Fatty acid metabolism in human breast cancer cells (MCF7) transfected with heart-type fatty acid binding protein

The human breast cancer cell line MCF7 does not express heart-type fatty acid binding protein (H-FABP), a marker protein for differentiated mammary gland. MCF7 cells transfected with the bovine H-FABP cDNA expressed the corresponding protein and were characterized by growth inhibition and lower tumorgenicity in nude mice [22]. By enzyme linked immunoassay we now determined the amount of bovine H-FABP in these cells as 638 ± 80 ng/mg protein and used the transfected cells to study the role of H-FABP in fatty acid metabolism. Compared to control cells the uptake of radioactively labelled palmitic acid and oleic acid into MCF7 cells after 30 or 60 min was increased by 67% in H-FABP expressing transfectants, demonstrating a stimulatory role for this FABP-type in fatty acid metabolism. However, preferential targeting of [¹⁴C]oleic acid into neutral or phospholipid classes was not observed by the criterion of high performance thin layer chromatography followed by autoradiography. A reason for the modest increase of fatty acid uptake in H-FABP transfected MCF7 cells may be the basal expression of epidermal-type FABP, which was detected for the first time in these cells. It appears that the small amount of E-FABP expressed in MCF7 cells fulfils the need of the cells for a cytosolic fatty acid carrier under culture conditions and that even high concentrations of another FABP do only slightly increase the uptake due to limitations of fatty acid transport through the plasma membrane or of metabolism.     

Novel non-peptide lead structures forBradykinin B2-receptor antagonists

Summary A series of new non-peptide Bradykinin (BK) B₂-receptor antagonists is reported. These new leads belong to a larger set including both commercially or otherwise available potential candidates found by proprietary database searches using 3D-pharmacophore models as query, and several bis-benzamidino compounds selected from our tryptase-like protease inhibitor library on the basis of topological considerations, derived from the same models. Some of these compounds show functional competitive antagonistic activity, inhibiting in vitro the BK-induced contraction of isolated guinea-pig ileum (GPI) and rat uterus with a pA₂ up to 5.3 and 7.0, respectively. They display also binding affinity (IC₅₀ up to 0.56 μM) to the BK B₂-receptor in radioligand binding assays on GPI membrane preparations and on human IMR-90 fetal lung fibroblast cells expressing this receptor subtype. Furthermore, the results with the commercially available compounds, in some cases developed as therapeutics, show that the used 3D-pharmacophore model allows to predict to some certainty possible side actions of potential drugs.     

Specificity of chemical cues used by a specialist egg parasitoid during host location

Plant synomones and host kairomones are known to guide the egg parasitoid Oomyzus gallerucae to its specific host, the elm leaf beetle Xanthogaleruca luteola (= Pyrrhalta) (Muller) feeding upon elm leaves (Ulmus spp.). In this study, we investigated whether the activities of these plant synomones and kairomones are specific for the plant and herbivore species, respectively. Olfactometer and contact bioassays were used. In habitat location, O. gallerucae (Fonscolombe) is known to use synomones from Ulmus minor (Miller) that are induced by egg depositions of X. luteola. The attractiveness of such induced volatiles was shown to be specific both for the Ulmus species and the herbivore species depositing eggs. Neither leaves of U. glabra Hudson (= U. montana) carrying eggs of X. luteola nor leaves of U. minor (= U. campestris = U. procera) carrying eggs of the chrysomelid species Galeruca tanaceti L. emitted attractive synomones. O. gallerucae is also known to be attracted by volatile kairomones from faeces of X. luteola feeding on U. minor and to show prolonged antennal drumming when contacting substrates contaminated with these faeces. The kairomonal activity of the faeces was proved to be independent of the Ulmus species, since also faeces from elm leaf beetles feeding upon U. glabra emitted attractive volatiles. However, the faecal kairomones were specific for the herbivorous species, since faeces from a lepidopteran larva (Opisthograptis luteolata L.) feeding upon elm hardly elicited any antennal drumming in O. gallerucae. The egg parasitoid studied is known to recognize host eggs of X. luteola by contact kairomones extractable from the egg shell. O. gallerucae clearly differentiated between host eggs and eggs of another closely related chrysomelid species, Galerucella lineola L., as was shown by comparing duration of antennal drumming on host eggs and eggs of G. lineola.     

Rapid prototyping of arbitrary 2D and 3D wireframe DNA origami

Wireframe DNA origami assemblies can now be programmed automatically from the top-down using simple wireframe target geometries, or meshes, in 2D and 3D, using either rigid, six-helix bundle (6HB) or more compliant, two-helix bundle (DX) edges. While these assemblies have numerous applications in nanoscale materials fabrication due to their nanoscale spatial addressability and high degree of customization, no easy-to-use graphical user interface software yet exists to deploy these algorithmic approaches within a single, standalone interface. Further, top-down sequence design of 3D DX-based objects previously enabled by DAEDALUS was limited to discrete edge lengths and uniform vertex angles, limiting the scope of objects that can be designed. Here, we introduce the open-source software package ATHENA with a graphical user interface that automatically renders single-stranded DNA scaffold routing and staple strand sequences for any target wireframe DNA origami using DX or 6HB edges, including irregular, asymmetric DX-based polyhedra with variable edge lengths and vertices demonstrated experimentally, which significantly expands the set of possible 3D DNA-based assemblies that can be designed. ATHENA also enables external editing of sequences using caDNAno, demonstrated using asymmetric nanoscale positioning of gold nanoparticles, as well as providing atomic-level models for molecular dynamics, coarse-grained dynamics with oxDNA, and other computational chemistry simulation approaches.     

