Merchant J L, Shiotani A, Mortensen E R, Shumaker D K, Abraczinskas D R
Department of Internal Medicine, University of Michigan, Ann Arbor.
J Biol Chem. 1995 Mar 17;270(11):6314-9. doi: 10.1074/jbc.270.11.6314.
Growth factors coordinately regulate a variety of different genes to stimulate cellular proliferation. In the stomach, gastrin, epidermal growth factor (EGF), and transforming growth factor-alpha all mediate gastric mucosal homeostasis by promoting cell renewal. We have previously shown that EGF and phorbol esters stimulate the human gastrin promoter through a novel GC-rich DNA element 5'-(68)GGGGCGGGGTGGGGGG-53 called gERE (gastrin EGF response element). In this report, we show that three factors bind to this element, the transcription factor Sp1 and two fast migrating complexes designated gastrin EGF response proteins (gERP 1 and 2). To understand how these factors bind and confer EGF responsiveness, mutations of gERE were tested in vitro for protein binding and in vivo for promoter activation. Both gel shift assays and UV cross-linking studies revealed that the factors bind to overlapping domains, Sp1 to the 5' half-site and gERP 1 and 2 to the 3' half-site. Placing either the 5' or 3' mutations upstream of a minimal gastrin promoter abolished EGF induction. Therefore both the 5' and 3' domains were required to confer EGF induction. Collectively, these results demonstrate that complex interactions between Sp1 and other factors binding to overlapping gERE half-sites confer EGF responsiveness to the gastrin promoter.
生长因子协同调节多种不同基因以刺激细胞增殖。在胃中，胃泌素、表皮生长因子（EGF）和转化生长因子-α均通过促进细胞更新来介导胃黏膜稳态。我们之前已经表明，EGF和佛波酯通过一个名为gERE（胃泌素EGF反应元件）的富含GC的新型DNA元件5'-(68)GGGGCGGGGTGGGGGG-53刺激人胃泌素启动子。在本报告中，我们表明有三种因子与该元件结合，即转录因子Sp1和两种快速迁移的复合物，称为胃泌素EGF反应蛋白（gERP 1和2）。为了了解这些因子如何结合并赋予EGF反应性，我们在体外测试了gERE的突变体与蛋白质的结合情况，并在体内测试了其对启动子激活的作用。凝胶迁移实验和紫外线交联研究均表明，这些因子结合于重叠区域，Sp1结合于5'半位点，gERP 1和2结合于3'半位点。将5'或3'突变置于最小胃泌素启动子上游会消除EGF诱导作用。因此，5'和3'区域对于赋予EGF诱导作用都是必需的。总的来说，这些结果表明，Sp1与其他结合于重叠gERE半位点的因子之间的复杂相互作用赋予了胃泌素启动子对EGF的反应性。
