Publications by authors named "Shuichiro Tomita"

16 Publications

  • Page 1 of 1

Multicomponent structures in camouflage and mimicry in butterfly wing patterns.

J Morphol 2019 01;280(1):149-166

Transgenic Silkworm Research Unit, Division of Biotechnology, Institute of Agrobiological Sciences, NARO, Ibaraki, Japan.

Understanding how morphological structures are built is essential for appreciating the morphological complexity and divergence of organisms. One representative case of morphological structures is the camouflage and mimicry of butterfly wing patterns. Some previous studies have questioned whether camouflage and mimicry are truly structures, considering that they rely on coloration. Nevertheless, our recent study revealed that the leaf pattern of Kallima inachus butterfly wings evolved through the combination of changes in several pigment components in a block-wise manner; it remains unclear whether such block-wise structures are common in other cases of camouflage and mimicry in butterflies and how they come about. Previous studies focused solely on a set of homologous components, termed the nymphalid ground plan. In the present study, we extended the scope of the description of components by including not only the nymphalid ground plan but also other common components (i.e., ripple patterns, dependent patterns, and color fields). This extension allowed us to analyze the combinatorial building logic of structures and examine multicomponent structures of camouflage and mimicry in butterfly wing patterns. We investigated various patterns of camouflage and mimicry (e.g., masquerade, crypsis, Müllerian mimicry, Batesian mimicry) in nine species and decomposed them into an assembly of multiple components. These structural component analyses suggested that camouflage and mimicry in butterfly wing patterns are built up by combining multiple types of components. We also investigated associations between components and the kinds of camouflage and mimicry. Several components are statistically more often used to produce specific types of camouflage or mimicry. Thus, our work provides empirical evidence that camouflage and mimicry patterns of butterfly wings are mosaic structures, opening up a new avenue of studying camouflage, and mimicry from a structural perspective.
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http://dx.doi.org/10.1002/jmor.20927DOI Listing
January 2019

A Hox Gene, Antennapedia, Regulates Expression of Multiple Major Silk Protein Genes in the Silkworm Bombyx mori.

J Biol Chem 2016 Mar 26;291(13):7087-96. Epub 2016 Jan 26.

From the Transgenic Silkworm Research Unit, National Institute of Agrobiological Sciences, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, Japan,

Hoxgenes play a pivotal role in the determination of anteroposterior axis specificity during bilaterian animal development. They do so by acting as a master control and regulating the expression of genes important for development. Recently, however, we showed that Hoxgenes can also function in terminally differentiated tissue of the lepidopteranBombyx mori In this species,Antennapedia(Antp) regulates expression of sericin-1, a major silk protein gene, in the silk gland. Here, we investigated whether Antpcan regulate expression of multiple genes in this tissue. By means of proteomic, RT-PCR, and in situ hybridization analyses, we demonstrate that misexpression of Antpin the posterior silk gland induced ectopic expression of major silk protein genes such assericin-3,fhxh4, and fhxh5 These genes are normally expressed specifically in the middle silk gland as is Antp Therefore, the evidence strongly suggests that Antpactivates these silk protein genes in the middle silk gland. The putativesericin-1 activator complex (middle silk gland-intermolt-specific complex) can bind to the upstream regions of these genes, suggesting that Antpdirectly activates their expression. We also found that the pattern of gene expression was well conserved between B. moriand the wild species Bombyx mandarina, indicating that the gene regulation mechanism identified here is an evolutionarily conserved mechanism and not an artifact of the domestication of B. mori We suggest that Hoxgenes have a role as a master control in terminally differentiated tissues, possibly acting as a primary regulator for a range of physiological processes.
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http://dx.doi.org/10.1074/jbc.M115.699819DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4807290PMC
March 2016

Gradual and contingent evolutionary emergence of leaf mimicry in butterfly wing patterns.

BMC Evol Biol 2014 Nov 25;14:229. Epub 2014 Nov 25.

Transgenic Silkworm Research Unit, Genetically Modified Organism Research Center, National Institute of Agrobiological Sciences, 1-2 Oowashi, 305-8634, Tsukuba, Ibaraki, Japan.

