Interaction of ethidium bromide with the transport system for monovalent cations in yeast

Summary Ethidium was found to be taken up by yeast cells in a process that, at certain concentrations has the main following characteristics: a) a substrate is required; b) it presents cooperative kinetics, withn, according to the Hill equation 3; c) ethidium can be concentrated more than 100-fold; d) the uptake is inhibited by Ca²⁺; e) the uptake of the dye is inhibited by monovalent cations with a selectivity pattern similar to that observed in their transport by yeast; f) ethidium inhibits the uptake of K⁺, and, at concentrations up to about 250 m produces a competitive inhibition on the uptake of Rb⁺; and g) ethidium produces the same effects as K⁺ on respiration and the extrusion of H⁺. It is concluded that ethidium is taken up by yeast cells in a selective way by the same transport system normally employed for monovalent cation uptake.     

Assessment of Real-time PCR Based Methods for Quantification of Pollen-mediated Gene Flow from GM to Conventional Maize in a Field Study

Maize is one of the main crops worldwide and an increasing number of genetically modified (GM) maize varieties are cultivated and commercialized in many countries in parallel to conventional crops. Given the labeling rules established e.g. in the European Union and the necessary coexistence between GM and non-GM crops, it is important to determine the extent of pollen dissemination from transgenic maize to other cultivars under field conditions. The most widely used methods for quantitative detection of GMO are based on real-time PCR, which implies the results are expressed in genome percentages (in contrast to seed or grain percentages). Our objective was to assess the accuracy of real-time PCR based assays to accurately quantify the contents of transgenic grains in non-GM fields in comparison with the real cross-fertilization rate as determined by phenotypical analysis. We performed this study in a region where both GM and conventional maize are normally cultivated and used the predominant transgenic maize Mon810 in combination with a conventional maize variety which displays the characteristic of white grains (therefore allowing cross-pollination quantification as percentage of yellow grains). Our results indicated an excellent correlation between real-time PCR results and number of cross-fertilized grains at Mon810 levels of 0.1–10%. In contrast, Mon810 percentage estimated by weight of grains produced less accurate results. Finally, we present and discuss the pattern of pollen-mediated gene flow from GM to conventional maize in an example case under field conditions.     

Butterfly diversity at the ecotone between agricultural and semi‐natural habitats across a climatic gradient

Aim Understanding the response of species to ecotones and habitat edges is essential to designing conservation management, especially in mosaic agricultural landscapes. This study examines how species diversity and composition change with distance from semi‐natural habitats, over ecotones into agricultural fields, and how within‐site patterns of community transition change across a climatic gradient and differ between crop types. Location A total of 19 sites in Israel where semi‐natural habitats border agricultural fields (wheat fields or olive groves) distributed along a sharp climatic gradient ranging between 100 and 800 mm mean annual rainfall. Methods  We performed butterfly surveys in 2006. We analysed species richness (α‐diversity), diversity, community nestedness and species turnover (β‐diversity) within sites and between sites (γ‐diversity). We also assessed where species of conservation concern occurred. Results In wheat sites, richness and diversity declined abruptly from ecotones to fields and remained homogenously poor throughout the fields, regardless of climate. In olive sites, despite the sharp structural boundary, richness and diversity remained high from the semi‐natural habitat to the grove margins and then declined gradually into groves. Species of conservation concern occurred across all habitats at olive sites, but none were found inside wheat fields or at their ecotones. The contrast in community structure between semi‐natural habitats and fields was affected by both climate and field type. Irrigation in arid regions did not augment species diversity. Main conclusions Our results indicate that consideration of crop type, within a climatic context, should receive high priority in biodiversity conservation in agricultural areas. In ‘hostile’ crops, such as wheat, we suggest favouring a combination of high‐intensity management and wide margins over less intensive management without margins, which may merely aid generalist butterfly species. The scarcity of butterflies in arid irrigated fields suggests a need to carefully assess the effects of irrigation and agrochemicals on species’ communities.     

Early metabolic effects and mechanism of ammonium transport in yeast

Studies were performed to define the effects and mechanism of NH+4 transport in yeast. The following results were obtained. Glucose was a better facilitator than ethanol-H2O2 for ammonium transport; low concentrations of uncouplers or respiratory inhibitors could inhibit the transport with ethanol as the substrate. With glucose, respiratory inhibitors showed only small inhibitory effects, and only high concentrations of azide or trifluoromethoxy carbonylcyanide phenylhydrazone could inhibit ammonium transport. Ammonium in the free state could be concentrated approximately 200-fold by the cells. Also, the addition of ammonium produced stimulation of both respiration and fermentation; an increased rate of H+ extrusion and an alkalinization of the interior of the cell; a decrease of the membrane potential, as monitored by fluorescent cyanine; an immediate decrease of the levels of ATP and an increase of ADP, which may account for the stimulation of both fermentation and respiration; and an increase of the levels of inorganic phosphate. Ammonium was found to inhibit 86Rb+ transport much less than K+. Also, while K+ produced a competitive type of inhibition, that produced by NH4+ was of the noncompetitive type. From the distribution ratio of ammonium and the pH gradient, an electrochemical potential gradient of around -180 mV was calculated. The results indicate that ammonium is transported in yeast by a mechanism similar to that of monovalent alkaline cations, driven by a membrane potential. The immediate metabolic effects of this cation seem to be due to an increased [H+]ATPase, to which its transport is coupled. However, the carriers seem to be different. The transport system studied in this work was that of low affinity.     

