Vaccination of BALB/c mice with Escherichia coli-expressed vaccinia virus proteins A27L, B5R, and D8L protects mice from lethal vaccinia virus challenge

The potential threat of smallpox use in a bioterrorist attack has heightened the need to develop an effective smallpox vaccine for immunization of the general public. Vaccination with the current smallpox vaccine, Dryvax, produces protective immunity but may result in adverse reactions for some vaccinees. A subunit vaccine composed of protective vaccinia virus proteins should avoid the complications arising from live-virus vaccination and thus provide a safer alternative smallpox vaccine. In this study, we assessed the protective efficacy and immunogenicity of a multisubunit vaccine composed of the A27L and D8L proteins from the intracellular mature virus (IMV) form and the B5R protein from the extracellular enveloped virus (EEV) form of vaccinia virus. BALB/c mice were immunized with Escherichia coli-produced A27L, D8L, and B5R proteins in an adjuvant consisting of monophosphoryl lipid A and trehalose dicorynomycolate or in TiterMax Gold adjuvant. Following immunization, mice were either sacrificed for analysis of immune responses or lethally challenged by intranasal inoculation with vaccinia virus strain Western Reserve. We observed that three immunizations either with A27L, D8L, and B5R or with the A27L and B5R proteins alone induced potent neutralizing antibody responses and provided complete protection against lethal vaccinia virus challenge. Several linear B-cell epitopes within the three proteins were recognized by sera from the immunized mice. In addition, protein-specific cellular responses were detected in spleens of immunized mice by a gamma interferon enzyme-linked immunospot assay using peptides derived from each protein. Our data suggest that a subunit vaccine incorporating bacterially expressed IMV- and EEV-specific proteins can be effective in stimulating anti-vaccinia virus immune responses and providing protection against lethal virus challenge.     

Intra-annual tree-ring parameters indicating differences in drought stress of Pinus sylvestris forests within the Erico-Pinion in the Valais (Switzerland)

The effects of drought on radial growth of Pinussylvestris were investigated by comparing sites along hydricgradients. The gradients were located in Valais, an inner Alpine dry valley inSwitzerland, with each consisting of two site types, an extreme dry, xeric siteand a less dry, moderate site. The two site types were assigned tophytosociological associations within the Erico-Pinion. The investigationcovered the responses of tree growth to climate and particularly concentratedonintra-annual features of tree-rings such as earlywood/latewood ratio,intra-annual density fluctuations (IADFs) and traumatic tissues (TTs) as wellasthe sapwood/heartwood ratio. Radial growth differed according to the sitetypes,with trees on dry sites generally showing more missing rings, lower mean ringwidths, lower autocorrelation, higher mean sensitivities, reduced latewoodproportions and lower sapwood areas than trees on moderate sites. Therelationships between climate and tree-ring width, studied using responsefunction analysis, varied strongly between the site types within theErico-Pinion: Tree growth on dry sites was positively influenced byprecipitation at the end of the winter and the beginning of the growing seasonand negatively influenced by temperature in June. Winter precipitation waspositively correlated with radial growth, demonstrating its importance for thesuccessful root and shoot growth of the plants in spring on dry sites. Onmoderate sites, tree growth was less controlled by climate than by priorgrowth.The intra-annual density fluctuations (IADFs) provided a valuable means todifferentiate between the site types. In comparison to the moderate sites, thetrees on dry sites contained more IADFs, and their frequency was increased.Moist-cool conditions in the middle of the growing season were the triggeringfactor for IADFs on dry sites, whereas on moderate sites, there must be anadditional warm period in early summer in order to initiate IADFs. Most IADFswere found in latewood. We found no relationship between climate and traumatictissues (TTs). It is unclear whether other abiotic or biotic factors such aswounding by insects or birds are responsible for the development of TTs. Theassignment of these differences in tree growth behaviour to phytosociologicalassociations will enable a deeper understanding of the site types and willfacilitate the comparison with similar studies. Furthermore, the results can becombined with studies from other scientific disciplines concerning thesephytosociological associations. The ecological indicator values of thevegetation was a precise method for the distinction of site types.     

Phospholipids: synthesis, sorting, subcellular traffic - the yeast approach

Most of the enzymes and genes required for lipid biosynthesis and degradation in the budding yeast Saccharomyces cerevisiae have now been identified and the global mechanisms that regulate their activity are being established. Synthesis of phospholipids is restricted to specific subcellular compartments, and the lipids migrate from their site of formation to their final destination. In addition to synthesis, remodelling and degradation of phospholipids controls the content of the lipid portion of cellular membranes, while highly specific phospholipases catalyse the release of lipid-based second messengers. In this review, we describe the current understanding of the organization and regulation of phospholipid metabolism in yeast, and discuss the mechanisms that have been proposed for intracellular lipid transport.     

The nuclear GTPase cycle: promoting peripheralization?

Numerous Ras-like GTPases function as molecular switches in the cytoplasm, but only one has been identified in the nucleus. This nuclear GTPase and its homologues are known in both yeasts and higher organisms and in all cases they are regulated by guanine-nucleotide-exchange factors. The 'nuclear GTPase cycle' created by these components is implicated in mRNA transport from and protein import to the nucleus, as well as in DNA replication, RNA processing and the regulation of the cell cycle. In this article, Alan Tartakoff and Roger Schneiter propose that this GTPase cycle regulates dispersive functions in the nucleoplasm, an idea that explains many of the observed effects of disrupting the cycle.     

