Regulation of the human glucocorticoid receptor by long-term and chronic treatment with glucocorticoid.
HeLa S3 cells that contain endogenous glucocorticoid receptors (GR) were treated with dexamethasone (DEX) for periods of time ranging from 24 h to 2 weeks or chronically over a 2-year period. Regulation of GR protein and mRNA were examined by affinity labeling, Western blotting, and Northern blotting. Relatively short-term treatment of cells with DEX for 24 or 48 h revealed more profound down-regulation of GR protein than of GR mRNA. However, by 2 weeks of DEX treatment, the levels of both receptor protein and mRNA were both maximally down-regulated. Cells that had been chronically DEX treated (for up to 2 years) had no measurable GR protein or mRNA. The down-regulation of receptor protein and RNA that occurred after 2 weeks of DEX treatment is completely reversible upon DEX removal, whereas reversibility did not occur with cells that had been chronically treated with DEX. Furthermore, transfection of a glucocorticoid responsive reporter plasmid into these chronically DEX-treated cells demonstrated that these cells were no longer responsive to steroid treatment. However, cotransfection of a plasmid encoding the human GR into these chronically DEX-treated cells resulted in restored production of GR and responsiveness to hormone, indicating that the defect in these cells occurs only at the receptor level.
Department of Physiology, University of North Carolina at Chapel Hill, 27599.
C M
CM
Silva
F E
FE
Powell-Oliver
C M
CM
Jewell
M
M
Sar
V E
VE
Allgood
J A
JA
Cidlowski
D015152:Blotting, Northern; D015153:Blotting, Western; D003907:Dexamethasone / Q000494:pharmacology*; D015536:Down-Regulation; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D012333:RNA, Messenger / Q000378:metabolism; D011965:Receptors, Glucocorticoid / Q000187:drug effects*; D013997:Time Factors; D014162:Transfection
D012333:RNA, Messenger; D011965:Receptors, Glucocorticoid; D003907:Dexamethasone