Dose-response characteristics of glomerular activity in the moth antennal lobe

Odours are represented as unique combinations of activated glomeruli in the antennal lobes of insects. Receptor neurons arborizing in the glomeruli are not only qualitatively selective, but in addition respond to variations in stimulus concentration. As each glomerulus likely represents a single receptor neuron type, optical recordings of calcium changes in insect antennal lobes show how concentration variations affect a large population of afferents. We measured the glomerular responses in the moth Spodoptera littoralis to different concentrations of plant-related odorants. Localized calcium responses were shown to correspond to individual glomeruli. We found that the dynamic range of glomerular responses spanned 3-4 log units of concentration and the most strongly responding glomeruli often reached a plateau at high stimulus doses. Further, we showed that the single most active glomerulus was often not the same across concentrations. However, if the principal glomerulus moved, it was generally to an adjacent or proximal glomerulus. As concentration increased, a higher number of glomeruli became activated. Correlations of glomerular representations of the same compound at different doses decreased as the difference in concentration increased. Moreover, representations evoked by different odorants were more correlated at high than at low doses, which means that the uniqueness of activity patterns decreased with increasing concentration. Thus, if odours are coded as spatial patterns of glomerular activity, as has been suggested, these olfactory codes are not persistent across concentrations.     

A two [4Fe-4S]-cluster-containing ferredoxin as an alternative electron donor for 2-hydroxyglutaryl-CoA dehydratase from<Emphasis Type="Italic"> Acidaminococcus fermentans</Emphasis>

The key step in the fermentation of glutamate by Acidaminococcus fermentans is a reversible syn-elimination of water from (R)-2-hydroxyglutaryl-CoA to (E)-glutaconyl-CoA catalyzed by 2-hydroxyglutaryl-CoA dehydratase, a two-component enzyme system. The actual dehydration is mediated by component D, which contains 1.0 [4Fe-4S]²⁺ cluster, 1.0 reduced riboflavin-5′-phosphate and about 0.1 molybdenum (VI) per heterodimer. The enzyme has to be activated by the extremely oxygen-sensitive [4Fe-4S]^1+/2+-cluster-containing homodimeric component A, which generates Mo(V) by an ATP/Mg²⁺-induced one-electron transfer. Previous experiments established that the hydroquinone state of a flavodoxin (m=14.6 kDa) isolated from A. fermentans served as one-electron donor of component A, whereby the blue semiquinone is formed. Here we describe the isolation and characterization of an alternative electron donor from the same organism, a two [4Fe-4S]^1+/2+-cluster-containing ferredoxin (m=5.6 kDa) closely related to that from Clostridium acidiurici. The protein was purified to homogeneity and almost completely sequenced; the magnetically interacting [4Fe-4S] clusters were characterized by EPR and Mössbauer spectroscopy. The redox potentials of the ferredoxin were determined as ?405 mV and ?340 mV. Growth experiments with A. fermentans in the presence of different iron concentrations in the medium (7–45 μM) showed that flavodoxin is the dominant electron donor protein under iron-limiting conditions. Its concentration continuously decreased from 3.5 μmol/g protein at 7 μM Fe to 0.02 μmol/g at 45 μM Fe. In contrast, the concentration of ferredoxin increased stepwise from about 0.2 μmol/g at 7–13 μM Fe to 1.1±0.1 μmol/g at 17–45 μM Fe.     

The frequency of gene targeting in Trypanosoma brucei is independent of target site copy number

We have investigated the effect of target copy number on the efficiency of stable transformation of the protozoan parasite Trypanosoma brucei. Using a single strain of the organism, we targeted integrative vectors to several different genomic sequences, occurring at copy numbers ranging from 1 to ~30 000 per diploid genome, and undertook a systematic assessment of both transformation and integration efficiencies. Even over this vast copy number range, frequency of gene targeting was the same for all sites. An independence of targeting frequency and target copy number is characteristic of mammalian homologous recombination and is unlike the situation in budding yeast. It is also not seen in the related parasite Leishmania, a distinction that may be the consequence of the different usage of recombination within the mechanisms of pathogenicity in the two species.     

Neopterin and 7,8-dihydroneopterin interfere with low density lipoprotein oxidation mediated by peroxynitrite and/or copper

Low density lipoprotein (LDL) oxidation within the artery wall likely represents a key event in the formation of atherosclerotic lesions. Oxidatively modified LDL particles exert chemotactic properties on macrophages, and the uncontrolled uptake of modified LDL by macrophages leads to the formation of lipid-loaded foam cells, a hallmark of early stage atherosclerosis. Human macrophages stimulated by interferon- γgenerate reactive oxygen species (ROS), neopterin, and 7,8-dihydroneopterin. Higher concentrations of neopterin were found in atherosclerosis, and earlier studies have provided evidence that these neopterin derivatives are able to interfere with reactive species. We therefore investigated whether they also modulate LDL oxidation mediated by Cu(II) and/or peroxynitrite (ONOO -). By means of UV-absorption recording the formation of conjugated dienes in the course of lipid oxidation as well as by measuring the relative electrophoretic mobility of oxidized LDL, we found that neopterin is capable of enhancing ONOO -- as well as Cu(II)-mediated LDL oxidation, whereas 7,8-dihydroneopterin mainly protects LDL from oxidation. However, in case of Cu(II)-mediated LDL oxidation, an initial prooxidative effect of 7,8-dihydroneopterin could be observed. We hypothesize that 7,8-dihydroneopterin may chemically reduce Cu(II) to Cu(I) thereby increasing its oxidative capacity. After total reduction of Cu(II), excess 7,8-dihydroneopterin may block the oxidative potential of Cu(I) and thus decrease the oxidation of LDL. These findings confirm the general behavior of pteridines in redox processes and suggest an in vivo contribution to the process of LDL oxidation.     

