AJP - Lung Cellular and Molecular Physiology, Vol 268, Issue 4 674-L682, Copyright © 1995 by American Physiological Society
Characterization of the promoter of human pulmonary surfactant protein B gene
V. C. Venkatesh, B. C. Planer, M. Schwartz, J. N. Vanderbilt, R. T. White and P. L. Ballard
Department of Pediatrics, University of Pennsylvania, Children's Hospital of Philadelphia 19104, USA.
Pulmonary surfactant protein B (SP-B) is required for normal surfactant
function and for survival at birth. To further study SP-B gene expression,
we sequenced genomic clones and examined promoter activity of SP-B DNA
fragments by transient transfection. A plasmid construct containing human
SP-B fragment -1039/+431 linked to chloramphenicol acetyltransferase (CAT)
reporter gene was readily expressed in H441 cells, which are derived from a
human lung adenocarcinoma, but was < 4% as active in Hep G2, HeLa, and
Calu 6 cell lines. SP-B promoter activity in H441 cells was orientation
dependent and increased by linked Rous sarcoma virus (RSV) enhancer and was
stronger than for thymidine kinase (tk) and RSV promoters. Deletional
mapping of the 5' flanking region with exonuclease III suggested
nonspecific negative (-811/-1039)- and positive (-453/-641)-control regions
and a cell-specific enhancer region at -208 to -54. When a fragment from
-403 to -35 base pairs (bp) was placed upstream or downstream of tkCAT, in
either orientation, expression in H441 cells but not other cell lines was
increased 4- to 28-fold relative to tkCAT. Deletional analysis of the 3'
terminus indicated a requirement for at least 7 bp 3' of the transcription
start site. Promoter activity was strongly inhibited in a dose-dependent
fashion by phorbol ester, with responsiveness mapped to bp -208/-54, but
was not responsive to glucocorticoid.(ABSTRACT TRUNCATED AT 250 WORDS)
