Compensatory Interactions between Sir3p and the Nucleosomal LRS Surface Imply Their Direct Interaction

Norris, Anne; Bianchet, M.A.; Boeke, Jef D.

doi:10.1371/journal.pgen.1000301

Cited by 41 publications

(82 citation statements)

References 60 publications

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“…If H3 K79 methylation were critical in antagonizing silencing, then the H3 K79R mutant would be expected to phenocopy dot1Δ cells with respect to the kinetics of silencing. For comparison, mutating H3 K79 to alanine abrogates silencing potentially by disrupting the charge-based interaction between either H3 and Sir3 or H3 and H4 (32,44). We replaced plasmids carrying HHT2::HHF2 alleles with plasmids carrying the appropriate K79R mutation (hht2K79R::HHF2).…”

Section: Resultsmentioning

confidence: 99%

“…First, Dot1 catalyzes the mono-, di-, and trimethylation states of H3 K79 (27,28,30,31). This particular lysine is located on the loss of rDNA silencing (LRS) face of H3, a surface whose electrostatic properties are important for association between the nucleosome and the BAH domain of Sir3 (32)(33)(34)(35)(36). H3 K79 methylation, therefore, interferes with the nucleosome's ability to adequately bind Sir3 (9).…”

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confidence: 99%

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Symmetry, asymmetry, and kinetics of silencing establishment in Saccharomyces cerevisiae revealed by single-cell optical assays

Osborne

Hiraoka²,

Rine³

2011

Proc. Natl. Acad. Sci. U.S.A.

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In Saccharomyces cerevisiae, silent chromatin inhibits the expression of genes at the HML, HMR, and telomeric loci. When silent chromatin forms de novo, the rate of its establishment is influenced by different chromatin states. In particular, loss of the enzyme Dot1, an H3 K79 methyltransferase, leads to rapid silencing establishment. We tested whether silencing establishment was antagonized by H3 K79 methylation or by the Dot1 protein itself competing with Sir3 for binding sites on nucleosomes. To do so, we monitored fluorescence activity in cells containing a GFP gene within the HML locus during silencing establishment in a series of dot1 and histone mutant backgrounds. Silencing establishment rate was correlated with Dot1's enzymatic function rather than with the Dot1 protein itself. In addition, histone mutants that mimicked the conformation of unmethylated H3 K79 increased the rate of silencing establishment, indicating that the H3 K79 residue affected silencing independently of Dot1 abundance. Using fluorophore-based reporters, we confirmed that mother and daughter cells often silence in concert, but in instances where asymmetric silencing occurs, daughter cells established silencing earlier than their mothers. This noninvasive technique enabled us to demonstrate an asymmetry in silencing establishment of a key regulatory locus controlling cell fate.

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Symmetry, asymmetry, and kinetics of silencing establishment in Saccharomyces cerevisiae revealed by single-cell optical assays

Osborne

Hiraoka²,

Rine³

2011

Proc. Natl. Acad. Sci. U.S.A.

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“…Genetic suppressor studies over the course of many years suggested that the domain mediates binding of nucleosomes. Mutations that mapped to the BAH domain suppressed mutations of either the H4 tail or a patch on the nucleosome surface known as the LRS domain (Johnson et al 1990;Thompson et al 2003;Norris et al 2008;Sampath et al 2009). The BAH domain was sufficient for partial silencing when forced to dimerize, strengthening the notion that the BAH domain was sufficient for nucleosome recognition (Connelly et al 2006).…”

Section: Sir3 Protein Structurementioning

confidence: 99%

“…Obtaining cocrystals required the use of a SIR3 hypermorphic allele D205N, which suppresses silencing defects caused by mutations in histones and other silencing factors (Johnson et al 1990;Liu and Lustig 1996;Norris et al 2008). The mutation increases the affinity of Sir3 for nucleosomes in vitro (Connelly et al 2006).…”

Section: Histone Binding By Sir3 Bah Domainmentioning

confidence: 99%

The Nuts and Bolts of Transcriptionally Silent Chromatin in Saccharomyces cerevisiae

