Molecular and functional comparison of 1,25‐dihydroxyvitamin D₃ and the novel vitamin D receptor ligand, lithocholic acid, in activating transcription of cytochrome P450 3A4

Abstract

The vitamin D receptor (VDR) binds to and mediates the effects of the 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) hormone to alter gene transcription. A newly recognized VDR ligand is the carcinogenic bile acid, lithocholic acid (LCA). We demonstrate that, in HT-29 colon cancer cells, both LCA and 1,25(OH)(2)D(3) induce expression of cytochrome P450 3A4 (CYP3A4), an enzyme involved in cellular detoxification. We also show that LCA-VDR stimulates transcription of gene reporter constructs containing DR3 and ER6 vitamin D responsive elements (VDREs) from the human CYP3A4 gene. Utilizing gel mobility shift, pulldown, and mammalian two-hybrid assays, we observe that: (i) 1,25(OH)(2)D(3) enhances retinoid X receptor (RXR) heterodimerization with VDR more effectively than LCA, (ii) the 1,25(OH)(2)D(3)-liganded VDR-RXR heterodimer recruits full-length SRC-1 coactivator, whereas this interaction is minimal with LCA unless LXXLL-containing fragments of SRC-1 are employed, and (iii) both 1,25(OH)(2)D(3) and LCA enhance the binding of VDR to DRIP205/mediator, but unlike 1,25(OH)(2)D(3)-VDR, LCA-VDR does not interact detectably with NCoA-62 or TRIP1/SUG1, suggesting a different pattern of LCA-VDR comodulator association. Finally, residues in the human VDR (hVDR) ligand binding domain (LBD) were altered to create mutants unresponsive to 1,25(OH)(2)D(3)- and/or LCA-stimulated transactivation, identifying S237 and S225/S278 as critical for 1,25(OH)(2)D(3) and LCA action, respectively. Therefore, these two VDR ligands contact distinct residues in the binding pocket, perhaps generating unique receptor conformations that determine the degree of RXR and comodulator binding. We propose that VDR is a bifunctional regulator, with the 1,25(OH)(2)D(3)-liganded conformation facilitating high affinity endocrine actions, and the LCA-liganded configuration mediating local, lower affinity cellular detoxification by upregulation of CYP3A4 in the colon.

Intestinal cell-specific vitamin D receptor (VDR)-mediated transcriptional regulation of CYP3A4 gene

“…2A). In the case of VDR, this observation could be explained based on the different recruitment of coactivators to VDR by 1,25OHvitD 3 and LCA [37]. In support of this, the structure-function relationships of VDR and LCA revealed that LCA interacts with different amino acids in the ligand binding domain (LBD) of VDR in comparison with 1,25OHvitD 3 [38,39].…”

CYP2R1 is a major, but not exclusive, contributor to 25-hydroxyvitamin D production in vivo

“…However, Cyp27a1-null mice showed a normal phenotype in the vitamin D activation pathway (15). Over the years, the identification of the nuclear hormone response elements in the Cyp3a4 gene promoter has drawn attention to the most abundant P450 in human liver and intestine as a possible vitamin D 25-hydroxylase (46)(47)(48)(49)(50), although the vitamin D receptor is undetectable in the liver (51). CYP3A4 is a well-known xenobiotic-metabolizing enzyme and can act on approximately half of the drugs in use.…”

Intestinal CYP3A4 protects against lithocholic acid-induced hepatotoxicity in intestine-specific VDR-deficient mice

“…It was suggested that LCA enters the colonic enterocyte, activates VDR, and thereby induces its own detoxifi cation ( 15 ). LCA was also shown to induce hepatic CYP3A4, a target gene regulated by VDR that is involved in bile acid metabolism, notably the oxidation of bile acids ( 16 ).…”