Isolation, characterization and developmental expression of the ecdysteroid-induced E75 gene of the wax moth Galleria mellonella.
Using the cDNA for the Drosophila ecdysteroid-induced member of the steroid-hormone-receptor superfamily, E75A, we isolated a genomic clone from Galleria mellonella that revealed 77% similarity with the region of E75A cDNA encoding the C-terminal zinc-finger motif. A Galleria cDNA clone was isolated that encoded a complete DNA-binding domain composed of two zinc fingers and designated GmE75A. Its deduced amino acid sequence showed 100% and 85% identities within the DNA-binding and ligand-binding domains of Drosophila E75A, respectively. The Galleria genomic clone did not encode the N-terminal zinc finger, but included a sequence similar to the B1 exon, which is unique to the B isoform of E75. Thus, the cDNA and genomic DNA sequences indicated that the Galleria gene E75 encoded at least two isoforms, GmE75A and GmE75B, which differed in their N-termini. Probes specific for GmE75A and B hybridized to two distinct transcripts of 2.6 kb. Both GmE75A and B mRNA levels correlated closely with the ecdysteroid titer during development. At the onset of larval/pupal transformation, both transcripts appeared in high amounts within 4 h of the ecdysteroid rise, then declined concurrently with the hormone titer decline. At the time of pupal ecdysis, there was another peak of GmE75A expression but not GmE75B expression, coincident with a minor ecdysteroid pulse. In isolated abdomens of final instar larvae, GmE75A mRNA was induced by 20-hydroxyecdysone within 20 min of the injection; the mRNA levels were maximal at 1 h and declined by 3 h following the treatment.
Jindra
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M
Institute of Entomology, Czech Academy of Sciences, Ceské Budĕjovice.
Sehnal
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F
Riddiford
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LM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D004330:Drosophila / Q000235:genetics; D026461:Ecdysteroids; D004441:Ecdysterone / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D017344:Genes, Insect; D006458:Hemolymph / Q000737:chemistry*; D007301:Insect Hormones / Q000097:blood; D019476:Insect Proteins; D007447:Invertebrate Hormones / Q000097:blood*; D008675:Metamorphosis, Biological; D008969:Molecular Sequence Data; D009036:Moths / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D011987:Receptors, Steroid / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D013256:Steroids / Q000097:blood*
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; C087173:E75 protein, insect; D026461:Ecdysteroids; D007301:Insect Hormones; D019476:Insect Proteins; D007447:Invertebrate Hormones; D011987:Receptors, Steroid; D013256:Steroids; D004441:Ecdysterone