Isotype-restricted corepressor recruitment:: a constitutively closed helix 12 conformation in retinoic acid receptors β and γ interferes with corepressor recruitment and prevents transcriptional repression

Retinoic acid receptors (RARs) are ligand-regulated transcription factors that play multiple roles in vertebrate development and differentiation. RARs as a class are capable of both repressing and activating target gene expression. Transcriptional repression is mediated through the recruitment of corepressor proteins such as SMRT. Notably, vertebrates encode three major forms of RARs, alpha, beta, and gamma, and these distinct RAR isotypes differ in the ability to recruit a corepressor. RARalpha strongly interacts with SMRT and can repress target gene transcription, whereas RARbeta and -gamma interact with SMRT only weakly and fail to repress. We report here the use of a genetic suppressor approach, based on a yeast two-hybrid interaction assay using Saccharomyces cerevisiae, for the isolation of RARbeta mutants that have gained the RARalpha-like corepressor phenotype, i.e., a strong interaction with SMRT and the ability to repress gene expression in vertebrate cells. Analysis of these gain-of-function mutants indicates that the different corepressor interaction properties of RARalpha, beta and gamma are determined by a gating mechanism through which amino acid differences in the helix 3 region of these receptors influence the position of the receptor C-terminal helix 12 domain. As a consequence, the RARbeta and RARgamma receptors appear to adopt a constitutively closed helix 12 conformation in the absence of hormone that may approximate the conformation of RARalpha when bound to hormone agonist. This closed helix 12 conformation in RARbeta and RARgamma blocks corepressor binding, prevents repression, and permits significant levels of target gene activation even in the absence of hormone. We refer to this phenomenon as a gate-latch model of corepressor regulation.