Rapid electrocatalytic procedure for hydrogenase kinetic determination in the H2 evolution direction

The linear sweep voltammetric method is used as a new approach for kinetic determination with enzymes accepting reversible redox couples as cosubstrate. A monolayer of hydrogenase molecules is grafted onto a glassy carbon electrode which is both the support of the enzyme and the detector of the activity. Reduced viologen concentration in the enzyme microenvironment is controlled by the electrode potential. The catalytic current produced by the enzyme allows an easy kinetic constant determination without the classical constraints found in hydrogenase kinetic measurements.     

Membrane chloride transport measured using a chloride-sensitive fluorescent probe

Transport of chloride across cell membranes through exchange, cotransport, or conductive pathways is a subject of great biological importance. Current methods of measurement are restricted in their sensitivity, time resolution, and applicability. A new transport measurement technique has been developed on the basis of the fluorescence quenching by chloride of the dye 6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ). SPQ fluorescence quenching by chloride is rapid (less than 1 ms) and sensitive, with a greater than 50% decrease in fluorescence at 10 mM chloride. SPQ fluorescence is not altered by other physiological anions or by pH and can be used to measure both neutral and conductive transport processes. The high water solubility and membrane permeability properties of SPQ make it ideal for use in both membrane vesicles and cells. Chloride transport determined with SPQ was validated by measurement of erythrocyte chloride/anion exchange and membrane vesicle chloride conductance.     

Cat satellite DNA. Isolation using netropsin with CsCl gradients

The absence of centromeric bands in the karyotype of Felis catus is confirmed. It is also confirmed that no satellite band is visible in CsCl density gradients. However, a satellite is observed both by recentrifuging the fraction of the DNA that bands at high density in CsCl and by using netropsin to enhance the resolution of a CsCl gradient containing total F. catus DNA. The satellite, about 0.5% of total DNA, was isolated by repeated centrifugation in CsCl alone and in CsCl with netropsin. Netropsin was removed and a pure satellite DNA obtained. The reassociation kinetics (C0t1/2 less than 10(-3) M . s) show that the satellite is of the simple sequence type and hence a candidate for centromeric heterochromatin. Its cytological localisation awaits in situ hybridisation experiments.     

The Sodium/Proton Exchanger NHE8 Regulates Late Endosomal Morphology and Function

The pH and lumenal environment of intracellular organelles is considered essential for protein sorting and trafficking through the cell. We provide the first evidence that a mammalian NHE sodium (potassium)/proton exchanger, NHE8, plays a key role in the control of protein trafficking and endosome morphology. At steady state, the majority of epitope-tagged NHE8 was found in the trans-Golgi network of HeLa M-cells, but a proportion was also localized to multivesicular bodies (MVBs). Depletion of NHE8 in HeLa M-cells with siRNA resulted in the perturbation of MVB protein sorting, as shown by an increase in epidermal growth factor degradation. Additionally, NHE8-depleted cells displayed striking perinuclear clustering of endosomes and lysosomes, and there was a ninefold increase in the cellular volume taken up by LAMP1/LBPA-positive, dense MVBs. Our data points to a role for the ion exchange activity of NHE8 being required to maintain endosome morphology, as overexpression of a nonfunctional point mutant protein (NHE8 E225Q) resulted in phenotypes similar to those seen after siRNA depletion of endogenous NHE8. Interestingly, we found that depletion of NHE8, despite its function as a sodium (potassium)/proton antiporter, did not affect the overall pH inside dense MVBs.     

Unraveling the importance of rice fields for waterbird populations in Europe

Rice fields are an alternative habitat for waterbirds and provide food and shelter for many avian species, but there is a lack of information about how the use of rice fields translates into population level effects. The aim of this study was to test the relationship between the use of rice fields by European waterbirds and trends in their populations. We tested this relationship during the autumn migration season and during the breeding season. Based on counts conducted over the last 23 years in natural marshes and areas of rice fields in Doñana (SW Spain), an index of rice field use was constructed for 76 bird species, which was then compared to these species’ European population trends obtained from the literature. A positive relationship was found between waterbird population trends and the use of rice fields during autumn migration season. Our study suggests that changes in the Common Agriculture Policy in Europe leading to reductions in areas of rice cultivation may have important effects on waterbirds. The restoration of former marsh areas and the maintenance of rice cultivation would seem to be more environmentally friendly approaches than the use of these areas to grow alternative crops or solar farms.     

Protective reaction of the myocardium in poisoning

Intensive synthesis of collagen-like substance was revealed in the rabbit myocardium during experimental diphtheria intoxication. It was more marked in the right ventricle 24 hours after the injection of diphtheria toxin. Since similar changes (the substance was mainly formed around blood vessels) have been observed in other cases of toxic myocardial alterations (i.e. ethanol intoxication, injection of pharmacological agents, etc.), it can be assumed that it is a standard protective reaction of the altered heart to the penetration of toxic agents from the blood into the myocardial tissue.     

Influence of exogenous NO donator and variations in the Ca<Superscript>2+</Superscript> content on transport activity related to tonoplast proton pumps in ontogenesis and under hyperosmotic stress

The changes in transport activity of tonoplast proton pumps under the influence of exogenous NO donator and modulation of Ca²⁺ concentration jointly and separately were investigated at different stages of ontogenesis and under hyperosmotic stress. The results suggest that both exogenous NO donator and Ca²⁺ ions can influence the activity of transport processes related to tonoplast and this influence is especially evident in the period of growth and accumulation of metabolites. Under hyperosmotic stress, H⁺-pyrophosphatase plays a more important role than H⁺-ATPase: the activity of the former increases 2.3-fold compared to the control osmotic conditions, whereas the activity of H⁺-ATPase is practically unchanged. H⁺-pyrophosphatase was more responsive to the presence of exogenous NO donator and to variations in Ca²⁺ concentration. The effects of exogenous NO donator on tonoplast proton pumps depended on the concentration of Ca²⁺, which apparently can mediate NO action.