Sequence specific protein binding to and activation of the TGF-beta 3 promoter through a repeated TCCC motif.
We have previously characterized the TGF-beta 3 promoter and shown that the activity of this promoter is highly variable in different cell types. Although the promoter contains a proximal cAMP responsive element, which is critical to basal and forskolin-induced promoter activity, this element is not responsible for the variable, cell-specific regulation of the promoter. In this paper, we identify a 25 base pair sequence in the proximal region of the TGF-beta 3 promoter that binds a novel DNA-binding protein. This region includes the sequence T-CCCTCCCTCCC, (3 x TCCC), and mutation of these T-CCC repeats inhibits protein binding. Further, we show that in the cell line A375, which we have previously shown expresses high levels of TGF-beta 3 mRNA, this region is responsible for mediating high level TGF-beta 3 promoter activity. Immediately 3' to the 3 x TCCC sequence is a consensus AP-2 binding site, however, we show that this region does not bind AP-2, and AP-2 does not transactivate the TGF-beta 3 promoter. Therefore, we provide strong evidence that high level expression of TGF-beta 3 in A375 cells results from transactivation of the TGF-beta 3 promoter by a protein that binds to a repeated TCCC motif in the promoter and suggest that this DNA-binding protein likely also regulates aspects of developmental and tissue-specific expression of this cytokine.
Lafyatis
R
R
Laboratory of Chemoprevention, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.
Denhez
F
F
Williams
T
T
Sporn
M
M
Roberts
A
A
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D004247:DNA; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004330:Drosophila; D029721:Drosophila Proteins; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation; D016212:Transforming Growth Factor beta / Q000235:genetics*
D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; C493674:TfAP-2 protein, Drosophila; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D004247:DNA