Project description:In the budding yeast, HMR, HML, telomere and rDNA domain are known as a silencing region. Sir2 need to make it at rDNA and, HMR, HML and the telomere need to Sir2, Sir3, Sir4 complex to control internal gene repression. In this report, we found a newly Sir3 binding domain, CN domain (Chromosome New region) 1~14, by the ChIP on chip analysis on S.cerevisiae chromosome. In addition, we also performed ChIP on chip analysis with anti-Sir3 antibody using G1 phase synchronized cell to find Sir3 distribution difference of stage of cell cycle and we found CN15~CN25 which was G1 phase specific Sir3 binding region. Furthermore, we analyzed difference of gene expression at CN region in sir3 strain, and some regions did not change level of gene expression. In the conventional report, Sir3 had recruited by Sir2 and Sir4 on chromosome, but recruit of Sir3 was independent on Sir2 and Sir4 at some CN regions. These data suggested that we found a newly Sir3 function and Sir3 recruited system on chromosome.
Project description:As part of a study of establishment of silencing in Saccharomyces cerevisiae, we performed ChIP-seq on myc-tagged Sir4 in several conditions. Included in those conditions are wild-type cycling cells, cycling sir3∆ cells, and various experiments during which silencing establishment was controlled using the inducible SIR3-EBD allele. Silencing establishment experiments were performed in both wild-type and dot1∆ cells.
Project description:Single-stranded DNA breaks (nicks) were tagged by biotin-dCTP in wild-type budding yeast cells and mapped by nick-ChIP coupled to whole-genomic microarrays.
Project description:The formation of heterochromatin at HML, HMR, and telomeres in Saccharomyces cerevisiae involves two main steps: Recruitment of Sir proteins to silencers and their spread throughout the silenced domain. For the following datasets, we created a fusion protein between the heterochromatin protein Sir3 and the non-site-specific bacterial adenine methyltransferase M.EcoGII, with or without a 3xV5 epitope at the C-terminus. We performed ChIP-seq experiments (immunoprecipitated Sir3-M.EcoGII-3xV5) and MeDIP-seq experiments (immunoprecipitated m6A methylated DNA). We also used a temperature-sensitive allele of SIR3 (sir3-8) fused to M.ECOGII to induce m6A methylation for MeDIP-seq.
Project description:Paired-end sequencing study of nucleosomal DNA prepared from budding yeast by micrococcal nuclease digestion. Comparison of control cells with cells treated with 10 mM 3-aminotriazole for 20 minutes
