Evidence that a complex of SIR proteins interacts with the silencer and telomere-binding protein RAP1.
The maintenance of transcriptional silencing at HM mating-type loci and telomeres in yeast requires the SIR2, SIR3, and SIR4 proteins, none of which appear to be DNA-binding proteins. Here we show that SIR3 and SIR4 interact with a carboxy-terminal domain of the silencer, telomere, and UAS-binding protein RAP1. We identified SIR3 and SIR4 in a two-hybrid screen for RAP1-interacting factors and showed that SIR3 interacts both with itself and with SIR4. The interaction between RAP1 and SIR3 can be observed in vitro in the absence of other yeast proteins. Consistent with the notion that native SIR proteins interact with the RAP1 carboxyl terminus, we show that mutation of the endogenous SIR3 and SIR4 genes increases transcriptional activation by LexA/RAP1 hybrids. To test the importance of the RAP1-SIR3 interaction for silencing, we identified mutations in the RAP1 carboxyl terminus that either diminish or abolish this interaction. When introduced into the native RAP1 protein, these mutations cause corresponding defects in silencing at both HMR and telomeres. We propose that RAP1 acts in the initiation of transcriptional silencing by recruiting a complex of SIR proteins to the chromosome via protein-protein interactions. These data are consistent with a model in which SIR3 and SIR4 play a structural role in the maintenance of silent chromatin and indicate that their action is initiated at the silencer itself.
Moretti
P
P
Department of Microbiology, Columbia University College of Physicians & Surgeons, New York, New York 10032.
Freeman
K
K
Coodly
L
L
Shore
D
D
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D004271:DNA, Fungal / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D049770:Genes, Mating Type, Fungal; D006655:Histone Deacetylases; D009154:Mutation; D011485:Protein Binding; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D012697:Serine Endopeptidases; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; D056565:Sirtuin 2; D037761:Sirtuins; D016615:Telomere / Q000378:metabolism; D034501:Telomere-Binding Proteins; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D000099160:LexA Repressor Protein
D001426:Bacterial Proteins; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D006655:Histone Deacetylases; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D012697:Serine Endopeptidases; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; D056565:Sirtuin 2; D037761:Sirtuins; D034501:Telomere-Binding Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D000099160:LexA Repressor Protein; C084122:SIR3 protein, S cerevisiae; C061573:SIR4 protein, S cerevisiae; C074461:RIF1 protein, S cerevisiae; C087778:SIR2 protein, S cerevisiae