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