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The permeation pore of K+ channels is formed by four copies of the pore domain. AtKCO3 is the only putative voltage-independent K+ channel subunit of Arabidopsis thaliana with a single pore domain. KCO3-like proteins recently emerged in evolution and, to date, have been found only in the genus Arabidopsis (A. thaliana and A. lyrata). We show that the absence of KCO3 does not cause marked changes in growth under various conditions. Only under osmotic stress we observed reduced root growth of the kco3-1 null-allele line. This phenotype was complemented by expressing a KCO3 mutant with an inactive pore, indicating that the function of KCO3 under osmotic stress does not depend on its direct ability to transport ions. Constitutively overexpressed AtKCO3 or AtKCO3::G FP are efficiently sorted to the tonoplast indicating that the protein is approved by the endoplasmic reticulum quality control. However, vacuoles isolated from transgenic plants do not have significant alterations in current density. Consistently, both AtKCO3 and AtKCO3::GFP are detected as homodimers upon velocity gradient centrifugation, an assembly state that would not allow for activity. We conclude that if AtKCO3 ever functions as a K+ channel, active tetramers are held by particularly weak interactions, are formed only in unknown specific conditions and may require partner proteins.
As heritage from early evolution, potassium (K+) is absolutely necessary for all living cells. It plays significant roles as stabilizer in metabolism and is important for enzyme activation, stabilization of protein synthesis, and neutralization of negative charges on cellular molecules as proteins and nucleic acids. Land plants even enlarged this spectrum of K+ utilization after having gone ashore, despite the fact that K+ is far less available in their new oligotrophic habitats than in sea water. Inevitably, plant cells had to improve and to develop unique transport systems for K+ accumulation and distribution. In the past two decades a manifold of K+ transporters from flowering plants has been identified at the molecular level. The recently published genome of the fern ally Selaginella moellendorffii now helps in providing a better understanding on the molecular changes involved in the colonization of land and the development of the vasculature and the seeds. In this article we present an inventory of K+ transporters of this lycophyte and pigeonhole them together with their relatives from the moss Physcomitrella patens, the monocotyledon Oryza sativa, and two dicotyledonous species, the herbaceous plant Arabidopsis thaliana, and the tree Populus trichocarpa. Interestingly, the transition of green plants from an aqueous to a dry environment coincides with a dramatic reduction in the diversity of voltage-gated potassium channels followed by a diversification on the basis of one surviving K+ channel class. The first appearance of K+ release (K-out) channels in S. moellendorffii that were shown in Arabidopsis to be involved in xylem loading and guard cell closure coincides with the specialization of vascular plants and may indicate an important adaptive step.
The regulation of protein function by modulating the surface charge status via sequence-locally enriched phosphorylation sites (P-sites) in so called phosphorylation "hotspots" has gained increased attention in recent years. We set out to identify P-hotspots in the model plant Arabidopsis thaliana. We analyzed the spacing of experimentally detected P-sites within peptide-covered regions along Arabidopsis protein sequences as available from the PhosPhAt database. Confirming earlier reports (Schweiger and Lanial, 2010), we found that, indeed, P-sites tend to cluster and that distributions between serine and threonine P-sites to their respected closest next P-site differ significantly from those for tyrosine P-sites. The ability to predict P-hotspots by applying available computational P-site prediction programs that focus on identifying single P-sites was observed to be severely compromised by the inevitable interference of nearby P-sites. We devised a new approach, named HotSPotter, for the prediction of phosphorylation hotspots. HotSPotter is based primarily on local amino acid compositional preferences rather than sequence position-specific motifs and uses support vector machines as the underlying classification engine. HotSPotter correctly identified experimentally determined phosphorylation hotspots in A. thaliana with high accuracy. Applied to the Arabidopsis proteome, HotSPotter-predicted 13,677 candidate P-hotspots in 9,599 proteins corresponding to 7,847 unique genes. Hotspot containing proteins are involved predominantly in signaling processes confirming the surmised modulating role of hotspots in signaling and interaction events. Our study provides new bioinformatics means to identify phosphorylation hotspots and lays the basis for further investigating novel candidate P-hotspots. All phosphorylation hotspot annotations and predictions have been made available as part of the PhosPhAt database at http://phosphat.mpimp-golm.mpg.de.
Glycolate oxidase (GO) catalyses the oxidation of glycolate to glyoxylate, thereby consuming O-2 and producing H2O2. In this work, Arabidopsis thaliana plants expressing GO in the chloroplasts (GO plants) were used to assess the expressional behavior of reactive oxygen species (ROS)-responsive genes and transcription factors (TFs) after metabolic induction of H2O2 formation in chloroplasts. In this organelle, GO uses the glycolate derived from the oxygenase activity of RubisCO. Here, to identify genes responding to an abrupt production of H2O2 in chloroplasts we used quantitative real-time PCR (qRT-PCR) to test the expression of 187 ROS-responsive genes and 1880 TFs after transferring GO and wild-type (WT) plants grown at high CO2 levels to ambient CO2 concentration. Our data revealed coordinated expression changes of genes of specific functional networks 0.5 h after metabolic induction of H2O2 production in GO plants, including the induction of indole glucosinolate and camalexin biosynthesis genes. Comparative analysis using available microarray data suggests that signals for the induction of these genes through H2O2 may originate in the chloroplast. The TF profiling indicated an up-regulation in GO plants of a group of genes involved in the regulation of proanthocyanidin and anthocyanin biosynthesis. Moreover, the upregulation of expression of IF and IF interacting proteins affecting development (e.g., cell division, stem branching, flowering time, flower development) would impact growth and reproductive capacity, resulting in altered development under conditions that promote the formation of H2O2.
Red, orange or green snow is the macroscopic phenomenon comprising different eukaryotic algae. Little is known about the ecology and nutrient regimes in these algal communities. Therefore, eight snow algal communities from five intensively tinted snow fields in western Spitsbergen were analysed for nutrient concentrations and fatty acid (FA) composition. To evaluate the importance of a shift from green to red forms on the FA-variability of the field samples, four snow algal strains were grown under nitrogen replete and moderate light (+N+ML) or N-limited and high light (-N+HL) conditions. All eight field algal communities were dominated by red and orange cysts. Dissolved nutrient concentration of the snow revealed a broad range of NH4+ (<0.005-1.2 mg NI-1) and only low PO43- (< 18 mu g P I-1) levels. The external nutrient concentration did not reflect cellular nutrient ratios as C:N and C:P ratios of the communities were highest at locations containing relatively high concentrations of NH4- and PO43-. Molar N:P ratios ranged from 11 to 21 and did not suggest clear limitation of a single nutrient. On a per carbon basis, we found a 6-fold difference in total FA content between the eight snow algal communities, ranging from 50 to 300 mg FA g C-1. In multivariate analyses total FA content opposed the cellular N:C quota and a large part of the FA variability among field locations originated from the abundant FAs C181n-9, C18 2n-6, and C183n-3. Both field samples and snow algal strains grown under -N+HL conditions had high concentrations of C181n-9. FAs possibly accumulated due to the cessation of growth. Differences in color and nutritional composition between patches of snow algal communities within one snow field were not directly related to nutrient conditions. We propose that the highly patchy distribution of snow algae within and between snow fields may also result from differences in topographical and geological parameters such as slope, melting water rivulets, and rock formation.