Alarming Proportions of Methicillin-Resistant Staphylococcus aureus (MRSA) in Wound Samples from Companion Animals,Germany 2010–2012

Staphylococcus (S.) aureus is an important cause of wound infections in companion animals, and infections with methicillin-resistant S. aureus (MRSA) are of particular concern due to limited treatment options and their zoonotic potential. However, comparable epidemiological data on MRSA infections in dogs, cats and horses is scarce, also limiting the knowledge about possible links to MRSA isolates from human populations. To gain more knowledge about the occurrence and genotypic variation of MRSA among wound swabs of companion animal origin in Germany we performed a survey (2010–2012) including 5,229 samples from 1,170 veterinary practices. S. aureus was identified in 201 (5.8%) canine, 140 (12.2%) feline and 138 (22.8%) equine swabs from a total of 3,479 canine, 1,146 feline and 604 equine wounds, respectively. High MRSA rates were identified with 62.7%, 46.4% and 41.3% in S. aureus of canine, feline and equine origin, respectively. Further genotyping including spa typing and multilocus sequence typing (MLST) revealed a comparable distribution of spa types among canine and feline MRSA with CC22 (47.6%; 49.2%) and CC5 (30.2%; 29.2%) as predominant lineages followed by CC398 (13.5%; 7.7%) and CC8 (4.0%; 9.2%). In contrast, the majority of equine MRSA belonged to CC398 (87.7%). Our data highlight the importance of S. aureus and MRSA as a cause of wound infections, particularly in cats and horses in Germany. While “human-associated” MRSA lineages were most common in dogs and cats, a remarkable number of CC398-MRSA was detected in horses, indicating a replacement of CC8-MRSA as the predominant lineage within horses in Germany. These data enforce further longitudinal epidemiological approaches to examine the diversity and temporal relatedness of MRSA populations in humans and animals to assess probable sources of MRSA infections. This would enable a sound risk assessment and establishment of intervention strategies to limit the additional spread of MRSA.     

Ancient death-grip leaf scars reveal ant–fungal parasitism

Parasites commonly manipulate host behaviour, and among the most dramatic examples are diverse fungi that cause insects to die attached to leaves. This death-grip behaviour functions to place insects in an ideal location for spore dispersal from a dead body following host death. Fossil leaves record many aspects of insect behaviour (feeding, galls, leaf mining) but to date there are no known examples of behavioural manipulation. Here, we document, to our knowledge, the first example of the stereotypical death grip from 48 Ma leaves of Messel, Germany, indicating the antiquity of this behaviour. As well as probably being the first example of behavioural manipulation in the fossil record, these data support a biogeographical parallelism between mid Eocene northern Europe and recent southeast Asia.     

Assessing population and environmental effects on thermal resistance in Drosophila melanogaster using ecologically relevant assays

To make laboratory studies of thermal resistance in ectotherms more ecologically relevant, temperature changes that reflect conditions experienced by individuals in nature should be used. Here we describe an assay that is useful for quantifying multiple measures of thermal resistance of individual adult flies. We use this approach to assess upper and lower thermal limits and functional thermal scope for Drosophila melanogaster and also show that the method can be used to (1) detect a previously described latitudinal cline for cold tolerance in D. melanogaster populations collected along the east coast of Australia, (2) demonstrate that acclimation at variable temperatures during development increases tolerance to both low and high thermal stresses and therefore increases thermal scope compared to acclimation at a constant temperature, (3) show that temperate populations adapted to variable thermal environments have wider thermal limits compared to those from the less variable tropics, at least when flies were reared under constant temperature conditions and (4) demonstrate that different measures of cold resistance are often not strongly correlated. Based on our findings, we suggest that the method could be routinely used in evaluating thermal responses potentially linked to ecological processes and evolutionary adaptation.     

Lysozyme-lysozyme self-interactions as assessed by the osmotic second virial coefficient: Impact for physical protein stabilization

The purpose of the presented study is to understand the physicochemical properties of proteins in aqueous solutions in order to identify solution conditions with reduced attractive protein-protein interactions, to avoid the formation of protein aggregates and to increase protein solubility. This is assessed by measuring the osmotic second virial coefficient (B²²), a parameter of solution non-ideality, which is obtained using self-interaction chromatography. The model protein is lysozyme. The influence of various solution conditions on B²² was investigated: protonation degree, ionic strength, pharmaceutical relevant excipients and combinations thereof. Under acidic solution conditions B²² is positive, favoring protein repulsion. A similar trend is observed for the variation of the NaCl concentration, showing that with increasing the ionic strength protein attraction is more likely. B²² decreases and becomes negative. Thus, solution conditions are obtained favoring attractive protein-protein interactions. The B²² parameter also reflects, in general, the influence of the salts of the Hofmeister series with regard to their salting-in/salting-out effect. It is also shown that B²² correlates with protein solubility as well as physical protein stability.