Correction to: Linear relation between leaf xylem water potential and transpiration in pearl millet during soil drying

Plant and Soil - In the original version of this article, equations 4 and 9 unfortunately contained errors     

Robust RNAi enhancement via human Argonaute-2 overexpression from plasmids,viral vectors and cell lines

As the only mammalian Argonaute protein capable of directly cleaving mRNAs in a small RNA-guided manner, Argonaute-2 (Ago2) is a keyplayer in RNA interference (RNAi) silencing via small interfering (si) or short hairpin (sh) RNAs. It is also a rate-limiting factor whose saturation by si/shRNAs limits RNAi efficiency and causes numerous adverse side effects. Here, we report a set of versatile tools and widely applicable strategies for transient or stable Ago2 co-expression, which overcome these concerns. Specifically, we engineered plasmids and viral vectors to co-encode a codon-optimized human Ago2 cDNA along with custom shRNAs. Furthermore, we stably integrated this Ago2 cDNA into a panel of standard human cell lines via plasmid transfection or lentiviral transduction. Using various endo- or exogenous targets, we demonstrate the potential of all three strategies to boost mRNA silencing efficiencies in cell culture by up to 10-fold, and to facilitate combinatorial knockdowns. Importantly, these robust improvements were reflected by augmented RNAi phenotypes and accompanied by reduced off-targeting effects. We moreover show that Ago2/shRNA-co-encoding vectors can enhance and prolong transgene silencing in livers of adult mice, while concurrently alleviating hepatotoxicity. Our customizable reagents and avenues should broadly improve future in vitro and in vivo RNAi experiments in mammalian systems.     

Middle Pleistocene ecology and Neanderthal subsistence: Insights from stable isotope analyses in Payre (Ardèche,southeastern France)

The Middle Palaeolithic site of Payre in southeastern France yields abundant archaeological material associated with fossil hominid remains. With its long sequence of Middle Pleistocene deposits, Payre is a key site to study the Middle Palaeolithic chronology of this region. This study is the first to investigate carbon and oxygen isotope contents of Neanderthal tooth enamel bioapatite, together with a wide range of herbivorous and carnivorous species. The aim is to contribute to the understanding of hunting behaviour, resource partitioning, diet and habitat use of animals and Neanderthals through a palaeoecological reconstruction.     

Formaldehyde metabolism and formaldehyde‐induced stimulation of lactate production and glutathione export in cultured neurons

Formaldehyde is endogenously produced in the human body and brain levels of this compound are elevated in neurodegenerative conditions. Although the toxic potential of an excess of formaldehyde has been studied, little is known on the molecular mechanisms underlying its neurotoxicity as well as on the ability of neurons to metabolize formaldehyde. To address these topics, we have used cerebellar granule neuron cultures as model system. These cultures express mRNAs of various enzymes that are involved in formaldehyde metabolism and were remarkably resistant toward acute formaldehyde toxicity. Cerebellar granule neurons metabolized formaldehyde with a rate of around 200 nmol/(h × mg) which was accompanied by significant increases in the cellular and extracellular concentrations of formate. In addition, formaldehyde application significantly increased glucose consumption, almost doubled the rate of lactate release from viable neurons and strongly accelerated the export of the antioxidant glutathione. The latter process was completely prevented by inhibition of the known glutathione exporter multidrug resistance protein 1. These data indicate that cerebellar granule neurons are capable of metabolizing formaldehyde and that the neuronal glycolysis and glutathione export are severely affected by the presence of formaldehyde.     

DNase I and Proteinase K eliminate DNA from injured or dead bacteria but not from living bacteria in microbial reference systems and natural drinking water biofilms for subsequent molecular biology analyses

Molecular techniques, such as polymerase chain reaction (PCR) and quantitative PCR (qPCR), are very sensitive, but may detect total DNA present in a sample, including extracellular DNA (eDNA) and DNA coming from live and dead cells. DNase I is an endonuclease that non-specifically cleaves single- and double-stranded DNA. This enzyme was tested in this study to analyze its capacity of digesting DNA coming from dead cells with damaged cell membranes, leaving DNA from living cells with intact cell membranes available for DNA-based methods. For this purpose, an optimized DNase I/Proteinase K (DNase/PK) protocol was developed. Intact Staphylococcus aureus cells, heat-killed Pseudomonas aeruginosa cells, free genomic DNA of Salmonella enterica, and a mixture of these targets were treated according to the developed DNase/PK protocol. In parallel, these samples were treated with propidium monoazide (PMA) as an already described assay for live-dead discrimination. Quantitative PCR and PCR-DGGE of the eubacterial 16S rDNA fragment were used to test the ability of the DNase/PK and PMA treatments to distinguish DNA coming from cells with intact cell membranes in the presence of DNA from dead cells and free genomic DNA. The methods were applied to three months old autochthonous drinking water biofilms from a pilot facility built at a German waterworks. Shifts in the DNA patterns observed after DGGE analysis demonstrated the applicability of DNase/PK as well as of the PMA treatment for natural biofilm investigation. However, the DNase/PK treatment demonstrated some practical advantages in comparison with the PMA treatment for live/dead discrimination of bacterial targets in drinking water systems.     

