Wang K S, Hodgetts R B
Department of Biological Sciences, University of Alberta, Edmonton, Canada.
J Mol Endocrinol. 1998 Feb;20(1):129-39. doi: 10.1677/jme.0.0200129.
The alpha 2u-globulins are the major urinary proteins in adult male rats. They are encoded by a gene family, the expression of which is under multihormonal control in the liver. Glucocorticoids are positive regulatory hormones and we have analyzed the contribution of 5'-upstream sequences to the induction by dexamethasone of two cloned members of the family transfected into mouse L-cells. The results demonstrate that sequences from -762 bp to -226 bp of clone 91 are required for the 24-fold level of induction that was observed. Addition of 5.5 kb of upstream sequence beyond -762 bp did not alter the level of induction significantly, whereas deletion of the DNA between -762 bp and -226 bp reduced inducibility to about 4-fold. Sequencing of this region revealed that an element, 5'-AGAACAggtTTCAAA-3', similar to the 15 bp consensus glucocorticoid response element 5'-AGAACAnnnTGTACC-3', is situated 513 bp upstream of the transcription start site. We infer that this element or its left half site is necessary for the dexamethasone-induced expression of clone 91 from the observation that a second gene, clone 2, that contained a base substitution at position 5 in the left half site was not inducible. It now appears that at least three distinct cis-acting regulatory regions, all of which bind to the glucocorticoid receptor in vitro, may contribute to the full induction of clone 91 by dexamethasone. These are: the distal upstream region identified by this study, a proximal upstream region that binds not only the receptor but also alpha 2uNF1, a constitutively expressed nuclear protein required for induction and a region within the fourth intron that contains five tandem receptor binding sites.
α2u球蛋白是成年雄性大鼠尿液中的主要蛋白质。它们由一个基因家族编码，该基因家族的表达在肝脏中受多种激素的控制。糖皮质激素是正向调节激素，我们分析了5′上游序列对转染到小鼠L细胞中的该家族两个克隆成员受地塞米松诱导的作用。结果表明，克隆91中从-762 bp到-226 bp的序列是观察到的24倍诱导水平所必需的。在-762 bp以外添加5.5 kb的上游序列并没有显著改变诱导水平，而删除-762 bp和-226 bp之间的DNA则将诱导性降低到约4倍。对该区域进行测序发现，一个元件5′-AGAACAggtTTCAAA-3′，类似于15 bp的糖皮质激素反应元件共有序列5′-AGAACAnnnTGTACC-3′，位于转录起始位点上游513 bp处。我们从以下观察推断该元件或其左半位点对于地塞米松诱导克隆91的表达是必需的：第二个基因克隆2，其左半位点第5位存在碱基替换，是不可诱导的。现在看来，至少有三个不同的顺式作用调节区域，所有这些区域在体外都能与糖皮质激素受体结合，可能对地塞米松对克隆91的完全诱导有作用。这些区域是：本研究确定的远端上游区域、一个近端上游区域，该区域不仅能结合受体，还能结合α2uNF1，α2uNF1是诱导所需的一种组成型表达的核蛋白，以及第四个内含子内的一个区域，该区域包含五个串联的受体结合位点。
