CANINE H-K+-ATPASE ALPHA-SUBUNIT GENE PROMOTER - STUDIES WITH CANINE PARIETAL-CELLS IN PRIMARY CULTURE()

H+-K+-adenosinetriphosphatase (H+-K+-ATPase) is the principal enzyme r esponsible for the process of gastric acid secretion. This enzyme is e xpressed in a cell-type-specific manner in gastric parietal cells. To explore the mechanisms regulating its expression, we transfected diffe rentiated canine parietal cells in primary culture with H+-K+-ATPase-l uciferase reporter genes and assessed transcriptional activities. Dele tional analysis of the 5'-flanking region of this gene demonstrated a remarkable increment in transcriptional activity associated with a seg ment between bases -54 to -45 (5' GCTCCGCCTC 3') relative to the trans criptional initiation site. Gel shift assays with competition and supe rshift analysis demonstrated that this segment is specifically bound b y the transcription factor Sp1.A point mutation, eliminating Sp1 bindi ng, diminished basal transcriptional activity by 80%, indicating that this Sp1 binding site is important for constitutive transcriptional ac tivity. Although these studies indicate that Sp1 is required to mainta in 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.