Nature, evolution and characterisation of rhizospheric chemical exudates affecting root herbivores

Similar to aboveground herbivores, root-feeding insects must locate and identify suitable resources. In the darkness of soil, they mainly rely on root chemical exudations and, therefore, have evolved specific behaviours. Because of their impact on crop yield, most of our knowledge in belowground chemical ecology is biased towards soil-dwelling insect pests. Yet the increasing literature on volatile-mediated interactions in the ground underpins the great importance of chemical signalling in this ecosystem and its potential in pest control. Here, we explore the ecology and physiology of these chemically based interactions. An evolutionary approach reveals interesting patterns in the response of insects to particular classes of volatile or water-soluble organic compounds commonly emitted by roots. Food web analyses reasonably support that volatiles are used as long-range cues whereas water-soluble molecules serve in host acceptance/rejection by the insect; however, data are still scarce. As a case study, the chemical ecology of Diabrotica virgifera virgifera is discussed and applications of belowground signalling in pest management are examined. Soil chemical ecology is an expanding field of research and will certainly be a hub of our understanding of soil communities and subsequently of the management of belowground ecosystem services.

Sex-mediated herbivory by galling insects on Baccharis concinna (Asteraceae)

The interaction patterns between the dioecious shrub Baccharis concinna Barroso (Asteraceae) and its speciose galling insect community were studied in southeastern Brazil. Two hypotheses were tested in this study: "the differential reproduction and growth hypothesis" that predicts that male plants present fewer reproductive structures and are larger than female plants; and the 'sex-biased herbivory hypothesis' that predicts that male plants support a larger abundance of insect galls than female plants. Plants did not show sexual dimorphism in growth (= mean leaf number). However, male plants had longer shoots and a lower average number of inflorescences than female plants. These results corroborate the hypothesis that male plants grow more and reproduce less than female plants. No statistically significant difference was found in the number of galls between male and female plants, but a sex by environmental effect on gall number was detected. When each species of galling insect was individually analyzed per population of the host plant, the rates of attack varied between sex and population of the host plant, and they were highly variable among the species of galling insects. These results highlight the importance of the interaction between sex and environment in the community structure of galling insects and indicate that other variables besides host sex may influence the patterns of attack by galling herbivores.

RecA-mediated strand exchange traverses substitutional heterologies more easily than deletions or insertions.

RecA protein in bacteria and its eukaryotic homolog Rad51 protein are responsible for initiation of strand exchange between homologous DNA molecules. This process is crucial for homologous recombination, the repair of certain types of DNA damage and for the reinitiation of DNA replication on collapsed replication forks. We show here, using two different types of in vitro assays, that in the absence of ATP hydrolysis RecA-mediated strand exchange traverses small substitutional heterologies between the interacting DNAs, whereas small deletions or insertions block the ongoing strand exchange. We discuss evolutionary implications of RecA selectivity against insertions and deletions and propose a molecular mechanism by which RecA can exert this selectivity.

Activated group 3 innate lymphoid cells promote T-cell-mediated immune responses.

Group 3 innate lymphoid cells (ILC3s) have emerged as important cellular players in tissue repair and innate immunity. Whether these cells meaningfully regulate adaptive immune responses upon activation has yet to be explored. Here we show that upon IL-1β stimulation, peripheral ILC3s become activated, secrete cytokines, up-regulate surface MHC class II molecules, and express costimulatory molecules. ILC3s can take up latex beads, process protein antigen, and consequently prime CD4(+) T-cell responses in vitro. The cognate interaction of ILC3s and CD4(+) T cells leads to T-cell proliferation both in vitro and in vivo, whereas its disruption impairs specific T-cell and T-dependent B-cell responses in vivo. In addition, the ILC3-CD4(+) T-cell interaction is bidirectional and leads to the activation of ILC3s. Taken together, our data reveal a novel activation-dependent function of peripheral ILC3s in eliciting cognate CD4(+) T-cell immune responses.

Hypotensive effect of the active fragment derived from factor XII is mediated by an activation of the plasma kallikrein-kinin system.

