Gil B, Pajares M A, Mato J M, Alvarez L
Institute of Biomedical Investigation, Consejo Superior de Investigaciones Cientificas, Madrid, Spain.
Endocrinology. 1997 Mar;138(3):1251-8. doi: 10.1210/endo.138.3.4967.
The effects of glucocorticoids on the regulation of rat liver S-adenosylmethionine synthetase were studied in vivo and in two culture systems. Livers from adrenalectomized animals were examined for enzyme activity, immunoreactive protein, and messenger RNA (mRNA) content. All three parameters showed a similar trend, i.e. they decreased 3-fold after adrenalectomy and increased over the control values upon triamcinolone replacement. These results suggested that glucocorticoid regulation of hepatic S-adenosylmethionine synthetase was mediated at the mRNA level. Triamcinolone and dexamethasone increased S-adenosylmethionine synthetase mRNA content in a time- and dose-dependent manner in both rat hepatoma H35 cells and primary cultures of adult rat hepatocytes. The kinetics of mRNA induction were identical in both culture systems, indicating that the hormone-mediated response is independent of the differentiated state of the cell. Insulin blocked the inducing effect of glucocorticoids on S-adenosylmethionine synthetase mRNA in a dose-dependent manner. On the other hand, the triamcinolone-dependent increase in mRNA levels was completely abolished by treatment with actinomycin D, whereas cycloheximide did not affect this response. The transcription rate of the gene, as measured by run-on assay, increased 3-fold after hormone addition. Transient transfections of H35 cells with 1.4 kilobases of the 5'-flanking region of the hepatic S-adenosylmethionine synthetase gene fused to a luciferase reporter gene showed that promoter activity is also increased 3-fold after triamcinolone treatment, suggesting that this promoter region contains the sequence elements necessary to confer glucocorticoid responsiveness. In addition to the transcriptional control of the hepatic S-adenosylmethionine synthetase gene, our results suggest that glucocorticoids may be acting at a posttranscriptional level.
在体内以及两种培养系统中研究了糖皮质激素对大鼠肝脏S-腺苷甲硫氨酸合成酶调节的影响。检测了肾上腺切除动物肝脏的酶活性、免疫反应性蛋白和信使核糖核酸（mRNA）含量。所有这三个参数都呈现出相似的趋势，即肾上腺切除后它们下降了3倍，而在使用曲安西龙替代后又高于对照值。这些结果表明，糖皮质激素对肝脏S-腺苷甲硫氨酸合成酶的调节是在mRNA水平介导的。曲安西龙和地塞米松在大鼠肝癌H35细胞和成年大鼠肝细胞原代培养物中均以时间和剂量依赖的方式增加了S-腺苷甲硫氨酸合成酶mRNA的含量。两种培养系统中mRNA诱导的动力学是相同的，这表明激素介导的反应与细胞的分化状态无关。胰岛素以剂量依赖的方式阻断了糖皮质激素对S-腺苷甲硫氨酸合成酶mRNA的诱导作用。另一方面，用放线菌素D处理可完全消除曲安西龙依赖的mRNA水平升高，而环己酰亚胺则不影响这种反应。通过连续转录分析测定，激素添加后基因的转录速率增加了3倍。用与荧光素酶报告基因融合的肝脏S-腺苷甲硫氨酸合成酶基因5'-侧翼区的1.4千碱基对瞬时转染H35细胞表明，曲安西龙处理后启动子活性也增加了3倍，这表明该启动子区域包含赋予糖皮质激素反应性所需的序列元件。除了肝脏S-腺苷甲硫氨酸合成酶基因的转录控制外，我们的结果表明糖皮质激素可能在转录后水平起作用。
