Biology subjects

Niimi, T.

Publications and source records attributed to Niimi, T..

16 recordsLinked to original sources

bioRxiv2026

Static allometry of the horn and pronotal depression in adult Oryctes rhinoceros supports continuous nonlinearity, sexual dimorphism, and size-independent covariation

Exaggerated insect traits often show positive allometry, yet nonlinear scaling can reflect either continuous curvature or discrete morphs. Distinguishing between these alternatives is important because they imply different developmental and evolutionary scenarios. Using cross-sectional data from 1,000 adult Oryctes rhinoceros, we analyzed the static allometry of nine traits with pronotum width as the primary body-size proxy and body length for sensitivity analyses. Cross-validated comparisons among linear, continuous nonlinear, and two-component mixture models showed that continuous nonlinear models improved predictive performance over linear models for horn length and pronotal depression width in both sexes. By contrast, mixture regression did not outperform the best continuous model in either sex, providing no positive support for discrete within-sex dimorphism under the tested model set. After accounting for body size, horn length remained positively associated with pronotal depression width, indicating size-independent covariation. These associations were retained when body length was used instead of pronotum width, supporting robustness to body-size proxy choice. Together, these results support continuous nonlinear adult scaling of the horn and pronotal depression in O. rhinoceros and indicate covariation not attributable solely to body size under the tested model set.

zoology↗

bioRxiv2025

Expression dynamics and functional characterization of the pigmented and non-diapausing egg gene (pnd) and pnd-2, responsible for the initiation of embryonic diapause in the silkworm, Bombyx mori

Diapause in insects is a developmental arrest that serves as an adaptation to severe environmental conditions. In the silkworm Bombyx mori, embryonic diapause in the next generation is maternally induced by the diapause hormone (DH). However, the downstream factors responsible for initiating diapause have remained unidentified. The pnd and pnd-2 mutants produce pigmented and non-diapausing eggs despite intact DH signaling, indicating that the pnd and pnd-2 genes act downstream as potential initiators of diapause. Here, we analyzed their expression dynamics and functions. Expression analyses showed that pnd was strongly upregulated in diapause-destined eggs in a DH-signal-dependent manner and was suppressed by HCl treatment that prevents diapause initiation. In contrast, pnd-2 exhibited a transient expression peak independent of DH-signal and was unaffected by HCl treatment. RNAi-mediated knockdown of either gene inhibited the diapause initiation, and injection of pnd mRNA into non-diapause-destined eggs induced a diapause-like state in embryos. These findings demonstrate that pnd and pnd-2 are required downstream of the DH-signal to initiate diapause, providing key insights into the molecular mechanism underlying embryonic diapause induction in B. mori. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=128 SRC="FIGDIR/small/667111v1_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@7e9516org.highwire.dtl.DTLVardef@6dddc2org.highwire.dtl.DTLVardef@fa11baorg.highwire.dtl.DTLVardef@1a2054e_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2025

Sex-specific doublesex regulation targeting the color-patterning gene h underlies the evolution of wing sexual dimorphism in the harlequin ladybug Harmonia axyridis

Organisms on Earth show various forms of sexual dimorphism, including ornaments, weapon traits, and pheromone glands, which has been acquired through sexual selection during evolution. Although the genetic basis of sexual trait has been investigated in diverse species, its evolutionary origin, evolutionary mode, and evolvability remain poorly understood. To address these issues, we investigated the strain-specific sexual dimorphism in elytral color patterns of the harlequin ladybug, Harmonia axyridis, a species with over 200 color morphs. The most basal Red-nSpots type color morph exhibits sexual dimorphism, whereas other derived color morphs have lost it. To investigate how this loss of sexual dimorphism is associated with the emergence of novel color morphs, we investigated the genetic basis of sexual dimorphism by focusing on the master sex differentiation gene, doublesex (dsx). We show that dsx regulates color pattern dimorphism by negatively modulating black spot size in males. This modulation is primarily mediated by the transcriptional regulation of the color patterning gene, h (Drosophila pannier ortholog). Intraspecific comparative ATAC-seq analysis of the pupal wings revealed that, at the h locus, not the absolute number of Dsx-binding motifs but the proportion of open chromatin regions containing Dsx-binding motifs relative to those lacking such motifs was reduced in strains that had lost sexual dimorphism and acquired novel color patterns, implying that sexual dimorphism evolves based on the balance between novel CREs and Dsx-binding motif density. The present study provides a fundamental molecular framework for understanding how a secondary sexual trait evolves within H. axyridis.

