Two forms, CtUSP-1 and CtUSP-2, of the Chironomus tentans homolog of Ultraspiracle
(new nomenclature: Chironomus NR2B4) were described and verified as components of
the functional ecdysteroid receptor. The two forms differed from each other in the
most N-terminal regions of the A/B domain and were tested for several properties.
Both forms showed the ability to heterodimerize with CtEcR and interact with a variety
of direct repeat and palindromic EcREs, and both conferred specific ligand binding
when heterodimerized with EcR. CtUSP-2 showed a twofold higher ponasterone-binding
potential than CtUSP-1. Both USP forms demonstrated the ability to activate ecdysteroid-inducible
transcription in HeLa cells and the variations in the A/B domain of these forms were
not associated with detectable differences in transcriptional activation. Thus, the
two forms function similarly. Among species for which USP forms have been reported,
Chironomus is the most closely related one evolutionarily to Drosophila. Despite this
proximity, a variety of structural differences were noted in both the A/B and E domains
of USP between the two species. The Chironomus USP forms lack many of the amino acid
residues associated with the ligand-dependent AF2 transactivation function found in
all other RXRs and USPs reported so far.