Activation state of alpha4beta1 integrin on sickle red blood cells is linked to the duffy antigen receptor for chemokines (DARC) expression

In sickle cell anemia, reticulocytes express enhanced levels of α4β1 integrin that interact mainly with vascular cell adhesion molecule-1 and fibronectin, promoting vaso-occlusion. These interactions are known to be highly sensitive to the inflammatory chemokine IL-8. The Duffy antigen receptor for chemokines (DARC) modulates the function of inflammatory processes. However, the link between α4β1 activation by chemokines and DARC erythroid expression is not or poorly explored. Therefore, the capacity of α4β1 to mediate Duffy-negative and Duffy-positive sickle reticulocyte (SRe) adhesion to immobilized vascular cell adhesion molecule-1 and fibronectin was evaluated. Using static adhesion assays, we found that, under basal conditions, Duffy-positive SRe adhesion was 2-fold higher than that of Duffy-negative SRes. Incubating the cells with IL-8 or RANTES (regulated on activation normal T cell expressed and secreted) increased Duffy-positive SRe adhesion only, whereas Mn(2+) increased cell adhesion independently of the Duffy phenotype. Flow cytometry analyses performed with anti-β1 and anti-α4 antibodies, including a conformation-sensitive one, in the presence or absence of IL-8, revealed that Duffy-positive and Duffy-negative SRes displayed similar erythroid α4β1 expression levels, but with distinct activation states. IL-8 did not affect α4β1 affinity in Duffy-positive SRes but induced its clustering as corroborated by immunofluorescence microscopy. Our results indicate that in Duffy-negative SRes α4β1 integrin is constitutively expressed in a low affinity state, whereas in Duffy-positive SRes α4β1 is expressed in a higher chemokine-sensitive affinity state. This activation state associated with DARC RBC expression may influence the intensity of the inflammatory responses encountered in sickle cell anemia and participate in its interindividual clinical expression variability.     

Spectrin-like repeats 11-15 of human dystrophin show adaptations to a lipidic environment

Dystrophin is essential to skeletal muscle function and confers resistance to the sarcolemma by interacting with cytoskeleton and membrane. In the present work, we characterized the behavior of dystrophin 11-15 (DYS R11-15), five spectrin-like repeats from the central domain of human dystrophin, with lipids. DYS R11-15 displays an amphiphilic character at the liquid/air interface while maintaining its secondary α-helical structure. The interaction of DYS R11-15 with small unilamellar vesicles (SUVs) depends on the lipid nature, which is not the case with large unilamellar vesicles (LUVs). In addition, switching from anionic SUVs to anionic LUVs suggests the lipid packing as a crucial factor for the interaction of protein and lipid. The monolayer model and the modulation of surface pressure aim to mimic the muscle at work (i.e. dynamic changes of muscle membrane during contraction and relaxation) (high and low surface pressure). Strikingly, the lateral pressure modifies the protein organization. Increasing the lateral pressure leads the proteins to be organized in a regular network. Nevertheless, a different protein conformation after its binding to monolayer is revealed by trypsin proteolysis. Label-free quantification by nano-LC/MS/MS allowed identification of the helices in repeats 12 and 13 involved in the interaction with anionic SUVs. These results, combined with our previous studies, indicate that DYS R11-15 constitutes the only part of dystrophin that interacts with anionic as well as zwitterionic lipids and adapts its interaction and organization depending on lipid packing and lipid nature. We provide strong experimental evidence for a physiological role of the central domain of dystrophin in sarcolemma scaffolding through modulation of lipid-protein interactions.     

Interacting regions of CD81 and two of its partners, EWI-2 and EWI-2wint, and their effect on hepatitis C virus infection

CD81 is a tetraspanin protein that is involved in several essential cellular functions, as well as in the hepatitis C virus (HCV) infection. CD81 interacts with a high stoichiometry with its partner proteins EWI-2, EWI-2wint, and EWI-F. These latter proteins modify the functions of CD81 and can thereby potentially inhibit infection or modulate cell migration. Here, we characterized the cleavage of EWI-2 leading to the production of EWI-2wint, which has been shown to inhibit HCV infection. We determined the regions of EWI-2/EWI-2wint and CD81 that are important for their interaction and their functionality. More precisely, we identified a glycine zipper motif in the transmembrane domain of EWI-2/EWI-2wint that is essential for the interaction with CD81. In addition, we found that palmitoylation on two juxtamembranous cysteines in the cytosolic tail of EWI-2/EWI-2wint is required for their interaction with CD81 as well as with CD9, another tetraspanin. Thus, we have shown that palmitoylation of a tetraspanin partner protein can influence the interaction with a tetraspanin. We therefore propose that palmitoylation not only of tetraspanins, but also of their partner proteins is important in regulating the composition of complexes in tetraspanin networks. Finally, we identified the regions in CD81 that are necessary for its functionality in HCV entry and we demonstrated that EWI-2wint needs to interact with CD81 to exert its inhibitory effect on HCV infection.     