Alkylation damage in DNA and RNA--repair mechanisms and medical significance

Alkylation lesions in DNA and RNA result from endogenous compounds, environmental agents and alkylating drugs. Simple methylating agents, e.g. methylnitrosourea, tobacco-specific nitrosamines and drugs like temozolomide or streptozotocin, form adducts at N- and O-atoms in DNA bases. These lesions are mainly repaired by direct base repair, base excision repair, and to some extent by nucleotide excision repair (NER). The identified carcinogenicity of O(6)-methylguanine (O(6)-meG) is largely caused by its miscoding properties. Mutations from this lesion are prevented by O(6)-alkylG-DNA alkyltransferase (MGMT or AGT) that repairs the base in one step. However, the genotoxicity and cytotoxicity of O(6)-meG is mainly due to recognition of O(6)-meG/T (or C) mispairs by the mismatch repair system (MMR) and induction of futile repair cycles, eventually resulting in cytotoxic double-strand breaks. Therefore, inactivation of the MMR system in an AGT-defective background causes resistance to the killing effects of O(6)-alkylating agents, but not to the mutagenic effect. Bifunctional alkylating agents, such as chlorambucil or carmustine (BCNU), are commonly used anti-cancer drugs. DNA lesions caused by these agents are complex and require complex repair mechanisms. Thus, primary chloroethyl adducts at O(6)-G are repaired by AGT, while the secondary highly cytotoxic interstrand cross-links (ICLs) require nucleotide excision repair factors (e.g. XPF-ERCC1) for incision and homologous recombination to complete repair. Recently, Escherichia coli protein AlkB and human homologues were shown to be oxidative demethylases that repair cytotoxic 1-methyladenine (1-meA) and 3-methylcytosine (3-meC) residues. Numerous AlkB homologues are found in viruses, bacteria and eukaryotes, including eight human homologues (hABH1-8). These have distinct locations in subcellular compartments and their functions are only starting to become understood. Surprisingly, AlkB and hABH3 also repair RNA. An evaluation of the biological effects of environmental mutagens, as well as understanding the mechanism of action and resistance to alkylating drugs require a detailed understanding of DNA repair processes.     

Effects of endogenous and exogenous processes on aboveground biomass stocks and dynamics in Andean forests

Tropical forests are paramount in regulating the global carbon cycle due to the storage of large amounts of carbon in their biomass. Using repeat censuses of permanent plots located at 15 sites in the Andes Mountains of northwest Colombia, we evaluate: (1) the relationship between aboveground biomass (AGB) stocks, AGB dynamics (mortality, productivity, and net change), and changes in temperature across a ca. 3000-m elevational gradient (≈?16.1 °C); (2) how AGB mortality and AGB productivity interact to determine net AGB change; and (3) the extent to which either fine-grain (0.04-ha) or coarse-grain (1-ha) processes determine the AGB dynamics of these forests. We did not find a significant relationship between elevation/temperature and biomass stocks. The net AGB sequestered each year by these forests (2.21?±?0.51 Mg ha^?1 year^?1), equivalent to approximately 1.09% of initial AGB, was primarily determined by tree growth. Both forest structural properties and global warming influenced AGB mortality and net change. AGB productivity increases with greater inequality of tree sizes, a pattern characteristic of forest patches recovering from disturbances. Overall, we find that global warming is triggering directional changes in species composition by thermophilization via increased tree mortality of species in the lower portions of their thermal ranges and that the inclusion of small-scale forest structural changes can effectively account for endogenous processes such as changes in forest structure. The inclusion of fine-grain processes in assessments of AGB dynamics could provide additional insights about the effects that ongoing climate change has on the functioning of tropical montane forests.     

A constitutive formulation of vascular tissue mechanics including viscoelasticity and softening behaviour

Nearly all soft tissues, among which the vascular tissue is included, present a certain degree of viscoelastic response. This behaviour may be attributed in part to fluid transport within the solid matrix, and to the friction between its fluid and solid constituents. After being preconditioned, the tissue displays highly repetitive behaviour, so that it can be considered pseudo-elastic, that is, elastic but behaving differently in loading and unloading. Because of this reason, very few constitutive laws accounting for the viscoelastic behaviour of the tissue have been developed. Nevertheless, the consideration of this inelastic effect is of crucial importance in surgeries—like vascular angioplasty—where the mentioned preconditioning cannot be considered since non-physiological deformation is applied on the vessel which, in addition, can cause damage to the tissue. A new constitutive formulation considering the particular features of the vascular tissue, such as anisotropy, together with these two inelastic phenomena is presented here and used to fit experimental stress–stretch curves from simple tension loading–unloading tests and relaxation test on porcine and ovine vascular samples.     

Effect of small doses of roentgen radiation on the spontaneous impulse activity of the hippocampus in vitro

X-Irradiation of rat hippocampus in vitro with low doses accelerated spontaneous impulse passage without concomitant changes in synaptic activity. There was a negative correlation between the original frequency of neuron discharges and the degree of quickening the impulses in response to the effect of radiation. Perfusion of slices by a noncalcium solution blocked the synaptic transmission but did not influence the response to the effect of ionizing radiation.