A multifunctional DNA-binding protein that promotes the formation of serum response factor/homeodomain complexes: identity to TFII-I.
The human homeodomain protein Phox1 interacts functionally with serum response factor (SRF) to impart serum responsive transcriptional activity to SRF-binding sites in a HeLa cell cotransfection assay. However, stable ternary complexes composed of SRF, Phox1, and DNA, which presumably mediate the transcriptional effects of Phox1 in vivo, have not been observed in vitro. Here, we report the identification, purification, and molecular cloning of a human protein that promotes the formation of stable higher-order complexes of SRF and Phox1. We show that this protein, termed SPIN, interacts with SRF and Phox1 in vitro and in vivo. Furthermore, SPIN binds specifically to multiple sequences in the c-fos promoter and interacts cooperatively with Phox1 to promote serum-inducible transcription of a reporter gene driven by the c-fos serum response element (SRE). SPIN is identical to the initiator-binding protein TFII-I. Consistent with this hypothesis, SPIN exhibits modest affinity for a characterized initiator sequence in vitro. We propose that this multifunctional protein coordinates the formation of an active promoter complex at the c-fos gene, including the linkage of specific signal responsive activator complexes to the general transcription machinery.
Grueneberg
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DA
ARIAD Pharmaceuticals, Cambridge, Massachusetts 02139, USA. gruenebe@ariad.com
Henry
R W
RW
Brauer
A
A
Novina
C D
CD
Cheriyath
V
V
Roy
A L
AL
Gilman
M
M
D000595:Amino Acid Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D015342:DNA Probes; D018076:DNA, Complementary; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D016762:Genes, fos / Q000235:genetics; D006367:HeLa Cells; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D026362:Serum Response Factor; D014157:Transcription Factors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*
D015342:DNA Probes; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; C082157:PRRX1 protein, human; D011994:Recombinant Proteins; D026362:Serum Response Factor; D014157:Transcription Factors; D004247:DNA