The structures of the liver X receptor LXR␤ (NR1H2) have been determined in complexes with two synthetic ligands, T0901317 and GW3965, to 2.1 and 2.4 Å, respectively. Together with its isoform LXR␣ (NR1H3) it regulates target genes involved in metabolism and transport of cholesterol and fatty acids. The two LXR␤ structures reveal a flexible ligand-binding pocket that can adjust to accommodate fundamentally different ligands. The ligand-binding pocket is hydrophobic but with polar or charged residues at the two ends of the cavity. T0901317 takes advantage of this by binding to His-435 close to H12 while GW3965 orients itself with its charged group in the opposite direction. Both ligands induce a fixed "agonist conformation" of helix H12 (also called the AF-2 domain), resulting in a transcriptionally active receptor.Liver X receptors (LXR) 1 are members of the superfamily of nuclear receptors. These transcription factors regulate target genes through a dynamic series of interactions with specific DNA response elements as well as transcriptional coregulators. The binding of ligand has profound effects on these interactions and has the potential to trigger both gene activation and, in some cases, gene silencing. There are 48 sequence-related nuclear receptors in humans and the family comprises receptors that recognize hormones, both steroidal and non-steroidal, but also receptors responding to metabolic intermediates and to xenobiotics. There are also a number of so-called orphan receptors where the natural ligand is unknown. Some of the receptors show a very specific and high affinity ligand binding, like the thyroid hormone receptors, whereas others have a substantially lower affinity for their ligands and are less discriminating in their ligand selectivity. Like many of the other nonsteroid hormone receptors, LXR functions as a heterodimer with the retinoid X receptor (RXR) to regulate gene expression (1, 2). Together with peroxisome proliferator-activated receptor (PPAR) and farnesoid X receptor (FXR), LXRs represent a subclass of so-called permissive RXR heterodimers. In this subclass, the RXR heterodimers can be activated independently by either the RXR ligand, the partner's ligand, or synergistically by both (3).LXRs consist of two closely related receptor isoforms encoded by separate genes, LXR␣ (NR1H3) and LXR␤ (NR1H2). LXR␣ shows tissue-restricted expression with the highest mRNA levels in the liver and somewhat lower levels in the kidney, small intestine, spleen, and adrenal gland (4, 5). In contrast, LXR␤ is ubiquitously expressed (6, 7). Both LXR isoforms can be activated by specific oxysterols that are formed in vivo (2,8,9). In view of the high degree of homology between the LXR isoforms (75% identity in the ligand-binding domain (LBD), 54% identity overall), it is perhaps not surprising that few subtypespecific biological responses have been described and that information on subtype selective ligands is limited. LXRs have been shown to regulate several genes involved in cholesterol and lipid homeos...

The dioxin receptor is a gene regulatory protein which exhibits many structural and functional similarities to steroid hormone receptors. In this study we compare the subunit composition of two forms of the dioxin receptor, sedimenting at approximately 9S and approximately 6S respectively, which are present in nuclear extract from wild‐type Hepa 1c1c7 mouse hepatoma cells following treatment in vivo with dioxin. The nuclear approximately 9S receptor form contained the 90 kd heat shock protein, hsp90. As assessed by a gel mobility shift assay, this receptor form did not bind to the xenobiotic response element (XRE) of the target gene cytochrome P‐450 IA1. In contrast, the smaller approximately 6S receptor form did not contain any immunochemically detectable hsp90. Moreover, this receptor form specifically bound to the XRE recognition sequence. Thus, the specific DNA binding activity of the dioxin receptor was inhibited by association with hsp90, and the approximately 9S dioxin receptor species could be regarded as a nonactive receptor form. Neither the approximately 9S nor the approximately 6S receptor forms were detected in nuclear extract from a dioxin treated mutant clone of Hepa 1 that expresses a nuclear translocation deficient receptor phenotype. We conclude that activation of the dioxin receptor is, at least, a two step process involving binding of the ligand and dissociation of hsp90 from the ligand‐binding receptor protein. Inhibition of the DNA binding activity of transcription factors by protein‐‐protein interaction has also been described for several steroid hormone receptors and for the NF kappa B factor.(ABSTRACT TRUNCATED AT 250 WORDS)

