Sp1 and CREB mediate gastrin-dependent regulation of chromogranin A promoter activity in gastric carcinoma cells

Chromogranin A (CgA) is a multifunctional acidic protein that in the stomac h is expressed predominantly in enterochromaffin-like cells (ECL cells) whe re it is regulated by gastrin, In order to investigate the transcriptional response of the mouse CgA (mCgA) promoter to gastrin stimulation, we studie d a 4.8-kilobase mCgA promoter-luciferase reporter gene construct in transi ently transfected AGS-B cells, 5'-Deletion analysis and scanning mutagenesi s 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 stimulati on 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 t o -64 bp. Gastrin also stimulated CREB Ser-133 phosphorylation, and abundan ce of cellular CREB protein levels. Overexpression of either Sp1 or phospho rylated CREB transactivated the mCgA promoter dose dependently, while coexp ression 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 mo tif, conferred gastrin responsiveness to a heterologous thymidine kinase pr omoter system, and therefore functions as a "true" enhancer element. This r eport demonstrates that Sp1 and CREB mediate CCK-B/gastrin receptor-depende nt gene regulation, and that the effect of gastrin on the CgA gene is broug ht about by cooperative action of both transcription factors.