Background: Special resemblance of animals to natural objects such as leaves provides a representative example of evolutionary adaptation. The existence of such sophisticated features challenges our understanding of how complex adaptive phenotypes evolved. Leaf mimicry typically consists of several pattern elements, the spatial arrangement of which generates the leaf venation-like appearance. However, the process by which leaf patterns evolved remains unclear.

Results: In this study we show the evolutionary origin and process for the leaf pattern in Kallima (Nymphalidae) butterflies. Using comparative morphological analyses, we reveal that the wing patterns of Kallima and 45 closely related species share the same ground plan, suggesting that the pattern elements of leaf mimicry have been inherited across species with lineage-specific changes of their character states. On the basis of these analyses, phylogenetic comparative methods estimated past states of the pattern elements and enabled reconstruction of the wing patterns of the most recent common ancestor. This analysis shows that the leaf pattern has evolved through several intermediate patterns. Further, we use Bayesian statistical methods to estimate the temporal order of character-state changes in the pattern elements by which leaf mimesis evolved, and show that the pattern elements changed their spatial arrangement (e.g., from a curved line to a straight line) in a stepwise manner and finally establish a close resemblance to a leaf venation-like appearance.

Conclusions: Our study provides the first evidence for stepwise and contingent evolution of leaf mimicry.  Leaf mimicry patterns evolved in a gradual, rather than a sudden, manner from a non-mimetic ancestor. Through a lineage of Kallima butterflies, the leaf patterns evolutionarily originated through temporal accumulation of orchestrated changes in multiple pattern elements.
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http://dx.doi.org/10.1186/s12862-014-0229-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4261531PMC
November 2014

Soaking RNAi in Bombyx mori BmN4-SID1 cells arrests cell cycle progression.

J Insect Sci 2013 ;13:155

1 Laboratory of Silkworm Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, Hakozaki 6-10-1, Fukuoka 812-8581.

RNA interference (RNAi) is an evolutionarily conserved mechanism for sequence-specific gene silencing. Previously, the BmN4-SID1 cell expressing Caenorhabditis ele gans SID-1 was established, in which soaking RNAi could induce effective gene silencing. To establish its utility, 6 cell cycle progression related cDNAs, CDK1, MYC, MYB, RNRS, CDT1, and GEMININ, were isolated from the silkworm, Bombyx mori L. (Lepidoptera: Bombycidae), and their expressions were further silenced by soaking RNAi in the BmN4-SID1 cells. The cell cycle progression analysis using flow cytometer demonstrated that the small amount of double stranded RNA was enough to arrest cell cycle progression at the specific cell phases. These data suggest that RNAi in the BmN4-SID1 cells can be used as a powerful tool for loss-of-function analysis of B. mori genes.
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http://dx.doi.org/10.1673/031.013.15501DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4015410PMC
January 2015

Novel gene rearrangements in the mitochondrial genome of a webspinner, Aposthonia japonica (Insecta: Embioptera).

Genome 2012 Mar 29;55(3):222-33. Epub 2012 Feb 29.

National Institute of Agrobiological Sciences, Tsukuba, Ibaraki 305-8634, Japan.

Webspinners (order Embioptera) are polyneopteran insects characterized by enlarged foretarsi with silk glands, whose silk is used to produce galleries in which the insects live gregariously. The phylogenetic position of webspinners has been debated. In the present study, an almost complete mitochondrial DNA (mtDNA) sequence of Embioptera is reported for the first time. The mtDNA of a webspinner, Aposthonia japonica , has the 13 protein-coding genes (PCGs) generally found in metazoan mtDNA sequences. There is a translocation of a large region including atp6, atp8, cox3, nad3, and nad5 as well as a duplication of the 12S rRNA gene. The rearrangement does not seem to affect nucleotide composition, although amino acid composition in some parts of the mtDNA is biased compared with other Polyneoptera species. Based on phylogenetic analyses using nucleotide sequences of all PCGs concatenated with two rRNA genes and the amino acid sequences of all PCGs, A. japonica is sister to Verophasmatodea, a suborder of typical stick and leaf insects.
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http://dx.doi.org/10.1139/g2012-007DOI Listing
March 2012

Effective RNA interference in cultured silkworm cells mediated by overexpression of Caenorhabditis elegans SID-1.