M-ficolin interacts with the long pentraxin PTX3: a novel case of cross-talk between soluble pattern-recognition molecules

Ficolins and pentraxins are soluble oligomeric pattern-recognition molecules that sense danger signals from pathogens and altered self-cells and might act synergistically in innate immune defense and maintenance of immune tolerance. The interaction of M-ficolin with the long pentraxin pentraxin 3 (PTX3) has been characterized using surface plasmon resonance spectroscopy and electron microscopy. M-ficolin was shown to bind PTX3 with high affinity in the presence of calcium ions. The interaction was abolished in the presence of EDTA and inhibited by N-acetyl-D-glucosamine, indicating involvement of the fibrinogen-like domain of M-ficolin. Removal of sialic acid from the single N-linked carbohydrate of the C-terminal domain of PTX3 abolished the interaction. Likewise, an M-ficolin mutant with impaired sialic acid-binding ability did not interact with PTX3. Interaction was also impaired when using the isolated recognition domain of M-ficolin or the monomeric C-terminal domain of PTX3, indicating requirement for oligomerization of both proteins. Electron microscopy analysis of the M-ficolin-PTX3 complexes revealed that the M-ficolin tetramer bound up to four PTX3 molecules. From a functional point of view, immobilized PTX3 was able to trigger M-ficolin-dependent activation of the lectin complement pathway. These data indicate that interaction of M-ficolin with PTX3 arises from its ability to bind sialylated ligands and thus differs from the binding to the short pentraxin C-reactive protein and from the binding of L-ficolin to PTX3. The M-ficolin-PTX3 interaction described in this study represents a novel case of cross-talk between soluble pattern-recognition molecules, lending further credit to the integrated view of humoral innate immunity that emerged recently.     

Resistance level to an aphid potato pest varies between genotypes from the same Solanum accession

Plant resistance to aphids can be improved by introgressing resistant traits from wild Solanum species into the potato germplasm. Breeding parents are commonly selected from the different accessions of each species. Accessions originate from several seeds collected in a restricted area and are conserved as seeds in genebanks. Genetic heterogeneity may be expected between genotypes from the same accession and could influence resistance level. Working with the potato aphid, Macrosiphum euphorbiae (Thomas), and the accession PI243340 of Solanum chomatophilum (Bitter), which has been previously rated as resistant to M. euphorbiae, we genetically identified and assessed the resistance level of genotypes within the accession. A combination of two multilplex polymerase chain reactions (PCRs) discriminated the 13 plant genotypes assessed. Survival of M. euphorbiae, measured using clip cages, varied significantly between five genotypes, randomly selected among the 13 previously assessed, but did not differ between same-genotype plants. Survival among genotypes ranged from 0 to > 60% 12 d after adult molt, and the least resistant genotype exhibited survival close to the susceptible standard, Solanum tuberosum L. Our results support the use of PCR multiplex methods to assess genetic heterogeneity in wild Solanum, and suggest that within accession genetic heterogeneity is sufficient to influence resistance level to aphids. Fine screening at the genotype level is preferable when assessing resistance to aphids.     

