ABSTRACT
Sir2 protein has been reported to be recruited to dicentric chromosomes under tension, and such chromosomes are reported to be especially vulnerable to breakage in sir2Δ mutants. We found that the loss of viability in such mutants was an indirect effect of the repression of nonhomologous end joining in Sir(-) mutants and that the apparent recruitment of Sir2 protein to chromosomes under tension was likely due to methodological weakness in early chromatin immunoprecipitation studies.
Subject(s)
Saccharomyces cerevisiae/genetics , Silent Information Regulator Proteins, Saccharomyces cerevisiae/metabolism , Centromere/genetics , Centromere/metabolism , Chromatin/genetics , Chromatin/metabolism , Chromosomes, Fungal/metabolism , DNA Repair/physiology , DNA-Binding Proteins/metabolism , Gene Expression Regulation, Fungal , Saccharomyces cerevisiae/metabolism , Silent Information Regulator Proteins, Saccharomyces cerevisiae/genetics , Telomere/genetics , Telomere/metabolism , Transcription, GeneticABSTRACT
Multiple mechanisms have been proposed for gene silencing in Saccharomyces cerevisiae, ranging from steric occlusion of DNA binding proteins from their recognition sequences in silenced chromatin to a specific block in the formation of the preinitiation complex to a block in transcriptional elongation. This study provided strong support for the steric occlusion mechanism by the discovery that RNA polymerase of bacteriophage T7 could be substantially blocked from transcribing from its cognate promoter when embedded in silenced chromatin. Moreover, unlike previous suggestions, we found no evidence for stalled RNA polymerase II within silenced chromatin. The effectiveness of the Sir protein-based silencing mechanism to block transcription activated by Gal4 at promoters in the domain of silenced chromatin was marginal, yet it improved when tested against mutant forms of the Gal4 protein, highlighting a role for specific activators in their sensitivity to gene silencing.
