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Distinguishing between recruitment and spread of silent chromatin structures in Saccharomyces cerevisiae [I]

ABSTRACT: 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.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE189038 | GEO | 2021/11/24

REPOSITORIES: GEO

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Distinguishing between recruitment and spread of silent chromatin structures in Saccharomyces cerevisiae [II]

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. We mapped sites of Sir3-chromatin interactions genome-wide using long-read Nanopore sequencing to detect adenines methylated by the fusion protein. We also used a temperature-sensitive allele of SIR3 (sir3-8) fused to M.ECOGII to induce m6A methylation over time. Time courses involved a switch from restrictive temperature (37°C) to permissive temperature (25°C).

2022-01-17 | GSE190136 | GEO

Analysis of a novel existence region of Sir3p in S. cerevisiae

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.

2015-11-18 | GSE75075 | GEO

Histone H3 N-terminus Regulates Higher Order Structure of Yeast Heterochromatin

Project description:In budding yeast, telomeres and the mating type (HM) loci are found in a heterochromatin-like silent structure initiated by Rap1 and extended by the interaction of Sir (Silencing Information Regulator) proteins with histones. Binding data demonstrate that both the H3 and H4 N terminal domains required for silencing in vivo interact directly with Sir3 and Sir4 in vitro. The role of H4 lysine 16 deacetylation is well established in Sir3 protein recruitment, however that of the H3 N terminal tail has remained unclear. In order to characterize the role of H3 in silent chromatin formation and compare it to H4 we have generated comprehensive high resolution genome-wide binding maps of heterochromatin proteins. We find that H4 lysine 16 deacetylation is required for the recruitment and spreading of heterochromatin proteins at all telomeres and HM loci. In contrast the H3 N terminus is required for neither recruitment nor spreading of Sir proteins. Instead, deletion of the H3 tail leads to increased accessibility within heterochromatin of an ectopic bacterial dam methylase and the decreased mobility of an HML heterochromatic fragment in sucrose gradients. These findings indicate an altered chromatin structure. We propose that Sir proteins recruited by the H4 tail then interact with the H3 tail to form a higher order silent chromatin structure.

2009-08-01 | GSE16742 | GEO

Context dependent function of transcriptional regulator Rap1 in gene silencing and activation [ChIPseq I]

Project description:As part of a study on the role of Rap1 at the promoters of HML and MAT in S cerevisiae, we endogenously tagged Rap1 with a 3xV5 tag and performed ChIPseq in a variety of genetic backgrounds including: silenced (SIR); unsilenced (sir1∆/sir3∆/sir4∆); in strains witha wild type alpha2 promoter (wtpromoter); or with a point mutation at the promoter (mutpromoter).

2023-05-09 | GSE227757 | GEO

Cooperative Associations with Sites on Different Nucleosomes Mediate Heterochromatin Spreading

Project description:Sir3wH and Sir4CC are required for SIR complex spreading in vivo but play partially redundant roles when Sir3 is overexpressed.

2017-01-04 | GSE76553 | GEO

γH2A is a component of yeast heterochromatin required for telomere elongation

Project description:Histones of heterochromatin are deacetylated in yeast and methylated in more complex eukaryotes to regulate heterochromatin structure and gene silencing. Here, we report that histone H2A phosphorylated at serine 129 (γH2A) in Saccharomyces cerevisiae is a conceptually new type of heterochromatin modification that functions downstream of silent chromatin assembly. We show that γH2A is enriched throughout yeast telomeric and silent mating locus (HM) heterochromatin where γH2A results from the action of kinases Tel1 and Mec1. Interestingly, mutation of γH2A has no apparent effect on the binding of Sir (silent information regulator) complex or on gene silencing. In contrast, deletion of SIR3 abolishes the formation of γH2A at heterochromatin. To address the function of γH2A, we used a ∆rif1 mutant strain in which telomeres are excessively elongated to show that γH2A is required for the optimal recruitment of Cdc13, a regulator of telomere elongation, and for telomere elongation itself. Thus, a histone modification that parallels Sir3 protein binding is shown here to be dispensable for the formation of a silent structure but is important for a crucial heterochromatin-specific downstream function in telomere homeostasis.

2011-01-05 | GSE21814 | GEO

ChIP-seq analysis of yeast Ty1 integrase reveal the role of the AC40 interacting domain in the recruitment at both Pol I and Pol III-transcribed genes

Project description:In budding yeast, the selective integration of the Ty1 LTR retrotransposon upstream of RNA polymerase III (Pol III)-transcribed genes requires the interaction between the AC40, a common subunit of Pol III and Pol I, and Ty1 integrase (IN1). The AC40/IN1 interaction involves a short sequence, the targeting domain (TD), present in the C-terminal part of IN1. Chip-seq analysis using WT or mutated Ty1 integrase demonstrated that TD is responsible for the recruitment of IN1 at both Pol I and Pol III-transcribed genes. Moreover, the introduction of the C-terminal residues of Ty1 in the Ty5 retrotransposon, which preferentially integrates in heterochromatin at silent mating loci (HMR and HML) and near telomeres, leads to its retargeting at Pol III-transcribed genes.

2020-05-11 | E-MTAB-9038 | biostudies-arrayexpress

Context dependent function of transcriptional regulator Rap1 in gene silencing and activation [ChIP-seq III]

Project description:As part of a study on the mechanism of Sir-based silencing at HML in S.cerevisiae, and the contribution of Rap1 to transcription of the locus, we performed ChIP-seq of multiple components of RNA Pol II machinery in different genetic backgrounds including: sir4∆, SIR+, and with or without a mutation that abbrogated Rap1 binding at the alpha promoter.

2023-05-09 | GSE227760 | GEO

GTP-dependent regulation of heterochromatin fluctuations at subtelomeric regions in S. cerevisiae

Project description:Analysis of gene expression changes following deletion of SIR genes at subtelomeric region. A total of 4 samples were analyzed : Wild type (BY4742) strain, SIR2 deletion strain, SIR3 deletion strain, SIR4 deletion strain.

2023-12-28 | GSE230739 | GEO

Nucleosome profiles in strains with different numbers of nucleosomes at HML and HMR

Project description:We report nucleosome positions by using native Mnase-seq. The results strongly suggest that nucleosome positions don't change at the heterochromatic locus HMR upon removal of adjacent nucleosomes via deletion of nucleosomal DNA. Positions of nucleosomes at HML are also quantified.

2019-09-06 | GSE136897 | GEO

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