A Reporter Gene Assay for High-Throughput Screening of G-Protein-Coupled Receptors Stably or Transiently Expressed in HEK293 EBNA Cells Grown in Suspension Culture

Durocher, Yves; Perret, Sylvie; Thibaudeau, Eric; Gaumond, Marie‐Hélène; Kamen, Amine; Stocco, Rino; Abramovitz, Mark

doi:10.1006/abio.2000.4698

Cited by 112 publications

(82 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We constructed cDNA of PLAP regulated by CRE in the vasoactive intestinal peptide promoter (5,9); the cDNA was incorporated into a retrovirus vector (18).…”

Section: Construction Of a Reporter Vectormentioning

confidence: 99%

A novel truncated glucagon-like peptide 2 (GLP-2) as a tool for analyzing GLP-2 receptor agonists

et al. 2013

View full text Add to dashboard Cite

Glucagon-like peptide 2 (GLP-2) is an intestinotropic peptide that binds to GLP-2 receptor (GLP-2R), a class-B G protein-coupled receptor. The GLP-2R antagonist GLP-2(3-33) has relatively high partial agonistic activity, and there are as yet no ideal known potent GLP-2R antagonists. We therefore prepared several truncated forms of human GLP-2 and characterized them by binding and reporter assays to find antagonists more potent than GLP-2(3-33). We found that GLP-2(11-33) was the most potent orthosteric GLP-2R antagonist, with binding activity almost equal to those of GLP-2 and GLP-2(3-33) and weaker intrinsic agonistic activity than GLP-2(3-33). GLP-2(11-33) retained weak agonistic activity toward human, cynomolgus monkey, dog, and Syrian hamster GLP-2Rs. However, it had no agonistic activity toward rat GLP-2R. GLP-2(11-33) potentiated the agonistic activity of an ago-allosteric modulator of GLP-2R, compound 1 (N-[1-(2,5-dichlorothiophen-3-yl)-2-(phenylsulfanyl)ethylidene]hydroxylamine), synergistically toward human GLP-2R. In the case of rat GLP-2R, GLP-2(11-33) decreased the agonistic activity of compound 1, although GLP-2 and GLP-2(3-33) increased this activity additively. These findings suggest that the binding sites of the ago-allosteric modulator and GLP-2 overlap, at least in rat GLP-2R. GLP-2(11-33) is a novel, useful tool for analyzing the mode of action of agonists and ago-allosteric modulators of GLP-2R.

show abstract

“…We constructed cDNA of PLAP regulated by CRE in the vasoactive intestinal peptide promoter (5,9); the cDNA was incorporated into a retrovirus vector (18).…”

Section: Construction Of a Reporter Vectormentioning

confidence: 99%

A novel truncated glucagon-like peptide 2 (GLP-2) as a tool for analyzing GLP-2 receptor agonists

et al. 2013

View full text Add to dashboard Cite

show abstract

“…We subjected this short peptide to an assay with HEK293 cells transiently expressing AQ27 and a reporter gene (cAMP-responsive element-luciferase). This assay was modified from the method of Chen et al (20,21). They demonstrated that the activation of GPCRs coupled with G s and/or G q causes an increase in the transcription of the cAMPresponsive element promoter.…”

Section: Identification Of a Novel Rfamide Peptide Gene-wementioning

confidence: 99%

A New Peptidic Ligand and Its Receptor Regulating Adrenal Function in Rats

Fukusumi

Yoshida

Fujii

et al. 2003

Journal of Biological Chemistry

170

290

View full text Add to dashboard Cite

We searched for peptidic ligands for orphan G protein-coupled receptors utilizing a human genome data base and identified a new gene encoding a preproprotein that could generate a peptide. This peptide consisted of 43 amino acid residues starting from Nterminal pyroglutamic acid and ending at C-terminal arginine-phenylalanine-amide. We therefore named it QRFP after pyroglutamylated arginine-phenylalanineamide peptide. We subsequently searched for its receptor and found that Chinese hamster ovary cells expressing an orphan G protein-coupled receptor, AQ27, specifically responded to QRFP. We analyzed tissue distributions of QRFP and its receptor mRNAs in rats utilizing quantitative reverse transcription-polymerase chain reaction and in situ hybridization. QRFP mRNA was highly expressed in the hypothalamus, whereas its receptor mRNA was highly expressed in the adrenal gland. The intravenous administration of QRFP caused the release of aldosterone, suggesting that QRFP and its receptor have a regulatory function in the rat adrenal gland.

show abstract

“…These CREBs bind to the CRE of the EGFP promoter, which results in increased EGFP transcription and green fluorescence emission [9,10] . The measurement of CRE-regulated reporter gene activity provides a sensitive assay for assessing intracellular changes in cAMP levels [11][12][13] in order to study ligand affinity and efficacy for β 2 -AR.…”

Section: Introductionmentioning

confidence: 99%

Identification of anti-asthmatic compounds in Pericarpium citri reticulatae and evaluation of their synergistic effects

et al. 2009

View full text Add to dashboard Cite

Aim:To investigate the anti-asthmatic mechanisms of the traditional Chinese medicine Pericarpium citri reticulatae (PCR). Methods: The alkaloid section (AS) of PCR was extracted using an ion exchange resin, separated, and purified into different fractions by semi-preparative HPLC. These fractions were screened for beta2-adrenergic receptor (β 2 AR) agonistic activity using rat β 2 AR-transfected CHO-CRE-EGFP cells. AS and its isolated components were characterized by ultra-performance liquid chromatography/quadrupole time-of-flight MS (UPLC/Q-Tof MS) and were evaluated for their spasmolytic and antitussive activities both in vitro and in vivo in a guinea pig model. Results: We demonstrated that the AS component responsible for activating β 2 AR signaling was synephrine. Both AS and synephrine showed significant spasmolytic effects on acetylcholine chloride (ACh)-induced contractions in isolated guinea pig trachea, and they protected against histamine-induced experimental asthma by prolonging the latent period. We further identified stachydrine as the antitussive component that could significantly reduce citric acid-induced coughing. The combination of these two bioactive compounds had a more potent spasmolytic activity in comparison with the single use of synephrine or stachydrine. Conclusion: We conclude that synephrine and stachydrine are the key components of AS that mediate asthma relief due to their synergism when used in combination.

show abstract

A Reporter Gene Assay for High-Throughput Screening of G-Protein-Coupled Receptors Stably or Transiently Expressed in HEK293 EBNA Cells Grown in Suspension Culture

Cited by 112 publications

References 32 publications

A novel truncated glucagon-like peptide 2 (GLP-2) as a tool for analyzing GLP-2 receptor agonists

A novel truncated glucagon-like peptide 2 (GLP-2) as a tool for analyzing GLP-2 receptor agonists

A New Peptidic Ligand and Its Receptor Regulating Adrenal Function in Rats

Identification of anti-asthmatic compounds in Pericarpium citri reticulatae and evaluation of their synergistic effects

Contact Info

Product

Resources

About