Molecular mechanisms of nuclear receptor regulation of bile acid synthesis
transactivation; feedback-regulation; identification; cholesterol 7-alpha-hydroxylase; transcription; heterodimer partner; repression; shp; factor 4-alpha; 7-alpha-hydroxylase gene cyp7a1
Chiang J Y L; Zhang M; Chen W; Norlin M; Owsley E
2003
2003
Book/Monograph
n/a
Structure and functions of human oxysterol 7 alpha-hydroxylase cDNAs and gene CYP7B1
27-hydroxycholesterol; 7-alpha-hydroxylase; bile-acid synthesis; bile-acid synthesis; Biochemistry & Molecular Biology; brain; cholesterol; cholesterol 7 alpha-hydroxylase; cytochrome P450; cytochrome P450; dehydroepiandrosterone (DHEA); dehydroepiandrosterone (DHEA); human liver-microsomes; identification; neurosteroids; transcription
Oxysterol 7 alpha-hydroxylase has broad substrate specificity for sterol metabolites and may be involved in many metabolic processes including bile acid synthesis and neurosteroid metabolism, The cloned human oxysterol 7 alpha-hydroxylase (CYP7B1) cDNA encodes a polypeptide of 506 amino acid residues that shares 40% sequence identity to human cholesterol 7 alpha-hydroxylase (CW7A1), the rate-limiting enzyme in the conversion of cholesterol to bile acids in the liver. In contrast to the liver-specific expression of CYP7A1, CYP7B1 mRNA transcripts were detected in human tissues involved in steroid genesis (brain, testes, ovary, and prostate) and in bile acid synthesis (liver) and reabsorption (colon, kidney, and small intestine). The human oxysterol 7 alpha-hydroxylase transiently expressed in 293/T cells was able to catalyze 7 alpha-hydroxylation of 27-hydroxycholesterol and dehydroepiandrosterone (DHEA), The human CYP7A1 and CYP7B1 both contain six exons and five introns. However, CPP7B1 spans at least 65 kb of the genome and is about 6-fold longer than CYP7A1. The transcription start site (+1) was localized 204 bp upstream of the initiation codon, No TATA box-like sequence was found near the transcription start site. Transient transfection assays of CYP7B1 promoter/luciferase reporter constructs in HepG2 cells revealed that the promoter was highly active. The 5' upstream region from nt -83 to +189 is the core promoter of the gene.
Wu Z L; Martin K O; Javitt N B; Chiang J Y L
Journal of Lipid Research
1999
1999-12
Journal Article
n/a
Aldosterone stimulates proliferation of cardiac fibroblasts by activating Ki-RasA and MAPK1/2 signaling
angiotensin-ii; Cardiovascular System & Cardiology; differentiation; fibrosis; gene-expression; Heart failure; identification; induction; Kirsten Ras; mineralocorticoid; mitogen-activated protein kinase; myofibroblast; na+ reabsorption; Physiology; receptor; spironolactone; transcription
Aldosterone plays a pathological role in cardiac fibrosis by directly affecting cardiac fibroblasts. Understanding of the cellular mechanisms of aldosterone action in cardiac fibroblasts, however, is rudimentary. One possibility is that aldosterone promotes proliferation of cardiac fibroblasts by activating specific cellular signaling cascades. The current study tests whether aldosterone stimulates proliferation of isolated adult rat cardiac myofibroblasts (RCF) by activating Kirsten Ras (Ki-RasA) and its effector, the MAPK1/2 cascade. Aldosterone (10 nM) significantly increased RCF proliferation. This action was sensitive to the mineralocorticoid receptor (MR) antagonist spironolactone. Expression of MR in RCF and the whole rat heart was confirmed by immunoblotting. Aldosterone significantly increased absolute and active (GTP bound) Ki-RasA levels in RCF. Aldosterone, in addition, significantly increased phospho-c-Raf and phospho-MAPK1/2. The effects of aldosterone on Ki-RasA and phospho-c-Raf proteins were inhibited by spironolactone but not RU-486, suggesting that aldosterone acts via MR. Inhibitors of MEK1/2 and c-Raf prevented aldosterone-induced activation of MAPK1/2 and proliferation. These results show that aldosterone directly increases RCF proliferation through MR-dependent activation of Ki-RasA and its effector, the MAPK1/2 cascade. Activation of cardiac fibroblasts through such a cascade may play a role in the pathological actions exerted by aldosterone on the heart.
Stockand J D; Meszaros J G
American Journal of Physiology-Heart and Circulatory Physiology
2003
2003-01
Journal Article
<a href="http://doi.org/10.1152/ajpheart.00421.2002" target="_blank" rel="noreferrer noopener">10.1152/ajpheart.00421.2002</a>
The stimulatory effect of LXR alpha is blocked by SHP despite the presence of a LXR alpha binding site in the rabbit CYP7A1 promoter
alpha-fetoprotein transcription; bile-acid biosynthesis; Biochemistry & Molecular Biology; cholesterol 7 alpha-hydroxylase; cholesterol 7-alpha-hydroxylase gene; dietary-cholesterol; dietary-cholesterol; factor; farnesoid X receptor; farnesoid X receptor; inhibition; liver X receptor; messenger-rna levels; nuclear receptor; orphan; rat; SHP; taurocholate; transcription
The transcription of the cholesterol 7 alpha-hydroxylase gene (CYP7A1) is greatly decreased in cholesterol-fed rabbits. To determine whether the molecular structure of the promoter is responsible for this downregulation, we cloned the rabbit CYP7A1 promoter, identified the binding sites for a-fetoprotein transcription factor (FTF) and liver X receptor (LXR alpha), and studied the effects of FTF, LXR alpha, and SHP on its transcription. Adding LXR alpha/retinoid X receptor together with their ligands (L/R) to the promoter/reporter construct transfected into HepG2 cells greatly increased its activity. FTF did not increase promoter activity, nor did it enhance the stimulatory effect of L/R. Mutating the FTF binding site abolished the promoter baseline activity. Increasing amounts of SHP abolished the effect of L/R, and FTF enhanced the ability of SHP to decrease promoter activity below baseline levels. Thus, downregulation of CYP7A1 in cholesterol-fed rabbits is attributable secondarily to the activation of farnesoid X receptor, which increases SHP expression to override the positive effects of LXR alpha. Although FFF is a competent factor for maintaining baseline activity, it does not further enhance and may suppress CYP7A1 transcription.
Shang Q; Pan L X; Saumoy M; Chiang J Y L; Tint G S; Salen G; Xu G R
Journal of Lipid Research
2006
2006-05
Journal Article
<a href="http://doi.org/10.1194/jlr.M500449-JLR200" target="_blank" rel="noreferrer noopener">10.1194/jlr.M500449-JLR200</a>
On the mechanism of bile acid inhibition of rat sterol 12alpha-hydroxylase gene (CYP8B1) transcription: roles of alpha-fetoprotein transcription factor and hepatocyte nuclear factor 4alpha.
Animals; Rats; Gene Expression Regulation; Base Sequence; Cattle; Cytochrome P-450 Enzyme System/*genetics; Hepatocyte Nuclear Factor 4; *DNA-Binding Proteins; Bile Acids and Salts/*pharmacology; Steroid Hydroxylases/*genetics; alpha-Fetoproteins/genetics/*physiology; DNA; Phosphoproteins/genetics/*physiology; Steroid 12-alpha-Hydroxylase; Transcription Factors/genetics/*physiology; Sprague-Dawley; RNA; Transcription; Genetic; Promoter Regions; Messenger/genetics; Enzymologic/*drug effects/physiology; Genetic/*drug effects
The sterol 12alpha-hydroxylase (CYP8B1) is a key enzyme of the bile acid biosynthetic pathway. It regulates the composition of bile acids in bile, i.e. ratio between cholic acid (CA) and chenodeoxycholic acid (CDCA). In similarity with cholesterol 7alpha-hydroxylase (CYP7A1), this enzyme is subjected to a negative feedback regulation by bile acids. It has been recently reported that bile acid-activated farnesoid X receptor (FXR) induces the small heterodimer partner (SHP) that interacts with alpha-fetoprotein transcription factor (FTF) and down-regulates CYP7A1 transcription. We studied whether the same mechanism also regulated rat CYP8B1 gene transcription. Feeding rats with CDCA caused a
Yang Yizeng; Zhang Ming; Eggertsen Gosta; Chiang John Y L
Biochimica et biophysica acta
2002
2002-06
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
Transcriptional regulation of human oxysterol 7 alpha-hydroxylase gene (CYP7B1) by Sp1.
Humans; Protein Binding; Gene Expression Regulation; Cell Line; Transfection; Base Sequence; Binding Sites/genetics; Molecular Sequence Data; Cytochrome P450 Family 7; Mutagenesis; Luciferases/genetics/metabolism; Recombinant Fusion Proteins/genetics/metabolism; CpG Islands/genetics; Cytochrome P-450 Enzyme System/*genetics/metabolism; DNA/genetics; Sequence Deletion; Sp1 Transcription Factor/metabolism/*physiology; Steroid Hydroxylases/*genetics/metabolism; Cultured; Binding; Competitive; Transcription; Genetic; Enzymologic; Tumor Cells; Site-Directed; Regulatory Sequences; Nucleic Acid/genetics
Oxysterol 7 alpha-hydroxylase catalyzes hydroxylation of oxysterols and neurosterols and plays a role in the alternative bile acid synthesis pathway. This gene is widely expressed in many organs and peripheral tissues and may protect tissues from the toxicity of oxysterols. Mutation in CYP7B1 caused neonatal cholestasis. To examine the regulatory mechanisms governing CYP7B1 expression, the 5' flanking sequence of the CYP7B1 was analyzed and revealed a CpG island of about 1.2 kb. Transient transfection assays of deletion mutants of the CYP7B1 promoter-luciferase reporter gene in human liver-derived HepG2, fibroblast NT1088, and human embryonic kidney 293 cell lines revealed that the region from -291 to +189 was critical for gene transcription. Three GC box sequences located between -25 and +10 were essential for basal transcription because mutations of these sequences markedly reduced promoter activity. Sp1 and Sp3 bound to these sequences as demonstrated by DNase I footprinting assays and electrophoretic mobility shift assay. Thus, regulation of CYP7B1 transcription by Sp1 may play a pivotal role in regulating oxysterol levels, which regulate cholesterol metabolism.
