Description of Euroleptochromus gen.n. (Coleoptera,Staphylinidae, Scydmaeninae) from Baltic amber,with discussion of biogeography and mouthpart evolution within Clidicini

A new extinct species of the ant‐like stone beetle supertribe Mastigitae, Euroleptochromus sabathi gen. & sp.n. is described from Eocene Baltic amber. A phylogenetic analysis of Clidicini, with representatives of Leptomastacini and Mastigini as out‐group taxa, provided strong support for a sister‐group relationship between the Neotropical Leptochromus and the new genus. The monophyly of Clidicini is questioned because of an alternative placement of Nearctic Papusus as a sister taxon to Leptomastacini + [Clidicus + (Palaeoleptochromus + (Euroleptochromus + Leptochromus))]. A dispersal‐vicariance analysis provided three alternative scenarios for the evolution of Mastigitae; with Laurasia as the ancestral area of the supertribe, major branching events occurring within either Eurasia or Laurentia and two trans‐Beringia dispersals in Late Cretaceous and Eocene. Euroleptochromus, Palaeoleptochromus and Leptochromus share highly derived structures on postgenae and maxillary palps, probably as part of a specialised feeding or prey capture mechanism. The formation of these modifications in Clidicini is demonstrated to involve a process (traced back to the Campanian, 79 Ma) of elongation and narrowing of maxillary palps and forming a cuticular setal projection from a broadened insertion site of sensory setae.     

De novo production of the flavonoid naringenin in engineered Saccharomyces cerevisiae

Background

Specific strains of Lactobacillus plantarum are marketed as health-promoting probiotics. The role and interplay of cell-wall compounds like wall- and lipo-teichoic acids (WTA and LTA) in bacterial physiology and probiotic-host interactions remain obscure. L. plantarum WCFS1 harbors the genetic potential to switch WTA backbone alditol, providing an opportunity to study the impact of WTA backbone modifications in an isogenic background.

Results

Through genome mining and mutagenesis we constructed derivatives that synthesize alternative WTA variants. The mutants were shown to completely lack WTA, or produce WTA and LTA that lack D-Ala substitution, or ribitol-backbone WTA instead of the wild-type glycerol-containing backbone. DNA micro-array experiments established that the tarIJKL gene cluster is required for the biosynthesis of this alternative WTA backbone, and suggest ribose and arabinose are precursors thereof. Increased tarIJKL expression was not observed in any of our previously performed DNA microarray experiments, nor in qRT-PCR analyses of L. plantarum grown on various carbon sources, leaving the natural conditions leading to WTA backbone alditol switching, if any, to be identified. Human embryonic kidney NF-κB reporter cells expressing Toll like receptor (TLR)-2/6 were exposed to purified WTAs and/or the TA mutants, indicating that WTA is not directly involved in TLR-2/6 signaling, but attenuates this signaling in a backbone independent manner, likely by affecting the release and exposure of immunomodulatory compounds such as LTA. Moreover, human dendritic cells did not secrete any cytokines when purified WTAs were applied, whereas they secreted drastically decreased levels of the pro-inflammatory cytokines IL-12p70 and TNF-α after stimulation with the WTA mutants as compared to the wild-type.

Conclusions

The study presented here correlates structural differences in WTA to their functional characteristics, thereby providing important information aiding to improve our understanding of molecular host-microbe interactions and probiotic functionality.

     

Cell cycle population effects in perturbation studies

Growth condition perturbation or gene function disruption are commonly used strategies to study cellular systems. Although it is widely appreciated that such experiments may involve indirect effects, these frequently remain uncharacterized. Here, analysis of functionally unrelated Saccharyomyces cerevisiae deletion strains reveals a common gene expression signature. One property shared by these strains is slower growth, with increased presence of the signature in more slowly growing strains. The slow growth signature is highly similar to the environmental stress response (ESR), an expression response common to diverse environmental perturbations. Both environmental and genetic perturbations result in growth rate changes. These are accompanied by a change in the distribution of cells over different cell cycle phases. Rather than representing a direct expression response in single cells, both the slow growth signature and ESR mainly reflect a redistribution of cells over different cell cycle phases, primarily characterized by an increase in the G1 population. The findings have implications for any study of perturbation that is accompanied by growth rate changes. Strategies to counter these effects are presented and discussed.     

The chemokine,CXCL16, and its receptor,CXCR6, are constitutively expressed in human annulus fibrosus and expression of CXCL16 is up-regulated by exposure to IL-1ß in vitro

Chemokines are an important group of soluble molecules with specialized functions in inflammation. The roles of many specialized chemokines and their receptors remain poorly understood in the human intervertebral disc. We investigated CXCL16 and its receptor, CXCR6, to determine their immunolocalization in disc tissue and their presence following exposure of cultured human annulus fibrosus cells to proinflammatory cytokines. CXCL16 is a marker for inflammation; it also can induce hypoxia-inducible factor 1α (HIF-1α), which is a phenotypic marker of heathy nucleus pulposus tissue. We found CXCL16 and CXCR6 immunostaining in many cells of the annulus portion of the disc. Molecular studies showed that annulus fibrosus cells exposed to IL-1ß, but not TNF-α, exhibited significant up-regulation of CXCL16 expression vs. control cells. There was no significant difference in the percentage of annulus cells that exhibited immunolocalization of CXCL16 in grade I/II, grade III or grade IV/V specimens. The presence of CXCL16 and its receptor, CXCR6, in the annulus in vivo suggests the need for future research concerning the role of this chemokine in proinflammatory functions, HIF-1α expression and disc vascularization.     

