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Multiple sequence-directed possibilities provide a pool of nucleosome position choices in different states of activity of a gene.

Vinayachandran V, Pusarla RH, Bhargava P - Epigenetics Chromatin (2009)

Bottom Line: While the DNA sequences may help decide their locations, the observed positions in vivo are end-results of chromatin remodeling, the state of gene activity and binding of the sequence-specific factors to the DNA, all of which influence nucleosome positions.Thus, the observed nucleosome locations in vivo do not reflect the true contribution of DNA sequence to the mapped position.On a gene locus, multiple nucleosome positions are directed by a gene sequence to provide a pool of possibilities, out of which the preferred ones are selected by the chromatin remodeler and transcription factor of the gene under different states of activity of the gene.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Cellular and Molecular Biology, (Council of Scientific and Industrial Research), Uppal Road, Hyderabad-500007, India. vinesh@ccmb.res.in

ABSTRACT

Background: Genome-wide mappings of nucleosome occupancy in different species have shown presence of well-positioned nucleosomes. While the DNA sequences may help decide their locations, the observed positions in vivo are end-results of chromatin remodeling, the state of gene activity and binding of the sequence-specific factors to the DNA, all of which influence nucleosome positions. Thus, the observed nucleosome locations in vivo do not reflect the true contribution of DNA sequence to the mapped position. Moreover, the naturally occurring nucleosome-positioning sequences are known to guide multiple translational positionings.

Results: We show that yeast SNR6, a gene transcribed by RNA polymerase III, constitutes nucleosome-positioning sequence. In the absence of a chromatin remodeler or any factor binding, the gene sequence confers a unique rotational phase to nucleosomes in the gene region, and directs assembly of several translationally positioned nucleosomes on approximately 1.2 kb DNA from the gene locus, including the short approximately 250 bp gene region. Mapping of all these gene sequence-directed nucleosome positions revealed that the array of nucleosomes in the gene upstream region occupy the same positions as those observed in vivo but the nucleosomes on the gene region can be arranged in three distinct registers. Two of these arrangements differ from each other in the position of only one nucleosome, and match with the nucleosome positions on the gene in repressed and active states in vivo, where the gene-specific factor is known to occupy the gene in both the states. The two positions are interchanged by an ATP-dependent chromatin remodeler in vivo. The third register represents the positions which block the access of the factor to the gene promoter elements.

Conclusion: On a gene locus, multiple nucleosome positions are directed by a gene sequence to provide a pool of possibilities, out of which the preferred ones are selected by the chromatin remodeler and transcription factor of the gene under different states of activity of the gene.

No MeSH data available.


Related in: MedlinePlus

Nucleosome positioning properties of two SNR6 halves are different. (A) Schematic representation of the selected gene regions PCR-amplified from pCS6 and cloned into a plasmid along with their names and sizes is given. (B), (C) and (D) Indirect end-labeling analysis of the chromatin assembled over the plasmids 601c, 5' half and 3' half respectively. Arrows mark the positions of the genomic DNA ends while ovals mark the positioned nucleosomes. Lanes 1 and 2 show digestion pattern of naked DNA N, while digestion pattern of the chromatin samples C are shown in the lanes 3 and 4 in each panel. Probe was same as that in the Figure 1C while lane M shows molecular size markers.
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Figure 4: Nucleosome positioning properties of two SNR6 halves are different. (A) Schematic representation of the selected gene regions PCR-amplified from pCS6 and cloned into a plasmid along with their names and sizes is given. (B), (C) and (D) Indirect end-labeling analysis of the chromatin assembled over the plasmids 601c, 5' half and 3' half respectively. Arrows mark the positions of the genomic DNA ends while ovals mark the positioned nucleosomes. Lanes 1 and 2 show digestion pattern of naked DNA N, while digestion pattern of the chromatin samples C are shown in the lanes 3 and 4 in each panel. Probe was same as that in the Figure 1C while lane M shows molecular size markers.

Mentions: The complete SNR6 gene region from TATA box at -30 to box B up to +242 bp can probably be occupied by two contiguous nucleosomes (Figure 1C). The nucleosome positioning signal on the sea urchin 5S rRNA gene is centered around the +1 position, giving a nucleosome positioned from -78 to +78 bp [54]. As box B of SNR6 is further downstream, similar positioning on SNR6 would allow formation of one more nucleosome on the gene region, downstream of +90 bp position. Therefore, we separated the SNR6 gene sequence into two halves and cloned the bp regions -87 to +83 (5' half of the gene) as well as +62 to +256 (3' half of the gene) into two different plasmids (Figure 4A). We used the IEL technique to further confirm the nucleosome-positioning properties of both the halves in the context of the flanking plasmid vector sequences.


