Wavelet Analysis of DNA Bending Profiles reveals Structural Constraints on the Evolution of Genomic Sequences

Analyses of genomic DNA sequences have shown in previous works that base pairs are correlated at large distances with scale-invariant statistical properties. We show in the present study that these correlations between nucleotides (letters) result in fact from long-range correlations (LRC) between sequence-dependent DNA structural elements (words) involved in the packaging of DNA in chromatin. Using the wavelet transform technique, we perform a comparative analysis of the DNA text and of the corresponding bending profiles generated with curvature tables based on nucleosome positioning data. This exploration through the optics of the so-called `wavelet transform microscope' reveals a characteristic scale of 100-200 bp that separates two regimes of different LRC. We focus here on the existence of LRC in the small-scale regime ( 200 bp). Analysis of genomes in the three kingdoms reveals that this regime is specifically associated to the presence of nucleosomes. Indeed, small scale LRC are observed in eukaryotic genomes and to a less extent in archaeal genomes, in contrast with their absence in eubacterial genomes. Similarly, this regime is observed in eukaryotic but not in bacterial viral DNA genomes. There is one exception for genomes of Poxviruses, the only animal DNA viruses that do not replicate in the cell nucleus and do not present small scale LRC. Furthermore, no small scale LRC are detected in the genomes of all examined RNA viruses, with one exception in the case of retroviruses. Altogether, these results strongly suggest that small-scale LRC are a signature of the nucleosomal structure. Finally, we discuss possible interpretations of these small-scale LRC in terms of the mechanisms that govern the positioning, the stability and the dynamics of the nucleosomes along the DNA chain. This paper is maily devoted to a pedagogical presentation of the theoretical concepts and physical methods which are well suited to perform a statistical analysis of genomic sequences. We review the results obtained with the so-called wavelet-based multifractal analysis when investigating the DNA sequences of various organisms in the three kingdoms. Some of these results have been announced in B. Audit et al. [1, 2].     

Does Manganese Protect Cultured Human Skin Fibroblasts Against Oxidative Injury by Uva, Dithranol and Hydrogen Peroxide?

Reactive oxygen species (ROS) are involved in the mechanism of photoaging and carcinogenesis. Skin is endowed with antioxidant enzymes including superoxide dismutases (SOD): cytosolic copper zinc SOD and mitochondrial manganese SOD. The aim of our study was to estimate the protective effect of manganese against oxidative injury on cultured human skin fibroblasts. Dithranol, hydrogen peroxide and UV-A radiation (375 nm) were employed as oxidative stressors. The supply of manganese chloride produced an increase in cellular content of this element up to 24 fold without concomitant elevation of MnSOD activity. Nevertheless, manganese protects cells against two of the three ROS generating systems assessed, namely hydrogen peroxyde and UV-A. This protective effect depends on the concentration of manganese in the medium, 0.1 mM and 0.2 mM protect against UVA cytotoxicity, only 0.2 mM protects against H₂O₂ cytotoxicity.     

Modulation of the phase heterogeneity of aminoglycerophospholipid mixtures by sphingomyelin and monovalent cations: maintenance of the lamellar arrangement in the biological membranes

The phase behaviour of mixed molecular species of phosphatidylethanolamine, phosphatidylserine and sphingomyelin of biological origin were examined in aqueous co-dispersions using synchrotron X-ray diffraction. The co-dispersions of phospholipids studied were aimed to model the mixing of lipids populating the cytoplasmic and outer leaflets in the resting or scrambled activated cell membrane. Mixtures enriched with phosphatidylethanolamine and phosphatidylserine were characterized by a phase separation of non-lamellar phases (cubic and inverted hexagonal) with a lamellar gel phase comprising the most saturated molecular species. Inclusion of sphingomyelin in the mixture resulted in a suppression of the hexagonal-II phase in favour of lamellar phases at temperatures where a proportion of the phospholipid was fluid. The effect was also dependent on the total amount of sphingomyelin in ternary mixtures, and the lamellar phase dominated in mixtures containing more than 30 mol%, irrespective of the relative proportions of phosphatidylserine/sphingomyelin. A transition from gel to liquid-crystal phase was detected by wide-angle scattering during heating scans of ternary mixtures enriched in sphingomyelin and was shown by thermal cycling experiments to be coupled with a hexagonal-II phase to lamellar transition. In such samples there was evidence of a coexistence of non-lamellar phases with a lamellar gel phase. A transition of the gel phase to the fluid state on heating from 35 to 41 °C was evidenced by a progressive increase in the lamellar d-spacing. The presence of calcium enhanced the phase separation of a lamellar gel phase from a hexagonal-II phase in mixtures enriched in phosphatidylserine. This effect was counteracted by charge screening with 150 mM NaCl. The effect of sphingomyelin on stabilizing the lamellar phase is discussed in the context of an altered composition in the cytoplasmic/outer leaflets of the plasma membrane resulting from scrambling of the phospholipid distribution. The results suggest that a lamellar structure can be retained by the inward translocation of sphingomyelin in biological membranes. The presence of monovalent cations serves also to stabilize the bilayer in activated cells where a translocation of aminoglycerophospholipids and an influx of calcium occur simultaneously.Abbreviations PC phosphatidylcholine - PE phosphatidylethanolamine - PS phosphatidylserine - SAXS small-angle X-ray scattering - SM sphingomyelin - WAXS wide-angle X-ray scattering - XRD X-ray diffraction     

Halimeda contribution to organic and inorganic production in a Tahitian reef system

