Cis-active elements controlling lung cell-specific expression of human pulmonary surfactant protein B gene.
Human surfactant protein B (SPB) is a 79-amino-acid hydrophobic protein that enhances the surface active properties of pulmonary surfactant. SPB is expressed in nonciliated bronchiolar and alveolar type II cells of the respiratory epithelium, and its expression increases markedly late in gestation. In the present study, a human pulmonary adenocarcinoma cell line, H441, was used in both functional and biochemical assays to identify DNA sequences controlling lung cell-specific expression of the SPB gene. DNase I hypersensitive studies demonstrated two distinct regions of lung cell-specific hypersensitivity located proximal to the SPB promoter and within the eighth intron of the gene. To functionally define these DNA sequences, a series of plasmid vectors were constructed in which segments of the human SPB gene and 5'-flanking sequence were linked to a CAT reporter gene and assayed for expression in lung and nonlung cell lines. Whereas far upstream and intronic sequences did not contain enhancer-like elements, a 259-base pair DNA segment (base pair -218 to +41) was sufficient to support lung cell-specific expression. DNase I footprinting demonstrated that this pulmonary epithelial cell-specific promoter fragment contained five nuclear protein-binding sites, two of which bound lung cell-specific nuclear protein complexes. These results suggest that the pulmonary epithelial cell-specific expression of SPB is determined, in part, by both ubiquitous and cell type-specific protein-DNA interactions within the proximal promoter region.
Division of Pulmonary Biology, Children's Hospital Medical Center, Cincinnati, Ohio 45229-2899.
R J
RJ
Bohinski
J A
JA
Huffman
J A
JA
Whitsett
D L
DL
Lattier
D000230:Adenocarcinoma; D001483:Base Sequence; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D015342:DNA Probes; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008168:Lung / Q000378:metabolism*; D008175:Lung Neoplasms; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods; D011401:Promoter Regions, Genetic; D011510:Proteolipids / Q000235:genetics*; D011663:Pulmonary Surfactants / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism; D015183:Restriction Mapping; D017384:Sequence Deletion; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D015342:DNA Probes; D009838:Oligodeoxyribonucleotides; D011510:Proteolipids; D011663:Pulmonary Surfactants; D011994:Recombinant Proteins; D015500:Chloramphenicol O-Acetyltransferase