Late Replication Domains in Polytene and Non-Polytene Cells of Drosophila melanogaster

In D. melanogaster polytene chromosomes, intercalary heterochromatin (IH) appears as large dense bands scattered in euchromatin and comprises clusters of repressed genes. IH displays distinctly low gene density, indicative of their particular regulation. Genes embedded in IH replicate late in the S phase and become underreplicated. We asked whether localization and organization of these late-replicating domains is conserved in a distinct cell type. Using published comprehensive genome-wide chromatin annotation datasets (modENCODE and others), we compared IH organization in salivary gland cells and in a Kc cell line. We first established the borders of 60 IH regions on a molecular map, these regions containing underreplicated material and encompassing ∼12% of Drosophila genome. We showed that in Kc cells repressed chromatin constituted 97% of the sequences that corresponded to IH bands. This chromatin is depleted for ORC-2 binding and largely replicates late. Differences in replication timing between the cell types analyzed are local and affect only sub-regions but never whole IH bands. As a rule such differentially replicating sub-regions display open chromatin organization, which apparently results from cell-type specific gene expression of underlying genes. We conclude that repressed chromatin organization of IH is generally conserved in polytene and non-polytene cells. Yet, IH domains do not function as transcription- and replication-regulatory units, because differences in transcription and replication between cell types are not domain-wide, rather they are restricted to small “islands” embedded in these domains. IH regions can thus be defined as a special class of domains with low gene density, which have narrow temporal expression patterns, and so displaying relatively conserved organization

Diversity of Eukaryotic DNA Replication Origins Revealed by Genome-Wide Analysis of Chromatin Structure

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

Separation of DNA Replication from the Assembly of Break-Competent Meiotic Chromosomes

The meiotic cell division reduces the chromosome number from diploid to haploid to form gametes for sexual reproduction. Although much progress has been made in understanding meiotic recombination and the two meiotic divisions, the processes leading up to recombination, including the prolonged pre-meiotic S phase (meiS) and the assembly of meiotic chromosome axes, remain poorly defined. We have used genome-wide approaches in Saccharomyces cerevisiae to measure the kinetics of pre-meiotic DNA replication and to investigate the interdependencies between replication and axis formation. We found that replication initiation was delayed for a large number of origins in meiS compared to mitosis and that meiotic cells were far more sensitive to replication inhibition, most likely due to the starvation conditions required for meiotic induction. Moreover, replication initiation was delayed even in the absence of chromosome axes, indicating replication timing is independent of the process of axis assembly. Finally, we found that cells were able to install axis components and initiate recombination on unreplicated DNA. Thus, although pre-meiotic DNA replication and meiotic chromosome axis formation occur concurrently, they are not strictly coupled. The functional separation of these processes reveals a modular method of building meiotic chromosomes and predicts that any crosstalk between these modules must occur through superimposed regulatory mechanisms

Behavioural and Developmental Interventions for Autism Spectrum Disorder: A Clinical Systematic Review

Background: Much controversy exists regarding the clinical efficacy of behavioural and developmental interventions for improving the core symptoms of autism spectrum disorders (ASD). We conducted a systematic review to summarize the evidence on the effectiveness of behavioural and developmental interventions for ASD. Methods and Findings: Comprehensive searches were conducted in 22 electronic databases through May 2007. Further information was obtained through hand searching journals, searching reference lists, databases of theses and dissertations, and contacting experts in the field. Experimental and observational analytic studies were included if they were written in English and reported the efficacy of any behavioural or developmental intervention for individuals with ASD. Two independent reviewers made the final study selection, extracted data, and reached consensus on study quality. Results were summarized descriptively and, where possible, meta-analyses of the study results were conducted. One-hundred-and-one studies at predominantly high risk of bias that reported inconsistent results across various interventions were included in the review. Meta-analyses of three controlled clinical trials showed that Lovaas treatment was superior to special education on measures of adaptive behaviour, communication and interaction, comprehensive language, daily living skills, expressive language, overall intellectual functioning and socialization. High-intensity Lovaas was superior to low-intensity Lovaas on measures of intellectual functioning in two retrospective cohort studies. Pooling the results of two randomized controlle

Transcription initiation activity sets replication origin efficiency in mammalian cells

Genomic mapping of DNA replication origins (ORIs) in mammals provides a powerful means for understanding the regulatory complexity of our genome. Here we combine a genome-wide approach to identify preferential sites of DNA replication initiation at 0.4% of the mouse genome with detailed molecular analysis at distinct classes of ORIs according to their location relative to the genes. Our study reveals that 85% of the replication initiation sites in mouse embryonic stem (ES) cells are associated with transcriptional units. Nearly half of the identified ORIs map at promoter regions and, interestingly, ORI density strongly correlates with promoter density, reflecting the coordinated organisation of replication and transcription in the mouse genome. Detailed analysis of ORI activity showed that CpG island promoter-ORIs are the most efficient ORIs in ES cells and both ORI specification and firing efficiency are maintained across cell types. Remarkably, the distribution of replication initiation sites at promoter-ORIs exactly parallels that of transcription start sites (TSS), suggesting a co-evolution of the regulatory regions driving replication and transcription. Moreover, we found that promoter-ORIs are significantly enriched in CAGE tags derived from early embryos relative to all promoters. This association implies that transcription initiation early in development sets the probability of ORI activation, unveiling a new hallmark in ORI efficiency regulation in mammalian cellsWork in María Gómez’s laboratory is supported by grants from the Spanish Ministry of Education and Science (BFU2007-66827) and the Consejería de Sanidad of the Junta de Castilla y León (SAN196/SA12/07). JSM was supported by a grant from the Portuguese Foundation for Science and Technology (SFRH/BD/ 11824/2003

Genome-wide characterization of fission yeast DNA replication origins

Eukaryotic DNA replication is initiated from multiple origins of replication, but little is known about the global regulation of origins throughout the genome or in different types of cell cycles. Here, we identify 401 strong origins and 503 putative weaker origins spaced in total every 14 kb throughout the genome of the fission yeast Schizosaccharomyces pombe. The same origins are used during premeiotic and mitotic S-phases. We found that few origins fire late in mitotic S-phase and that activating the Rad3 dependent S-phase checkpoint by inhibiting DNA replication had little effect on which origins were fired. A genome-wide analysis of eukaryotic origin efficiencies showed that efficiency was variable, with large chromosomal domains enriched for efficient or inefficient origins. Average efficiency is twice as high during mitosis compared with meiosis, which can account for their different S-phase lengths. We conclude that there is a continuum of origin efficiency and that there is differential origin activity in the mitotic and meiotic cell cycles