Ciliate epibionts associated with crustacean zooplankton are widespread in aquatic systems, but their ecological roles are little known. We studied the occurrence of ciliate epibionts on crustacean zooplankton in nine German lakes with different limnological features during the summer of 2011. We also measured the detachment and re-attachment rates of the ciliates, changes in their motility, and the feeding rates of attached vs. detached ciliate epibionts. Epibionts were found in all lakes sampled except an acidic lake with large humic inputs. Epibiont prevalence was as high as 80.96% on the cladoceran Daphnia cucullata, 67.17% on the cladoceran Diaphanosoma brachyurum, and 46.67% on the calanoid copepod Eudiaptomus gracilis. Both cladoceran groups typically had less than 10 epibionts per individual, while the epibiont load on E. gracilis ranged from 1 to >30 epibionts per individual. After the death of the zooplankton host, the peritrich ciliate epibiont Epistylis sp. detached in an exponential fashion with a half-life of 5 min, and 98% detached within 30 min, leaving behind the stalks used for attachment. Immediately after detachment, the ciliates were immotile, but 62% became motile within 60 min. When a new host was present, only 27% reattached after 120 min. The average measured ingestion rate and clearance rate of Epistylis were 11,745 bacteria ciliate(-1) h(-1) and 24.33 mu l ciliate(-1) h(-1), respectively. Despite their high feeding rates, relatively low epibiont abundances were observed in the field, which suggests either diversion of energy to stalk formation, high metabolic loss by the epibionts, or high mortality among the epibiont populations.
Population fragmentation is often correlated with loss of genetic diversity and reduced fitness. Obligate out-crossing (dioecy) is expected to enhance genetic diversity, reduce genetic differentiation, and avoid inbreeding depression through frequent gene flow. However, in highly fragmented populations dioecy has only diminishing effects upon genetic structure as pollination limitations (e.g. flight distance of pollinators) most often restrict inter-population gene flow in insect pollinated species. In fragmented dry grasslands in northeastern Germany, we analysed genetic structure, fitness, and habitat quality of the endangered dioecious Silene otites (Caryophyllaceae). Using AFLP markers, a high level of differentiation among ten populations was found (F (st) = 0.36), while the intra-population genetic diversities (H (E) = 0.165-0.240) were similar as compared to hermaphroditic species. There was neither a correlation between geographic and genetic distance nor between genetic diversity and population size, which indicates reduced gene flow among populations and random genetic drift. Plant size was positively correlated with genetic diversity. Seed set and number of juveniles were positively related to population size. Higher total coverage resulted in reduced plant fitness, and the number of juveniles was negatively correlated to cryptogam cover. Additionally, we found a sex ratio bias towards more male plants in larger populations. Overall, our results indicate that on a regional geographic scale dioecy does not necessarily prevent genetic erosion in the case of habitat fragmentation, especially in the absence of long distance seed and pollen dispersal capacity.
The tricarboxylic acid (TCA) cycle is a crucial component of respiratory metabolism in both photosynthetic and heterotrophic plant organs. All of the major genes of the tomato TCA cycle have been cloned recently, allowing the generation of a suite of transgenic plants in which the majority of the enzymes in the pathway are progressively decreased. Investigations of these plants have provided an almost complete view of the distribution of control in this important pathway. Our studies suggest that citrate synthase, aconitase, isocitrate dehydrogenase, succinyl CoA ligase, succinate dehydrogenase, fumarase and malate dehydrogenase have control coefficients flux for respiration of -0.4, 0.964, -0.123, 0.0008, 0.289, 0.601 and 1.76, respectively; while 2-oxoglutarate dehydrogenase is estimated to have a control coefficient of 0.786 in potato tubers. These results thus indicate that the control of this pathway is distributed among malate dehydrogenase, aconitase, fumarase, succinate dehydrogenase and 2-oxoglutarate dehydrogenase. The unusual distribution of control estimated here is consistent with specific non-cyclic flux mode and cytosolic bypasses that operate in illuminated leaves. These observations are discussed in the context of known regulatory properties of the enzymes and some illustrative examples of how the pathway responds to environmental change are given.
Syntheses of thiazolidine-fused heterocycles via exo-mode cyclizations of vinylogous N-acyliminium ions incorporating heteroatom-based nucleophiles have been examined and discussed. The formation of (5,6)-membered systems was feasible with all nucleophiles tried (O, S and N), while the closing of the five-membered ring was restricted to O- and S-nucleophiles. The closure of a four-membered ring failed. Instead, the bicyclic (5,6)-membered acetal derivative and the tricyclic system with an eight-membered central ring were obtained from the substrates containing O and S nucleophilic moieties, respectively. The reaction outcome and stereochemistry are rationalized using quantum chemical calculations at B3LYP/6-31G(d) level. The exclusive cis-stereoselectivity in the formation of (5,6)- and (5,5)-membered systems results from thermodynamic control, whereas the formation of the eight-membered ring was kinetically controlled.
Ex situ cultivation in botanic gardens could be one possibility to preserve plant species diversity and genetic variation. However, old ex situ populations are often sparsely documented. We were able to retrieve three different ex situ populations and their source in situ populations of the endangered plant species Silene otites after 20-36 years of isolation. Furthermore, three additional wild populations were included in the analysis. Population genetic diversity and differentiation were analysed using AFLP markers. Genetic variation in the ex situ populations was lower than the variation found in the in situ populations. Strong differentiation (F-ST = 0.21-0.36) between corresponding in situ and ex situ populations was observed. Bayesian clustering approach also showed a distinct genetic separation between in situ and ex situ populations. The high genetic differentiation and loss of genetic diversity during spatial and temporal isolation in the ex situ populations can be attributable to small population sizes and unconscious selection during cultivation. Therefore, adequate sampling prior to ex situ cultivation and large effective population sizes are important to preserve genetic diversity. Near-natural cultivation allowing for generation overlap and interspecific competition without artificial selection is recommended as being best for the maintenance of the genetic constitution.
