Immunodetection of disease-associated mutant PrP, which accelerates disease in GSS transgenic mice

The absence of infectivity-associated, protease-resistant prion protein (PrP^Sc) in the brains of spontaneously sick transgenic (Tg) mice overexpressing PrP linked to Gerstmann–Sträussler Scheinker syndrome, and the failure of gene-targeted mice expressing such PrP to develop disease spontaneously, challenged the concept that mutant PrP expression led to spontaneous prion production. Here, we demonstrate that disease in overexpressor Tg mice is associated with accumulation of protease-sensitive aggregates of mutant PrP that can be immunoprecipitated by the PrP^Sc-specific monoclonal antibody designated 15B3. Whereas Tg mice expressing multiple transgenes exhibited accelerated disease when inoculated with disease-associated mutant PrP, Tg mice expressing mutant PrP at low levels failed to develop disease either spontaneously or following inoculation. These studies indicate that inoculated mutant PrP from diseased mice promotes the aggregation and accumulation of pre-existing pathological forms of mutant PrP produced as a result of transgene overexpression. Thus, while pathological mutant PrP possesses a subset of PrP^Sc characteristics, we now show that the attribute of prion transmission suggested by previous studies is more accurately characterized as disease acceleration.     

Defence of Rhizobium etli bacteroids against oxidative stress involves a complexly regulated atypical 2-Cys peroxiredoxin

In general, oxidative stress, the consequence of an aerobic lifestyle, induces bacterial antioxidant defence enzymes. Here we report on a peroxiredoxin of Rhizobium etli, prxS, strongly expressed under microaerobic conditions and during the symbiotic interaction with Phaseolus vulgaris. The microaerobic induction of the prxS-rpoN2 operon is mediated by the alternative sigma factor RpoN and the enhancer-binding protein NifA. The RpoN-dependent promoter is also active under low-nitrogen conditions through the enhancer-binding protein NtrC. An additional symbiosis-specific weak promoter is located between prxS and rpoN2. Constitutive expression of prxS confers enhanced survival and growth to R. etli in the presence of H2O2. Single prxS mutants are not affected in their symbiotic abilities or defence response against oxidative stress under free-living conditions. In contrast, a prxS katG double mutant has a significantly reduced (>40%) nitrogen fixation capacity, suggesting a functional redundancy between PrxS and KatG, a bifunctional catalase-peroxidase. In vitro assays demonstrate the reduction of PrxS protein by DTT and thioredoxin. PrxS displays substrate specificity towards H2O2 (Km = 62 microM) over alkyl hydroperoxides (Km > 1 mM). Peroxidase activity is abolished in both the peroxidatic (C56) and resolving (C156) cysteine PrxS mutants, while the conserved C81 residue is required for proper folding of the protein. Resolving of the R. etli PrxS peroxidatic cysteine is probably an intramolecular process and intra- and intersubunit associations were observed. Taken together, our data support, for the first time, a role for an atypical 2-Cys peroxiredoxin against oxidative stress in R. etli bacteroids.     

Rab8, POSH,and TAK1 regulate synaptic growth in a Drosophila model of frontotemporal dementia

Mutations in genes essential for protein homeostasis have been identified in frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) patients. Why mature neurons should be particularly sensitive to such perturbations is unclear. We identified mutations in Rab8 in a genetic screen for enhancement of an FTD phenotype associated with ESCRT-III dysfunction. Examination of Rab8 mutants or motor neurons expressing a mutant ESCRT-III subunit, CHMP2B^Intron5, at the Drosophila melanogaster neuromuscular junction synapse revealed synaptic overgrowth and endosomal dysfunction. Expression of Rab8 rescued overgrowth phenotypes generated by CHMP2B^Intron5. In Rab8 mutant synapses, c-Jun N-terminal kinase (JNK)/activator protein-1 and TGF-β signaling were overactivated and acted synergistically to potentiate synaptic growth. We identify novel roles for endosomal JNK-scaffold POSH (Plenty-of-SH3s) and a JNK kinase kinase, TAK1, in regulating growth activation in Rab8 mutants. Our data uncover Rab8, POSH, and TAK1 as regulators of synaptic growth responses and point to recycling endosome as a key compartment for synaptic growth regulation during neurodegenerative processes.     

