Gawky is a component of cytoplasmic mRNA processing bodies required for early Drosophila development

In mammalian cells, the GW182 protein localizes to cytoplasmic bodies implicated in the regulation of messenger RNA (mRNA) stability, translation, and the RNA interference pathway. Many of these functions have also been assigned to analogous yeast cytoplasmic mRNA processing bodies. We have characterized the single Drosophila melanogaster homologue of the human GW182 protein family, which we have named Gawky (GW). Drosophila GW localizes to punctate, cytoplasmic foci in an RNA-dependent manner. Drosophila GW bodies (GWBs) appear to function analogously to human GWBs, as human GW182 colocalizes with GW when expressed in Drosophila cells. The RNA-induced silencing complex component Argonaute2 and orthologues of LSm4 and Xrn1 (Pacman) associated with 5'-3' mRNA degradation localize to some GWBs. Reducing GW activity by mutation or antibody injection during syncytial embryo development leads to abnormal nuclear divisions, demonstrating an early requirement for GWB-mediated cytoplasmic mRNA regulation. This suggests that gw represents a previously unknown member of a small group of genes that need to be expressed zygotically during early embryo development.     

Roles of glutamate receptors and the mammalian target of rapamycin (mTOR) signaling pathway in activity-dependent dendritic protein synthesis in hippocampal neurons

Local protein synthesis in neuronal dendrites is critical for synaptic plasticity. However, the signaling cascades that couple synaptic activation to dendritic protein synthesis remain elusive. The purpose of this study is to determine the role of glutamate receptors and the mammalian target of rapamycin (mTOR) signaling in regulating dendritic protein synthesis in live neurons. We first characterized the involvement of various subtypes of glutamate receptors and the mTOR kinase in regulating dendritic synthesis of a green fluorescent protein (GFP) reporter controlled by alphaCaMKII 5' and 3' untranslated regions in cultured hippocampal neurons. Specific antagonists of N-methyl-d-aspartic acid (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), and metabotropic glutamate receptors abolished glutamate-induced dendritic GFP synthesis, whereas agonists of NMDA and metabotropic but not AMPA glutamate receptors activated GFP synthesis in dendrites. Inhibitions of the mTOR signaling, as well as its upstream activators, phosphatidylinositol 3-kinase and AKT, blocked NMDA receptor-dependent dendritic GFP synthesis. Conversely, activation of mTOR signaling stimulated dendritic GFP synthesis. In addition, we also found that inhibition of the mTOR kinase blocked dendritic synthesis of the endogenous alphaCaMKII and MAP2 proteins induced by tetanic stimulations in hippocampal slices. These results identify critical roles of NMDA receptors and the mTOR signaling pathway for control of synaptic activity-induced dendritic protein synthesis in hippocampal neurons.     

Pharmacological targeting of secondary brain damage following ischemic or hemorrhagic stroke,traumatic brain injury,and bacterial meningitis - a systematic review and meta-analysis

Background

The effectiveness of pharmacological strategies exclusively targeting secondary brain damage (SBD) following ischemic stroke, aneurysmal subarachnoid hemorrhage, aSAH, intracerebral hemorrhage (ICH), traumatic brain injury (TBI) and bacterial meningitis is unclear. This meta-analysis studied the effect of SBD targeted treatment on clinical outcome across the pathological entities.

Methods

Randomized, controlled, double-blinded trials on aforementioned entities with ‘death’ as endpoint were identified. Effect sizes were analyzed and expressed as pooled risk ratio (RR) estimates with 95% confidence intervals (CI). 123 studies fulfilled the criteria, with data on 66,561 patients.

Results

In the pooled analysis, there was a minor reduction of mortality for aSAH [RR 0.93 (95% CI:0.85–1.02)], ICH [RR 0.92 (95% CI:0.82–1.03)] and bacterial meningitis [RR 0.86 (95% CI:0.68–1.09)]. No reduction of mortality was found for ischemic stroke [RR 1.05 (95% CI:1.00–1.11)] and TBI [RR 1.03 (95% CI:0.93–1.15)]. Additional analysis of “poor outcome” as endpoint gave similar results. Subgroup analysis with respect to effector mechanisms showed a tendency towards a reduced mortality for the effector mechanism category “oxidative metabolism/stress” for aSAH with a risk ratio of 0.86 [95% CI: 0.73–1.00]. Regarding specific medications, a statistically significant reduction of mortality and poor outcome was confirmed only for nimodipine for aSAH and dexamethasone for bacterial meningitis.

Conclusions

Our results show that only a few selected SBD directed medications are likely to reduce the rate of death and poor outcome following aSAH, and bacterial meningitis, while no convincing evidence could be found for the usefulness of SBD directed medications in ischemic stroke, ICH and TBI. However, a subtle effect on good or excellent outcome might remain undetected. These results should lead to a new perspective of secondary reactions following cerebral injury. These processes should not be seen as suicide mechanisms that need to be fought. They should be rather seen as well orchestrated clean-up mechanisms, which may today be somewhat too active in a few very specific constellations, such as meningitis under antibiotic treatment and aSAH after surgical or endovascular exclusion of the aneurysm.