2016

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Transcriptional silencing in Saccharomyces cerevisiae occurs at several genomic sites including the silent mating-type loci, telomeres, and the ribosomal DNA (rDNA) tandem array. Epigenetic silencing at each of these domains is characterized by the absence of nearly all histone modifications, including most prominently the lack of histone H4 lysine 16 acetylation. In all cases, silencing requires Sir2, a highly-conserved NAD + -dependent histone deacetylase. At locations other than the rDNA, silencing also requires additional Sir proteins, Sir1, Sir3, and Sir4 that together form a repressive heterochromatin-like structure termed silent chromatin. The mechanisms of silent chromatin establishment, maintenance, and inheritance have been investigated extensively over the last 25 years, and these studies have revealed numerous paradigms for transcriptional repression, chromatin organization, and epigenetic gene regulation. Studies of Sir2-dependent silencing at the rDNA have also contributed to understanding the mechanisms for maintaining the stability of repetitive DNA and regulating replicative cell aging. The goal of this comprehensive review is to distill a wide array of biochemical, molecular genetic, cell biological, and genomics studies down to the "nuts and bolts" of silent chromatin and the processes that yield transcriptional silencing.

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“…Sir2p, which does not bind chromatin directly, is a histone deacetylase responsible for the hypoacetylation of histones in heterochromatin (Moazed 2001). Sir3p and Sir4p interact with the amino (N)-terminal tails of histones H3 and H4, and Sir3p also binds a region called LRS (loss of rDNA silencing) on the solvent-accessible surface of the nucleosome (Hecht et al 1995;Park et al 2002;Liou et al 2005;Norris et al 2008;Sampath et al 2009). These interactions are believed to be key to the binding/propagation of SIR complex along chromatin.…”

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confidence: 99%

Differential Contributions of Histone H3 and H4 Residues to Heterochromatin Structure

Olsen

Zhang

et al. 2011

Genetics

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Transcriptional silencing in Saccharomyces cerevisiae is mediated by heterochromatin. There is a plethora of information regarding the roles of histone residues in transcriptional silencing, but exactly how histone residues contribute to heterochromatin structure is not resolved. We address this question by testing the effects of a series of histone H3 and H4 mutations involving residues in their aminoterminal tails, on the solvent-accessible and lateral surfaces of the nucleosome, and at the interface of the H3/H4 tetramer and H2A/H2B dimer on heterochromatin structure and transcriptional silencing. The general state, stability, and conformational heterogeneity of chromatin are examined with a DNA topology-based assay, and the primary chromatin structure is probed by micrococcal nuclease. We demonstrate that the histone mutations differentially affect heterochromatin. Mutations of lysine 16 of histone H4 (H4-K16) and residues in the LRS (loss of rDNA silencing) domain of nucleosome surface markedly alter heterochromatin structure, supporting the notion that H4-K16 and LRS play key roles in heterochromatin formation. Deletion of the aminoterminal tail of H3 moderately alters heterochromatin structure. Interestingly, a group of mutations in the globular domains of H3 and H4 that abrogate or greatly reduce transcriptional silencing increase the conformational heterogeneity and/or reduce the stability of heterochromatin without affecting its overall structure. Surprisingly, yet another series of mutations abolish or reduce silencing without significantly affecting the structure, stability, or conformational heterogeneity of heterochromatin. Therefore, histone residues may contribute to the structure, stability, conformational heterogeneity, or other yet-to-be-characterized features of heterochromatin important for transcriptional silencing.

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Compensatory Interactions between Sir3p and the Nucleosomal LRS Surface Imply Their Direct Interaction

Cited by 41 publications

References 60 publications

Symmetry, asymmetry, and kinetics of silencing establishment in Saccharomyces cerevisiae revealed by single-cell optical assays

Symmetry, asymmetry, and kinetics of silencing establishment in Saccharomyces cerevisiae revealed by single-cell optical assays

The Nuts and Bolts of Transcriptionally Silent Chromatin in Saccharomyces cerevisiae

Differential Contributions of Histone H3 and H4 Residues to Heterochromatin Structure

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