Assessment of Efficacy and Quality of Two Albendazole Brands Commonly Used against Soil-Transmitted Helminth Infections in School Children in Jimma Town,Ethiopia

BackgroundThere is a worldwide upscale in mass drug administration (MDA) programs to control the morbidity caused by soil-transmitted helminths (STHs): Ascaris lumbricoides, Trichuris trichiura and hookworm. Although anthelminthic drugs which are used for MDA are supplied by two pharmaceutical companies through donation, there is a wide range of brands available on local markets for which the efficacy against STHs and quality remain poorly explored. In the present study, we evaluated the drug efficacy and quality of two albendazole brands (Bendex and Ovis) available on the local market in Ethiopia.Conclusion/SignificanceThe study revealed that differences in efficacy between the two brands of albendazole (ABZ) tablets against hookworm are linked to the differences in the in-vitro drug release profile. Differences in uptake and metabolism of this benzimidazole drug among different helminth species may explain that this efficacy difference was only observed in hookworms and not in the two other species. The results of the present study underscore the importance of assessing the chemical and physicochemical quality of drugs before conducting efficacy assessment in any clinical trials to ensure appropriate therapeutic efficacy and to exclude poor drug quality as a factor of reduced drug efficacy other than anthelminthic resistance. Overall, this paper demonstrates that “all medicines are not created equal”.     

Tree Height Reduction After Selective Logging in a Tropical Forest

By harvesting scattered large trees, selective logging increases light availability and thereby stimulates growth and crown expansion at early‐life stage among remnant trees. We assessed the effects of logging on total and merchantable bole (i.e., lowest branch at crown base) heights on 952 tropical canopy trees in French Guiana. We observed reductions in both total (mean, ?2.3 m) and bole (mean, ?2.0 m) heights more than a decade after selective logging. Depending on local logging intensity, height reductions resulted in 2–13 percent decreases in aboveground tree biomass and 3–17 percent decreases in bole volume. These results highlight the adverse effects of logging at both tree and stand levels. This decrease in height is a further threat to future provision of key environmental services, such as timber production and carbon sequestration.     

Performance of LBSap Vaccine after Intradermal Challenge with L. infantum and Saliva of Lu. longipalpis: Immunogenicity and Parasitological Evaluation

In the last decade, the search for new vaccines against canine visceral leishmaniasis has intensified. However, the pattern related to immune protection during long periods after experimental infection in vaccine trials is still not fully understood. Herein, we investigated the immunogenicity and parasitological levels after intradermal challenge with Leishmania infantum plus salivary gland extract in dogs immunized with a vaccine composed of L. braziliensis antigens plus saponin as an adjuvant (LBSap vaccine). The LBSap vaccine elicited higher levels of total anti-Leishmania IgG as well as both IgG1 and IgG2. Furthermore, dogs vaccinated had increased levels of lymphocytes, particularly circulating B cells (CD21⁺) and both CD4⁺ and CD8⁺ T lymphocytes. LBSap also elicited an intense in vitro cell proliferation associated with higher levels of CD4⁺ T lymphocytes specific for vaccine soluble antigen and soluble lysate of L. infantum antigen even 885 days after experimental challenge. Furthermore, LBSap vaccinated dogs presented high IFN-γ and low IL-10 and TGF-β1 expression in spleen with significant reduction of parasite load in this tissue. Overall, our results validate the potential of LBSap vaccine to protect against L. infantum experimental infection and strongly support further evaluation of efficiency of LBSap against CVL in natural infection conditions.     

