Structure of the Murine Constitutive Androstane Receptor Complexed to Androstenol

Abstract

The nuclear receptor CAR is a xenobiotic responsive transcription factor that plays a central role in the clearance of drugs and bilirubin while promoting cocaine and acetaminophen toxicity. In addition, CAR has established a "reverse" paradigm of nuclear receptor action where the receptor is active in the absence of ligand and inactive when bound to inverse agonists. We now report the crystal structure of murine CAR bound to the inverse agonist androstenol. Androstenol binds within the ligand binding pocket, but unlike many nuclear receptor ligands, it makes no contacts with helix H12/AF2. The transition from constitutive to basal activity (androstenol bound) appears to be associated with a ligand-induced kink between helices H10 and H11. This disrupts the previously predicted salt bridge that locks H12 in the transcriptionally active conformation. This mechanism of inverse agonism is distinct from traditional nuclear receptor antagonists thereby offering a new approach to receptor modulation.

Evolutionary Trace-based Peptides Identify a Novel Asymmetric Interaction That Mediates Oligomerization in Nuclear Receptors

“…Trace residues in H11 are generally ranked higher than their counterparts in H3, consistent with a more widespread pattern of homodimerization and heterodimerization via helix 10/11 interactions (16,17,53,54). The lower, still significant, ranked residues of H3 are consistent with the more diverse functions among different NR subfamilies in that helix (20,28,(55)(56)(57). ET analysis suggests that both H3 and H11 have important biological roles among the nuclear receptors analyzed, including GCNF, and that mutations of trace residues in these helices should disrupt their normal interactions and reveal their specific function.…”

Evolution and Function of the NR1I Nuclear Hormone Receptor Subfamily (VDR, PXR, and CAR) with Respect to Metabolism of Xenobiotics and Endogenous Compounds

“…CAR differs from PXR and VDR in having high constitutive activity in the absence of ligand [42,[45][46][47]. CAR genes have so far been detected only in mammals ( Table 1).…”

“…Recently, three groups independently reported high-resolution crystallographic structures of human or mouse CARs bound to various ligands [45][46][47]. These structures provide insight into the high level of constitutive activity of CAR and to the ability of certain compounds, such as androstanol or androstenol in the mouse CAR [45], to act as inverse agonists. Fig.…”

“…The sequence identities of PXR and CAR across species are unusually low compared to the rest of vertebrates NRs, which tend to have comparable sequences identities between species 10-15% higher [1,27]. PXR and CAR even show considerable sequence variation at amino acid positions corresponding to residues that interact directly with ligands in X-ray crystallographic structures of human PXR [67,68,70,93], human CAR [47], and mouse CAR [45,46] bound to various ligands ( Fig. (3B,C)).…”

Interaction of PER2 with the Constitutive Androstane Receptor Possibly Links Circadian Rhythms to Metabolism

“…Both receptors seem to bind LXXLL motifs at the »end of helix 5« to the »end of helix 3«, which coincides with the lysine positioning. [31][32][33][34][35] Even though this could be projected onto many nuclear receptors, the position of the lysine at helix 5 could additionally explain the favouring of co-activators with a serine before LXXLL. 9,29,36 The hypothesis that PER2 and CAR interact directly was confirmed with co-immunoprecipitation (Figure 3).…”