     

Recombinant flagellin-PAL fusion protein of <Emphasis Type="Italic">Legionella pneumophila</Emphasis> induced cell-mediated and protective immunity against bacteremia in BALB/c mice

We report a new recombinant fusion protein composed of full-length Legionella pneumophila flagellin A and peptidoglycan-associated lipoprotein (PAL), rFLA-PAL, capable of inducing protective immunity against L. pneumophila. The recombinant protein was over expressed in Escherichia coli strain BL21 (DE3) using pET-28a (+) expression vector (pET28a-flaA-pal) and purified by Ni²⁺ exchange chromatography. Immunological properties of rFLA-PAL were assessed in a mouse model. Female BALB/c mice, immunized with rFLA-PAL, exhibited a rapid increase in serum antibody concentration against each of its protein portions. Furthermore, a strong activation of both innate and adaptive cell-mediated immunity was observed as indicated by antigen-specific splenocyte proliferation, IFN-γ and IL-12 production, and early production of TNF-α in the serum and in splenocyte cultures which were separately assessed against PAL and FLA. BALB/c mice were challenged with a lethal dose of L. pneumophila intravenously. In a 10-days follow-up after intravenous lethal challenge with L. pneumophila, a 100% survival rate was observed for mice immunized with rFLA-PAL, same as for those immunized with a sublethal dose of L. pneumophila. Based on the potent immune responses observed in mice immunized with rFLA-PAL, this recombinant fusion protein could be a potential vaccine candidate against the intracellular pathogen L. pneumophila.     

Investigation into the structural, morphological, mechanical and thermal behaviour of bacterial cellulose after a two-step purification process

Bacterial cellulose (BC) is a natural hydrogel, which is produced by Acetobacter xylinum (recently renamed Gluconacetobacter xylinum) in culture and constitutes of a three-dimensional network of ribbon-shaped bundles of cellulose microfibrils. Here, a two-step purification process is presented that significantly improves the structural, mechanical, thermal and morphological behaviour of BC sheet processed from these hydrogels produced in static culture. Alkalisation of BC using a single-step treatment of 2.5 wt.% NaOH solution produced a twofold increase in Young's modulus of processed BC sheet over untreated BC sheet. Further enhancements are achieved after a second treatment with 2.5 wt.% NaOCl (bleaching). These treatments were carefully designed in order to prevent any polymorphic crystal transformation from cellulose I to cellulose II, which can be detrimental for the mechanical properties. Scanning electron microscopy and thermogravimetric analysis reveals that with increasing chemical treatment, morphological and thermal stability of the processed films are also improved.     

Association of a Genetic Variant in the Cyclin-Dependent Kinase Inhibitor 2B with Risk of Pancreatic Cancer

Background:Pancreatic cancer (PC) is among the most aggressive tumors with a poor prognosis, indicating the need for the identification of a novel prognostic biomarker for risk stratifications. Recent genome-wide association studies have demonstrated common genetic variants in a region on chromosome 9p21 associated with an increased risk of different malignancies.Methods:In the present study, we explore the possible relationship between genetic variant, rs10811661, and gene expression of CDKN2B in 75 pancreatic cancer patients, and 188 healthy individuals. DNAs were extracted and genotyping and gene expression were performed by TaqMan real-time PCR and RT-PCR, respectively. Logistic regression was used to assess the association between risk and genotypes, while the significant prognostic variables in the univariate analysis were included in multivariate analyses.Results:The patients with PDAC had a higher frequency of a TT genotype for rs10811661 than the control group. Also, PDAC patients with dominant genetic model, (TT + TC), was associated with increased risk of developing PDAC (OR= 14.71, 95% CI [1.96-110.35], p= 0.009). Moreover, patients with CC genotype had a higher expression of CDKN2B, in comparison with TT genotype.Conclusion:Our findings demonstrated that CDKN2A/B was associated with the risk of developing PDAC, supporting further investigations in the larger and multicenter setting to validate the potential value of this gene as an emerging marker for PDAC. Key Words: CDKN2A/B, Rs10811661, Pancreatic cancer, Prognostic biomarker     

Multivariate analysis of digital gene expression profiles identifies a xylem signature of the vascular tissue of white spruce (Picea glauca)

A collection of cDNA libraries from white spruce (Picea glauca) and interior spruce (P. glauca × engelmanii) vascular tissue were analyzed to identify a set of genes that could serve as tissue-related markers within the coniferous vascular system. Multivariate exploratory methods identified up to 128 genes co-expressed similarly among three xylem libraries. The majority (87) of these genes formed three distinctive meta-clusters, denoting putative gene cliques in xylem tissue. Of the selected genes, 33 (25%) exhibited no significant sequence homology in queries against any public databases, indicating the possibility of their unique expression in the xylem tissue of conifers. Another 38 genes (30%) had ambiguous annotation. Validation of the annotated genes with analog data, obtained from a wet-lab study utilizing microarray slides with 18,881 spots, resulted in a screened list of 29 genes as xylem-related markers. Response to stress was the predominant category to which the screened genes corresponded. Among the screened genes, elements of the phenolics biosynthesis, cinnamyl alcohol dehydrogenase and laccase, together with the fundamental enzyme of the cell wall biosynthesis, cellulose synthase, prominently delineated characteristics of the wood-forming tissue, xylem.     

Synthesis of some 3-hydroxynaphthalene-2-carbonylamino acid and dipeptide derivatives

Synthesis of a series of 3-hydroxynaphthalene-2-carbonylamino acid methyl esters (II-XI) and some of their corresponding hydrazides (XII-XXI), dipeptide methyl esters (XXII-XXXV) and dipeptide hydrazides (XXXVI-XXXIX) is described. 3-Hydroxynaphthalene-2-CO-L-Tyr-N2H3 (XX) and the corresponding L-Val-L-Ala-N2H3 (XXXVI) were found to be active against a number of micro-organisms.