Plasma protein fraction (PPF) contaminated by factor XII active fragment (XIIf) may cause hypotensive reactions when infused to patients. This study was planned to assess in conscious normotensive rats whether the blood pressure response to the factor XIIf is mediated by an activation of the plasma kallikrein-kinin system or by stimulation of prostaglandin synthesis. To test whether the factor XIIf-induced blood pressure fall is due partially to an enhanced generation of vasodilating prostaglandins, the blood pressure effect of XIIf (1 microgram i.v.) was investigated 15 min after treatment with indomethacin (5 mg i.v.), an inhibitor of cyclo-oxygenase. Factor XIIf reduced mean blood pressure similarly in indomethacin- and vehicle-treated rats (-23 +/- 4 mmHg, n = 5, and -23 +/- 5 mmHg, n = 4, respectively). Other rats received factor XIIf 15 min after depletion of circulating prekallikrein by the administration of dextran sulfate. Thirty minutes after a 0.25 mg i.v. dose of this agent, plasma prekallikrein activity averaged 0.12 +/- 0.015 mumol/min/ml (n = 6) as compared to 2.48 +/- 0.31 mumol/min/ml in control rats (n = 4, P less than .001). Factor XIIf decreased mean blood pressure by only 4 +/- 2 mm Hg in rats pretreated with dextran sulfate. Thus, it was possible to blunt the acute hypotensive effect of factor XIIf by depleting circulating prekallikrein, but not by inhibiting prostaglandin production. This strongly suggests that the blood pressure effects of factor XIIf is mediated by a stimulation of the plasma kallikrein-kinin system.

Dicarboxylic amino acids and glycine-betaine regulate chaperone-mediated protein-disaggregation under stress.

Active protein-disaggregation by a chaperone network composed of ClpB and DnaK + DnaJ + GrpE is essential for the recovery of stress-induced protein aggregates in vitro and in Escherichia coli cells. K-glutamate and glycine-betaine (betaine) naturally accumulate in salt-stressed cells. In addition to providing thermo-protection to native proteins, we found that these osmolytes can strongly and specifically activate ClpB, resulting in an increased efficiency of chaperone-mediated protein disaggregation. Moreover, factors that inhibited the chaperone network by impairing the stability of the ClpB oligomer, such as natural polyamines, dilution, or high salt, were efficiently counteracted by K-glutamate or betaine. The combined protective, counter-negative and net activatory effects of K-glutamate and betaine, allowed protein disaggregation and refolding under heat-shock temperatures that otherwise cause protein aggregation in vitro and in the cell. Mesophilic organisms may thus benefit from a thermotolerant osmolyte-activated chaperone mechanism that can actively rescue protein aggregates, correctly refold and maintain them in a native state under heat-shock conditions.

Uptake of particulate vaccine adjuvants by dendritic cells activates the NALP3 inflammasome.

Many currently used and candidate vaccine adjuvants are particulate in nature, but their mechanism of action is not well understood. Here, we show that particulate adjuvants, including biodegradable poly(lactide-co-glycolide) (PLG) and polystyrene microparticles, dramatically enhance secretion of interleukin-1beta (IL-1beta) by dendritic cells (DCs). The ability of particulates to promote IL-1beta secretion and caspase 1 activation required particle uptake by DCs and NALP3. Uptake of microparticles induced lysosomal damage, whereas particle-mediated enhancement of IL-1beta secretion required phagosomal acidification and the lysosomal cysteine protease cathepsin B, suggesting a role for lysosomal damage in inflammasome activation. Although the presence of a Toll-like receptor (TLR) agonist was required to induce IL-1beta production in vitro, injection of the adjuvants in the absence of TLR agonists induced IL-1beta production at the injection site, indicating that endogenous factors can synergize with particulates to promote inflammasome activation. The enhancement of antigen-specific antibody production by PLG microparticles was independent of NALP3. However, the ability of PLG microparticles to promote antigen-specific IL-6 production by T cells and the recruitment and activation of a population of CD11b(+)Gr1(-) cells required NALP3. Our data demonstrate that uptake of microparticulate adjuvants by DCs activates the NALP3 inflammasome, and this contributes to their enhancing effects on innate and antigen-specific cellular immunity.

Evaluation of a topical cyclosporine A prodrug on corneal graft rejection in rats.