Multiple sequence-directed possibilities provide a pool of nucleosome position choices in different states of activity of a gene.

Vinayachandran V, Pusarla RH, Bhargava P - Epigenetics Chromatin (2009)

Nucleosome positioning properties of two SNR6 halves are different. (A) Schematic representation of the selected gene regions PCR-amplified from pCS6 and cloned into a plasmid along with their names and sizes is given. (B), (C) and (D) Indirect end-labeling analysis of the chromatin assembled over the plasmids 601c, 5' half and 3' half respectively. Arrows mark the positions of the genomic DNA ends while ovals mark the positioned nucleosomes. Lanes 1 and 2 show digestion pattern of naked DNA N, while digestion pattern of the chromatin samples C are shown in the lanes 3 and 4 in each panel. Probe was same as that in the Figure 1C while lane M shows molecular size markers.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2667510&req=5

Figure 4: Nucleosome positioning properties of two SNR6 halves are different. (A) Schematic representation of the selected gene regions PCR-amplified from pCS6 and cloned into a plasmid along with their names and sizes is given. (B), (C) and (D) Indirect end-labeling analysis of the chromatin assembled over the plasmids 601c, 5' half and 3' half respectively. Arrows mark the positions of the genomic DNA ends while ovals mark the positioned nucleosomes. Lanes 1 and 2 show digestion pattern of naked DNA N, while digestion pattern of the chromatin samples C are shown in the lanes 3 and 4 in each panel. Probe was same as that in the Figure 1C while lane M shows molecular size markers.
Mentions: The complete SNR6 gene region from TATA box at -30 to box B up to +242 bp can probably be occupied by two contiguous nucleosomes (Figure 1C). The nucleosome positioning signal on the sea urchin 5S rRNA gene is centered around the +1 position, giving a nucleosome positioned from -78 to +78 bp [54]. As box B of SNR6 is further downstream, similar positioning on SNR6 would allow formation of one more nucleosome on the gene region, downstream of +90 bp position. Therefore, we separated the SNR6 gene sequence into two halves and cloned the bp regions -87 to +83 (5' half of the gene) as well as +62 to +256 (3' half of the gene) into two different plasmids (Figure 4A). We used the IEL technique to further confirm the nucleosome-positioning properties of both the halves in the context of the flanking plasmid vector sequences.

Bottom Line: While the DNA sequences may help decide their locations, the observed positions in vivo are end-results of chromatin remodeling, the state of gene activity and binding of the sequence-specific factors to the DNA, all of which influence nucleosome positions.Thus, the observed nucleosome locations in vivo do not reflect the true contribution of DNA sequence to the mapped position.On a gene locus, multiple nucleosome positions are directed by a gene sequence to provide a pool of possibilities, out of which the preferred ones are selected by the chromatin remodeler and transcription factor of the gene under different states of activity of the gene.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Cellular and Molecular Biology, (Council of Scientific and Industrial Research), Uppal Road, Hyderabad-500007, India. vinesh@ccmb.res.in

ABSTRACT

Background: Genome-wide mappings of nucleosome occupancy in different species have shown presence of well-positioned nucleosomes. While the DNA sequences may help decide their locations, the observed positions in vivo are end-results of chromatin remodeling, the state of gene activity and binding of the sequence-specific factors to the DNA, all of which influence nucleosome positions. Thus, the observed nucleosome locations in vivo do not reflect the true contribution of DNA sequence to the mapped position. Moreover, the naturally occurring nucleosome-positioning sequences are known to guide multiple translational positionings.

Results: We show that yeast SNR6, a gene transcribed by RNA polymerase III, constitutes nucleosome-positioning sequence. In the absence of a chromatin remodeler or any factor binding, the gene sequence confers a unique rotational phase to nucleosomes in the gene region, and directs assembly of several translationally positioned nucleosomes on approximately 1.2 kb DNA from the gene locus, including the short approximately 250 bp gene region. Mapping of all these gene sequence-directed nucleosome positions revealed that the array of nucleosomes in the gene upstream region occupy the same positions as those observed in vivo but the nucleosomes on the gene region can be arranged in three distinct registers. Two of these arrangements differ from each other in the position of only one nucleosome, and match with the nucleosome positions on the gene in repressed and active states in vivo, where the gene-specific factor is known to occupy the gene in both the states. The two positions are interchanged by an ATP-dependent chromatin remodeler in vivo. The third register represents the positions which block the access of the factor to the gene promoter elements.

Conclusion: On a gene locus, multiple nucleosome positions are directed by a gene sequence to provide a pool of possibilities, out of which the preferred ones are selected by the chromatin remodeler and transcription factor of the gene under different states of activity of the gene.

No MeSH data available.


Related in: MedlinePlus