Wu Z; Chiang J Y
Gene
2001
2001-07
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
Structure and functions of human oxysterol 7alpha-hydroxylase cDNAs and gene CYP7B1.
Humans; Animals; Mice; Cell Line; Transfection; Base Sequence; Molecular Sequence Data; Chromosome Mapping; Cytochrome P450 Family 7; DNA; Luciferases/genetics; Cytochrome P-450 Enzyme System/*genetics/metabolism; Steroid Hydroxylases/*genetics/metabolism; Hydroxycholesterols/metabolism; Codon; Northern; Blotting; Transcription; Genetic; Cloning; Molecular; Genetic/genetics; Promoter Regions; Nucleic Acid; Complementary/biosynthesis/*isolation & purification; Initiator; Regulatory Sequences
Oxysterol 7alpha-hydroxylase has broad substrate specificity for sterol metabolites and may be involved in many metabolic processes including bile acid synthesis and neurosteroid metabolism. The cloned human oxysterol 7alpha-hydroxylase (CYP7B1) cDNA encodes a polypeptide of 506 amino acid residues that shares 40% sequence identity to human cholesterol 7alpha-hydroxylase (CYP7A1), the rate-limiting enzyme in the conversion of cholesterol to bile acids in the liver. In contrast to the liver-specific expression of CYP7A1, CYP7B1 mRNA transcripts were detected in human tissues involved in steroid genesis (brain, testes, ovary, and prostate) and in bile acid synthesis (liver) and reabsorption (colon, kidney, and small intestine). The human oxysterol 7alpha-hydroxylase transiently expressed in 293/T cells was able to catalyze 7alpha-hydroxylation of
Wu Z; Martin K O; Javitt N B; Chiang J Y
Journal of lipid research
1999
1999-12
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
Peroxisome proliferator-activated receptor alpha (PPARalpha) and agonist inhibit cholesterol 7alpha-hydroxylase gene (CYP7A1) transcription.
Humans; Animals; Binding Sites; Protein Binding; Rats; Gene Expression Regulation; Species Specificity; Liver/metabolism; Transcriptional Activation; Hepatocyte Nuclear Factor 4; Response Elements; *DNA-Binding Proteins; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; Retinoid X Receptors; Anticholesteremic Agents/pharmacology; Cholesterol 7-alpha-Hydroxylase/biosynthesis/*genetics; Clofibric Acid/pharmacology; Peroxisome Proliferators/pharmacology; Phosphoproteins/metabolism; Pyrimidines/pharmacology; Transcription Factors/*agonists/metabolism; Genes; Receptors; Models; Transcription; Genetic; Enzymologic; Reporter; Retinoic Acid/metabolism; Promoter Regions; Nucleic Acid; Cytoplasmic and Nuclear/*agonists; *Regulatory Sequences
Fibrates are widely used hypolipidemic drugs that regulate the expression of many genes involved in lipid metabolism by activating the peroxisome proliferator-activated receptor alpha (PPARalpha). The objective of this study was to investigate the mechanism of action of peroxisome proliferators and PPARalpha on the transcription of cholesterol 7alpha-hydroxylase, the rate-limiting enzyme in the conversion of cholesterol to bile acids in the liver. When cotransfected with the expression vectors for PPARalpha and RXRalpha, Wy14,643 reduced human and rat cholesterol 7alpha-hydroxylase gene (CYP7A1)/luciferase reporter activities by 88% and 43%, respectively, in HepG2 cells, but not in CV-1 or CHO cells. We have mapped the peroxisome proliferator response element (PPRE) to a conserved sequence containing the canonical AGGTCA direct repeats separated by one nucleotide (DR1). This DR1 sequence was mapped previously as a binding site for the hepatocyte nuclear factor 4 (HNF-4) which stimulates CYP7A1 transcription. Electrophoretic mobility shift assay (EMSA) showed no direct binding of in vitro synthesized PPARalpha/RXRalpha heterodimer to the DR1 sequence. PPARalpha and Wy14,643 did not affect HNF-4 binding to the DR1. However, Wy14,643 and PPARalpha/RXRalpha significantly reduced HNF-4 expression in HepG2 cells. These results suggest that PPARalpha and agonist repress cholesterol 7alpha-hydroxylase activity by reducing the availability of
Marrapodi M; Chiang J Y
Journal of lipid research
2000
2000-04
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
Regulation of cholesterol 7alpha-hydroxylase gene (CYP7A1) transcription by the liver orphan receptor (LXRalpha).
Humans; Animals; Binding Sites; Rats; Species Specificity; Transfection; Gene Expression Regulation/drug effects; Organ Specificity; Transcription Factors/genetics/metabolism; Cricetinae; Response Elements; Luciferases/genetics/metabolism; Retinoid X Receptors; Cholesterol 7-alpha-Hydroxylase/drug effects/*genetics/metabolism; Hydroxycholesterols; Liver/physiology; Lovastatin/pharmacology; Mevalonic Acid/metabolism/pharmacology; Nicotinic Acids/pharmacology; Polyisoprenyl Phosphates/pharmacology; Tetrahydronaphthalenes/pharmacology; Cells; Cultured; Receptors; Transcription; Genetic; Retinoic Acid/genetics/metabolism; Steroid/genetics/*metabolism
The cholesterol 7alpha-hydroxylase gene (CYP7A1) plays an important role in regulation of bile acid biosynthesis and cholesterol homeostasis. Oxysterol receptor, LXR, stimulates, whereas the bile acid receptor, FXR, inhibits CYP7A1 transcription. The goal of this study was to investigate the role of LXRalpha on the regulation of rat, human and hamster CYP7A1 transcription in its native promoter and cellular context. Cotransfection with LXRalpha and RXRalpha expression plasmids strongly stimulated rat CYP7A1/luciferase reporter activity in HepG2 cells and oxysterol was not required. However, LXRalpha had much less effect on hamster and no significant effect on human CYP7A1 promoter activity in HepG2 cells. In Chinese hamster ovary cells, cotransfection with LXRalpha stimulated reporter activity by less than 2-fold and addition of 22(R)-hydroxycholesterol caused a small but significant stimulation of rat, human and hamster CYP7A1 promoter activity. At least two direct repeats of AGGTCA-like sequences with 4-base spacing (DR4) and five-base spacing (DR5), in previously identified bile acid response elements of the rat CYP7A1 were able to bind LXRalpha/RXRalpha and confer LXRalpha stimulation. However, LXRalpha did not bind to the corresponding sequences of the human gene and bound weakly to hamster and mouse DR4 sequences. Therefore, rats and mice have the unusual capacity to convert cholesterol to bile acids by LXRalpha-mediated stimulation of CYP7A1 transcription, whereas other species do not respond to cholesterol and develop hypercholesterolemia on a diet high in cholesterol.
Chiang J Y; Kimmel R; Stroup D
Gene
2001
2001-01
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
Nuclear receptor-mediated repression of human cholesterol 7alpha-hydroxylase gene transcription by bile acids.
Humans; Animals; Rats; Cell Line; Transfection; Liver/metabolism; Reverse Transcriptase Polymerase Chain Reaction; DNA/metabolism; CHO Cells; Cricetinae; Cholesterol 7-alpha-Hydroxylase/*genetics; Bile Acids and Salts/*pharmacology; *Membrane Glycoproteins; *Hydroxysteroid Dehydrogenases; Caco-2 Cells; Carrier Proteins/genetics/physiology; DNA-Binding Proteins/drug effects/genetics/physiology; Gene Expression/*drug effects; Kidney; Luciferases/genetics; Recombinant Fusion Proteins/metabolism; Retinoid X Receptors; Taurocholic Acid/pharmacology; Transcription Factors/drug effects/genetics/physiology; Cultured; Receptors; RNA; Genetic/drug effects; Messenger/analysis; Transcription; Genetic; Tumor Cells; Promoter Regions; Embryo; Cytoplasmic and Nuclear/genetics/*physiology; Mammalian; Retinoic Acid/genetics/physiology
Hydrophobic bile acids strongly repressed transcription of the human cholesterol 7alpha-hydroxylase gene (CYP7A1) in the bile acid biosynthetic pathway in the liver. Farnesoid X receptor (FXR) repressed CYP7A1/Luc reporter activity in a transfection assay in human liver-derived HepG2 cells, but not in human embryonic kidney (HEK) 293 cells. FXR-binding activity was required for bile acid repression of CYP7A1 transcription despite the fact that FXR did not bind to the CYP7A1 promoter. FXR-induced liver-specific factors must be required for mediating bile acid repression. Bile acids and FXR repressed endogenous CYP7A1 but stimulated alpha-fetoprotein transcription factor (FTF) and small heterodimer partner (SHP) mRNA expression in HepG2 cells. Feeding of rats with chenodeoxycholic acid repressed CYP7A1, induced FTF, but had no effect on SHP mRNA expression in the liver. FTF strongly repressed CYP7A1 transcription in a dose-dependent manner, and SHP further inhibited CYP7A1 in HepG2 cells, but not in HEK 293 cells. FXR only moderately stimulated SHP transcription, whereas FTF strongly inhibited SHP transcription in HepG2 cells. Results revealed that FTF was a dominant negative factor that was induced by bile acid-activated FXR to inhibit both CYP7A1 and SHP transcription. Differential regulation of FTF and SHP expression by bile acids may explain the wide variation in CYP7A1 expression and the rate of bile acid synthesis and regulation in different species.
Chen W; Owsley E; Yang Y; Stroup D; Chiang J Y
Journal of lipid research
2001
2001-09
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
In situ hybridization study of obesity-associated alteration in growth hormone mRNA levels.
