Regulation of the alpha 1(I) collagen promoter via a transforming growth factor-beta activation element.
A transforming growth factor-beta (TGF-beta) activating element (TAE), with a nuclear factor-1 (NF-1)-like sequence, was previously located 1.6 kilobases upstream from the transcription start site in the alpha 1(I) collagen promoter (Ritzenthaler, J. D., Goldstein, R. H., Fine, A., Lichtler, A., Rowe, D. W., and Smith, B. D. (1991) Biochem. J. 280, 157-162). Double-stranded TAE, but not NF-1 consensus sequences, abrogated TGF-beta stimulation of co-transfected collagen promoter-chloramphenicol acetyltransferase constructs. Mutations in non-NF-1 binding sites, located by methylation interference, eliminated activity of the TAE oligonucleotide. However, TAE sequences failed to bind in vitro expressed NF-1 protein, to compete for NF-1-binding proteins, and to bind with protein which reacts with antibodies to NF-1 family of proteins. Within the TAE there was an activator protein 2 (AP-2) binding site. Although AP-2 protein bound to TAE, antibodies to AP-2 did not react with nuclear protein-TAE complexes. TAE bound to a 34,000-Da protein on Southwestern analysis. However, the UV-cross-linked TAE-nuclear protein complex was 82,000 Da. Finally, a dose-response study demonstrated that TGF-beta increased TAE nuclear binding proteins at lower doses with a different response curve than NF-1 nuclear binding proteins. Taken together these data demonstrated that TGF-beta functions in human lung fibroblasts to activate collagen transcription through TAE sites by protein complexes independent of NF-1 or AP-2 protein.
Ritzenthaler
J D
JD
Department of Biochemistry, Boston University Medical Center, Massachusetts.
Goldstein
R H
RH
Fine
A
A
Smith
B D
BD
D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis / Q000235:genetics*; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008969:Molecular Sequence Data; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000235:genetics; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D051840:Y-Box-Binding Protein 1
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D051840:Y-Box-Binding Protein 1; C070856:YBX1 protein, human; D003094:Collagen; D004247:DNA