The wide knowledge gaps in invasion biology research that exist in the developing world are crucial impediments to the scientific management and global policymaking on biological invasions. In an effort to fill such knowledge gaps, we present here an inventory of the alien flora of India, based on systematic reviews and rigorous analyses of research studies (ca. 190) published over the last 120 years (1890-2010 AD), and updated with field records of the last two decades. Currently, the inventory comprises of 1,599 species, belonging to 842 genera in 161 families, and constitutes 8.5% of the total Indian vascular flora. The three most species-rich families are Asteraceae (134 spp.), Papilionaceae (114 spp.) and Poaceae (106 spp.), and the three largest genera are Eucalyptus (25 spp.), Ipomoea (22 spp.), and Senna (21 spp.). The majority of these species (812) have no report of escaping from cultivation. Of the remaining subset of 787 species, which have either escaped from intentional cultivation, or spread after unintentional introduction, casuals are represented by 57 spp., casual/naturalised by 114 spp., naturalised by 257 spp., naturalised/invasive by 134 spp., and invasive by 225 spp. Biogeographically, more than one-third (35%) of the alien flora in India has its native ranges in South America, followed by Asia (21%), Africa (20%), Europe (11%), Australia (8%), North America (4%); and cryptogenic (1%). The inventory is expected to serve as the scientific baseline on plant invasions in India, with implications for conservation of global biodiversity.
The aromatic peroxygenase (APO; EC 1.11.2.1) from the agraric basidomycete Marasmius rotula (MroAPO) immobilized at the chitosan-capped gold-nanoparticle-modified glassy carbon electrode displayed a pair of redox peaks with a midpoint potential of -278.5 mV vs. AgCl/AgCl (1 M KCl) for the Fe(2+)/Fe(3+) redox couple of the heme-thiolate-containing protein. MroAPO oxidizes aromatic substrates such as aniline, p-aminophenol, hydroquinone, resorcinol, catechol, and paracetamol by means of hydrogen peroxide. The substrate spectrum overlaps with those of cytochrome P450s and plant peroxidases which are relevant in environmental analysis and drug monitoring. In M. rotula peroxygenase-based enzyme electrodes, the signal is generated by the reduction of electrode-active reaction products (e.g., p-benzoquinone and p-quinoneimine) with electro-enzymatic recycling of the analyte. In these enzyme electrodes, the signal reflects the conversion of all substrates thus representing an overall parameter in complex media. The performance of these sensors and their further development are discussed.
Are gastropods, rather than ants, important dispersers of seeds of myrmecochorous forest herbs?
(2012)
Seed dispersal by ants (myrmecochory) is widespread, and seed adaptations to myrmecochory are common, especially in the form of fatty appendices (elaiosomes). In a recent study, slugs were identified as seed dispersers of myrmecochores in a central European beech forest. Here we used 105 beech forest sites to test whether myrmecochore presence and abundance is related to ant or gastropod abundance and whether experimentally exposed seeds are removed by gastropods. Myrmecochorous plant cover was positively related to gastropod abundance but was negatively related to ant abundance. Gastropods were responsible for most seed removal and elaiosome damage, whereas insects (and rodents) played minor roles. These gastropod effects on seeds were independent of region or forest management. We suggest that terrestrial gastropods can generally act as seed dispersers of myrmecochorous plants and even substitute myrmecochory, especially where ants are absent or uncommon.
Multidirectional communicative interactions in social networks can have a profound effect on mate choice behavior. Male Atlantic molly Poecilia mexicana exhibit weaker mating preferences when an audience male is presented. This could be a male strategy to reduce sperm competition risk: interacting more equally with different females may be advantageous because rivals might copy mate choice decisions. In line with this hypothesis, a previous study found males to show a strong audience effect when being observed while exercising mate choice, but not when the rival was presented only before the choice tests. Audience effects on mate choice decisions have been quantified in poeciliid fishes using association preference designs, but it remains unknown if patterns found from measuring association times translate into actual mating behavior. Thus, we created five audience treatments simulating different forms of perceived sperm competition risk and determined focal males' mating preferences by scoring pre-mating (nipping) and mating behavior (gonopodial thrusting). Nipping did not reflect the pattern that was found when association preferences were measured, while a very similar pattern was uncovered in thrusting behavior. The strongest response was observed when the audience could eavesdrop on the focal male's behavior. A reduction in the strength of focal males' preferences was also seen after the rival male had an opportunity to mate with the focal male's preferred mate. In comparison, the reduction of mating preferences in response to an audience was greater when measuring association times than actual mating behavior. While measuring direct sexual interactions between the focal male and both stimulus females not only the male's motivational state is reflected but also females' behavior such as avoidance of male sexual harassment.
Nuclear proteins play a central role in regulating gene expression. Their identification is important for understanding how the nuclear repertoire changes over time under different conditions. Nuclear proteins are often underrepresented in proteomic studies due to the frequently low abundance of proteins involved in regulatory processes. So far, only few studies describing the nuclear proteome of plant species have been published. Recently, the genome sequence of the unicellular green alga Chlamydomonas reinhardtii has been obtained and annotated, allowing the development of further detailed studies for this organism. However, a detailed description of its nuclear proteome has not been reported so far. Here, we present an analysis of the nuclear proteome of the sequenced Chlamydomonas strain cc503. Using LC-MS/MS, we identified 672 proteins from nuclei isolates with a maximum 1% peptide spectrum false discovery rate. Besides well-known proteins (e.g. histones), transcription factors and other transcriptional regulators (e.g. tubby and HMG) were identified. The presence of protein motifs in nuclear proteins was investigated by computational tools, and specific over-represented protein motifs were identified. This study provides new insights into the complexity of the nuclear environment and reveals novel putative protein targets for further studies of nuclear mechanisms.
