Canine Recombinant Adenovirus Vector Induces an Immunogenicity-Related Gene Expression Profile in Skin-Migrated CD11b+ -Type DCs

Gene expression profiling of the blood cell response induced early after vaccination has previously been demonstrated to predict the immunogenicity of vaccines. In this study, we evaluated whether the analysis of the gene expression profile of skin-migrated dendritic cells (DCs) could be informative for the in vitro prediction of immunogenicity of vaccine, using canine adenovirus serotype 2 (CAV2) as vaccine vector. CAV2 has been shown to induce immunity to transgenes in several species including sheep and is an interesting alternative to human adenovirus-based vectors, based on the safety records of the parental strain in dogs and the lack of pre-existing immunity in non-host species. Skin-migrated DCs were collected from pseudo-afferent lymph in sheep. Both the CD11b⁺ -type and CD103⁺ -type skin-migrated DCs were transduced by CAV2. An analysis of the global gene response to CAV2 in the two skin DC subsets showed that the gene response in CD11b⁺ -type DCs was far higher and broader than in the CD103⁺ -type DCs. A newly released integrative analytic tool from Ingenuity systems revealed that the CAV2-modulated genes in the CD11b⁺ -type DCs clustered in several activated immunogenicity-related functions, such as immune response, immune cell trafficking and inflammation. Thus gene profiling in skin-migrated DC in vitro indicates that the CD11b⁺ DC type is more responsive to CAV2 than the CD103⁺ DC type, and provides valuable information to help in evaluating and possibly improving viral vector vaccine effectiveness.     

Rapeseed rotation, compost and biocontrol amendments reduce soilborne diseases and increase tuber yield in organic and conventional potato production systems

Aims

Integrating multiple soil and disease management practices may improve crop productivity and disease control, but potential interactions and limitations need to be determined.

Methods

Three different potential disease-suppressive management practices, including a Brassica napus (rapeseed) green manure rotation crop, conifer-based compost amendment, and three biological control organisms (Trichoderma virens, Bacillus subtilis, and Rhizoctonia solani hypovirulent isolate Rhs1A1) were evaluated alone and in combination at sites with both organic and conventional management histories for their effects on soilborne diseases and tuber yield.

Results

Rapeseed rotation reduced all observed soilborne diseases (stem canker, black scurf, common scab, and silver scurf) by 10 to 52 % in at least one year at both sites. Compost amendment had variable effects on tuber diseases, but consistently increased yield (by 9 to 15 %) at both sites. Biocontrol effects on disease varied, though Rhs1A1 decreased black scurf at the conventional site and T. virens reduced multiple diseases at the organic site in at least one year. Combining rapeseed rotation with compost amendment both reduced disease and increased yield, whereas biocontrol additions produced only marginal additive effects.

Conclusions

Use of these treatments alone, and in combination, can be effective at reducing disease and increasing yield under both conventional and organic production practices.     

Phosphorylation of ADF/cofilin abolishes EGF-induced actin nucleation at the leading edge and subsequent lamellipod extension

In metastatic rat mammary adenocarcinoma cells, cell motility can be induced by epidermal growth factor. One of the early events in this process is the massive generation of actin barbed ends, which elongate to form filaments immediately adjacent to the plasma membrane at the tip of the leading edge. As a result, the membrane moves outward and forms a protrusion. To test the involvement of ADF/cofilin in the stimulus-induced barbed end generation at the leading edge, we inhibited ADF/cofilin's activity in vivo by increasing its phosphorylation level using the kinase domain of LIM-kinase 1 (GFP-K). We report here that expression of GFP-K in rat cells results in the near total phosphorylation of ADF/cofilin, without changing either the G/F-actin ratio or signaling from the EGF receptor in vivo. Phosphorylation of ADF/cofilin is sufficient to completely inhibit the appearance of barbed ends and lamellipod protrusion, even in the continued presence of abundant G-actin. Coexpression of GFP-K, together with an active, nonphosphorylatable mutant of cofilin (S3A cofilin), rescues barbed end formation and lamellipod protrusion, indicating that the effects of kinase expression are caused by the phosphorylation of ADF/cofilin. These results indicate a direct role for ADF/cofilin in the generation of the barbed ends that are required for lamellipod extension in response to EGF stimulation.     

Size-selective oviposition by parasitoids and the evolution of life-history timing in hosts: fixed preferences vs frequency-dependent host selection

The influence of size-selective oviposition behaviour by parasitoids on the evolution of life-history timing in their hosts was examined using an optimization model of a two-stage life history similar to a genetic algorithm. Host populations with varying durations of early-larval development were subjected to selection in scenarios where parasitoids had fixed preferences for oviposition on late-stage larvae, or those where parasitoid attack was dependent on the relative frequencies of the two life stages present in the population. Fixed preference for oviposition on late-stage larvae caused positive directional selection on the duration of early-larval development. Surviving individuals remained for as long as possible in the first stage of development in order to avoid parasitoid attack. Frequency-dependent parasitoid attack, in contrast, caused maintenance of variation in the duration of early-larval development. The influence of the fitness payoffs of different life stages on the plasticity of size-selective oviposition behaviour is discussed, as are possible implications of the model results for parasitoid-host population dynamics.     

