Activation mechanism of the multifunctional transcription factor repressor-activator protein 1 (Rap1p).

AUTOR(ES)

Drazinic, C M

RESUMO

Transcriptional activation in eukaryotic organisms normally requires combinatorial interactions of multiple transcription factors. In most cases, the precise role played by each transcription factor is not known. The upstream activating sequence (UAS) elements of glycolytic enzyme genes in Saccharomyces cerevisiae are excellent model systems for the study of combinatorial interactions. The yeast protein known as Rap1p acts as both a transcriptional repressor and an activator, depending on sequence context. Rap1p-binding sites are found adjacent to Gcr1p-binding sites in the UAS elements of glycolytic enzyme genes. These UAS elements constitute some of the strongest activating sequences known in S. cerevisiae. In this study, we have investigated the relationship between Rap1p- and Gcr1p-binding sites and the proteins that bind them. In vivo DNA-binding studies with rap1ts mutant strains demonstrated that the inability of Rap1p to bind at its site resulted in the inability of Gcr1p to bind at adjacent binding sites. Synthetic oligonucleotides, modeled on the UAS element of PYK1, in which the relative positions of the Rap1p- and Gcr1p-binding sites were varied prepared and tested for their ability to function as UAS elements. The ability of the oligonucleotides to function as UAS elements was dependent not only on the presence of both binding sites but also on the relative distance between the binding sites. In vivo DNA-binding studies showed that the ability of Rap1p bind its site was independent of Gcr1p but that the ability of Gcr1p to bind its site was dependent on the presence of an appropriately spaced and bound Rap1p-binding site. In vitro binding studies showed Rap1p-enhanced binding of Gcr1p on oligonucleotides modeled after the native PYK1 UAS element but not when the Rap1p- and Gcr1p-binding sites were displaced by 5 nucleotides. This work demonstrates that the role of the Rap1p in the activation of glycolytic enzyme genes is to bind in their UAS elements and to facilitate the binding of Gcr1p at adjacent binding sites.

Documentos Relacionados

• Interaction of yeast repressor-activator protein Ume6p with glycogen synthase kinase 3 homolog Rim11p.
• Separation of transcriptional activation and silencing functions of the RAP1-encoded repressor/activator protein 1: isolation of viable mutants affecting both silencing and telomere length.
• Cloning and characterization of tfdS, the repressor-activator gene of tfdB, from the 2,4-dichlorophenoxyacetic acid catabolic plasmid pJP4.
• The catabolite repressor/activator (Cra) protein of enteric bacteria.
• Repression by SSN6-TUP1 is directed by MIG1, a repressor/activator protein.