An arginine to lysine mutation in the vicinity of the heme propionates affects the redox potentials of the hemes and associated electron and proton transfer in cytochrome c oxidase

Cytochrome c oxidase pumps protons across a membrane using energy from electron transfer and reduction of oxygen to water. It is postulated that an element of the energy transduction mechanism is the movement of protons to the vicinity of the hemes upon reduction, to favor charge neutrality. Possible sites on which protons could reside, in addition to the conserved carboxylate E286, are the propionate groups of heme a and/or heme a(3). A highly conserved pair of arginines (R481 and R482) interact with these propionates through ionic and hydrogen bonds. This study shows that the conservative mutant, R481K, although as fully active as the wild type under many conditions, exhibits a significant decrease in the midpoint redox potential of heme a relative to Cu(A) (DeltaE(m)) of approximately equal 40 mV, has lowered activity under conditions of high pH or in the presence of a membrane potential, and has a slowed heme a(3) reduction with dithionite. Another mutant, D132A, which strongly inhibits proton uptake from the internal side of the membrane, has <4% of the activity of the wild type and appears to be dependent on proton uptake from the outside. A double mutation, D132A/R481K, is even more strongly inhibited ( approximately 1% of that of the wild type). The more-than-additive effect supports the concept that R481K not only lowers the midpoint potential of heme a but also limits a supply route for protons from the outside of the membrane used by the D132 mutant. The results are consistent with an important role of R481 and heme a/a(3) propionates in proton movement in a reversible exit path.     

Multilocus analysis of hypertension: a hierarchical approach

While hypertension is a complex disease with a well-documented genetic component, genetic studies often fail to replicate findings. One possibility for such inconsistency is that the underlying genetics of hypertension is not based on single genes of major effect, but on interactions among genes. To test this hypothesis, we studied both single locus and multilocus effects, using a case-control design of subjects from Ghana. Thirteen polymorphisms in eight candidate genes were studied. Each candidate gene has been shown to play a physiological role in blood pressure regulation and affects one of four pathways that modulate blood pressure: vasoconstriction (angiotensinogen, angiotensin converting enzyme - ACE, angiotensin II receptor), nitric oxide (NO) dependent and NO independent vasodilation pathways and sodium balance (G protein-coupled receptor kinase, GRK4). We evaluated single site allelic and genotypic associations, multilocus genotype equilibrium and multilocus genotype associations, using multifactor dimensionality reduction (MDR). For MDR, we performed systematic reanalysis of the data to address the role of various physiological pathways. We found no significant single site associations, but the hypertensive class deviated significantly from genotype equilibrium in more than 25% of all multilocus comparisons (2,162 of 8,178), whereas the normotensive class rarely did (11 of 8,178). The MDR analysis identified a two-locus model including ACE and GRK4 that successfully predicted blood pressure phenotype 70.5% of the time. Thus, our data indicate epistatic interactions play a major role in hypertension susceptibility. Our data also support a model where multiple pathways need to be affected in order to predispose to hypertension.     

Ecology of antarctic marine sponges: an overview

Sponges are important components of marine benthic communitiesof Antarctica. Numbers of species are high, within the lowerrange for tropical latitudes, similar to those in the Arctic,and comparable or higher than those of temperate marine environments.Many have circumpolar distributions and in some habitats hexactinellidsdominate benthic biomass. Antarctic sponge assemblages contributeconsiderable structural heterogeneity for colonizing epibionts.They also represent a significant source of nutrients to prospectivepredators, including a suite of spongivorous sea stars whoseselective foraging behaviors have important ramifications uponcommunity structure. The highly seasonal plankton blooms thattypify the Antarctic continental shelf are paradoxical whenconsidering the planktivorous diets of sponges. Throughout muchof the year Antarctic sponges must either exploit alternatesources of nutrition such as dissolved organic carbon or bephysiologically adapted to withstand resource constraints. Incontrast to predictions that global patterns of predation shouldselect for an inverse correlation between latitude and chemicaldefenses in marine sponges, such defenses are not uncommon inAntarctic sponges. Some species sequester their defensive metabolitesin the outermost layers where they are optimally effective againstsea star predation. Secondary metabolites have also been shownto short-circuit molting in sponge-feeding amphipods and preventfouling by diatoms. Coloration in Antarctic sponges may be theresult of relict pigments originally selected for aposematismor UV screens yet conserved because of their defensive properties.This hypothesis is supported by the bioactive properties ofpigments examined to date in a suite of common Antarctic sponges.     

Synaptotagmins are trafficked to distinct subcellular domains including the postsynaptic compartment

The synaptotagmin family has been implicated in calcium-dependent neurotransmitter release, although Synaptotagmin 1 is the only isoform demonstrated to control synaptic vesicle fusion. Here, we report the characterization of the six remaining synaptotagmin isoforms encoded in the Drosophila genome, including homologues of mammalian Synaptotagmins 4, 7, 12, and 14. Like Synaptotagmin 1, Synaptotagmin 4 is ubiquitously present at synapses, but localizes to the postsynaptic compartment. The remaining isoforms were not found at synapses (Synaptotagmin 7), expressed at very low levels (Synaptotagmins 12 and 14), or in subsets of putative neurosecretory cells (Synaptotagmins alpha and beta). Consistent with their distinct localizations, overexpression of Synaptotagmin 4 or 7 cannot functionally substitute for the loss of Synaptotagmin 1 in synaptic transmission. Our results indicate that synaptotagmins are differentially distributed to unique subcellular compartments. In addition, the identification of a postsynaptic synaptotagmin suggests calcium-dependent membrane-trafficking functions on both sides of the synapse.