DAX-1 Functions as an LXXLL-containing Corepressor for Activated Estrogen Receptors

Abstract

We have discovered that the orphan receptor DAX-1 (NROB1) interacts with the estrogen receptors ER␣ and ER␤. Interaction occurs with ligand-activated ERs in solution and on DNA and is mediated by the unique DAX-1 N-terminal repeat domain. Each of the three repeats contains a leucine-rich receptor-binding motif, known as the LXXLL motif, which is usually found in nuclear receptor coactivators. We have demonstrated that DAX-1 functions as an inhibitor of ER activation in mammalian cells and suggest a mechanism involving two sequential events, occupation of the ligand-induced coactivator-binding surface and subsequent recruitment of corepressors. Accordingly, we propose that DAX-1 itself acts as a corepressor for ERs. Because DAX-1 is coexpressed with ERs in reproductive tissues, these interactions could play significant roles by influencing estrogen signaling pathways. Our results point at functional similarities between DAX-1 and the orphan receptor SHP (NROB2) in that they have acquired features of transcriptional coregulators that are unique for members of the nuclear receptor family.To transduce hormone and metabolic signaling to target genes, nuclear receptors require transcriptional cofactors that are collectively referred to as coregulators (1). These proteins exist in multiple complexes, possess multiple enzymatic activities, and bridge receptors to chromatin components or to the basal transcription machinery or to both. Multiple candidate proteins exist that are believed to be critical for the proper function of nuclear receptor signaling (1-3). The majority of coregulators bind to the receptor ligand-binding domain (LBD), 1 which is able to adopt different conformations depending on the ligand status and thereby discriminate between coactivators and corepressors. Functional and structural studies in particular elucidated the precise mechanisms of coactivator interaction with the ligand-inducible activation domain (AF-2) via short leucine motifs known as LXXLL or NR-Box (4 -8).We became interested in coregulators in particular that influence the transcriptional activity of estrogen receptors (ER). Two related subtypes, ER␣ and ER␤, play crucial roles in sex development and reproduction in multiple physiological processes as well as in cancer (9 -11). Previous research has provided detailed insights into structural and functional aspects of their interplay with coregulators (8,(12)(13)(14). Although agonist binding usually is associated with ER activation caused by coactivator recruitment, regulatory mechanisms have been proposed that could play a role in modulation and feedback control of estrogen signaling (15, 16). Recent work has revealed an unexpected role of the orphan receptor SHP (NROB2) in inhibiting transactivation of ERs (17, 18). Particularly, we have provided evidence that SHP, which consists only of an LBD and thus cannot bind target genes directly, has instead acquired a novel coregulator function by antagonizing the interactions of ERs with associated coactivators (18,19).The closest relati...

The structure of corepressor Dax-1 bound to its target nuclear receptor LRH-1

“…The data described in this work demonstrate that the Dax-1 LBD interacts directly with the LBD of LRH-1. Our finding that Dax-1 binds in the coactivator groove of LRH-1 is consistent with previous data showing that repression by Dax-1 could be through competitive inhibition (9). A novel twist that our analysis introduces to this hypothesis is that the repression site is located in the Dax-1 LBD and contains the subfamily conserved sequence motif PCFXXLP (Fig.…”

“…Because multiple regions of Dax-1 have been reported to bind nuclear receptors (4,5,(7)(8)(9)12), we assessed the binding of five different fragments of Dax-1 to LRH-1 LBD: its N-terminal region (aa 1-208), the LBD (aa 205-472), the LBD with preceding LXXL/ML repeats (aa 138-472 and 70-472), and full-length Dax-1 (aa 1-472). Of these fragments, only the putative Dax-1 LBD produced a stable complex with LRH-1 (Fig.…”

The orphan nuclear receptor DAX1 is up-regulated by the EWS/FLI1 oncoprotein and is highly expressed in Ewing tumors

“…44,45 Moreover, the nuclear localization of DAX1 in Ewing tumor cells is also in agreement with a transcriptional repressor function in these cells. In addition, DAX1 functions extend beyond the regulation of SF-1 target genes, since ligand-dependent transactivation by other nuclear receptors, such as oestrogen receptor, 46 androgen receptor, 47 and retinoic acid receptor 37 are also negatively regulated by DAX1.…”

Linking dioxins to diabetes: epidemiology and biologic plausibility.

“…Another possible pathway for cross-talk between Ah receptor and PPARγ signaling is through LXXLL domains (nuclear receptor boxes), which are present on numerous coactivators and corepressors that bind to members of the nuclear receptor superfamily (56)(57)(58)(59)(60)(61)(62)(63). Of these, SRC-1 was found to bind Ah receptor via LXXLL motifs (64), whereas RIP140 was shown to bind Ah receptor via an LXXLL-independent mechanism (65).…”