Biology subjects

Tanaka, K. M.

Publications and source records attributed to Tanaka, K. M..

6 recordsLinked to original sources

bioRxiv2025

Alternative Splicing in TRPA1 Drives Sensory Adaptation to Electrophiles in Drosophilids

Behaviors are among the first traits to evolve as animals enter new niches, but their molecular bases are poorly understood. To address this gap, we used the mustard-feeding drosophilid fly Scaptomyza flava, which feeds on toxic Brassicales plants like wasabi that produce noxious, electrophilic isothiocyanates (ITCs or mustard oils). We found that S. flava exhibits dramatically reduced behavioral sensitivity to allyl isothiocyanate (AITC) compared to its microbe-feeding relatives Scaptomyza pallida and Drosophila melanogaster. We hypothesized that molecular evolution of the "wasabi receptor" TRPA1, known to detect ITCs in flies, could explain this loss of aversion. Our experiments revealed three interconnected evolutionary genetic changes consistent with this hypothesis. First, TRPA1 was expressed in labellar tissues of S. flava at the lowest levels among the three species, at a nearly four-fold lower level than in D. melanogaster. Second, S. flava expressed a higher proportion of TRPA1 splice variants previously reported to be less sensitive to chemical stimulus. Third, we identified amino acid substitutions in S. flava that could influence the structure of intracellular domains of TRPA1. To test the functional salience of these mechanisms in vitro and in vivo, we validated TRPA1 splicing isoforms using Xenopus oocyte electrophysiology and the GAL4/UAS system in D. melanogaster. Single TRPA1 isoform electrophysiology in vitro revealed evolution of the channel in the S. flava lineage towards reduced electrophile sensitivity. Ectopic expression of S. flava TRPA1 in D. melanogaster also consistently conferred weaker AITC sensitivity in vivo than expression of its orthologues, although this did not fully recapitulate differences in wild-type phenotypes between species, suggesting other molecular mechanisms were involved. To address this, we explored the consequences of isoform co-expression using oocyte electrophysiology. We found that enrichment of electrophile-insensitive TRPA1 splicing isoforms as observed in the salient S. flava sensory organs additively reduced cellular responses to AITC, which could further contribute to reduced electrophile aversion. Our findings illuminate how expression differences, protein structural changes, and especially alternative splicing, together can drive sensory evolution as animals behaviorally adapt to toxic new niches.

evolutionary biology↗

bioRxiv2022

Trichomes on female reproductive tract: rapid diversification and underlying gene regulatory network in Drosophila suzukii and its related species

BackgroundThe ovipositors of some insects are external female genitalia, which have their primary function to deliver eggs. Drosophila suzukii and its sibling species D. subpulchrella are known to have acquired highly sclerotized and enlarged ovipositors upon their shifts in oviposition sites from rotting to ripening fruits. Inside the ovipositor plates, there are scale-like polarized protrusions termed "oviprovector scales" that are likely to aid the mechanical movement of the eggs. The size and spatial distribution of the scales need to be rearranged following the divergence of the ovipositors. In this study, we examined the features of the oviprovector scales in D. suzukii and its closely related species. We also investigated whether the scales are single-cell protrusions comprised of F-actin under the same conserved gene regulatory network as the well-characterized trichomes on the larval cuticular surface. ResultsThe oviprovector scales of D. suzukii and D. subpulchrella were distinct in size and spatial arrangement compared to those of a closely related species D. biarmipes and D. melanogaster. The comparisons of the size of the scales suggested that the apical cell area of the oviprovector has expanded upon the elongation of the ovipositor plates in these species. Our transcriptome analysis revealed that 43 out of the 46 genes known to be involved in the trichome gene regulatory network are expressed in the developing female genitalia of D. suzukii and D. subpulchrella. An antibody staining depicted the presence of Shavenbaby (Svb) in the inner cavity of the developing ovipositors of D. melanogaster at 44-48 h after puparium formation (APF). Also, shavenoid (sha) was expressed in the corresponding patterns in the developing ovipositors and showed differential expression levels between D. suzukii and D. subpulchrella at 48 h APF. ConclusionsThe oviprovector scales have divergent size and spatial arrangements among species. Therefore, these scales may represent a rapidly diversifying morphological trait of the female reproductive tract reflecting ecological contexts. Furthermore, our results showed that the gene regulatory network underlying trichome formation is adopted to develop the rapidly evolving trichomes on the oviprovectors of these flies.

developmental biology↗

bioRxiv2021

A silencer repressing redundant enhancer activities revealed by deleting endogenous cis-regulatory element of ebony in Drosophila melanogaster

