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Distal chromatin structure influences local nucleosome positions and gene expression.

Jansen A, van der Zande E, Meert W, Fink GR, Verstrepen KJ - Nucleic Acids Res. (2012)

Bottom Line: In addition, we show that changes in the nucleosome positions in the URA3 promoter strongly affect the promoter activity.Most interestingly, in addition to demonstrating the effect of the local DNA sequence, our study provides novel in vivo evidence that nucleosome positions are also affected by the position of neighboring nucleosomes.Nucleosome structure may therefore be an important selective force for conservation of gene order on a chromosome, because relocating a gene to another genomic position (where the positions of neighboring nucleosomes are different from the original locus) can have dramatic consequences for the gene's nucleosome structure and thus its expression.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Systems Biology, VIB, Bio-Incubator, Gaston Geenslaan 1, B-3001, Leuven, Belgium.

ABSTRACT
The positions of nucleosomes across the genome influence several cellular processes, including gene transcription. However, our understanding of the factors dictating where nucleosomes are located and how this affects gene regulation is still limited. Here, we perform an extensive in vivo study to investigate the influence of the neighboring chromatin structure on local nucleosome positioning and gene expression. Using truncated versions of the Saccharomyces cerevisiae URA3 gene, we show that nucleosome positions in the URA3 promoter are at least partly determined by the local DNA sequence, with so-called 'anti-nucleosomal elements' like poly(dA:dT) tracts being key determinants of nucleosome positions. In addition, we show that changes in the nucleosome positions in the URA3 promoter strongly affect the promoter activity. Most interestingly, in addition to demonstrating the effect of the local DNA sequence, our study provides novel in vivo evidence that nucleosome positions are also affected by the position of neighboring nucleosomes. Nucleosome structure may therefore be an important selective force for conservation of gene order on a chromosome, because relocating a gene to another genomic position (where the positions of neighboring nucleosomes are different from the original locus) can have dramatic consequences for the gene's nucleosome structure and thus its expression.

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The nucleosome profile of the URA3 promoter is determined by the positions of the −1 and +1 nucleosomes. (A) Using tiling qPCR, we determined the nucleosome profile of the full-length URA3 promoter at the native URA3 locus. Peaks of PCR signal represent enrichment of DNA segments protected by nucleosomes from micrococcal nuclease digestion, and valleys represent nucleosome-free segments. Peak positions corresponding to the centers of the −1 and +1 nucleosomes were calculated (see ‘Materials and Methods’ section), as well as the distance d between the −1 and +1 nucleosome centers (Table 1). (B) The URA3 gene including the (truncated) promoter was inserted at various locations p in the LYS2 gene. Before insertion, n is the position of the center of the first nucleosome upstream of the insertion site p. This upstream nucleosome might influence the positions of the −1 and +1 nucleosomes of the URA3 promoter after insertion. Coordinates for n are relative to the insertion site p. After insertion, the positions of the centers of the −1 and +1 nucleosomes were determined, as well as the distance d between the −1 and +1 nucleosome centers (Table 1). Coordinates for −1 and +1 are relative to the translation start site (ATG) of URA3.
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gkr1311-F2: The nucleosome profile of the URA3 promoter is determined by the positions of the −1 and +1 nucleosomes. (A) Using tiling qPCR, we determined the nucleosome profile of the full-length URA3 promoter at the native URA3 locus. Peaks of PCR signal represent enrichment of DNA segments protected by nucleosomes from micrococcal nuclease digestion, and valleys represent nucleosome-free segments. Peak positions corresponding to the centers of the −1 and +1 nucleosomes were calculated (see ‘Materials and Methods’ section), as well as the distance d between the −1 and +1 nucleosome centers (Table 1). (B) The URA3 gene including the (truncated) promoter was inserted at various locations p in the LYS2 gene. Before insertion, n is the position of the center of the first nucleosome upstream of the insertion site p. This upstream nucleosome might influence the positions of the −1 and +1 nucleosomes of the URA3 promoter after insertion. Coordinates for n are relative to the insertion site p. After insertion, the positions of the centers of the −1 and +1 nucleosomes were determined, as well as the distance d between the −1 and +1 nucleosome centers (Table 1). Coordinates for −1 and +1 are relative to the translation start site (ATG) of URA3.

Mentions: To investigate the influence of chromatin context on local nucleosome positioning, we inserted the URA3 gene at various locations in the S. cerevisiae genome and explored the consequences for URA3 nucleosome distribution. URA3 was chosen because the nucleosome structure of this gene is well characterized and reflects the nucleosome pattern of a typical yeast gene, containing a 5′ NFR that is surrounded by two highly localized nucleosomes (Figure 1A, 2A) (27,32,43). Moreover, expression of URA3 can be easily estimated by growth on two different substrates: SC-ura, a growth medium lacking uracil and allowing growth only if URA3 is induced, and 5-FOA, allowing growth in the absence of basal URA3 transcription (44,45).Figure 1.


