Reporter cell lines are useful tools for monitoring biological activity of nuclear receptor ligands

Abstract

To characterize the specificity of synthetic compounds for nuclear receptors, we established stable cell lines expressing the luciferase gene and different wild-type or chimaeric receptors. MCF-7 cells, which express the oestrogen receptor alpha (ER alpha), and HeLa cells, which do not express the oestrogen receptor, were transfected with a plasmid containing the luciferase gene downstream from a minimum promoter (beta-globin) and an oestrogen-responsive element, generating the MELN and the HELN cell lines, respectively. MELN cells enabled the detection of compounds that bind to the ER alpha or interfere with its pathway. HELN cells were used to establish stable transfectants expressing different nuclear receptors containing the DNA-binding domain of the oestrogen receptors. We thus established ER alpha or ER beta reporter cell lines by transfecting ER alpha or ER beta expression plasmids, and also retinoic acid receptor alpha, beta or gamma reporter cell lines by transfecting the chimaeric RAR gene, in which the DNA-binding domain was replaced by the ER alpha DNA-binding domain.

Protection of mammalian cell used in biosensors by coating with a polyelectrolyte shell

“…Cell line MELN was derived from the MCF-7 breast cancer cells that express estrogen receptor ␣ (ER ␣). MCF-7 cells were stably transfected with a construct expressing the luciferase gene under the control of an estrogen-regulated promoter (Balaguer et al, 1999(Balaguer et al, , 2001.…”

“…For example, exposure to environmental estrogens has been associated with an increase in incidence of testicular and breast cancer, and compromised reproductive capacity (Byford et al, 2002;Diel et al, 2002). To realize a biosensor able to detect molecules displaying a hormone or anti-hormone-like activity on mammals, we used a mammalian cell line: MELN derived from MCF-7 human breast cancer cells stably transfected with an estrogen regulated luciferase gene (Balaguer et al, 1999(Balaguer et al, , 2001. Realization of in vitro or in vivo biosensors using mammalian cells require to isolate them from environment.…”

Development of a high-performance reporter plasmid for detection of chemicals with androgenic activity

“…Moreover, among 56 test chemicals consisting of several types of steroids, only the androgen derivatives selectively triggered a high response in the reporter gene assay using the new reporter plasmid, ARE-AUGLuc+. Recently several reporter gene systems for detecting endocrine-active chemicals have been described (Gagne et al 1994;Vinggaad et al 1999;Balaguer et al 1999Balaguer et al , 2001Miller et al 2000) but most of them were intended for detecting estrogenic activity of chemicals, whereas our new plasmid construct is suitable for detecting the androgenic activity of chemicals. Adoption of high-throughput screening for the androgenic and anti-androgenic activity of chemicals will surely lead to safer chemicals, and the reporter gene assay system should provide a powerful tool to screen for endocrinedisrupting chemicals Yamasaki et al 2002).…”

Highly Potent Naphthofuran‐Based Retinoic Acid Receptor Agonists

“…Both systems are based on fusion proteins, which consist of the ligand binding domain of the corresponding receptor and the DNA binding domain of either the human estrogen receptor (ER DBD) or the yeast Gal4 transcription factor. The cells also contain stably integrated luciferase reporters, which are controlled by estrogen-responsive elements (for ER DBD chimeras) or five Gal4 response elements (for Gal4 DBD chimeras) in front of a minimal b-globin promoter; these are referred to as "ERE-bGlob-Luc-SV-Neo" [46] or "A C H T U N G T R E N N U N G (17m) 5 -bGLuc". [47,48] The Monod, Changeux, and Wyman model was adopted for the description of the activity of the ligands.…”