Biochemical and Genetic Characterization of Coagulin, a New Antilisterial Bacteriocin in the Pediocin Family of Bacteriocins, Produced by Bacillus coagulans I4

A plasmid-linked antimicrobial peptide, named coagulin, produced by Bacillus coagulans I₄ has recently been reported (B. Hyronimus, C. Le Marrec and M. C. Urdaci, J. Appl. Microbiol. 85:42–50, 1998). In the present study, the complete, unambiguous primary amino acid sequence of the peptide was obtained by a combination of both N-terminal sequencing of purified peptide and the complete sequence deduced from the structural gene harbored by plasmid I₄. Data revealed that this peptide of 44 residues has an amino acid sequence similar to that described for pediocins AcH and PA-1, produced by different Pediococcus acidilactici strains and 100% identical. Coagulin and pediocin differed only by a single amino acid at their C terminus. Analysis of the genetic determinants revealed the presence, on the pI₄ DNA, of the entire 3.5-kb operon of four genes described for pediocin AcH and PA-1 production. No extended homology was observed between pSMB74 from P. acidilactici and pI₄ when analyzing the regions upstream and downstream of the operon. An oppositely oriented gene immediately dowstream of the bacteriocin operon specifies a 474-amino-acid protein which shows homology to Mob-Pre (plasmid recombination enzyme) proteins encoded by several small plasmids extracted from gram-positive bacteria. This is the first report of a pediocin-like peptide appearing naturally in a non-lactic acid bacterium genus.     

Diterpenes from the leaves of Croton zambesicus

Two new trachylobane- and one isopimarane-type diterpenoids: ent-18-hydroxy-trachyloban-3-one; ent-trachyloban-3-one; isopimara-7,15-dien-3beta-ol, were isolated from the leaves of Croton zambesicus, together with trans-phytol, beta-sitosterol, alpha-amyrin and stigmasterol. The structures were determined by extensive NMR techniques and X-ray analysis. The cytotoxicity of these compounds has been evaluated on cancer and non-cancer cell-lines.     

The WISER metadatabase: the key to more than 100 ecological datasets from European rivers, lakes and coastal waters

In ecological sciences, the role of metadata (i.e. key information about a dataset) to make existing datasets visible and discoverable has become increasingly important. Within the EU-funded WISER project (Water bodies in Europe: Integrative Systems to assess Ecological status and Recovery), we designed a metadatabase to allow scientists to find the optimal data for their analyses. An online questionnaire helped to collect metadata from the data providers and an online query tool (http://www.wiser.eu/results/meta-database/) facilitated data evaluation. The WISER metadatabase currently holds information on 114 datasets (22 river, 71 lake, 1 general freshwater and 20 coastal/transitional datasets), which also can be accessed by external scientists. We evaluate if generally used metadata standards (e.g. Darwin Core, ISO 19115, CSDGM, EML) are suitable for such specific purposes as WISER and suggest at least the linkage with standard metadata fields. Furthermore, we discuss whether the simple metadata documentation is enough for others to reuse a dataset and why there is still reluctance to publish both metadata and primary research data (i.e. time and financial constraints, misuse of data, abandoning intellectual property rights). We emphasise that metadata publication has major advantages as it makes datasets detectable by other scientists and generally makes a scientist’s work more visible.     

The human polycomb group EED protein interacts with the integrase of human immunodeficiency virus type 1

Human EED, a member of the superfamily of WD-40 repeat proteins and of the Polycomb group proteins, has been identified as a cellular partner of the human immunodeficiency virus type 1 (HIV-1) matrix (MA) protein (R. Peytavi et al., J. Biol. Chem. 274:1635-1645, 1999). In the present study, EED was found to interact with HIV-1 integrase (IN) both in vitro and in vivo in yeast. In vitro, data from mutagenesis studies, pull-down assays, and phage biopanning suggested that EED-binding site(s) are located in the C-terminal domain of IN, between residues 212 and 264. In EED, two putative discrete IN-binding sites were mapped to its N-terminal moiety, at a distance from the MA-binding site, but EED-IN interaction also required the integrity of the EED last two WD repeats. EED showed an apparent positive effect on IN-mediated DNA integration reaction in vitro, in a dose-dependent manner. In situ analysis by immunoelectron microscopy (IEM) of cellular distribution of IN and EED in HIV-1-infected cells (HeLa CD4(+) cells or MT4 lymphoid cells) showed that IN and EED colocalized in the nucleus and near nuclear pores, with maximum colocalization events occurring at 6 h postinfection (p.i.). Triple colocalizations of IN, EED, and MA were also observed in the nucleoplasm of infected cells at 6 h p.i., suggesting the ocurrence of multiprotein complexes involving these three proteins at early steps of the HIV-1 virus life cycle. Such IEM patterns were not observed with a noninfectious, envelope deletion mutant of HIV-1.     

Mitochondrial protein import motor: differential role of Tim44 in the recruitment of Pam17 and J-complex to the presequence translocase

The presequence translocase of the mitochondrial inner membrane (TIM23 complex) mediates the import of preproteins with amino-terminal presequences. To drive matrix translocation the TIM23 complex recruits the presequence translocase-associated motor (PAM) with the matrix heat shock protein 70 (mtHsp70) as central subunit. Activity and localization of mtHsp70 are regulated by four membrane-associated cochaperones: the adaptor protein Tim44, the stimulatory J-complex Pam18/Pam16, and Pam17. It has been proposed that Tim44 serves as molecular platform to localize mtHsp70 and the J-complex at the TIM23 complex, but it is unknown how Pam17 interacts with the translocase. We generated conditional tim44 yeast mutants and selected a mutant allele, which differentially affects the association of PAM modules with TIM23. In tim44-804 mitochondria, the interaction of the J-complex with the TIM23 complex is impaired, whereas unexpectedly the binding of Pam17 is increased. Pam17 interacts with the channel protein Tim23, revealing a new interaction site between TIM23 and PAM. Thus, the motor PAM is composed of functional modules that bind to different sites of the translocase. We suggest that Tim44 is not simply a scaffold for binding of motor subunits but plays a differential role in the recruitment of PAM modules to the inner membrane translocase.