evolutionary biology↗

bioRxiv2025

A highly efficient knock-in strategy using fast-annealed 3'-extended dsDNA templates for emerging model insects

CRISPR-Cas genome editing toolkits have expanded the scope of genetic studies in various emerging model organisms. However, their applications are limited mainly to knockout experiments due to technical difficulties in establishing knock-in strains, which enable in vivo molecular tagging-based experiments. Here, we investigated knock-in strategies in the harlequin ladybug Harmonia axyridis, a model insect for evolutionary developmental biology, which shows more than 200 color pattern variations within a species. We tested several knock-in strategies using synthetic DNA templates. We found that ssDNA templates generated founder knock-in strains efficiently (2.5-11%), whereas the 5 regions of ssDNA templates were frequently deleted when the insert length exceeded [~]40 bases. To overcome this limitation, we designed several 3 extended DNA templates. Fast-annealed 3-extended double-stranded DNA templates, which were designed for tagging endogenous proteins with epitope tags, showed high founder generation efficiency (9.9-20.9%) and accuracy (30.8-85.7%). This strategy is also applicable to the two-spotted cricket Gryllus bimaculatus, suggesting that the fast-annealed 3-extended dsDNA template is a versatile DNA template for generating knock-in strains in emerging model insects for developmental genetic studies. Summary statementFast-annealed 3-extended dsDNA templates facilitate efficient CRISPR-Cas9-mediated knock-in in emerging model insects.

bioengineering↗

bioRxiv2024

Morphological Study of Left-right Head Asymmetry in Doubledaya bucculenta (Coleoptera: Erotylidae: Languriinae)

Left-right asymmetry in paired organs is well-documented across various species, including the claws of fiddler crabs and snail-eating snakes dentition. However, the mechanisms underlying these asymmetries remain largely elusive. This study investigates Doubledaya bucculenta (Coleoptera: Erotylidae), a lizard beetle species known for pronounced left-sided asymmetry in adult female mandible and gena. Given that insect mouthparts comprise multiple functionally significant appendages, we aimed to clarify the degree of asymmetry extending beyond the mandibles and genae. Phenotypic morphology was assessed through trait measurement and asymmetry index calculations. Our detailed morphometric analyses revealed left-longer asymmetry not only in mandibles and genae but also in maxillae and labium. Notably, the degree of asymmetry in other mouthparts was generally less pronounced compared to outer mandibles, suggesting a potential influence of left mandible development on other mouthparts. Additionally, male mandibles exhibited region-specific asymmetry, potentially indicative of constrained evolutionary adaptations. This study enhances a comprehensive understanding of adult phenotype morphology and offers insights into the developmental basis of asymmetrical mouthparts.

zoology↗

bioRxiv2023

Evolution of horn length and lifting strength in the Japanese rhinoceros beetle Trypoxylus dichotomus

Rhinoceros beetle (Trypoxylus dichotomus) males have pitchfork-shaped head horns, which they use to pry rival males from the trunks of trees. In the largest males these horns can be three times the length of horns in the two closest sister species. Because this weapon functions as a lever, longer horns should lift with less force than shorter horns (the paradox of the weakening combatant) unless other elements of the weapon system (e.g., input lever length, muscle mass) evolve to compensate. We used next-generation sequencing approaches to consolidate 23 sample locations into 8 genetically distinguishable populations, reconstructing their historical relationships and providing a comprehensive picture of the evolution of this horn lever system. We show that head horns likely increased in length independently in the Northern and Southern lineages. In both instances this resulted in weaker lifting forces, but this mechanical disadvantage was later ameliorated, to some extent and in some locations, by subsequent reductions to horn length, changes in muscle size, or by an increase in input lever length (head height). Our results reveal an exciting geographic mosaic of differences in weapon size, weapon force, and in the extent and nature of mechanical compensation. Reconstructing the evolution of this weapon system offers critical insights towards meaningfully linking mating system dynamics, selection patterns, and diversity in sexually selected traits.

