Genetic characterization of rbt mutants that enhance basal transcription from core promoters in Saccharomyces cerevisiae.
While this Saccharomyces cerevisiae SIN4 gene product is a component of a mediator complex associated with RNA polymerase II, various studies suggest the involvement of Sin4 in the alteration of higher-order chromatin structure. Our previous analysis of a sin4 mutant suggested that the mechanisms of transcriptional repression by Sin4 (mediator) and the Tup1-Ssn6 complex (general repressor) are different. To elucidate the way in which these two repression systems are interrelated, we isolated mutants that exhibit enhanced transcription of a reporter gene harboring the upstream activation sequence (UAS), but still are subject to Tup1-Ssn6-mediated repression. Besides sin4, rgr1, tup1, and ssn6 mutants, we also obtained new mutants that enhance basal transcription even from a core promoter without UAS. Such mutants, designated rbt for regulator of basal transcription, can be classified into at least six complementation groups, i.e., four single (rbt1 to rbt4) and two apparently double (rbt5 rbt6 and rbt7 rbt8) mutations. The phenotype of rbt mutants is dependent on the TATA box and not specific to the integration site or kind of core promoter. No significant difference in micrococcal nuclease (MNase) accessibility to the core promoter of test genes was observed between rbt mutants and the wild-type strain, indicating that the higher-order chromatin structure of the core promoter region is not significantly altered in these mutants. The rbt1 to rbt4 mutations are suppressed by the Dgal11 mutation as in the case of the sin4 mutation, but give rise to a different profile from the sin4 mutation with regard to the activity of some of the promoters. From these observations, we suggest that RBT gene product(s) could be novel mediators that act with or in close association with Sin4 but have a function distinct from that of Sin4. Moreover, the fact that rbt mutations nullify Tup1-Ssn6 general repressor-mediated repression is consistent with the idea that the mechanisms of Rbt (mediator)- and Tup1-Ssn6 (general repressor)-mediated repression are interconnected but substantially different.
Kunoh
T
T
Department of Biotechnology, Graduate School of Engineering, Osaka University, Yamadaoka, Suita, Osaka 565-0871, Japan.
Sakuno
T
T
Furukawa
T
T
Kaneko
Y
Y
Harashima
S
S
D000135:Acid Phosphatase / Q000378:metabolism; D001483:Base Sequence; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal / Q000235:genetics*; D017930:Genes, Reporter; D005816:Genetic Complementation Test; D005838:Genotype; D056892:Mediator Complex; D008836:Micrococcal Nuclease / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D009687:Nuclear Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D016385:TATA Box / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
C065455:CYC8 protein, S cerevisiae; D002843:Chromatin; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D056892:Mediator Complex; D009687:Nuclear Proteins; D012333:RNA, Messenger; D012097:Repressor Proteins; C076878:SIN4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C066502:TUP1 protein, S cerevisiae; D015534:Trans-Activators; D000135:Acid Phosphatase; D008836:Micrococcal Nuclease