La transformation en ondelettes continue : un microscope mathématique adapté à l'étude des propriétés d'invariance d'échelle et de corrélations à longue portée des séquences d'ADN

Depuis le début des années 90, l'intérêt des mathématiciens, physiciens et informaticiens pour l'analyse statistique des séquences d'ADN n'a pas cessé de croître. En effet, les immenses progrès de la biologie moléculaire et les grands projets de séquençage ont révélé l'extraordinaire complexité des génomes. Afin de mieux comprendre l'organisation et l'évolution des génomes, il est apparu nécessaire d'introduire de nouveaux concepts et de nouvelles techniques d'analyse du signal. Ainsi la possibilité que les séquences d'ADN présentent des propriétés d'invariance d'échelle associées à l'existence de corrélations à longue portée (CLP) a été le sujet d'une longue controverse. La raison principale de ce malentendu est le caractère non stationnaire des séquences d'ADN résultant de l'hétérogénéité de composition des génomes. Cette observation nous a conduit à proposer l'utilisation de la transformation en ondelettes continue (TO) comme outil naturel d'analyse des séquences d'ADN : par un choix adéquat de l'ondelette analysatrice, on peut s'affranchir de la "structure mosaïque" de ces séquences et quantifier l'existence de CLP associées à des propriétés d'invariance d'échelle monofractales. L'exploration de séquences d'ADN du génome humain sous l'optique du microscope TO, nous a permis de démontrer l'existence de CLP dans les séquences exoniques (codantes pour les protéines) comme dans les séquences introniques (non codantes), remettant par là en cause les différentes interprétations de ces corrélations à l'aide de modèles de dynamique évolutive (plasticité) des génomes. En profitant de la disponibilité de génomes complets (levure, E. coli,...) et en utilisant différentes tables expérimentales associant des di- ou tri- nucléotides à des grandeurs de nature structurelle (courbure, flexibilité), nous avons montré récemment qu'il est possible d'extraire des séquences d'ADN des informations sur l'organisation spatiale et dynamique de la double hélice dans les cellules via l'interaction avec certaines protéines de structure telles que les histones pour la formation du nucléosome eucaryote. En particulier, l'existence et la nature des CLP jusqu'à des distances de l'ordre de 3 104 nucléotides dans certains profils de courbure et/ou de flexibilité locales permettent de diagnostiquer la présence de nucléosomes dans les génomes étudiés. L'observation de certaines CLP dans tous les organismes eucaryotes ainsi que dans les organismes des deux autres règnes (eubactéries et archaeabactéries) suggère fortement que ces corrélations pourraient être essentielles aux phénomènes de condensation-décondensation de la chromatine en relation avec les processus de réplication, transcription et division cellulaire

Open chromatin encoded in DNA sequence is the signature of ‘master’ replication origins in human cells

For years, progress in elucidating the mechanisms underlying replication initiation and its coupling to transcriptional activities and to local chromatin structure has been hampered by the small number (approximately 30) of well-established origins in the human genome and more generally in mammalian genomes. Recent in silico studies of compositional strand asymmetries revealed a high level of organization of human genes around 1000 putative replication origins. Here, by comparing with recently experimentally identified replication origins, we provide further support that these putative origins are active in vivo. We show that regions ∼300-kb wide surrounding most of these putative replication origins that replicate early in the S phase are hypersensitive to DNase I cleavage, hypomethylated and present a significant enrichment in genomic energy barriers that impair nucleosome formation (nucleosome-free regions). This suggests that these putative replication origins are specified by an open chromatin structure favored by the DNA sequence. We discuss how this distinctive attribute makes these origins, further qualified as ‘master’ replication origins, priviledged loci for future research to decipher the human spatio-temporal replication program. Finally, we argue that these ‘master’ origins are likely to play a key role in genome dynamics during evolution and in pathological situations

Neurodevelopment Genes in Lampreys Reveal Trends for Forebrain Evolution in Craniates

