The farnesoid X receptor FXRα/NR1H4 acquired ligand specificity for bile salts late in vertebrate evolution

Abstract

There is an extra sequence in the middle of the sFxr ligand binding domain (LBD) compared with the LBD of FXR␣. Luciferase reporter assays demonstrated that sFxr responds weakly to scymnol sulfate, bile salts, and synthetic FXR␣ ligands, in striking difference from human FXR␣ (hFXR␣). Interestingly, all-trans retinoic acid was capable of transactivating both hFXR␣ and sFxr. When the extra amino acids in the sFxr LBD were deleted and replaced with the corresponding sequence from hFXR␣, the mutant sFxr gained responsiveness to ursodeoxycholic acid, GW4064, and fexaramine. Surprisingly, chenodeoxycholic acid antagonized this activation. Together, these results indicate that FXR is an ancient nuclear receptor and suggest that FXR␣ may have acquired ligand specificity for bile acids later in evolution by deletion of a sequence from its LBD. Acquisition of this property may be an example of molecular exploitation, where an older molecule is recruited for a new functional role. nuclear receptor; structure/function relationship NUCLEAR RECEPTORS PLAY CRITICAL roles in development and physiology by sensing cellular levels of steroid hormones and dietary metabolites, although the true ligands for many of these receptors have not yet been identified or may even not exist. It has been demonstrated that nuclear receptors have shifted or gained their ligand specificity during evolution (4, 10); however, it remains to be determined how most nuclear receptors have acquired their ligand specificity and when this occurred in the course of evolution.FXR␣ (NR1H4) and FXR␤ (NR1H5) represent two forms of the farnesoid X receptor (FXR) in mammals, although FXR␤ is a pseudogene in humans and other primates (23). FXR␣ and FXR␤ are members of the class I nuclear receptors, which heterodimerize with the retinoid X receptors (RXR, NR2B) and regulate gene expression. FXRs share the typical structure of other members in this family, including an activation function domain 1 (AF1) at the N-terminus, followed by a conserved DNA binding domain (DBD), a ligand binding domain (LBD) that also contains an activation function domain 2 (AF2) at the C-terminus, as well as a hinge region that links the DBD and LBD (11). FXR␣ and FXR␤ share ϳ50% amino acid identity, but their ligand specificity does not overlap (23).

Modification of the cysteine residues in IκBα kinase and NF-κB (p65) by xanthohumol leads to suppression of NF-κB–regulated gene products and potentiation of apoptosis in leukemia cells

“…44,45 XN has been shown to be a natural ligand for FXR. 37 However, we found that XN inhibits TNF-induced NF-B activation in both FXR-positive and -negative cell lines, suggesting that FXR is not required for the NF-B-inhibitory activity of XN. However, this chalcone also inhibited bile acid-induced NF-B activation, which is linked with FXR.…”

“…37 We found that bile acids can activate NF-B in MCF-7 cells and that XN can suppress NF-B activation ( Figure 7D). Like XN, we found that guggulsterone also suppressed bile acid-induced NF-B activation.…”

Retinoic acid represses CYP7A1 expression in human hepatocytes and HepG2 cells by FXR/RXR-dependent and independent mechanisms

“…However, as a permissive nuclear receptor, FXR can also be activated through the ligand of its obligated heterodimeric partner, the retinoid X receptor (RXR, NR2B) ( 29,30 ), where 9-cis retinoic acid (9cRA) is its specifi c ligand ( 31 ( 32 ). While characterizing a primitive Fxr from the marine skate Leucoraja erinacea, we have also observed that atRA is a conserved activator of FXR in a reporter assay using cotransfection of skate Fxr and human RXR ␣ expression constructs ( 33 ). These observations raise two questions: (1) Is atRA or its metabolite a true ligand for FXR?…”

“…Human FXR ( hFXR ), RXR ␣ (hRXR ␣ ) and I-BABP promoter reporter constructs are described in a previous article ( 33 ). Mouse HNF4 ␣ and PGC-1 ␣ expression plasmids were provided by Dr. Peter Edwards (University of California at Los Angeles, Los Angeles, CA).…”

“…However, numerous reports have documented that hyperlipidemia is often a major side effect, including hypercholesterolemia and hypertriglyceridemia (34)(35)(36)(37)(38)(39). Increased total serum cholesterol levels described ( 33 ). The fi refl y luciferase activity was normalized by Renilla luciferase activity.…”