Incrimination of Anopheles (Anopheles) intermedius Peryassú, An. (Nyssorhynchus) nuneztovari Gabaldón, An. (Nys.) oswaldoi Peryassú as natural vectors of Plasmodium falciparum in French Guiana

Anopheles darlingi Root is the major vector of human malaria in the Neotropics and has been considered to be the sole malaria vector in French Guiana. The presence of other potential vectors suggests that malaria may be transmitted by other species under certain conditions. From 2006-2011, all anopheline specimens collected from 11 localities were assayed to determine if the Plasmodium circumsporozoite protein was present. In addition to An. darlingi, we found Anopheles oswaldoi, Anopheles intermedius and Anopheles nuneztovari specimens that were infected with Plasmodium sp. Further investigations on the behaviour and ecology of An. oswaldoi, An. intermedius and An. nuneztovari are necessary to determine their role in malaria transmission in French Guiana.     

A Predictive Computational Model of the Dynamic 3D Interphase Yeast Nucleus

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Highlights? We describe a predictive computational model of dynamic chromosomes in the yeast nucleus ? The model quantitatively recapitulates experimental data on nuclear organization ? The model predicts nuclear reorganization in response to treatment by rapamycin ? Large-scale nuclear organization is dominated by unspecific effects of crowded polymers     

TM9/Phg1 and SadA proteins control surface expression and stability of SibA adhesion molecules in Dictyostelium

TM9 proteins form a family of conserved proteins with nine transmembrane domains essential for cellular adhesion in many biological systems, but their exact role in this process remains unknown. In this study, we found that genetic inactivation of the TM9 protein Phg1A dramatically decreases the surface levels of the SibA adhesion molecule in Dictyostelium amoebae. This is due to a decrease in sibA mRNA levels, in SibA protein stability, and in SibA targeting to the cell surface. A similar phenotype was observed in cells devoid of SadA, a protein that does not belong to the TM9 family but also exhibits nine transmembrane domains and is essential for cellular adhesion. A contact site A (csA)-SibA chimeric protein comprising only the transmembrane and cytosolic domains of SibA and the extracellular domain of the Dictyostelium surface protein csA also showed reduced stability and relocalization to endocytic compartments in phg1A knockout cells. These results indicate that TM9 proteins participate in cell adhesion by controlling the levels of adhesion proteins present at the cell surface.     

Modulating Uranium Binding Affinity in Engineered Calmodulin EF-Hand Peptides: Effect of Phosphorylation

To improve our understanding of uranium toxicity, the determinants of uranyl affinity in proteins must be better characterized. In this work, we analyzed the contribution of a phosphoryl group on uranium binding affinity in a protein binding site, using the site 1 EF-hand motif of calmodulin. The recombinant domain 1 of calmodulin from A. thaliana was engineered to impair metal binding at site 2 and was used as a structured template. Threonine at position 9 of the loop was phosphorylated in vitro, using the recombinant catalytic subunit of protein kinase CK2. Hence, the T₉TKE₁₂ sequence was substituted by the CK2 recognition sequence TAAE. A tyrosine was introduced at position 7, so that uranyl and calcium binding affinities could be determined by following tyrosine fluorescence. Phosphorylation was characterized by ESI-MS spectrometry, and the phosphorylated peptide was purified to homogeneity using ion-exchange chromatography. The binding constants for uranyl were determined by competition experiments with iminodiacetate. At pH 6, phosphorylation increased the affinity for uranyl by a factor of ∼5, from K_d = 25±6 nM to K_d = 5±1 nM. The phosphorylated peptide exhibited a much larger affinity at pH 7, with a dissociation constant in the subnanomolar range (K_d = 0.25±0.06 nM). FTIR analyses showed that the phosphothreonine side chain is partly protonated at pH 6, while it is fully deprotonated at pH 7. Moreover, formation of the uranyl-peptide complex at pH 7 resulted in significant frequency shifts of the ν_as(P-O) and ν_s(P-O) IR modes of phosphothreonine, supporting its direct interaction with uranyl. Accordingly, a bathochromic shift in ν_as(UO₂)²⁺ vibration (from 923 cm⁻¹ to 908 cm⁻¹) was observed upon uranyl coordination to the phosphorylated peptide. Together, our data demonstrate that the phosphoryl group plays a determining role in uranyl binding affinity to proteins at physiological pH.     

