bluuebunny/crossref_metadata_2025 huggingface dataset:
Relationships of intercropped maize, stem borer damage to maize yield and land-use efficiency in the humid forest of Cameroon
Authors: Chabi-Olaye A. , Nolte C. , Schulthess F. , Borgemeister C.
Published: September 2005

Stem borers are the most important maize pests in the humid forest zone of Cameroon. Field trials were conducted in the long and short rainy seasons of 2002 and 2003 to assess the level of damage and yield reductions caused by stem borers in monocropped maize and in maize intercropped with non-host plants such as cassava, cowpea and soybean. The intercrops were planted in two spatial arrangements, i.e. alternating hills or alternating rows. All intercrops and the maize monocrop were grown with and without insecticide treatment for assessment of maize yield loss due to borer attacks. The land-use efficiency of each mixed cropping system was evaluated by comparing it with the monocrop. The temporal fluctuation of larval infestations followed the same pattern in all cropping systems, but at the early stage of plant growth, larval densities were 21.3–48.1% higher in the monocrops than in intercrops, and they tended to be higher in alternating rows than alternating hills arrangements. At harvest, however, pest densities did not significantly vary between treatments. Maize monocrops had 3.0–8.8 times more stems tunnelled and 1.3–3.1 times more cob damage than intercrops. Each percentage increase in stem tunnelling lowered maize grain yield by 1.10 and 1.84 g per plant, respectively, during the long and short rainy season in 2002, and by 5.39 and 1.41 g per plant, respectively, in 2003. Maize yield losses due to stem borer were 1.8–3.0 times higher in monocrops than in intercrops. Intercrops had generally a higher land-use efficiency than monocrops, as indicated by land-equivalent-ratios and area-time-equivalent-ratios of >1.0. Land-use efficiency was similar in both spatial arrangements. At current price levels, the net production of mixed cropping systems was economically superior to controlling stem borers with insecticide in monocropped maize. The maize–cassava intercrop yielded the highest land equivalent ratios and the highest replacement value of the intercrop. At medium intensity cropping this system is thus recommended for land-constrained poor farmers who do not use external inputs such as fertilizer and insecticides.
Role of host feeding niches and host refuges in habitat-related behaviour of Hyssopus pallidus (Hymenoptera: Eulophidae), a larval parasitoid of the codling moth
Authors: Hausmann C. , Mattiacci L. , Dorn S.
Published: September 2005

Parasitoid fitness depends largely on the capability to locate a host in an ecosystem. A parasitoid of a polyphagous host might not be able to find or to access the host in all its feeding niches. This study evaluated the niche selection of Hyssopus pallidus (Askew), a larval parasitoid of Cydia pomonella (Linnaeus), at the plant level with the goal of assessing its potential for biological control on different fruit crops throughout the plant cycle. Parasitoid behaviour during host location and reproduction rate were investigated on host caterpillars actively feeding on apple, pear, apricot or plum, and on caterpillars diapausing under the bark. Under laboratory conditions, the host searching behaviour of H. pallidus varied depending on the fruit species offered and the infestation of the fruits. Parasitoid females searched longer on apples than on other fruit species, and they searched longer on infested than on uninfested apples. Female wasps were able to locate and parasitize host caterpillars under the tree bark, and their behaviour did not vary with host accessibility. The numbers of caterpillars attacked by H. pallidus depended on the fruit species. The highest numbers of caterpillars were parasitized in apples and apricots. Their accessibility (i.e. position) within the fruit or on the branch did not influence parasitism success. Although hosts were parasitized throughout the season, the best results were achieved with early and late releases. Therefore, the host niche selection behaviour of H. pallidus most likely co-evolved with the host C. pomonella on apples, which renders H. pallidus a valuable biocontrol agent for successful release at different times of the season into apple orchards.
Behavioural responses of diamondback mothPlutella xylostella(Lepidoptera: Plutellidae) to extracts derived fromMelia azedarachandAzadirachta indica
Authors: Charleston D.S. , Kfir R. , Vet L.E.M. , Dicke M.
Published: September 2005

The impact of three different doses of botanical insecticide derived from the syringa tree,Melia azedarachand the neem tree,Azadirachta indicawas tested on the behaviour of the diamondback moth,Plutella xylostella(Linnaeus). Both botanical insecticides had a significant impact on larval behaviour. At higher doses the extracts showed feeding deterrent activity, with larvae preferring the untreated sides of cabbage leaves and consuming less of the treated half of cabbage leaves. The botanical insecticides had less of an effect on the oviposition behaviour ofP. xylostellamoths. In laboratory and glasshouse trials, significantly fewer eggs were oviposited on the plants that had been treated with syringa extracts. Therefore, the syringa extracts appear to have a repellent effect. In contrast, when exposed to the neem extracts the moths did not discriminate between control plants and treated plants. Behavioural observation indicated that, despite the lower number of eggs oviposited on cabbage treated with syringa extracts, the moths chose cabbage treated with the highest dose of syringa more often than they chose control cabbage plants. Similar observations were found in cabbage plants treated with neem, moths chose the medium dose more often than they chose the control. Oviposition and feeding deterrent properties are important factors in pest control, and results from this study indicate that botanical insecticides have the potential to be incorporated into control programmes forP. xylostellain South Africa.
Biological control of the coffee berry borerHypothenemus hampei(Coleoptera: Curculionidae) byPhymastichus coffea(Hymenoptera: Eulophidae) in Colombia
Authors: Jaramillo J. , Bustillo A.E. , Montoya E.C. , Borgemeister C.
Published: September 2005

