Pergamon 09651748(95)00097-6 Insect Biochem. M&c. Bid. Vol. 26, No. 3, pp. 217-221, 1996 Copyright 0 1996 Elsevier Science Ltd Printed in Great Britain. All rights reserved 0965-1748/96 $15.00 + 0.00 Rapid Communication Bombyx EcR (BmEcR) and Bombyx USP (BmCFl) Combine to Form a Functional Ecdysone Receptor LUC SWEVERS,* LUCY CHERBASJ PETER CHERBAS,? KOSTAS IATROU*$ Received 11 October 1!>95; revised and accepted 16 November 1995 The Drosophila ecdysone receptor (DmEcR) is a member of the nuclear receptor superfamily; it functions as an obligate heterodimer with another nuclear receptor, DmUSP. EcR homologs have now been cloned from several other insects. We report here that one such homolog, BmEcR from the commercial silkmoth, Bombyx mori, is a functional ecdysone receptor. Upon dimerization with BmCFl, the silkmoth homolog of DmUSP, BmEcR binds the radiolabeled steroid ligand ‘Z51-iodoponasterone A with K,, = 1.1 nM, indistinguishable from that exhibited by DmEcR/DmUSP. BmEcR/BmCFl forms a specific complex with an ecdysone response element (E&E) derived from the heat shock protein 27 (hsp27) gene promoter of Drosophila; and, as with DmEcR/DmUSP, formation of this complex is stimulated by the presence of 20- hydroxyecdysone. Finally, BmEcR can substitute for DmEcR in an EcR-deficient Drosophila tissue culture line, stimulating trans-activation of an ecdysone-inducible reporter gene con- struct. Thus, BmEcR and BmCFl are the functional counterparts of DmEcR and DmUSP, respectively and, despite considerable sequence divergence between the Drosophila and Bom- byx proteins, the counterparts are-at least qualitatively-functionally equivalent. Ecdysone receptor BmEcR Ultraspiracle BmCFl Bombyx mori Silkmoth Lepidoptera Nuclear receptors INTRODUCTION In Drosophila, functional ecdysone receptors are hetero- dimers containing the polypeptides EcR (DmEcR, Koelle et al., 1991) and USP, (Ultraspiracle, DmUSP) (Yao et al., 1992, 1993; Thomas et al., 1993). Homologs of DmEcR have been isolated recently from several insect species (Chironomus tentans: Imhof et al., 1993; Aedes aegypti: Cho et al., 1995; Manduca sexta: Fujiwara et al., 1995; Bombyx mori: Swevers et al., 1995). The sequences of these EcR homologs are remarkably diverse, particularly when compared with those of ver- tebrate nuclear hormone receptors (see Cherbas and Cherbas, 1995, for review). For example, the ligand- *Department of Medical Biochemistry, The University of Calgary, 3330 Hospital Dr. N. W., Calgary AB T2N 4N1, Canada. TDepartment of Biology, Indiana University, Bloomington, Indiana 47405, U.S.A. $Author for correspondence. binding domains of the thyroid hormone receptors of Xenopus and humans are 93% identical in amino acid sequence (Brooks et al., 1989) and differ only by an extended insertion/deletion; similarly, the ligand-binding domains of the (Y forms of retinoic acid receptors in sala- manders and mammals are 97% identical (Ragsdale et al., 1989). In contrast, the known EcRs from Diptera and Lepidoptera are only 70-73% identical in their putative ligand-binding domains and show barely detectable simi- larities in their N-terminal (putative transactivation) domains. Moreover, DmEcR is also distinguished from all the other known EcRs by the presence of a long (about 150 residues) tail at its carboxy-terminus; a shorter, unre- lated tail is present in the Aedes EcR homolog, while the I&R homologs from Bombyx, Manduca and Chironomus have no such structure. Only within their DNA-binding domains (DBDs) do the insect receptors show consist- ently high levels of sequence conservation; for example, BmEcR and DmEcR are identical at 62/66 positions within their DBDs. In sum, although the ancestors of the 217