Consensus structural features of purified bacterial TatABC complexes

The twin-arginine translocation (Tat) system transports folded proteins across bacterial plasma membranes and the chloroplast thylakoid membrane. Here, we investigate the composition and structural organization of three different purified Tat complexes from Escherichia coli, Salmonella typhimurium and Agrobacterium tumefaciens. First, we demonstrate the functional activity of these Tat systems in vivo, since expression of the tatABC operons from S.typhimurium or A.tumefaciens in an E.coli tat null mutant strain resulted in efficient Tat-dependent export of an E.coli cofactor-containing substrate, TMAO reductase. The three isolated, affinity-tagged Tat complexes comprised TatA, TatB and TatC in each case, demonstrating a strong interaction between these three subunits. Single-particle electron microscopy studies of all three complexes revealed approximately oval-shaped, asymmetric particles with maximal dimensions up to 13 nm. A common feature is a number of stain-excluding densities surrounding more or less central pools of stain, suggesting protein-lined pores or cavities. The characteristics of size variation among the particles suggest a modular form of assembly and/or the recruitment of varying numbers of TatBC/TatA units. Despite low levels of sequence homology, the combined data indicate structural and functional conservation in the Tat systems of these three bacterial species.     

Function of Rx, but not Pax6, is essential for the formation of retinal progenitor cells in mice

Rx plays a critical role in eye formation. Targeted elimination of Rx results in embryos that do not develop eyes. In this study, we have investigated the expression of Otx2, Six3, and Pax6 in Rx deficient embryos. We find that these genes show normal activation in the anterior neural plate in Rx-/- embryos, but they are not upregulated in the area of the neural plate that would form the primordium of the optic vesicle. In contrast, in homozygous Small eye embryos that lack Pax6 function, Rx shows normal activation in the anterior neural plate and normal upregulation in the optic vesicle/retinal progenitor cells. This suggests that neither Rx expression nor the formation of retinal progenitor cells is dependent on a functional copy of the Pax6 gene, but that Pax6 expression and the formation of the progenitor cells of the optic cup is dependent on a functional copy of the Rx gene.     

Polycationic lipids translocate lipopolysaccharide into HeLa cells

We have investigated the ability of LIPOFECTAMINE, a polycationic lipid reagent used in DNA transfection, to translocate E. coli lipopolysaccharide (LPS) into HeLa cells. Although HeLa cells did not spontaneously take up fluorescein isothiocyanate-labelled LPS (FITC-LPS) from the culture medium, the cells that were co-incubated with greater than 1 g/mL FITC-LPS and LIPOFECTAMINE showed punctate fluorescence. Virtually all cells were loaded on incubation with 100 micrograms/mL FITC-LPS. Confocal scanning laser microscopy showed extensive FITC-LPS loading in the cytoplasm of HeLa cells, but no label was evident in the nuclear regions of these cells. Loading with LPS for up to six hours had no effect on the viability of HeLa cells, beyond the 30% reduction in live cells that is attributable to the toxic effect of LIPOFECTAMINE itself. In contrast to cells treated with etoposide for six hours, LPS-loaded cells did not display apoptotic bodies. Exposure of cells to 4 beta-phorbol 12-myristate 13-acetate led to the induction of the immediate early gene c-fos and resulted in an enhanced c-Fos signal, detected by Western blot analysis. In contrast, LPS loading did not alter the c-fos expression in HeLa cells. The loading of LPS into HeLa cells by means of polycationic lipids results in relatively low acute toxicity, as judged from cell viability, morphology and c-fos expression. Therefore, our method appears well suited to the study of acute actions of LPS in the intracellular compartment of mammalian cells.     

The role of fine sediment characteristics and body size on the vertical movement of a freshwater amphipod

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Dietary Patterns and Socioeconomic Status in the Very Old: The Newcastle 85+ Study

BackgroundDietary patterns (DP) are associated with health outcomes in younger adults but there is a lack of evidence in the very old (aged 85+) on DP and their association with sociodemographic factors, lifestyle, health and functioning measures. Higher socioeconomic status (SES) has been linked with healthier DP but it is not known whether these associations are sustained in the very old.ObjectiveWe aimed to (a) characterise DP in the very old and (b) assess the relationships between three SES indicators (education, occupational class and area-deprivation index [IMD]) and DP.MethodsComplete dietary data at baseline (2006/07) for 793 participants in the Newcastle 85+ Study were established through 24-hr multiple pass recall. We used Two-Step clustering and 30 food groups to derive DP, and multinomial logistic regression models to assess the association with SES.ResultsWe identified three distinct DP (characterised as ‘High Red Meat’, ‘Low Meat’, and ‘High Butter’) that varied with key sociodemographic, health and functioning measures. ‘Low Meat’ participants were more advantaged (i.e. higher education and occupational class, and lived in more affluent areas in owned homes), were least disabled, cognitively impaired, and depressed, and were more physically active than those in the other DP. After adjusting for other lifestyle factors, cognitive status and BMI, lower educational attainment remained a significant predictor of ‘High Red Meat’ and ‘High Butter’ membership compared with ‘Low Meat’ (‘High Red Meat’: OR [95% CI] for 0–9 and 10–11 years of education vs. ≥12 years: 5.28 [2.85–9.79], p<0.001 and 3.27 [1.65–6.51], p = 0.001, respectively; ‘High Butter’: 3.32 [1.89–5.82], p<0.001 and 2.83 [1.52–5.28], p = 0.001).ConclusionsIn this cohort of very old adults, we detected a favourable DP (‘Low Meat’), which was associated with better health and functioning and higher SES.     

Human adaptations to meat eating

It is argued that Homo sapiens is a habitual rather than a facultative meat eater. Quantitative similarity of human gut morphology to guts of carnivorous mammals, preferential absorption of haem rather than iron of plant origin, and the exclusive use of humans as the definitive host by Taenia saginata and the almost complete human specificity of T. solium are used to support the argument.     

Expert opinion as 'validation' of risk assessment applied to calf welfare

Background  

Recently, a Risk Assessment methodology was applied to animal welfare issues in a report of the European Food Safety Authority (EFSA) on intensively housed calves.