Distal chromatin structure influences local nucleosome positions and gene expression.

Jansen A, van der Zande E, Meert W, Fink GR, Verstrepen KJ - Nucleic Acids Res. (2012)

The nucleosome profile of the URA3 promoter is determined by the positions of the −1 and +1 nucleosomes. (A) Using tiling qPCR, we determined the nucleosome profile of the full-length URA3 promoter at the native URA3 locus. Peaks of PCR signal represent enrichment of DNA segments protected by nucleosomes from micrococcal nuclease digestion, and valleys represent nucleosome-free segments. Peak positions corresponding to the centers of the −1 and +1 nucleosomes were calculated (see ‘Materials and Methods’ section), as well as the distance d between the −1 and +1 nucleosome centers (Table 1). (B) The URA3 gene including the (truncated) promoter was inserted at various locations p in the LYS2 gene. Before insertion, n is the position of the center of the first nucleosome upstream of the insertion site p. This upstream nucleosome might influence the positions of the −1 and +1 nucleosomes of the URA3 promoter after insertion. Coordinates for n are relative to the insertion site p. After insertion, the positions of the centers of the −1 and +1 nucleosomes were determined, as well as the distance d between the −1 and +1 nucleosome centers (Table 1). Coordinates for −1 and +1 are relative to the translation start site (ATG) of URA3.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3351160&req=5

gkr1311-F2: The nucleosome profile of the URA3 promoter is determined by the positions of the −1 and +1 nucleosomes. (A) Using tiling qPCR, we determined the nucleosome profile of the full-length URA3 promoter at the native URA3 locus. Peaks of PCR signal represent enrichment of DNA segments protected by nucleosomes from micrococcal nuclease digestion, and valleys represent nucleosome-free segments. Peak positions corresponding to the centers of the −1 and +1 nucleosomes were calculated (see ‘Materials and Methods’ section), as well as the distance d between the −1 and +1 nucleosome centers (Table 1). (B) The URA3 gene including the (truncated) promoter was inserted at various locations p in the LYS2 gene. Before insertion, n is the position of the center of the first nucleosome upstream of the insertion site p. This upstream nucleosome might influence the positions of the −1 and +1 nucleosomes of the URA3 promoter after insertion. Coordinates for n are relative to the insertion site p. After insertion, the positions of the centers of the −1 and +1 nucleosomes were determined, as well as the distance d between the −1 and +1 nucleosome centers (Table 1). Coordinates for −1 and +1 are relative to the translation start site (ATG) of URA3.
Mentions: To investigate the influence of chromatin context on local nucleosome positioning, we inserted the URA3 gene at various locations in the S. cerevisiae genome and explored the consequences for URA3 nucleosome distribution. URA3 was chosen because the nucleosome structure of this gene is well characterized and reflects the nucleosome pattern of a typical yeast gene, containing a 5′ NFR that is surrounded by two highly localized nucleosomes (Figure 1A, 2A) (27,32,43). Moreover, expression of URA3 can be easily estimated by growth on two different substrates: SC-ura, a growth medium lacking uracil and allowing growth only if URA3 is induced, and 5-FOA, allowing growth in the absence of basal URA3 transcription (44,45).Figure 1.

Bottom Line: In addition, we show that changes in the nucleosome positions in the URA3 promoter strongly affect the promoter activity.Most interestingly, in addition to demonstrating the effect of the local DNA sequence, our study provides novel in vivo evidence that nucleosome positions are also affected by the position of neighboring nucleosomes.Nucleosome structure may therefore be an important selective force for conservation of gene order on a chromosome, because relocating a gene to another genomic position (where the positions of neighboring nucleosomes are different from the original locus) can have dramatic consequences for the gene's nucleosome structure and thus its expression.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Systems Biology, VIB, Bio-Incubator, Gaston Geenslaan 1, B-3001, Leuven, Belgium.

ABSTRACT
The positions of nucleosomes across the genome influence several cellular processes, including gene transcription. However, our understanding of the factors dictating where nucleosomes are located and how this affects gene regulation is still limited. Here, we perform an extensive in vivo study to investigate the influence of the neighboring chromatin structure on local nucleosome positioning and gene expression. Using truncated versions of the Saccharomyces cerevisiae URA3 gene, we show that nucleosome positions in the URA3 promoter are at least partly determined by the local DNA sequence, with so-called 'anti-nucleosomal elements' like poly(dA:dT) tracts being key determinants of nucleosome positions. In addition, we show that changes in the nucleosome positions in the URA3 promoter strongly affect the promoter activity. Most interestingly, in addition to demonstrating the effect of the local DNA sequence, our study provides novel in vivo evidence that nucleosome positions are also affected by the position of neighboring nucleosomes. Nucleosome structure may therefore be an important selective force for conservation of gene order on a chromosome, because relocating a gene to another genomic position (where the positions of neighboring nucleosomes are different from the original locus) can have dramatic consequences for the gene's nucleosome structure and thus its expression.

Show MeSH