scholarly journals Orphan Nuclear Receptor RORα Regulates Enzymatic Metabolism of Cerebral 24S-Hydroxycholesterol through CYP39A1 Intronic Response Element Activation

2020 ◽  
Vol 21 (9) ◽  
pp. 3309
Author(s):  
Hiroshi Matsuoka ◽  
Miyu Katayama ◽  
Ami Ohishi ◽  
Kaoruko Miya ◽  
Riki Tokunaga ◽  
...  
Keyword(s):  
Nuclear Receptor ◽  
Cholesterol Homeostasis ◽  
Orphan Receptor ◽  
Hek293 Cells ◽  
Luciferase Reporter ◽  
Metabolic Enzyme ◽  
Orphan Nuclear Receptor ◽  
Human Hepatoma Cells ◽  
Mobility Shift ◽  
The Brain

Oxysterols, important regulators of cholesterol homeostasis in the brain, are affected by neurodegenerative diseases. Early-onset Alzheimer’s disease is associated with higher levels of circulating brain-derived 24S-hydroxycholesterol (24S-OHC). Conversion of cholesterol to 24S-OHC is mediated by cholesterol 24S-hydroxylase in the brain, which is the major pathway for oxysterol elimination, followed by oxidation through hepatic first-pass metabolism by CYP39A1. Abnormal CYP39A1 expression results in accumulation of 24S-OHC, influencing neurodegenerative disease-related deterioration; thus, it is important to understand the normal elimination of 24S-OHC and the system regulating CYP39A1, a selective hepatic metabolic enzyme of 24S-OHC. We examined the role of transcriptional regulation by retinoic acid receptor-related orphan receptor α (RORα), a nuclear receptor that responds to oxysterol ligands. In humans, the promoter and first intronic regions of CYP39A1 contain two putative RORα response elements (ROREs). RORα binding and responses of these ROREs were assessed using electrophoretic mobility shift, chromatin immunoprecipitation, and luciferase reporter assays. CYP39A1 was upregulated by RORα overexpression in HEK293 cells, while RORα knockdown by siRNA significantly downregulated CYP39A1 expression in human hepatoma cells. Additionally, CYP39A1 was induced by RORα agonist treatment, suggesting that CYP39A1 expression is activated by RORα nuclear receptors. This may provide a way to increase CYP39A1 activity using RORα agonists, and help halt 24S-OHC accumulation in neurodegenerative illnesses.

scholarly journals Differential regulation of aldosterone synthase and 11beta-hydroxylase transcription by steroidogenic factor-1

2002 ◽  
Vol 28 (2) ◽  
pp. 125-135 ◽  
Author(s):  
MH Bassett ◽  
Y Zhang ◽  
C Clyne ◽  
PC White ◽  
WE Rainey
Keyword(s):  
Electrophoretic Mobility ◽  
Cell Model ◽  
Orphan Receptor ◽  
Luciferase Reporter ◽  
Zona Fasciculata ◽  
Genetic Linkage Analysis ◽  
Mobility Shift ◽  
Steroidogenic Factor 1 ◽  
Aldosterone Synthase ◽  
H295r Cells

