Two monomers of yeast transcription factor ADR1 bind a palindromic sequence symmetrically to activate ADH2 expression.
ADR1 is a transcription factor from Saccharomyces cerevisiae that regulates ADH2 expression through a 22-bp palindromic sequence (UAS1). Size fractionation studies revealed that full-length ADR1 and a truncated ADR1 protein containing the first 229 amino acids, which has the complete DNA-binding domain, ADR1:17-229, exist as monomers in solution. However, two complexes were formed with target DNA-binding sites. UV-cross-linking studies suggested that these two complexes represent one and two molecules of ADR1 bound to DNA. Studies of ADR1 complexes formed with wild-type UAS1, asymmetrically altered UAS1, and one half of UAS1 showed that ADR1 can bind to one half of UAS1 and gives rise to a complex containing one molecule of ADR1. Dimethyl sulfate interference studies were consistent with this interpretation and in addition indicated that purine contact sites in each half of UAS1 were identical. Increasing the distance between the two halves of UAS1 had at most a minor effect of the thermodynamics of formation of the two complexes. These data are more consistent with ADR1 binding as two independent monomers, one to each half of UAS1. However, binding of two ADR1 monomers at UAS1 is apparently essential for transactivation in vivo. Further, we have identified a stretch of 18 amino acid residues amino terminal to the zinc two-finger domains of ADR1 which is essential for DNA-binding activity. Single amino acid substitutions of residues in this region resulted in severely reduced DNA-binding activity.
Thukral
S K
SK
Department of Biochemistry, University of Washington, Seattle 98195.
Eisen
A
A
Young
E T
ET
D000426:Alcohol Dehydrogenase / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D046911:Macromolecular Substances; D008745:Methylation; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry; D011485:Protein Binding; D012045:Regulatory Sequences, Nucleic Acid; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014466:Ultraviolet Rays
C074682:ADR1 protein, S cerevisiae; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D046911:Macromolecular Substances; D009841:Oligonucleotides; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D000426:Alcohol Dehydrogenase