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J. Biol. Chem., Vol. 275, Issue 50, 39061-39072, December 15, 2000
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From the An evolutionarily conserved vertebrate homologue
of the Drosophila NK-3 homeodomain gene
bagpipe, Nkx3-1, is expressed in vascular and visceral
mesoderm-derived muscle tissues and may influence smooth muscle cell
differentiation. Nkx3-1 was evaluated for mediating smooth muscle
The Smooth Muscle
-Actin Gene Promoter Is a Molecular
Target for the Mouse bagpipe Homologue, mNkx3-1, and
Serum Response Factor*
§,
, and Department of Cellular and Molecular
Biology, Baylor College of Medicine, Houston, Texas 77030 and the
¶ Department of Structural and Cellular Biology, University of
South Alabama, Mobile, Alabama 36688
-actin (SMGA) gene activity, a specific marker of smooth muscle
differentiation. Expression of mNkx3-1 in heterologous CV-1 fibroblasts
was unable to elicit SMGA promoter activity but required the
coexpression of serum response factor (SRF) to activate robust SMGA
transcription. A novel complex element containing a juxtaposed
Nkx-binding site (NKE) and an SRF-binding element (SRE) in the proximal
promoter region was found to be necessary for the Nkx3-1/SRF
coactivation of SMGA transcription. Furthermore, Nkx3-1 and SRF
associate through protein-protein interactions and the homeodomain
region of Nkx3-1 facilitated SRF binding to the complex
NKE·SRE. Mutagenesis of Nkx3-1 revealed an inhibitory domain
within its C-terminal segment. In addition, mNkx3-1/SRF cooperative
activity required an intact Nkx3-1 homeodomain along with the MADS box
of SRF, which contains DNA binding and dimerization structural domains,
and the contiguous C-terminal SRF activation domain. Thus, SMGA is a
novel target for Nkx3-1, and the activity of Nkx3-1 on the SMGA
promoter is dependent upon SRF.
*
This work was supported in part by National Institutes of
Health Grants R01HL50422, P01HL49953 (to R. J. S.), 5P60HL38639, and
HL59956 (to W. E. Z.), United States Department of Agriculture Grant
9404341, and American Heart Association Grant AL-G-940003 (to
W. E. Z.).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 Cellular
and Molecular Biology, Baylor College of Medicine, One Baylor Plaza,
Houston, TX 77030. Tel.: 713-798-6649; Fax: 713-798-7799; E-mail:
schwartz@bcm.tmc.edu.
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