Quantitative four-dimensional tracking of cytoplasmic and nuclear HIV-1 complexes

Emerging real-time techniques for imaging viral infections provide powerful tools for understanding the dynamics of virus-host cell interactions. Here we labeled human immunodeficiency virus-1 (HIV-1) integrase with a small tetracysteine tag, which preserved the virus' infectivity while allowing it to be labeled with the bis-arsenical fluorescein derivative FlAsH. This labeling allowed us to image both intracytoplasmic and intranuclear HIV-1 complexes in three dimensions over time (4D) in human cells and enabled us to analyze HIV-1 kinetics by automated 4D quantitative particle tracking. In the cytoplasm, HIV-1 complexes underwent directed movements toward the nuclear compartment, kinetically characteristic of both microtubule- and actin-dependent transport. The complexes then adopted smaller movements in a very confined volume once associated with the nuclear membrane and more diffuse movements once inside the nucleus. This work contributes new insight into the various movements of HIV-1 complexes within infected cells and provides a useful tool for the study of virus-host cell interactions during infection.     

Sensitivity of methods for estimating breeding values using genetic markers to the number of QTL and distribution of QTL variance

The objective of this simulation study was to compare the effect of the number of QTL and distribution of QTL variance on the accuracy of breeding values estimated with genomewide markers (MEBV). Three distinct methods were used to calculate MEBV: a Bayesian Method (BM), Least Angle Regression (LARS) and Partial Least Square Regression (PLSR). The accuracy of MEBV calculated with BM and LARS decreased when the number of simulated QTL increased. The accuracy decreased more when QTL had different variance values than when all QTL had an equal variance. The accuracy of MEBV calculated with PLSR was affected neither by the number of QTL nor by the distribution of QTL variance. Additional simulations and analyses showed that these conclusions were not affected by the number of individuals in the training population, by the number of markers and by the heritability of the trait. Results of this study show that the effect of the number of QTL and distribution of QTL variance on the accuracy of MEBV depends on the method that is used to calculate MEBV.     

Unusual entropy-driven affinity of Chromobacterium violaceum lectin CV-IIL toward fucose and mannose

The purple pigmented bacterium Chromobacterium violaceum is a dominant component of tropical soil microbiota that can cause rare but fatal septicaemia in humans. Its sequenced genome provides insight into the abundant potential of this organism for biotechnological and pharmaceutical applications and allowed an ORF encoding a protein that is 60% identical to the fucose binding lectin (PA-IIL) from Pseudomonas aeruginosa and the mannose binding lectin (RS-IIL) from Ralstonia solanacearum to be identified. The lectin, CV-IIL, has recently been purified from C. violaceum [Zinger-Yosovich, K., Sudakevitz, D., Imberty, A., Garber, N. C., and Gilboa-Garber, N. (2006) Microbiology 152, 457-463] and has been confirmed to be a tetramer with subunit size of 11.86 kDa and a binding preference for fucose. We describe here the cloning of CV-IIL and its expression as a recombinant protein. A complete structure-function characterization has been made in an effort to analyze the specificity and affinity of CV-IIL for fucose and mannose. Crystal structures of CV-IIL complexes with monosaccharides have yielded the molecular basis of the specificity. Each monomer contains two close calcium cations that mediate the binding of the monosaccharides, which occurs in different orientations for fucose and mannose. The thermodynamics of binding has been analyzed by titration microcalorimetry, giving dissociation constants of 1.7 and 19 microM for alpha-methyl fucoside and alpha-methyl mannoside, respectively. Further analysis demonstrated a strongly favorable entropy term that is unusual in carbohydrate binding. A comparison with both PA-IIL and RS-IIL, which have binding preferences for fucose and mannose, respectively, yielded insights into the monosaccharide specificity of this important class of soluble bacterial lectins.     

