The upstream region of the SP-B gene: intrinsic promoter activity and glucocorticoid responsiveness related to a new DNA-binding protein.
We identified and cloned the rabbit SP-B gene, encoding the pulmonary surfactant-associated protein, and sequenced its upstream region from -2635 to +428, including a much larger fragment of the upstream region than has previously been reported for an SP-B for any species. Rabbit SP-B showed substantial homology to its human counterpart in the coding and noncoding regions immediately upstream from the TATAA box. Using a luciferase (Luc) reporter gene (luc) construct we measured promoter activity with a 212-bp fragment (SPB212) from nucleotides (nt) -41 to -252, inclusive. SPB212 functioned as an active promoter in this assay. Further, we identified, cloned and sequenced the cDNA encoding a unique DNA-binding protein, N, that bound SPB212 at approx. -195. When the N cDNA was cloned into the expression vector pKC4 and cotransfected with the luc reporter construct, N significantly enhanced Luc production, but only in the presence of dexamethasone. Therefore, we identified and sequenced a functional promoter region upstream from rabbit SP-B, and isolated and characterized a DNA-binding protein that confers enhanced glucocorticoid responsiveness on this promoter.
Department of Pathology, Anatomy and Cell Biology, Jefferson Medical College, Philadelphia, PA 19107, USA.
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects*; D005796:Genes / Q000235:genetics*; D005938:Glucocorticoids / Q000494:pharmacology*; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011510:Proteolipids / Q000235:genetics*; D011663:Pulmonary Surfactants / Q000235:genetics*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid
D004268:DNA-Binding Proteins; C097169:DNA-binding protein N; D005938:Glucocorticoids; D011510:Proteolipids; D011663:Pulmonary Surfactants; D011993:Recombinant Fusion Proteins; D003907:Dexamethasone; D004247:DNA; D008156:Luciferases