Male; Animals; Rats; *Gene Expression Regulation; Autoradiography; Growth Hormone/*genetics/metabolism; Nucleic Acid Hybridization; Obesity/*etiology/physiopathology; Pituitary Gland/*chemistry/metabolism; Northern; Blotting; RNA; Transcription; Genetic; Zucker; Messenger/*analysis
In order to investigate whether the impaired GH secretion associated with obesity is due to a pituitary disorder we studied GH mRNA levels by in situ hybridization in genetically obese and lean Zucker rats. The levels of GH mRNA were at least two fold lower in obese rats in comparison to that in lean controls as quantified by both the scanning of autoradiographs of tissue sections and Northern blot analysis. Quantification of somatotrophs revealed no significant difference in their number between lean and obese rat pituitaries. It is therefore likely that the attenuated GH mRNA levels in genetically obese Zucker rats are due to a decrease in GH transcripts per somatotroph rather than a result of a pituitary defect involving a preferential decrease in somatotroph population.
Ahmad I; Steggles A W; Finkelstein J A
International journal of obesity and related metabolic disorders : journal of the International Association for the Study of Obesity
1992
1992-06
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
PXR induces CYP27A1 and regulates cholesterol metabolism in the intestine.
*Lipid Metabolism; ATP Binding Cassette Transporter; ATP Binding Cassette Transporter 1; ATP-Binding Cassette Transporters/genetics; Base Sequence; Cell Line; Cholestanetriol 26-Monooxygenase/*metabolism; Cholesterol; Cholesterol/*metabolism; Fluorinated; Genes; Genetic/drug effects; Genetic/genetics; HDL/metabolism; Hepatocytes/drug effects/enzymology/metabolism; Humans; Hydrocarbons; Hydroxycholesterols/metabolism/pharmacology; Intestinal Mucosa/metabolism; Intestines/cytology/drug effects/enzymology; Member 1; Messenger/genetics/metabolism; Molecular Sequence Data; Pregnane X Receptor; Promoter Regions; Receptors; Reporter; Response Elements/genetics; Rifampin/pharmacology; RNA; Steroid/*metabolism; Subfamily G; Sulfonamides/pharmacology; Transcription; Up-Regulation/drug effects
Mitochondrial sterol 27-hydroxylase (CYP27A1) catalyzes oxidative cleavage of the sterol side chain in the bile acid biosynthetic pathway in the liver and
Li Tiangang; Chen Wenling; Chiang John Y L
Journal of lipid research
2007
2007-02
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1194/jlr.M600282-JLR200" target="_blank" rel="noreferrer noopener">10.1194/jlr.M600282-JLR200</a>
A putative role of micro RNA in regulation of cholesterol 7alpha-hydroxylase expression in human hepatocytes.
3' Untranslated Regions/genetics; Base Sequence; Chenodeoxycholic Acid/pharmacology; Cholesterol 7-alpha-Hydroxylase/*genetics; Enzymologic/drug effects/*genetics; Fibroblast Growth Factors/pharmacology; Gene Expression Regulation; Genetic/drug effects/genetics; Hep G2 Cells; Hepatocytes/drug effects/enzymology/*metabolism; Humans; Isoxazoles/pharmacology; MicroRNAs/*genetics/*metabolism; Oligonucleotide Array Sequence Analysis; Post-Transcriptional/drug effects/genetics; RNA Processing; Transcription
Cholesterol 7alpha-hydroxylase (CYP7A1) plays a critical role in regulation of bile acid synthesis in the liver. CYP7A1 mRNAs have very short half-lives, and bile acids destabilize CYP7A1 mRNA via the 3'-untranslated region (3'-UTR). However, the underlying mechanism of translational regulation of CYP7A1 mRNA remains unknown. Screening of a human micro RNA (miRNA) microarray has identified five differentially expressed miRNAs in human primary hepatocytes treated with chenodeoxycholic acid, GW4064, or fibroblast growth factor (FGF)19. These compounds also significantly induced the expression of miR-122a, a liver-specific and the predominant miRNA in human hepatocytes. The putative recognition sequences for miR-122a and miR-422a were localized in the 3'-UTR of human CYP7A1 mRNA. The miR-122a and miR-422a mimics inhibited, whereas their inhibitors stimulated CYP7A1 mRNA expression. These miRNAs specifically inhibited the activity of the CYP7A1-3'-UTR reporter plasmids, and mutations of miRNA binding sites in 3'-UTR abrogated miRNA inhibition of reporter activity. These results suggest that miR-122a and miR-422a may destabilize CYP7A1 mRNA to inhibit CYP7A1 expression. However, these miRNAs did not play a role in mediating FGF19 inhibition of CYP7A1 transcription. Under certain conditions, miRNA may reduce CYP7A1 mRNA stability to inhibit bile acid synthesis, and the miR-122a antagomirs may stimulate bile acid synthesis to reduce serum cholesterol and triglycerides.
Song Kwang-Hoon; Li Tiangang; Owsley Erika; Chiang John Y L
Journal of lipid research
2010
2010-08
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1194/jlr.M004531" target="_blank" rel="noreferrer noopener">10.1194/jlr.M004531</a>
Mechanism of rifampicin and pregnane X receptor inhibition of human cholesterol 7 alpha-hydroxylase gene transcription.
Bile Acids and Salts/pharmacology; Cholestasis/*physiopathology; Cholesterol 7-alpha-Hydroxylase/*biosynthesis/pharmacology; Cultured; Cytoplasmic and Nuclear/*physiology; DNA-Binding Proteins/pharmacology; Enzyme Inhibitors/*pharmacology; Genetic; Glucocorticoid; Hepatoblastoma/pathology; Hepatocyte Nuclear Factor 4; Humans; Liver Neoplasms/pathology; Messenger/analysis/biosynthesis; Phosphoproteins/pharmacology; Pregnane X Receptor; Receptors; Rifampin/*pharmacology; RNA; Steroid/*physiology; Transcription; Transcription Factors/pharmacology; Tumor Cells; Up-Regulation
Bile acids, steroids, and drugs activate steroid and xenobiotic receptor pregnane X receptor (PXR; NR1I2), which induces human cytochrome P4503A4 (CYP3A4) in drug metabolism and cholesterol 7 alpha-hydroxylase (CYP7A1) in bile acid synthesis in the liver. Rifampicin, a human PXR agonist, inhibits bile acid synthesis and has been used to treat cholestatic diseases. The objective of this study is to elucidate the mechanism by which PXR inhibits CYP7A1 gene transcription. The mRNA expression levels of CYP7A1 and several nuclear receptors known to regulate the CYP7A1 gene were assayed in human primary hepatocytes by quantitative real-time PCR (Q-PCR). Rifampicin reduced CYP7A1 and small heterodimer partner (SHP; NR02B) mRNA expression suggesting that SHP was not involved in PXR inhibition of CYP7A1. Rifampicin inhibited CYP7A1 reporter activity and a PXR binding site was localized to the bile acid response element-I. Mammalian two-hybrid assays revealed that PXR interacted with hepatic nuclear factor 4 alpha (HNF4 alpha, NR2A1) and rifampicin was required. Coimmunoprecipitation assay confirmed PXR interaction with HNF4 alpha. PXR also interacted with peroxisome proliferator-activated receptor gamma coactivator (PGC-1 alpha), which interacted with HNF4 alpha and induced CYP7A1 gene transcription. Rifampicin enhanced PXR interaction with HNF4 alpha and reduced PGC-1 alpha interaction with HNF4 alpha. Chromatin immunoprecipitation assay showed that PXR, HNF4 alpha, and PGC-1 alpha bound to CYP7A1 chromatin, and rifampicin dissociated PGC-1 alpha from chromatin. These results suggest that activation of PXR by rifampicin promotes PXR interaction with HNF4 alpha and blocks PGC-1 alpha activation with HNF4 alpha and results in inhibition of CYP7A1 gene transcription. Rifampicin inhibition of bile acid synthesis may be a protective mechanism against drug and bile acid-induced cholestasis.
Li Tiangang; Chiang John Y L
American journal of physiology. Gastrointestinal and liver physiology
2005
2005-01
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1152/ajpgi.00258.2004" target="_blank" rel="noreferrer noopener">10.1152/ajpgi.00258.2004</a>
Cytokine regulation of human sterol 12alpha-hydroxylase (CYP8B1) gene.
Cell Line; Chenodeoxycholic Acid/pharmacology; Chromosome Mapping; DNA-Binding Proteins/genetics/metabolism; Enzyme Inhibitors/pharmacology; Gene Expression Regulation/*drug effects; Genetic/drug effects; Genetic/physiology; Hepatocyte Nuclear Factor 4; Hepatocytes/metabolism; Humans; Interleukin-1/*pharmacology; MAP Kinase Signaling System/drug effects/physiology; Messenger/antagonists & inhibitors; Mitogen-Activated Protein Kinase 8/metabolism; Mitogen-Activated Protein Kinases/antagonists & inhibitors; Phosphoproteins/genetics/metabolism; Phosphorylation; Promoter Regions; Response Elements/genetics; RNA; Steroid 12-alpha-Hydroxylase/antagonists & inhibitors/*genetics; Transcription; Transcription Factors/genetics/metabolism
Sterol 12alpha-hydroxylase (CYP8B1) catalyzes cholic acid synthesis in the liver and is feedback inhibited by bile acids. In addition to activating farnesoid X receptor (nuclear receptor subfamily 1H4), bile acids also induce inflammatory cytokines in hepatocytes. The objective of this study was to investigate the mechanism by which inflammatory cytokines inhibit human CYP8B1 gene transcription. Real-time PCR assays revealed that both chenodeoxycholic acid (CDCA) and interleukin-1beta (IL-1beta) markedly reduced CYP8B1, cholesterol 7alpha-hydroxylase CYP7A1 and hepatic nuclear factor 4alpha (HNF4alpha) mRNA expression levels in human primary hepatocytes. However, CDCA induced, but
Jahan Asmeen; Chiang John Y L
American journal of physiology. Gastrointestinal and liver physiology
2005
2005-04
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1152/ajpgi.00207.2004" target="_blank" rel="noreferrer noopener">10.1152/ajpgi.00207.2004</a>
Trivalent chromium inhibits TSP-1 expression, proliferation, and O-GlcNAc signaling in vascular smooth muscle cells in response to high glucose in vitro.
