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Diversity of eukaryotic DNA replication origins revealed by genome-wide analysis of chromatin structure.

ABSTRACT: Eukaryotic DNA replication origins differ both in their efficiency and in the characteristic time during S phase when they become active. The biological basis for these differences remains unknown, but they could be a consequence of chromatin structure. The availability of genome-wide maps of nucleosome positions has led to an explosion of information about how nucleosomes are assembled at transcription start sites, but no similar maps exist for DNA replication origins. Here we combine high-resolution genome-wide nucleosome maps with comprehensive annotations of DNA replication origins to identify patterns of nucleosome occupancy at eukaryotic replication origins. On average, replication origins contain a nucleosome depleted region centered next to the ACS element, flanked on both sides by arrays of well-positioned nucleosomes. Our analysis identified DNA sequence properties that correlate with nucleosome occupancy at replication origins genome-wide and that are correlated with the nucleosome-depleted region. Clustering analysis of all annotated replication origins revealed a surprising diversity of nucleosome occupancy patterns. We provide evidence that the origin recognition complex, which binds to the origin, acts as a barrier element to position and phase nucleosomes on both sides of the origin. Finally, analysis of chromatin reconstituted in vitro reveals that origins are inherently nucleosome depleted. Together our data provide a comprehensive, genome-wide view of chromatin structure at replication origins and suggest a model of nucleosome positioning at replication origins in which the underlying sequence occludes nucleosomes to permit binding of the origin recognition complex, which then (likely in concert with nucleosome modifiers and remodelers) positions nucleosomes adjacent to the origin to promote replication origin function.

SUBMITTER: Berbenetz NM

PROVIDER: S-EPMC2932696 | biostudies-literature | 2010 Sep

REPOSITORIES: biostudies-literature

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Diversity of eukaryotic DNA replication origins revealed by genome-wide analysis of chromatin structure.

Berbenetz Nicolas M NM Nislow Corey C Brown Grant W GW

PLoS genetics 20100902 9

Eukaryotic DNA replication origins differ both in their efficiency and in the characteristic time during S phase when they become active. The biological basis for these differences remains unknown, but they could be a consequence of chromatin structure. The availability of genome-wide maps of nucleosome positions has led to an explosion of information about how nucleosomes are assembled at transcription start sites, but no similar maps exist for DNA replication origins. Here we combine high-reso ...[more]

PMID: 20824081

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Project description:This study examines the chromatin structure encompassing replication origins in transformed and normal cells. Analysis of the global levels of histone H3 acetylated at K9&14 (open chromatin) and histone H3 trimethylated at K9 (closed chromatin) revealed a higher ratio of open to closed chromatin in the transformed cells. Also, the trithorax and polycomb group proteins, Brg-1 and Bmi-1, respectively, were overexpressed and more abundantly bound to chromatin in the transformed cells. Quantitative comparative analyses of episomal and in situ chromosomal replication origin activity as well as chromatin immunoprecipitation (ChIP) assays, using specific antibodies targeting members of the pre-replication complex (pre-RC) as well as open/closed chromatin markers encompassing both episomal and chromosomal origins, revealed that episomal origins had similar levels of in vivo activity, nascent DNA abundance, pre-RC protein association, and elevated open chromatin structure at the origin in both cell types. In contrast, the chromosomal origins corresponding to 20mer1, 20mer2, and c-myc displayed a 2- to 3-fold higher activity and pre-RC protein abundance as well as higher ratios of open to closed chromatin and of Brg-1 to Bmi-1 in the transformed cells, whereas the origin associated with the housekeeping lamin B2 gene exhibited similar levels of activity, pre-RC protein abundance, and higher ratios of open to closed chromatin and of Brg-1 to Bmi-1 in both cell types. Nucleosomal positioning analysis, using an MNase-Southern blot assay, showed that all the origin regions examined were situated within regions of inconsistently positioned nucleosomes, with the nucleosomes being spaced farther apart from each other prior to the onset of S phase in both cell types. Overall, the results indicate that cellular transformation is associated with differential epigenetic regulation, whereby chromatin structure is more open, rendering replication origins more accessible to initiator proteins, thus allowing increased origin activity.

Dataset Information

Diversity of eukaryotic DNA replication origins revealed by genome-wide analysis of chromatin structure.

Publications

Diversity of eukaryotic DNA replication origins revealed by genome-wide analysis of chromatin structure.

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