Small RNA binding is a common strategy to suppress RNA silencing by several viral suppressors

RNA silencing is an evolutionarily conserved system that functions as an antiviral mechanism in higher plants and insects. To counteract RNA silencing, viruses express silencing suppressors that interfere with both siRNA- and microRNA-guided silencing pathways. We used comparative in vitro and in vivo approaches to analyse the molecular mechanism of suppression by three well-studied silencing suppressors. We found that silencing suppressors p19, p21 and HC-Pro each inhibit the intermediate step of RNA silencing via binding to siRNAs, although the molecular features required for duplex siRNA binding differ among the three proteins. None of the suppressors affected the activity of preassembled RISC complexes. In contrast, each suppressor uniformly inhibited the siRNA-initiated RISC assembly pathway by preventing RNA silencing initiator complex formation.     

dsRNA-mediated resistance to Beet Necrotic Yellow Vein Virus infections in sugar beet (Beta vulgaris L. ssp. vulgaris)

Rhizomania, one of the most devastating diseases in sugar beet, is caused by Beet Necrotic Yellow Vein Virus (BNYVV) belonging to the genus Benyvirus. Use of sugar beet varieties with resistance to BNYVV is generally considered as the only way to maintain a profitable yield on rhizomania-infested fields. As an alternative to natural resistance, we explored the transgenic expression of viral dsRNA for engineering resistance to rhizomania. Transgenic plants expressing an inverted repeat of a 0.4 kb fragment derived from the BNYVV replicase gene displayed high levels of resistance against different genetic strains of BNYVV when inoculated using the natural vector, Polymyxa betae. The resistance was maintained under high infection pressures and over prolonged growing periods in the greenhouse as well as in the field. Resistant plants accumulated extremely low amounts of transgene mRNA and high amounts of the corresponding siRNA in the roots, illustrative of RNA silencing as the underlying mechanism. The transgenic resistance compared very favourably to natural sources of resistance to rhizomania and thus offers an attractive alternative for breeding resistant sugar beet varieties.     

Trichosporon mycotoxinivorans sp. nov., a new yeast species useful in biological detoxification of various mycotoxins

A yeast strain isolated from the hindgut of the lower termite Mastotermes darwiniensis (Mastotermitidae) was found to represent a new member of the genus Trichosporon. Trichosporon mycotoxinivorans is closely related to T. loubieri on the basis of the phylogenetic trees based on the D1/D2 region of 26S rDNA, an approx. 600 bp fragment of the 18S rDNA and both ITS regions. However, the two species differ at nine positions in the D1/D2 region of 26S rDNA. The IGS1 region of T. mycotoxinivorans is 401 bp long. T. mycotoxinivorans is distinguished from T. loubieri by its ability to assimilate inulin and galactitol, and its inability to grow at 40 °C. The name of this newly isolated strain refers to an important characteristics of T. mycotoxinivorans to detoxify mycotoxins such as ochratoxin A and zearalenone. Therefore this strain can be used for the deactivation of the respective mycotoxins in animal feeds.     

Aphid–willow interactions in a high Arctic ecosystem: responses to raised temperature and goose disturbance

Recently, there have been several studies using open top chambers (OTCs) or cloches to examine the response of Arctic plant communities to artificially elevated temperatures. Few, however, have investigated multitrophic systems, or the effects of both temperature and vertebrate grazing treatments on invertebrates. This study investigated trophic interactions between an herbivorous insect (Sitobion calvulum, Aphididae), a woody perennial host plant (Salix polaris) and a selective vertebrate grazer (barnacle geese, Branta leucopsis). In a factorial experiment, the responses of the insect and its host to elevated temperatures using open top chambers (OTCs) and to three levels of goose grazing pressure were assessed over two summer growing seasons (2004 and 2005). OTCs significantly enhanced the leaf phenology of Salix in both years and there was a significant OTC by goose presence interaction in 2004. Salix leaf number was unaffected by treatments in both years, but OTCs increased leaf size and mass in 2005. Salix reproduction and the phenology of flowers were unaffected by both treatments. Aphid densities were increased by OTCs but unaffected by goose presence in both years. While goose presence had little effect on aphid density or host plant phenology in this system, the OTC effects provide interesting insights into the possibility of phenological synchrony disruption. The advanced phenology of Salix effectively lengthens the growing season for the plant, but despite a close association with leaf maturity, the population dynamics of the aphid appeared to lack a similar phenological response, except for the increased population observed.     

Diglyceride kinase from Escherichia coli: Modulation of enzyme activity by glycosphingolipids

Diglyceride kinase was purified from membranes of Escherichia coli K-12 using organic solvents. The enzyme apoprotein depended on lipids, such as cardiolipin (diphosphatidylglycerol), phosphatidylcholine or 1-monooleoylglycerol, for activity with 1,2-dipalmitoylglycerol. Mixed brain cerebrosides and gangliosides as well as defined ganglioside fractions and synthetic lactocerebroside were devoid of lipid cofactor activity. However, all these glycosphingolipids were strong inhibitors of activation by phosphatidylcholine. When cardiolipin was used as lipid activator with the detergent, Triton X-100, as solubilizing agent, the addition of mixed or purified gangliosides first (at about 0.4 mM) resulted in additional activation, but higher ganglioside concentrations were strongly inhibitory. Both effects were absolutely dependent on the presence of lipid-bound sialic acid and were not given by cerebrosides, by free sialic acid or by sialyl-lactose. The stimulating and inhibitory effects of glycosphingolipids could also be demonstrated when 1-monooleoylglycerol was used as substrate, lipid activator and solubilizing agent at the same time. The modulation of kinase activity by glycosphingolipids is discussed at the level of lipid/protein interactions.     

