Monitoring ligand-mediated nuclear receptor-coregulator interactions by noncovalent mass spectrometry.
Fiche publication
Date publication
décembre 2004
Auteurs
Membres identifiés du Cancéropôle Est :
Dr CIANFERANI Sarah, Dr GRONEMEYER Hinrich, Dr MORAS Dino, Dr VAN DORSSELAER Alain
Tous les auteurs :
Sanglier S, Bourguet W, Germain P, Chavant V, Moras D, Gronemeyer H, Potier N, Van Dorsselaer A
Lien Pubmed
Résumé
Retinoid receptors are ligand-dependent transcription factors belonging to the nuclear receptor superfamily. Retinoic acid (RARalpha, beta, gamma) and retinoid X (RXRalpha, beta, gamma) receptors mediate the retinoid/rexinoid signal to the transcriptional machineries by interacting at the first level with coactivators or corepressors, which leads to the recruitment of enzymatically active noncovalent complexes at target gene promoters. It has been shown that the interaction of corepressors with nuclear receptors involves conserved LXXI/HIXXXI/L consensus sequences termed corepressor nuclear receptor (CoRNR) boxes. Here we describe the use of nondenaturing electrospray ionization mass spectrometry (ESI-MS) to determine the characteristics of CoRNR box peptide binding to the ligand binding domains of the RARalpha-RXRalpha heterodimer. The stability of the RARalpha-RXRalpha-CoRNR ternary complexes was monitored in the presence of different types of agonists or antagonists for the two receptors, including inverse agonists. These results show unambiguously the differential impact of distinct retinoids on corepressor binding. We show that ESI-MS is a powerful technique that complements classical methods and allows one to: (a) obtain direct evidence for the formation of noncovalent NR complexes; (b) determine ligand binding stoichiometries and (c) monitor ligand effects on these complexes.
Référence
Eur J Biochem. 2004 Dec;271(23-24):4958-67.