Aorta/drug effects/metabolism; Cell Proliferation/*drug effects/genetics; Cells; Chromium/*pharmacology; Cultured; Fructosephosphates/metabolism; Genetic/drug effects/genetics; Glucose/*metabolism; Glutamine/genetics; Glycosylation/drug effects; Hexosamines/metabolism; Humans; Hyperglycemia/metabolism; Muscle; Myocytes; N-Acetylglucosaminyltransferases/genetics; O-glycosylation; Promoter Regions; reactive oxygen species; Reactive Oxygen Species/metabolism; Signal Transduction/*drug effects/genetics; Smooth; Smooth Muscle/*drug effects/metabolism; Thrombospondin 1/*antagonists & inhibitors/genetics; thrombospondin-1; Transcription; trivalent chromium; vascular smooth muscle cells; Vascular/*drug effects/metabolism
Trivalent chromium (Cr(3+)) is a mineral nutrient reported to have beneficial effects in glycemic and cardiovascular health. In vitro and in vivo studies suggest that Cr(3+) supplementation reduces the atherogenic potential and lowers the risk of vascular inflammation in diabetes. However, effects of Cr(3+) in vascular cells under conditions of hyperglycemia, characteristic of diabetes, remain unknown. In the present study we show that a therapeutically relevant concentration of Cr(3+) (100 nM) significantly downregulates a potent proatherogenic matricellular protein, thrombospondin-1 (TSP-1), in human aortic smooth muscle cells (HASMC) stimulated with high glucose in vitro. Promoter-reporter assays reveal that this downregulation of TSP-1 expression by Cr(3+) occurs at the level of transcription. The inhibitory effects of Cr(3+) on
Ganguly Rituparna; Sahu Soumyadip; Chavez Ronaldo J; Raman Priya
American journal of physiology. Cell physiology
2015
2015-01
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1152/ajpcell.00256.2014" target="_blank" rel="noreferrer noopener">10.1152/ajpcell.00256.2014</a>
Leptin augments recruitment of IRF-1 and CREB to thrombospondin-1 gene promoter in vascular smooth muscle cells in vitro.
*cAMP response element-binding protein; *interferon regulatory factor-1; *leptin; *thrombospondin-1; *transcription; *vascular smooth muscle cells; Binding Sites/genetics; Cells; Chromatin Immunoprecipitation/methods; Cultured; Cyclic AMP Response Element-Binding Protein/*metabolism; Gene Expression Regulation/genetics; Genetic/genetics; Humans; Interferon Regulatory Factor-1/*metabolism; Leptin/*metabolism; Muscle; Mutagenesis; Myocytes; Promoter Regions; Response Elements/genetics; Site-Directed/methods; Smooth; Smooth Muscle/*metabolism; Thrombospondin 1/*genetics/*metabolism; Transcription; Transcriptional Activation/genetics; Transfection/methods; Up-Regulation/genetics; Vascular/*metabolism
We previously reported that high pathophysiological concentrations of leptin, the adipocyte-secreted peptide, upregulate the expression of a potent proatherogenic matricellular protein, thrombospondin-1 (TSP-1), in vascular smooth muscle cells. Moreover, this regulation was found to occur at the level of transcription; however, the underlying molecular mechanisms remain unknown. The goal of the present study was to investigate the specific transcriptional mechanisms that mediate upregulation of TSP-1 expression by leptin. Primary human aortic smooth muscle cell cultures were transiently transfected with different TSP-1 gene (THBS1) promoter-linked luciferase reporter constructs, and luciferase activity in response to leptin (100 ng/ml) was assessed. We identified a long THBS1 promoter (-1270/+750) fragment with specific leptin response elements that are required for increased TSP-1 transcription by leptin. Promoter analyses, protein/DNA array and gel shift assays demonstrated activation and association of transcription factors, interferon regulatory factor-1 (IRF-1) and cAMP response element-binding protein (CREB), to the distal fragment of the THBS1 promoter in response to leptin. Supershift, chromatin immunoprecipitation, and coimmunoprecipitation assays revealed formation of a single complex between IRF-1 and CREB in response to leptin; importantly, recruitment of this complex to the THBS1 promoter mediated leptin-induced TSP-1 transcription. Finally, binding sequence decoy oligomer and site-directed mutagenesis revealed that regulatory elements for both IRF-1 (-1019 to -1016) and CREB (-1198 to -1195), specific to the distal THBS1 promoter, were required for leptin-induced TSP-1 transcription. Taken together, these findings demonstrate that leptin promotes a cooperative association between IRF-1 and CREB on the THBS1 promoter driving TSP-1 transcription in vascular smooth muscle cells.
Sahu Soumyadip; Ganguly Rituparna; Raman Priya
American journal of physiology. Cell physiology
2016
2016-08
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1152/ajpcell.00068.2016" target="_blank" rel="noreferrer noopener">10.1152/ajpcell.00068.2016</a>
Estrogen-related receptor gamma controls hepatic CB1 receptor-mediated CYP2E1 expression and oxidative liver injury by alcohol.
Alcohol-Induced Injury; Alcoholic Liver Disease; Alcoholic/genetics/*metabolism/prevention & control; Animals; Cannabinoid; CB1/*physiology; Cytochrome P-450 CYP2E1 Inhibitors; Cytochrome P-450 CYP2E1/genetics/*metabolism; Enzyme Inhibitors/pharmacology/therapeutic use; Enzymologic/drug effects/physiology; Estrogen/deficiency/genetics/*physiology; Ethanol/pharmacology; Gene Expression Profiling/methods; Gene Expression Regulation; Gene Regulation; Genetic/physiology; Inbred C57BL; Knockout; Liver; Liver Diseases; Liver Metabolism; Liver/metabolism; Male; Mice; Oxidation-Reduction; Oxidative Stress/physiology; Receptor; Receptors; Signal Transduction/drug effects/physiology; Tamoxifen/analogs & derivatives/pharmacology/therapeutic use; Transcription
BACKGROUND: The hepatic endocannabinoid system and cytochrome P450 2E1 (CYP2E1), a key enzyme causing alcohol-induced reactive oxygen species (ROS) generation, are major contributors to the pathogenesis of alcoholic liver disease. The nuclear hormone receptor oestrogen-related receptor gamma (ERRgamma) is a constitutively active transcriptional activator regulating gene expression. OBJECTIVE: To investigate the role of ERRgamma in the alcohol-mediated regulation of CYP2E1 and to examine the possibility to control alcohol-mediated oxidative stress and liver injury through an ERRgamma inverse agonist. DESIGN: For chronic alcoholic hepatosteatosis study, C57BL/6J wild-type and CB1(-/-) mice were administered alcohol for 4 weeks. GSK5182 and chlormethiazole (CMZ) were given by oral gavage for the last 2 weeks of alcohol feeding. Gene expression profiles and biochemical assays were performed using the liver or blood of mice. RESULTS: Hepatic ERRgamma gene expression induced by alcohol-mediated activation of CB1 receptor results in induction of CYP2E1, while liver-specific ablation of ERRgamma gene expression blocks alcohol-induced expression of CYP2E1 in mouse liver. An ERRgamma inverse agonist significantly ameliorates chronic alcohol-induced liver injury in mice through inhibition of CYP2E1-mediated generation of ROS, while inhibition of CYP2E1 by CMZ abrogates the beneficial effects of the inverse agonist. Finally, chronic alcohol-mediated ERRgamma and CYP2E1 gene expression, ROS generation and liver injury in normal mice were nearly abolished in CB1(-/-) mice. CONCLUSIONS: ERRgamma, as a previously unrecognised transcriptional regulator of hepatic CB1 receptor, controls alcohol-induced oxidative stress and liver injury through CYP2E1 induction, and its inverse agonist could ameliorate oxidative liver injury due to chronic alcohol exposure.
Kim Don-Kyu; Kim Yong-Hoon; Jang Hyun-Hee; Park Jinyoung; Kim Jung Ran; Koh Minseob; Jeong Won-Il; Koo Seung-Hoi; Park Tae-Sik; Yun Chul-Ho; Park Seung Bum; Chiang John Y L; Lee Chul-Ho; Choi Hueng-Sik
Gut
2013
2013-07
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1136/gutjnl-2012-303347" target="_blank" rel="noreferrer noopener">10.1136/gutjnl-2012-303347</a>
Farnesoid X Receptor Agonist Represses Cytochrome P450 2D6 Expression by Upregulating Small Heterodimer Partner.
Animals; Cytochrome P-450 CYP2D6/*biosynthesis; Cytochrome P-450 Enzyme Inhibitors/*pharmacology; Cytoplasmic and Nuclear/*agonists; Enzymologic/drug effects; Gene Expression Regulation; Genetic/drug effects/genetics; HEK293 Cells; Hepatocytes/drug effects/enzymology/metabolism; Humans; Isoxazoles/*pharmacology; Knockout; Mice; Receptors; Transcription; Transgenic
Cytochrome P450 2D6 (CYP2D6) is a major drug-metabolizing enzyme responsible for eliminating approximately 20% of marketed drugs. Studies have shown that differential transcriptional regulation of CYP2D6 may contribute to large interindividual variability in CYP2D6-mediated drug metabolism. However, the factors governing CYP2D6 transcription are largely unknown. We previously demonstrated small heterodimer partner (SHP) as a novel transcriptional repressor of CYP2D6 expression. SHP is a representative target gene of the farnesoid X receptor (FXR). The objective of this study is to investigate whether an agonist of FXR,
Pan Xian; Lee Yoon-Kwang; Jeong Hyunyoung
Drug metabolism and disposition: the biological fate of chemicals
2015
2015-07
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1124/dmd.115.064758" target="_blank" rel="noreferrer noopener">10.1124/dmd.115.064758</a>
Mechanism of vitamin D receptor inhibition of cholesterol 7alpha-hydroxylase gene transcription in human hepatocytes.
