Kim Y K, Lee A S
Department of Biochemistry, University of Southern California School of Medicine, Los Angeles 90033.
J Biol Chem. 1992 Feb 5;267(4):2723-7.
When quiescent cells are stimulated to enter the cell cycle, the thymidine kinase (TK) gene is transcriptionally activated at the border of G1 and S. In this report we show that the human TK promoter contains multiple protein-binding sites. By site-directed mutagenesis, we identified a protein-binding site on the human TK promoter required for conferring G1-S-regulated transcription to a heterologous promoter and dissociated it functionally from an adjacent protein-binding domain containing an inverted CCAAT motif required for high basal level expression. Substitution-mutation of this site results in constitutive expression of the neo reporter gene in serum-stimulated fibroblasts, as well as in cells arrested in mid-G1 by a temperature-sensitive mutation. The regulatory domains for the human TK promoter exhibit interesting symmetrical features, including a set of CCAAT motifs and sites similar to the novel Yi protein-binding site recently discovered in the mouse TK promoter.
当静止细胞受到刺激进入细胞周期时，胸苷激酶（TK）基因在G1期和S期交界处被转录激活。在本报告中，我们表明人类TK启动子包含多个蛋白质结合位点。通过定点诱变，我们在人类TK启动子上鉴定出一个蛋白质结合位点，该位点是将G1-S期调控转录赋予异源启动子所必需的，并在功能上使其与一个相邻的蛋白质结合结构域分离，该结构域包含高基础水平表达所需的反向CCAAT基序。该位点的取代突变导致新霉素报告基因在血清刺激的成纤维细胞中组成性表达，以及在因温度敏感突变而停滞在G1期中期的细胞中组成性表达。人类TK启动子的调控结构域呈现出有趣的对称特征，包括一组CCAAT基序和与最近在小鼠TK启动子中发现的新型Yi蛋白结合位点相似的位点。
