Perspective CpG Islands: Starting Blocks for Replication and Transcription Marie-Noe¨ lle Prioleau* Institut Jacques Monod, CNRS, Universite´ Paris 7, Universite´ Pierre et Marie Curie, Paris, France The ongoing release of numerous ge- nome sequences constitutes a scientific landmark that has transformed our way of thinking about the organization of living matter. The complete list of annotated genes that is finally at hand pushes us to a stage where we can now explore how genomes harmonize the regulation of gene expression with other processes that must concurrently take place along the genome. A major unresolved question pertaining to mammalian genomes is how their replica- tion is precisely coordinated so that exactly one copy is made at each cell cycle. Despite extensive efforts aimed at under- standing the molecular mechanisms in- volved in the prevention of abnormal replication (i.e., over- or under-replica- tion), the molecular determinants for selecting the starting points (termed ‘‘ori- gins,’’ or ORIs) of replication remain poorly understood [1]. Until recently, only a very small number of ORIs had been mapped, hindering the development of an accurate and global view of origin specification. Indeed, what is true for a specific ORI can be misleading for the majority of ORIs because of the flexibility of mechanisms that are involved in origin selection. As a result, the lack of a statistically relevant dataset has led to precocious and incorrect conclusions. In the past few years, ge- nome-wide studies have been used, at a massive scale, to study transcription and chromatin structure, providing an inte- grated view of genome organization and gene regulation. Several genome-wide studies focusing on the analysis of DNA replication timing have shown a global coordination between gene regulation and replication timing, with GC-rich regions replicating early and AT-rich replicating late [2,3]. Moreover, a recent study showed that 20% of the mouse genome displayed important changes in replication timing upon differentiation of embryonic stem (ES) cells, suggesting a highly dy- namic regulation [4]. Until very recently, however, large-scale mapping of replica- tion origins had been prevented by the technical difficulties inherent to obtaining enough samples with good enrichment in ORI sequences. A few months ago, common features shared by large groups of ORIs were extracted from a large-scale mapping of replication origins based on the resistance of short nascent strands (SNS) to l-exonuclease digestion [5]. This study, conducted in HeLa cells and covering approximately 1% of the human genome, showed that origin density is strongly correlated with genomic land- scapes, with clusters of closely spaced origins in GC-rich regions and no or only few origins in GC-poor regions. Moreover, half of the origins were mapped within or near CpG islands. In this issue of PLoS Genetics, a study by Sequeira-Mendes et al. extends this obser- vation to ORIs in mouse ES cells and shows the conservation of origin specifica- tion mechanisms across species and cell types [6]. In this study, Sequeira-Mendes et al. also explored the relative strength of mapped ORIs. Indeed, in contrast to the first study, in which amplified SNS were hybridized on microarrays, Sequeira- Mendes et al. used nonamplified SNS extracted from a large population of cells, giving them semi-quantitative information on ORI efficiency. They observed that, in general, origins that lie within CpG island promoter are more efficient than non- CpG origins. They confirmed this result on a subset of origins by quantitative analysis in ES cells and fibroblasts. An- other strength of this study is the use of short-size SNS, allowing the authors to precisely localize ORIs. They observed a strong correlation between the 59 end of annotated genes and highly efficient ORIs. This correlation was especially remarkable in the case of promoters containing alternative transcription initiation sites: here, distinct points of replication initia- tion were located immediately adjacent to the mapped transcription start sites. This result raises the question of whether the spatial coincidence between replication and transcription initiation has a function- al significance. It remains to be seen whether the transcriptional and replication machineries recognize similar features. However, it is also important to point out that in both studies, a significant fraction of ORIs are associated neither with known promoters nor with histone modifications indicative of transcriptional activity. Therefore ORI specification, at least for a subset of origins, is not tightly linked to transcription. This subtype of origins could possibly share molecular mechanisms with highly efficient CpG island promoter origins for origin specifi- cation. Alternatively, they might have developed another strategy to trigger replication initiation with lower efficiency. Origin sequences found in human cells are evolutionarily conserved among mam- mals, suggesting the maintenance of spe- cific cis-elements at replication origins [5,7]. The identification of a large collec- tion of origins now gives the opportunity to search for specific over-represented sequence motifs using computational tools for de novo prediction of regulatory elements. Such studies should provide new information on origin selection mech- anisms. It should be noted that methods used in both studies were designed to identify focused origins. Although these studies could detect weak origins of replication active in only 10% of cell cycles, the sensitivity of the analyses was Citation: Prioleau M-N (2009) CpG Islands: Starting Blocks for Replication and Transcription. PLoS Genet 5(4): e1000454. doi:10.1371/journal.pgen.1000454 Editor: Wendy A. Bickmore, Medical Research Council Human Genetics Unit, United Kingdom Published April 10, 2009 Copyright: 2009 Marie-Noe¨lle Prioleau. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The author received no specific funding for this article. Competing Interests: The author has declared that no competing interests exist. * E-mail: prioleau@ijm.jussieu.fr PLoS Genetics | www.plosgenetics.org 1 April 2009 | Volume 5 | Issue 4 | e1000454