The tumor suppressor Apc controls planar cell polarities central to gut homeostasis

The stem cells (SCs) at the bottom of intestinal crypts tightly contact niche-supporting cells and fuel the extraordinary tissue renewal of intestinal epithelia. Their fate is regulated stochastically by populational asymmetry, yet whether asymmetrical fate as a mode of SC division is relevant and whether the SC niche contains committed progenitors of the specialized cell types are under debate. We demonstrate spindle alignments and planar cell polarities, which form a novel functional unit that, in SCs, can yield daughter cell anisotropic movement away from niche-supporting cells. We propose that this contributes to SC homeostasis. Importantly, we demonstrate that some SC divisions are asymmetric with respect to cell fate and provide data suggesting that, in some SCs, mNumb displays asymmetric segregation. Some of these processes were altered in apparently normal crypts and microadenomas of mice carrying germline Apc mutations, shedding new light on the first stages of progression toward colorectal cancer.     

Variation in phenotypic plasticity and selection patterns in blue tit breeding time: between- and within-population comparisons

1.?Phenotypic plasticity, the response of individual phenotypes to their environment, can allow organisms to cope with spatio-temporal variation in environmental conditions. Recent studies have shown that variation exists among individuals in their capacity to adjust their traits to environmental changes and that this individual plasticity can be under strong selection. Yet, little is known on the extent and ultimate causes of variation between populations and individuals in plasticity patterns. 2.?In passerines, timing of breeding is a key life-history trait strongly related to fitness and is known to vary with the environment, but few studies have investigated the within-species variation in individual plasticity. 3.?Here, we studied between- and within-population variation in breeding time, phenotypic plasticity and selection patterns for this trait in four Mediterranean populations of blue tits (Cyanistes caeruleus) breeding in habitats varying in structure and quality. 4.?Although there was no significant warming over the course of the study, we found evidence for earlier onset of breeding in warmer years in all populations, with reduced plasticity in the less predictable environment. In two of four populations, there was significant inter-individual variation in plasticity for laying date. Interestingly, selection for earlier laying date was significant only in populations where there was no inter-individual differences in plasticity. 5.?Our results show that generalization of plasticity patterns among populations of the same species might be challenging even at a small spatial scale and that the amount of within-individual variation in phenotypic plasticity may be linked to selective pressures acting on these phenotypic traits.     

Cross-talk between Staphylococcus aureus leukocidins-intoxicated macrophages and lung epithelial cells triggers chemokine secretion in an inflammasome-dependent manner

Staphylococcus aureus is a major pathogen responsible for both nosocomial and community-acquired infections. Central to its virulence is its ability to secrete haemolysins, pore-forming toxins and cytolytic peptides. The large number of membrane-damaging toxins and peptides produced during S. aureus infections has hindered a precise understanding of their specific roles in diseases. Here, we used comprehensive libraries of recombinant toxins and synthetic cytolytic peptides, of S. aureus mutants and clinical strains to investigate the role of these virulence factors in targeting human macrophages and triggering IL-1β release. We found that the Panton Valentine leukocidin (PVL) is the major trigger of IL-1β release and inflammasome activation in primary human macrophages. The cytolytic peptides, δ-haemolysin and PSMα3; the pore-forming toxins, γ-haemolysin and LukDE; and β-haemolysin synergize with PVL to amplify IL-1β release, indicating that these factors cooperate with PVL to trigger inflammation. PVL(+) S. aureus causes necrotizing pneumonia in children and young adults. The severity of this disease is due to the massive recruitment of neutrophils that cause lung damage. Importantly, we demonstrate that PVL triggers IL-1β release in human alveolar macrophages. Furthermore, IL-1β released by PVL-intoxicated macrophages stimulates the secretion of the neutrophil attracting chemokines, IL-8 and monocyte chemotactic protein-1, by lung epithelial cells. Finally, we show that PVL-induced IL-8/monocyte chemotactic protein-1 release is abolished by the inclusion of IL-1 receptor antagonist (IL-1Ra) in a mixed culture of lung epithelial cells and macrophages. Together, our results identify PVL as the predominant S. aureus secreted factor for triggering inflammasome activation in human macrophages and demonstrate how PVL-intoxicated macrophages orchestrate inflammation in the lung. Finally, our work suggests that anakinra, a synthetic IL-1Ra, may be an effective therapeutic agent to reduce the massive neutrophils infiltration observed during necrotizing pneumonia and decrease the resulting host-mediated lung injury.     