The intracellular basic region/helix-loop-helix (bHLH) dioxin receptor mediates signal transduction by dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin) and functions as a ligand-activated DNA binding protein directly interacting with target genes by binding to dioxin response elements. Here we show that the partially purified, ligand-bound receptor alone could not bind target DNA. In contrast, DNA binding by the receptor could be induced by addition of a cytosolic auxiliary activity which functionally and biochemically corresponded to the bHLH factor Arnt. While Arnt exhibited no detectable affinity for the dioxin response element in the absence of the dioxin receptor, it strongly promoted the DNA binding function of the ligand-activated but not the ligand-free receptor forms. Arnt also functionally reconstituted in vitro the DNA binding activity of a mutant, nuclear translocation-deficient dioxin receptor phenotype in cytosolic extracts from a dioxin-resistant hepatoma cell line. Importantly, coimmunoprecipitation experiments showed that Arnt physically interacted in solution with the ligand-activated dioxin receptor but failed to heterodimerize with the ligand-free, hsp9O-associated receptor form. Mutational analysis suggested that the functional interaction between these two factors occurred via the bHLH motif of Arnt. These data suggest that dioxin receptor activity is governed by a complex pattern of combinatorial regulation involving repression by hsp9O and then by ligand-dependent recruitment of the positive coregulator Arnt. The dioxin receptor system also provides the first example of signal-controlled dimerization of bHLH factors.Signal transduction by dioxins (most notably 2,3,7,8-tetrachlorodibenzo-p-dioxin [TCDD]) is mediated by the intracellular dioxin (or aryl hydrocarbon) receptor. The receptor binds dioxin and its planar aromatic congeners in a saturable manner with high affinity (for reviews, see references 22 and 49). The potent toxicity of dioxins is well established in animal models but is a matter of debate in humans (for a recent review, see reference 19). In animals the toxic effects are typified by thymic wasting and immune suppression, severe epithelial disorders, and tumor promotion (for a review, see reference 51). At the molecular level, dioxins are very potent inducers of transcription of a battery of target genes encoding xenobiotic metabolizing enzymes such as cytochrome P-450IA1, glutathione S-transferase Ya, aldehyde dehydrogenase, and quinone oxidoreductase (for a review, see reference 34). In addition, dioxin appears to transcriptionally regulate the expression of the growth modulatory genes for interleukin-1 and plasminogen activator inhibitor-2 (59). The dioxin induction response is mediated by single or multiple copies of dioxin-inducible transcriptional control elements (xenobiotic response elements [XREs]) in target promoters (16,18,47). In analogy to the current model of action of steroid hormone receptors (for a review, see reference 3), the ligand-activated dioxin recepto...

A series of substituted 2-benzyl-3-aryl-7-trifluoromethylindazoles were prepared as LXR modulators. These compounds were partial agonists in transactivation assays when compared to 1 (T0901317) and were slightly weaker with respect to potency and efficacy on LXRalpha than on LXRbeta. Lead compounds in this series 12 (WAY-252623) and 13 (WAY-214950) showed less lipid accumulation in HepG2 cells than potent full agonists 1 and 3 (WAY-254011) but were comparable in efficacy to 1 and 3 with respect to cholesterol efflux in THP-1 foam cells, albeit weaker in potency. Compound 13 reduced aortic lesion area in LDLR knockout mice equivalently to 3 or positive control 2 (GW3965). In a 7-day hamster model, compound 13 showed a lesser propensity for plasma TG elevation than 3, when the compounds were compared at doses in which they elevated ABCA1 and ABCG1 gene expression in duodenum and liver at equal levels. In contrast to results previously published for 2, the lack of TG effect of 13 correlated with its inability to increase liver fatty acid synthase (FAS) gene expression, which was up-regulated 4-fold by 3. These results suggest indazoles such as 13 may have an improved profile for potential use as a therapeutic agent.