Despite the great agricultural and ecological importance of efficient use of urea-containing nitrogen fertilizers by crops, molecular and physiological identities of urea transport in higher plants have been investigated only in Arabidopsis. We performed short-time urea-influx assays which have identified a low-affinity and high-affinity (Km of 7.55 mu M) transport system for urea-uptake by rice roots (Oryza sativa). A high-affinity urea transporter OsDUR3 from rice was functionally characterized here for the first time among crops. OsDUR3 encodes an integral membrane-protein with 721 amino acid residues and 15 predicted transmembrane domains. Heterologous expression demonstrated that OsDUR3 restored yeast dur3-mutant growth on urea and facilitated urea import with a Km of c. 10 mu M in Xenopus oocytes. Quantitative reverse-transcription polymerase chain reaction (qPCR) analysis revealed upregulation of OsDUR3 in rice roots under nitrogen-deficiency and urea-resupply after nitrogen-starvation. Importantly, overexpression of OsDUR3 complemented the Arabidopsis atdur3-1 mutant, improving growth on low urea and increasing root urea-uptake markedly. Together with its plasma membrane localization detected by green fluorescent protein (GFP)-tagging and with findings that disruption of OsDUR3 by T-DNA reduces rice growth on urea and urea uptake, we suggest that OsDUR3 is an active urea transporter that plays a significant role in effective urea acquisition and utilisation in rice.
In this study we used molecular markers to screen for the occurrence and prevalence of the three most common haemosporidian genera (Haemoproteus, Plasmodium, and Leucocytozoon) in blood samples of the Philippine Bulbul (Hypsipetes philippinus), a thrush-size passerine bird endemic to the Philippine Archipelago. We then used molecular data to ask whether the phylogeographic patterns in this insular host-parasite system might follow similar evolutionary trajectories or not. We took advantage of a previous study describing the pattern of genetic structuring in the Philippine Bulbul across the Central Philippine Archipelago (6 islands, 7 populations and 58 individuals; three mitochondrial DNA genes). The very same birds were here screened for the occurrence of parasites by species-specific PCR assays of the mitochondrial cytochrome b gene (471 base pairs). Twenty-eight out of the 58 analysed birds had Haemoproteus (48%) infections while just 2% of the birds were infected with either Leucocytozoon or Plasmodium. Sixteen of the 28 birds carrying Haemoproteus had multiple infections. The phylogeography of the Philippine Bulbul mostly reflects the geographical origin of samples and it is consistent with the occurrence of two different subspecies on (1) Semirara and (2) Carabao, Boracay, North Gigante, Panay, and Negros, respectively. Haemoproteus phylogeography shows very little geographical structure, suggesting extensive gene flow among locations. While movements of birds among islands seem very sporadic, we found co-occurring evolutionary divergent parasite lineages. We conclude that historical processes have played a major role in shaping the host phylogeography, while they have left no signature in that of the parasites. Here ongoing population processes, possibly multiple reinvasions mediated by other hosts, are predominant.
The present cross-sectional anthropometric study evaluates new approaches in preventing overweight in children. Anthropometric data were obtained in 289 German children (146 boys, 143 girls) aged 6-11 years and 41 20-29 year old German female students for describing the type of body shape by Metric-Index (thoracic breadth, thoracic depth, height). The data were correlated with 15 fat layers measured with a lipometer with Principal Component Analyses and ANOVA. In females the android respectively gynoid type of body shape was significantly associated with the pyknomorphic respectively leptomorphic type. The study demonstrates that this pattern is established by an age of 8 years, when the earliest maturing children start to enter puberty. This pattern can be used to check the individual's risk of overweight and to recommend preventive healthcare interventions.
The levels of cellular organization, from gene transcription to translation to protein-protein interaction and metabolism, operate via tightly regulated mutual interactions, facilitating organismal adaptability and various stress responses. Characterizing the mutual interactions between genes, transcription factors, and proteins involved in signaling, termed crosstalk, is therefore crucial for understanding and controlling cells' functionality. We aim at using high-throughput transcriptomics data to discover previously unknown links between signaling networks. We propose and analyze a novel method for crosstalk identification which relies on transcriptomics data and overcomes the lack of complete information for signaling pathways in Arabidopsis thaliana. Our method first employs a network-based transformation of the results from the statistical analysis of differential gene expression in given groups of experiments under different signal-inducing conditions. The stationary distribution of a random walk (similar to the PageRank algorithm) on the constructed network is then used to determine the putative transcripts interrelating different signaling pathways. With the help of the proposed method, we analyze a transcriptomics data set including experiments from four different stresses/signals: nitrate, sulfur, iron, and hormones. We identified promising gene candidates, downstream of the transcription factors (TFs), associated to signaling crosstalk, which were validated through literature mining. In addition, we conduct a comparative analysis with the only other available method in this field which used a biclustering-based approach. Surprisingly, the biclustering-based approach fails to robustly identify any candidate genes involved in the crosstalk of the analyzed signals. We demonstrate that our proposed method is more robust in identifying gene candidates involved downstream of the signaling crosstalk for species for which large transcriptomics data sets, normalized with the same techniques, are available. Moreover, unlike approaches based on biclustering, our approach does not rely on any hidden parameters.
We use substituted polyanilines for the construction of new polymer electrodes for interaction studies with the redox protein cytochrome c (cyt c) and the enzyme sulfite oxidase (SO). For these purposes four different polyaniline copolymers are chemically synthesized. Three of them are copolymers, containing 2-methoxyaniline-5-sulfonic acid with variable ratios of aniline; the fourth copolymer consists of 3-amino-benzoic acid and aniline. The results show that all polymers are suitable for being immobilized as thin stable films on gold wire and indium tin oxide (ITO) electrode surfaces from DMSO solution. This can be demonstrated by cyclic voltammetry and UV-Vis spectroscopy measurements. Moreover, cyt c can be electrochemically detected not only in solution, but also immobilized on top of the polymer films. Furthermore, the appearance of a significant catalytic current has been demonstrated for the sulfonated polyanilines, when the polymer-coated protein electrode is being measured upon addition of sulfite oxidase, confirming the establishment of a bioanalytical signal chain. Best results have been obtained for the polymer with highest sulfonation grade. The redox switching of the polymer by the enzymatic reaction can also be analyzed by following the spectral properties of the polymer electrode.
The trade-off between predation risk and the need to feed is one of the major constraints animals have to cope with. Virtually all animals have a higher risk of being preyed upon when being active (e.g., searching for food or mating partners), compared with being inactive (e.g., staying at their burrows, nests, etc.). Yet, staying safe leads to a higher risk of starvation and may reduce reproductive success and body growth. Hence selection on behaviour optimizing the search, handling and digestion of food while avoiding the risk of predation is strong and should lead to strategies maximising survival chances and inclusive fitness. These facts call for integrative studies manipulating both, abundance of food and predation risk in a factorial set up, analysing the effects of both factors on behaviour and physiological parameters. We present results of a 2 x 2 factorial experiment, manipulating risk of predation and food abundance in guppies. We found that the two factors have an additive effect on body growth, but that predation risk by a pike cichlid is the main factor affecting the expression of behavioural strategies in guppies. Low food availability and high predation risk led to lower body growth. High predation risk affected swimming depth and risk sensitivity of guppies and might represent adaptive behavioural changes to the environmental context experienced in early life. Our study shows that integrative studies, analysing multiple interdependent and interconnected factors in the wild and in the lab are needed to further understand animal behaviour and development.
