Effect of temperature on the pathways of NADH-oxidation in broad-bean mitochondria

1. Respiration rates of broad-bean (Vicia faba) mitochondria were studied as a function of temperature. Arrhenius plots of all membrane-bound enzymes, as obtained with saturating substrate concentrations, revealed a break in the lower temperature range. That break was considered to indicate a phase transition of membrane phospholipids, characteristic for chilling-sensitive plants. A second discontinuity at 30°C occurred only with activities linked to energy conservation. — 2. The activation energies for the oxidation of NAD⁺-linked substrates differ between states 3 and 4. State 3 respiration of NAD⁺-linked substrates is the result a superimposition of two branches of electron transport, which can be separated by different sensibilities to rotenone. A characteristic temperature dependency of the respiratory control, as well as a shift of the low temperature break in the Arrhenius plot toward a higher temperature after state 4 to state 3 transition, are calculated to be caused by the superimposition of the two branches. — 3. The temperature dependency of the oxidation of extra-mitochondrial NADH and of succinate differs remarkably from that of the oxidation of matrix-NADH. It has been concluded that the rotenone-resistant oxidation of matrix-NADH and the oxidation of external NADH are mediated via different pathways with individual regulation sites.Abbreviations BSA bovine serum albumin - CCCP carbonylcyanide-m-chlorophenylhydrazone - TPP thiaminepyrophosphate     

Heavy metal tolerance in Festuca ovina L. from contaminated sites in the Eifel Mountains,Germany

Extremely high degrees of lead tolerance, measured by comparing rates of root extension in culture solutions, are reported from populations of Festuca ovina growing at two lead-mining sites (Westschacht and Keldenich-II) near Mechernich in the Eifel Mountains, Germany. Other populations from nearby heavy metal-contaminated areas show a considerably smaller degree of lead tolerance. Samples of Festuca ovina collected in the field at Westschacht and Keldenich-II contain higher levels of lead in their aerial organs than do those from other lead-contaminated sites. The main soil factor determining the high degree of lead tolerance is the high Pb/Ca ratio. Populations from soils with a low Pb/Ca ratio display a very low degree of tolerance. It is therefore concluded that in Westschacht and Keldenich-II plants, a genuine intracellular tolerance mechanism is present, allowing the accumulation of lead in aerial organs.Leaf samples of zinc-tolerant Festuca contain higher levels of zinc than do samples of non-tolerant plants. Lead and zinc amounts in leaves are correlated with the soil ratios of Pb/Ca and Zn/Ca, respectively, rather than with the absolute soil-metal levels.In a slightly lead-tolerant, but highly zinc-tolerant clone of Festuca ovina from a site contaminated with large amounts of lead and zinc (Plombières), lead was found to be the major factor affecting the inhibition of root extension with combined treatments of lead and zinc in culture solutions. In the highly lead-tolerant, zinc-sensitive population from Westschacht, zinc governs the response of root growth to combinations of the two metals. The results are discussed in terms of discriminating distinct types of heavy-metal tolerance.     

Correction to: Integrative transcriptomics reveals genotypic impact on sugar beet storability

Plant Molecular Biology - In the above mentioned publication, part of Fig. 6B was distorted (extra diagonal lines appeared). The original article has been corrected and the proper version...     

Galleria mellonella as a model for studying Enterococcus faecium host persistence

Enterococcus faecium is an opportunistic pathogen responsible for numerous outbreaks worldwide. The basis for the colonization capacities, host persistence and environmental stress response of the hospital-adapted clones emerging from E. faecium are poorly understood. In this study, we propose the use of Galleria mellonella as a simple nonmammalian model to assess E. faecium host persistence. Various strains (n = 10), including hospital-adapted, commensal or animal isolates and a SodA-deficient strain were used to assess the relevance of this model. Compared to Enterococcus faecalis, E. faecium strains do not appear very lethal in a Galleria killing assay. The ability of E. faecium strains to overcome host-immune responses and multiply within the host system was evaluated by monitoring bacterial loads following Galleria infection. Among the E. faecium strains, two hospital-adapted isolates displayed increased colonization ability. In contrast, inactivation of sodA, encoding a putative manganese-dependent superoxide dismutase, significantly reduced survival of E. faecium to Galleria defenses. Galleria appears to be a suitable and convenient surrogate model to study E. faecium survival to host defenses and the role of suspected virulence factors in the colonization process.     