RNA Biol 2012 Jan 1;9(1):40-6. Epub 2012 Jan 1.

Laboratory of Silkworm Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, Hakozaki, Fukuoka, Japan.

RNA interference (RNAi) is a conserved mechanism that catalyzes sequence-specific gene silencing and has been used for loss-of-function genetic screens in many organisms. Here, we demonstrated that the expression of Caenorhabditis elegans SID-1 (CeSID-1) could trigger effective gene silencing in the cultured silkworm cell line, BmN4 (BmN4-SID1). Soaking the BmN4-SID1 in dsRNA corresponding to endogenous target genes induced a significant decrease of the amount of mRNA or protein. A small amount of dsRNA was enough to silence the target gene in a few days. Overexpression of CeSID-1 did not affect the cell viability. Our results suggest that BmN4-SID1 can be used in many applications in silkworm cells and will become a valuable resource for gene analysis.
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http://dx.doi.org/10.4161/rna.9.1.18084DOI Listing
January 2012

SID-1 protein of Caenorhabditis elegans mediates uptake of dsRNA into Bombyx cells.

Insect Biochem Mol Biol 2012 Feb 8;42(2):148-54. Epub 2011 Dec 8.

Transgenic Silkworm Research Unit, National Institute of Agrobiological Sciences, 1-2 Ohwashi, Tsukuba, Ibaraki 305-8634, Japan.

RNA interference is one of the most revolutionary tools in the study of gene function, particularly in non-model systems. However, in Bombyx mori, as with many lepidopteran species, attempts at systemic RNAi have had mixed success. Gene identification and phylogenetic analyses suggest that Bombyx has the core RNAi machinery, which is necessary to undergo RNAi as a cellular response. We introduced sid genes from Caenorhabditis elegans into Bombyx BmN4 cells to enhance the uptake of dsRNA and revealed that the SID-1 protein, but not SID-2, has the ability to endow the RNAi effect with the addition of dsRNA to the medium. Observed RNAi effect was dependent on both the levels of sid-1 expression and the concentration of the dsRNA. These results suggest that SID-1 promotes the uptake of dsRNA from the medium into Bombyx cells. We generated transgenic animals that express sid-1 but have not detected significant enhancements of in vivo phenotype in response to the injection of the dsRNA into hemocoel.
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http://dx.doi.org/10.1016/j.ibmb.2011.11.007DOI Listing
February 2012

RNA interference in Lepidoptera: an overview of successful and unsuccessful studies and implications for experimental design.

J Insect Physiol 2011 Feb 20;57(2):231-45. Epub 2010 Nov 20.

Department of Ecology, Swedish University of Agricultural Sciences, Uppsala, Sweden.

Gene silencing through RNA interference (RNAi) has revolutionized the study of gene function, particularly in non-model insects. However, in Lepidoptera (moths and butterflies) RNAi has many times proven to be difficult to achieve. Most of the negative results have been anecdotal and the positive experiments have not been collected in such a way that they are possible to analyze. In this review, we have collected detailed data from more than 150 experiments including all to date published and many unpublished experiments. Despite a large variation in the data, trends that are found are that RNAi is particularly successful in the family Saturniidae and in genes involved in immunity. On the contrary, gene expression in epidermal tissues seems to be most difficult to silence. In addition, gene silencing by feeding dsRNA requires high concentrations for success. Possible causes for the variability of success in RNAi experiments in Lepidoptera are discussed. The review also points to a need to further investigate the mechanism of RNAi in lepidopteran insects and its possible connection to the innate immune response. Our general understanding of RNAi in Lepidoptera will be further aided in the future as our public database at http://insectacentral.org/RNAi will continue to gather information on RNAi experiments.
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http://dx.doi.org/10.1016/j.jinsphys.2010.11.006DOI Listing
February 2011

Exploring the molecular phylogeny of phasmids with whole mitochondrial genome sequences.

Mol Phylogenet Evol 2011 Jan 26;58(1):43-52. Epub 2010 Oct 26.

Division of Insect Science, National Institute of Agrobiological Sciences, 1-2 Ohwashi, Tsukuba, Ibaraki, Japan.

