Dataset Information

Nucleosome-positioning sequence repeats impact chromatin silencing in yeast minichromosomes.

ABSTRACT: Eukaryotic gene expression occurs in the context of structurally distinct chromosomal domains such as the relatively open, gene-rich, and transcriptionally active euchromatin and the condensed and gene-poor heterochromatin where its specific chromatin environment inhibits transcription. To study gene silencing by heterochromatin, we created a minichromosome reporter system where the gene silencer elements were used to repress the URA3 reporter gene. The minichromosome reporters were propagated in yeast Saccharomyces cerevisiae at a stable copy number. Conduction of gene silencing through nucleosome arrays was studied by placing various repeats of clone-601 DNA with high affinity for histones between the silencer and reporter in the yeast minichromosomes. High-resolution chromatin mapping with micrococcal nuclease showed that the clone-601 nucleosome positioning downstream of the HML-E gene silencing element was not significantly altered by chromatin silencing. Using URA3 reporter assays, we observed that gene silencing was conducted through arrays of up to eight nucleosomes. We showed that the shorter nucleosome repeat lengths, typical of yeast (167 and 172 bp), were more efficient in conducting silencing in vivo compared to the longer repeats (207 bp) typical of higher eukaryotes. Both the longer and the shorter repeat lengths were able to conduct silencing in minichromosomes independently of clone-601 nucleosome positioning orientations vs. the silencer element. We suggest that the shorter nucleosome linkers are more suitable for conducting gene silencing than the long repeats in yeast due to their higher propensity to support native-like chromatin higher-order folding.

SUBMITTER: Chakraborty SA

PROVIDER: S-EPMC4224149 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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Nucleosome-positioning sequence repeats impact chromatin silencing in yeast minichromosomes.

Chakraborty Sangita A SA Kazi Abid A AA Khan Tamreen M TM Grigoryev Sergei A SA

Genetics 20140903 3

Eukaryotic gene expression occurs in the context of structurally distinct chromosomal domains such as the relatively open, gene-rich, and transcriptionally active euchromatin and the condensed and gene-poor heterochromatin where its specific chromatin environment inhibits transcription. To study gene silencing by heterochromatin, we created a minichromosome reporter system where the gene silencer elements were used to repress the URA3 reporter gene. The minichromosome reporters were propagated i ...[more]

PMID: 25189873

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Project description:To gain a better understanding of the sequence patterns that characterize positioned nucleosomes, we first performed an analysis of the periodicities of the 256 tetranucleotides in a yeast genome-wide library of nucleosomal DNA sequences that was prepared by in vitro reconstitution. The approach entailed the identification and analysis of 24 unique tetranucleotides that were defined by 8 consensus sequences. These consensus sequences were shown to be responsible for most if not all of the tetranucleotide and dinucleotide periodicities displayed by the entire library, demonstrating that the periodicities of dinucleotides that characterize the yeast genome are, in actuality, due primarily to the 8 consensus sequences. A novel combination of experimental and bioinformatic approaches was then used to show that these tetranucleotides are important for preferred formation of nucleosomes at specific sites along DNA in vitro. These results were then compared to tetranucleotide patterns in genome-wide in vivo libraries from yeast and C. elegans in order to assess the contributions of DNA sequence in the control of nucleosome residency in the cell. These comparisons revealed striking similarities in the tetranucleotide occurrence profiles that are likely to be involved in nucleosome positioning in both in vitro and in vivo libraries, suggesting that DNA sequence is an important factor in the control of nucleosome placement in vivo. However, the strengths of the tetranucleotide periodicities were 3-4 fold higher in the in vitro as compared to the in vivo libraries, which implies that DNA sequence plays less of a role in dictating nucleosome positions in vivo. The results of this study have important implications for models of sequence-dependent positioning since they suggest that a defined subset of tetranucleotides is involved in preferred nucleosome occupancy and that these tetranucleotides are the major source of the dinucleotide periodicities that are characteristic of positioned nucleosomes.

Dataset Information

Nucleosome-positioning sequence repeats impact chromatin silencing in yeast minichromosomes.

Publications

Nucleosome-positioning sequence repeats impact chromatin silencing in yeast minichromosomes.

Similar Datasets

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