The potential of the eulophid parasitoidPhymastichus coffeaLaSalle to control coffee berry borerHypothenemus hampei(Ferrari) populations under field conditions in Colombia was evaluated. Parasitoid adults were released one, five and nine days after artificial infestations of 90-, 150- and 210-day-old coffee berries withH. hampeifemales. The position of the beetle inside the berry and the parasitism levels were assessed ten days after eachP. coffearelease. Parasitism ofH. hampeibyP. coffeawas significantly affected by the age of the berries at the time of infestation, and by the position of the beetle inside the berries. Highest levels of parasitism were recorded in 150-day-old berries (75–85%) and in 90-day-old berries (75%) whenP. coffeawere released one day after the artificial infestation withH. hampei. In 150-day-old berries, highest levels of parasitism were recorded forH. hampeifound in the outer layer of the endosperm followed by beetles penetrating the exocarp. Increasing the time ofP. coffeareleases after the artificial infestations withH. hampeiled to decreased levels of parasitism in beetles attacking 90- and 150-day-old coffee berries. Low levels of parasitism were recorded inH. hampeifemales infesting older coffee berries because most of the beetles had already constructed galleries deep in the endosperm of the berries, i.e. out of reach of the parasitoid. The potential ofP. coffeafor biological control of coffee berry borer in Colombia is discussed.
Screening of entomopathogenic nematodes for virulence against the invasive western corn rootworm, Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae) in Europe
Authors: Toepfer S. , Gueldenzoph C. , Ehlers R.-U. , Kuhlmann U.
Published: September 2005

Entomopathogenic nematode species available in Europe were screened for their efficacy against both the root-feeding larvae and silk-feeding adults of the western corn rootworm, Diabrotica virgifera virgifera LeConte. Laboratory screening tests were aimed at the selection of candidate biological control agents for the management of this invasive alien pest in Europe. Steinernema glaseri, S. arenarium, S. abassi, S. bicornutum, S. feltiae, S. kraussei, S. carpocapsae and Heterorhabditis bacteriophora were studied to determine their virulence against third instar larvae and adults of D. v. virgifera in small-volume arenas (using nematode concentrations of 0.5, 0.8, 7.9 and 15.9 infective juveniles cm–2). All nematode species were able to invade and propagate in D. v. virgifera larvae, but adults were rarely infected. At concentrations of 7.9 and 15.9 cm–2, S. glaseri, S. arenarium, S. abassi and H. bacteriophora caused the highest larval mortality of up to 77%. Steinernema bicornutum, S. abassi, S. carpocapsae and H. bacteriophora appeared to have a high propagation level, producing 5970±779, 5595±811, 5341±1177 and 4039±1025 infective juveniles per larva, respectively. Steinernema glaseri, S. arenarium, S. feltiae, S. kraussei and H. bacteriophora were further screened at a concentration of 16.7 nematodes cm–2 against third instar larvae in medium-volume arenas (sand-filled trays with maize plants). Heterorhabditis bacteriophora, S. arenarium and S. feltiae caused the highest larval mortality with 77±16.6%, 67±3.5%, and 57±17.1%, respectively. In a next step, criteria for rating the entomopathogenic nematode species were applied based on results obtained for virulence and propagation, and for current production costs and availability in Europe. These criteria were then rated to determine the potential of the nematodes for further field testing. Results showed the highest potential in H. bacteriophora, followed by S. arenarium and S. feltiae, for further testing as candidate biological control agents.
The attack of the clones: tracking the movement of insecticide-resistant peach–potato aphids Myzus persicae (Hemiptera: Aphididae)
Authors: Fenton B. , Malloch G. , Woodford J.A.T. , Foster S.P. , Anstead J. , Denholm I. , King L. , Pickup J.
Published: September 2005

Myzus persicae (Sulzer) collected in Scotland were characterized for four microsatellite loci, intergenic spacer fingerprints and the resistance mechanisms modified acetylcholinesterase (MACE), overproduced carboxylesterase and knockdown resistance (kdr). Microsatellite polymorphisms were used to define a limited number of clones that were either fully susceptible to insecticides or possessed characteristic combinations of resistance mechanisms. Within these clones, intergenic spacer fingerprints could either be very consistent or variable, with the latter indicating ongoing evolution within lineages, most likely derived from the same zygote. Two clones (termed A and B) possessed all three resistance mechanisms and predominated at sites treated with insecticides. Their appearance on seed potatoes and oilseed rape in Scotland in 2001 coincided with extensive insecticide use and severe control failures. Clones C, I and J, with no or fewer resistance mechanisms, were found in samples from 1995 and were dominant at untreated sites in 2001. A comparison of Scottish collections with those from other UK and non-UK sites provides insight into the likely origins, distribution and dynamics of M. persicae clones in a region where asexual (anholocyclic) reproduction predominates, but is vulnerable to migration by novel genotypes from areas of Europe where sexual (holocyclic) reproduction occurs.
Effect of folic acid supplementation on mood and serotonin response in healthy males
Authors: Williams Emma , Stewart-Knox Barbara , Bradbury Ian , Rowland Ian , Pentieva Kristina , Helander Anders , McNulty Helene
Published: September 2005

