Höcker M, Raychowdhury R, Plath T, Wu H, O'Connor D T, Wiedenmann B, Rosewicz S, Wang T C
Medizinische Klink mit Schwerpunkt Gastroenterologie und Hepatologie, Universitätsklinikum Charitè, Campus Virchow-Klinikum, Humboldt Universität Berlin, Germany.
J Biol Chem. 1998 Dec 18;273(51):34000-7. doi: 10.1074/jbc.273.51.34000.
Chromogranin A (CgA) is a multifunctional acidic protein that in the stomach is expressed predominantly in enterochromaffin-like cells (ECL cells) where it is regulated by gastrin. In order to investigate the transcriptional response of the mouse CgA (mCgA) promoter to gastrin stimulation, we studied a 4.8-kilobase mCgA promoter-luciferase reporter gene construct in transiently transfected AGS-B cells. 5'-Deletion analysis and scanning mutagenesis of mCgA 5'-flanking DNA showed that a Sp1/Egr-1 site spanning -88 to -77 base pairs (bp) and a cyclic AMP-responsive element (CRE) at -71 to -64 bp are essential for gastrin-dependent mCgA transactivation. Gastrin stimulation increased cellular Sp1 protein levels and Sp1-binding to the mCgA -88 to -77 bp element, as well as binding of CREB to its consensus motif at -71 to -64 bp. Gastrin also stimulated CREB Ser-133 phosphorylation, and abundance of cellular CREB protein levels. Overexpression of either Sp1 or phosphorylated CREB transactivated the mCgA promoter dose dependently, while coexpression of both transcription factors resulted in an additive mCgA promoter response. mCgA -92 to -64 bp, comprising the Sp1/Egr-1 site and the CRE motif, conferred gastrin responsiveness to a heterologous thymidine kinase promoter system, and therefore functions as a "true" enhancer element. This report demonstrates that Sp1 and CREB mediate CCK-B/gastrin receptor-dependent gene regulation, and that the effect of gastrin on the CgA gene is brought about by cooperative action of both transcription factors.
嗜铬粒蛋白A（CgA）是一种多功能酸性蛋白，在胃中主要表达于肠嗜铬样细胞（ECL细胞），并受胃泌素调节。为了研究小鼠CgA（mCgA）启动子对胃泌素刺激的转录反应，我们在瞬时转染的AGS - B细胞中研究了一个4.8千碱基的mCgA启动子 - 荧光素酶报告基因构建体。mCgA 5'侧翼DNA的5'缺失分析和扫描诱变表明，跨越 - 88至 - 77碱基对（bp）的Sp1 / Egr - 1位点和 - 71至 - 64 bp处的环磷酸腺苷反应元件（CRE）对于胃泌素依赖性mCgA反式激活至关重要。胃泌素刺激增加了细胞Sp1蛋白水平以及Sp1与mCgA - 88至 - 77 bp元件的结合，以及CREB与其在 - 71至 - 64 bp处的共有基序的结合。胃泌素还刺激了CREB丝氨酸133的磷酸化以及细胞CREB蛋白水平的丰度。Sp1或磷酸化CREB的过表达均以剂量依赖性方式反式激活mCgA启动子，而两种转录因子的共表达导致mCgA启动子反应的累加。包含Sp1 / Egr - 1位点和CRE基序的mCgA - 92至 - 64 bp赋予了胃泌素对异源胸苷激酶启动子系统的反应性，因此作为一个“真正的”增强子元件发挥作用。本报告表明，Sp1和CREB介导CCK - B /胃泌素受体依赖性基因调控，并且胃泌素对CgA基因的作用是由两种转录因子的协同作用实现的。
