A role for the Saccharomyces cerevisiae RENT complex protein Net1 in HMR silencing.
Silencing in the yeast Saccharomyces cerevisiae is known in three classes of loci: in the silent mating-type loci HML and HMR, in subtelomeric regions, and in the highly repetitive rDNA locus, which resides in the nucleolus. rDNA silencing differs markedly from the other two classes of silencing in that it requires a DNA-associated protein complex termed RENT. The Net1 protein, a central component of RENT, is required for nucleolar integrity and the control of exit from mitosis. Another RENT component is the NAD(+)-dependent histone deacetylase Sir2, which is the only silencing factor known to be shared among the three classes of silencing. Here, we investigated the role of Net1 in HMR silencing. The mutation net1-1, as well as NET1 expression from a 2micro-plasmid, restored repression at silencing-defective HMR loci. Both effects were strictly dependent on the Sir proteins. We found overexpressed Net1 protein to be directly associated with the HMR-E silencer, suggesting that Net1 could interact with silencer binding proteins and recruit other silencing factors to the silencer. In agreement with this, Net1 provided ORC-dependent, Sir1-independent silencing when artificially tethered to the silencer. In contrast, our data suggested that net1-1 acted indirectly in HMR silencing by releasing Sir2 from the nucleolus, thus shifting the internal competition for Sir2 from the silenced loci toward HMR.
Kasulke
Daniela
D
Max-Planck-Institute for Molecular Genetics, 14195 Berlin, Germany.
Seitz
Stefanie
S
Ehrenhofer-Murray
Ann E
AE
D018797:Cell Cycle Proteins; D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; D015966:Gene Expression Regulation, Fungal / Q000502:physiology*; D020868:Gene Silencing / Q000502:physiology*; D006655:Histone Deacetylases / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D000089804:Shelterin Complex; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae / Q000378:metabolism / Q000502:physiology; D056565:Sirtuin 2; D037761:Sirtuins / Q000378:metabolism; D016615:Telomere / Q000235:genetics; D034501:Telomere-Binding Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism
D018797:Cell Cycle Proteins; D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; C052130:GAL4 protein, S cerevisiae; C119300:Net1 protein, S cerevisiae; D009687:Nuclear Proteins; C440829:RAP1 protein, S cerevisiae; C084121:SIR1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D000089804:Shelterin Complex; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; D034501:Telomere-Binding Proteins; D014157:Transcription Factors; C087778:SIR2 protein, S cerevisiae; D056565:Sirtuin 2; D037761:Sirtuins; D006655:Histone Deacetylases