The Universal Stress Protein UspC Scaffolds the KdpD/KdpE Signaling Cascade of Escherichia coli under Salt Stress

The sensor kinase KdpD and the response regulator KdpE control induction of the kdpFABC operon encoding the high-affinity K⁺-transport system KdpFABC in response to K⁺ limitation or salt stress. Under K⁺ limiting conditions the Kdp system restores the intracellular K⁺ concentration, while in response to salt stress K⁺ is accumulated far above the normal content. The kinase activity of KdpD is inhibited at high concentrations of K⁺, so it has been puzzling how the sensor can be activated in response to salt stress. Here, we demonstrate that the universal stress protein UspC acts as a scaffolding protein of the KdpD/KdpE signaling cascade by interacting with a Usp domain in KdpD of the UspA subfamily under salt stress. Escherichia coli encodes three single domain proteins of this subfamily, UspA, UspC, and UspD, whose expression is up-regulated under various stress conditions. Among these proteins only UspC stimulated the in vitro reconstructed signaling cascade (KdpD→KdpE→DNA) resulting in phosphorylation of KdpE at a K⁺ concentration that would otherwise almost prevent phosphorylation. In agreement, in a ΔuspC mutant KdpFABC production was down-regulated significantly when cells were exposed to salt stress, but unchanged under K⁺ limitation. Biochemical studies revealed that UspC interacts specifically with the Usp domain in the stimulus perceiving N-terminal domain of KdpD. Furthermore, UspC stabilized the KdpD/KdpE∼P/DNA complex and is therefore believed to act as a scaffolding protein. This study describes the stimulation of a bacterial two-component system under distinct stress conditions by a scaffolding protein, and highlights a new role of the universal stress proteins.     

Development and validation of a prototype 16S rRNA-based taxonomic microarray for Alphaproteobacteria

The microarray approach has been proposed for high throughput analysis of the microbial community by providing snapshots of the microbial diversity under different environmental conditions. For this purpose, a prototype of a 16S rRNA-based taxonomic microarray was developed and evaluated for assessing bacterial community diversity. The prototype microarray is composed of 122 probes that target bacteria at various taxonomic levels from phyla to species (mostly Alphaproteobacteria). The prototype microarray was first validated using bacteria in pure culture. Differences in the sequences of probes and potential target DNAs were quantified as weighted mismatches (WMM) in order to evaluate hybridization reliability. As a general feature, probes having a WMM > 2 with target DNA displayed only 2.8% false positives. The prototype microarray was subsequently tested with an environmental sample, which consisted of an Agrobacterium-related polymerase chain reaction amplicon from a maize rhizosphere bacterial community. Microarray results were compared to results obtained by cloning-sequencing with the same DNA. Microarray analysis enabled the detection of all 16S rRNA gene sequences found by cloning-sequencing. Sequences representing only 1.7% of the clone library were detected. In conclusion, this prototype 16S rRNA-based taxonomic microarray appears to be a promising tool for the analysis of Alphaproteobacteria in complex ecosystems.     

Halimeda contribution to organic and inorganic production in a Tahitian reef system

Of the seven species of Halimeda inhabiting a lagoon on Moorea island, three representing 10% of the algal covering and averaging 111 g of dry weight m^-2, have been studied in the course of a year. The biomass, measured bimonthly, stresses a slight seasonal variability in the species life. The main decrease was reported for H. opuntia after a fruiting event, which happened in October. The primary production was assessed, in situ, periodically over a year, by measuring oxygen variations in enclosures. Either expressed on specific-weight basis or in area units, the highest primary productions were recorded for H. opuntia. Productions and biomasses vary simultaneously during the year. The three species produce all together about 6 g C m^-2y^-1. The growth rate of the sand-dwelling H. incrassata f. ovata was followed during the year by staining, in the field, individuals with alizarin Red-S. The average rates measured were 3.3 segments ind^-1d^-1 and 0.17 gdw d^-1m^-2. The contribution of the three species to the carbonate budget of the reef was estimated by total alkalinity measurements during 24-h cycles. H. opuntia had the highest CaCO₃ production. For the three species studied, CaCO₃ production of 1.4 kg m^-2y^-1, which could correspond to a 0.4mm/year accretion of the studied reefal system, was estimated.     

Complete elimination of hostplant quinolizidine alkaloids by larvae of a polyphagous lycaenid butterfly,Callophrys rubi

Caterpillars of the lycaenid butterfly Callophrys rubi accept a variety of hostplants. When fed inflorescences or leaves of Genista tinctoria (a natural hostplant) or Lupinus polyphyllus (a non-host), the larvae completely eliminate quinolizidine alkaloids ingested from their food in their frass. No alkaloids are stored. Infestation by the parasitoid wasp Distatrix sancus (Braconidae) did not affect alkaloid elimination. The presence of an effective anti-toxin system is discussed with reference to the evolution of hostplant relationships in the genus Callophrys. There is no evidence that in the secondarily myrmecoxenous larvae of C. rubi hostplantderived chemical defense takes the place of former myrmecophily.