Physical interaction between the MADS box of serum response factor and the TEA/ATTS DNA-binding domain of transcription enhancer factor-1.
Serum response factor is a MADS box transcription factor that binds to consensus sequences CC(A/T)(6)GG found in the promoter region of several serum-inducible and muscle-specific genes. In skeletal myocytes serum response factor (SRF) has been shown to heterodimerize with the myogenic basic helix-loop-helix family of factors, related to MyoD, for ... control of muscle gene regulation. Here we report that SRF binds to another myogenic factor, TEF-1, that has been implicated in the regulation of a variety of cardiac muscle genes. By using different biochemical assays such as affinity precipitation of protein, GST-pulldown assay, and coimmunoprecipitation of proteins, we show that SRF binds to TEF-1 both in in vitro and in vivo assay conditions. A strong interaction of SRF with TEF-1 was seen even when one protein was denatured and immobilized on nitrocellulose membrane, indicating a direct and stable interaction between SRF and TEF-1, which occurs without a cofactor. This interaction is mediated through the C-terminal subdomain of MADS box of SRF encompassing amino acids 204-244 and the putative 2nd and 3rd alpha-helix/beta-sheet configuration of the TEA/ATTS DNA-binding domain of TEF-1. In the transient transfection assay, a positive cooperative effect of SRF and TEF-1 was observed when DNA-binding sites for both factors, serum response element and M-CAT respectively, were intact; mutation of either site abolished their synergistic effect. Similarly, an SRF mutant, SRFpm-1, defective in DNA binding failed to collaborate with TEF-1 for gene regulation, indicating that the synergistic trans-activation function of SRF and TEF-1 occurs via their binding to cognate DNA-binding sites. Our results demonstrate a novel association between SRF and TEF-1 for cardiac muscle gene regulation and disclose a general mechanism by which these two super families of factors are likely to control diversified biological functions.
Mesh Terms:
Animals, Binding Sites, Blotting, Western, COS Cells, Cell Nucleus, Collodion, Conserved Sequence, DNA, DNA-Binding Proteins, Gene Expression, Glutathione Transferase, Models, Genetic, Myocardium, Nuclear Proteins, Plasmids, Precipitin Tests, Protein Binding, Protein Isoforms, Protein Structure, Tertiary, Recombinant Fusion Proteins, Serum Response Factor, Transcription Factors, Transfection
Animals, Binding Sites, Blotting, Western, COS Cells, Cell Nucleus, Collodion, Conserved Sequence, DNA, DNA-Binding Proteins, Gene Expression, Glutathione Transferase, Models, Genetic, Myocardium, Nuclear Proteins, Plasmids, Precipitin Tests, Protein Binding, Protein Isoforms, Protein Structure, Tertiary, Recombinant Fusion Proteins, Serum Response Factor, Transcription Factors, Transfection
J. Biol. Chem.
Date: Mar. 30, 2001
PubMed ID: 11136726
View in: Pubmed Google Scholar
Download Curated Data For This Publication
7750
Switch View:
- Interactions 3