Functional characterization of chicken glucocorticoid and mineralocorticoid receptors

Abstract

Glucocorticoid (GR) and mineralocorticoid (MR) receptors are ligand-activated transcription factors that belong to the nuclear hormone receptor superfamily. Little is known about the function of GR and MR in avian species. Recently, the chicken homologue of the GR (cGR) gene was cloned, and its tissue-specific expression was characterized, whereas the full-length sequence of the chicken MR (cMR) gene remains unknown. Therefore, the aims of this project were to clone the full-length cMR and to functionally characterize both chicken receptors. Cos-7 cells were transiently transfected with cGR or cMR expression vectors along with a glucocorticoid response element-luciferase (GRE-Luc) reporter construct. Transfected cells were then treated with increasing doses of corticosterone (CORT) or aldosterone (ALDO) alone and with GR or MR antagonists (ZK98299 and spironolactone, respectively). Transactivation of cGR or cMR was evaluated by luciferase assay. CORT and ALDO induced cGR- and cMR-driven transcriptional activity in a dose-dependent manner. Each receptor responded to both steroids, but cMR transcriptional activity was induced by lower levels of CORT and ALDO than cGR. Coexpression of both chicken corticosteroid receptors in Cos-7 cells had no synergistic or additive effect on CORT- or ALDO-induced transcriptional activity. Corticosteroid-dependent transactivation of cGR and cMR was partially blocked by antagonists. ZK98299 showed high specificity to cGR, while spironolactone had agonist properties toward both receptors. Immunocytochemistry was used to assess the cellular localization of both receptors. Corticosteroids induced translocation of both receptors into the nucleus. The functional properties of cGR and cMR determined in this study will be helpful in defining the physiological roles of GR and MR in avian species.