Fig. 6.

Activation of FXR mutants by oxysterol 22(R)-OHC (A) and bile acid CDCA (B). Amino acid residues R331, L340, R351, and I352 in the FXR LBD were mutated to leucine, tyrosine, leucine, and tyrosine, respectively. After sequence verification, each of the resultant FXR mutants (100 ng) was cotransfected into Huh 7 cells on 24-well plates with 100 ng of pBSEP-Luc and 10 ng of the null-Renilla luciferase plasmid. Sixteen hours post-transfection, cells were treated with 10 μM 22(R)-OHC and 1% ETOH or 10 μM CDCA and 1% DMSO. The treatments were continued for 30 h, followed by detection of luciferase activities with the Dual-Luciferase Reporter assay system. Data are presented as mean ± S.D. of at least three separate experiments.