Active genes in budding yeast display enhanced in vivo accessibility to foreign DNA methylases: a novel in vivo probe for chromatin structure of yeast.

Singh, Jagmohan; Klar, Amar J. S.

doi:10.1101/gad.6.2.186

Cited by 188 publications

(136 citation statements)

References 44 publications

(53 reference statements)

Supporting

Mentioning

132

Contrasting

Order By: Relevance

“…This characteristic together with the compact organization of the genome suggests that yeast gene regulation has evolved efficient mechanisms for insulating genes from each other. Some sequences named STAR (subtelomeric antisilencing region) are able to counteract silencer-driven repression at the matingtype HML locus and act with antisilencing properties against the spreading of silenced chromatin (Singh and Klar, 1992;Fourel et al, 1999Fourel et al, , 2001.…”

Section: Partition Of Distinct Chromosomal Regionsmentioning

confidence: 99%

Insulators are fundamental components of the eukaryotic genomes

Brasset

Vaury

2005

Heredity

View full text Add to dashboard Cite

The properties of cis-regulatory elements able to influence gene transcription over large distances have led to the hypothesis that elements called insulators should exist to limit the action of enhancers and silencers. During the last decades, insulators have been identified in many eukaryotes from yeast to human. Insulators possess two main properties: (i) they can block enhancer-promoter communication ('enhancer blocker activity'), and (ii) they can prevent the spread of repressive chromatin ('barrier activity'). This review focuses on recent studies designed to elucidate the molecular mechanisms of the insulator function, and gives an overview of the critical role of insulators in nuclear organization and functional identity of chromatin. Heredity (2005) 94, 571-576.

show abstract

Section: Partition Of Distinct Chromosomal Regionsmentioning

confidence: 99%

Insulators are fundamental components of the eukaryotic genomes

Brasset

Vaury

2005

Heredity

View full text Add to dashboard Cite

show abstract

“…A sir2⌬ mutant strain exhibits complete derepression at these loci. Derepression has been correlated with increased accessibility to DNA-modifying enzymes and psoralen, indicating that these loci have a more relaxed chromatin structure in the absence of Sir2p (Nasmyth, 1982;Gottschling, 1992;Singh and Klar, 1992;Loo and Rine, 1994;Fritze and Esposito, 1997;Smith and Boeke, 1997;Smith et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

The Conserved Core of a HumanSIR2Homologue Functions in Yeast Silencing

Sherman

Stone

Freeman-Cook

et al. 1999

MBoC

View full text Add to dashboard Cite

Silencing is a universal form of transcriptional regulation in which regions of the genome are reversibly inactivated by changes in chromatin structure. Sir2 (Silent Information Regulator) protein is unique among the silencing factors in Saccharomyces cerevisiae because it silences the rDNA as well as the silent mating-type loci and telomeres. Discovery of a gene family of Homologues of Sir Two (HSTs) in organisms from bacteria to humans suggests that SIR2's silencing mechanism might be conserved. The Sir2 and Hst proteins share a core domain, which includes two diagnostic sequence motifs of unknown function as well as four cysteines of a putative zinc finger. We demonstrate by mutational analyses that the conserved core and each of its motifs are essential for Sir2p silencing. Chimeras between Sir2p and a human Sir2 homologue (hSir2Ap) indicate that this human protein's core can substitute for that of Sir2p, implicating the core as a silencing domain. Immunofluorescence studies reveal partially disrupted localization, accounting for the yeast-human chimeras' ability to function at only a subset of Sir2p's target loci. Together, these results support a model for the involvement of distinct Sir2p-containing complexes in HM/telomeric and rDNA silencing and that HST family members, including the widely expressed hSir2A, may perform evolutionarily conserved functions.

show abstract

“…Few techniques are capable of demonstrating these interactions in the context of native chromatin in living cells, and these methods have limitations (1). For example, with footprinting techniques, protection against chemical (e.g., dimethyl sulfate) or enzymatic probes expressed in cells, e.g., DNA methyltransferases (MTases) (2)(3)(4)(5)(6) or DNase I (7), requires close proximity of the interacting factor to DNA sites that are modified or cleaved by the footprinting agent. Footprinting methods also require that the factor resists displacement by the enzymatic or chemical probe.…”

mentioning

confidence: 99%

Targeted cytosine methylation for in vivo detection of protein–DNA interactions

Carvin

Dhasarathy

Friesenhahn

et al. 2003

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

View full text Add to dashboard Cite

We report a technique, named targeted gene methylation (TAGM), for identifying in vivo protein-binding sites in chromatin. M.CviPI, a cytosine-5 DNA methyltransferase recognizing GC sites, is fused to a DNA-binding factor enabling simultaneous detection of targeted methylation, factor footprints, and chromatin structural changes by bisulfite genomic sequencing. Using TAGM with the yeast transactivator Pho4, methylation enrichments of up to 34-fold occur proximal to native Pho4-binding sites. Additionally, significant selective targeting of methylation is observed several hundred nucleotides away, suggesting the detection of long-range interactions due to higher-order chromatin structure. In contrast, at an extragenic locus lacking Pho4-binding sites, methylation levels are at the detection limit at early times after Pho4 transactivation. Notably, substantial amounts of methylation are targeted by Pho4-M.CviPI under repressive conditions when most of the transactivator is excluded from the nucleus. Thus, TAGM enables rapid detection of DNA-protein interactions even at low occupancies and has potential for identifying factor targets at the genome-wide level. Extension of TAGM from yeast to vertebrates, which use methylation to initiate and propagate repressed chromatin, could also provide a valuable strategy for heritable inactivation of gene expression.T he interaction of proteins with chromosomal target sites, either directly or through recruitment by DNA-bound factors, is central to many processes, including transcriptional activation and repression, replication and repair of DNA, recombination, and chromosome segregation. Therefore, strategies are needed that can efficiently identify specific chromosomal sites at which factors act. Few techniques are capable of demonstrating these interactions in the context of native chromatin in living cells, and these methods have limitations (1). For example, with footprinting techniques, protection against chemical (e.g., dimethyl sulfate) or enzymatic probes expressed in cells, e.g., DNA methyltransferases (MTases) (2-6) or DNase I (7), requires close proximity of the interacting factor to DNA sites that are modified or cleaved by the footprinting agent. Footprinting methods also require that the factor resists displacement by the enzymatic or chemical probe. Moreover, because many proteins can exclude probe access, a footprint does not provide an unequivocal identity of the bound protein (8). To circumvent this latter problem, proteins have been fused to an endonuclease (9); however, the resulting DNA damage alters chromatin structure and activates checkpoint controls. Another method, chromatin immunoprecipitation (ChIP), uses in situ fixation with formaldehyde followed by immunoselection of DNA-bound complexes. The requirements for large numbers of cells and highly specific antibodies as well as low fixation efficiencies (Ϸ0.1-0.5%) (10, 11) present distinct disadvantages of ChIP analysis. The approach of tethering chromatin proteins to the Dam MTase, which methylates ...

show abstract

Active genes in budding yeast display enhanced in vivo accessibility to foreign DNA methylases: a novel in vivo probe for chromatin structure of yeast.

Cited by 188 publications

References 44 publications

Insulators are fundamental components of the eukaryotic genomes

Insulators are fundamental components of the eukaryotic genomes

The Conserved Core of a HumanSIR2Homologue Functions in Yeast Silencing

Targeted cytosine methylation for in vivo detection of protein–DNA interactions

Contact Info

Product

Resources

About