Ligand specificity and developmental expression of RXR and ecdysone receptor in the migratory locust

The ecdysone receptor1<fn id="fn1"><no>1</no>The avoid ambiguity in the terminology relating to this system, we have followed the usage suggested by under which the heterodimer is designated the ecdysone receptor and one of its components is referred to as EcR.</fn>, which is a heterodimer of EcR and the retinoic acid receptor (RXR) homolog, Ultraspiracle (USP), has been well studied in the evolutionarily advanced and derived insects, the flies and moths. It is less well characterized in more primitive insect orders such as the Orthoptera, which include the grasshoppers and locusts. Following our previous isolation from Locusta migratoria (Lm) of a shorter RXR isoform (now called LmRXR-S), the isolation of a second, longer isoform (LmRXR-L) that appears to have characteristics of a ligand-modulated nuclear receptor is reported here. Transcripts for both isoforms, as well as LmEcR, were detected in embryos and in females during oocyte maturation. After expression in E. coli, both LmRXR-S and LmRXR-L form heterodimers with recombinant LmEcR in vitro which bind the active ecdysteroid, ponasterone A. Binding was only weakly competed for by ecdysone agonists that are known to be toxic to more advanced insects, suggesting functionally significant divergence in EcR ligand binding domains. In contrast, the DNA binding domain of LmEcR is less divergent and a protein complex, presumably LmEcR/LmRXR, that bound the ecdysone response element, IR-1, was detected in locust nuclear extracts. Because of reports of juvenile hormone (JH III) binding to Drosophila USP and the observed in silico RXR-like ligand-binding site in LmRXR-L, the recombinant proteins were also tested for binding to JH III. Neither LmRXR isoform, alone or in combination with LmEcR, bound JH III at nanomolar concentrations. [Copyright &y& Elsevier]