PURPOSE: To assess the efficacy of a topical cyclosporine A (CsA), water-soluble prodrug, for promoting the survival of allogenic rat corneal grafts after penetrating keratoplasty (PKP). METHODS: Corneas of Brown-Norway rats (donors) were transplanted to Lewis rats (recipients). Transplanted rats were divided in three treatment groups: group I (PBS) and group II (0.26% Debio088) received drops five times per day. Group III received a daily intramuscular CsA injection (10 mg/kg/day). Blood CsA concentrations were measured on days 2 and 14. On day 4, 10, 13 after PKP, grafts were scored for corneal transparency, edema and extent of neovascularization. An opacity score of greater than or equal to 3 was considered as a nonreversible graft rejection process. On day 14, the experimental eyes were processed for histology. RESULTS: On day 13, 12 of the 18 corneal transplants (67%) in group I showed irreversible graft rejection. Three of 18 transplants (19%) in group II and 5 of 16 transplants (28%) in group III showed irreversible graft rejection (p=0.013/p=0.019, OR=0.14/0.06 versus vehicle). Each mean clinical score for edema, opacity, and neovessels in group II were significantly lower than those of the grafts in group I (respectively p=0.010, p=0.013, p=0.024) and III except for neovessels (respectively p=0.002, p=0.001, p=0.057). Histology confirmed the clinical results. The mean CsA blood levels for groups II and III were, respectively 54+/-141 mug/l and 755+/-319 mug/l on day 2 and 14+/-34 mug/l and 1318+/-463 mug/l on day 14. CONCLUSIONS: Debio088 CsA prodrug drops given five times daily are as effective as intramuscular injection of 10 mg/kg/day for the prevention of acute corneal graft rejection in rats.

Allergen-specific antibody and cytokine responses, mast cell reactivity and intestinal permeability upon oral challenge of sensitized and tolerized mice.

BACKGROUND: Food allergy has reached an epidemic level in westernized countries and although central mechanisms have been described, the variability associated with genetic diversity underscores the still unresolved complexity of these disorders. OBJECTIVE: To develop models of food allergy and oral tolerance, both strictly induced by the intestinal route, and to compare antigen-specific responses. METHODS: BALB/c mice were mucosally sensitized to ovalbumin (OVA) in the presence of the mucosal adjuvant cholera toxin, or tolerized by intra-gastric administrations of OVA alone. Antibody titres and cytokines were determined by ELISA, and allergic status was determined through several physiologic parameters including decline in temperature, diarrhoea, mast cell degranulation and intestinal permeability. RESULTS: OVA-specific antibodies (IgE, IgGs and IgA in serum and feces) were produced in sensitized mice exclusively. Upon intra-gastric challenge with OVA, sensitized mice developed anaphylactic reactions associated with a decline of temperature, diarrhoea, degranulation of mast cells, which were only moderately recruited in the small intestine, and increased intestinal permeability. Cytokines produced by immune cells from sensitized mice included T-helper type 2 cytokines (IL-5, IL-13), but also IL-10, IFN-gamma and IL-17. In contrast, all markers of allergy were totally absent in tolerized animals, and yet the latter were protected from subsequent sensitization, demonstrating that oral tolerance took place efficiently. CONCLUSION: This work allows for the first time an appropriate comparison between sensitized and tolerized BALB/c mice towards OVA. It highlights important differences from other models of allergy, and thus questions some of the generally accepted notions of allergic reactions, such as the protective role of IFN-gamma, the importance of antigen-specific secretory IgA and the role of mucosal mast cells in intestinal anaphylaxis. In addition, it suggests that IL-17 might be an effector cytokine in food allergy. Finally, it demonstrates that intestinal permeability towards the allergen is increased during challenge.

Biosphere/lithosphere interface : microbially-mediated CaCO3 precipitation in hypersaline and freshwater environments

