A novel bile acid-activated vitamin D receptor signaling in human hepatocytes.
Vitamin D receptor (VDR) is activated by natural ligands, 1alpha, 25-dihydroxy-vitamin D(3) [1alpha,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 7alpha-hydroxylase (CYP7A1). Primary human hepatocytes were used to study LCA and 1alpha,25(OH)(2)-D(3) activation of VDR signaling. Confocal immunofluorescent microscopy imaging and immunoblot analysis showed that LCA and 1alpha, 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-4alpha, 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 (RXRalpha). Chromatin immunoprecipitation assays revealed that an ERK1/2 inhibitor reversed VDR ligand-induced recruitment of VDR, RXRalpha, 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/RXRalpha 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.
Han
Shuxin
S
Department of Integrative Medical Sciences, Northeastern Ohio University's Colleges of Medicine and Pharmacy, 4209 State Route 44, Rootstown, Ohio 44272, USA.
Li
Tiangang
T
Ellis
Ewa
E
Strom
Stephen
S
Chiang
John Y L
JY
D002117:Calcitriol / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002790:Cholesterol 7-alpha-Hydroxylase / Q000037:antagonists & inhibitors / Q000235:genetics; D004789:Enzyme Activation / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors; D056945:Hep G2 Cells; D051557:Hepatocyte Nuclear Factor 4 / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D008024:Ligands; D008095:Lithocholic Acid / Q000494:pharmacology*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D021381:Protein Transport / Q000187:drug effects; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D018167:Receptors, Calcitriol / Q000378:metabolism*; D047490:Retinoid X Receptor alpha / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013250:Steroid Hydroxylases / Q000235:genetics / Q000378:metabolism; D065668:Vitamin D3 24-Hydroxylase; D019061:src-Family Kinases / Q000378:metabolism
C496592:HNF4A protein, human; D051557:Hepatocyte Nuclear Factor 4; D008024:Ligands; D047428:Protein Kinase Inhibitors; D018167:Receptors, Calcitriol; D047490:Retinoid X Receptor alpha; D019000:Phosphotyrosine; D008095:Lithocholic Acid; D013250:Steroid Hydroxylases; D002790:Cholesterol 7-alpha-Hydroxylase; D065668:Vitamin D3 24-Hydroxylase; D019061:src-Family Kinases; D019908:Proto-Oncogene Proteins c-raf; D048049:Extracellular Signal-Regulated MAP Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; D002117:Calcitriol