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Review
. 2016 Aug 1;30(15):1683-97.
doi: 10.1101/gad.285114.116.

DNA Replication Origins-Where Do We Begin?

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Free PMC article
Review

DNA Replication Origins-Where Do We Begin?

Marie-Noëlle Prioleau et al. Genes Dev. .
Free PMC article

Abstract

For more than three decades, investigators have sought to identify the precise locations where DNA replication initiates in mammalian genomes. The development of molecular and biochemical approaches to identify start sites of DNA replication (origins) based on the presence of defining and characteristic replication intermediates at specific loci led to the identification of only a handful of mammalian replication origins. The limited number of identified origins prevented a comprehensive and exhaustive search for conserved genomic features that were capable of specifying origins of DNA replication. More recently, the adaptation of origin-mapping assays to genome-wide approaches has led to the identification of tens of thousands of replication origins throughout mammalian genomes, providing an unprecedented opportunity to identify both genetic and epigenetic features that define and regulate their distribution and utilization. Here we summarize recent advances in our understanding of how primary sequence, chromatin environment, and nuclear architecture contribute to the dynamic selection and activation of replication origins across diverse cell types and developmental stages.

Keywords: DNA replication; chromatin; epigenetics; nuclear architecture; origins of DNA replication.

Figures

Figure 1.
Figure 1.
Mapping molecular intermediates of replication initiation. Schematic representation of replication intermediates and their detection by enrichment for short nascent strands (SNSs), replication bubbles, and Okazaki fragments. Three potential origins (two highly efficient and one inefficient) are depicted in a 25-kb region. For simplicity, initiation events are represented on the same molecule as an aggregate across the entire population; however, in reality, initiation events are stochastic and sparse, with the likelihood of more than one origin being activated in this region being very low. Restriction sites used for fragmentation for the bubble-seq method are indicated (restriction sites). Detection of replication bubbles is very sensitive but limited by the resolution of the restriction fragments and subject to false negatives when a restriction site is within the replication bubble. The distribution of Okazaki fragments and the DNA strand (Watson or Crick) to which they map provides information about replication fork direction (RFD); sharp transitions in strandedness mark origins and termination zones. However, this method cannot discriminate between a zone of initiation and a region containing several specific sites used in a stochastic manner. SNS mapping by treatment with λ-exonuclease provides increased resolution but can detect only the most efficient origins of DNA replication.
Figure 2.
Figure 2.
Defining a replication origin. Several features have been proposed to contribute to local origin activity. Nucleosome-free regions (NFRs) may be preferential sites for ORC binding, pre-RC formation, and/or activation. G4 motifs may also favor the formation of NFRs and/or recruitment of specific factors involved in origin licensing or activation. Cooperation with neighboring cis-regulatory sequences and chromatin modifications may also increase the capacity to form a NFR or contribute to the recruitment of trans-factors, influencing origin selection and activation. The site of initiation (ORI) is located ∼220 bp 3′ of the G4 structure, which may also function to transiently impede fork progression on the leading strand.
Figure 3.
Figure 3.
Hierarchy of temporal regulation. Active early, mid-, and late S-phase replication origins are represented as small green, blue, and red dots, respectively. Constant early-replicating domains are enriched for GC content, genes, regulatory elements (transcription factor-binding and enhancer sites), DNase I-hypersensitive sites, and origins of DNA replication. Early-replicating domains are found in the interior of the nucleus and are frequently associated with activating chromatin modifications and histone variants, including H3K4me3, H2AZ, and H3K9ac. Timing transition zones are also origin-rich and frequently associated with polycomb-repressive chromatin environments (H3K27me3). In contrast, late-replicating regions are AT-rich and gene-poor and exhibit a decreased density of origins. Late-replicating domains are associated with nuclear lamins and Rif1 (Rap1-interacting factor 1), which may lead to the formation of a microenvironment at the periphery of the nucleus or the nucleolus that restricts the activity of critical kinases (DDK and CDK) and/or limiting initiation factors.

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