FIGURE 8.

XBP1s transactivates the transcriptional activity of the Fgf21 promoter. A, luciferase reporter assay. 293T cells were transfected with the Luc construct under the control of the mouse Fgf21 promoter that spans the region from −1983 to +5. Cells were then treated with DMSO, thapsigargin (Tg), or Tm for 6 h. Data were normalized to the DMSO control and are shown as the mean ± S.E. (n = 3 independent experiments). **, p < 0.01 by one-way ANOVA. B, sequence comparison of mouse, rat, and human Fgf21 promoter. Shown is the region that contains the signature XBP1s-binding element (ERSE) indicated by black boxes. C, luciferase reporter assay. 293T cells were co-transfected with the empty control vector or pCMV-XBP1s plasmid together with Luc constructs under the control of the mouse Fgf21 promoter (WT) or that with the CCACG element deleted (ΔCCACG). Data are shown as the mean ± S.E. (n = 3 independent experiments). **, p < 0.01 by two-way ANOVA. D, chromatin immunoprecipitation (IP) assay. Primary hepatocytes were infected for 2 days with adenoviruses expressing EGFP or XBP1s. ChIP was performed using control IgG or XBP1s antibody prior to PCR amplification of the indicated region (nucleotide −280 to −24) of the promoter. E, ChIP assay of extracts from 293T cells co-transfected with the empty control vector or pCMV-XBP1s plasmid together with the WT or ΔCCACG mutant reporter construct. ChIP was performed using anti-XBP1s antibody. F, ChIP assay of liver nuclear extracts from male C57BL/6 mice treated with PBS (vehicle) or Tm (1 mg/kg body weight) using control IgG or anti-XBP1s antibody. Quantitative PCR results are shown as the mean ± S.E. (n = 5/group). **, p < 0.01 by two-way ANOVA.