Defining cis and trans factors that regulate the cell cycle-dependent establishment of gene silencing at the yeast mating type loci

Gene silencing in the budding yeast Saccharomyces cerevisiae is mediated by the formation of a repressive chromatin structure, and is seen at three loci: the mating type loci, telomeres, and the rDNA repeats. Independent studies have determined that progression through the cell cycle is required for the establishment of silencing in S. cerevisiae. However, the cell cycle specific event that is required for the establishment of silencing is unknown. Understanding the cell-cycle requirement for the establishment of silencing will help answer basic questions about epigenetics in general and the regulation of gene expression in cell development.;To examine the generality of the requirement for cell cycle progression in the establishment of silencing we regulated the availability or activity of the Sir3 silencing factor, using strains bearing either galactose inducible or temperature sensitive alleles of the SIR3 gene. Using these two systems we observed locus specific differences in the establishment of silencing at the mating type loci: consistent with prior studies cell cycle progression is required in the establishment of silencing at HMR; however, it has only a minor influence on the establishment of silencing at HML. Investigation of cis-elements that could be regulating the establishment of silencing at the mating type loci led us to identify a tRNA gene present adjacent to HMR, but not at HML, that imposes cell cycle dependence at HMR. In addition, translocating this tRNA gene to HML imposes cell cycle dependence at this locus. This tRNA gene has previously been shown to act as a barrier to the spread of heterochromatin from the HMR locus, and also to be necessary for the recruitment of the Scc1 cohesin to HMR. We went on to show that inactivating Scc1 abolished the requirement for cell cycle progression at HMR. Our studies indicate that the cell cycle progression requirement is not an intrinsic feature of silent chromatin in yeast, but is instead a cis-limited property that may involve localized recruitment of the cohesin complex.