Cloning of a Chironomus tentans cDNA encoding a protein (cEcRH) homologous to the Drosophila melanogaster ecdysteroid receptor (dEcR)

Imhof, Markus O.; Rusconi, Sandro; Lezzi, Markus

doi:10.1016/0965-1748(93)90089-b

Cited by 118 publications

(51 citation statements)

References 40 publications

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“…The splicing sites that generated the EcR isoforms were conserved among these three insects. Some similarity was also observed with the EcR of Chironomus tentans (Imhof et al, 1993).…”

Section: Resultsmentioning

confidence: 51%

Tissue‐Specific and Stage‐Specific Expression of Two Silkworm Ecdysone Receptor Isoforms

Kamimura

Tomita

Kiuchi

et al. 1997

European Journal of Biochemistry

105

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Previously, we isolated a cDNA clone for the ecdysone receptor B1 isoform of the silkworm, Bombyx mori (BmEcR-Bl). Here we report the cloning of a cDNA that encodes the Bombyx ecdysone receptor A isoform (BmEcR-A) and mRNA expression of the two BmEcR isoforms during molting and metamorphosis. At larval-pupal transformation, mRNA expression of BmEcR-B 1 was predominant in most tissues examined, including three larval tissues (midgut, epidermis, and fat body) and the wing imaginal disc. The anterior silk gland was the only tissue where BmEcR-A was predominant. These expression patterns were different from observations demonstrated in Drosophila. In the anterior silk gland, both EcR isoforms were expressed synchronously during the fifth larval instar, while expression of the A isoform preceded that of the B1 isoform by two days in the fourth instar. Precedence of BmEcR-A during the fourth instar and synchronization of both isoforms during the fifth instar were also observed in the middle and posterior silk glands, suggesting that transcription of BmEcR in the silk gland is regulated differently in these two instars. In the cultured anterior silk glands of day 0 of the fifth instar, transcription of BmEcR-A and BmEcR-B1 was induced dose dependently by more than 5 ng/ml 20-hydroxyecdysone. BmEcR-A and BmEcR-B1 mRNAs were induced within 2 h and 1 h, respectively, of the addition of 20-hydroxyecdysone. These results suggest that the increase of BmEcR mRNAs during the fifth instar is induced in vivo by a small increase in ecdysteroids.Keywords: ecdysone receptor; Bombyx mori; ecdysteroid ; silk gland ; isoform.Steroid hormones coordinate a wide array of developmental and physiological processes in higher organisms, through binding with receptor proteins to regulate the stage-specific and tissue-specific transcription of target genes. In insects, ecdysteroids, particularly 20-hydroxyecdysone, are the key steroid hormones. 20-Hydroxyecdysone plays a central role in the orchestration of development during molts and metamorphosis. Ashburner et al. (1974) proposed a model for 20-hydroxyecdysone action based on puffing responses of the polytene chromosomes in the Drosophila salivary gland. In this model, 20-hydroxyecdysone binds with an ecdysone receptor protein and directly activates the transcription of a small set of early genes, which then activate many late genes. Recent molecular analyses characterized the ecdysone receptor gene and some of the early genes, most of which are putative transcriptional factors as predicted by Ashburner's model (reviewed in Henrich and Brown, 1995;Thummel, 1995). The ecdysone receptor (EcR) identified from Drosophila is a member of the nuclear receptor superfamily and has three isoforms (A, B1, and B2) with common DNA and

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“…The splicing sites that generated the EcR isoforms were conserved among these three insects. Some similarity was also observed with the EcR of Chironomus tentans (Imhof et al, 1993).…”

Section: Resultsmentioning

confidence: 51%

Tissue‐Specific and Stage‐Specific Expression of Two Silkworm Ecdysone Receptor Isoforms

Kamimura

Tomita

Kiuchi

et al. 1997

European Journal of Biochemistry

105

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“…DmEcR, CfEcR, BaEcR, and BmEcR are underlined as their proteins contain a Tyr at the critical ligand specificity site. Data bank search yielded EcR protein sequences from 16 species: 6 Diptera species, the Mediterranean fruit fly Ceratitis capitata (CcEcR [67]), the sheep blowfly Lucilia cuprina (LcEcR [23]), the yellow fever mosquitoes A. aegypti (AaEcR, 12) and A. albopictus (not shown, as its EcR I box is 100% identical to AaEcR [28]), the midge Chironomus tentans (CtEcR [27]), and D. melanogaster (DmEcR [34]); 6 Lepidoptera species, the spruce budworm C. fumiferana (CfEcR [35]), squinting bush brown Bicyclus anynana (BaEcR [R. K. Reinhardt, P. Weber, and P. B. Koch, submitted to GenBank, accession no. CAB63236]), the silkworm B. mori (BmEcR [32,60]), the tobacco budworm Heliothis virescens (HvEcR [41]), the tobacco hornworm M. sexta (MsEcR [18]), and the buckeye Junonia coenia (JcEcR [R. K. Reinhardt, P. Weber, and P. B. Koch submitted to GenBank, accession no.…”

