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Papers In Press, published online ahead of print March 23, 2002
Dept.of Pathology, 90-31-649, Washington University School of Medicine, St. Louis, MO 63110
Corresponding Author: crouch{at}path.wustl.edu
Surfactant protein D plays roles in pulmonary host defense and surfactant homeostasis, and is increased following acute lung injury. Given the importance of C/EBP binding elements in the systemic acute phase response and lung development, and the expression of C/EBP isoforms by lung epithelial cells, we hypothesized that conserved C/EBP motifs in the near distal and proximal promoter, contribute to the regulation of SP-D expression by C/EBPs. Five SP-D motifs (-432, -340, -319, -140, and -90) homologous to the C/EBP consensus specifically bound to C/EBPs in gel shift assays, and four of the five sites (-432, -340,-319, and -90) efficiently competed for the binding of C/EBPalpha, C/EBPbeta, or C/EBPdelta to consensus oligomers. Co-transfection of CEBPalpha, C/EBPbeta, or C/EBPdelta cDNA in H441 lung adenocarcinoma cells significantly increased the luciferase activity of a wild-type SP-D promoter construct containing 698 bp of upstream sequence (SS698). Transfection of C/EBP also increased the level of endogenous SP-D mRNA in H441 cells. Transactivation of the reporter construct was abrogated by the deletion of sequences upstream of -205. Independent site-directed mutagenesis of the sites at -432, -340, and -319 reduced C/EBP-mediated activation by approximately 50%, and mutagenesis of the site at -432 in combination with either of the tandem sites at –340 and –319 blocked activation. The conserved AP-1 element at -109 was required for maximal promoter activity, but not for the transactivation of SS698 by C/EBPs. Thus, interactions among C/EBP elements in the near distal promoter can modulate the promoter activity of SP-D.
J. Biol. Chem, 10.1074/jbc.M201126200
Submitted on February 4, 2002
Revised on March 13, 2002
Accepted on March 22, 2002
Surfactant protein D (SP-D) gene regulation: Interactions among the conserved C/EBP elements
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