Base Sequence; Calcitriol/drug effects/genetics/*physiology; Cell Line; Cells; Cholesterol 7-alpha-Hydroxylase/*genetics; Cultured; DNA Primers; Electrophoretic Mobility Shift Assay; Gene Knockdown Techniques; Genetic/*physiology; Hepatocytes/*drug effects/enzymology; Humans; Immunoprecipitation; Lithocholic Acid/pharmacology; Messenger/genetics; Polymerase Chain Reaction; Receptors; RNA; Small Interfering; Transcription; Tumor; Two-Hybrid System Techniques
Lithocholic acid (LCA) is a potent endogenous vitamin D receptor (VDR) ligand. In cholestasis, LCA levels increase in the liver and intestine. The objective of this study is to test the hypothesis that VDR plays a role in inhibiting cholesterol 7alpha-hydroxylase (CYP7A1) gene expression and bile acid synthesis in human hepatocytes. Immunoblot analysis has detected VDR proteins in the nucleus of the human hepatoma cell line HepG2 and human primary hepatocytes. 1alpha, 25-Dihydroxy-vitamin D(3) or LCA acetate-activated VDR inhibited CYP7A1 mRNA expression and bile acid synthesis, whereas small interfering RNA to VDR completely abrogated VDR inhibition of CYP7A1 mRNA expression in HepG2 cells. Electrophoretic mobility shift assay and mutagenesis analyses have identified the negative VDR response elements that bind VDR/retinoid X receptor alpha in the human CYP7A1 promoter. Mammalian two-hybrid, coimmunoprecipitation, glutathione
Han Shuxin; Chiang John Y L
Drug metabolism and disposition: the biological fate of chemicals
2009
2009-03
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1124/dmd.108.025155" target="_blank" rel="noreferrer noopener">10.1124/dmd.108.025155</a>
A Prospero-related homeodomain protein is a novel co-regulator of hepatocyte nuclear factor 4alpha that regulates the cholesterol 7alpha-hydroxylase gene.
*Gene Expression Regulation; Aged; Amino Acid Motifs; Bile Acids and Salts/metabolism; Cell Line; Cell Nucleus/metabolism; Cells; Cholesterol 7-alpha-Hydroxylase/*chemistry/*genetics; Cultured/metabolism; Enzymologic; Female; Genes; Genetic; Gluconeogenesis; Glutathione Transferase/metabolism; Hepatocyte Nuclear Factor 4/metabolism/*physiology; Hepatocytes/metabolism; Homeodomain Proteins/metabolism/*physiology; Humans; Immunoprecipitation; Liver/metabolism; Luciferases/metabolism; Male; Messenger/metabolism; Middle Aged; Phosphoenolpyruvate Carboxykinase (ATP)/metabolism; Plasmids/metabolism; Protein Structure; Reporter; Response Elements; Reverse Transcriptase Polymerase Chain Reaction; RNA; Small Interfering/metabolism; Tertiary; Time Factors; Transcription; Transcriptional Activation; Transfection; Tumor Suppressor Proteins; Two-Hybrid System Techniques
Prox1, an early specific marker for developing liver and pancreas in foregut endoderm has recently been shown to interact with alpha-fetoprotein transcription factor and repress cholesterol 7alpha-hydroxylase (CYP7A1) gene transcription. Using a yeast two-hybrid assay, we found that Prox1 strongly and specifically interacted with hepatocyte nuclear factor (HNF)4alpha, an important transactivator of the human CYP7A1 gene in bile acid synthesis and phosphoenolpyruvate carboxykinase (PEPCK) gene in gluconeogenesis. A real time PCR assay detected Prox1 mRNA expression in human primary hepatocytes and HepG2 cells. Reporter assay, GST pull-down, co-immunoprecipitation, and yeast two-hybrid assays identified a specific interaction between the N-terminal LXXLL motif of Prox1 and the activation function 2 domain of HNF4alpha. Prox1 strongly inhibited HNF4alpha and peroxisome proliferators-activated receptor gamma coactivator-1alpha co-activation of the CYP7A1 and PEPCK genes. Knock down of the endogenous Prox1 by small interfering RNA resulted in significant increase of CYP7A1 and PEPCK mRNA expression and the rate of bile acid synthesis in HepG2 cells. These results suggest that Prox1 is a novel co-regulator of HNF4alpha that may play a key role in the regulation of bile acid synthesis and gluconeogenesis in the liver.
Song Kwang-Hoon; Li Tiangang; Chiang John Y L
The Journal of biological chemistry
2006
2006-04
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1074/jbc.M513420200" target="_blank" rel="noreferrer noopener">10.1074/jbc.M513420200</a>
Bcl-2 positively regulates Sox9-dependent chondrocyte gene expression by suppressing the MEK-ERK1/2 signaling pathway.
*Gene Expression Regulation; Adenoviridae/genetics; Animals; Apoptosis; beta-Galactosidase/metabolism; Blotting; Butadienes/pharmacology; Caspase Inhibitors; Cell Differentiation; Cell Line; Chondrocytes/*metabolism; Collagen Type II/metabolism; Down-Regulation; Enzyme Inhibitors/pharmacology; Fibroblasts/metabolism; Fluorescence; Genetic; High Mobility Group Proteins/*metabolism; Lac Operon; Luciferases/metabolism; MAP Kinase Kinase Kinases/*metabolism; Messenger/metabolism; Microscopy; Mitogen-Activated Protein Kinase 1/*metabolism; Mitogen-Activated Protein Kinase 3/*metabolism; NF-kappa B/metabolism; Nitriles/pharmacology; Phenotype; Phosphorylation; Promoter Regions; Protein Kinase C-alpha; Protein Kinase C/antagonists & inhibitors; Proteoglycans/metabolism; Proto-Oncogene Proteins c-bcl-2/*metabolism; Rats; Reverse Transcriptase Polymerase Chain Reaction; RNA; Signal Transduction; Small Interfering/metabolism; SOX9 Transcription Factor; Sprague-Dawley; Time Factors; Transcription; Transcription Factors/*metabolism; Transfection; Western
Bcl-2 is an anti-apoptotic protein that has recently been shown to regulate other cellular functions. We previously reported that Bcl-2 regulates chondrocyte matrix gene expression, independent of its anti-apoptotic function. Here, we further investigate this novel function of Bcl-2 and examine three intracellular signaling pathways likely to be associated with this function. The present study demonstrates that the activity of Sox9, a master transcription factor that regulates the gene expression of chondrocyte matrix proteins, is suppressed by Bcl-2 small interference RNA in the presence of caspase inhibitors. This effect was attenuated by prior exposure of chondrocytes to an adenoviral vector expressing sense Bcl-2. In addition, the down-regulation of Bcl-2, Sox9, and chondrocyte-specific gene expression by serum withdrawal in primary chondrocytes was reversed by expressing Bcl-2. Inhibition of the protein kinase C alpha and NFkappaB pathways had no effect on the maintenance of Sox9-dependent gene expression by Bcl-2. In contrast, whereas the MEK-ERK1/2 pathway negatively regulated the differentiated phenotype in wild type chondrocytes, inhibition of this pathway reversed the loss of differentiation markers and fibroblastic phenotype in Bcl-2-deficient chondrocytes. In conclusion, the present study identifies a specific signaling pathway, namely, MEK-ERK1/2, that is downstream of Bcl-2 in the regulation of Sox9-dependent chondrocyte gene expression and phenotype.
Yagi Rieko; McBurney Denise; Horton Walter E Jr
The Journal of biological chemistry
2005
2005-08
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1074/jbc.M502751200" target="_blank" rel="noreferrer noopener">10.1074/jbc.M502751200</a>
A novel role of transforming growth factor beta1 in transcriptional repression of human cholesterol 7alpha-hydroxylase gene.
Bile Acids and Salts/metabolism; Carcinoma; Cell Line; Cells; Cholesterol 7-alpha-Hydroxylase/*genetics/*metabolism; Cultured; Enzyme Inhibitors/pharmacology; Genetic/drug effects/*physiology; Hepatocellular/*metabolism/pathology; Hepatocyte Nuclear Factor 4/metabolism; Hepatocytes/drug effects/*metabolism/pathology; Humans; Hydroxamic Acids/pharmacology; Liver Neoplasms/*metabolism/pathology; Messenger/metabolism; RNA; Signal Transduction/physiology; Smad3 Protein/metabolism; Transcription; Transforming Growth Factor beta1/*metabolism; Tumor
BACKGROUND & AIMS: Inhibition of cholesterol 7alpha-hydroxylase (CYP7A1) by bile acids and inflammatory cytokines provides an important mechanism to protect hepatocytes from bile acid toxicity during cholestasis. Transforming growth factor beta1 (TGFbeta1) released by hepatic stellate cells during chronic liver injury plays a critical role in liver inflammation and fibrogenesis. The objective of this study is to investigate the role of TGFbeta1 in hepatic bile acid synthesis. METHODS: mRNA expressions in primary human hepatocytes and HepG2 cells were measured by quantitative real-time polymerase chain reaction. Reporter assay, glutathione-S-transferase pull-down assay, adenovirus-mediated gene transduction, and chromatin immunoprecipitation assay were used to study the mechanism of TGFbeta1 regulation of CYP7A1 gene transcription. RESULTS: TGFbeta1 inhibited the mRNA expression of CYP7A1 and bile acid synthesis in HepG2 cells and primary human hepatocytes. Mothers against decapentaplegic homolog (Smad3) inhibited both CYP7A1 promoter activity and mRNA expression by inhibiting
Li Tiangang; Chiang John Y L
Gastroenterology
2007
2007-11
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1053/j.gastro.2007.08.042" target="_blank" rel="noreferrer noopener">10.1053/j.gastro.2007.08.042</a>
Orphan nuclear receptor oestrogen-related receptor gamma (ERRgamma) plays a key role in hepatic cannabinoid receptor type 1-mediated induction of CYP7A1 gene expression.