Section: Discussionmentioning

confidence: 99%

“…The molecular basis of 20E action has been elucidated in great detail (10,24,26,46,51,56,64,65). The two nuclear hormone receptors that play a central role in the initiation of the 20E-induced gene regulatory hierarchy are the ecdysteroid receptor (EcR) and the retinoid X receptor (RXR) homologue Ultraspiracle (USP) (12,18,25,27,34,43,57,60). It has been shown that USP is an obligatory heterodimeric partner of EcR, required for both ligand and DNA binding (63,74,75).…”

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confidence: 99%

Molecular Determinants of Differential Ligand Sensitivities of Insect Ecdysteroid Receptors

Wang

Ayer

Segraves

et al. 2000

Molecular and Cellular Biology

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The functional receptor for insect ecdysteroid hormones is a heterodimer consisting of two nuclear hormone receptors, ecdysteroid receptor (EcR) and the retinoid X receptor homologue Ultraspiracle (USP). Although ecdysone is commonly thought to be a hormone precursor and 20-hydroxyecdysone (20E), the physiologically active steroid, little is known about the relative activity of ecdysteroids in various arthropods. As a step toward characterization of potential differential ligand recognition, we have analyzed the activities of various ecdysteroids using gel mobility shift assays and transfection assays in Schneider-2 (S2) cells. Ecdysone showed little activation of the Drosophila melanogaster receptor complex (DmEcR-USP). In contrast, this steroid functioned as a potent ligand for the mosquito Aedes aegypti receptor complex (AaEcR-USP), significantly enhancing DNA binding and transactivating a reporter gene in S2 cells. The mosquito receptor also displayed higher hormoneindependent DNA binding activity than the Drosophila receptor. Subunit-swapping experiments indicated that the EcR protein, not the USP protein, was responsible for ligand specificity. Using domain-swapping techniques, we made a series of Aedes and Drosophila EcR chimeric constructs. Differential ligand responsiveness was mapped near the C terminus of the ligand binding domain, within the identity box previously implicated in the dimerization specificity of nuclear receptors. This region includes helices 9 and 10, as determined by comparison with available crystal structures obtained from other nuclear receptors. Site-directed mutagenesis revealed that Phe529 in Aedes EcR, corresponding to Tyr611 in Drosophila EcR, was most critical for ligand specificity and hormone-independent DNA binding activity. These results demonstrated that ecdysone could function as a bona fide ligand in a species-specific manner.

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“…In C. tentans, EcR mRNA abundance parallels changes in ecdysteroid titer (78). In this species, the EcR mRNA has been observed to be relatively uncommon during diapause (68).…”

Section: Ecdysone Receptor (Ecr)mentioning

confidence: 89%

Differential Gene Expression in Insects: Transcriptional Control

Harshman¹,

James

1998

Annu. Rev. Entomol.

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Studies on transcriptional control of gene expression play a pivotal role in many areas of biology. In non-Drosophilid insects, the cuticle, chorion, immune response, silk gland, storage proteins, and vitellogenin are foci for advances in basic research on promoter elements and transcription factors. Insects offer other advantages for gene regulation studies, including the availability of applied problems. In non-Drosophilid insects, the most serious problem for transcriptional control studies is the lack of homologous in vivo expression systems. Once this deficiency is addressed, the full impact of research on transcription control will be realized throughout the field of entomology.

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Cloning of a Chironomus tentans cDNA encoding a protein (cEcRH) homologous to the Drosophila melanogaster ecdysteroid receptor (dEcR)

Cited by 118 publications

References 40 publications

Tissue‐Specific and Stage‐Specific Expression of Two Silkworm Ecdysone Receptor Isoforms

Tissue‐Specific and Stage‐Specific Expression of Two Silkworm Ecdysone Receptor Isoforms

Molecular Determinants of Differential Ligand Sensitivities of Insect Ecdysteroid Receptors

Differential Gene Expression in Insects: Transcriptional Control

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