Ko M S, Nakauchi H, Takahashi N
Fursawa MorphoGene Project, Research Development Corporation of Japan, Tsukuba.
EMBO J. 1990 Sep;9(9):2835-42. doi: 10.1002/j.1460-2075.1990.tb07472.x.
Glucocorticoid hormones induce the transcription of genes having glucocorticoid response elements in a dose dependent manner. To determine whether this dose dependence represents a response of individual templates or of the mass of templates, we introduced a bacterial beta-galactosidase gene linked to the glucocorticoid-inducible enhancer/promoter of the mouse mammary tumor virus (MTV) into Ltk- cells and obtained stable transformants containing a single or a few templates per cell. Visual inspection and flow cytometry analysis by enzyme histochemistry assay for beta-galactosidase revealed that individual cells showed very heterogeneous beta-galactosidase activity after 48 h induction with dexamethasone. When the glucocorticoid concentration was increased, an increasing cell population producing beta-galactosidase was observed. These phenomena were probably not due to heterogeneity of template copy number or to a predetermined cellular state among individual cells, since cells forming a single small colony gave similar results. This was also supported by data showing that recloned cells retained both their responsiveness to the glucocorticoid hormone and their digestion pattern in Southern blotting analyses. These results indicate that the dose dependent increase of glucocorticoid-inducible gene expression is caused by an increase in the number of transcriptionally active templates.
糖皮质激素以剂量依赖的方式诱导具有糖皮质激素反应元件的基因转录。为了确定这种剂量依赖性是代表单个模板还是大量模板的反应，我们将与小鼠乳腺肿瘤病毒（MTV）的糖皮质激素诱导型增强子/启动子相连的细菌β-半乳糖苷酶基因导入Ltk-细胞，并获得每个细胞含有单个或少数几个模板的稳定转化体。通过β-半乳糖苷酶的酶组织化学测定进行的目视检查和流式细胞术分析表明，在用地塞米松诱导48小时后，单个细胞显示出非常异质的β-半乳糖苷酶活性。当糖皮质激素浓度增加时，观察到产生β-半乳糖苷酶的细胞群体增加。这些现象可能不是由于模板拷贝数的异质性或单个细胞之间预先确定的细胞状态，因为形成单个小菌落的细胞给出了相似的结果。这也得到了数据的支持，这些数据表明在Southern印迹分析中，再克隆的细胞保留了它们对糖皮质激素的反应性和它们的消化模式。这些结果表明，糖皮质激素诱导型基因表达的剂量依赖性增加是由转录活性模板数量的增加引起的。