Evidence suggests that low folate status may be detrimental to mood and associated with depleted cerebrospinal fluid levels of the neurotransmitter serotonin (5-hydroxytryptamine; 5-HT). A placebo-controlled trial was carried out to determine the effect of folic acid supplementation (100 μg for 6 weeks followed by 200 μg for a further 6 weeks) upon subjective mood (Positive and Negative Affect Schedule) and biochemical markers of mood (5-HT) in healthy males (n23). Blood samples were obtained at baseline (week 0) and during the intervention at week 6 and week 12. Subjective mood assessments were obtained at week 0 and week 12. The results showed an increase in serum and erythrocyte folate concentrations (P=0·02 andP=0·003, respectively) and a corresponding decrease in plasma homocysteine (P=0·015) in response to the folic acid intervention. Neither subjective mood nor 5-HT levels, however, were significantly altered in response to the change in folate status. Folic acid given at physiological doses did not appear to improve the mood of healthy folate-replete individuals over a 12-week period. Further research is needed to address the effect of folic acid supplementation or of longer duration or increased dose, particularly in the face of sub-optimal folate status.
Intake of alkylresorcinols from wheat and rye in the United Kingdom and Sweden
Authors: Ross Alastair B. , Becker Wulf , Chen Yan , Kamal-Eldin Afaf , Åman Per
Published: September 2005

Alkylresorcinols, phenolic lipids present in high amounts in whole-grain wheat and rye but not present in appreciable amounts in other foods, are candidates as biomarkers of whole-grain intake from these cereals. We estimated the intake of alkylresorcinols in Sweden and the UK using two different methods: food supply data (FSD); food consumption data (FCD; based on individual and household survey data). The average per capita intake of alkylresorcinols in Sweden was 17·5 mg/d v(FSD) and 22·9 (sd 16·6) mg/d (FCD), while in the UK it was 11·9 mg/d (FSD) and 11·8 (sd 18·62) mg/d (FCD). Ninety-six per cent of all Swedes consumed some alkylresorcinols, compared with 50 % of British people surveyed. Both women and men over the age of 40 years had a higher alkylresorcinol intake than younger people. The average results from the two methods were similar, but the FCD data provided more detail about the range of alkylresorcinol intake, and indicate that because the intake of alkylresorcinols varies so widely, they may be good markers of diets rich or poor in whole-grain wheat/rye products.
Sodium bicarbonated mineral water decreases postprandial lipaemia in postmenopausal women compared to a low mineral water
Authors: Schoppen Stefanie , Pérez-Granados Ana M. , Carbajal Ángeles , Sarriá Beatriz , Sánchez-Muniz Francisco J. , Gómez-Gerique Juan A. , Vaquero M. Pilar
Published: September 2005

The role of bicarbonated mineral waters on lipid metabolism and lipoprotein concentrations in man has scarcely been investigated. The present study aimed to investigate whether drinking sodium bicarbonated mineral water affects postprandial cholesterol and triacylglycerol metabolism in postmenopausal women. In a three-way, randomised, crossover study, eighteen healthy postmenopausal women consumed two sodium bicarbonated mineral waters (bicarbonated mineral water 1 and bicarbonated mineral water 2) and a low mineral water (500 ml of each) with a standard fat-rich meal (4552 kJ; 75·3 g fat). The bicarbonated waters were rich in sodium and bicarbonate and bicarbonated mineral water 1 contained 5·7 times more fluoride than bicarbonated mineral water 2. Fasting blood samples and postprandial blood samples were taken at 30, 60, 120, 240, 360 and 420 min after the end of the meal consumption. Cholesterol and triacylglycerols were determined in serum and chylomicrons. A significant water consumption effect was observed in the total area under the curve (TAUC) of serum and chylomicron triacylglycerols (ANOVA, P=0·008 and P=0·027, respectively). TAUC of serum triacylglycerols for bicarbonated mineral water 2 was significantly lower compared to low mineral water (Bonferroni, P=0·039). Peak concentration of serum triacylglycerols showed a significant water effect (P=0·025). Changes in chylomicron cholesterol were not significantly affected by the type of water. Bicarbonated mineral waters 1 and 2 did not show any significant differences. Drinking sodium bicarbonate-rich mineral waters reduces postprandial lipaemia in healthy postmenopausal women compared to drinking a low mineral water.
Seeds and seasons: interpreting germination timing in the field
Authors: Donohue Kathleen
Published: September 2005

This paper discusses how field and laboratory experiments, using a variety of genetic material, can be combined to investigate the genetic basis of germination under realistic ecological conditions, and it reviews some of our recent work on germination phenology ofArabidopsis thalianain the field. Our results indicate that the genetic basis of germination depends on the environment. In particular, the conditions during seed maturation interact with post-dispersal environmental factors to determine germination phenology, and these interactions have a genetic basis. Therefore genetic studies of germination need to consider carefully the environment – both during seed maturation and after dispersal – in which the experiments are conducted in order to characterize genetic pathways involved with germination in the field. Laboratory studies that explicitly manipulate ecologically relevant environmental factors can be combined with manipulative field studies. These studies can identify the particular environmental cues to which seeds respond in the field and characterize the genetic basis of germination responses to those cues. In addition, a variety of genetic material – including mutant and transgenic lines, intact natural genotypes, recombinant genotypes, and near isogenic lines – can be used in field studies as tools to characterize genetic pathways involved in germination schedules under natural ecological conditions.
Testing for adventitious presence of transgenic material in conventional seed or grain lots using quantitative laboratory methods: statistical procedures and their implementation
Authors: Laffont Jean-Louis , Remund Kirk M. , Wright Deanne , Simpson Robert D. , Grégoire Sylvain
Published: September 2005

