Canine H(+)-K(+)-ATPase alpha-subunit gene promoter: studies with canine parietal cells in primary culture

Abstract

H(+)-K(+)-adenosinetriphosphatase (H(+)-K(+)-ATPase) is the principal enzyme responsible for the process of gastric acid secretion. This enzyme is expressed in a cell-type-specific manner in gastric parietal cells. To explore the mechanisms regulating its expression, we transfected differentiated canine parietal cells in primary culture with H(+)-K(+)-ATPase-luciferase reporter genes and assessed transcriptional activities. Deletional analysis of the 5'-flanking region of this gene demonstrated a remarkable increment in transcriptional activity associated with a segment between bases -54 to -45 (5' GCTCCGCCTC 3') relative to the transcriptional initiation site. Gel shift assays with competition and supershift analysis demonstrated that this segment is specifically bound by the transcription factor Sp1. A point mutation, eliminating Sp1 binding, diminished basal transcriptional activity by 80%, indicating that this Sp1 binding site is important for constitutive transcriptional activity. Although these studies indicate that Sp1 is required to maintain a high concentration of the H(+)-K(+)-ATPase gene in the parietal cell, its cell-type-specific expression must rely on other elements because Sp1 is a ubiquitously expressed transcription factor.