Today, serious health problems as overweight and obesity are not just constricted to the developed world, but also increase in the developing countries (Prentice 2006, Ramachandram et al. 2002). Focusing on this issue, BMI and percentage of body fat were compared in 2094 schoolchildren from two cross-sectional studies from India and Germany investigated in 2008 and 2009. The German children are in all age groups significantly taller, whereas the Indian children show higher values in BMI (e.g. 12 years: Indian: around 22 kg/m(2); German: around 19 kg/m(2)) and in the percentage of body fat (e.g. 12 years: Indian: around 27 %; German: around 18-20%) in most of the investigated age groups. The Indian children have significantly higher BMI between 10 and 13 (boys) respectively 14 years (girls). Indian children showed significant higher percentage of body fat between 10 and 15 years (boys) and between 8 and 16 years (girls). The difference in overweight between Indian and German children was strongest at 11 (boys) and 12 (girls) years: 70 % of the Indian but 20% of the German children were classified as overweight. In countries such as India that undergo nutritional transition, a rapid increase in obesity and overweight is observed. In contrast to the industrialized countries, the risk of overweight in developing countries is associated with high socioeconomic status. Other reasons of the rapid increase of overweight in the developing countries caused by different environmental or genetic factors are discussed.
Auxology has developed from mere describing child and adolescent growth into a vivid and interdisciplinary research area encompassing human biologists, physicians, social scientists, economists and biostatisticians. The meeting illustrated the diversity in auxology, with the various social, medical, biological and biostatistical aspects in studies on child growth and development.
Phytoplankton development affects the community structure and dynamics of freshwater bacteria by changing the availability of nutrients, algal exudates and biological surfaces. To elucidate these effects of phytoplankton development in spring in oligotrophic Lake Stechlin (Germany), we measured limnological and biological parameters, including the bacterial community composition (BCC), at the depth of the highest chlorophyll a concentration. To increase the resolution of BCC measurements, we separated particle-associated (PA) and free-living (FL) bacteria using serial filtration through 5.0 and 0.2 mu m pore size filters, respectively. The BCC of ultramicrobacteria was also determined by collecting the 0.2 mu m filtrate on 0.1 mu m filters. Changes in the community composition of Bacteria and particularly of Actinobacteria, one of the most important bacterial groups in temperate freshwater habitats, were studied via DGGE analysis of PCR-amplified 16S rRNA gene fragments. Patterns in BCC dynamics of FL Bacteria and Actinobacteria remained fairly constant throughout the study period, while patterns of PA Bacteria were more variable over time. At the breakdown of the diatom spring bloom, bacterial production and abundance sharply increased, indicating a close coupling between heterotrophic bacteria and algal detritus. The succession in BCC revealed life-style dependent patterns related to specific environmental variables. Our results indicate independent dynamics of PA and FL Bacteria as well as Actinobacteria during succession of phytoplankton spring blooms. These differences in bacterial lifestyle can only be resolved when the PA and FL fractions of microorganisms are separated.
The Antarctic terrestrial environment is under increasing pressure from human activities. The Fildes Region is characterized by high biodiversity, but is also a major logistic centre for the northern Antarctic Peninsula. Different interests, from scientific research, nature conservation, protection of geological and historical values, station operations, transport logistics and tourism, regularly overlap in space and time. This has led to increasing conflict among the multiple uses of the region and breaches of the legal requirements for environmental protection that apply in the area. The aim of this study was to assess the impacts of human activities in the Fildes Region by monitoring the distribution of bird and seal breeding sites and recording human activities and their associated environmental impacts. Data from an initial monitoring period 2003-06 were compared with data from 2008-10. We observed similar or increased levels of air, land and ship traffic, but fewer violations of overflight limits near Antarctic Specially Protected Area No. 150 Ardley Island. Open waste dumping and oil contamination are still major environmental impacts. Scientific and outdoor leisure activities undertaken by station personnel are more frequent than tourist activities and are likely to have a commensurate level of environmental impact. Despite the initial success of some existing management measures, it is essential that scientific and environmental values continue to be safeguarded, otherwise environmental impacts will increase and the habitat will be further degraded. We argue that the Fildes Region should be considered for designation as an Antarctic Specially Managed Area, a measure that has proven effective for environmental management of vulnerable areas of the Antarctic.
Disturbances are characteristic for many ecosystems. However, we still lack generalizations concerning their role in shaping communities, particularly when disturbances co-occur. To study such effects, we used a novel modeling approach that is unrestricted by a priori tradeoffs among specific plant traits, except for those generated by allocation principles. Thus, trait combinations were emergent properties associated with biotic and abiotic constraints. Specifically, we asked which traits dominate under specific disturbance regimes, whether single and combined disturbance regimes promote similar trait tradeoffs and how complex disturbance regimes affect species richness and functional diversity. Overall, disturbances' temporal properties governed the outcome of combined disturbances and were a stronger assortative force than spatial disturbance properties: low temporal predictability decreased seed-dispersability and dormancy, but increased competitive ability and disturbance tolerance. Evidence for tradeoffs between different colonization modes and between dormancy and disturbance tolerance were found, while surprisingly, the widely accepted colonization-competition tradeoff was not generated. Diversity was highest at intermediate disturbance intensity, but decreased monotonically with increasing unpredictability. In accordance with our results, future models should avoid restrictive assumptions about tradeoffs to generate robust and more general predictions about the role of disturbances for community dynamics.
Rates of multiple paternities were investigated in the sailfin molly (Poecilia latipinna), using eight microsatellite loci. Genotyping was performed for offspring and mothers in 40 broods from four allopatric populations from the south-eastern U.S.A. along a geographic stretch of 1200 km in west-east direction and approximately 200 km from north to south. No significant differences regarding rates of multiple paternities were found between populations despite sample populations stemming from ecologically divergent habitats. Even the most conservative statistical approach revealed a minimum of 70% of the broods being sired by at least two males, with an average of 1.80-2.95 putative fathers per brood. Within broods, one male typically sired far more offspring than would be expected under an assumed equal probability of all detected males siring offspring.