The forebrain is the brain region which has undergone the most dramatic changes through vertebrate evolution. Analyses conducted in lampreys are essential to gain insight into the broad ancestral characteristics of the forebrain at the dawn of vertebrates, and to understand the molecular basis for the diversifications that have taken place in cyclostomes and gnathostomes following their splitting. Here, we report the embryonic expression patterns of 43 lamprey genes, coding for transcription factors or signaling molecules known to be involved in cell proliferation, stemcellness, neurogenesis, patterning and regionalization in the developing forebrain. Systematic expression patterns comparisons with model organisms highlight conservations likely to reflect shared features present in the vertebrate ancestors. They also point to changes in signaling systems –pathways which control the growth and patterning of the neuroepithelium-, which may have been crucial in the evolution of forebrain anatomy at the origin of vertebrates

Replication Fork Polarity Gradients Revealed by Megabase-Sized U-Shaped Replication Timing Domains in Human Cell Lines

In higher eukaryotes, replication program specification in different cell types remains to be fully understood. We show for seven human cell lines that about half of the genome is divided in domains that display a characteristic U-shaped replication timing profile with early initiation zones at borders and late replication at centers. Significant overlap is observed between U-domains of different cell lines and also with germline replication domains exhibiting a N-shaped nucleotide compositional skew. From the demonstration that the average fork polarity is directly reflected by both the compositional skew and the derivative of the replication timing profile, we argue that the fact that this derivative displays a N-shape in U-domains sustains the existence of large-scale gradients of replication fork polarity in somatic and germline cells. Analysis of chromatin interaction (Hi-C) and chromatin marker data reveals that U-domains correspond to high-order chromatin structural units. We discuss possible models for replication origin activation within U/N-domains. The compartmentalization of the genome into replication U/N-domains provides new insights on the organization of the replication program in the human genome

Evidence for Sequential and Increasing Activation of Replication Origins along Replication Timing Gradients in the Human Genome

Genome-wide replication timing studies have suggested that mammalian chromosomes consist of megabase-scale domains of coordinated origin firing separated by large originless transition regions. Here, we report a quantitative genome-wide analysis of DNA replication kinetics in several human cell types that contradicts this view. DNA combing in HeLa cells sorted into four temporal compartments of S phase shows that replication origins are spaced at 40 kb intervals and fire as small clusters whose synchrony increases during S phase and that replication fork velocity (mean 0.7 kb/min, maximum 2.0 kb/min) remains constant and narrowly distributed through S phase. However, multi-scale analysis of a genome-wide replication timing profile shows a broad distribution of replication timing gradients with practically no regions larger than 100 kb replicating at less than 2 kb/min. Therefore, HeLa cells lack large regions of unidirectional fork progression. Temporal transition regions are replicated by sequential activation of origins at a rate that increases during S phase and replication timing gradients are set by the delay and the spacing between successive origin firings rather than by the velocity of single forks. Activation of internal origins in a specific temporal transition region is directly demonstrated by DNA combing of the IGH locus in HeLa cells. Analysis of published origin maps in HeLa cells and published replication timing and DNA combing data in several other cell types corroborate these findings, with the interesting exception of embryonic stem cells where regions of unidirectional fork progression seem more abundant. These results can be explained if origins fire independently of each other but under the control of long-range chromatin structure, or if replication forks progressing from early origins stimulate initiation in nearby unreplicated DNA. These findings shed a new light on the replication timing program of mammalian genomes and provide a general model for their replication kinetics

Transcription-coupled and splicing-coupled strand asymmetries in eukaryotic genomes

