Strobl J S, van Eys G J, Thompson E B
Department of Pharmacology and Toxicology, West Virginia University, Morgantown 26506.
Mol Cell Endocrinol. 1989 Sep;66(1):71-82. doi: 10.1016/0303-7207(89)90050-6.
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.
为阐明糖皮质激素在GH3垂体细胞中激活生长激素（GH）基因的机制，在有无放线菌酮的情况下，测定了核和细胞质区室中GH mRNA的积累情况。在地塞米松处理的细胞中，核内GH mRNA水平在6小时时升高，细胞质内GH mRNA水平在12小时时升高。地塞米松处理导致48 - 72小时内总GH mRNA水平升高5至24倍。相对于细胞质中GH mRNA的量，核内GH mRNA水平的差异升高持续了48小时。在地塞米松和放线菌酮同时处理的GH3细胞中，核内GH mRNA短暂积累后，细胞质中GH mRNA水平短暂升高。在先用放线菌酮预处理2小时的GH3细胞中，地塞米松介导的核和细胞质中GH mRNA水平的升高被完全阻断。糖皮质激素受体水平不受放线菌酮影响。这些数据表明，糖皮质激素对GH mRNA水平的刺激始于细胞核内，且放线菌酮敏感事件对于这种刺激的发生至关重要。为评估糖皮质激素对GH基因转录激活的重要性，进行了核转录连续反应以及在稳定转化的GH3细胞内的氨基糖苷3'-磷酸转移酶（Neo）融合基因中对GH启动子活性的测定。在核转录连续试验中获得了地塞米松对GH基因进行微弱、短暂转录激活的证据。与此观点一致，在GH - Neo GH3细胞转化体中，暴露于地塞米松30 - 72小时使Neo mRNA水平升高不到或等于2倍。我们得出结论，糖皮质激素对GH3细胞中GH mRNA的刺激需要持续的蛋白质合成，并且在很大程度上可以独立于GH基因转录激活而发生。