When the laboratory methods employed are qualitative, the statistical methodologies used in testing for the adventitious presence (AP) of transgenic material in conventional seed and grain lots are well defined. However, when the response from the method used by the laboratory is quantitative (e.g. percent transgenic DNA), the statistical methodologies developed for qualitative laboratory methods are not fully appropriate. In this paper, we present the details of procedures specific to quantitative laboratory methods. In particular we consider: (1) the assessment of quantitative laboratory method errors using linear modelling; and (2) the process of deciding whether or not a lot meets pre-specified purity standards, including the development of probability calculations needed to develop operating characteristic curves and estimate consumer and producer risks for a given lower quality limit (LQL), acceptable quality limit (AQL) and testing plan. We also describe implementation of this approach in a useful spreadsheet application.
Endo-β-mannanase and β-mannosidase activities in rice grains during and following germination, and the influence of gibberellin and abscisic acid
Authors: Wang Aoxue , Wang Xiaofeng , Ren Yanfang , Gong Xuemei , Bewley J. Derek
Published: September 2005

Grains of indica rice (Oryza sativa cv. Peiza 67) exhibit an increase in endo-β-mannanase activity, mostly after the completion of germination. According to tissue blots, the initial increase occurs in association with the embryo, and possibly the scutellum, although the largest sustained increase in activity is in the peripheral regions of the endosperm. The aleurone layer, being the only living region of the endosperm, is presumably the site of synthesis and secretion of the enzyme into the non-living, starch-laden region. β-Mannosidase activity is low throughout germination and subsequent seedling growth, particularly in the endosperm regions. Its activity profile does not mimic that of endo-β-mannanase. In the intact grain, gibberellin (GA) causes a relatively small increase in endo-β-mannanase activity, while abscisic acid (ABA) causes a large decrease; this inhibition is overcome to a considerable extent when GA is supplied along with ABA. β-Mannosidase activity is little affected by either GA or ABA. Embryoless half-grains imbibed in water exhibit only a small increase in endo-β-mannanase activity with time of imbibition, showing the necessity for a stimulus from the embryo for this to occur. Incubating half-grains in the presence of GA results in a large increase in enzyme activity; ABA reduces the amount of activity compared to the water controls. GA is capable of reversing the inhibitory effect of ABA with respect to endo-β-mannanase activity. As in the intact grains, β-mannosidase activity in the half-grains is unaffected by either GA or ABA. It is concluded that the major site for the production of endo-β-mannanase activity is the aleurone layer, and this event is influenced by the presence of the embryo; in the absence of the latter, the increase in enzyme activity is stimulated by GA. β-Mannosidase activity is low throughout germination and post-germination, it is not influenced by GA and ABA, and thus its activity is not regulated in a coordinated manner with that of endo-β-mannanase.
Seed bank of an invasive alien, Heracleum mantegazzianum, and its seasonal dynamics
Authors: Krinke Lukáš , Moravcová Lenka , Pyšek Petr , Jarošík Vojtěch , Pergl Jan , Perglová Irena
Published: September 2005

The seed bank ofHeracleum mantegazzianum, native to the western Caucasus and invasive in Europe, was studied in the Slavkovský les Protected Landscape Area, the Czech Republic, during the course of two growing seasons. In each of seven study sites with dominating stands of the species, soil samples were taken in October (after the fruit release), April (before germination) and June–July (before the fruit release). Ten samples were taken from each site using a soil hand bore, 4.6 cm in diameter. Seeds elutriated from the samples were immediately germinated in laboratory conditions (10°C, 12 h light/12 h dark); those that germinated during 1 month were considered as non-dormant. Non-germinated but viable seeds, tested for viability by tetrazolium, were considered as dormant. The number of dead seeds was also recorded. The total number of seeds significantly increased with mean density of flowering plants at a site. Of the total variation in seed-bank size, about four-fifths were attributed to that among sites, and one-fifth to that within sites. The number and proportion of living seeds differed significantly between years in summer, but not in autumn and spring. Total number and the numbers of dormant, non-dormant, living and dead seeds significantly differed among the autumn, spring and summer samples. The percentage of living seeds in the total seed bank decreased from 56% in autumn to 42% in spring to 15% in summer. The percentage of non-dormant seeds among those living was 0.3% in autumn, it increased to 87.5% in the spring sample, and decreased to 3% in summer. Pooled across all sites, the average seed numbers (expressed per m2) were 6719±4119 (mean±SD) in autumn, 4907±2278 in spring and 1301±1036 in summer for the total number of seeds, and 3759±2906, 2044±1198 and 192±165, respectively, for living seeds. The majority of seeds (95%) were concentrated in the upper 5 cm soil layer. However, some seeds were present in lower soil layers, which indicates a short-term persistent seed bank. The present data are the first quantitative estimate of the seed bank inH. mantegazzianum, and show that the reproductive potential of the species is enormous, which seems to be a crucial feature of its invasion success.
Induction of Dedifferentiation, Genomewide Transcriptional Programming, and Epigenetic Reprogramming by Extracts of Carcinoma and Embryonic Stem Cells
Authors: Taranger Christel K. , Noer Agate , Sørensen Anita L. , Håkelien Anne-Mari , Boquest Andrew C. , Collas Philippe
Published: September 2005