Land use is increasingly recognized as a major driver of biodiversity and ecosystem functioning in many current research projects. In grasslands, land use is often classified by categorical descriptors such as pastures versus meadows or fertilized versus unfertilized sites. However, to account for the quantitative variation of multiple land-use types in heterogeneous landscapes, a quantitative, continuous index of land-use intensity (LUI) is desirable. Here we define such a compound, additive LUI index for managed grasslands including meadows and pastures. The LUI index summarizes the standardized intensity of three components of land use, namely fertilization, mowing, and livestock grazing at each site. We examined the performance of the LUI index to predict selected response variables on up to 150 grassland sites in the Biodiversity Exploratories in three regions in Germany(Alb, Hainich, Schorlheide). We tested the average Ellenberg nitrogen indicator values of the plant community, nitrogen and phosphorus concentration in the aboveground plant biomass, plant-available phosphorus concentration in the top soil, and soil C/N ratio, and the first principle component of these five response variables.
The LUI index significantly predicted the principal component of all five response variables, as well as some of the individual responses. Moreover, vascular plant diversity decreased significantly with LUI in two regions (Alb and Hainich).
Inter-annual changes in management practice were pronounced from 2006 to 2008, particularly due to variation in grazing intensity. This rendered the selection of the appropriate reference year(s) an important decision for analyses of land-use effects, whereas details in the standardization of the index were of minor importance. We also tested several alternative calculations of a LUI index, but all are strongly linearly correlated to the proposed index.
The proposed LUI index reduces the complexity of agricultural practices to a single dimension and may serve as a baseline to test how different groups of organisms and processes respond to land use. In combination with more detailed analyses, this index may help to unravel whether and how land-use intensities, associated disturbance levels or other local or regional influences drive ecological processes.
The understanding of environmentally induced changes in the biochemical composition of phytoplankton species is of great importance in both physiological studies and ecological food web research. In extensive laboratory experiments we tested the influence of two different temperatures (10 degrees C and 25 degrees C) and a phosphorus supply gradient on the sterol concentrations of the three freshwater phytoplankton species Scenedesmus quadricauda, Cryptomonas ovata and Cyclotella meneghiniana. The diatom C. meneghiniana was additionally exposed to a silicate gradient. In two separate experiments we analysed (1) possible interactive effects of temperature and phosphorus supply and (2) the effect of four phosphorus levels and three silicate levels on algal sterol concentrations. We observed that sterol concentrations were higher at 25 degrees C than at 10 degrees C in S. quadricauda and C. meneghiniana, but were not affected by temperature in C. ovata. Interactive effects of temperature and phosphorus supply on sterol concentrations were found in C. meneghiniana. This presumably was due to the bioconversion of one sterol (24-methylenecholesterol) into another (22-dihydrobrassicasterol). Increasing phosphorus supply resulted in species-specific effects on sterol concentrations, viz. an optimum curve response in S. quadricauda, a saturation curve response in C. meneghiniana and no change in sterol concentration in C. ovata. Effects of silicate supply on the sterols of C. meneghiniana equalled the effects of phosphorus supply. Albeit we did not observe a general trend in the three phytoplankton species tested, we conclude that sterol concentrations of phytoplankton are strongly affected by temperature and nutrient supply. Interactive effects point out the importance of taking into account more than just one environmental factor when assessing the effects of environmentally induced changes on phytoplankton sterol concentrations.
Background: The multidrug transporter P-glycoprotein (PGP) is expressed in the human placenta. In particular the C3435T ABCB1 polymorphism was associated with altered tissue expression of PGP in the human placenta. However, the potential functional impact of this polymorphism on the offspring is unknown so far.
Methods: We analyzed the impact of the ABCB1/C3435T polymorphism on fetal growth in 262 mother/child pairs. Fetal growth was assessed by differential ultrasound examination of the fetal body prior to birth and by measuring birth weight.
Results: The maternal ABCB1/C3435T polymorphism showed no trend for an association with birth weight or any ultrasound parameter describing late gestational fetal body shape. Genotyping the newborns, however, demonstrated that newborns carrying two copies of the T allele had a birth weight of 3176.39 g, whereas CT and CC newborns had a birth weight of 3345.04 g (p = 0.022). Adjusting for gestational age at delivery, child's gender, maternal BM1, maternal age and body weight at delivery confirmed this finding (p = 0.009). Considering gestational day of late ultrasound examination, gestational age at delivery, child's gender, maternal BMI, maternal age and maternal body weight at delivery, the fetal ABCB1/C3435T genotype revealed likewise a significant negative correlation with abdominal diameter and abdominal circumference (R-2 = 0.538, p = 0.010 and R-2 = 0.534, p = 0.005, respectively).
Conclusions: Low birth weight may be a risk factor for cardiovascular diseases in later life. The fetal ABCB1/C3435T gene polymorphism may contribute to this risk. Since PGP controls transport of various biological agents, we suggest that PGP is involved in the transport of biological agents to the fetus that are important for normal fetal growth.
Restoration is gaining importance in the management of plant invasions. As the success of restoration projects is frequently determined by factors other than ecological ones, we explored the ecological and financial feasibility of active restoration on three different invaded sites in South Africa's Cape Floristic Region. The aim of our study was to identify cost-effective ways of restoring functional native ecosystems following invasion by alien plants. Over three years we evaluated different restoration approaches using field trials and experimental manipulations (i.e. mechanical clearing, burning, different soil restoration techniques and sowing of native species) to reduce elevated soil nutrient levels and to re-establish native fynbos communities. Furthermore we investigated the possibility of introducing native fynbos species that can be used for sustainable harvesting to create an incentive for restoration on private land.
Diversity and evenness of native plant species increased significantly after restoration at all three sites, whereas cover of alien plants decreased significantly, confirming that active restoration was successful. However, sowing of native fynbos species had no significant effect on native cover, species richness, diversity or evenness in the Acacia thicket and Kikuyu field, implying that the ecosystem was sufficiently resilient to allow autogenic recovery following clearing and burning of the invasive species. Soil restoration treatments resulted in an increase of available nitrogen in the Acacia thicket, but had no significant effects in the Eucalyptus plantation. However, despite elevated available soil nitrogen levels, native species germinated irrespective whether sown or unsown (i.e. regeneration from the soil seed bank).
Without active introduction of native species, native grasses, forbs and other shrubs would have dominated, and proteoids and ericoids (the major fynbos growth forms) would have been under-represented.