11beta-Hydroxylase (hCYP11B1) and aldosterone synthase (hCYP11B2) are closely related isozymes with distinct roles in cortisol and aldosterone production respectively. Aldosterone synthase catalyzes the final step in aldosterone biosynthesis and is expressed only in the zona glomerulosa of the normal adrenal. 11beta-Hydroxylase catalyzes the final reaction in the production of cortisol and is expressed at higher levels in the zona fasciculata. The mechanisms causing differential expression of these genes are not well defined. Herein, we demonstrate contrasting roles for the orphan receptor steroidogenic factor-1 (SF-1) in the regulation of human (h) CYP11B1 and hCYP11B2. Human NCI-H295R (H295R) or mouse Y-1 cells were transiently transfected with luciferase reporter constructs containing 5'-flanking regions of hCYP11B1, hCYP11B2, human 17alpha-hydroxylase (hCYP17), human cholesterol side-chain cleavage (hCYP11A1) or mouse (m) cyp11b2 (mcyp11b2). Co-transfection of vectors encoding SF-1 increased expression of hCYP11B1, hCYP11A1 and hCYP17 constructs, but inhibited hCYP11B2 reporter activity. Murine, bovine and human SF-1 were unable to increase transcription of hCYP11B2 in H295R cells. Both hCYP11B2 and mcyp11b2 promoter constructs were inhibited similarly by human SF-1. In mouse Y-1 cells, reporter expression of hCYP11B2 and mcyp11b2 was very low compared with hCYP11B1 constructs, suggesting that this adrenal cell model may not be appropriate for studies of CYP11B2. Electrophoretic mobility shift assay demonstrated that SF-1 interacted with an element from both hCYP11B1 and hCYP11B2. However, mutation of this element, termed Ad4, did not prevent agonist stimulation of hCYP11B2 by angiotensin II or forskolin but blocked activity of hCYP11B1. In some, but not all, reports of genetic linkage analysis, a naturally occurring single nucleotide polymorphism within the Ad4 element of hCYP11B2 (-344C/T) has been associated with cardiovascular disease. Herein, we have demonstrated that this polymorphism influenced binding of SF-1 in electrophoretic mobility shift assays, with the C allele binding SF-1 more strongly than the T allele. However, when hCYP11B2 constructs containing these alleles were transfected into H295R cells, there was no difference in agonist-stimulated expression or the response of either reporter construct to co-expression with human SF-1. Taken together, these data suggest that SF-1 and the Ad4 element are not major regulators of adrenal hCYP11B2 gene expression. Thus far, hCYP11B2 is the first steroid hydroxylase gene which is not positively regulated by SF-1.

scholarly journals The Orphan Nuclear Receptor 4A1: A Potential New Therapeutic Target for Metabolic Diseases

2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Lei Zhang ◽  
Qun Wang ◽  
Wen Liu ◽  
Fangyan Liu ◽  
Ailing Ji ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Lipid Metabolism ◽  
Nuclear Receptor ◽  
Metabolic Diseases ◽  
Early Response ◽  
Orphan Receptor ◽  
Special Focus ◽  
Orphan Nuclear Receptor ◽  
Physiological Functions ◽  
The Impact

Orphan nuclear receptor 4A1 (NR4A1) is a transcriptional factor of the nuclear orphan receptor (NR4A) superfamily that has sparked interest across different research fields in recent years. Several studies have demonstrated that ligand-independent NR4A1 is an immediate-early response gene and the protein product is rapidly induced by a variety of stimuli. Hyperfunction or dysfunction of NR4A1 is implicated in various metabolic processes, including carbohydrate metabolism, lipid metabolism, and energy balance, in major metabolic tissues, such as liver, skeletal muscle, pancreatic tissues, and adipose tissues. No endogenous ligands for NR4A1 have been identified, but numerous compounds that bind and activate or inactivate nuclear NR4A1 or induce cytoplasmic localization of NR4A1 have been identified. This review summarizes recent advances in our understanding of the molecular biology and physiological functions of NR4A1. And we focus on the physiological functions of NR4A1 receptor to the development of the metabolic diseases, with a special focus on the impact on carbohydrate and lipid metabolism in skeletal muscle, liver, adipose tissue, and islet.

The orphan nuclear receptor NGFI-B regulates expression of the gene encoding steroid 21-hydroxylase

1993 ◽  
Vol 13 (2) ◽  
pp. 861-868
Author(s):  
T E Wilson ◽  
A R Mouw ◽  
C A Weaver ◽  
J Milbrandt ◽  
K L Parker
Keyword(s):  
Nuclear Receptor ◽  
Transcriptional Activation ◽  
Core Promoter ◽  
Orphan Nuclear Receptor ◽  
Recognition Sequence ◽  
Mobility Shift ◽  
Adrenocortical Cells ◽  
Gene Encoding ◽  
Adult Rat ◽  
Gel Mobility Shift