Competing ParA Structures Space Bacterial Plasmids Equally over the Nucleoid

Low copy number plasmids in bacteria require segregation for stable inheritance through cell division. This is often achieved by a parABC locus, comprising an ATPase ParA, DNA-binding protein ParB and a parC region, encoding ParB-binding sites. These minimal components space plasmids equally over the nucleoid, yet the underlying mechanism is not understood. Here we investigate a model where ParA-ATP can dynamically associate to the nucleoid and is hydrolyzed by plasmid-associated ParB, thereby creating nucleoid-bound, self-organizing ParA concentration gradients. We show mathematically that differences between competing ParA concentrations on either side of a plasmid can specify regular plasmid positioning. Such positioning can be achieved regardless of the exact mechanism of plasmid movement, including plasmid diffusion with ParA-mediated immobilization or directed plasmid motion induced by ParB/parC-stimulated ParA structure disassembly. However, we find experimentally that parABC from Escherichia coli plasmid pB171 increases plasmid mobility, inconsistent with diffusion/immobilization. Instead our observations favor directed plasmid motion. Our model predicts less oscillatory ParA dynamics than previously believed, a prediction we verify experimentally. We also show that ParA localization and plasmid positioning depend on the underlying nucleoid morphology, indicating that the chromosomal architecture constrains ParA structure formation. Our directed motion model unifies previously contradictory models for plasmid segregation and provides a robust mechanistic basis for self-organized plasmid spacing that may be widely applicable.     

The virtual ecologist approach: simulating data and observers

Ecologists carry a well‐stocked toolbox with a great variety of sampling methods, statistical analyses and modelling tools, and new methods are constantly appearing. Evaluation and optimisation of these methods is crucial to guide methodological choices. Simulating error‐free data or taking high‐quality data to qualify methods is common practice. Here, we emphasise the methodology of the ‘virtual ecologist’ (VE) approach where simulated data and observer models are used to mimic real species and how they are ‘virtually’ observed. This virtual data is then subjected to statistical analyses and modelling, and the results are evaluated against the ‘true’ simulated data. The VE approach is an intuitive and powerful evaluation framework that allows a quality assessment of sampling protocols, analyses and modelling tools. It works under controlled conditions as well as under consideration of confounding factors such as animal movement and biased observer behaviour. In this review, we promote the approach as a rigorous research tool, and demonstrate its capabilities and practical relevance. We explore past uses of VE in different ecological research fields, where it mainly has been used to test and improve sampling regimes as well as for testing and comparing models, for example species distribution models. We discuss its benefits as well as potential limitations, and provide some practical considerations for designing VE studies. Finally, research fields are identified for which the approach could be useful in the future. We conclude that VE could foster the integration of theoretical and empirical work and stimulate work that goes far beyond sampling methods, leading to new questions, theories, and better mechanistic understanding of ecological systems.     

A role for voltage gated T-type calcium channels in mediating "capacitative" calcium entry?

Calcium entry through plasma membrane calcium channels is one of the most important cell signaling mechanism involved in such diverse functions as secretion, contraction and cell growth by regulating gene expression, proliferation and apoptosis. The identity of plasma membrane calcium channels, the main regulators of calcium entry, involved in cell proliferation has been thus extensively sought. Among these, a calcium entry pathway called capacitative calcium entry (CCE), activated by calcium store depletion, is particularly important in non-excitable cells. Though this capacitative calcium entry is generally supposed to occur through TRP channels there is some evidence that voltage-dependent T-type calcium channels may contribute to calcium entry after store depletion. Here we show that though mibefradil, a T-type calcium channel blocker, is able to reduce capacitative calcium entry induced by either thapsigargin or ATP, this was not mimicked by any other T-type calcium channel inhibitors even in cells overexpressing alpha(1H) T-type calcium channels, leading us to conclude that T-type calcium channels are not responsible for the capacitative calcium entry observed in different cancer cell lines. On the contrary, we show that the action of mibefradil on capacitative calcium entry is due to an action on store-operated calcium channels.     

Encapsulation of recombinant <Emphasis Type="Italic">E. coli</Emphasis> expressing cyclopentanone monooxygenase in polyelectrolyte complex capsules for Baeyer–Villiger biooxidation of 8-oxabicyclo[3.2.1]oct-6-en-3-one

Recombinant Escherichia coli cells, over-expressing cyclopentanone monooxygenase activity, were immobilized in polyelectrolyte complex capsules, made of sodium alginate, cellulose sulfate, poly(methylene-co-guanidine), CaCl₂ and NaCl. More than 90% of the cell viability was preserved during the encapsulation process. Moreover, the initial enzyme activity was fully maintained within encapsulated cells while it halved in free cells. Both encapsulated and free cells reached the end point of the Baeyer–Villiger biooxidation of 8-oxabicyclo[3.2.1]oct-6-en-3-one to 4,9-dioxabicyclo[4.2.1]non-7-en-3-one at the same time (48 h). Similarly, the enantiomeric excess above 94% was identical for encapsulated and free cells.