Identification of Glucocorticoid Response Element of the Rat TRH gene
- Publisher: Korean Association of Internal Medicine
The Korean Journal of Internal Medicine,
(issn: 1226-3303, eissn: 2005-6648)
Original Article | Transcription factors | Transfection | TRH promoter | GRE | HeLa cells
mesheuropmc: endocrine system | hormones, hormone substitutes, and hormone antagonists
Objectives It was suggested that glucocorticoid exerts cell-specific effects on thyrotropin-releasing hormone (TRH) gene expression at the transcriptional level. Althought there is no typical palindromic glucocorticoid response element (GRE) on the rat TRH gene promoter, a perfect GRE half-site is found between −205 bp and −211 bp. We investigated whether the segment between −242 bp and −200 bp of the rat TRH gene promoter is responsible for glucocorticoid response. Methods For the 5′ deletion study of the TRH gene, four differnt plasmid constructs, pTRH(−554/84), pTRH(−242/84), pTRH(−200/6), pTRH(−113/84) were used for transient transfection study. The plasmid pMAMneo-LUC was used as a positive control and pRSV-GR expression vector, as a co-transfection study. Transfection was performed by the modified calcium precipitation method with 2 μg of each lasmid on the HeLa cells. Results Dexamethasone (DEX) stimulated the transcriptional activity of pTRH(−554/84)-LUC and pTRH(−242/84)-LUC by approximately 3.2 fold at 10−8 M and 5.4 fold at 10−6 M. On the contrary, deletion of the region between −242 to −200 bp reduced the basal transcriptional activity by 90% and also completely abolished the DEX-induced transcriptional activation of the luciferase gene. The DEX-induced transcriptional activation of pTRH(−242/84)-LUC was in dose-dependent manner. While the co-transfection of glucocorticoid receptor expression vector (pRSV-GR) did not increase the basal transcriptional activity of pMAMneo-LUC, it increased the basal transcription of pTRH(−242/84)-LUC by 1.8 fold. The pRSV-GR co-transfection and DEX tratment further increased the transcription of pTRH(−242/84)-LUC by 2–4 fold at the concentration of 10−8 M. Conclusion These findings suggest that a cis-acting element(s) which is important for the basal transcriptional activation and glucocorticoid response of the rat TRH gene is located between −242 bp and −200 bp. The gene has a weak GRE glucocorticoid response and seems to be mediated by an interaction between glucocorticoid receptor and other transcriptional factor (s).