Generalization of Mimics Imperfect in Colour Patterns: The Point of View of Wild Avian Predators

Current research of imperfect mimicry brings ambiguous results. Experiments simulating more natural conditions rather than laboratory experiments show lower willingness of avian predators to attack less perfect mimics. We decided to simulate a natural situation by testing responses of wild‐caught adult avian predators (Great tit – Parus major) to variously perfect mimics of the red firebug (Pyrrhocoris apterus), which were in previous studies shown to elicit avoidance in Great tits. Presented mimics were perfect in all traits (firebug with its own colour pattern), imperfect in colour pattern (firebug with modified colour pattern), perfect in colour pattern, but imperfect in other visual traits (cockroach with firebug colour pattern), and imperfect in colour pattern as well as in other visual traits (cockroach with modified colour patterns). Modification of the pattern focused on the rounded spots on firebug's hemielytra, which is a conspicuous trait within the pattern. The pattern modification had no influence on the number of birds attacking the prey; nevertheless, birds spent more time observing the cockroaches that displayed the perfect firebug colour pattern than in the case of any other prey. Moreover, firebugs that displayed the perfect firebug colour pattern were observed for the shortest time (equal to that of the model – unmodified firebug). Cockroaches were attacked more often than firebugs, which suggest that birds were able to use additional visual cues (shape of legs and antennae) in prey recognition. Given these result, we conclude that differences in morphological traits characteristic for used prey taxa (true bugs, cockroaches) seem to be more important in the prey's protection than its colour pattern.     

Phosphorylation of C-terminal polycystin-2 influences the interaction with PIGEA14: A QCM study based on solid supported membranes

Polycystin-2 (PC2) trafficking has been proposed to be a result of the interaction of PIGEA14 with PC2 as a function of the phosphorylation state of PC2. Here, we investigated the interaction of PIGEA14 with the C-terminal part of polycystin-2 wild type (cPC2wt) and the pseudophosphorylated mutant (cPC2S812D) to first, quantify the binding affinity between cPC2 and PIGEA14 and second, to elucidate the influence of PC2 phosphorylation on PIGEA14 binding. Solid supported membranes composed of octanethiol/1,2-dioleoyl-sn-glycero-3-phosphocholine doped with the receptor lipid DOGS–NTA–Ni were used to attach PIGEA14 to the membrane via its hexahistidine tag. By means of the quartz crystal microbalance technique, binding affinities as well as kinetic constants of the interaction were extracted in a label-free manner by applying the scaled particle theory. The results show that the dissociation constant of cPC2 to PIGEA14 is in the 10 nM regime providing strong evidence of a very specific interaction of cPC2 with PIGEA14. The interaction of cPC2wt is twofold larger than that of cPC2S812D. The moderate higher binding affinity of cPC2wt to PIGEA14 is discussed in light of PC2 trafficking to the plasma membrane.     

Thermal and acido-basic stability of antioxidant properties of extracts from cereal and pseudocereal grains

Beneficial effects of whole grains of cereals and pseudocereals and their fractions to human physiology are well known and broadly published. Especially secondary metabolites, dominantly from the category of phenolics (or polyphenols), beneficially influence the health physiology and/or prevent disease progress. Within the frame of this study, ten genotypes of four cereals or pseudocereals, respectively, were chosen for their antioxidant activity, determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP) and β-carotene-linoleic acid bleaching model (BCLM) mechanisms. Tested genotypes were selected from primary collection based on their antioxidant activity values, as well as higher level of flavonoids or phenolic acids. The stability of antioxidant properties after thermic, acidic, and basic treatments was evaluated. The oat cultivar Sirene and buckwheat cultivar Bogatyr expressed high level of the antioxidant activity, but they lost it due to all types of treatment. Oppositely, treatments increased antioxidant activities in some samples, especially in oat cultivar Maris Oberon, wheat cultivar Ines and Karolinum, or partially in barley cultivars Kompakt (after basic treatment) and Jubilant (acidic and basic treatments). The lack of the antioxidant activity could be observed due to destruction of the key compounds responsible for the antioxidant effect, whereas the increasing activity could be seen due to release of the aglycons from glycosidic forms after treatment. The stability of antioxidant properties could be a valuable parameter of the raw material for manufacturing special foods with functional properties.