ChIP-on-chip
Characterizing Regulatory Control of DNA Replication in Eukaryotic Cells
A review of Genome-wide analysis of re-replication reveals inhibitory controls that target multiple stages of replication initiation.
Note: This is a review of the published article listed below. All information, quotes, figures, methods, and findings mentioned in this review are from that article, and are the property of its authors and/or the publication in which the article originally appeared.
Tanny and colleagues at the Howard Hughes Medical Institute (2006) use chromatin immunoprecipitation microarrays to perform genome-wide analysis of re-replication, revealing inhibitory controls critical to genome maintenance. The group has developed custom 44K Agilent microarrays to use with the model organism Saccharomyces cerevisiae to study how eukaryotic cells prevent replication origins from reinitiating during a single cell cycle. Using the Agilent microarrays, the group was able to map sites across the S. cerevisiae genome that are re-replicated as well as sites of pre-RC formation during re-replication, demonstrating that only a fraction of the genome is re-replicated by a subset of origins, some of which are capable of multiple re-initiation events. The findings of this group are valuable to the characterization of genomic re-replication, shedding new light on DNA replication processes in eukaryotic cells.
Figure 1. Analysis of genome-wide re-replication.
Re-replication is detected by copy number analysis using DNA microarrays. DNA from re-replicating cells and from G1-arrested cells was differentially labeled and cohybridized to a high-density DNA microarray. The log ratio re-replicated/unreplicated for each spot was plotted as a function of its position along the chromosome. Chromosome IV (see Supplementary Figure 2 for other chromosomes) is shown here as an example. (B) Sites of re-replication initiation are associated with G1 Mcm2-7 binding sites. A smoothing algorithm and a significance cutoff was applied to the re-replication data (see Materials and Methods) and plotted here for Chromosome IV (gray histogram). G1 Mcm2-7 binding sites (black histogram), determined by genome-wide location analysis, are superimposed on top of the re-replication data. Key sites discussed throughout the text, ARS1, ARS418, ARS428, and iYDR309C, are marked with a gray dashed line.
Figure 2. Subtelomeric regions have a high probability of re-replicating.
(A) There is a positive correlation between the proximity of a sequence to the telomere and its probability of re-replicating. The relative enrichment for each spot on the microarray was plotted as a function of its distance to the closest telomere for both the re-replicating strain (black) and wild-type strain (gray) 3 h after addition of galactose. (B) There is no correlation between re-replication and proximity to centromeres. The relative enrichment for each spot on the microarray was plotted as a function of its distance to the centromere for both the re-replicating strain (black) and wild-type strain (gray) 3 h after addition of galactose.
Title: Genome-wide Analysis of Re-replication Reveals Inhibitory Controls That Target Multiple Stages of Replication Initiation.
Authors: Tanny RE, MacAlpine DM, Blitzblau HG, Bell SP.
Journal: Mol Biol Cell. 2006 May; 17(5): 2415-2423.
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