Interaction of the N-terminus of ecdysone receptor isoforms with the ligand-binding domain

• Published in: Molecular and cellular endocrinology Vol. 332; no. 1-2; pp. 293 - 300
• Main Authors: Tremmel, Ch, Schaefer, M., Azoitei, A., Ruff, H., Spindler-Barth, M.
• Format: Journal Article
• Language: English
• Published: Ireland Elsevier Ireland Ltd 30.01.2011; Elsevier
• Subjects: Animals; CHO Cells; Corepressor; Cricetinae; Cricetulus; Cross talk; Drosophila; Drosophila melanogaster - genetics; Drosophila melanogaster - metabolism; Drosophila Proteins - chemistry; Drosophila Proteins - genetics; Drosophila Proteins - metabolism; ecdysone; fluorescence; Fluorescence Resonance Energy Transfer; FRET; Insect; Ligands; Mutation; N/C-interaction; Nuclear receptor; Protein Binding; Protein Isoforms - chemistry; Protein Isoforms - genetics; Protein Isoforms - metabolism; Protein Structure, Tertiary; Receptors, Steroid - chemistry; Receptors, Steroid - genetics; Receptors, Steroid - metabolism; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; transcription (genetics); Transcription, Genetic; Aa; PR; RAR; YFP; Nuclear receptor; AF; Drosophila; N/C-interaction; DBD; CFP; ER; RXR; Corepressor; LBD; AR; EcRE; Usp; PPAR; EcR; VP16; FRET; wt; Insect; NTD; insect; nuclear receptor
• ISSN: 0303-7207; 1872-8057; 0303-7207
• DOI: 10.1016/j.mce.2010.11.013

▶ Basal and hormone induced transcriptional activity of EcR are regulated in an isoform specific manner. ▶ The NTD can act as an intramolecular coprepressor in an isoform specific manner. ▶ Without hormone the NTD interacts with the LBD. ▶ With hormone the NTD interaction is interrrupted and a salt bride is formed in the LBD. Ecdysone receptor (EcR) isoforms exert different biological functions, although they vary only in their N-terminal domain. Despite identical C-termini, which mediate hormone-induced activity, the influence of ligand is isoform specific, which indicates an N/C-interaction. The position of helix 12 with and without hormone varies among isoforms and modifies N/C-interaction determined by fluorescence resonance-energy transfer (FRET), which depends on the salt bridge between helices 4 and 12 of the ligand-binding domain (LBD). Disruption of the salt bridge by mutation of K497 (helix 4) had no effect on basal N/C-interaction, but prevented the hormone-induced increase, which was partially restored by a salt bridge with reversed polarity. The heterodimerization partner Ultraspiracle (Usp) can compensate for the disruption of the salt bridge. Without ligand the AB-domains of EcR-A and EcR-B1, but not EcR-B2, interact with the LBD via K497 and repress transcriptional activity. This intramolecular cross talk between N- and C-terminus along with the position of helix 12 stabilized by K497 regulates transcriptional activity of EcR isoforms.