Expression of GCR1, the transcriptional activator of glycolytic enzyme genes in the yeast Saccharomyces cerevisiae, is positively autoregulated by Gcr1p.
When regulation of GCR1 expression was analysed using a GCR1-lacZ fusion, lacZ expression levels were decreased in the Deltagcr1 or Deltagcr2 mutant. RT-PCR analysis of genomic GCR1 transcript confirmed the dependency of GCR1 expression on the Gcr1p-Gcr2p complex. Examination of the 5' non-coding region of GCR1 identified three putative Gcr1p binding sites (CT-boxes) in the -100 to -200 region of GCR1, and the putative binding sites for Rap1p (RPG-box) and Abf1p were also identified nearby. The region containing putative cis-elements was analysed by cloning it upstream of the CYC1TATA-lacZ fusion. The GCR1(UAS)-CYC1TATA-lacZ fusion showed a moderate activity and, as expected, the activity was drastically reduced in the Deltagcr1 or Deltagcr2 mutant. Systematic deletion and mutation analyses of cis-elements in this region demonstrated that the putative binding sites for Rap1p and Abf1p were not involved in the promoter activity of GCR1(UAS) and only one of the three CT-boxes showed GCR1- and GCR2-dependent promoter activity. In contrast to the expression of glycolytic genes, where a RPG-box adjacent to the CT-box is required for strong promoter activities, CT-box-dependent expression of GCR1 did not require the RPG-box. Also, a contribution of Sgc1p, an E-box binding transcription factor, to the expression of GCR1 was suggested, based on its disruption analysis.
Sasaki
Hiromi
H
Institute for Biological Resources and Functions, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, 305-8566, Japan.
Kishimoto
Tomoko
T
Mizuno
Takayuki
T
Shinzato
Tomoko
T
Uemura
Hiroshi
H
D001483:Base Sequence; D001665:Binding Sites / Q000502:physiology; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000235:genetics; D015966:Gene Expression Regulation, Fungal / Q000502:physiology*; D006019:Glycolysis / Q000502:physiology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000502:physiology; D012331:RNA, Fungal / Q000737:chemistry / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D000089804:Shelterin Complex; D034501:Telomere-Binding Proteins / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C076227:GCR1 protein, S cerevisiae; C076228:GCR2 protein, S cerevisiae; C440829:RAP1 protein, S cerevisiae; D012331:RNA, Fungal; D029701:Saccharomyces cerevisiae Proteins; D000089804:Shelterin Complex; D034501:Telomere-Binding Proteins; D015534:Trans-Activators; D014157:Transcription Factors