Raychowdhury Raktima, Schäfer Georgia, Fleming John, Rosewicz Stefan, Wiedenmann Bertram, Wang Timothy C, Höcker Michael
Medizinische Klink mit Schwerpunkt Gastroenterologie, Hepatologie, Endokrinologie und Stoffwechsel, Universitätsklinikum Charité, Campus Virchow-Klinikum, Humboldt Universität, 13353 Berlin, Germany.
Mol Endocrinol. 2002 Dec;16(12):2802-18. doi: 10.1210/me.2001-0292.
Recently, binding of specific protein 1 (Sp1) and cAMP response element binding protein (CREB) to a GC-rich element at -92/-62 has been identified as a critical step in gastrin-dependent regulation of the chromogranin A (CgA) gene in gastric epithelial cells. Here we demonstrate that binding of early growth response protein 1 (Egr-1) to the distal part of the -92/-62 site is also required for gastrin-dependent CgA transactivation. Gastrin elevated cellular and nuclear Egr-1 levels in a time-dependent manner and also increased Egr-1 binding to the CgA -92/-73 region. Disruption of this site reduced gastrin responsiveness without influencing basal promoter activity, while loss of Sp1 and/or CREB binding sites diminished basal and gastrin-stimulated CgA promoter activity. Ectopic Egr-1 overexpression potently stimulated the CgA promoter, whereas coexpression of Egr-1 with Sp1 and/or CREB resulted in additive effects. Functional analysis of Sp1-, Egr-1-, or CREB-specific promoter mutations in transfection studies confirmed the tripartite organization of the CgA -92/-62 element. Signaling studies revealed that MAPK kinase 1 (MEK1)/ERK1/2 cascades are critical for gastrin-dependent Egr-1 protein accumulation as well as Egr-1 binding to the CgA promoter. Our studies for the first time identify Egr-1 as a nuclear target of gastrin and show that functional interplay of Egr-1, Sp1, and CREB is indispensable for gastrin-dependent CgA transactivation in gastric epithelial cells.
最近，已确定特异性蛋白1（Sp1）和环磷酸腺苷反应元件结合蛋白（CREB）与位于-92 / -62的富含GC的元件结合是胃上皮细胞中胃泌素依赖性调节嗜铬粒蛋白A（CgA）基因的关键步骤。在此，我们证明胃泌素依赖性CgA反式激活也需要早期生长反应蛋白1（Egr-1）与-92 / -62位点的远端部分结合。胃泌素以时间依赖性方式提高细胞和核Egr-1水平，并且还增加Egr-1与CgA -92 / -73区域的结合。该位点的破坏降低了胃泌素反应性，而不影响基础启动子活性，而Sp1和/或CREB结合位点的缺失则降低了基础和胃泌素刺激的CgA启动子活性。异位Egr-1过表达有力地刺激了CgA启动子，而Egr-1与Sp1和/或CREB的共表达产生了累加效应。转染研究中Sp1、Egr-1或CREB特异性启动子突变的功能分析证实了CgA -92 / -62元件的三方组织。信号研究表明，丝裂原活化蛋白激酶1（MEK1）/细胞外信号调节激酶1/2（ERK1/2）级联对于胃泌素依赖性Egr-1蛋白积累以及Egr-1与CgA启动子的结合至关重要。我们的研究首次将Egr-1鉴定为胃泌素的核靶点，并表明Egr-1、Sp1和CREB的功能相互作用对于胃上皮细胞中胃泌素依赖性CgA反式激活是必不可少的。