Functional reprogramming of a differentiated cell toward pluripotency may have long-term applications in regenerative medicine. We report the induction of dedifferentiation, associated with genomewide programming of gene expression and epigenetic reprogramming of an embryonic gene, in epithelial 293T cells treated with an extract of undifferentiated human NCCIT carcinoma cells. 293T cells exposed for 1 h to extract of NCCIT cells, but not of 293T or Jurkat T-cells, form defined colonies that are maintained for at least 23 passages in culture. Microarray and quantitative analyses of gene expression reveal that the transition from a 293T to a pluripotent cell phenotype involves a dynamic up-regulation of hundreds of NCCIT genes, concomitant with down-regulation of 293T genes and of indicators of differentiation such as A-type lamins. Up-regulated genes encompass embryonic and stem cell markers, including OCT4, SOX2, NANOG, and Oct4-responsive genes. OCT4 activation is associated with DNA demethylation in the OCT4 promoter and nuclear targeting of Oct4 protein. In fibroblasts exposed to extract of mouse embryonic stem cells, Oct4 activation is biphasic and RNA-PolII dependent, with the first transient rise of Oct4 up-regulation being necessary for the second, long-term activation of Oct4. Genes characteristic of multilineage differentiation potential are also up-regulated in NCCIT extract-treated cells, suggesting the establishment of “multilineage priming.” Retinoic acid triggers Oct4 down-regulation, de novo activation of A-type lamins, and nestin. Furthermore, the cells can be induced to differentiate toward neurogenic, adipogenic, osteogenic, and endothelial lineages. The data provide a proof-of-concept that an extract of undifferentiated carcinoma cells can elicit differentiation plasticity in an otherwise more developmentally restricted cell type.
Global regulation of X chromosomal genes by the MSL complex in Drosophila melanogaster
Authors: Hamada Fumika N. , Park Peter J. , Gordadze Polina R. , Kuroda Mitzi I.
Published: October 2005

A long-standing model postulates that X-chromosome dosage compensation in Drosophila occurs by twofold up-regulation of the single male X, but previous data cannot exclude an alternative model, in which male autosomes are down-regulated to balance gene expression. To distinguish between the two models, we used RNA interference to deplete Male-Specific Lethal (MSL) complexes from male-like tissue culture cells. We found that expression of many genes from the X chromosome decreased, while expression from the autosomes was largely unchanged. We conclude that the primary role of the MSL complex is to up-regulate the male X chromosome.
Identification of programmed translational -1 frameshifting sites in the genome of Saccharomyces cerevisiae
Authors: Bekaert Michaël , Richard Hugues , Prum Bernard , Rousset Jean-Pierre
Published: October 2005

Frameshifting is a recoding event that allows the expression of two polypeptides from the same mRNA molecule. Most recoding events described so far are used by viruses and transposons to express their replicase protein. The very few number of cellular proteins known to be expressed by a -1 ribosomal frameshifting has been identified by chance. The goal of the present work was to set up a systematic strategy, based on complementary bioinformatics, molecular biology, and functional approaches, without a priori knowledge of the mechanism involved. Two independent methods were devised. The first looks for genomic regions in which two ORFs, each carrying a protein pattern, are in a frameshifted arrangement. The second uses Hidden Markov Models and likelihood in a two-step approach. When this strategy was applied to the Saccharomyces cerevisiae genome, 189 candidate regions were found, of which 58 were further functionally investigated. Twenty-eight of them expressed a full-length mRNA covering the two ORFs, and 11 showed a -1 frameshift efficiency varying from 5% to 13% (50-fold higher than background), some of which corresponds to genes with known functions. From other ascomycetes, four frameshifted ORFs are found fully conserved. Strikingly, most of the candidates do not display a classical viral-like frameshift signal and would have escaped a search based on current models of frameshifting. These results strongly suggest that -1 frameshifting might be more widely distributed than previously thought.
Prediction of aggregation rate and aggregation‐prone segments in polypeptide sequences
Authors: Tartaglia Gian Gaetano , Cavalli Andrea , Pellarin Riccardo , Caflisch Amedeo
Published: September 2005

The reliable identification of β‐aggregating stretches in protein sequences is essential for the development of therapeutic agents for Alzheimer's and Parkinson's diseases, as well as other pathological conditions associated with protein deposition. Here, a model based on physicochemical properties and computational design of β‐aggregating peptide sequences is shown to be able to predict the aggregation rate over a large set of natural polypeptide sequences. Furthermore, the model identifies aggregation‐prone fragments within proteins and predicts the parallel or anti‐parallel β‐sheet organization in fibrils. The model recognizes different β‐aggregating segments in mammalian and nonmammalian prion proteins, providing insights into the species barrier for the transmission of the prion disease.
A mutation designed to alter crystal packing permits structural analysis of a tight‐binding fluorescein–scFv complex
Authors: Honegger Annemarie , Spinelli Silvia , Cambillau Christian , Plückthun Andreas
Published: September 2005

