title: Transcriptional regulator Leu3 of Saccharomyces cerevisiae: separation of activator and repressor functions.
abstract: The Leu3 protein of Saccharomyces cerevisiae binds to specific DNA sequences present in the 5' noncoding region of at least five RNA polymerase II-transcribed genes. Leu3 functions as a transcriptional activator only when the metabolic intermediate alpha-isopropylmalate is also present. In the absence of alpha-isopropylmalate, Leu3 causes transcription to be repressed below basal levels. We show here that different portions of the Leu3 protein are responsible for activation and repression. Fusion of the 30 C-terminal residues of Leu3 to the DNA-binding domain of the Gal4 protein created a strong cross-species activator, demonstrating that the short C-terminal region is not only required but also sufficient for transcriptional activation. Using a recently developed Leu3-responsive in vitro transcription assay as a test system for repression (J. Sze, M. Woontner, J. Jaehning, and G. B. Kohlhaw, Science 258:1143-1145, 1992), we show that mutant forms of the Leu3 protein that lack the activation domain still function as repressors. The shortest repressor thus identified had only about 15% of the mass of the full-length Leu3 protein and was centered on the DNA-binding region of Leu3. Implications of this finding for the mechanism of repression are discussed.
authors.affiliation: Department of Biochemistry, Purdue University, West Lafayette, Indiana 47907.
authors.forename: J Y
authors.initials: JY
authors.lastname: Sze
authors.forename: E
authors.initials: E
authors.lastname: Remboutsika
authors.forename: G B
authors.initials: GB
authors.lastname: Kohlhaw
mesh_terms: D000818:Animals; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal; D051379:Mice; D012097:Repressor Proteins / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D013045:Species Specificity; D013329:Structure-Activity Relationship; D015534:Trans-Activators / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; C063667:LEU3 protein, S cerevisiae; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors