Specific Protein Binding to Far Upstream Activating Sequences in Polymerase II Promoters

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 82; no. 1; pp. 43 - 47
• Main Authors: Bram, Richard J., Kornberg, Roger D.
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01-01-1985; National Acad Sciences
• Subjects: Base Sequence; Biological and medical sciences; Cell extracts; Cellulose nitrate; Deoxyribonuclease I; DNA; DNA, Fungal - genetics; DNA-Binding Proteins - genetics; Fundamental and applied biological sciences. Psychology; Galactose - genetics; Gels; Gene Expression Regulation; Genes; Genes, Fungal; Genetic vectors; Molecular and cellular biology; Molecular genetics; Oligonucleotides; Plasmids; Promoter regions; Promoter Regions, Genetic; promoters; Protein Binding; RNA Polymerase II - genetics; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Transcription Factors - genetics; Transcription. Transcription factor. Splicing. Rna processing; Yeasts; Binding protein; Regulation(control); Yeast; Transcription promotor; Transcription; Enzyme; Transferase; RNA polymerase 2; Activation; Saccharomyces cerevisiae
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.82.1.43

A binding activity specific for the upstream activating sequence of the GAL1-GAL10 promoter of Saccharomyces cerevisiae has been purified 220-fold on the basis of a nitrocellulose filter-binding assay. The binding activity is enriched in a nuclear preparation and is likely to be the GAL4 gene product. DNase I-protection mapping patterns reveal binding to two 30-base-pair regions at the boundaries of the sequence. A nearly identical mapping pattern is obtained with the coordinately regulated GAL7 promoter. The four 30-base-pair regions of binding in the two promoters are closely homologous, with a core consensus sequence of C-G-G
C-$_{\text{G}}^{\text{T}}$-C-A-A-C-A-G-T-G-C-T-C-C-G-A-A-$_{\text{C}}^{\text{G}}$-G-A-T. A synthetic oligonucleotide with such a sequence competes with the upstream activating sequence in the binding reaction.