As part of its trophic action to maintain the steroidogenic capacity of adrenocortical cells, corticotropin (ACTH) increases the transcription of the cytochrome P-450 steroid hydroxylase genes, including the gene encoding steroid 21-hydroxylase (21-OHase). We previously identified several promoter elements that regulate 21-OHase gene expression in mouse Y1 adrenocortical tumor cells. One of these elements, located at nucleotide -65, closely resembles the recognition sequence of the orphan nuclear receptor NGFI-B, suggesting that NGFI-B regulates this essential steroidogenic enzyme. To explore this possibility, we first used in situ hybridization to demonstrate high levels of NGFI-B transcripts in the adrenal cortex of the adult rat. In cultured mouse Y1 adrenocortical cells, treatment with ACTH, the major regulator of 21-OHase transcription, rapidly increased NGFI-B expression. Gel mobility shift and DNase I footprinting experiments showed that recombinantly expressed NGFI-B interacts specifically with the 21-OHase -65 element and identified one complex formed by Y1 extracts and the 21-OHase -65 element that contains NGFI-B. Expression of NGFI-B significantly augmented the activity of the intact 21-OHase promoter, while mutations of the -65 element that abolish NGFI-B binding markedly diminished NGFI-B-mediated transcriptional activation. Specific mutations of NGFI-B shown previously to impair either DNA binding or transcriptional activation diminished the effect of NGFI-B coexpression on 21-OHase expression. Finally, an oligonucleotide containing the NGFI-B response element conferred ACTH response to a core promoter from the prolactin gene, showing that this element is sufficient for ACTH induction. Collectively, these results identify a cellular promoter element that is regulated by NGFI-B and implicate NGFI-B in the transcriptional induction of 21-OHase by ACTH.

scholarly journals Bavachalcone Enhances RORαExpression, Controls Bmal1 Circadian Transcription, and Depresses Cellular Senescence in Human Endothelial Cells

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Yanqi Dang ◽  
Shuang Ling ◽  
Jing Ma ◽  
Rongzhen Ni ◽  
Jin-Wen Xu
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Replicative Senescence ◽  
Orphan Receptor ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Orphan Nuclear Receptor ◽  
Human Endothelial Cells ◽  
Peripheral Tissues ◽  
Natural Medicine

The circadian clock regulates many aspects of (patho)physiology in the central nervous system and in the peripheral tissues. RAR-related orphan receptorα(RORα), an orphan nuclear receptor, is involved in circadian rhythm regulation, including regulation of cardiovascular function. Bavachalcone, a prenylchalcone, is a major bioactive chalcone isolated fromPsoralea corylifolia. This natural ingredient activated RORα1 luciferase reporter activity on drug screening. In addition, bavachalcone induced RORα1 expression in mRNA and protein levels in a dose-dependent manner and enhanced the circadian amplitude of Bmal1 mRNA expression after serum shock. Moreover, bavachalcone suppressed senescence in human endothelial cells and mRNA expression ofp16ink4a(a marker of replicative senescence) and IL-1α(a proinflammatory cytokine of the senescence-associated secretory phenotype). These inhibitory effects were partially reversed by the RORαinhibitor VPR-66. Our results demonstrate that bavachalcone, as a natural medicine ingredient, has a pharmacological function in regulating RORα1.

scholarly journals Spermatogenesis and Testis Development Are Normal in Mice Lacking Testicular Orphan Nuclear Receptor 2

2002 ◽  
Vol 22 (13) ◽  
pp. 4661-4666 ◽  
Author(s):  
Chih-Rong Shyr ◽  
Loretta L. Collins ◽  
Xiao-Min Mu ◽  
Kenneth A. Platt ◽  
Chawnshang Chang
Keyword(s):  
Protein Expression ◽  
Nuclear Receptor ◽  
Expression Patterns ◽  
Orphan Receptor ◽  
Orphan Nuclear Receptor ◽  
Male Mice ◽  
Orphan Nuclear Receptors ◽  
Nuclear Receptor Superfamily ◽  
Testis Development

ABSTRACT Early in vitro cell culture studies suggested that testicular orphan nuclear receptor 2 (TR2), a member of the nuclear receptor superfamily, may play important roles in the control of several pathways including retinoic acids, vitamin D, thyroid hormones, and ciliary neurotrophic factor. Here we report the surprising results showing that mice lacking TR2 are viable and have no serious developmental defects. Male mice lacking TR2 have functional testes, including normal sperm number and motility, and both male and female mice lacking TR2 are fertile. In heterozygous TR2+/− male mice we found that β-galactosidase, the indicator of TR2 protein expression, was first detected at the age of 3 weeks and its expression pattern was restricted mainly in the spermatocytes and round spermatids. These protein expression patterns were further confirmed with Northern blot analysis of TR2 mRNA expression. Together, results from TR2-knockout mice suggest that TR2 may not play essential roles in spermatogenesis and normal testis development, function, and maintenance. Alternatively, the roles of TR2 may be redundant and could be played by other close members of the nuclear receptor superfamily such as testicular orphan receptor 4 (TR4) or unidentified orphan receptors that share many similar functions with TR2. Further studies with double knockouts of both orphan nuclear receptors, TR2 and TR4, may reveal their real physiological roles.