A comparison of anti-HER2 IgA and IgG1 in vivo efficacy is facilitated by high N-glycan sialylation of the IgA

Monomeric IgA has been proposed as an alternative antibody format for cancer therapy. Here, we present our studies on the production, purification and functional evaluation of anti-HER2 IgA antibodies as anti-cancer agents in comparison to the anti-HER2 IgG1 trastuzumab. MALDI-TOF MS analysis showed profound differences in glycosylation traits across the IgA isotypes and cell lines used for production, including sialylation and linkage thereof, fucosylation (both core and antennary) and the abundance of high-mannose type species. Increases in sialylation proved to positively correlate with in vivo plasma half-lives. The polymerization propensity of anti-HER2 IgA2m2 could be suppressed by an 18-aa deletion of the heavy chain tailpiece - coinciding with the loss of high-mannose type N-glycan species - as well as by 2 cysteine to serine mutations at positions 320 and 480. The HER2 F(ab')2-mediated anti-proliferative effect of the IgA2m1 and IgA2m2 subtypes was similar to IgG1, whereas the IgA1 isotype displayed considerably lower potency and efficacy. The Fc-mediated induction of antibody-dependent cell-mediated cytotoxicity (ADCC) using human whole blood ADCC assays did not demonstrate such clear differences between the IgA isotypes. However, the potency of the anti-HER2 IgA antibodies in these ADCC assays was found to be significantly lower than that of trastuzumab. In vivo anti-tumor activity of the anti-HER2 IgA antibodies was compared to that of trastuzumab in a BT-474 breast cancer xenograft model. Multiple dosing and sialylation of the IgA antibodies compensated for the short in vivo half-life of native IgA antibodies in mice compared to a single dose of IgG1. In the case of the IgA2m2 antibody, the resulting high plasma exposure levels were sufficient to cause clear tumor stasis comparable to that observed for trastuzumab at much lower plasma exposure levels.     

In vitro test systems supporting the development of improved pest control methods: a case study with chemical mixtures and bivalve biofoulers

Using the bivalve macrofouler Corbicula fluminea, the suitability of in vitro testing as a stepping stone towards the improvement of control methods based on chemical mixtures was addressed in this study. In vitro cholinesterase (ChE) activity inhibition following single exposure of C. fluminea tissue to four model chemicals (the organophosphates dimethoate and dichlorvos, copper and sodium dodecyl phosphate [SDS]) was first assessed. Consequently, mixtures of dimethoate with copper and dichlorvos with SDS were tested and modelled; mixtures with ChE revealed synergistic interactions for both chemical pairs. These synergic combinations were subsequently validated in vivo and the increased control potential of these selected combinations was verified, with gains of up to 50% in C. fluminea mortality relative to corresponding single chemical treatments. Such consistency supports the suitability of using time- and cost-effective surrogate testing platforms to assist the development of biofouling control strategies incorporating mixtures.     

The vocal expression of feeding motivation and frustration in the domestic laying hen, Gallus gallus domesticus

Thwarting of feeding behaviour in the laying hen results in an increase in stereotyped pacing, displacement preening, and the gakel-call. These behaviours therefore reflect the frustration arousal caused by the thwarting of feeding behaviour. This raises the question whether the level of frustration also varies with the intensity of the motivation to perform the thwarted behaviour. This study investigated the relationship between the intensity of the motivation and level of frustration on the one hand and the gakel-call on the other hand. In Experiment 1, the strength of the motivation to feed was varied by thwarting hens in their feeding behaviour in an operant procedure after different durations of food deprivation (0, 8, 23 and 47 h). Trend analysis showed that with increasing hunger state, an increasing number of gakel-calls was given. No effect of treatments on temporal characteristics of the gakel-call was found. In Experiment 2, the level of frustration was varied by reducing or increasing the duration of access to food for food-deprived hens compared to the duration of access during training. It was assumed that a shorter duration of access to food compared to training would elicit frustration, which in turn would affect the performance of behaviours indicative of thwarting. However, we found neither a relation between the number of gakel-calls nor the temporal features of the gakel-call and the duration of access to food. Possibly, the differences between treatments were not large enough to induce differences in frustration level. Also, other factors that might have influenced the motivation are discussed.     

Anatomical and chemical characteristics of the seed coat of <i>Medicago sativa</i> L. (alfalfa) cv. Baralfa 85 seeds and their association with seed dormancy

Seeds of alfalfa (Medicago sativa L.) can exhibit seedcoat imposed dormancy, which produces hard seeds within a seed lot. These seeds do not germinate because they do not imbibe water due to a barrier to water entry in the seed coat. The aim of this work was to analyze the anatomical and chemical characteristics of the testa of alfalfa seeds with respect to water permeability levels. The anatomy of seeds of the cv. Baralfa 85 was studied and structural substances, polyphenols, tannins and cutin present in the testa of seeds of different water permeability levels were determined. The anatomical characteristics of the seed coat and the proportions of components were found to determine the permeability level of the seed coat, an aspect that is associated with the physical seed dormancy level. Anatomically, increased thickness of the testa was associated with a lower permeability level. The difference may be attributed to the variation in cuticle thickness, length of macrosclereids and thickness of the cell wall, and presence and development of osteosclereids. From the physiological and chemical points of view, the mechanism of physical dormancy of the testa is explained by a greater amount of components that repel water and cement the cell wall, such as polyphenols, lignins, condensed tannins, pectic substances, and a lower proportion of cellulose and hemicellulose.