Abstract: Microbial mats very efficiently cycle elements, such as C, 0, N, S and H, which makes them key players of redox processes at the biosphere-lithosphere interface. They are characterized by high metabolic activities and high turnover rates (production and consumption) of biomass, which mainly consists of cell material and of extracellular organic matter (EOM). The EOM forms a matrix, embedding the microbial cells and fulfilling various functions within the microbial mat, including: mat attachment to surfaces; creation of micro-domains within the mat; physical stabilization under hy- drodynamic stress and the protection of the cells in multiple other stress conditions. EOM mainly consists of polysaccharides, amino acids, and a variety of chemical func-tional groups {e.g., -C00H, - SH -OH). These groups strongly bind cations such as Ca2+ and Mg2+ and thus exert a strong control on carbonate mineral formation within the microbial mat. A feedback mechanism between community metabolisms, their prod¬ucts, and the surrounding physicochemical microenvironment thus influences the de¬gree of carbonate saturation favoring either carbonate precipitation or dissolution. We investigated the driving forces and mechanisms of microbialite formation in the Sari ne River, FR, Switzerland, the hypersaline lake, Big Pond, Bahamas and in labo¬ratory experiments. The two fundamentally different natural systems allowed us to compare the geochemical conditions and microbial metabolisms, necessary for car¬bonate formation in microbial mats. Although carbonates are oversaturated in both environments, precipitation does not occur on physicochemical substrates (i.e. out¬side the microbial mats). In the Sarine a high crystal nucleation threshold exceeds the carbonate saturation, despite the high carbonate alkalinity in the water column. Cyanobacterial photosynthesis strongly locally enhances the carbonate alkalinity, whereas the EOM attract and immobilize calcium, which increases the saturation state and finally leads to carbonate precipitation within the EOM (in this case the cyanobacterial sheath) as nucleation template. In Big Pond, the presence of calcium- chelating anions (i.e. sulfate) and EOM, as well as the presence of magnesium, lowers the calcium activity in the water column and mat, and thus inhibits carbonate pre¬cipitation. Coupled with other heterotrophic metabolisms, sulfate reduction uses the EOM as carbon source, degrading it. The resulting EOM consumption creates alkalin¬ity, releases calcium and consumes sulfate in mat-micro domains, which leads to the formation of carbonate layers at the top of the microbial mat. Résumé: Interface biosphère/lithosphère: médiation microbienne de la précipitation de CaC03 dans des environnements en eaux douces et hypersalines Les tapis microbiens engendrent une circulation très efficace des éléments, tels que C, 0, N, S et H, ce qui en fait des acteurs clé pour les processus d'oxydoréduction à l'inter¬face biosphère-lithosphère. Ils sont caractérisés par des taux élevés d'activité méta¬bolique, ainsi que par la production et la consommation de biomasse, principalement constituée de cellules microbiennes et de matière organique extracellulaire (MOE). Dans un tapis microbien, les cellules microbiennes sont enveloppées par une matrice de MOE qui a différentes fonctions dont l'attachement du tapis aux surfaces, la créa¬tion de micro-domaines dans le tapis, la stabilisation physique en situation de stress hydrodynamique, et la protection des cellules dans de multiples autres conditions de stress. La MOE se compose principalement de polysaccharides, d'acides aminés, et d'une variété de groupes fonctionnels chimiques (par exemple, COOH, -SH et -OH). Ces groupes se lient fortement aux cations, tels que Ca2+ et Mg2+, et exercent ainsi un contrôle fort sur la formation de CaC03 dans le tapis microbien. Un mécanisme de rétroaction, entre les métabolismes de la communauté microbienne, leurs produits, et le microenvironnement physico-chimique, influence le degré de saturation de car¬bonate, favorisant soit leur précipitation, soit leur dissolution. Nous avons étudié le moteur et les mécanismes de minéralisation dans des tapis de la Sarine, FR, Suisse et du lac hypersalin, Big Pond, aux Bahamas, ainsi que durant des expériences en laboratoire. Les deux systèmes naturels, fondamentalement dif¬férents, nous ont permis de comparer les conditions géochimiques et les métabolis¬mes nécessaires à la formation des carbonates dans des tapis microbiens. Bien que les carbonates soient sursaturés dans les deux environnements, la précipitation ne se produit pas sur des substrats physico-chimiques (en dehors du tapis microbien). Dans la Sarine, malgré un taux d'alcalinité élevé, les valeurs de seuil pour la nucléa- tion de carbonates sont plus hautes que la saturation du carbonate. La photosynthèse cyanobactérienne augmente localement l'alcalinité, alors que la MOE attire et immo¬bilise le calcium, ce qui augmente l'état de saturation et conduit finalement à la pré¬cipitation des carbonates, en utilisant la MOE comme substrat de nucléation. À Big Pond, la présence de chélateurs de calcium, notamment les anions (p.ex. le sulfate) et la MOE, ainsi que la présence de magnésium, réduit l'activité du calcium et inhibe en conséquence la précipitation des carbonates. Couplée avec d'autres métabolismes hétérotrophes, la réduction des sulfates utilise la MOE comme source de carbone, en la dégradant. Cette consommation de MOE crée l'alcalinité, consomme des sulfates et libère du calcium dans des micro-domaines, conduisant à la formation de couches de carbonates dans le haut du tapis microbien.

Intralesional adenovirus-mediated interleukin-2 gene transfer for advanced solid cancers and melanoma.

