title: Construction of sensitive reporter assay yeasts for comprehensive detection of ligand activities of human corticosteroid receptors through inactivation of CWP and PDR genes.
abstract: BACKGROUND
  The glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) are members of the nuclear receptor superfamily and ligand-dependent transcription factors, whose major ligands are glucocorticoid and mineralocorticoid, so-called corticosteroids. The corticosteroids are a class of substances that include steroid hormones naturally produced in the adrenal cortex of vertebrates and analogues of these hormones that are synthesized in industry. They are involved in a wide range of physiological processes including stress and immune responses, and the regulation of carbohydrate metabolism, protein catabolism, sodium homeostasis, and inflammation. These substances are potential environmental contaminants because they are clinically consumed in large amounts worldwide. To develop a simple and sensitive bioassay to detect corticosteroids, we newly established reporter assay yeasts expressing human GR and MR.
  
  
  METHODS
  Ligand responses of the established assay yeasts were improved by forced expression of a human transcription coactivator SRC-1e. Further enhancement of the responses was achieved by inactivating the CWP and PDR genes that encode cell wall mannoproteins and plasma membrane efflux pumps, respectively, which may be attributable to an increased intracellular concentration of ligands.
  
  
  RESULTS
  These new assay yeasts were more responsive to both natural and synthetic agonist ligands than the conventional assay yeasts. They detected both agonistic and antagonistic activities of mifepristone, spironolactone, and eplerenone in a receptor-selective manner. They also detected ligand activities contained in oral pharmaceutical tablets and human urine.
  
  
  CONCLUSIONS
  This assay system will be a valuable tool to detect agonists as well as antagonists of corticosteroid receptors, in the fields of drug discovery and the assessment of environmental pollutants.
authors.affiliation: Department of Biological Science, Graduate School of Science, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Sayoko
authors.initials: S
authors.lastname: Ito-Harashima
authors.affiliation: Department of Biological Science, Graduate School of Science, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Shiizaki
authors.affiliation: Department of Biological Science, Graduate School of Science, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Masanobu
authors.initials: M
authors.lastname: Kawanishi
authors.affiliation: Department of Biological Science, Graduate School of Science, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Kakiuchi
authors.affiliation: Department of Biological Science, Graduate School of Science, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Kana
authors.initials: K
authors.lastname: Onishi
authors.affiliation: Department of Applied Life Science, Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Sakai, Osaka, Japan.
authors.forename: Ryoichi
authors.initials: R
authors.lastname: Yamaji
authors.affiliation: Department of Biological Science, Graduate School of Science, Osaka Prefecture University, Sakai, Osaka, Japan; Department of Life Science, Dongguk University, Seoul, Republic of Korea. Electronic address: yagi-t@riast.osakafu-u.ac.jp.
authors.forename: Takashi
authors.initials: T
authors.lastname: Yagi
mesh_terms: D001681:Biological Assay / Q000379:methods; D000077545:Eplerenone; D017930:Genes, Reporter / Q000235:genetics*; D006801:Humans; D008024:Ligands; D015735:Mifepristone / Q000494:pharmacology; D011965:Receptors, Glucocorticoid / Q000235:genetics*; D018161:Receptors, Mineralocorticoid / Q000235:genetics*; D011987:Receptors, Steroid / Q000235:genetics*; D013148:Spironolactone / Q000031:analogs & derivatives / Q000494:pharmacology; D015003:Yeasts / Q000187:drug effects / Q000235:genetics*
chemical_list: D008024:Ligands; D011965:Receptors, Glucocorticoid; D018161:Receptors, Mineralocorticoid; D011987:Receptors, Steroid; D013148:Spironolactone; D015735:Mifepristone; D000077545:Eplerenone
keywords: Agonist; Antagonist; Glucocorticoid receptors; Mineralocorticoid receptors; Reporter assay; Saccharomyces cerevisiae; Steroid hormone