evolutionary biology↗

bioRxiv2022

The draft genome sequence of Japanese rhinoceros beetle Trypoxylus dichotomus

Beetles are the largest insect order and one of the most successful animal groups in terms of number of species. The Japanese rhinoceros beetle Trypoxylus dichotomus (Coleoptera, Scarabaeidae, Dynastini) is a giant beetle with distinctive exaggerated horns present on the head and prothoracic regions of the male. T. dichotomus has been used as research model in various fields such as evolutionary developmental biology, ecology, ethology, biomimetics, and drug discovery. In this study, de novo assembly of 615 Mb, representing 80% of the genome estimated by flow cytometry, was obtained using the 10x Chromium platform. The scaffold N50 length of the genome assembly was 8.02 Mb, with repetitive elements predicted to comprise 49.5% of the assembly. In total, 23,987 protein-coding genes were predicted in the genome. In addition, de novo assembly of the mitochondrial genome yielded a contig of 20,217 bp. We also analyzed the transcriptome by generating 16 RNA-seq libraries from a variety of tissues of both sexes and developmental stages, which allowed us to identify 13 co-expressed gene modules. The detailed genomic and transcriptomic information of T. dichotomus is the most comprehensive among those reported for any species of Dynastinae. This genomic information will be an excellent resource for further functional and evolutionary analyses, including the evolutionary origin and genetic regulation of beetle horns and the molecular mechanisms underlying sexual dimorphism.

genomics↗

bioRxiv2021

Evolutionary history of sexual differentiation mechanism in insects

Gain of alternative splicing gives rise to functional diversity in proteins and underlies the complexity and diversity of biological aspects. However, it is still not fully understood how alternatively spliced genes develop the functional novelty. To this end, we infer the evolutionary history of the doublesex gene, the key transcriptional factor in the sexual differentiation of arthropods. doublesex is controlled by sex-specific splicing and promotes both male and female differentiation in some holometabolan insects. In contrast, doublesex promotes only male differentiation in some hemimetabolan insects. Here, we investigate ancestral states of doublesex using Thermobia domestica belonging to Zygentoma, the sister group of winged insects. We find that doublesex of T. domestica expresses sex-specific isoforms but is only necessary for male differentiation of sexual morphology. This result ensures the hypothesis that doublesex was initially only used to promote male differentiation during insect evolution. However, T. domestica doublesex has a short female-specific region and upregulates the expression of vitellogenin homologs in females, suggesting that doublesex may have already controlled some aspects of feminization in the common ancestor of winged insects. Reconstruction of the ancestral sequence and prediction of the protein structure show that the female-specific isoform of doublesex has a long C-terminal disordered region in holometabolan insects, but not in non-holometabolan species. We propose that doublesex acquired a female-specific isoform and then underwent a change in the protein motif structure, which became essential for female differentiation in sexual dimorphisms.

evolutionary biology↗

bioRxiv2021

Evolutionary origin of sex differentiation system in insects

The evolution of the functionality of genes and genetic systems is a major source of animal diversity. Its best example is insect sex differentiation systems: promoting male and female differentiation (dual-functionality) or only male differentiation (single-functionality). However, the evolutionary origin of such functional diversity is largely unknown. Here, we investigate the ancestral functions of doublesex, a key factor of insect sex differentiation system, using the apterygote insect, Thermobia domestica, and reveal that its doublesex is essential for only males at the phenotypic level, but contributes to promoting female-specific vitellogenin expression in females. This functional discordance between the phenotypic and transcription-regulatory levels in T. domestica shows a new type of functionality of animal sex differentiation systems. Then, we examine how the sex differentiation system transited from the single-functionality to the dual-functionality in phenotypes and uncover that a conserved female-specific motif of doublesex is detected in taxa with the dual-functional doublesex. It is estimated that the role of the sex differentiation system for female phenotypes may have evolved through accumulating mutations in the protein motif structures that led to the enhancement of its transcription-regulatory function.

evolutionary biology↗

bioRxiv2021

Genomic and transcriptomic analyses of the subterranean termite Reticulitermes speratus: gene duplication facilitates social evolution

