Design of a minimal silencer for the silent mating-type locus HML of Saccharomyces cerevisiae.
The silent mating-type loci HML and HMR of Saccharomyces cerevisiae contain mating-type information that is permanently repressed. This silencing is mediated by flanking sequence elements, the E- and I-silencers. They contain combinations of binding sites for the proteins Rap1, Abf1 and Sum1 as well as for the origin recognition complex (ORC). Together, they recruit other silencing factors, foremost the repressive Sir2/Sir3/Sir4 complex, to establish heterochromatin-like structures at the HM loci. However, the HM silencers exhibit considerable functional redundancy, which has hampered the identification of further silencing factors. In this study, we constructed a synthetic HML-E silencer (HML-SS ΔI) that lacked this redundancy. It consisted solely of Rap1 and ORC-binding sites and the D2 element, a Sum1-binding site. All three elements were crucial for minimal HML silencing, and mutations in these elements led to a loss of Sir3 recruitment. The silencer was sensitive to a mutation in RAP1, rap1-12, but less sensitive to orc mutations or sum1Δ. Moreover, deletions of SIR1 and DOT1 lead to complete derepression of the HML-SS ΔI silencer. This fully functional, minimal HML-E silencer will therefore be useful to identify novel factors involved in HML silencing.
Weber
Jan M
JM
Zentrum für Medizinische Biotechnologie, Abteilung Genetik, Universität Duisburg-Essen, 45117 Essen, Germany.
Ehrenhofer-Murray
Ann E
AE
D001665:Binding Sites; D015966:Gene Expression Regulation, Fungal; D020868:Gene Silencing; D049770:Genes, Mating Type, Fungal; D056426:Genetic Loci; D011495:Histone-Lysine N-Methyltransferase / Q000235:genetics; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics; D009841:Oligonucleotides / Q000737:chemistry; D051738:Origin Recognition Complex / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D000089804:Shelterin Complex; D037941:Silencer Elements, Transcriptional; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae / Q000235:genetics; D034501:Telomere-Binding Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism
D009687:Nuclear Proteins; D009841:Oligonucleotides; D051738:Origin Recognition Complex; 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; C554167:URA3 protein, S cerevisiae; C416235:Dot1 protein, S cerevisiae; D011495:Histone-Lysine N-Methyltransferase