The financial analysis shows that income from flower harvesting following active restoration consistently outweighs income following passive restoration, but that the associated increase in income does not always justify the higher costs. We conclude that active restoration can be effective and financially feasible when compared to passive restoration, depending on the density of invasion. Active restoration of densely invaded sites may therefore only be justifiable if the target area is in a region of high conservation priority.
Coastal regions of the North East Atlantic and Mediterranean Seas have four known species of driftwood talitrids. Records are extremely scanty, often limited to the type locality and dating to 1950. We were able to study three of them, all belonging to the genus Macarorchestia, using fresh and archived samples including type material. Allometric and molecular analyses support: (1) a close relationship among all the three classically defined Macarorchestia species, (2) Macarorchestia was well separated from non-driftwood taxa, and (3) a putative new driftwood talitrid discovered during this study was not closely related to Macarorchestia. Genetic divergence between the new species and Macarorchestia remyi is as high as the average distance among a number of talitrid species included in the study for comparison. A key is provided to identify all three of the presently known species of Macarorchestia, using morphological characters employed in the allometric study.
The interaction of diverse biomaterials with surfaces is more crucial than ever for biomedical applications to ensure efficiency and reproducibility. Very interesting surface materials are micrometer-thick polyelectrolyte multilayers. Not only their surface but also the bulk can be loaded with biomaterials like proteins or DNA for various purposes. Therefore, we established a method to analyze the lateral and vertical distribution of fluorescently labelled proteins of various size and charge in polyelectrolyte films composed of poly(L-lysine) and hyaluronic acid by confocal laser scanning microscopy. This approach enables us to measure the diffusion coefficients of the proteins via fluorescence recovery after photobleaching as a function of their vertical position in the film and facilitates the understanding of molecular interactions in the film with a high resolution in both space and time. As a result, we confirm that protein loading in the film is driven by electrostatic interactions - uncharged dextran molecules of 10 and 500 kDa do not diffuse into the film. Proteins of different sizes (3-11 nm) can diffuse relatively fast (D = 2-4 mm(2) s(-1)) independent of their net charge, indicating complex interpolymer interactions. This approach is a new powerful experimental tool to design the polyelectrolyte multilayers for bio-applications by finding a relationship between intermolecular interactions and mobility and availability of biomolecules to biological samples (e.g. cells) or detection units (e.g. biosensors).
Degradable polymers with a tailorable degradation rate might be promising candidate materials for biomaterial-based cartilage repair. In view of the poor intrinsic healing capability of cartilage, implantation of autologous chondrocytes seeded on a biocompatible slow degrading polymer might be an encouraging approach to improve cartilage repair in the future. This study was undertaken to test if the fiber orientation (random versus aligned) of two different degradable polymers and a polymer intended for long term applications could influence primary articular chondrocytes growth and ultrastructure.
A degradable copoly(ether) esterurethane (PDC) was synthesized via co-condensation of poly(p-dioxanone) diol and poly(epsilon-caprolactone) diol using an aliphatic diisocyanate as linker. Poly(p-dioxanone) (PPDO) was applied as commercially available degradable polymer, while polyetherimide (PEI) was chosen as biomaterial enabling surface functionalization. The fibrous scaffolds of PDC and PPDO were obtained by electrospinning using 1,1,1,3,3,3 hexafluoro-2-propanol (HFP), while for PEI dimethyl acetamide (DMAc) was applied as solvent. Primary porcine articular chondrocytes were seeded at different cell densities on the fibrous polymer scaffolds and analyzed for viability (fluorescein diacetate/ethidiumbromide staining), for type II collagen synthesis (immunolabelling), ultrastructure and orientation on the fibers (SEM: scanning electron microscopy).
Vital chondrocytes adhered on all electrospun scaffolds irrespective of random and aligned topologies. In addition, the chondrocytes produced the cartilage-specific type II collagen on all tested polymer topologies suggesting their differentiated functions. SEM revealed an almost flattened chondrocytes shape on scaffolds with random fiber orientation: whereby chondrocytes growth remained mainly restricted to the scaffold surface. On aligned fibers the chondrocytes exhibited a more spindle-shaped morphology with rougher cell surfaces but only a minority of the cells aligned according to the fibers. As a next step the reduction of the fiber diameter of electrospun scaffolds should be addressed as an important parameter to mimic cartilage ECM structure.
Background: Triblock copolymers from hydrophilic oligo(ethylene glycol) segment A and oligo(propylene glycol) segment B, providing an ABA structure (OEG-OPG-OEG triblock), are known to be biocompatible and are used as self-solidifying gels in drug depots. A complete removal of these depots would be helpful in cases of undesired side effects of a drug, but this remains a challenge as they liquefy below their transition temperature. Therefore we describe the synthesis of covalently cross-linked hydrogel networks.
Method: Triblock copolymer-based hydrogels were created by irradiating aqueous solutions of the corresponding macro-dimethacrylates with UV light. The degree of swelling, swelling kinetics, mechanical properties and morphology of the networks were investigated.
Results: Depending on precursor concentration, equilibrium degree of swelling of the films ranged between 500% and 880% and was reached in 1 hour. In addition, values for storage and loss moduli of the hydrogel networks were in the 100 Pa to 10 kPa range.
Conclusion: Although OEG-OPG-OEG triblocks are known for their micellization, which could hamper polymer network formation, reactive OEG-OPG-OEG triblock oligomers could be successfully polymerized into hydrogel networks. The degree of swelling of these hydrogels depends on their molecular weight and on the oligomer concentration used for hydrogel preparation. In combination with the temperature sensitivity of the ABA triblock copolymers, it is assumed that such hydrogels might be beneficial for future medical applications -e.g., removable drug release systems.
Transcription factor OsHsfC1b regulates salt tolerance and development in Oryza sativa ssp japonica
(2012)
Background and aims Salt stress leads to attenuated growth and productivity in rice. Transcription factors like heat shock factors (HSFs) represent central regulators of stress adaptation. Heat shock factors of the classes A and B are well established as regulators of thermal and non-thermal stress responses in plants; however, the role of class C HSFs is unknown. Here we characterized the function of the OsHsfC1b (Os01g53220) transcription factor from rice.
Methodology We analysed the expression of OsHsfC1b in the rice japonica cultivars Dongjin and Nipponbare exposed to salt stress as well as after mannitol, abscisic acid (ABA) and H2O2 treatment. For functional characterization of OsHsfC1b, we analysed the physiological response of a T-DNA insertion line (hsfc1b) and two artificial micro-RNA (amiRNA) knock-down lines to salt, mannitol and ABA treatment. In addition, we quantified the expression of small Heat Shock Protein (sHSP) genes and those related to signalling and ion homeostasis by quantitative real-time polymerase chain reaction in roots exposed to salt. The subcellular localization of OsHsfC1b protein fused to green fluorescent protein (GFP) was determined in Arabidopsis mesophyll cell protoplasts.