Animals; bile acid; Bile Acids and Salts/metabolism; Cannabinoid; cannabinoid receptors; CB1/agonists/genetics/*metabolism; Cells; Cholesterol 7-alpha-Hydroxylase/*biosynthesis/genetics; cholesterol 7alpha-hydroxylase (CYP7A1); Cultured; Cytoplasmic and Nuclear/metabolism; Drug Inverse Agonism; Estrogen/genetics/*metabolism; Ethanol/pharmacology; Gene Expression; Genetic; Glycerides/pharmacology; GSK5182; HEK293 Cells; Hepatocytes/metabolism; Humans; Inbred C57BL; Knockout; Liver/*metabolism; Mice; oestrogen-related receptor gamma (ERRgamma); orphan nuclear receptor; Promoter Regions; Rats; Receptor; Receptors; small heterodimer partner (SHP); Sprague-Dawley; Transcription
Bile acids are primarily synthesized from cholesterol in the liver and have important roles in dietary lipid absorption and cholesterol homoeostasis. Detailed roles of the orphan nuclear receptors regulating cholesterol 7alpha-hydroxylase (CYP7A1), the rate-limiting enzyme in bile acid synthesis, have not yet been fully elucidated. In the present study, we report that oestrogen-related receptor gamma (ERRgamma) is a novel transcriptional regulator of CYP7A1 expression. Activation of cannabinoid receptor type 1 (CB1 receptor) signalling induced ERRgamma-mediated transcription of the CYP7A1 gene. Overexpression of ERRgamma increased CYP7A1 expression in vitro and in vivo, whereas knockdown of ERRgamma attenuated CYP7A1 expression. Deletion analysis of the CYP7A1 gene promoter and a ChIP assay revealed an ERRgamma-binding site on the CYP7A1 gene promoter. Small heterodimer partner (SHP) inhibited the transcriptional activity of ERRgamma and thus regulated CYP7A1 expression. Overexpression of ERRgamma led to increased bile acid levels, whereas an inverse agonist of ERRgamma, GSK5182, reduced CYP7A1 expression and bile acid synthesis. Finally, GSK5182 significantly reduced hepatic CB1 receptor-mediated induction of CYP7A1 expression and bile acid synthesis in alcohol-treated mice. These results provide the molecular mechanism linking ERRgamma and bile acid metabolism.
Zhang Yaochen; Kim Don-Kyu; Lee Ji-Min; Park Seung Bum; Jeong Won-Il; Kim Seong Heon; Lee In-Kyu; Lee Chul-Ho; Chiang John Y L; Choi Hueng-Sik
The Biochemical journal
2015
2015-09
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1042/BJ20141494" target="_blank" rel="noreferrer noopener">10.1042/BJ20141494</a>
Sensory nerves and neuropeptides in uterine cervical ripening.
Animal; Animals; Calcitonin Gene-Related Peptide/biosynthesis; Capsaicin/pharmacology; Cervical Ripening/*metabolism; Cervix Uteri/*innervation/*metabolism; Complementary/metabolism; Female; Genetic; Immunohistochemistry; Labor; Messenger/metabolism; Neurokinin-1/biosynthesis; Neurons/metabolism; Neuropeptides/*biosynthesis; Nitric Oxide Synthase/biosynthesis; Obstetric; Plasmids/metabolism; Postpartum Period; Pregnancy; Rats; Receptors; RNA; Secretogranin II; Sprague-Dawley; Substance P/biosynthesis; Transcription
At the time of parturition (fetal delivery) the uterine cervix must "ripen," becoming soft, pliable, and dilated to accommodate the fetus' delivery. The fundamental processes underlying cervical ripening remain poorly understood. Knowledge that abundant autonomic and sensory nerves supply the uterine cervix, that transection of afferent nerves supplying the cervix blocks parturition, and that some of the changes in the cervix resemble those seen in inflammatory reactions suggests nerves may have a role in the cervical ripening changes. The present study utilized immunohistochemistry, plasma extravasation, and solution hybridization-nuclease protection assay to elucidate the complement of primary afferent nerves and some receptors in the rat cervix during pregnancy, and to determine if they may have roles in the ripening process at term. This study revealed an abundance of nerves associated with the cervical vasculature and myometrial smooth muscle containing immunoreactivity for substance P, calcitonin gene-related peptide, secretoneurin, and nitric oxide synthase throughout pregnancy. Many of these are small unmyelinated capsaicin-sensitive C-fibers. Substance P- (NK1-) and calcitonin gene-related peptide receptors were apparent on uterine cervix vasculature from pregnant, parturient, and postpartum rats. NK1 receptor mRNA was maximal at 20 days of pregnancy. Plasma extravasation of i.v. administered Evans Blue or Monastral Blue was most pronounced at parturition (shortly after NK1 mRNA is maximal); this was similar to plasma extravasation evoked by i.v. administration of substance P or capsaicin-treatment. This study revealed new data about the nervous system of the rat uterine cervix and that these nerves and their transmitters could very well be part of a neurogenic inflammatory process involved in cervical ripening.
Collins J J; Usip S; McCarson K E; Papka R E
Peptides
2002
2002-01
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1016/s0196-9781(01)00593-9" target="_blank" rel="noreferrer noopener">10.1016/s0196-9781(01)00593-9</a>
Interleukin IL-12 blocks a specific subset of the transcriptional profile responsive to UVB in epidermal keratinocytes.
*Keratinocytes/metabolism/radiation effects; *Ultraviolet Rays; Cell Culture Techniques; Cells; Cultured; Epidermal Cells; Gene Expression Regulation/drug effects; Genetic/*radiation effects; Humans; Interleukin-12/*genetics/*metabolism/pharmacology; Transcription
Interleukin-12 (IL-12) is a proinflammatory and immunomodulatory cytokine that plays a critical role it in innate and adaptive immunity by inducing production of interferon-gamma and other cytokines. IL-12 was shown to block the ultraviolet light-induced immunosuppression, important in cancer immunosurveillance, cutaneous allergies and inflammation. To characterize the molecular effects of
Molenda Matthew; Mukkamala Lakshmi; Blumenberg Miroslav
Molecular immunology
2006
2006-05
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1016/j.molimm.2005.12.003" target="_blank" rel="noreferrer noopener">10.1016/j.molimm.2005.12.003</a>
APeg3, a novel paternally expressed gene 3 antisense RNA transcript specifically expressed in vasopressinergic magnocellular neurons in the rat supraoptic nucleus.
3' Untranslated Regions/genetics; Amino Acid; Animals; Antisense/*genetics/isolation & purification/*metabolism; Conserved Sequence/genetics; DNA-Binding Proteins/genetics/*metabolism; Gene Expression Regulation/*genetics; Genetic/genetics; Genomic Imprinting/genetics; Hypothalamo-Hypophyseal System/cytology/metabolism; Kruppel-Like Transcription Factors; Male; Messenger/genetics/metabolism; Molecular Sequence Data; Neurons/*metabolism; Nucleic Acid; Protein Kinases/genetics/*metabolism; Rats; RNA; Sequence Homology; Sprague-Dawley; Supraoptic Nucleus/cytology/*metabolism; Transcription; Transcription Factors/genetics/*metabolism; Vasopressins/*metabolism; Water-Electrolyte Balance/genetics
Vasopressin (VP) and oxytocin (OT) play critical roles in the regulation of salt and water balance, lactation, and various behaviors and are expressed at very high levels in specific magnocellular neurons (MCNs) in the hypothalamo-neurohypophysial system (HNS). In addition to the cell-specific expression of the VP and OT genes in these cells, there are other transcripts that are preferentially expressed in the VP or OT MCNs. One such gene, paternally expressed gene 3 (Peg3), is an imprinted gene expressed exclusively from the paternal allele that encodes a Kruppel-type zinc finger-containing protein involved in maternal behavior and is abundantly expressed in the VP-MCNs. We report here the robust expression in the VP-MCNs of an RNA, which we designate APeg3 that is transcribed in the antisense direction to the 3' untranslated region of the Peg3 gene. The APeg3 mRNA is about 1 kb in size, and the full-length sequence of APeg3, as determined by 5' and 3' RACE, contains an open reading frame that predicts a protein of 93 amino acids and is predominantly expressed in VP-MCNs. Both Peg3 and APeg3 gene expression in the VP-MCNs increase during systemic hyperosmolality in vivo, demonstrating that both of these genes are osmoregulated.
Glasgow Eric; Ryu Seung-Lim; Yamashita Mitsuo; Zhang Bing-Jun; Mutsuga Noriko; Gainer Harold
Brain research. Molecular brain research
2005
2005-06
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1016/j.molbrainres.2005.02.030" target="_blank" rel="noreferrer noopener">10.1016/j.molbrainres.2005.02.030</a>
Impact of circadian nuclear receptor REV-ERBalpha on midbrain dopamine production and mood regulation.
*Affect; *Circadian Rhythm; Animals; Bipolar Disorder/genetics; CLOCK Proteins/genetics/metabolism; Cytoplasmic and Nuclear/genetics/*metabolism; Dopamine/*metabolism; Genetic; Group A; Histones/metabolism; Humans; Inbred C57BL; Knockout; Male; Member 2/metabolism; Mesencephalon/*metabolism; Mice; Mood Disorders/genetics/metabolism; Nuclear Receptor Subfamily 4; Rats; Receptors; Repressor Proteins/genetics/*metabolism; Transcription; Tyrosine 3-Monooxygenase/genetics
The circadian nature of mood and its dysfunction in affective disorders is well recognized, but the underlying molecular mechanisms are still unclear. Here, we show that the circadian nuclear receptor REV-ERBalpha, which is associated with bipolar disorder, impacts midbrain dopamine production and mood-related behavior in mice. Genetic deletion of the Rev-erbalpha gene or pharmacological inhibition of REV-ERBalpha activity in the ventral midbrain induced mania-like behavior in association with a central hyperdopaminergic state. Also, REV-ERBalpha repressed tyrosine hydroxylase (TH) gene transcription via competition with nuclear receptor-related 1 protein (NURR1), another nuclear receptor crucial for dopaminergic neuronal function, thereby driving circadian TH expression through a target-dependent antagonistic mechanism. In conclusion, we identified a molecular connection between the circadian timing system and mood regulation, suggesting that REV-ERBalpha could be targeting in the treatment of circadian rhythm-related affective disorders.
Chung Sooyoung; Lee Eun Jeong; Yun Seongsik; Choe Han Kyoung; Park Seong-Beom; Son Hyo Jin; Kim Kwang-Soo; Dluzen Dean E; Lee Inah; Hwang Onyou; Son Gi Hoon; Kim Kyungjin
Cell
2014
2014-05
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1016/j.cell.2014.03.039" target="_blank" rel="noreferrer noopener">10.1016/j.cell.2014.03.039</a>
Transcriptional regulation of human oxysterol 7alpha-hydroxylase by sterol response element binding protein.
