A Reb1p-binding site is required for efficient activation of the yeast RAP1 gene, but multiple binding sites for Rap1p are not essential.
The Saccharomyces cerevisiae RAP1 protein (Rap1p) is a key multifunctional transcription factor. Using gel retardation analysis, four binding sites for Rap1p have been identified within the promoter of the RAP1 gene. These sites are located downstream of a binding site for the transcription factor Reb1p. The Reb1p site and an associated AT-rich region are important for transcriptional activation, but deletion of three of the Rap1p-binding sites had little effect on promoter activity. The activity of the RAP1 promoter has been analysed in a yeast strain (YDS410) that contains a temperature-sensitive mutation in the RAP1 gene. This mutation renders the DNA-binding activity of Rap1p temperature dependent. When YDS410 was grown at a semi-permissive temperature (30 degrees C), the activity of the RAP1 promoter increased by approximately 170%, compared with the same strain grown at the permissive temperature (25 degrees C). A RAP1 promoter in which three of the four Rap1p-binding sites had been deleted, showed only a small increase in activity in the same experiment. These data confirm that Rap1p is not required for activation of the RAP1 gene, and suggest a role for Rap1p in negative autoregulation.
Graham
I R
IR
Department of Genetics, University of Nottingham, Queen's Medical Centre, UK.
Chambers
A
A
D001483:Base Sequence; D001665:Binding Sites; D004271:DNA, Fungal / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012331:RNA, Fungal / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation / Q000235:genetics*
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C065681:REB1 protein, S cerevisiae; D012331:RNA, Fungal; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors