How do "Zn2 cys6" proteins distinguish between similar upstream activation sites? Comparison of the DNA-binding specificity of the GAL4 protein in vitro and in vivo.
The GAL4 protein of Saccharomyces cerevisiae is the prototype of a family of transcription factors that contain a "Zn2Cys6" coordination complex in the DNA-binding domain. GAL4 activates the transcription of genes involved in galactose and melibiose metabolism by binding to sites that contain one or more copies of a sequence 5'-CGGN5TN5CCG-3'. Other Zn2Cys6 proteins in S. cerevisiae also recognize sequences containing two CGG triplets, but with different spacings between them. In this report we investigate the mechanism by which GAL4 distinguishes its bona fide binding site from similar sequences as well as from bulk genomic DNA. In vitro, GAL4 recognizes with moderate to high affinity a variety of sites of the general formula (A/C)GGN10-12CCG. This level of specificity is apparently insufficient for the activator to carry out its biological role. However, many of the sites to which GAL4 binds in vitro do not support GAL4-activated transcription in vivo. In most cases there is not a quantitative correlation between the relative affinity of a site for GAL4 in vitro and the level of GAL4-dependent transcription supported by it in vivo. These data imply that there is some mechanism in vivo by which the intrinsic binding specificity of GAL4 is modified.
Vashee
S
S
Department of Chemistry and Biochemistry, University of Texas, Austin 78712.
Xu
H
H
Johnston
S A
SA
Kodadek
T
T
D001482:Base Composition; D001483:Base Sequence; D001665:Binding Sites; D003545:Cysteine; D004247:DNA / Q000138:chemical synthesis / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors; D014158:Transcription, Genetic; D015032:Zinc
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D004247:DNA; D015032:Zinc; D003545:Cysteine