NATURE OF STEROID-RECEPTOR INTERACTIONS

本项目的目标是定义初始的，细胞内的 类固醇激素作用的事件。 这些事件包括类固醇 与细胞内受体分子的结合， 受体类固醇复合物的DNA结合和核结合 物种，以及活化的复合物与那些核 参与转录调控的受体位点 特定基因 一种用于检查这些的方法 步骤是比较各种类固醇在不同的属性 细胞系 因此，以前的研究的诱导量， 酪氨酸氨基转移酶（达特）的几种糖皮质激素在两个 大鼠肝癌组织培养系（HTC和Fu 5 -5）显示， 最大达特的50%所需的类固醇浓度 HTC细胞中的诱导比Fu 5 -5细胞中高约6倍。 现在在达特酶的诱导中也看到了同样的差异 和mRNA水平的稳定的cAMP衍生物，（8-（4- 氯苯硫基）cAMP）。 这些数据表明，一个共同的前- 翻译事件决定达特的不同敏感性 HTC和Fu 5 -5细胞中糖皮质激素和cAMP诱导。 第二种方法是检查 不可逆抗糖皮质激素（和亲和标记）地塞米松 21-甲磺酸酯（Dex-Mes）。 Dex-Mes 在碱性水溶液中与蛋白质的半胱氨酸特异性反应， 解决方案 与糖皮质激素受体的Dex-Mes反应 在类固醇结合位点的Cys-656处唯一发生。 这 鉴定与生物活性相关的第一个氨基酸， 糖皮质激素受体的特性将有助于未来的研究。 结构-功能研究

The objective of this project is to define the initial, intracellular events of steroid hormone action. These events include steroid binding to the intracellular receptor molecule, "activation" of the receptor-steroid complex to a DNA-binding and nuclear-binding species, and binding of the activated complex to those nuclear acceptor sites involved in the regulation of transcription of specific genes. One approach that has been used to examine these steps is to compare the properties of various steroids in different cell lines. Thus previous studies of the amount of induction of tyrosine aminotransferase (TAT) by several glucocorticoids in two rat hepatoma tissue culture lines (HTC and Fu5-5) revealed that the steroid concentration required for 50% of maximal TAT induction in HTC cells was about 6-fold higher than in Fu5-5 cells. The same difference is now seen in the induction of TAT enzyme and mRNA levels by the stable cAMP derivative, (8-(4- chlorophenylthio)cAMP). These data suggest that a common pre- translational event determines the different sensitivity of TAT induction by glucocorticoids and by cAMP in HTC and Fu5-5 cells. A second approach has been to examine the properties of the irreversible antiglucocorticoid (and affinity label) dexamethasone 21-mesylate (Dex-Mes) at a molecular level. Dex-Mes specifically reacts with the cysteines of proteins in basic aqueous solutions. Dex-Mes reaction with the glucocorticoid receptor occurs uniquely at Cys-656 in the steroid binding site. This identification of the first amino acid associated with a biological property of the glucocorticoid receptor should facilitate future structure-function studies.