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J. Biol. Chem., Vol. 276, Issue 24, 21686-21691, June 15, 2001
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From the Division of Pulmonary Biology, Children's Hospital
Medical Center, Cincinnati, Ohio 45229-3039
Surfactant protein B (SP-B) is a 79-amino acid
peptide critical to postnatal respiratory adaptation and is
developmentally regulated. Previous studies demonstrated that retinoic
acid receptors (RARs) and thyroid transcription factor 1 (TTF-1)
stimulated SP-B gene expression in respiratory epithelial cells.
Clustered retinoic acid-responsive element and TTF-1 binding sites were
identified in the enhancer region of the SP-B gene and were
required for retinoic acid stimulation of the human SP-B (hSP-B)
promoter. In addition, RAR and TTF-1 were colocalized in mouse
bronchiolar and alveolar type II epithelial cells, the cellular site of
SP-B synthesis. In the present studies, RAR and TTF-1 were colocalized in the nucleus of H441 cells. RAR and TTF-1 synergistically stimulated the hSP-B promoter in H441 cells. Direct protein-protein interactions between RAR and TTF-1 were demonstrated by the glutathione
S-transferase pull-down assay and the mammalian cell
two hybrid assay. Truncation/deletion studies showed that the RAR-TTF-1
interaction was mediated through the RAR DNA binding domain (DBD) and
the TTF-1 homeodomain. RAR DBD greatly enhanced TTF-1 homeodomain DNA
binding activity to a hSP-B enhancer oligonucleotide, in which retinoic
acid-responsive element and TTF-1 DNA binding sites overlap. Chromatin
immunoprecipitation assay demonstrated that retinoic acid treatment of
H441 cells greatly stimulated both RAR and TTF-1 DNA binding to the
hSP-B enhancer region in H441 cells. These findings support a
model in which RAR/retinoid X receptor, TTF-1, and coactivators (p160 members and CBP) form an enhanceosome in the enhancer region of the
hSP-B gene.
Protein-Protein Interaction of Retinoic Acid Receptor
and Thyroid Transcription Factor-1 in Respiratory Epithelial
Cells*
,
*
This work was supported by the NHLBI, National Institutes of
Health Grant HL-61803 and Specialized Center for Research Grant HL56387 (to C. Y.).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: Children's Hospital
Medical Center, Division of Pulmonary Biology, TCHRF, 3333 Burnet Ave.,
Cincinnati, OH 45229-3039. Tel.: 513-636-7990; Fax: 513-636-7868;
E-mail: Cong.Yan@chmcc.org.
Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.
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