title: DNA binding specificity determinants in MADS-box transcription factors.
abstract: The MADS box is a conserved sequence motif found in the DNA binding domain of a family of transcription factors which possess related but distinct DNA binding specificities. We investigated the basis of differential sequence recognition by the MADS-box proteins serum response factor (SRF), MCM1, and MEF2A, using chimeric proteins and site-directed mutants in conjunction with gel mobility shift and binding site selection assays. Deletion of sequences immediately N terminal to the SRF MADS box alters its preferred binding site to that of MEF2A, although the resulting protein still weakly binds SRF-specific sites: exclusive binding to MEF2 sites requires further mutations, at MADS-box residues 11 to 15. In contrast to SRF, the sequence specificity of MCM1 (and of MEF2A) is determined entirely by sequences within its MADS box, and mutation of only SRF MADS-box residue 1 is sufficient to alter its binding specificity to that of MCM1. However, changes at both MADS-box positions 1 and 11 to 15 are necessary and sufficient to alter the specificity of the MCM1 MADS box to that of MEF2, and vice versa. The role of SRF MADS-box residues which differ from those present in the other proteins was investigated by selection of functional SRF variants in yeast cells. SRF MADS-box position 1 was always a glycine in the variants, but many different sequences at the other nonconserved MADS-box residues were compatible with efficient DNA binding. We discuss potential mechanisms of DNA recognition by MADS-box proteins.
authors.affiliation: Transcription Laboratory, Imperial Cancer Research Fund Laboratories, London, United Kingdom.
authors.forename: S J
authors.initials: SJ
authors.lastname: Nurrish
authors.forename: R
authors.initials: R
authors.lastname: Treisman
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D016384:Consensus Sequence; D004247:DNA / Q000378:metabolism*; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D014644:Genetic Variation; D064326:MEF2 Transcription Factors; D026341:Minichromosome Maintenance 1 Protein; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018006:Myogenic Regulatory Factors; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012641:Selection, Genetic; D026362:Serum Response Factor; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D064326:MEF2 Transcription Factors; D026341:Minichromosome Maintenance 1 Protein; D018006:Myogenic Regulatory Factors; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D026362:Serum Response Factor; D014157:Transcription Factors; D004247:DNA