HA binding to sialic acid, and NA cleavage of sialic acid. The inhibitory effect of the drug "zanamivir" on NA-sialic acid interactions is monitored in real-time; the reaction rate constant of NA-sialic acid reaction was successfully determined. We demonstrate that graphene-FETs are powerful platforms for measurement of biomolecular interactions and contribute to future deployment of solid-state devices in drug discovery/biosensing.

bioengineering↗