title: The negative effects of bile acids and tumor necrosis factor-alpha on the transcription of cholesterol 7alpha-hydroxylase gene (CYP7A1) converge to hepatic nuclear factor-4: a novel mechanism of feedback regulation of bile acid synthesis mediated by nuclear receptors.
abstract: Bile acids regulate the cholesterol 7alpha-hydroxylase gene (CYP7A1), which encodes the rate-limiting enzyme in the classical pathway of bile acid synthesis. Here we report a novel mechanism whereby bile acid feedback regulates CYP7A1 transcription through the nuclear receptor hepatocyte nuclear factor-4 (HNF-4), which binds to the bile acid response element (BARE) at nt -149/-118 relative to the transcription start site. Using transient transfection assays of HepG2 cells with Gal4-HNF-4 fusion proteins, we show that chenodeoxycholic acid (CDCA) dampened the transactivation potential of HNF-4. Overexpression of a constitutive active form of MEKK1, an upstream mitogen-activated protein kinase (MAPK) module triggered by stress signals, strongly repressed the promoter activity of CYP7A1 via the consensus sequence for HNF-4 embedded in the BARE. Similarly, MEKK1 inhibited the activity of HNF-4 in the Gal4-based assay. The involvement of the MEKK1-dependent pathway in the bile acid-mediated repression of CYP7A1 was confirmed by co-transfecting a dominant negative form of the stress-activated protein kinase kinase, SEK, which abolished the effect of CDCA upon CYP7A1 transcription. Treatment of transfected HepG2 cells with tumor necrosis factor alpha (TNF-alpha), an activator of the MEKK1 pathway, led to the repression of CYP7A1 via the HNF-4 site in the BARE. TNF-alpha also inhibited the transactivation potential of HNF-4. Collectively, our results demonstrate for the first time that HNF-4, in combination with a MAPK signaling pathway, acts as a bile acid sensor in the liver. Furthermore, the effects of CDCA and TNF-alpha converge to HNF-4, which binds to the BARE of CYP7A1, suggesting a link between the cascades elicited by bile acids and pro-inflammatory stimuli in the liver.
authors.affiliation: Dipartimento di Scienze Farmacologiche, Facoltà di Farmacia, Università degli Studi di Milano, Milano 20133, Italy.
authors.forename: E
authors.initials: E
authors.lastname: De Fabiani
authors.forename: N
authors.initials: N
authors.lastname: Mitro
authors.forename: A C
authors.initials: AC
authors.lastname: Anzulovich
authors.forename: A
authors.initials: A
authors.lastname: Pinelli
authors.forename: G
authors.initials: G
authors.lastname: Galli
authors.forename: M
authors.initials: M
authors.lastname: Crestani
mesh_terms: D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D001647:Bile Acids and Salts / Q000096:biosynthesis / Q000494:pharmacology*; D002478:Cells, Cultured; D002790:Cholesterol 7-alpha-Hydroxylase / Q000235:genetics*; D004268:DNA-Binding Proteins; D005246:Feedback; D051557:Hepatocyte Nuclear Factor 4; D006801:Humans; D048728:MAP Kinase Kinase Kinase 1; D010750:Phosphoproteins / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D020218:Response Elements; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic / Q000187:drug effects*; D015533:Transcriptional Activation; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D001647:Bile Acids and Salts; D004268:DNA-Binding Proteins; D051557:Hepatocyte Nuclear Factor 4; C403128:MLX protein, human; D010750:Phosphoproteins; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; D002790:Cholesterol 7-alpha-Hydroxylase; D017346:Protein Serine-Threonine Kinases; D048728:MAP Kinase Kinase Kinase 1; C482576:MAP3K1 protein, human