CIS-ACTIVE ELEMENTS CONTROLLING LUNG CELL-SPECIFIC EXPRESSION OF HUMAN PULMONARY SURFACTANT PROTEIN-B GENE

Human surfactant protein B (SPB) is a 79-aminoacid 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 adenocarcin oma cell line, H441, was used in both functional and biochemical assay s to identify DNA sequences controlling lung cell-specific expression of the SPB gene. DNase I hypersensitive studies demonstrated two disti nct regions of lung cell-specific hypersensitivity located proximal to the SPB promoter and within the eighth intron of the gene. To functio nally define these DNA sequences, a series of plasmid vectors were con structed in which segments of the human SPB gene and 5'-flanking seque nce were linked to a CAT reporter gene and assayed for expression in l ung and nonlung cell lines. Whereas far upstream and intronic sequence s did not contain enhancer-like elements, a 259-base pair DNA segment (base pair -218 to +41) was sufficient to support lung cell-specific e xpression. DNase I footprinting demonstrated that this pulmonary epith elial cell-specific promoter fragment contained five nuclear protein-b inding sites, two of which bound lung cell-specific nuclear protein co mplexes. These results suggest that the pulmonary epithelial cell-spec ific expression of SPB is determined, in part, by both ubiquitous and cell type-specific protein-DNA interactions within the proximal promot er region.