Lefebvre Y A, Venkatraman J T, Golsteyn E J, Howell G M
Biochem Cell Biol. 1986 Jun;64(6):594-600. doi: 10.1139/o86-082.
Three approaches have been taken to determine the molecular mechanism by which steroid hormones traverse the nuclear envelope on their way to the genome. The first approach involved characterization of steroid binding to nuclear envelope preparations. We have characterized androgen binding to nuclear envelopes isolated from the rat ventral prostate, the rat liver, and androgen-responsive and androgen-unresponsive cell lines of the Shionogi mouse mammary carcinoma and glucocorticoid binding to rat liver. Relatively high affinity binding sites for steroids have been identified on nuclear envelopes. Importantly, the number and specificity of the sites correlates with the responsiveness of the tissue to the steroid. In the second approach, we have undertaken to identify the steroid binding site directly. As the characteristics of the rat ventral prostate site resembled those of the nuclear androgen receptor, we have begun purifying that receptor and have found fast protein liquid chromatography to be very effective. By affinity labelling studies, the dexamethasone binding site on the rat liver nuclear envelope has been identified as a peptide of molecular weight of approximately 90,000. The third approach we have used is to identify androgen-dependent peptides in nuclear envelope preparations. In both the rat ventral prostate and an androgen-responsive cell line of the Shionogi mouse mammary carcinoma, we have identified abundant androgen-dependent peptides. The relationship of these peptides to the binding sites identified by the first two approaches and their role in steroid transport is being investigated.
为了确定甾体激素在前往基因组的途中穿过核膜的分子机制，人们采取了三种方法。第一种方法涉及对甾体与核膜制剂结合的特性进行表征。我们已经对雄激素与从大鼠腹侧前列腺、大鼠肝脏以及狮王小鼠乳腺癌的雄激素反应性和雄激素非反应性细胞系中分离出的核膜的结合进行了表征，以及糖皮质激素与大鼠肝脏的结合。在核膜上已鉴定出对甾体具有相对高亲和力的结合位点。重要的是，这些位点的数量和特异性与组织对甾体的反应性相关。在第二种方法中，我们已着手直接鉴定甾体结合位点。由于大鼠腹侧前列腺位点的特性类似于核雄激素受体的特性，我们已开始纯化该受体，并发现快速蛋白质液相色谱法非常有效。通过亲和标记研究，已确定大鼠肝脏核膜上的地塞米松结合位点是一种分子量约为90,000的肽。我们使用的第三种方法是在核膜制剂中鉴定雄激素依赖性肽。在大鼠腹侧前列腺和狮王小鼠乳腺癌的雄激素反应性细胞系中，我们都鉴定出了丰富 的雄激素依赖性肽。正在研究这些肽与前两种方法鉴定出的结合位点的关系及其在甾体转运中的作用。
