Dexamethasone control of growth hormone mRNA levels in GH3 pituitary cells is cycloheximide-sensitive and primarily posttranscriptional.
To clarify the mechanism of growth hormone (GH) gene activation by glucocorticoids in GH3 pituitary cells, GH mRNA accumulation in nuclear and cytoplasmic compartments was measured in the presence and absence of cycloheximide. In dexamethasone-treated cells, levels of GH mRNA were increased in the nucleus by 6 h and in the cytoplasm by 12 h. Dexamethasone treatment caused a 5- to 24-fold rise in total GH mRNA levels by 48-72 h. The differential elevation of nuclear levels of GH mRNA relative to the amount of cytoplasmic GH mRNA persisted for 48 h. A transient accumulation of GH mRNA in the nucleus was followed by a brief rise in cytoplasmic GH mRNA levels in GH3 cells treated simultaneously with dexamethasone and cycloheximide. In GH3 cells pretreated for 2 h with cycloheximide, the rise in nuclear and cytoplasmic GH mRNA levels mediated by dexamethasone was blocked completely. Levels of glucocorticoid receptor were unaffected by cycloheximide. These data suggest that the stimulation of GH mRNA levels by glucocorticoids is initiated within the nucleus and that cycloheximide-sensitive events are essential for this stimulation to occur. To assess the importance of GH gene transcriptional activation by glucocorticoids, nuclear transcription run-on reactions and assays of GH promoter activity in an aminoglycoside 3'-phosphotransferase (Neo) fusion gene within stably transformed GH3 cells were performed. Evidence for a weak, transient transcriptional activation of the GH gene by dexamethasone in nuclear run-on assays was obtained. Consistent with this idea, a 30-72 h exposure to dexamethasone raised levels of Neo mRNA in GH-Neo GH3 cell transformants by less than or equal to 2-fold. We conclude that glucocorticoid stimulation of GH mRNA in GH3 cells requires ongoing protein synthesis and can occur largely independently of GH gene transcriptional activation.
Strobl
J S
JS
Department of Pharmacology and Toxicology, West Virginia University, Morgantown 26506.
van Eys
G J
GJ
Thompson
E B
EB
D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000187:drug effects; D003513:Cycloheximide / Q000494:pharmacology*; D003593:Cytoplasm / Q000032:analysis; D004247:DNA / Q000235:genetics; D003907:Dexamethasone / Q000494:pharmacology*; D013006:Growth Hormone / Q000235:genetics*; D008969:Molecular Sequence Data; D010902:Pituitary Gland / Q000166:cytology / Q000187:drug effects*; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012323:RNA Processing, Post-Transcriptional / Q000187:drug effects*; D012333:RNA, Messenger / Q000187:drug effects*; D015138:RNA, Nuclear / Q000032:analysis; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000032:analysis; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D015138:RNA, Nuclear; D011965:Receptors, Glucocorticoid; D003907:Dexamethasone; D013006:Growth Hormone; D004247:DNA; D003513:Cycloheximide