James R M, Neil C, Webster J, Roos S, Clark A J, Whitelaw C B
Division of Molecular Biology, Roslin Institute (Edinburgh), Midlothian, United Kingdom.
Biochem Biophys Res Commun. 2000 May 27;272(1):284-9. doi: 10.1006/bbrc.2000.2766.
Increasing the number of transcription factor binding sites within a construct can enhance expression. In an attempt to create a synthetic locus control region for mammary expression, we have generated beta-lactoglobulin-reporter constructs with multiple copies of the cluster of transcription sites normally located within the proximal promoter. These constructs were functionally tested by stable transfection of mammary epithelial cells in vitro and in transgenic mice in vivo. Rather than enhancing expression, multimerisation of the promoter region acted neither in vivo nor in vitro to enhance expression. Indeed, its presence reduced expression. This failure to enhance expression was reflected in the inability of this region to form a DNaseI hypersensitive site autonomously in mammary chromatin in vivo. It is implicit from our study that not all combinations of transcription factor binding sites will enhance transcription.
增加构建体中转录因子结合位点的数量可增强表达。为了创建一个用于乳腺表达的合成基因座控制区，我们构建了带有通常位于近端启动子内的转录位点簇多个拷贝的β-乳球蛋白报告基因构建体。通过体外稳定转染乳腺上皮细胞和体内转基因小鼠对这些构建体进行了功能测试。启动子区域的多聚化非但没有增强表达，在体内和体外均未起到增强表达的作用。实际上，它的存在降低了表达。这种增强表达的失败体现在该区域在体内乳腺染色质中无法自主形成DNaseI超敏位点。我们的研究暗示并非所有转录因子结合位点的组合都会增强转录。
