Inhibition of ligand induced promoter occupancy in vivo by a dominant negative RXR

Abstract

Background: Retinoid X receptors (RXRs) heterodimerize with other nuclear hormone receptors and control ligand mediated transcription. To address how RXRs function as heterodimers, we investigated activities of truncated RXR® and RXR¯that lack 20 conserved C-terminal amino acids.

Interaction between the Retinoid X Receptor and Transcription Factor IIB Is Ligand-dependent in Vivo

“…Since the ⌬390 -410 RXR deletion mutant has previously been shown to bind ligand (71), this suggests that ligand binding alone is not sufficient to promote interaction of RXR with TFIIB. Our results are consistent with dominant negative activity reported for the ⌬390 -410 mutant (55,56,66,69,70,71) and support other studies (66,69) suggesting that the mechanism involves binding of the receptor to DNA. In addition, our results suggest the mutant is unable to interact properly with TFIIB in vivo.…”

“…Also against an indirect or silent partner role for RXR is the observation that a dominant negative RXR mutant (e.g. ⌬AF-2 or c) could abrogate the ability of a heterodimer to augment gene activation (64,66,69,70,71); since RXRs associate weakly, or not at all with SMRT (16) or N-CoR (17,18), trans-repression is not easily explained. Similarly, an indirect mechanism of action does not explain the ability of RXR to function as a homodimer (68).…”

Distinct retinoid X receptor activation function-2 residues mediate transactivation in homodimeric and vitamin D receptor heterodimeric contexts

“…Such unmasking of the RXR corepressor site may be responsible for the reported dominant negative phenotype exhibited by an RXR AF-2 deletion mutant in inhibiting VDR-mediated, 1,25(OH) 2 D 3 -dependent transcription (Blanco et al 1996). To exclude the possible repressor activity that would result from the use of an RXR total AF-2 deletion mutant, the contribution of unliganded RXR to the overall transcriptional responsiveness of the RXR-VDR heterodimer was assessed in the current study using a series of RXR AF-2 point mutations.…”

“…This would not occur efficiently with the binding of the 9-cis RA natural ligand to RXR, because of the lack of direct AF-2 ligand contact, rendering it the subordinate receptor in most heterodimeric settings. Consequently, in this secondary role, RXR likely depends upon allosteric activation by the liganded primary partner followed by coactivator recruitment to create a stable supercomplex that remains bound to the DNA recognition site (Blanco et al 1996, Jurutka et al 2000a.…”

“…However, it has also been observed in other experimental systems that the presence of an RXR ligand, in combination with 1,25(OH) 2 D 3 , results in a synergistic transcriptional response , Kephart et al 1996, Li et al 1997. Moreover, a C-terminally truncated RXR lacking the AF-2 domain has been reported to exert a dominant negative effect on VDR-mediated transactivation, suggesting that an intact RXR AF-2 region is required for 1,25(OH) 2 D 3 -stimulated transcription (Blanco et al 1996). Interpretation of these latter results warrants caution, however, as the generation of such C-terminal deletions could also conceivably expose a constitutive corepressor binding site (Schulman et al 1996), thereby eliciting the observed dominant negative phenotype in RXR.…”