Termites are model social organisms characterized by a polyphenic caste system. Subterranean termites (Rhinotermitidae) are ecologically and economically important species, including acting as destructive pests. Rhinotermitidae occupies an important evolutionary position within the clade representing an intermediate taxon between the higher (Termitidae) and lower (other families) termites. Here, we report the genome, transcriptome and methylome of the Japanese subterranean termite Reticulitermes speratus. The analyses highlight the significance of gene duplication in social evolution in this termite. Gene duplication associated with caste-biased gene expression is prevalent in the R. speratus genome. Such duplicated genes encompass diverse categories related to social functions, including lipocalins (chemical communication), cellulases (wood digestion and social interaction), lysozymes (social immunity), geranylgeranyl diphosphate synthase (social defense) and a novel class of termite lineage-specific genes with unknown functions. Paralogous genes were often observed in tandem in the genome, but the expression patterns were highly variable, exhibiting caste biases. Some duplicated genes assayed were expressed in caste-specific organs, such as the accessory glands of the queen ovary and frontal glands in soldier heads. We propose that gene duplication facilitates social evolution through regulatory diversification leading to caste-biased expression and subfunctionalization and/or neofunctionalization that confers caste-specialized functions. Significance StatementTermites are model social organisms characterized by a sophisticated caste system, where distinct castes arise from the same genome. Our genomics data of Japanese subterranean termite provides insights into the evolution of the social system, highlighting the significance of gene duplication. Gene duplication associated with caste-biased gene expression is prevalent in the termite genome. Many of the duplicated genes were related to social functions, such as chemical communication, social immunity and defense, and they often expressed in caste-specific organs. We propose that gene duplication facilitates social evolution through regulatory diversification leading to caste-biased expression and functional specialization. In addition, since subterranean termites are ecologically and economically important species including destructive pests in the world, our genomics data serves as a foundation for these studies.

genomics↗

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

A wing growth organizer in a hemimetabolous insect suggests wing origin

The origin and evolution of insect wings remain enigmatic after a century-long discussion. Molecular dissection of wing development in hemimetabolous insects, in which the first functional wings evolved, is key to understand genetic changes required for wing evolution. We investigated Drosophila wing marker genes in the cricket, Gryllus bimaculatus, and found apterous and vestigial show critical functions in nymphal tergal identity and margin formation, respectively. We further demonstrate that margin cells in the lateral-anterior tergal region constitute a growth organizer of wing blades. Transcriptome and RNAi analyses unveiled that Wnt, Fat-Dachsous, and Hippo pathways are involved in disproportional growth of Gryllus wings. Our data collectively support the idea that tergal margin cells of a wingless ancestor gave rise to the body wall extension required for evolution of the first powered flight.

developmental biology↗

bioRxiv2020

Genetical control of 2D pattern and depth of the primordial furrow that codes 3D shape of the rhinoceros beetle horn

The head horn of the Asian rhinoceros beetle develops as extensively folded primordia before unfurling into its final 3D shape at the pupal molt. The information of the final 3D structure of the beetle horn is encoded in the folding pattern of the developing primordia. However, the developmental mechanism underlying epithelial folding of the primordia is unknown. In this study, we addressed this gap in our understanding of the developmental patterning of the 3D horn shape of beetles by focusing on the formation of surficial furrows that become the bifurcated 3D shape of the horn. By gene knockdown screening via RNAi, we found that knockdown of the gene Notch disturbed overall horn primordia furrow depth without affecting 2D furrow pattern. In contrast, knockdown of CyclinE altered 2D horn primordia furrow pattern without affecting furrow depth. From these results, depth and 2D pattern of primordial surficial furrow are likely to be regulated independently during the development and both of change can alter the final 3D shape. Author SummaryIn insects, some large structure is made under the old exoskeleton before the molting. Long horn of rhino-beetle is one of extreme cases. The beetle horn is compactly packed as furrowed primordia under the larval exoskeleton. At molting, the primordia is extended to form its final 3D horn shape as blowing up furrows like a balloon. This transformation from primordia to final horn does not required any living cell activities. Thus, characteristics of furrows of primordia actually determine the final 3D shape. However, molecular mechanisms and genetic basis of furrow formation is not well understood not only in beetle horn but also in any other insects. In this study, by using beetle horn as a model, we addressed what kind of genetic factors are contributed to primordial furrow formation. By gene knockdown screening, we found that knockdown of the gene Notch disturbed primordial furrow depth without affecting 2D furrow pattern. In contrast, knockdown of CyclinE altered 2D furrow pattern without affecting furrow depth. In both case, final horn shapes were disturbed. From these results, we concluded that both of the depth and 2D pattern of primordial furrow can contribute final shape, but their development is controlled independently.

