Helicobacter pylori infection of the gastric body induces transient hypochlorhydria and contributes to mucosal progression towards gastric carcinoma. Acid secretion is mediated by parietal cell H,K-ATPase whose catalytic subunit (HK) promoter activity in transfected gastric epithelial AGS cells is repressed by H. pylori through NF-B p50 homodimer binding to the promoter. IL-1, an acid secretory inhibitor whose mucosal level is increased by H. pylori, up-regulates HK promoter activity in AGS cells. Because IL-1 also activates NF-B signaling, we investigated disparate HK regulation by H. pylori and IL-1, testing the hypothesis that IL-1-induced HK promoter activation is mediated by the transcription factor Sp1. DNase I footprinting revealed Sp1 binding to HK promoter at -56 to -39 bp. IL-1 stimulated activity of three HK promoter constructs containing NF-B and Sp1 sites transfected into AGS cells, and also stimulated a construct containing only an Spl site. This stimulation was abrogated by mutating the HK promoter Sp1 binding site. Gelshift assays showed that IL-1 increased Sp1 but not p50 binding to cognate HK probes, and that Sp1 also interacts with an HK NF-B site when bound to it's cognate HK cis-response element. H. pylori did not augment Sp1 binding to an HK Sp1 probe, and siRNA-mediated knock-down of Sp1 expression abrogated IL-1-induced HK promoter stimulation. We conclude that IL-1 up-regulates HK gene transcription by inducing Sp1 binding to HK Sp1 and NF-B sites, and that H. pylori perturbation of HK gene expression is independent of Sp1-mediated basal HK transcription.