Engineering seed dormancy by the modification of zeaxanthin epoxidase gene expression

Abscisic acid (ABA) is a plant hormone synthesized during seed development that is involved in the induction of seed dormancy. Delayed germination due to seed dormancy allows long-term seed survival in soil but is generally undesirable in crop species. Freshly harvested seeds of wild-type Nicotiana plumbaginifolia plants exhibit a clear primary dormancy that results in delayed germination, the degree of primary dormancy being influenced by environmental culture conditions of the mother plant. In contrast, seeds, obtained either from ABA-deficient mutant aba2-s1 plants directly or aba2-s1 plants grafted onto wild-type plant stocks, exhibited rapid germination under all conditions irrespective of the mother plant culture conditions. The ABA biosynthesis gene ABA2 of N. plumbaginifolia, encoding zeaxanthin epoxidase, was placed under the control of the constitutive 35S promoter. Transgenic plants overexpressing ABA2 mRNA exhibited delayed germination and increased ABA levels in mature seeds. Expression of an antisense ABA2 mRNA, however, resulted in rapid seed germination and in a reduction of ABA abundance in transgenic seeds. It appears possible, therefore, that seed dormancy can be controlled in this Nicotiana model species by the manipulation of ABA levels.     

Association between reduced bronchodilatory effect of deep inspiration and loss of alveolar attachments

Background

We have previously shown that the bronchodilatory effect of deep inspiration is attenuated in individuals with COPD. This study was designed to investigate whether the impairment in this effect is associated with loss of alveolar attachments.

Methods

We measured deep inspiration (DI)-induced bronchodilation in 15 individuals with and without COPD (67 ± 2.2 yrs of age, mean ± SEM) undergoing lobar resection for peripheral pulmonary nodule. Prior to surgery, we measured TLCO and determined the bronchodilatory effect of deep inspiration after constricting the airways with methacholine. The number of destroyed alveolar attachments, as well as airway wall area and airway smooth muscle area, were determined in tumor-free, peripheral lung tissue.

Results

The bronchodilatory effect of deep inspiration correlated inversely with the % destroyed attachments (r = -0.51, p = 0.05) and directly with the airway smooth muscle area (r = 0.59, p = 0.03), but not with the total wall area (r = 0.39, p = 0.15).

Conclusion

We postulate that attenuation of airway stretch due to loss of alveolar attachments contributes to the loss of the bronchodilatory effect of lung inflation in COPD.     

Crystal structure determination and site-directed mutagenesis of the Pyrococcus abyssi aCBF5-aNOP10 complex reveal crucial roles of the C-terminal domains of both proteins in H/ACA sRNP activity

In archaeal rRNAs, the isomerization of uridine into pseudouridine (Ψ) is achieved by the H/ACA sRNPs and the minimal set of proteins required for RNA:Ψ-synthase activity is the aCBF5–aNOP10 protein pair. The crystal structure of the aCBF5–aNOP10 heterodimer from Pyrococcus abyssi was solved at 2.1 Å resolution. In this structure, protein aNOP10 has an extended shape, with a zinc-binding motif at the N-terminus and an α-helix at the C-terminus. Both motifs contact the aCBF5 catalytic domain. Although less efficiently as does the full-length aNOP10, the aNOP10 C-terminal domain binds aCBF5 and stimulates the RNA-guided activity. We show that the C-terminal domain of aCBF5 (the PUA domain), which is wrapped by an N-terminal extension of aCBF5, plays a crucial role for aCBF5 binding to the guide sRNA. Addition of this domain in trans partially complement particles assembled with an aCBF5ΔPUA truncated protein. In the crystal structure, the aCBF5–aNOP10 complex forms two kinds of heterotetramers with parallel and perpendicular orientations of the aNOP10 terminal α-helices, respectively. By gel filtration assay, we showed that aNOP10 can dimerize in solution. As both residues Y41 and L48 were needed for dimerization, the dimerization likely takes place by interaction of parallel α-helices.