*Transcription Factors; Base Sequence; CCAAT-Enhancer-Binding Proteins/*metabolism; Cytochrome P-450 Enzyme System/*genetics/physiology; Cytochrome P450 Family 7; DNA-Binding Proteins/*metabolism; Enzyme Repression; Genetic; Humans; Molecular Sequence Data; Mutagenesis; Promoter Regions; Response Elements; Sp1 Transcription Factor/antagonists & inhibitors; Steroid Hydroxylases/*genetics/physiology; Sterol Regulatory Element Binding Protein 1; Sterols/metabolism; Transcription
Oxysterol 7alpha-hydroxylase (CYP7B1) metabolizes oxysterols, potent regulators of lipid homeostasis. Very little is known about transcriptional regulation of human CYP7B1. The present results indicate that sterol response element binding protein (SREBP), a family of oxysterol-responsive transcription factors that stimulates cholesterol synthesis, may be an important regulator of CYP7B1. SREBP suppressed a human CYP7B1 luciferase reporter gene in several cell lines, most markedly in rat hepatoma McA-RH7777 cells. An SREBP-1-responsive region was mapped to a GC-rich sequence in the proximal CYP7B1 promoter, containing binding sites for the basal transcriptional activator Sp1. Mutagenesis of this sequence abolished SREBP-1-mediated suppression. Data indicated that SREBP does not bind this sequence but affects the gene indirectly, probably via interaction with Sp1. Our findings indicate that CYP7B1 transcription is controlled by SREBP and reveal a link between oxysterol-sensitive regulators and oxysterol metabolism. We propose that CYP7B1 is important for regulating cellular sterol content and protects against oxysterol-mediated toxicity.
Norlin Maria; Chiang John Y L
Biochemical and biophysical research communications
2004
2004-03
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1016/j.bbrc.2004.02.029" target="_blank" rel="noreferrer noopener">10.1016/j.bbrc.2004.02.029</a>
Serotonergic involvement in the regulation of prolactin and vasoactive intestinal peptide mRNA expression in the rat anterior pituitary.
5-Hydroxytryptophan/pharmacology; alpha-Methyltyrosine; Animals; Anterior/*chemistry/drug effects/metabolism; Blotting; Bromocriptine/pharmacology; Gene Expression Regulation/drug effects/physiology; Genetic; Haloperidol/pharmacology; Ketanserin/pharmacology; Male; Messenger/*analysis/genetics; Methiothepin/pharmacology; Methoxydimethyltryptamines/pharmacology; Methyltyrosines/pharmacology; Northern; Pituitary Gland; Prolactin/analysis/*genetics/metabolism; Quipazine/pharmacology; Rats; RNA; Serotonin/*physiology; Sprague-Dawley; Time Factors; Transcription; Vasoactive Intestinal Peptide/analysis/*genetics/metabolism
These studies examined the contribution of serotonin (5-HT) to the control of prolactin (PRL) and vasoactive intestinal peptide (VIP) messenger RNA expression in rat anterior pituitary. Daily injection of rats with the biosynthetic precursor to serotonin, 5-hydroxytryptophan (5-HTP; 25 mg/kg, q.i.d.), resulted on day 5 in a 50% increase in the expression of PRL mRNA in the pituitary while at the same time reducing the levels of both the 1.0 and 1.7 kb VIP mRNA transcripts. Co-treatment of rats with 5-HTP plus the catecholamine biosynthesis inhibitor, alpha-methyl-tyrosine (alpha-MT; 150 mg/kg, q.d. x 2 days), or the dopamine receptor antagonist haloperidol (1.25 mg/kg, b.i.d. x 5 days), resulted in increases in pituitary PRL message levels that were greater than those observed with either anti-dopaminergic agent alone. In contrast, 5-HTP was unable to reverse the inhibition of PRL mRNA expression caused by treatment with the dopamine receptor agonist bromocriptine (2.5 mg/kg, b.i.d. x 5 days). Neither alpha-MT, haloperidol nor bromocriptine had a significant effect on pituitary VIP mRNA expression. Administration of the direct-acting 5-HT receptor agonist quipazine (5 mg/kg, b.i.d.) for 14 consecutive days caused a significant increase in pituitary PRL mRNA levels on day 1 and reached a plateau of 90% above control levels on days 7 and 14. VIP mRNA levels rose significantly on day 1 of quipazine treatment but thereafter fell to a minimum of 22% (1.0 kb) and 52% (1.7 kb) of control by day 14.(ABSTRACT TRUNCATED AT 250 WORDS)
Signs S A; Liu B; Wolford J; Carrillo A J
Molecular and cellular endocrinology
1994
1994-11
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1016/0303-7207(94)90168-6" target="_blank" rel="noreferrer noopener">10.1016/0303-7207(94)90168-6</a>
The isolation and characterization of the bovine cytochrome b5 gene, and a transcribed pseudogene.
*Pseudogenes; Amino Acid Sequence; Animals; Base Sequence; Blotting; Cattle/*genetics; Cytochromes b5/*genetics; DNA/genetics/isolation & purification; Exons; Gene Expression; Genetic; Genomic Library; Humans; Introns; Messenger/biosynthesis/metabolism; Molecular Sequence Data; Nucleic Acid; Nucleic Acid Conformation; Oligodeoxyribonucleotides; Rabbits; Restriction Mapping; Reticulocytes/metabolism; RNA; RNA Precursors/chemistry/metabolism; Sequence Homology; Southern; Transcription
This is the first isolation and characterization of a cytochrome b5(b5) gene. The bovine b5 gene is quite large, spanning about 28 kb and contains six exons. One of these exons appears to code for a reticulocyte-specific sequence similar to that described for human and rabbit b5. All of the splicing junctions conform to the GT-AG consensus rule. The 5' flanking sequence has no obvious TATA box, two CAAT boxes, and contains several G:C-rich SpI motifs indicative of a house-keeping gene. In reticulocyte mRNA we found evidence for a transcribed b5 pseudogene, but could not detect sequences coding for the soluble form of b5. We conclude that the soluble form of b5 is derived from the membrane-bound b5 by a post-translational mechanism.
Cristiano R J; Giordano S J; Steggles A W
Genomics
1993
1993-08
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1006/geno.1993.1331" target="_blank" rel="noreferrer noopener">10.1006/geno.1993.1331</a>
Effects of bile acids and steroid/thyroid hormones on the expression of cholesterol 7 alpha-hydroxylase mRNA and the CYP7 gene in HepG2 cells.
Animals; Bile Acids and Salts/*pharmacology; Cells; Cholesterol 7-alpha-Hydroxylase/*genetics; Cultured; Enzymologic/*drug effects; Gene Expression Regulation; Genetic; Genetic/genetics; Humans; Luciferases/biosynthesis/genetics; Messenger/*biosynthesis; Promoter Regions; Rats; Recombinant Fusion Proteins/biosynthesis; RNA; Steroids/*pharmacology; Thyroid Hormones/*pharmacology; Transcription; Transfection
The expression of cholesterol 7 alpha-hydroxylase mRNA levels in confluent HepG2 cultures was reduced by tauro- or glyco-conjugates of deoxycholate and chenodeoxycholate, but not by cholate. Ursodeoxycholates, on the other hand, stimulated the mRNA level. The 5'-upstream regions of rat cholesterol 7 alpha-hydroxylase gene (CYP7) were fused to luciferase reporter gene and the constructs, p-3616/Luc, p-224/Luc and p-160/Luc, were transiently transfected into HepG2 cells. Tauro-conjugates of deoxycholate and chenodeoxycholate inhibited the transcriptional activities of the gene constructs in the confluent cells, but not in subconfluent cells. These results reveal that bile acid responsive elements are located in the -160 fragment and also between nt -3616 and -224. Thyroid and steroid hormones stimulated transcriptional activity expressed in the confluent cells and their responsive elements are located upstream of nt -224. It appears that adult phenotypes are responsible for bile acid feedback and hormone response in HepG2 cells.
Crestani M; Karam W G; Chiang J Y
Biochemical and biophysical research communications
1994
1994-01
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1006/bbrc.1994.1080" target="_blank" rel="noreferrer noopener">10.1006/bbrc.1994.1080</a>
Promoter activity and regulation of the CYP4F2 leukotriene B(4) omega-hydroxylase gene by peroxisomal proliferators and retinoic acid in HepG2 cells.
*Gene Expression Regulation; *Promoter Regions; Amino Acid Sequence; Base Sequence; Cell Line; Cloning; Cytochrome P-450 CYP4A; Cytochrome P-450 Enzyme System/*genetics/metabolism; Cytochrome P450 Family 4; Cytoplasmic and Nuclear/metabolism; DNA; DNA Footprinting; Enzymologic; Exons; Genes; Genetic; Genetic/drug effects; Humans; Introns; Leukotriene B4/metabolism; Mixed Function Oxygenases/metabolism; Models; Molecular; Molecular Sequence Data; Peroxisome Proliferators/*metabolism; Receptors; Reporter; Retinoic Acid Receptor alpha; Retinoic Acid/metabolism; Sequence Analysis; Transcription; Transcription Factors/metabolism; Transfection; Tretinoin/*metabolism
The human liver CYP4F2 gene (Accession No. AF221943) encodes a leukotriene B(4) omega-hydroxylase that metabolizes leukotriene B(4) (LTB(4)) to a less potent proinflammatory eicosanoid, 20-OH-LTB(4). We sequenced a 6.7-kb genomic fragment of the human CYP4F2 gene that has the first five exons and 500 bp of the 5'-flanking region. The major transcription start site was found to be 49 bp upstream of the 3' end of exon 1 and the ATG translation initiation codon was located in exon 2. Besides the TATA box at -39 bp and basal transcription factor binding sites, the promoter region and 412-bp intron 1 have several putative binding sites for nuclear factors that may mediate the inflammatory response and lipid homeostasis. We found two DR1 elements in the 5' promoter, a DR2 element in intron 1, and RXR/RAR binding sites in both intron 1 and the 5' promoter. DNase I footprinting revealed three protected sequences, with the region containing two CAATT boxes at -71 and -111 bp important in CYP4F2 gene expression. Luciferase reporter assays showed that the 500-bp upstream sequence has strong promoter activity. Transient transfection experiments identified two sites in the 5' promoter and intron 1 that cooperate in gene transcription while exon 1 and a
Zhang X; Chen L; Hardwick J P
Archives of biochemistry and biophysics
2000
2000-06
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1006/abbi.2000.1836" target="_blank" rel="noreferrer noopener">10.1006/abbi.2000.1836</a>
A plant lignan, 3'-O-methyl-nordihydroguaiaretic acid, suppresses papillomavirus E6 protein function, stabilizes p53 protein, and induces apoptosis in cervical tumor cells.