The structure of the scFv fragment FITC‐E2, obtained from a naive phage antibody scFv library derived from human donors, was determined at 2.1 Å resolution in the free form and at 3.0 Å in the complexed form. The wild‐type (wt) scFv binds fluorescein with a KD of 0.75 nM. The free scFv readily crystallizes by compacting its 18 amino acid‐long CDR‐H3, partially occluding the binding site and further blocking access by binding to the ”bottom” of a neighboring scFv molecule with a cluster of exposed aromatic residues within CDR‐H3. Only upon mutating one of the residues involved in this dominant crystal contact, an exposed tryptophan in the middle of CDR‐H3, crystals of the complex could be obtained. A series of alanine mutants within the putative antigen binding site, covering a range of binding affinities, were used to relate macroscopic thermodynamic and kinetic binding parameters to single‐molecule disruption forces measured by AFM. The effects of the mutations on the binding properties, particularly on the fraction of binding‐competent molecules within the population, cannot be fully explained by changes in the strength of local interactions. The significant conformational change of CDR‐H3 between the free and the liganded form illustrates the plasticity of the binding site. An accompanying study in this issue by Curcio and colleagues presents the molecular dynamics simulation of the forced unbinding experiments and explores possible effects of the mutations on the unbinding pathway of the hapten.
Two crystal forms of the restriction enzyme MspI–DNA complex show the same novel structure
Authors: Xu Qian Steven , Roberts Richard J. , Guo Hwai‐Chen
Published: September 2005

The crystal structure of the Type IIP restriction endonuclease MspI bound to DNA containing its cognate recognition sequence has been determined in both monoclinic and orthorhombic space groups. Significantly, these two independent crystal forms present an identical structure of a novel monomer–DNA complex, suggesting a functional role for this novel enzyme–DNA complex. In both crystals, MspI interacts with the CCGG DNA recognition sequence as a monomer, using an asymmetric mode of recognition by two different structural motifs in a single polypeptide. In the crystallographic asymmetric unit, the two DNA molecules in the two MspI–DNA complexes appear to stack with each other forming an end‐to‐end pseudo‐continuous 19‐mer duplex. They are primarily B‐form and no major bends or kinks are observed. For DNA recognition, most of the specific contacts between the enzyme and the DNA are preserved in the orthorhombic structure compared with the monoclinic structure. A cation is observed near the catalytic center in the monoclinic structure at a position homologous to the 74/45 metal site of EcoRV, and the orthorhombic structure also shows signs of this same cation. However, the coordination ligands of the metal are somewhat different from those of the 74/45 metal site of EcoRV. Combined with structural information from other solved structures of Type II restriction enzymes, the possible relationship between the structures of the enzymes and their cleavage behaviors is discussed.
Crystal structures of oxidized and reduced forms of human mitochondrial thioredoxin 2
Authors: Smeets Aude , Evrard Christine , Landtmeters Marie , Marchand Cécile , Knoops Bernard , Declercq Jean‐Paul
Published: September 2005

Mammalian thioredoxin 2 is a mitochondrial isoform of highly evolutionary conserved thioredoxins. Thioredoxins are small ubiquitous protein–disulfide oxidoreductases implicated in a large variety of biological functions. In mammals, thioredoxin 2 is encoded by a nuclear gene and is targeted to mitochondria by a N‐terminal mitochondrial presequence. Recently, mitochondrial thioredoxin 2 was shown to interact with components of the mitochondrial respiratory chain and to play a role in the control of mitochondrial membrane potential, regulating mitochondrial apoptosis signaling pathway. Here we report the first crystal structures of a mammalian mitochondrial thioredoxin 2. Crystal forms of reduced and oxidized human thioredoxin 2 are described at 2.0 and 1.8 Å resolution. Though the folding is rather similar to that of human cytosolic/nuclear thioredoxin 1, important differences are observed during the transition between the oxidized and the reduced states of human thioredoxin 2, compared with human thioredoxin 1. In spite of the absence of the Cys residue implicated in dimer formation in human thioredoxin 1, dimerization still occurs in the crystal structure of human thioredoxin 2, mainly mediated by hydrophobic contacts, and the dimers are associated to form two‐dimensional polymers. Interestingly, the structure of human thioredoxin 2 reveals possible interaction domains with human peroxiredoxin 5, a substrate protein of human thioredoxin 2 in mitochondria.
The leukocidin pore: Evidence for an octamer with four LukF subunits and four LukS subunits alternating around a central axis
Authors: Jayasinghe Lakmal , Bayley Hagan
Published: September 2005

The staphylococcal α‐hemolysin (αHL) and leukocidin (Luk) polypeptides are members of a family of related β‐barrel pore‐forming toxins. Upon binding to susceptible cells, αHL forms water‐filled homoheptameric transmembrane pores. By contrast, Luk pores are formed by two classes of subunit, F and S, rendering a heptameric structure displeasing on symmetry grounds at least. Both the subunit stoichiometry and arrangement within the Luk pore have been contentious issues. Here we use chemical and genetic approaches to show that (1) the predominant, or perhaps the only, form of the Luk pore is an octamer; (2) the subunit stoichiometry is 1:1; and (3) the subunits are arranged in an alternating fashion about a central axis of symmetry, at least when a fused LukS‐LukF construct is used. The experimental approaches we have used also open up new avenues for engineering the arrangement of the subunits of β‐barrel pore‐forming toxins.
Delineation of xenobiotic substrate sites in rat glutathione S‐transferase M1‐1
Authors: Hearne Jennifer L. , Colman Roberta F.
Published: September 2005