A phloem-sap feeder mixes phloem and xylem sap to regulate osmotic potential

Phloem-sap feeders (Hemiptera) occasionally consume the dilute sap of xylem, a behaviour that has previously been associated with replenishing water balance following dehydration. However, a recent study reported that non-dehydrated aphids ingested xylem sap. Here, we tested the hypothesis that the consumption of xylem sap, which has a low osmolality, is a general response to osmotic stresses other than dehydration. Alate aphids were subjected to different treatments and subsequently transferred onto a plant, where electrical penetration graph (EPG) was used to estimate durations of passive phloem sap consumption and active sucking of xylem sap. The proportion of time aphids fed on xylem sap (i.e., time spent feeding on xylem sap/total time spent feeding on phloem plus xylem sap) was used as a proxy of the solute concentration of the uptake. The proportion of time alate aphids fed on xylem sap increased: (1) with the time spent imbibing an artificial diet containing a solution of sucrose, which is highly concentrated in phloem sap and is mainly responsible for the high osmotic potential of phloem sap; (2) with the osmotic potential of the artificial diet, when osmotic potential excess was not related to sucrose concentration; and (3) when aphids were deprived of primary symbionts, a condition previously shown to lead to a higher haemolymph osmotic potential. All our results converge to support the hypothesis that xylem sap consumption contributes to the regulation of the osmotic potential in phloem-sap feeders.     

Deciphering sulfoglycolipids of Mycobacterium tuberculosis

For 4 decades, in vivo and in vitro studies have suggested that sulfoglycolipids (SGLs) play a role in the virulence or pathogenesis of the tubercle bacilli. However, the SGL structure and biosynthesis pathway remain only partially elucidated. Using the modern tools of structural analysis, including MALDI-time-of-flight MS, MS/MS, and two-dimensional NMR, we reevaluated the structure of the different SGL acyl (di-, tri-, and tetra-acylated) forms of the reference strain Mycobacterium tuberculosis H37Rv, as well as those produced by the mmpL8 knockout strains previously described to intracellularly accumulate di-acylated SGL. We report here the identification of new acyl forms: di-acylated SGL esterified by simple fatty acids only, as well as mono-acylated SGL bearing a hydroxyphthioceranoic acid, which were characterized in the wild-type strain. In a clinical strain, a complete family of mono-acylated SGLs was characterized in high abundance for the first time. For the mmpL8 mutant, SGLs were found to be esterified i) by an oxophthioceranoic acid, never observed so far, and ii) at nonconventional positions in the case of the unexpected tri-acylated forms. Our results further confirm the requirement of MmpL8 for the complete assembly of the tetra-acylated forms of SGL and also provide, by the discovery of new intermediates, insights in terms of the possible SGL biosynthetic pathways.     

Effect of media-induced social distancing on disease transmission in a two patch setting

We formulate an SIS epidemic model on two patches. In each patch, media coverage about the cases present in the local population leads individuals to limit the number of contacts they have with others, inducing a reduction in the rate of transmission of the infection. A global qualitative analysis is carried out, showing that the typical threshold behavior holds, with solutions either tending to an equilibrium without disease, or the system being persistent and solutions converging to an endemic equilibrium. Numerical analysis is employed to gain insight in both the analytically tractable and intractable cases; these simulations indicate that media coverage can reduce the burden of the epidemic and shorten the duration of the disease outbreak.     

Oral vaccination against raccoon rabies: landscape heterogeneity and timing of distribution influence wildlife contact rates with the ONRAB vaccine bait

Aerial distribution of oral vaccine baits is one of the available strategies for controlling the spread of infectious wildlife diseases. This technique has commonly been used to control rabies in wild carnivores and, together with other techniques, was used to immunize wild populations of raccoons (Procyon lotor) and striped skunks (Mephitis mephitis) after the detection of the first rabid raccoon in the province of Quebec, Canada, in 2006. Vaccine bait distribution was conducted over large areas where agricultural land is dominant but interspersed with residual forest patches. Our objective was to evaluate the effect of habitat (forest vs. agricultural crops) in space and time on the contact rate between wildlife and the ONRAB(?) vaccine bait, a recent alternative to the V-RG(?). Four transects of eight vaccine baits each were installed parallel to, and at different distances from, the forest's edge (under forest cover, at field-forest edge, and at 50 and 200 m from forest edge in agricultural crops) at three sites composed of various crop types interspersed with forest patches. This experiment was conducted during three periods (late spring, 1-7 June; summer, 27 July-2 August; and fall, 24-30 October) in 2009. Contact rates with vaccine baits were monitored for 7 days in each period to evaluate the potential temporal variations generated within the habitat types. Contact rates with ONRAB vaccine baits were highest under forest cover and in the fall. Of 13 species observed in proximity to the vaccine baits, raccoons were the most frequent (49.5%, n=55 visits). Our study underlines the importance of taking into account landscape heterogeneity and timing of distribution when planning the distribution of vaccine baits to control rabies in raccoons.