The heterodimeric ecdysteroid receptor complex in the brown shrimp Crangon crangon: EcR and RXR isoform characteristics and sensitivity towards the marine pollutant tributyltin

Decapod crustaceans are characterized by multiple ecdysteroid receptor (EcR) and retinoid-X-receptor (RXR) isoforms, which likely exhibit variant dimerization and transactivation interactions. In the brown shrimp C. crangon we cloned C-terminally truncated CrcEcR and CrcRXR isoforms and isoforms exhibiting deletions within the hinge region. For the former, in silico modeling of the CrcEcR indicated that, where the conserved helices H10 and H11 of the ligand-binding domain (LBD) are missing, an alternative C-terminal α-helix repairs the ligand-binding pocket (LBP). The truncated CrcRXR isoforms lack a major part of the LBD (H4–H12), thereby compromising ligand binding and dimerization. Through an in vitro ecdysteroid responsive reporter assay, we showed that these natural receptor variations do not impair receptor functioning but probably alter the receptor dimerization preferences. By the same in vitro assay, using full-length CrcEcR and CrcRXR, the effect of tributyltin (TBT) on ecdysteroid-induced transactivation was evaluated. The transactivation by 10 nM PonA was reduced with 64% by 20 nM TBT. In silico modeling confirmed that TBT fits in the full-length CrcRXR–LBD. Furthermore, semi-quantitative PCR indicated altered expression of CrcEcR and CrcRXR isoforms after in vivo acute exposure to TBT, especially in the ovaries.

Putative pathway of tributyltin endocrine disruption through the retinoid-X-receptor (RXR) in Crangon crangon. TBT fits the ligand binding pocket of RXR, influencing the transactivational activity of other nuclear receptors (NRs) such as the ecdysteroid-receptor. DBD: RXR DNA binding domain; Hinge: RXR hinge region; LBD: RXR ligand binding domain.Figure optionsDownload as PowerPoint slideHighlights
► The brown shrimp Crangon crangon has multiple ecdysteroid receptor (CrcEcR) and retinoid-X-receptor (CrcRXR) isoforms.
► Truncation of the CrcEcR and CrcRXR receptors does not impair but alters transactivational activity.
► In silico modeling predicts tributyltin (TBT) fits the ligand binding pocket of CrcRXR.
► In vitro exposure shows that TBT affects ecdysteroid transactivation.
► TBT leads to altered in vivo expression of CrcEcR and CrcRXR, especially in the ovaries.