The spatiotemporal regulation of gene expression is essential to ensure robust phenotypic outcomes. Pigmentation patterns in Drosophila are formed by the deposition of different pigments synthesized in the developing epidermis and the role of cis-regulatory elements (CREs) of melanin biosynthesis pathway-related genes is well-characterized. These CREs typically exhibit modular arrangement in the regulatory region of the gene with each enhancer regulating a specific spatiotemporal expression of the gene. However, recent studies have suggested that multiple enhancers of a number of developmental genes as well as those of yellow (involved in dark pigment synthesis) exhibit redundant activities. Here we report the redundant enhancer activities in the cis-regulatory region of another gene in the melanin biosynthesis pathway, ebony, in the developing epidermis of Drosophila melanogaster. The evidence was obtained by introducing an approximately 1 kbp deletion at the endogenous primary epidermis enhancer (priEE) by genome editing. The effect of the priEE deletion on pigmentation and on the endogenous expression pattern of a mCherry-tagged ebony allele was examined in the thoracic and abdominal segments. The expression level of ebony in the priEE-deleted strains was similar to that of the control strain, indicating the presence of redundant enhancer activities that drive the broad expression of ebony in the developing epidermis. Additionally, the priEE fragment contained a silencer that suppresses ebony expression in the dorsal midline of the abdominal tergites, which is necessary for the development of the subgenus Sophophora-specific dark pigmentation patterns along the midline. The endogenous expression pattern of ebony in the priEE-deleted strains and the reporter assay examining the autonomous activity of the priEE fragment indicated that the silencer is involved in repressing the activities of both proximal and distant enhancers. These results suggest that multiple silencers are dispensable in the regulatory system of a relatively stable taxonomic character. The prevalence of other redundant enhancers and silencers in the genome can be investigated using a similar approach. Author summaryGenes are expressed at the right timing and place to give rise to diverse phenotypes. The spatiotemporal regulation is usually achieved through the coordinated activities of transcription-activating and transcription-repressing proteins that bind to the DNA sequences called enhancers and silencers, respectively, located near the target gene. Most studies identified the locations of enhancers by examining the ability of the sequence fragments to regulate the expression of fused reporters. Various short enhancers have been identified using this approach. This study employed an alternative approach in which the previously identified enhancer that regulates expression of ebony (a gene involved in body color formation) was deleted in a fruitfly, Drosophila melanogaster, using the genome-editing technique. The knockout of this enhancer did not affect the transcription level of the gene to a large extent. This indicated the presence of transcription-activating elements with redundant functions outside the deleted enhancer. Additionally, the transcription of ebony at the midline of the abdomen, which is repressed in the normal flies, were derepressed in the enhancer-deleted flies, which indicated that the deleted enhancer fragment contained a silencer that negatively regulates multiple enhancer activities in a spatially restricted manner.

genetics↗

bioRxiv2020

Drosophila suzukii avoidance of microbes in oviposition choice

While the majority of Drosophila species lay eggs onto fermented fruits, females of D. suzukii pierce the skin and lay eggs into ripening fruits using their serrated ovipositors. The changes of oviposition site preference must have accompanied this niche exploitation. In this study, we established an oviposition assay to investigate the effects of commensal microbes deposited by conspecific and heterospecific individuals, and showed that presence of microbes on the oviposition substrate enhances egg-laying of D. melanogaster and D. biarmipes, but discourages that of D. suzukii. This result suggests that a drastic change has taken place in the lineage leading to D. suzukii in how females respond to chemical cues produced by microbes. We also found that hardness of the substrate affects the response to microbial growth, indicating that mechanosensory stimuli interact with chemosensory invoked decisions to select or avoid oviposition sites.

evolutionary biology↗

bioRxiv2020

Unravelling the genetic basis for the rapid diversification of male genital structures between Drosophila species

In the last 240,000 years, males of the Drosophila simulans species clade have evolved striking differences in the morphology of their epandrial posterior lobes and claspers (surstyli). These changes have most likely been driven by sexual selection and mapping studies indicate a highly polygenic and generally additive genetic basis. However, we have limited understanding of the gene regulatory networks that control the development of genital structures and how they evolved to result in this rapid phenotypic diversification. Here, we used new D. simulans / D. mauritiana introgression lines on chromosome 3L to generate higher resolution maps of posterior lobe and clasper differences between these species. We then carried out RNA-seq on the developing genitalia of both species to identify the genes expressed during this process and those that are differentially expressed between the two species. This allowed us to test the function of expressed positional candidates during genital development in D. melanogaster. We identified several new genes involved in the development and possibly the evolution of these genital structures, including the transcription factors Hairy and Grunge. Furthermore, we discovered that during clasper development Hairy negatively regulates tartan, a gene known to contribute to divergence in clasper morphology. Taken together our results provide new insights into the regulation of genital development and how this evolves between species.

evolutionary biology↗

bioRxiv2018

tartan underlies the evolution of male genital morphology

Male genital structures are among the most rapidly evolving morphological traits and are often the only features that can distinguish closely related species. This process is thought to be driven by sexual selection and may reinforce species separation. However, while the genetic basis of many phenotypic differences have been identified, we still lack knowledge about the genes underlying evolutionary differences in male genital organs and organ size more generally. The claspers (surstyli) are periphallic structures that play an important role in copulation in insects. Here we show that natural variation in clasper size and bristle number between Drosophila mauritiana and D. simulans is caused by evolutionary changes in tartan (trn), which encodes a transmembrane leucine-rich repeat domain protein that mediates cell-cell interactions and affinity differences. There are no fixed amino acid differences in trn between D. mauritiana and D. simulans but differences in the expression of this gene in developing genitalia suggest cis-regulatory changes in trn underlie the evolution of clasper morphology in these species. Finally, analysis of reciprocal hemizyotes that are genetically identical, except for which species the functional allele of trn is from, determined that the trn allele of D. mauritiana specifies larger claspers with more bristles than the allele of D. simulans. Therefore we have identified the first gene underlying evolutionary change in the size of a male genital organ, which will help to better understand the rapid diversification of these structures and the regulation and evolution of organ size more broadly.\n\nSignificance StatementThe morphology of male genital organs evolves rapidly driven by sexual selection. However, little is known about the genes underlying genitalia differences between species. Identifying these genes is key to understanding how sexual selection acts on development to produce rapid phenotypic change. We have found that the gene tartan underlies differences between male Drosophila mauritiana and D. simulans in the size and bristle number of the claspers - genital projections that grasp the female during copulation. Moreover, since tartan encodes a protein that is involved in cell affinity, this may represent a new developmental mechanism for morphological change. Therefore, our study provides new insights into genetic and developmental bases for the rapid evolution of male genitalia and organ size more generally.

evolutionary biology↗