Gastrin gene expression is regulated by developmental cues, pH, and inflammation. These processes are mediated by various extracellular ligands, e.g., growth factors, cytokines, and neuropeptides that also stimulate c-fos gene expression. Therefore, to determine whether Fos is required for stimulation of the gastrin promoter, a c-fos sense expression vector was coexpressed with a gastrin reporter construct in a GH4 rat pituitary cell line. We found that epidermal growth factor (EGF) and tumor necrosis factor-alpha (TNF-alpha) transiently stimulate an increase in Fos protein that precedes stimulation of the gastrin promoter. However, the induction mediated by TNF-alpha was weaker than that mediated by EGF, indicating minimal overlap of the signaling pathways activated by EGF and TNF-alpha. Accordingly, overexpression of c-fos mRNA facilitated primarily EGF rather than TNF-alpha induction of the gastrin promoter. Expression of the c-fos gene in the absence of ligand did not stimulate the gastrin promoter. Thus c-fos gene expression is required but is not sufficient for induction of the gastrin promoter by EGF.