The LRH-1 site is required for FXR-induced expression of SHP.

(A) shows the location of IR-1-like half sites and an LRH-1 site in the −122/−69 region of the hSHP promoter. The latter was previously identified in the murine Shp promoter [35] and conserved in rat and human (see Figure S6). All 4 IR-1-like sites and the LRH-1 site were mutated and analyzed for the effect on FXR/CDCA-mediated induction of the −278/+10 hSHP promoter (B) mutating one of the IR-1 half-sites did not or only partially reduce FXR/CDCA-dependent activation of the −122/−69 hSHP promoter fragment, whereas mutations in the LRH-1 site strongly reduced the response of the −122/−69 hSHP promoter fragment to FXR/CDCA-stimulation. *) significantly different from CDCA treated 278/+10 WT (C) in the context of the −569/−10 hSHP promoter fragment the LRH-1 site is the dominant FXR/CDCA response element (over the previously identified IR-1). DLD-1 cells were transfected with hFXR and hRXRα expression plasmids and various hSHP promoter constructs as indicated. Cells were treated with or without 100 µmol/L CDCA. Luciferase activity was measured to determine the SHP promoter activity. a) significantly different from CDCA treated 569/+10 WT. b) significantly different from CDCA treated −569/+10 IR-1 KO. Data presented as means ± SD; n≥3. P≤0.05 for *), a) and b) in a pairwise comparison by Mann-Whitney U test.