Phasmids are remarkable mimics of twigs, sticks, and leaves. This extreme adaptation for crypsis can easily lead to the convergent evolution of morphology, making it difficult to establish a taxonomic system of phasmids. Accordingly, there are multiple phylogenetic hypotheses that conflict with each other. Phylogenetic arrangements suggested by molecular data disagree with the morphology-based taxonomy in some instances. We collected 13 phasmatodean species, sequenced their mitochondrial genomes, and recovered their molecular phylogeny. Our analyses did not support the monophyly of Areolatae or Anareolatae, two major infraorders of Phasmatodea. The position of Neohirasea was also quite different from the conventional taxonomic systems, thus challenging the previously assumed monophyly of the subfamily Lonchodinae. The enigmatic taxon, Timema, was shown to be distantly related to other phasmatodeans.
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http://dx.doi.org/10.1016/j.ympev.2010.10.013DOI Listing
January 2011

Abd-B suppresses lepidopteran proleg development in posterior abdomen.

Dev Biol 2009 Apr 6;328(2):403-9. Epub 2009 Feb 6.

Insect Genome Research Unit, National Institute of Agrobiological Sciences, 1-2 Ohwashi, Tsukuba, Ibaraki 305-8634, Japan.

Pterygotes lack abdominal appendages except for pleuropods and prolegs. The larvae of some holometabolous insects develop prolegs, which are used for locomotion. We analyzed the role of the homeotic genes abd-A and Abd-B in lepidopteran proleg development using mutant analysis and embryonic RNAi in the silkworm Bombyx mori. The E(Mu) mutant developed extra prolegs in its posterior abdomen and showed the misexpression of both genes, suggesting their involvement in proleg formation. The depletion of Abd-B by embryonic RNAi caused the development of extra prolegs on all segments posterior to A6, indicating the suppressive function of Abd-B. The abd-A RNAi animals failed to develop prolegs. These results indicate that abd-A and Abd-B are involved in proleg development in B. mori.
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http://dx.doi.org/10.1016/j.ydbio.2009.01.040DOI Listing
April 2009

Exploring systemic RNA interference in insects: a genome-wide survey for RNAi genes in Tribolium.

Genome Biol 2008 Jan 17;9(1):R10. Epub 2008 Jan 17.

Division of Biology, Kansas State University, Manhattan, Kansas 66506, USA.

Background: RNA interference (RNAi) is a highly conserved cellular mechanism. In some organisms, such as Caenorhabditis elegans, the RNAi response can be transmitted systemically. Some insects also exhibit a systemic RNAi response. However, Drosophila, the leading insect model organism, does not show a robust systemic RNAi response, necessitating another model system to study the molecular mechanism of systemic RNAi in insects.

Results: We used Tribolium, which exhibits robust systemic RNAi, as an alternative model system. We have identified the core RNAi genes, as well as genes potentially involved in systemic RNAi, from the Tribolium genome. Both phylogenetic and functional analyses suggest that Tribolium has a somewhat larger inventory of core component genes than Drosophila, perhaps allowing a more sensitive response to double-stranded RNA (dsRNA). We also identified three Tribolium homologs of C. elegans sid-1, which encodes a possible dsRNA channel. However, detailed sequence analysis has revealed that these Tribolium homologs share more identity with another C. elegans gene, tag-130. We analyzed tag-130 mutants, and found that this gene does not have a function in systemic RNAi in C. elegans. Likewise, the Tribolium sid-like genes do not seem to be required for systemic RNAi. These results suggest that insect sid-1-like genes have a different function than dsRNA uptake. Moreover, Tribolium lacks homologs of several genes important for RNAi in C. elegans.

Conclusion: Although both Tribolium and C. elegans show a robust systemic RNAi response, our genome-wide survey reveals significant differences between the RNAi mechanisms of these organisms. Thus, insects may use an alternative mechanism for the systemic RNAi response. Understanding this process would assist with rendering other insects amenable to systemic RNAi, and may influence pest control approaches.
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http://dx.doi.org/10.1186/gb-2008-9-1-r10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2395250PMC
January 2008

Isoform specific control of gene activity in vivo by the Drosophila ecdysone receptor.

Mech Dev 2003 Aug;120(8):909-18

Department of Biology, University of Washington, Seattle WA 98195, USA.

