To get insights into the regulation of replication initiation, we systematically mapped replication origins along 1% of the human genome in HeLa cells. We identified 283 origins, 10 times more than previously known. Origin density is strongly correlated with genomic landscapes, with clusters of closely spaced origins in GC-rich regions and no origins in large GC-poor regions. Origin sequences are evolutionarily conserved, and half of them map within or near CpG islands. Most of the origins overlap transcriptional regulatory elements, providing further evidence of a connection with gene regulation. Moreover, we identify c-JUN and c-FOS as important regulators of origin selection. Half of the identified replication initiation sites do not have an open chromatin configuration, showing the absence of a direct link with gene regulation. Replication timing analyses coupled with our origin mapping suggest that a relatively strict origintiming program regulates the replication of the human genome.chromatin structure ͉ DNA replication origin ͉ ENCODE regions ͉ genome-wide mapping ͉ CpG island C ontrolling the number of origins from which replication begins in a given chromosome is necessary to protect it from instability (1, 2). Mapping DNA replication starting points, known as ''origins of replication,'' would make a large contribution to understanding how genome replication is coordinated. Identification of a large number of replication origins is necessary to decipher the rules of origin specification. However, fewer than 30 origins have been identified in human cells (3), and they were mapped mostly in well-characterized transcribed regions, leaving gene-poor regions unexplored. The Encyclopedia of DNA Elements (ENCODE) project (4), launched to develop high-throughput methods for identifying functional elements, now provides a comprehensive view of gene expression and chromatin structure along 1% of the human genome (30 Mb, 44 regions) (5); it thus provides a powerful model for studying interactions among chromosome organization, gene regulation, and initiation of DNA replication. Origin selection is initiated by the binding of the origin recognition complex (ORC) to origin-proximal DNA sequences (6). In contrast to Saccharomyces cerevisiae, characterized metazoan origins do not conform to a clear consensus sequence, and the ORCs from higher eukaryotes exhibit no sequence specificity in vitro (7). Transcription factors at sites of replication initiation have been shown to stimulate replication in many systems, including viruses, yeast, Drosophila, and Xenopus (8, 9). This stimulation may be a consequence of direct interaction with components of the replication machinery or of facilitating the access of the replication complexes to DNA through recruitment of chromatin remodeling complexes (10-12). Here we present a high-resolution map of replication origins in HeLa cells based on hybridization of short nascent strands (SNS) on DNA microarrays covering ENCODE regions. To construct this map we use one of the most string...

DNA replication ensures the accurate duplication of the genome at each cell cycle. It begins at specific sites called replication origins. Genome-wide studies in vertebrates have recently identified a consensus G-rich motif potentially able to form G-quadruplexes (G4) in most replication origins. However, there is no experimental evidence to demonstrate that G4 are actually required for replication initiation. We show here, with two model origins, that G4 motifs are required for replication initiation. Two G4 motifs cooperate in one of our model origins. The other contains only one critical G4, and its orientation determines the precise position of the replication start site. Point mutations affecting the stability of this G4 in vitro also impair origin function. Finally, this G4 is not sufficient for origin activity and must cooperate with a 200-bp cis-regulatory element. In conclusion, our study strongly supports the predicted essential role of G4 in replication initiation.

Interleukin 6 (IL-6), a multifunctional cytokine, has been implicated in the pathophysiology of type 2 diabetes (T2D). The elevated circulating level of IL-6 is an independent predictor of T2D and is considered to be involved in the development of inflammation, insulin resistance and β-cell dysfunction. On the other hand, an increasing number of evidence suggests that IL-6 has an anti-inflammatory role and improves glucose metabolism. The complex signal transduction mechanism of IL-6 may help explain the pleiotropic nature of the cytokine. IL-6 acts via two distinct signalling pathways called classic signalling and trans-signalling. While both signalling modes lead to activation of the same receptor subunit, their final biological effects are completely different. The aim of this review is to summarize our current knowledge about the role of IL-6 in the development of T2D. We will also discuss the importance of specific blockade of IL-6 trans-signalling rather than inhibiting both signalling pathways as a therapeutic strategy for the treatment of T2D and its associated macrovascular complications.