Normalization of a chromosomal contact map

ABSTRACT: BACKGROUND: Chromatin organization has been increasingly studied in relation with its important influence on DNA-related metabolic processes such as replication or regulation of gene expression. Since its original design ten years ago, capture of chromosome conformation (3C) has become an essential tool to investigate the overall conformation of chromosomes. It relies on the capture of long-range trans and cis interactions of chromosomal segments whose relative proportions in the final bank reflect their frequencies of interactions, hence their average spatial proximity. The recent coupling of 3C with deep sequencing approaches now allows the generation of high resolution genome-wide chromosomal contact maps. Different protocols have been used to generate such maps in various organisms. This includes mammals, drosophila and yeast. The massive amount of raw data generated by the genomic 3C has to be carefully processed to alleviate the various biases and byproducts generated by the experiments. Our study aims at proposing a simple normalization procedure to take into account these unwanted but inevitable events. RESULTS: Careful analysis of the raw data generated previously for budding yeast Saccharomyces cerevisiae led to the identification of three main biases affecting the final datasets, including an original bias resulting from the circularization of DNA molecules exhibiting specific lengths in accordance with laws from polymer physics. We then developed a simple normalization procedure to process the data and allow the generation of a normalized, highly contrasted, chromosomal contact map for S. cerevisiae. The same method was then extended to the first human genome contact map. Using the normalized data, we revisited the preferential interactions originally described between subsets of discrete chromosomal features. Notably, the detection of preferential interactions between tRNA in yeast and CTCF, PolII binding sites in human can vary with the normalization procedure used. CONCLUSIONS: We quantitatively reanalyzed the genomic 3C data obtained for S. cerevisiae, identified some of the biases inherent to the technique and proposed a simple normalization procedure to analyze them. Such an approach can be easily generalized for genomic 3C experiments in other organisms. More experiments and analysis will be necessary to reach optimal resolution and accuracies of the maps generated through these approaches. Working with cell population presenting highest levels of homogeneity will prove useful in this regards.     

Ranging behaviour of roe deer in an experimental high-density population: Are females territorial?

We studied the ranging behaviour and spatial relationships between seven roe deer during more than 4 years in a partly wooded 14.2-ha enclosure. The animals (three young males, four adult females) were monitored with GPS telemetry collars. As expected, the surface area and overlap of the males’ bimonthly ranges decreased, and the distance between their arithmetic centres increased, as they became adult and, for two of them, territorial. Unexpectedly, females also tended to space out, the surface area and overlap of their bimonthly ranges being minimal in May to June, i.e. during the birth period. The distance between their arithmetic centres reached its maximum at the same time. Overlap between females’ ranges was consistently lower than those between males and females’ ranges, or between 1-year old males’ ranges. Our results raise the questions of female seasonal territoriality and of independence of the spacing systems of the two sexes in roe deer.     

Functional consequences of neuromyelitis optica-IgG astrocyte interactions on blood-brain barrier permeability and granulocyte recruitment

Autoantibody neuromyelitis optica-IgG (NMO-IgG) recognizing aquaporin-4 (AQP4) is implicated as playing a central role in the physiopathology of NMO. The aim of this in vitro-based study was to characterize functional consequences of interaction between NMO-IgG and cells of the neurovascular unit (astrocytes and brain endothelium) that would provide insight into recognized features of NMO, namely altered blood-brain barrier (BBB) permeability and granulocyte recruitment. We used sera from NMO and longitudinally extensive transverse myelitis cases shown to bind in a characteristic perivascular pattern to primate cerebellar slices. Using flow cytometry, we found that sera from NMO-IgG-positive patients reacted with CNS-derived human fetal astrocytes, whereas sera from multiple sclerosis patients did not. We demonstrated that NMO-IgG binding to astrocytes alters aquaporin-4 polarized expression and increases permeability of a human BBB endothelium/astrocyte barrier. We further demonstrated that NMO-IgG binding to human fetal astrocytes can result in NK cell degranulation, astrocyte killing by Ab-dependent cellular cytotoxicity and complement-dependent granulocyte attraction through the BBB model. Our study highlights important functional roles for NMO-IgG that could account for pathological lesions and BBB dysfunction observed in NMO.