The Orphan Nuclear Receptor DAX-1 Functions as a Potent Corepressor of the Constitutive Androstane Receptor (NR1I3)

Abstract

Regulation of gene transcription is controlled in part by nuclear receptors that function coordinately with coregulator proteins. The human constitutive androstane receptor (CAR; NR1I3) is expressed primarily in liver and regulates the expression of genes involved in xenobiotic metabolism as well as hormone, energy, and lipid homeostasis. In this report, DAX-1, a nuclear receptor family member with corepressor properties, was identified as a potent CAR regulator. Results of transaction and mutational studies demonstrated that both DAX-1's downstream LXXLL and its PCFQVLP motifs were critical contributors to DAX-1's corepression activities, although two other LXXM/LL motifs located nearer the N terminus had no impact on the CAR functional interaction. Deletion of DAX-1's C-terminal transcription silencing domain restored CAR1 transactivation activity in reporter assays to approximately 90% of control, demonstrating its critical function in mediating the CAR repression activities. Furthermore, results obtained from mammalian two-hybrid experiments assessing various domain configurations of the respective receptors showed that full-length DAX-1 inhibited the CAR-SRC1 interaction by approximately 50%, whereas the same interaction was restored to 90% of control when the DAX-1 transcription silencing domain was deleted. Direct interaction between CAR and DAX-1 was demonstrated with both alpha-screen and coimmunoprecipitation experiments, and this interaction was enhanced in the presence of the CAR activator 6-(4-chlorophenyl)imidazo[2,1-b]thiazole-5-carbaldehyde O- (3,4-dichlorobenzyl)oxime (CITCO). Results obtained in primary human hepatocytes further demonstrated DAX-1 inhibition of CAR-mediated CITCO induction of the CYP2B6 target gene. The results of this investigation identify DAX-1 as a novel and potent CAR corepressor and suggest that DAX-1 functions as a coordinate hepatic regulator of CAR's biological function.

MicroRNA-561 Promotes Acetaminophen-Induced Hepatotoxicity in HepG2 Cells and Primary Human Hepatocytes through Downregulation of the Nuclear Receptor Corepressor Dosage-Sensitive Sex-Reversal Adrenal Hypoplasia Congenital Critical Region on the X Chromosome, Gene 1 (DAX-1)

“…5A). Previous studies by other groups have shown that DAX-1 at mRNA and protein levels is clearly detected in HepG2 cells and primary human hepatocytes and mouse liver (Nedumaran et al, 2009(Nedumaran et al, , 2010Laurenzana et al, 2012). However, there is another published study in which DAX-1 protein and mRNA were not detected in human liver samples Fig.…”

“…DAX-1 is well known to have a key role in steroidogenesis, adrenogonadal development, and maintenance of stem cell pluripotency (McCabe, 2007). Although DAX-1 is expressed at a low level in the liver, it plays an important role in repressing diverse nuclear receptors such as HNF4a, CAR, liver X receptor-a (LXRa), and farnesoid X receptor (FXR) (Nedumaran et al, 2009(Nedumaran et al, , 2010Li et al, 2011;Laurenzana et al, 2012). DAX-1 appears to have a further role in hepatic lipogenesis and gluconeogenesis.…”

Deciphering the roles of the constitutive androstane receptor in energy metabolism

“…The LXXLL-binding pocket can also be targeted by some corepressors. Like many other nuclear receptors, CAR is also bound and repressed by the following common corepressors: nuclear receptor corepressor 1 (NCoR1), NCoR2/SMRT, and DAX-1 (NR0B1) [53,54,57,62] . Collectively, the LXXLL-mediated direct interaction of CAR and its cofactors is very important for the activity of CAR.…”

Role of Orphan Nuclear Receptor DAX-1/NR0B1 in Development, Physiology, and Disease

“…A mechanism employed by DAX-1 to exert its negative transcriptional effect involves direct protein-protein interaction with the C-terminal domain of SF-1 [16,29,45] and other nuclear receptors [46][47][48][49][50][51][52][53][54][55][56][57] (see the following list of transcription factors reported to interact with DAX-1: SF-1 [29,45]; LRH-1 [45]; ER /ER [46]; AR [47,48]; PR [48]; GR [51]; Nur77 [49]; ERR [50]; PPAR [52]; HNF4 [53]; LXR [54]; FXR [55]; CAR [56]; Esrrb [57]; Oct3/4 [58]; EWS/FLI1 [59]). In many cases protein-protein interactions between DAX-1 and other nuclear receptors were suggested to be mediated via the LXXLL motifs present in the DAX-1 N-terminal domain, which would occupy the coactivator-binding surface in the nuclear receptor LBD, competing with positive transcriptional cofactors and also recruiting the strong transcriptional silencing domain present in the DAX-1 C-terminus to the complex [46].…”