Female coloration indicates female reproductive capacity in blue tits

It is poorly understood whether female morphological and behavioural traits can be used as 'signals'. In particular, experimental tests of the hypothesis that female ornaments reflect quality are scarce. Here, we experimentally examine whether female plumage coloration might signal maternal quality in the blue tit, Cyanistes caeruleus by forcing half of the females breeding in our population to produce a replacement clutch. Using statistical models that controlled for the effects of male coloration, and the effects of age and condition of both parents, we found that carotenoid-based female coloration was positively linked to key proxies of bird lifetime reproductive success: clutch size, fledgling success and recruitment. Importantly, the relationships between maternal yellow carotenoid coloration and both clutch size and recruitment were stronger in the experimental group than in the control group, indicating that breeding females with higher values of yellow coloration were better able to handle the cost of producing a second clutch. Finally, UV-blue female coloration was positively linked to female survival and marginally linked to laying date. Taken together, these results show for the first time in a natural population that female coloration can indicate individual and maternal quality under natural and adverse reproductive conditions. They highlight the potential for the evolution of female ornamental traits through sexual selection.     

Chronic food shortage and seasonal modulations of daily torpor and locomotor activity in the grey mouse lemur (Microcebus murinus)

The extent to which seasonal plasticity in torpor displayed by one of the smallest Malagasy primates (Microcebus murinus) will help survival in the context of ongoing global change-induced chronic food shortage, is unknown. Body temperature (Tb) and locomotor activity were measured by telemetry in short- (SD, winter-acclimated) and long-days (LD, summer-acclimated) males (n = 24) during an experimental 35-day calorie restriction of 40 or 80%. Under SD exposure, regardless of calorie restriction intensity, mouse lemurs immediately increased torpor depth and duration by 4.6-fold, and showed greater phase-advanced entry into torpor (2.4-fold). Tb adjustments were efficient under 40% calorie restriction to maintain body mass, whereas they did not prevent a 0.71 +/- 0.11 g/day mass loss during 80% calorie restriction. The 40% food-deprived LD animals combined an early shallow deepening of torpor (1 degrees C) and a late 18% decrease in locomotor activity, resulting in a moderate 6% mass loss. After 15 days of 80% calorie restriction, LD animals exhibited a SD phenotype by increasing their torpor duration and phase-advancing the entry of torpor (16 min/day). Those adjustments had no impact on mass loss (0.93 +/- 0.07 g/day) as locomotor activity increased four-fold. Daily torpor allows M. murinus to face moderate food shortage whatever the photoperiod but poorly mitigates energy imbalance during severe food deprivation, especially under LD exposure. Although the behavioral thermoregulation role warrants further investigation in energy savings, M. murinus survival would be impaired during long-term food shortage in summer.     

Dynamics of Genome Architecture in Rhizobium sp. Strain NGR234

Bacterial genomes are usually partitioned in several replicons, which are dynamic structures prone to mutation and genomic rearrangements, thus contributing to genome evolution. Nevertheless, much remains to be learned about the origins and dynamics of the formation of bacterial alternative genomic states and their possible biological consequences. To address these issues, we have studied the dynamics of the genome architecture in Rhizobium sp. strain NGR234 and analyzed its biological significance. NGR234 genome consists of three replicons: the symbiotic plasmid pNGR234a (536,165 bp), the megaplasmid pNGR234b (>2,000 kb), and the chromosome (>3,700 kb). Here we report that genome analyses of cell siblings showed the occurrence of large-scale DNA rearrangements consisting of cointegrations and excisions between the three replicons. As a result, four new genomic architectures have emerged. Three consisted of the cointegrates between two replicons: chromosome-pNGR234a, chromosome-pNGR234b, and pNGR234a-pNGR234b. The other consisted of a cointegrate of the three replicons (chromosome-pNGR234a-pNGR234b). Cointegration and excision of pNGR234a with either the chromosome or pNGR234b were studied and found to proceed via a Campbell-type mechanism, mediated by insertion sequence elements. We provide evidence showing that changes in the genome architecture did not alter the growth and symbiotic proficiency of Rhizobium derivatives.