Apoptosis Regulatory Proteins/metabolism; Apoptosis/*drug effects; bcl-2-Associated X Protein/metabolism; Caspase 3/metabolism; Caspase 9/metabolism; Cultured/drug effects; Enzyme Activation/drug effects; Female; Genetic; Humans; Luciferases; Masoprocol/*analogs & derivatives/therapeutic use; Oncogene Proteins; Plants/chemistry; Proto-Oncogene Proteins/metabolism; Repressor Proteins/*metabolism; Transcription; Tumor Cells; Tumor Suppressor Protein p53/genetics/*metabolism; Uterine Cervical Neoplasms/*drug therapy/metabolism/pathology; Viral/*metabolism
Persistent infection with oncogenic human papillomaviruses (HPVs) is the most important factor in the induction of uterine cervical cancer, a leading cause of cancer mortality in women worldwide. Upon cell transformation, continual expression of the viral oncogenes is required to maintain the transformed phenotype. The viral E6 protein forms a ternary complex with the cellular E6-AP protein and p53 protein which promotes the rapid degradation of p53. Recent studies have revealed that lignans from the creosote bush (3'-O-methyl-nordihydroguaiaretic acid) can repress the viral promoter responsible for E6 gene expression. Work reported here shows that the lignan can subvert viral oncogene function resulting in stabilized p53 protein within treated HPV-containing tumor cells. The stabilized p53 is transcriptionally active as demonstrated by a luciferase reporter vector and induction of genes for Bax and PUMA proteins. Apoptosis is detected by annexin V binding to treated cells as analyzed by flow cytometry. Programmed cell death is confirmed by the induction of active caspases and TUNEL assay. Initiator caspase-9 is activated first, followed later by the effector caspase-3 enzyme. The stabilization and induced apoptosis are not observed within treated HPV-negative cervical tumor cells. Quantitative real time RT-PCR analysis of endogenous E6 gene transcription from the integrated HPV 16 promoter shows at least a fivefold repression of expression as compared to untreated cells. These results indicate that the loss of E6 protein in treated cells could be, in part, responsible for the stabilization of p53 within the lignan treated cells.
Allen Kristi L; Tschantz Deidra R; Awad Keytam S; Lynch William P; DeLucia Angelo L
Molecular carcinogenesis
2007
2007-07
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1002/mc.20305" target="_blank" rel="noreferrer noopener">10.1002/mc.20305</a>
All-trans-retinoic acid ameliorates hepatic steatosis in mice by a novel transcriptional cascade.
Animals; Basic Helix-Loop-Helix Transcription Factors/genetics; Blood Glucose/analysis; Cytoplasmic and Nuclear/*physiology; Fatty Liver/*drug therapy/metabolism; Gene Expression Regulation; Genetic; Inbred C57BL; Lipid Metabolism; Liver/metabolism; Male; Mice; Non-alcoholic Fatty Liver Disease; PPAR gamma/*genetics; Receptors; Repressor Proteins/genetics; Retinoic Acid Receptor alpha; Retinoic Acid/physiology; Transcription; Tretinoin/pharmacology/*therapeutic use
UNLABELLED: Mice deficient in small heterodimer partner (SHP) are protected from diet-induced hepatic steatosis resulting from increased fatty acid oxidation and decreased lipogenesis. The decreased lipogenesis appears to be a direct consequence of very low expression of peroxisome proliferator-activated receptor gamma 2 (PPAR-gamma2), a potent lipogenic transcription factor, in the SHP(-/-) liver. The current study focused on the identification of a SHP-dependent regulatory cascade that controls PPAR-gamma2 gene expression, thereby regulating hepatic fat accumulation. Illumina BeadChip array (Illumina, Inc., San Diego, CA) and real-time polymerase chain reaction were used to identify genes responsible for the linkage between SHP and PPAR-gamma2 using hepatic RNAs isolated from SHP(-/-) and SHP-overexpressing mice. The initial efforts identify that hairy and enhancer of split 6 (Hes6), a novel transcriptional repressor, is an important mediator of the regulation of PPAR-gamma2 transcription by SHP. The Hes6 promoter is specifically activated by the retinoic acid receptor (RAR) in response to its natural agonist ligand, all-trans retinoic acid (atRA), and is repressed by SHP. Hes6 subsequently represses hepatocyte nuclear factor 4 alpha (HNF-4alpha)-activated PPAR-gamma2 gene expression by direct inhibition of
Kim Seong-Chul; Kim Chun-Ki; Axe David; Cook Aaron; Lee Mikang; Li Tiangang; Smallwood Nicole; Chiang John Y L; Hardwick James P; Moore David D; Lee Yoon-Kwang
Hepatology (Baltimore, Md.)
2014
2014-05
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1002/hep.26699" target="_blank" rel="noreferrer noopener">10.1002/hep.26699</a>
Bile acids and cytokines inhibit the human cholesterol 7 alpha-hydroxylase gene via the JNK/c-jun pathway in human liver cells.
Bile Acids and Salts/*metabolism; Cells; Chenodeoxycholic Acid/pharmacology; Cholesterol 7-alpha-Hydroxylase/*genetics/metabolism; Cultured; Cytokines/*metabolism; Gene Expression Regulation; Genetic; Hepatocytes/*cytology/drug effects; Humans; Immunoblotting; In Vitro Techniques; Interleukin-1/pharmacology; Messenger/analysis; Probability; Proto-Oncogene Proteins c-jun/*metabolism; Reverse Transcriptase Polymerase Chain Reaction; RNA; Sensitivity and Specificity; Signal Transduction/genetics; Transcription
Cholesterol 7 alpha-hydroxylase (CYP7A1) of the bile acid biosynthesis pathway is suppressed by bile acids and inflammatory cytokines. Bile acids are known to induce inflammatory cytokines to activate the mitogen-activated protein kinase/c-Jun N-terminal kinase (JNK) signaling pathway that inhibits CYP7A1 gene transcription. c-Jun has been postulated to mediate bile acid inhibition of CYP7A1. However, the c-Jun target involved in the regulation of CYP7A1 is unknown. Human primary hepatocytes and HepG2 cells were used as models to study chenodeoxycholic acid (CDCA) and interleukin-1 beta (IL-1 beta) regulation of human CYP7A1 gene expression via real-time polymerase chain reaction, reporter assays, co-immunoprecipitation and chromatin immunocipitation (ChIP) assays. IL-1 beta and CDCA reduced CYP7A1 but induced c-Jun messenger RNA expression in human primary hepatocytes. IL-1beta inhibited human CYP7A1 reporter activity via the HNF4 alpha binding site. A JNK-specific inhibitor blocked the inhibitory effect of IL-1 beta on HNF4 alpha expression and CYP7A1 reporter activity. c-Jun inhibited HNF4 alpha and PPARgamma coactivator-1 alpha (PGC-1 alpha) coactivation of CYP7A1 reporter activity, whereas a dominant negative c-Jun did not. Co-immunoprecipitation and ChIP assays revealed that IL-1 beta and CDCA reduced HNF4 alpha bound to the CYP7A1 chromatin, and that c-Jun interacted with HNF4 alpha and blocked HNF4 alpha recruitment of PGC-1 alpha to the CYP7A1 chromatin. In conclusion, IL-1 beta and CDCA inhibit HNF4 alpha but induce c-Jun, which in turn blocks HNF 4 alpha recruitment of PGC-1 alpha to the CYP7A1 chromatin and results in inhibition of CYP7A1 gene transcription. The JNK/c-Jun signaling pathway inhibits bile acid synthesis and protects hepatocytes against the toxic effect of inflammatory agents.
Li Tiangang; Jahan Asmeen; Chiang John Y L
Hepatology (Baltimore, Md.)
2006
2006-06
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1002/hep.21183" target="_blank" rel="noreferrer noopener">10.1002/hep.21183</a>
Glucagon and cAMP inhibit cholesterol 7alpha-hydroxylase (CYP7A1) gene expression in human hepatocytes: discordant regulation of bile acid synthesis and gluconeogenesis.
*Gluconeogenesis; 8-Bromo Cyclic Adenosine Monophosphate/*pharmacology; Adolescent; Bile Acids and Salts/*biosynthesis; Cells; Cholesterol 7-alpha-Hydroxylase/*genetics; Chromatin/metabolism; Cultured; Cyclic AMP-Dependent Protein Kinases/physiology; Enzymologic/*drug effects; Female; Gene Expression Regulation; Genetic/drug effects; Glucagon/*pharmacology; Hepatocyte Nuclear Factor 4/genetics/metabolism; Hepatocytes/*enzymology; Humans; Male; Messenger/analysis; Middle Aged; Organ Specificity; Phosphorylation; RNA; Transcription
The gene encoding cholesterol 7alpha-hydroxylase (CYP7A1) is tightly regulated to control bile acid synthesis and maintain lipid homeostasis. Recent studies in mice suggest that bile acid synthesis is regulated by the fasted-to-fed cycle, and fasting induces CYP7A1 gene expression in parallel to the induction of peroxisome proliferators-activated receptor gamma co-activator 1alpha (PGC-1alpha) and phosphoenolpyruvate carboxykinase (PEPCK). How glucagon regulates CYP7A1 gene expression in the human liver is not clear. Here we show that glucagon and cyclic adenosine monophosphate (cAMP) strongly repressed CYP7A1 mRNA expression in human primary hepatocytes. Reporter assays confirmed that cAMP and protein kinase A (PKA) inhibited human CYP7A1 gene transcription, in contrast to their stimulation of the PEPCK gene. Mutagenesis analysis identified a
Song Kwang-Hoon; Chiang John Y L
Hepatology (Baltimore, Md.)
2006
2006-01
Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).
<a href="http://doi.org/10.1002/hep.20919" target="_blank" rel="noreferrer noopener">10.1002/hep.20919</a>