Numerous preclinical and clinical studies have shown that interleukin-2 (IL-2) induces regression of metastatic tumors. We have conducted a phase I/II, multicenter, open-label, dose-escalating study to evaluate the safety, efficacy, and biological effects of repeated intratumoral injections of adenovirus-IL-2 (TG1024) in patients with advanced solid tumors and melanoma. Thirty five patients (twenty-five with metastatic melanoma and ten with other solid tumors) were treated in eight successive cohorts at dose levels ranging from 3 x 10(8) to 3 x 10(11) viral particles (vp). Intratumoral TG1024 injections in combination with dacarbazine (DTIC) were tested in metastatic melanoma in one cohort. No clinical responses were observed at doses below 3 x 10(11) vp. Six local objective responses were recorded in patients receiving 3 x 10(11) vp per treatment [five in metastatic melanoma and one in metastatic squamous cell carcinoma (SCC) of the skin], of which two were complete responses (CRs). Most of the common side effects were injection site reactions and flu-like syndrome. TG1024 dose intensification across cohorts resulted in increased serum IL-2 levels after the injection. Intratumoral TG1024 injection induced pronounced inflammation of the treated lesion, with predominant CD8(+), TIA+ lymphocytic infiltrate. Our results show that intratumoral injections of TG1024 are safe and well tolerated. The clinical activity of TG1024 observed in this study warrants further investigations.

Transepithelial transport of HIV-1 by M cells is receptor-mediated.

Human colon carcinoma Caco-2 cell monolayers undergo conversion into cells that share morphological and functional features of M cells when allowed to interact with B lymphocytes. A lymphotropic (X4) HIV-1 strain crosses M cell monolayers and infects underlying CD4(+) target cells. Transport requires both lactosyl cerebroside and CXCR4 receptors, which are expressed on the apical surface of Caco-2 and M cells. Antibodies specific for each receptor block transport. In contrast, a monotropic (R5) HIV-1 strain is unable to cross M cell monolayers and infect underlying monocytes, despite efficient transport of latex beads. Caco-2 and M cells do not express CCR5, but transfection of these cells with CCR5 cDNA restores transport of R5 virus, which demonstrates that HIV-1 transport across M cells is receptor-mediated. The follicle-associated epithelium covering human gut lymphoid follicles expresses CCR5, but not CXCR4, and lactosyl cerebroside, suggesting that HIV-1 infection may occur through M cells and enterocytes at these sites.

LRF-mediated Dll4 repression in erythroblasts is necessary for hematopoietic stem cell maintenance.

Hematopoietic stem cells (HSCs) are the most primitive cells in the hematopoietic system and are under tight regulation for self-renewal and differentiation. Notch signals are essential for the emergence of definitive hematopoiesis in mouse embryos and are critical regulators of lymphoid lineage fate determination. However, it remains unclear how Notch regulates the balance between HSC self-renewal and differentiation in the adult bone marrow (BM). Here we report a novel mechanism that prevents HSCs from undergoing premature lymphoid differentiation in BM. Using a series of in vivo mouse models and functional HSC assays, we show that leukemia/lymphoma related factor (LRF) is necessary for HSC maintenance by functioning as an erythroid-specific repressor of Delta-like 4 (Dll4) expression. Lrf deletion in erythroblasts promoted up-regulation of Dll4 in erythroblasts, sensitizing HSCs to T-cell instructive signals in the BM. Our study reveals novel cross-talk between HSCs and erythroblasts, and sheds a new light on the regulatory mechanisms regulating the balance between HSC self-renewal and differentiation.

A link between FXYD3 (Mat-8)-mediated Na,K-ATPase regulation and differentiation of Caco-2 intestinal epithelial cells.

FXYD3 (Mat-8) proteins are regulators of Na,K-ATPase. In normal tissue, FXYD3 is mainly expressed in stomach and colon, but it is also overexpressed in cancer cells, suggesting a role in tumorogenesis. We show that FXYD3 silencing has no effect on cell proliferation but promotes cell apoptosis and prevents cell differentiation of human colon adenocarcinoma cells (Caco-2), which is reflected by a reduction in alkaline phosphatase and villin expression, a change in several other differentiation markers, and a decrease in transepithelial resistance. Inhibition of cell differentiation in FXYD3-deficient cells is accompanied by an increase in the apparent Na+ and K+ affinities of Na,K-ATPase, reflecting the absence of Na,K-pump regulation by FXYD3. In addition, we observe a decrease in the maximal Na,K-ATPase activity due to a decrease in its turnover number, which correlates with a change in Na,K-ATPase isozyme expression that is characteristic of cancer cells. Overall, our results suggest an important role of FXYD3 in cell differentiation of Caco-2 cells. One possibility is that FXYD3 silencing prevents proper regulation of Na,K-ATPase, which leads to perturbation of cellular Na+ and K+ homeostasis and changes in the expression of Na,K-ATPase isozymes, whose functional properties are incompatible with Caco-2 cell differentiation.