Characterization of the canine h+,k+-atpase [alpha]-subunit gene
promoter: studies with canine parietal cells in primary culture.
Muraoka, Akira, Mitsuru Kaise, Yi-Jun Guo, Junko Yamada, Il Song, John
Delvalle, Andrea Todisco, Tadataka Yamada.
Departments of Internal Medicine and Physiology, University of
Michigan, Medical Center, Ann Arbor, MI
APStracts 3:0141G, 1996.
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. In order 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 b 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 relie on other elements as Sp1 is an ubiquitously
expressed transcription factor.
Received 8 February 1996; accepted in final form 28 June 1996.
APS Manuscript Number G51-6.
Article publication pending Am. J. Physiol. (Gastrointest. Liver
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 25 July 1996