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J. Biol. Chem., Vol. 277, Issue 22, 19530-19537, May 31, 2002
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From the Department of Pathology and Immunology, Washington
University School of Medicine, St. Louis, Missouri 63110
Surfactant protein D (SP-D) plays roles in
pulmonary host defense and surfactant homeostasis and is increased
following acute lung injury. Given the importance of
CCAAT/enhancer-binding protein (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 promoters contribute to the
regulation of SP-D expression by C/EBPs. Five SP-D motifs (
Surfactant Protein D Gene Regulation
INTERACTIONS AMONG THE CONSERVED CCAAT/ENHANCER-BINDING PROTEIN
ELEMENTS*
432,
340, 319, 140, and 90) homologous to the C/EBP consensus
sequence 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/EBP
, C/EBP
, or C/EBP
to consensus oligomers.
Cotransfection of C/EBP, C/EBP, or C/EBP 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 deletion of sequences upstream of 205.
Independent site-directed mutagenesis of the sites at 432, 340, and
319 reduced C/EBP-mediated activation by ~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.
*
This work was supported by National Institutes of Health
Grants HL-44015 and HL-29594.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed: Dept. of Pathology and
Immunology, Barnes-Jewish Hospital, Rm. 2457, North Campus, Surgical
Pathology Mailstop 90-31-649, 216 S. Kingshighway Blvd., St.
Louis, MO 63110. Tel.: 314-454-8462; Fax: 314-454-5505; E-mail: crouch@path.wustl.edu.
Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.
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