Glutathione S‐transferases catalyze the conjugation of glutathione with endogenous and exogenous xenobiotics. Hu and Colman (1995) proposed that there are two distinct substrate sites in rat GST M1‐1, a 1‐chloro‐2,4‐dintrobenzene (CDNB) substrate site located in the vicinity of tyrosine‐115, and a monobromobimane (mBBr) substrate site. To determine whether the mBBr substrate site is distinguishable from the CDNB substrate site, we tested S‐(hydroxyethyl)bimane, a nonreactive derivative of mBBr, for its ability to compete kinetically with the substrates. We find that S‐(hydroxyethyl)bimane is a competitive inhibitor (KI = 0.36 μM) when mBBr is used as substrate, but not when CDNB is used as substrate, demonstrating that these two sites are distinct. Using site‐directed mutagenesis, we have localized the mBBr substrate site to an area midway through α‐helix 4 (residues 90–114) and have identified residues that are important in the enzymatic reaction. Substitution of alanine at positions along α‐helix 4 reveals that mutations at positions 103, 104, and 109 exhibit a greater perturbation of the enzymatic reaction with mBBr than with CDNB as substrate. Various other substitutions at positions 103 and 104 reveal that a hydrophobic residue is necessary at each of these positions to maintain optimal affinity of the enzyme for mBBr and preserve the secondary structure of the enzyme. Substitutions at position 109 indicate that this residue is important in the enzyme's affinity for mBBr but has a minimal effect on Vmax. These results demonstrate that the promiscuity of rat GST M1‐1 is in part due to at least two distinct substrate sites.
Effects of cytochrome P450 3A modulators ketoconazole and carbamazepine on quetiapine pharmacokinetics
Authors: Grimm Scott W. , Richtand Neil M. , Winter Helen R. , Stams Karen R. , Reele Stots B.
Published: September 2005

Aims

To explore the potential for drug interactions on quetiapine pharmacokinetics using in vitro and in vivo assessments.

Methods

The CYP enzymes responsible for quetiapine metabolite formation were assessed using recombinant expressed CYPs and CYP‐selective inhibitors. P‐glycoprotein (Pgp) transport was tested in MDCK cells expressing the human MDR1 gene. The effects of CYP3A4 inhibition were evaluated clinically in 12 healthy volunteers that received 25 mg quetiapine before and after 4 days of treatment with ketoconazole 200 mg daily. To assess CYP3A4 induction in vivo, 18 patients with psychiatric disorders were titrated to steady‐state quetiapine levels (300 mg twice daily), then titrated to 600 mg daily carbamazepine for 2 weeks.

Results

CYP3A4 was found to be responsible for formation of quetiapine sulfoxide and N‐ and O‐desalkylquetiapine and not a Pgp substrate. In the clinical studies, ketoconazole increased mean quetiapine plasma Cmax by 3.35‐fold, from 45 to 150 ng ml−1 (mean Cmax ratio 90% CI 2.51, 4.47) and decreased its clearance (Cl/F) by 84%, from 138 to 22 l h−1 (mean ratio 90% CI 0.13, 0.20). Carbamazepine decreased quetiapine plasma Cmax by 80%, from 1042 to 205 ng ml−1 (mean Cmax ratio 90% CI 0.14, 0.28) and increased its clearance 7.5‐fold, from 65 to 483 l h−1 (mean ratio 90% CI 6.04, 9.28).

Conclusions

Cytochrome P450 3A4 is a primary enzyme responsible for the metabolic clearance of quetiapine. Quetiapine pharmacokinetics were affected by concomitant administration of ketoconazole and carbamazepine, and therefore other drugs and ingested natural products that strongly modulate the activity or expression of CYP3A4 would be predicted to change exposure to quetiapine.
Strikingly higher interleukin (IL)-1α, IL-1β and soluble interleukin-1 receptor antagonist (sIL-1RA) but similar IL-2, sIL-2R, IL-3, IL-4, IL-6, sIL-6R, IL-10, tumour necrosis factor (TNF)-α, transforming growth factor (TGF)-β2 and interferon IFN-γ urine levels in healthy females compared to healthy males: protection against urinary tract injury?
Authors: Sadeghi M , Daniel V , Naujokat C , Weimer R , Opelz G
Published: September 2005

The aim of this prospective study was to examine gender-related differences of cytokines in the plasma and urine of healthy individuals that might provide a clue concerning the lower rate of chronic renal diseases in females. Soluble interleukin-1 receptor antagonist (sIL-1RA), interleukin (IL)-1α, IL-1β, IL-2, sIL-2R, IL-3, IL-4, IL-6, sIL-6R, IL-10, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β2 and interferon (IFN)-γ were determined using standard enzyme-linked immunosorbent assay (ELISA). Cytokine levels were determined in simultaneously obtained plasma and urine samples of 18 male and 28 female healthy members of our laboratory staff. Urine cytokine levels were studied three times at 1-month intervals. All individuals had a negative urine nitrite test and showed no symptoms of urinary tract infection (UTI). Plasma levels of all studied cytokines were similar in males and females (P = n.s.). However, females had significantly higher urine IL-1α (P < 0·0001; P < 0·0001; P < 0·0001) and sIL-1RA (P = 0·0001; P = 0·0003; P = 0·0002) than males at three and higher IL-1β at one of the three investigations (P = 0·098; P = 0·003; P = 0·073). Urine levels of the other cytokines were similar in males and females. Higher urine levels of IL-1α, IL-1β and sIL-1RA in females may result from stimulation of cells in the urinary tract. Increased sIL-1RA might block T lymphocyte activation. The elevated cytokines may play a role in the protection of the female urinary tract from certain renal diseases, such as pyelonephritis and other inflammatory and sclerotic kidney diseases.
Cytokine responses correlate differentially with age in infancy and early childhood
Authors: Härtel C , Adam N , Strunk T , Temming P , Müller-Steinhardt M , Schultz C
Published: September 2005

