Functional domains of rep2, a transcriptional activator subunit for Res2-Cdc10, controlling the cell cycle "start".
In the fission yeast Schizosaccharomyces pombe, passage from G1 to S-phase requires the execution of the transcriptional factor complex that consists of the Cdc10 and Res1/2 molecules. This complex activates the MluI cell cycle box cis-element contained in genes essential for S-phase onset and progression. The rep2(+) gene, isolated as a multicopy suppressor of a temperature-sensitive cdc10 mutant, has been postulated to encode a putative transcriptional activator subunit for the Res2-Cdc10 complex. To identify the rep2(+) function and molecularly define its domain organization, we reconstituted the Res2-Cdc10 complex-dependent transcriptional activation in Saccharomyces cerevisiae. Reconstitution experiments, deletion analyses using one and two hybrid systems, and in vivo Res2 coimmunoprecipitation assays show that the Res2-Cdc10 complex itself can recognize but cannot activate MluI cell cycle box without Rep2, and that consistent with its postulated function, Rep2 contains 45-amino acid Res2 binding and 22-amino acid transcriptional activation domains in the middle and C terminus of the molecule, respectively. The functional essentiality of these domains is also demonstrated by their requirement for rescue of the cold-sensitive rep2 deletion mutant of fission yeast.
Tahara
S
S
Department of Hygiene and Oncology, Tokyo Medical and Dental University, School of Medicine, Bunkyo-ku, Tokyo 113, Japan.
Tanaka
K
K
Yuasa
Y
Y
Okayama
H
H
D001665:Binding Sites; D002453:Cell Cycle / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D005656:Fungal Proteins / Q000235:genetics / Q000502:physiology*; D020558:GTP Phosphohydrolases; D015966:Gene Expression Regulation, Fungal; D008565:Membrane Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins; D012568:Schizosaccharomyces; D029702:Schizosaccharomyces pombe Proteins; D015534:Trans-Activators / Q000235:genetics / Q000502:physiology*; D014157:Transcription Factors; D015533:Transcriptional Activation
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D008565:Membrane Proteins; C490689:REP2 protein, S cerevisiae; C096254:Rep2 protein, S pombe; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C061178:cdc10 protein, S pombe; C087021:res2 protein, S pombe; C512892:CDC10 protein, S cerevisiae; D020558:GTP Phosphohydrolases