The steroid hormone 20-hydroxyecdysone induces metamorphosis in insects. The receptor for the hormone is the ecdysone receptor, a heterodimer of two nuclear receptors, EcR and USP. In Drosophila the EcR gene encodes 3 isoforms (EcR-A, EcR-B1 and EcR-B2) that vary in their N-terminal region but not in their DNA binding and ligand binding domains. The stage and tissue specific distribution of the isoforms during metamorphosis suggests distinct functions for the different isoforms. By over-expressing the three isoforms in animals we present results supporting this hypothesis. We tested for the ability of the different isoforms to rescue the lack of dendritic pruning that is characteristic of mutants lacking both EcR-B1 and EcR-B2. By expressing the different isoforms specifically in the affected neurons, we found that both EcR-B isoforms were able to rescue the neuronal defect cell autonomously, but that EcR-A was less effective. We also analyzed the effect of over-expressing the isoforms in a wild-type background. We determined a sensitive period when high levels of either EcR-B isoform were lethal, indicating that the low levels of EcR-B at this time are crucial to ensure normal development. Over-expressing EcR-A in contrast had no detrimental effect. However, high levels of EcR-A expressed in the posterior compartment suppressed puparial tanning, and resulted in down-regulation of some of the tested target genes in the posterior compartment of the wing disc. EcR-B1 or EcR-B2 over-expression had little or no effect.
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http://dx.doi.org/10.1016/s0925-4773(03)00134-5DOI Listing
August 2003

Transient in vivo reporter gene assay for ecdysteroid action in the Bombyx mori silk gland.

Comp Biochem Physiol B Biochem Mol Biol 2003 Jul;135(3):431-7

Developmental Biology Department, National Institute of Agrobiological Sciences, 1-2, Owashi, Tsukuba, Ibaraki 305-8634, Japan.

To analyze the molecular mechanisms underlying hormone-regulated gene expression during molt and metamorphosis, we developed a transient reporter gene assay system using the silkworm anterior silk gland. Reporter plasmids were delivered into dissected anterior silk glands by particle bombardment and bombarded glands transplanted into other larvae, to which hormones were then administered. When the green fluorescent protein gene, coupled with the constitutive cytoplasmic actin gene A3 promoter, was introduced into the anterior silk gland, strong green fluorescence was observed a few days later. Bombarded silk glands transplanted into other larvae showed the same morphological changes as intrinsic glands after 20-hydroxyecdysone (20E) alone or 20E plus juvenile hormone (JH) treatment, indicating that the transplanted gland received hormonal signals properly. When a 20E-responsive reporter construct containing four tandemly repeated pal-1 ecdysone response elements upstream from the luciferase gene was delivered into the gland, an approximately 50-fold increase in luciferase activity was detected 30 h after 20E injection. This induction was comparable to that in an ecdysteroid-responsive Bombyx cell line. This in vivo reporter assay system is thus a rapid, effective tool for analyzing gene expression regulated by 20E and probably by JH.
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http://dx.doi.org/10.1016/s1096-4959(03)00094-0DOI Listing
July 2003

Molecular cloning and expression analysis of ultraspiracle (USP) from the rice stem borer Chilo suppressalis.

Insect Biochem Mol Biol 2003 Jan;33(1):41-9

Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, 606-8502, Kyoto, Japan.

cDNA for ultraspiracle (USP) from the lepidopteran rice stem borer Chilo suppressalis was cloned using PCR techniques. The deduced amino acid sequence of C. suppressalis USP (CsUSP) was very similar to those of other lepidopteran USPs, especially to the Manduca sexta USP-2 isoform. Northern hybridization analysis detected a 6.5-kb message in the epidermis, fat body, and midgut of wandering larvae. CsUSP mRNA expression in the epidermis varied little during the last larval instar. Gel mobility shift assays showed that in vitro translated C. suppressalis ecdysone receptor (CsEcR) and CsUSP proteins bound to the Pal1 or Drosophila melanogaster hsp27 ecdysone response element as a heterodimer. In a ligand-receptor binding assay, [(3)H]ponasterone A ([(3)H]PoA) did not bind to individual CsEcR or CsUSP protein, but bound strongly to the CsEcR/CsUSP complex. [(3)H]PoA binding to CsEcR/CsUSP complex was competed by 20-hydroxyecdysone and a non-steroidal ecdysteroid agonist, RH-5992, but not by cholesterol, indicating that compounds with molting hormone activity against C. suppressalis can bind specifically to the CsEcR/CsUSP complex.
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http://dx.doi.org/10.1016/s0965-1748(02)00165-0DOI Listing
January 2003

A possible role of 20-hydroxyecdysone in embryonic development of the silkworm Bombyx mori.

