Abstract
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.
Highlights
The fundamental building block of the eukaryotic chromosome is the nucleosome, which consists of 147 bp of DNA, wrapped 1.65 times around an octamer of core histone proteins [reviewed in 1–3]
Reconstituted chromatin was digested with micrococcal nuclease (MNase) and the nucleosome core particle DNAs were sequenced by utilization of the Illumina Solexa technology
Tetranucleotides vs. Dinucleotides Perhaps the most distinguishing sequence characteristic of positioned nucleosomes is the periodic occurrences of certain dinucleotides, and this feature forms the basis of many models that have been used for predicting nucleosome occupancy from nucleotide sequence [7,16,28,29,35]
Summary
The fundamental building block of the eukaryotic chromosome is the nucleosome, which consists of 147 bp of DNA, wrapped 1.65 times around an octamer of core histone proteins [reviewed in 1–3]. During the past five years, large scale sequencing approaches and microarray hybridization technology have permitted the localization of the majority of nucleosomes in the genomes of yeast, worms, flies and humans, and these genome-wide studies have revealed that a surprisingly large fraction of nucleosomes are wellordered with respect to their positions along the chromosomes [3,4,5,6,7,8,9,10,11,12,13,14] These results are in agreement with a large body of earlier work, which has shown that nucleosomes are distributed in a nonrandom fashion along the eukaryotic chromosome [1]. Elucidation of the factors that govern nucleosome positioning is required for a better understanding of genome regulation
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