Mutations in nucleolar proteins lead to nucleolar accumulation of polyA+ RNA in Saccharomyces cerevisiae.

Synthesis of mRNA and rRNA occur in the chromatin-rich nucleoplasm and the nucleolus, respectively. Nevertheless, we here report that a Saccharomyces cerevisiae gene, MTR3, previously implicated in mRNA transport, codes for a novel essential 28-kDa nucleolar protein. Moreover, in mtr3-1 the accumulated polyA+ RNA actually colocalizes with nucleolar antigens, the nucleolus becomes somewhat disorganized, and rRNA synthesis and processing are inhibited. A strain with a ts conditional mutation in RNA polymerase I also shows nucleolar accumulation of polyA+ RNA, whereas strains with mutations in the nucleolar protein Nop1p do not. Thus, in several mutant backgrounds, when mRNA cannot be exported i concentrates in the nucleolus. mRNA may normally encounter nucleolar components before export and proteins such as Mtr3p may be critical for export of both mRNA and ribosomal subunits.     

Heligmosomoides polygyrus Venom Allergen-like Protein-4 (HpVAL-4) is a sterol binding protein

Heligmosomoides polygyrus bakeri is a model parasitic hookworm used to study animal and human helminth diseases. During infection, the parasite releases excretory/secretory products that modulate the immune system of the host. The most abundant protein family in excretory/secretory products comprises the venom allergen-like proteins (VALs), which are members of the SCP/TAPS (sperm-coating protein/Tpx/antigen 5/pathogenesis related-1/Sc7) superfamily. There are >30 secreted Heligmosomoides polygyrus VAL proteins (HpVALs) and these proteins are characterised by having either one or two 15?kDa CAP (cysteine-rich secretory protein (CRISP)/antigen 5/pathogenesis related-1) domains. The first known HpVAL structure, HpVAL-4, refined to 1.9?Å is reported. HpVAL-4 was produced as a homogeneously glycosylated protein in leaves of Nicotiana benthamiana infiltrated with recombinant plasmids, making this plant expression platform amenable for the production of biological products. The overall topology of HpVAL-4 is a three layered αβα sandwich between a short N-terminal loop and a C-terminal cysteine rich extension. The C-terminal cysteine rich extension has two strands stabilized by two disulfide bonds and superposes well with the previously reported extension from the human hookworm Necator americanus Ancylostoma secreted protein-2 (Na-ASP-2). The N-terminal loop is connected to alpha helix 2 via a disulfide bond previously observed in Na-ASP-2. HpVAL-4 has a central cavity that is more similar to the N-terminal CAP domain of the two CAP Na-ASP-1 from Necator americanus. Unlike Na-ASP-2, mammalian CRISP, and the C-terminal CAP domain of Na-ASP-1, the large central cavity of HpVAL-4 lacks the two histidines required to coordinate divalent cations. HpVAL-4 has both palmitate-binding and sterol-binding cavities and is able to complement the in vivo sterol export phenotype of yeast mutants lacking their endogenous CAP proteins. More studies are required to determine endogenous binding partners of HpVAL-4 and unravel the possible impact of sterol binding on immune-modulatory functions.     

Allometric equations to predict the total aboveground biomass of tree species in a planted forest in Egypt

The aim of this study is to estimate the total above‐ground biomass (TAGB), stem height (H), diameter at breast height (dbh) and basal area of five tree species (ages 7‐8 years) irrigated by municipal sewage water in the Egyptian‐Chinese friendship forest, Sadat City, Egypt. From the biomass data that obtained through destructive sampling, models for predicting aboveground biomass were developed. The highest values for stem density and height were estimated for Eucalyptus citriodora, while the lowest value for density was obtained for Dalbergia sissoo and stem height for Khaya senegalensis. The highest values for basal area and dbh were obtained for Casuarina spp., while the lowest values were recorded for Dalbergia sissoo. Eucalyptus camaldulensis had the highest stand stem biomass and TAGB (55.5, 83.9 t DW ha^‐1, respectively). In addition, Casuarina spp. had the highest leafy branches biomass (32.5 t DW ha^‐1) while Dalbergia sissoo had the lowest values for all tree components. All the generated allometric equations had high correlation coefficients at high probability levels. Moreover, the results revealed that not only the dbh data can be used as independent variable for biomass determination, but also stem height and size index are recommended for biomass estimation (© 2012 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

A highly efficient and selective turn‐on fluorescent sensor for Hg2+, Ag+ and Ag nanoparticles based on a coumarin dithioate derivative

Based on chelation‐enhanced fluorescence, a new fluorescent coumarin derivative probe 3(1‐(7‐hydroxy‐4‐methylcoumarin)ethylidene)hydrazinecarbodithioate for Hg²⁺, Ag⁺ and Ag nanoparticles is reported. Fluorescent probe acts as a rapid and highly selective “off–on” fluorescent probe and fluorescence enhancement by factors 5 to12 times was observed upon selective complexation with Hg²⁺, Ag⁺ and Ag nanoparticles. The molar ratio plots indicated the formation of 1:1 complexes between Hg²⁺ and Ag⁺ with the probe. The linear response range covers a concentration range 0.1 × 10^–5–1.9 × 10^–5 mol/L, 0.1 × 10^–5–2.3 × 10^–5 mol/L and 0.146 × 10^–12–2.63 × 10^–12 mol/L for Hg²⁺, Ag⁺ and Ag nanoparticles, respectively. The interference effect of some anions and cations was also tested. Copyright © 2013 John Wiley & Sons, Ltd.