Arch Insect Biochem Physiol 2002 Nov;51(3):111-20

Department of Life and Functional Material Science, Graduate School of Natural Sciences, Konan University, Kobe, Japan.

It has been well established that eggs of insects, including those of the silkworm Bombyx mori, contain various ecdysteroids and the amounts of these ecdysteroids fluctuate during embryonic development. In order to know the function of egg ecdysteroids in embryonic development of B. mori, we examined the biological activities of various egg ecdysteroids by in vitro ligand-binding assay and bioassay using B. mori eggs. First, using the ecdysteroid receptor of B. mori (BmEcR-B1/BmUSP heterodimer) prepared by yeast and Escherichia coli expression systems, the interaction between the ecdysteroid receptor and various egg ecdysteroids of B. mori was analyzed. The relative binding affinities of egg ecdysteroids to the BmEcR-B1/BmUSP heterodimer decreased in the order of 20-hydroxyecdysone > 2-deoxy-20-hydroxyecdysone > 22-deoxy-20-hydroxyecdysone > ecdysone > 2-deoxyecdysone > ecdysone 22-phosphate. Next, several egg ecdysteroids of B. mori were injected into the prospective diapause eggs, which show a very low level of free ecdysteroids at the onset of embryonic diapause (gastrula stage). Approximately 7% of them (P < 0.002, chi(2)-test) developed beyond the gastrula stage without entering diapause by the injection of 20-hydroxyecdysone (25 ng/egg). In contrast, the injection of other ecdysteroids was not effective in inducing embryonic development. These results suggest that 20-hydroxyecdysone, via the ecdysteroid receptor, is responsible for the developmental difference between diapause and non-diapause in B. mori embryos. Furthermore, it was suggested that continuous supply of 20-hydroxyecdysone may be required to induce embryonic development.
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http://dx.doi.org/10.1002/arch.10055DOI Listing
November 2002

Bombyx mori orphan receptor, BmHR78: cDNA cloning, testis abundant expression and putative dimerization partner for Bombyx ultraspiracle.

Mol Cell Endocrinol 2002 Mar;189(1-2):201-11

Department of Development and Differentiation, National Institute of Agrobiological Science, 1-2 Owashi, Tsukuba, 305-8634, Ibaraki, Japan.

We have identified a novel member of the nuclear receptor superfamily from the silkworm Bombyx mori, and named it as BmHR78, the B. mori hormone receptor. The DNA binding domain of BmHR78 shows high similarities to those of Tenebrio molitor hormone receptor 78, Drosophila hormone receptor 78, and mammalian testicular receptor 2, whereas the ligand binding domain is not well conserved. Northern blot analysis showed that BmHR78 gene was most abundantly expressed in the testis. From the fourth to fifth instar, BmHR78 gene was constantly expressed in the testis. In the anterior silk gland, the level of BmHR78 gene expression was developmentally changed. From day 10.0 to 11.0 in the fifth instar, another BmHR78 transcript with the smaller size appeared. Ultraspiracle (USP) isoform also appeared at the same stages in this tissue. BmHR78 forms not only a homodimer, but also a heterodimer with USP in a yeast two hybrid assay. The direct interaction between BmHR78 and USP was confirmed by pull down assay. Deletion mutant analysis showed that BmHR78 interacts with USP via the ninth heptad repeat in helix ten of the E region. This repeat is well conserved in RXR and its heterodimer partners, and shown to be an interface for their dimerization. In insect, only the ecdysone receptor and hormone receptor 38 are known thus far to dimerize with USP. Thus, BmHR78 is a third dimerization partner for USP and may modulate the molecular action of USP, including the ecdysone signal cascades.
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http://dx.doi.org/10.1016/s0303-7207(01)00604-9DOI Listing
March 2002
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