Trefoil factor 2 (TFF2)/spasmolytic polypeptide (SP) is a highly stable peptide which is abundantly expressed and secreted by mucous cells of the stomach and which functions in gastric cytoprotection. Previous studies from our group have shown that TFF2 is an immediate early gene capable of regulating its own expression through activation of the TFF2 promoter. We therefore aimed to investigate the cis-acting elements mediating this response in AGS cells transfected with TFF2 promoter-reporter gene constructs, using a TFF2-expression system resembling physiologic paracrine conditions. TFF2 peptide expression was achieved through stable transfection of AGS cells with a TFF2-expression construct. Stimulation of transiently transfected cells with this TFF2-containing conditioned media resulted in a significant increase in TFF2 promoter activity. Promoter stimulation was blocked by an anti-TFF2 antibody, indicating that it was mediated specifically by TFF2. Deletion analysis of the TFF2 promoter led to the identification of a specific response element located between - 191 and - 174 upstream of the transcriptional initiation site. This region of the promoter. which was designated SPRE (for spasmolytic polypeptide response element), was sufficient to confer responsiveness in a heterologous promoter system. Mutational analysis and electrophoretic mobility shift assays (EMSA) showed that a GAG motif was responsible for mediating promoter activation in response to TFF2 stimulation, Since auto- and cross-induction of TFF2 promoter is likely to be a means of rapid amplification of TFF2 expression in the critical first minutes following mucosal injury, these results should lead to insight into the molecular events initiating epithelial restitution and healing. (C) 2002 Elsevier Science (USA). All rights reserved.