The Orientation of the AP-1 Heterodimer on DNA Strongly Affects Transcriptional Potency

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 95; no. 24; pp. 14076 - 14081
• Main Authors: Chytil, Milan, Peterson, Blake R., Erlanson, Daniel A., Verdine, Gregory L.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 24.11.1998; National Acad Sciences; National Academy of Sciences; The National Academy of Sciences
• Subjects: Architecture; Arginine; Base Sequence; Binding Sites; Biochemistry; Biological Sciences; Deoxyribonucleic acid; Dimerization; DNA; DNA - chemistry; DNA - genetics; DNA - metabolism; Dyadic relations; Genetic mutation; Genetics; Isomers; Leucine Zippers; Models, Molecular; Mutation; Nucleic Acid Conformation; Oligodeoxyribonucleotides - chemistry; Point Mutation; Protein Multimerization; Protein Structure, Secondary; Proteins; Proto-Oncogene Proteins c-fos - chemistry; Proto-Oncogene Proteins c-fos - metabolism; Proto-Oncogene Proteins c-jun - chemistry; Proto-Oncogene Proteins c-jun - metabolism; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Stereochemistry; Transcription Factor AP-1 - chemistry; Transcription Factor AP-1 - metabolism; Transcription, Genetic; Transcriptional activation; Yeasts
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.95.24.14076

Activation of gene transcription in eukaryotes requires the cooperative assembly of an initiation complex containing many protein subunits. The necessity that these components contact each other and the promoter/enhancer in defined ways suggests that their spatial arrangement might influence the activation response. Indeed, growing evidence indicates that DNA architecture can profoundly affect transcriptional potency. Much less is known about the influence of protein architecture on transcriptional activation. Here, we examine the architectural dependence of activator function through the analysis of matched pairs of AP-1$\boldsymbol{\cdot}$DNA complexes differing only in their orientation. Mutation of a critical Arg residue in the basic-leucine zipper domain of either Fos or Jun yielded single point-mutant heterodimers that bind DNA in a single defined orientation, as determined directly by native chemical ligation/affinity cleavage; by contrast, the corresponding wild-type protein binds DNA as a roughly equal mixture of two isomeric orientations, which are related by subunit interchange. The stereochemistry of the point-mutant heterodimers could be switched by inversion of a C$\boldsymbol{\cdot}$G base pair in the center of the AP-1 site, thus providing access to both fixed orientational isomers. Yeast reporter gene assays consistently revealed that one orientational isomer activates transcription at least 10-fold more strongly than the other. These results suggest that protein architecture, especially the spatial relationship of the activation domain to the promoter, can exert a powerful influence on activator potency.