developmental biology↗

bioRxiv2019

Acute feeding disorder after oral RNAi exposure results in efficient pest control

Selecting an appropriate target gene is critical to the success of feeding RNA interference (f-RNAi)-based pest control. Gene targets have been chosen based on their ability to induce lethality. However, lethality induction by f-RNAi is slow-acting and crop damage can progress during this time. Here, we show that f-RNAi of death-associated inhibitor of apoptosis protein 1 (diap1), but not two conventional targets vacuolar ATPase subunit A and E, induces acute feeding cessation in the solanaceous pest, Henosepilachna vigintioctopunctata during 24-48 hours. We also found that the feeding cessation by diap1 f-RNAi has species-specificity and occurs with only 1.6 ng dsRNA. Our results suggest that diap1 is an appropriate target in the context of rapid reduction of crop damage. We propose that acute feeding disorder should be assessed as a novel criterion for selecting appropriate target genes for RNAi-based pest control in addition to the conventional criterion based on lethality.

molecular biology↗

bioRxiv2018

Repeated inversions at the pannier intron drive diversification of intraspecific colour patterns of ladybird beetles

How genetic information is modified to generate phenotypic variation within a species is one of the central questions in evolutionary biology. Here we focus on the striking intraspecific diversity of more than 200 aposematic elytral (forewing) colour patterns of the multicoloured Asian ladybird beetle, Harmonia axyridis, which is regulated by a tightly linked genetic locus h. Our loss-of-function analyses, genetic association studies, de novo genome assemblies, and gene expression data reveal that the GATA transcription factor gene pannier is the major regulatory gene located at the h locus, and suggest that repeated inversions and cis-regulatory modifications at pannier led to the expansion of colour pattern variation in H. axyridis. Moreover, we show that the colour patterning function of pannier is conserved in the seven spotted ladybird beetle, Coccinella septempunctata, suggesting that H. axyridis extraordinary intra-specific variation may have arisen from ancient modifications in a conserved elytral colour patterning mechanisms in ladybird beetles.

evolutionary biology↗

bioRxiv2018

doublesex regulates sexually dimorphic beetle horn formation by integrating spatial and temporal developmental contexts in the Japanese rhinoceros beetle Trypoxylus dichotomus

AbstractMany scarab beetles have sexually dimorphic exaggerated horns that are an evolutionary novelty. Since the shape, number, size, and location of horns are highly diverged within Scarabaeidae, beetle horns are an attractive model for studying the evolution of sexually dimorphic and novel traits. In beetles including the Japanese rhinoceros beetle Trypoxylus dichotomus, the sex determination gene doublesex (dsx) plays a crucial role in sexually dimorphic horn formation during larval-pupal development. However, knowledge of when and how dsx drives the gene regulatory network (GRN) for horn formation to form sexually dimorphic horns during development remains elusive. To address this issue, we identified a Trypoxylus-ortholog of the sex determination gene, transformer (tra), that regulates sex-specific splicing of the dsx pre-mRNA, and whose loss of function results in sex transformation. By knocking down tra function at multiple developmental timepoints during larval-pupal development, we estimated the onset when the sex-specific GRN for horn formation is driven. In addition, we also revealed that dsx regulates different aspects of morphogenetic activities during the prepupal and pupal developmental stages to form appropriate morphologies of pupal head and thoracic horn primordia as well as those of adult horns. Based on these findings, we discuss the evolutionary developmental background of sexually dimorphic trait growth in horned beetles.\n\nAuthor Summary:\n\nBeetle horns are highly enriched in a particular family Scarabaeidae, although the shape, size and number of horns are diversified within the group. In addition, many scarab beetle horns are sexually dimorphic. It has been questioned how a particular group of beetles has originated and diversified evolutionary novel horns. Here we found the exact time when morphological sexual dimorphism of horn primordia appeared, estimated the onset of the developmental program for sexually dimorphic horn formation driven by Doublesex, and revealed that Doublesex regulates different aspects of cell activities of horn primordia depending on the spatiotemporal contexts. Our study provides our understanding regarding regulatory shifts in these mechanisms during the evolution of sexually dimorphic traits in horned beetles.

developmental biology↗