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. Author manuscript; available in PMC: 2014 Mar 1.
Published in final edited form as: Hepatology. 2013 Feb 15;57(3):1098–1106. doi: 10.1002/hep.26146

Figure 3. FXR inhibits expression of gankyrin.

Figure 3

A. Gankyrin promoter contains two consensuses for C/EBP proteins. Upper. Two C/EBP consensuses within 1.4 kb gankyrin promoter are shown by red. Bottom image: Nucleotide sequence of the region containing C/EBP sites. The positions and sequences of primers used for ChIP assay are underlined. B. EMSA with C/EBP probes covering sites 1 and 2 using nuclear extracts from quiescent livers. Antibodies to C/EBPα and C/EBPβ were incorporated in binding reactions. Positions of C/EBPα, C/EBPβ, supershift (SS), non specific band (NS) and free probe are shown by arrows. C. C/EBPβ-HDAC1 complexes occupy gankyrin promoter in livers of WT mice, but are removed from the promoter in livers of FXR/SHP KO mice. ChIP assay was performed with chromatin solutions from WT quiescent livers and from livers of FXR/SHP KO mice. In; 1/100 of input, B; beads. D. Activation of FXR in mouse Hepa 1-6 cells increases levels of C/EBPβ and HDAC1 and inhibits expression of gankyrin. Western blotting was performed with protein extracts isolated from cells treated with DMSO (control) and with increasing concentrations of CDCA and GW4064. Bar graphs show levels of gankyrin, C/EBPβ and HDAC1 calculated as ratios to β-actin. E. FXR inhibits expression of gankyrin on the revel of mRNA. Expression of gankyrin mRNA was examined in mouse Hepa 1-6 cells treated with DMSO, CDCA and GW4064 by Q-RT-PCR.