The functional differentiation of immune cells at early age plays a central role in immune physiology, e.g. for the sufficient eradication of pathogens. However, imbalances in effector cell responses may also have an impact in the pathophysiology of childhood diseases such as atopy and autoimmune disorders. As information on immune cell responses in infancy and early childhood is scarce, we conducted an observational, cross-sectional study in healthy newborns (n = 18), infants and young children (n = 54) aged 1–96 months and adult controls (n = 19) to assess cytokine mRNA and protein expression upon phorbol 12-myristate 13-actate/ionomycin stimulation and LPS-induced IL-12 expression in monocytes. The intracellular expression of interferon (IFN)-γ, tumour necrosis factor (TNF)-α (R = 0·748, P < 0·0001; R = 0·784, P < 0·0001, respectively) and interleukin (IL)-2 protein expression (R = 0·384, P = 0·008) was demonstrated to increase progressively with age. While a correlation between IL-4 protein expression and age was noted (R = 0·342, P = 0·007), the levels of IL-5 and IL-10 protein expression tended to be regulated on an individual basis during infancy and early childhood. An age correlation was also observed for intracellular IL-12 expression (R = 0·331, P = 0·009) in monocytes. These findings are valuable for further assessment of normal variations and maturation processes in immune cell responses and for the clinical–therapeutic monitoring of immunological status in various childhood diseases.
Interferon-α2a is sufficient for promoting dendritic cell immunogenicity
Authors: Tamir A , Jordan W J , Ritter M , Habib N , Lechler R I , Foster G R , Lombardi G
Published: September 2005

Type I interferons (IFNs) are widely used therapeutically. IFN-α2a in particular is used as an antiviral agent, but its immunomodulatory properties are poorly understood. Dendritic cells (DCs) are the only antigen-presenting cells able to prime naive T cells and therefore play a crucial role in initiating the adaptive phase of the immune response. We studied the effects of IFN-α2a on DC maturation and its role in determining Th1/Th2 equilibrium. We found that IFN-α2a induced phenotypic maturation of DCs and increased their allostimulatory capacity. When dendritic cells were stimulated simultaneously by CD40 ligation and IFN-α2a, the production of interleukin (IL)-10 and IL-12 was increased. In contrast, lipopolysaccharide (LPS) stimulation in the presence of IFN-α2a mainly induced IL-10 release. The production of IFN-γ and IL-5 by the responder naive T cells was also amplified in response to IFN-α2a-treated DCs. Furthermore, IL-12 production by IFN-α2a-treated DCs was enhanced further in the presence of anti-IL-10 antibody. Different results were obtained when DCs were treated simultaneously with IFN-α2a and other maturation factors, in particular LPS, and then stimulated by CD40 ligation 36 h later. Under these circumstances, IFN-α2a did not modify the DC phenotype, and the production of IL-10/IL-12 and IFN-γ/IL-5 by DCs and by DC-stimulated naive T cells, respectively, was inhibited compared to the effects on DCs treated with maturation factors alone. Altogether, this work suggests that IFN-α2a in isolation is sufficient to promote DC activation, however, other concomitant events, such as exposure to LPS during a bacterial infection, can inhibit its effects. These results clarify some of the in vivo findings obtained with IFN-α2a and have direct implications for the design of IFN-α-based vaccines for immunotherapy.
Screening of anEchinococcus granulosuscDNA library with IgG4 from patients with cystic echinococcosis identifies a new tegumental protein involved in the immune escape
Authors: Ortona E , Margutti P , Delunardo F , Nobili V , Profumo E , Riganò R , Buttari B , Carulli G , Azzarà A , Teggi A , Bruschi F , Siracusano A
Published: September 2005

The worldwide problem of chronic Echinococcus granulosus disease calls for new parasite-derived immunomodulatory molecules. By screening an E. granulosus cDNA library with IgG4 from patients with active cystic echinococcosis, we identified a cDNA that encodes a predicted partial protein that immunofluorescence studies localized in the protoscolex tegument and on the germinal layer of cyst wall. We named this protein EgTeg because the 105 amino acid sequence scored highest against a family of Schistosoma tegumental proteins. Evaluating the role of EgTeg in the human early inflammatory response we found that EgTeg significantly inhibited polymorphonuclear cell (PMN) chemotaxis. Cytometric analysis of intracellular cytokines disclosed a significantly higher percentage of cells producing IL-4 than IFN-γ (P = 0·001, Student's t-test) in T lymphocytes from patients with cystic echinococcosis stimulated with EgTeg. EgTeg induced weak Th1-dependent proliferation in 42% of patients’ peripheral blood mononuclear cells. In immunoblotting (IB) analysis of total IgG and IgG subclass responses to EgTeg in patients with cystic echinococcosis, patients with other parasitoses, patients with cystic lesions and healthy controls, total IgG specific to EgTeg yielded high sensitivity (73%) but low specificity (44%) precluding its use in immunodiagnosis. Conversely, IgG4 specific to EgTeg gave acceptable sensitivity (65%) and high specificity (89%) suggesting its use in immunodiagnosis to confirm ultrasound documented cysts suggestive of E. granulosus. Because the new tegumental antigen EgTeg inhibits chemotaxis, induces IL-4-positive T lymphocytes and noncomplement fixing antibodies (IgG4) it is an immunomodulatory molecule associated with chronic infection.