Under no-strand bias conditions, each genomic DNA strand should present equimolarities of A and T and of G and C. Deviations from these rules are attributed to asymmetric properties intrinsic to DNA mutation–repair processes. In bacteria, strand biases are associated with replication or transcription. In eukaryotes, recent studies demonstrate that human genes present transcription-coupled biases that might reflect transcription-coupled repair processes. Here, we study strand asymmetries in intron sequences of evolutionarily distant eukaryotes, and show that two superimposed intron biases can be distinguished. (i) Biases that are maximum at intron extremities and decrease over large distances to zero values in internal regions, possibly reflecting interactions between pre-mRNA and splicing machinery; these extend over ∼0.5 kb in mammals and Arabidopsis thaliana, and over 1 kb in Caenorhabditis elegans and Drosophila melanogaster. (ii) Biases that are constant along introns, possibly associated with transcription. Strikingly, in C.elegans, these latter biases extend over intergenic regions that separate co-oriented genes. When appropriately examined, all genomes present transcription-coupled excess of T over A in the coding strand. On the opposite, GC skews are either positive (mammals, plants) or negative (invertebrates). These results suggest that transcription-coupled asymmetries result from mutation–repair mechanisms that differ between vertebrates and invertebrates

A novel strategy of transcription regulation by intragenic nucleosome ordering.

Numerous studies of chromatin structure showed that nucleosome free regions (NFRs) located at 5' gene ends contribute to transcription initiation regulation. Here, we determine the role of intragenic chromatin structure on gene expression regulation. We show that, along Saccharomyces cerevisiae genes, nucleosomes are highly organized following two types of architecture that depend only on the distance between the NFRs located at the 5' and 3' gene ends. In the first type, this distance constrains in vivo the positioning of n nucleosomes regularly organized in a "crystal-like" array. In the second type, this distance is such that the corresponding genes can accommodate either n or (n + 1) nucleosomes, thereby displaying two possible crystal-like arrays of n weakly compacted or n + 1 highly compacted nucleosomes. This adaptability confers "bi-stable" properties to chromatin and is a key to its dynamics. Compared to crystal-like genes, bi-stable genes present higher transcriptional plasticity, higher sensitivity to chromatin regulators, higher H3 turnover rate, and lower H2A.Z enrichment. The results strongly suggest that transcription elongation is facilitated by higher chromatin compaction. The data allow us to propose a new paradigm of transcriptional control mediated by the stability and the level of compaction of the intragenic chromatin architecture and open new ways for investigating eukaryotic gene expression regulation

DNA physical properties determine nucleosome occupancy from yeast to fly.

Epigénome er paléogénomieInternational audienceNucleosome positioning plays an essential role in cellular processes by modulating accessibility of DNA to proteins. Here, using only sequence-dependent DNA flexibility and intrinsic curvature, we predict the nucleosome occupancy along the genomes of Saccharomyces cerevisiae and Drosophila melanogaster and demonstrate the predictive power and universality of our model through its correlation with experimentally determined nucleosome occupancy data. In yeast promoter regions, the computed average nucleosome occupancy closely superimposes with experimental data, exhibiting a <200 bp region unfavourable for nucleosome formation bordered by regions that facilitate nucleosome formation. In the fly, our model faithfully predicts promoter strength as encoded in distinct chromatin architectures characteristic of strongly and weakly expressed genes. We also predict that nucleosomes are repositioned by active mechanisms at the majority of fly promoters. Our model uses only basic physical properties to describe the wrapping of DNA around the histone core, yet it captures a substantial part of chromatin's structural complexity, thus leading to a much better prediction of nucleosome occupancy than methods based merely on periodic curved DNA motifs. Our results indicate that the physical properties of the DNA chain, and not just the regulatory factors and chromatin-modifying enzymes, play key roles in eukaryotic transcription

Complete Sequence of the Intronless Mitochondrial Genome of the Saccharomyces cerevisiae Strain CW252

WOS:000452362800008The mitochondrial genomes of strains contain up to 13 introns. An intronless recombinant genome introduced into the nuclear background of strain W303 gave the CW252 strain, which is used to model mitochondrial respiratory pathologies. The complete sequence of this mitochondrial genome was obtained using a hybrid assembling methodology