scholarly journals A constitutive nuclear localization signal from the second zinc-finger of orphan nuclear receptor TR2

1998 ◽  
Vol 159 (1) ◽  
pp. 53-60 ◽  
Author(s):  
Z Yu ◽  
CH Lee ◽  
C Chinpaisal ◽  
LN Wei
Keyword(s):  
Nuclear Receptor ◽  
Zinc Finger ◽  
Nuclear Localization ◽  
Nuclear Localization Signal ◽  
Fluorescent Protein ◽  
Biologically Active ◽  
Binding Property ◽  
Orphan Nuclear Receptor ◽  
Mobility Shift ◽  
Localization Signal

The orphan nuclear receptor TR2 and its truncated isoform deleted in the ligand binding domain (LBD) were localized exclusively in the nuclei as revealed by two methods of detection. An anti-hemagglutinin (HA) antibody detected specific nuclear localization of HA-tagged receptors and the green fluorescent protein (GFP)-tagged receptors were found to be distributed in the nuclei of living cells. By deletion analyses, the sequence responsible for targeting this receptor into the nucleus was defined. A stretch of 20 amino acid residues (KDCVINKHHRNRCQYCRLQR) within the second zinc-finger of this receptor is required for its nuclear localization and this signal is constitutively active. No nuclear localization signal was found in the N-terminus or the LBD. The GFP-tagged receptor remained biologically active, as evidenced by its repressive activity on the reporter that carried a binding site for this receptor, a direct repeat-5 (DR5). An electrophoretic mobility shift assay was performed to characterize the binding property of TR2 and its truncated isoform. TR2 bound to the DR5 as dimers whereas its truncated isoform bound as monomers.

scholarly journals Orphan Nuclear Receptor ERRγ Is a Novel Transcriptional Regulator of IL-6 Mediated Hepatic BMP6 Gene Expression in Mice

2020 ◽  
Vol 21 (19) ◽  
pp. 7148
Author(s):  
Kamalakannan Radhakrishnan ◽  
Yong-Hoon Kim ◽  
Yoon Seok Jung ◽  
Jina Kim ◽  
Don-Kyu Kim ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcriptional Regulation ◽  
Molecular Mechanism ◽  
Nuclear Receptor ◽  
Iron Homeostasis ◽  
Luciferase Reporter ◽  
Orphan Nuclear Receptor ◽  
Knock Out

Bone morphogenetic protein 6 (BMP6) is a multifunctional growth factor involved in organ development and homeostasis. BMP6 controls expression of the liver hormone, hepcidin, and thereby plays a crucial role in regulating iron homeostasis. BMP6 gene transcriptional regulation in liver is largely unknown, but would be of great help to externally modulate iron load in pathologic conditions. Here, we describe a detailed molecular mechanism of hepatic BMP6 gene expression by an orphan nuclear receptor, estrogen-related receptor γ (ERRγ), in response to the pro-inflammatory cytokine interleukin 6 (IL-6). Recombinant IL-6 treatment increases hepatic ERRγ and BMP6 expression. Overexpression of ERRγ is sufficient to increase BMP6 gene expression in hepatocytes, suggesting that IL-6 is upstream of ERRγ. In line, knock-down of ERRγ in cell lines or a hepatocyte specific knock-out of ERRγ in mice significantly decreases IL-6 mediated BMP6 expression. Promoter studies show that ERRγ directly binds to the ERR response element (ERRE) in the mouse BMP6 gene promoter and positively regulates BMP6 gene transcription in IL-6 treatment conditions, which is further confirmed by ERRE mutated mBMP6-luciferase reporter assays. Finally, an inverse agonist of ERRγ, GSK5182, markedly inhibits IL-6 induced hepatic BMP6 expression in vitro and in vivo. Taken together, these results reveal a novel molecular mechanism on ERRγ mediated transcriptional regulation of hepatic BMP6 gene expression in response to IL-6.

