A Novel Bile Acid-Activated Vitamin D Receptor Signaling in Human Hepatocytes

Vitamin D receptor (VDR) is activated by natural ligands, 1 alpha, 25-dihydroxy-vitamin D(3) [1 alpha,25(OH)(2)D(3)] and lithocholic acid (LCA). Our previous study shows that VDR is expressed in human hepatocytes, and VDR ligands inhibit bile acid synthesis and transcription of the gene encoding cholesterol 7 alpha-hydroxylase (CYP7A1). Primary human hepatocytes were used to study LCA and 1 alpha,25(OH)(2)-D(3) activation of VDR signaling. Confocal immunofluorescent microscopy imaging and immunoblot analysis showed that LCA and 1 alpha, 25(OH)(2)-D(3) induced intracellular translocation of VDR from the cytosol to the nucleus and also plasma membrane where VDR colocalized with caveolin-1. VDR ligands induced tyrosine phosphorylation of c-Src and VDR and their interaction. Inhibition of c-Src abrogated VDR ligand-dependent inhibition of CYP7A1 mRNA expression. Kinase assays showed that VDR ligands specifically activated the c-Raf/MEK1/2/ extracellular signal-regulated kinase (ERK) 1/2 pathway, which stimulates serine phosphorylation of VDR and hepatocyte nuclear factor-4 alpha, and their interaction. Mammalian two-hybrid assays showed a VDR ligand-dependent interaction of nuclear receptor corepressor-1 and silencing mediator of retinoid and thyroid with VDR/retinoid X receptor-alpha(RXR alpha). Chromatin immunoprecipitation assays revealed that an ERK1/2 inhibitor reversed VDR ligand-induced recruitment of VDR, RXR alpha, and corepressors to human CYP7A1 promoter. In conclusion, VDR ligands activate membrane VDR signaling to activate the MEK1/2/ERK1/2 pathway, which stimulates nuclear VDR/RXR alpha recruitment of corepressors to inhibit CYP7A1 gene transcription in human hepatocytes. This membrane VDR-signaling pathway may be activated by bile acids to inhibit bile acid synthesis as a rapid response to protect hepatocytes from cholestatic liver injury. (Molecular Endocrinology 24: 1151-1164, 2010)