Of the seven species of Halimeda inhabiting a lagoon on Moorea island, three representing 10% of the algal covering and averaging 111 g of dry weight m^-2, have been studied in the course of a year. The biomass, measured bimonthly, stresses a slight seasonal variability in the species life. The main decrease was reported for H. opuntia after a fruiting event, which happened in October. The primary production was assessed, in situ, periodically over a year, by measuring oxygen variations in enclosures. Either expressed on specific-weight basis or in area units, the highest primary productions were recorded for H. opuntia. Productions and biomasses vary simultaneously during the year. The three species produce all together about 6 g C m^-2y^-1. The growth rate of the sand-dwelling H. incrassata f. ovata was followed during the year by staining, in the field, individuals with alizarin Red-S. The average rates measured were 3.3 segments ind^-1d^-1 and 0.17 gdw d^-1m^-2. The contribution of the three species to the carbonate budget of the reef was estimated by total alkalinity measurements during 24-h cycles. H. opuntia had the highest CaCO₃ production. For the three species studied, CaCO₃ production of 1.4 kg m^-2y^-1, which could correspond to a 0.4mm/year accretion of the studied reefal system, was estimated.     

FRET or no FRET: a quantitative comparison

Fluorescence resonance energy transfer (FRET) is a technique used to measure the interaction between two molecules labeled with two different fluorophores (the donor and the acceptor) by the transfer of energy from the excited donor to the acceptor. In biological applications, this technique has become popular to qualitatively map protein-protein interactions, and in biophysical projects it is used as a quantitative measure for distances between a single donor and acceptor molecule. Numerous approaches can be found in the literature to quantify and map FRET, but the measures they provide are often difficult to interpret. We propose here a quantitative comparison of these methods by using a surface FRET system with controlled amounts of donor and acceptor fluorophores and controlled distances between them. We support the system with a Monte Carlo simulation of FRET, which provides reference values for the FRET efficiency under various experimental conditions. We validate a representative set of FRET efficiencies and indices calculated from the different methods with different experimental settings. Finally, we test their sensitivity and draw conclusions for the preparation of FRET experiments in more complex and less-controlled systems.     

The role of spatio‐temporal heterogeneity in the establishment and maintenance of Suaeda maritima in salt marshes

Abstract. The effects of disturbance and microtopography on the organization and dynamics of plant communities were studied in a European salt marsh located in the Bay of Mont St. Michel, France. The existence of seed trapping mechanisms was also tested. The study took place in the lower and middle marsh plant communities dominated by the perennials Puccinellia maritima and Halimione portulacoides, respectively and associated with the annual Suaeda maritima. Three treatments were used in series of plots placed in each community: (1) vegetation removal and root destruction to a depth of 10 cm and refilling, (2) non‐remnant herbicide treatment without vegetation removal and (3) creation of depressions (20 cm deep). These treatments were compared with adjacent control plots. The first year of the experiment showed that the perennials facilitated the establishment of Suaeda by trapping its seeds. Estimation of cover, density and biomass over 5 yr following the disturbances showed that in the first 2 yr Suaeda dominated the disturbed plots. Thereafter Suaeda was gradually eliminated by competitive exclusion after ca. 3 yr in the zone originally dominated by Puccinellia maritima and after 4 yr in the zone occupied by Halimione portulacoides. Depressions constituted refuge habitats for Suaeda by limiting competition with the perennials but also led to a high risk of mortality with temporal fluctuations in density. Despite a period of investigation limited to 5 yr, our study demonstrated that natural disturbances of various types occurred and influenced the dynamics of Suaeda, Halimione and Puccinellia. We deduced that natural disturbances and microtopography are responsible for the maintenance of the habitat in a state of non‐equilibrium by favouring the establishment of both spatial and temporal environmental heterogeneity. These conditions appear to be particularly favourable for the maintenance of annual species such as Suaeda maritima.     

Evolution of Bombesin Conjugates for Targeted PET Imaging of Tumors

Bombesin receptors are under intense investigation as molecular targets since they are overexpressed in several prevalent solid tumors. We rationally designed and synthesized a series of modified bombesin (BN) peptide analogs to study the influence of charge and spacers at the N-terminus, as well as amino acid substitutions, on both receptor binding affinity and pharmacokinetics. This enabled development of a novel ^64/67Cu-labeled BN peptide for PET imaging and targeted radiotherapy of BN receptor-positive tumors. Our results show that N-terminally positively charged peptide ligands had significantly higher affinity to human gastrin releasing peptide receptor (GRPr) than negatively charged or uncharged ligands (IC₅₀: 3.2±0.5 vs 26.3±3.5 vs 41.5±2.5 nM). The replacement of Nle¹⁴ by Met, and deletion of D-Tyr⁶, further resulted in 8-fold higher affinity. Contrary to significant changes to human GRPr binding, modifications at the N-terminal and at the 6^th, 11^th, and 14^th position of BN induced only slight influences on affinity to mouse GRPr. [Cu^II]-CPTA-[βAla¹¹] BN(7–14) ([Cu^II]-BZH7) showed the highest internalization rate into PC-3 cells with relatively slow efflux because of its subnanomolar affinity to GRPr. Interestingly, [^64/67Cu]-BZH7 also displayed similar affinities to the other 2 human BN receptor subtypes. In vivo studies showed that [^64/67Cu]-BZH7 had a high accumulation in PC-3 xenografts and allowed for clear-cut visualization of the tumor in PET imaging. In addition, a CPTA-glycine derivative, forming a hippurane-type spacer, enhanced kidney clearance of the radiotracer. These data indicate that the species variation of BN receptor plays an important role in screening radiolabeled BN. As well, the positive charge from the metallated complex at the N-terminal significantly increases affinity to human GRPr. Application of these observations enabled the novel ligand [^64/67Cu]-BZH7 to clearly visualize PC-3 tumors in vivo. This study provides a strong starting point for optimizing radiopeptides for targeting carcinomas that express any of the BN receptor subtypes.