Orphan nuclear receptor SHP interacts with and represses hepatocyte nuclear factor-6 (HNF-6) transactivation

2008 ◽  
Vol 413 (3) ◽  
pp. 559-569 ◽  
Author(s):  
Yong-Soo Lee ◽  
Don-Kyu Kim ◽  
Yong Deuk Kim ◽  
Ki Cheol Park ◽  
Minho Shong ◽  
...  
Keyword(s):  
Nuclear Receptor ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Target Genes ◽  
Consensus Sequence ◽  
Nuclear Factor ◽  
Orphan Nuclear Receptor ◽  
Hepatocyte Nuclear Factor ◽  
Mobility Shift ◽  
Endocrine Differentiation

SHP (small heterodimer partner; NR0B2) is an atypical orphan NR (nuclear receptor) that functions as a transcriptional co-repressor by interacting with a diverse set of NRs and transcriptional factors. HNF-6 (hepatocyte nuclear factor-6) is a key regulatory factor in pancreatic development, endocrine differentiation and the formation of the biliary tract, as well as glucose metabolism. In this study, we have investigated the function of SHP as a putative repressor of HNF-6. Using transient transfection assays, we have shown that SHP represses the transcriptional activity of HNF-6. Confocal microscopy revealed that both SHP and HNF-6 co-localize in the nuclei of cells. SHP physically interacted with HNF-6 in protein–protein association assays in vitro. EMSAs (electrophoretic mobility-shift assays) and ChIP (chromatin immunoprecipitation) assays demonstrated that SHP inhibits the DNA-binding activity of HNF-6 to an HNF-6-response element consensus sequence, and the HNF-6 target region of the endogenous G6Pase (glucose 6-phosphatase) promoter respectively. Northern blot analysis of HNF-6 target genes in cells infected with adenoviral vectors for SHP and SHP siRNAs (small inhibitory RNAs) indicated that SHP represses the expression of endogenous G6Pase and PEPCK (phosphoenolpyruvate carboxykinase). Our results suggest that HNF-6 is a novel target of SHP in the regulation of gluconeogenesis.

Circadian clock regulates granulosa cell autophagy through NR1D1-mediated inhibition of ATG5

2021 ◽  
Author(s):  
Jing Zhang ◽  
Lijia Zhao ◽  
Yating Li ◽  
Hao Dong ◽  
Haisen Zhang ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Clock Genes ◽  
Expression Patterns ◽  
Current Data ◽  
Orphan Receptor ◽  
Luciferase Reporter ◽  
Mobility Shift ◽  
Rhythmic Expression ◽  
Clock System ◽  
Circadian Clock Genes

Autophagy of granulosa cells (GCs) is involved in follicular atresia, which occurs repeatedly during the ovarian development cycle. Several circadian clock genes are rhythmically expressed in both rodent ovarian tissues and GCs. Nuclear receptor subfamily 1 group D member 1 (NR1D1), an important component of the circadian clock system, is involved in the autophagy process through the regulation of autophagy-related genes. However, there are no reports illustrating the role of the circadian clock system in mouse GC autophagy. In the present study, we found that core circadian clock genes (Bmal1, Per2, Nr1d1, and Dbp) and an autophagy-related gene (Atg5) exhibited rhythmic expression patterns across 24 h in mouse ovaries and primary GCs. Treatment with SR9009, an agonist of NR1D1, significantly reduced the expression of Bmal1, Per2, and Dbp in mouse GCs. ATG5 expression was significantly attenuated by SR9009 treatment in mouse GCs. Conversely, Nr1d1 knockdown increased ATG5 expression in mouse GCs. Decreased NR1D1 expression at both the mRNA and protein levels was detected in the ovaries of Bmal1-/- mice, along with elevated expression of ATG5. Dual-luciferase reporter assay and electrophoretic mobility shift assay showed that NR1D1 inhibited Atg5 transcription by binding to two putative retinoic acid-related orphan receptor response elements within the promoter. In addition, rapamycin-induced autophagy and ATG5 expression were partially reversed by SR9009 treatment in mouse GCs. Taken together, our current data demonstrated that the circadian clock regulates GC autophagy through NR1D1-mediated inhibition of ATG5 expression, and thus, plays a role in maintaining autophagy homeostasis in GCs.

Sign in / Sign up

Export Citation Format

Share Document