Principal results Expression of OsHsfC1b was induced by salt, mannitol and ABA, but not by H2O2. Impaired function of OsHsfC1b in the hsfc1b mutant and the amiRNA lines led to decreased salt and osmotic stress tolerance, increased sensitivity to ABA, and temporal misregulation of salt-responsive genes involved in signalling and ion homeostasis. Furthermore, sHSP genes showed enhanced expression in knock-down plants under salt stress. We observed retarded growth of hsfc1b and knock-down lines in comparison with control plants under non-stress conditions. Transient expression of OsHsfC1b fused to GFP in protoplasts revealed nuclear localization of the transcription factor.
Conclusions OsHsfC1b plays a role in ABA-mediated salt stress tolerance in rice. Furthermore, OsHsfC1b is involved in the response to osmotic stress and is required for plant growth under non-stress conditions.
Influence of drying on the secondary structure of intrinsically disordered and globular proteins
(2012)
Circular dichroism (CD) spectroscopy of five Arabidopsis late embryogenesis abundant (LEA) proteins constituting the plant specific families LEA_5 and LEA_6 showed that they are intrinsically disordered in solution and partially fold during drying. Structural predictions were comparable to these results for hydrated LEA_6, but not for LEA_5 proteins. FTIR spectroscopy showed that verbascose, but not sucrose, strongly affected the structure of the dry proteins. The four investigated globular proteins were only mildly affected by drying in the absence, but strongly in the presence of sugars. These data highlight the larger structural flexibility of disordered compared to globular proteins and the impact of sugars on the structure of both disordered and globular proteins during drying.
Unique properties of eukaryote-type actin and profilin horizontally transferred to cyanobacteria
(2012)
A eukaryote-type actin and its binding protein profilin encoded on a genomic island in the cyanobacterium Microcystis aeruginosa PCC 7806 co-localize to form a hollow, spherical enclosure occupying a considerable intracellular space as shown by in vivo fluorescence microscopy. Biochemical and biophysical characterization reveals key differences between these proteins and their eukaryotic homologs. Small-angle X-ray scattering shows that the actin assembles into elongated, filamentous polymers which can be visualized microscopically with fluorescent phalloidin. Whereas rabbit actin forms thin cylindrical filaments about 100 mu m in length, cyanobacterial actin polymers resemble a ribbon, arrest polymerization at 510 lam and tend to form irregular multi-strand assemblies. While eukaryotic profilin is a specific actin monomer binding protein, cyanobacterial profilin shows the unprecedented property of decorating actin filaments. Electron micrographs show that cyanobacterial profilin stimulates actin filament bundling and stabilizes their lateral alignment into heteropolymeric sheets from which the observed hollow enclosure may be formed. We hypothesize that adaptation to the confined space of a bacterial cell devoid of binding proteins usually regulating actin polymerization in eukaryotes has driven the co-evolution of cyanobacterial actin and profilin, giving rise to an intracellular entity.
Background: Different immunotherapy approaches for the treatment of cancer and autoimmune diseases are being developed and tested in clinical studies worldwide. Their resulting complex experimental data should be properly evaluated, therefore reliable normal healthy control baseline values are indispensable.
Methodology/Principal Findings: To assess intra- and inter-individual variability of various biomarkers, peripheral blood of 16 age and gender equilibrated healthy volunteers was sampled on 3 different days within a period of one month. Complex "crossomics'' analyses of plasma metabolite profiles, antibody concentrations and lymphocyte subset counts as well as whole genome expression profiling in CD4(+)T and NK cells were performed. Some of the observed age, gender and BMI dependences are in agreement with the existing knowledge, like negative correlation between sex hormone levels and age or BMI related increase in lipids and soluble sugars. Thus we can assume that the distribution of all 39.743 analysed markers is well representing the normal Caucasoid population. All lymphocyte subsets, 20% of metabolites and less than 10% of genes, were identified as highly variable in our dataset.
Conclusions/Significance: Our study shows that the intra- individual variability was at least two-fold lower compared to the inter-individual one at all investigated levels, showing the importance of personalised medicine approach from yet another perspective.
Preferential accumulation of mutant proteins in the nucleus has been suggested to be the molecular culprit that confers cellular toxicity in the neurodegenerative disorders caused by polyglutamine (polyQ) expansion. Here, we use dynamic imaging approaches, orthogonal cross-seeding, and composition analysis to examine the dynamics and structure of nuclear and cytoplasmic inclusions of atrophin-1, implicated in dentatorubropallidoluysian atrophy, a polyQ-based disease with complex clinical features. Our results reveal a large heterogeneity in the dynamics of the nuclear inclusions compared with the compact and immobile cytoplasmic aggregates. At least two types of inclusions of expanded atrophin-1 with different mobility of the molecular species and ability to exchange with the surrounding monomer pool coexist in the nucleus. Intriguingly, the enrichment of nuclear inclusions with slow dynamics parallels changes in the aggregate core architecture that are dominated by the polyQ stretch. We propose that the observed complexity in the dynamics of the nuclear inclusions provides a molecular explanation for the enhanced cellular toxicity of the nuclear aggregates in polyQ-based neurodegeneration.
Characterization of NE81, the first lamin-like nucleoskeleton protein in a unicellular organism
(2012)
Lamins build the nuclear lamina and are required for chromatin organization, gene expression, cell cycle progression, and mechanical stabilization. Despite these universal functions, lamins have so far been found only in metazoans. We have identified protein NE81 in Dictyostelium, which has properties that justify its denomination as a lamin-like protein in a lower eukaryote. This is based on its primary structure, subcellular localization, and regulation during mitosis, and its requirement of the C-terminal CaaX box as a posttranslational processing signal for proper localization. Our knockout and overexpression mutants revealed an important role for NE81 in nuclear integrity, chromatin organization, and mechanical stability of cells. All our results are in agreement with a role for NE81 in formation of a nuclear lamina. This function is corroborated by localization of Dictyostelium NE81 at the nuclear envelope in human cells. The discovery of a lamin-like protein in a unicellular organism is not only intriguing in light of evolution, it may also provide a simple experimental platform for studies of the molecular basis of laminopathies.
Background: In trying to understand the evolutionary relationships of organisms, the current flood of sequence data offers great opportunities, but also reveals new challenges with regard to data quality, the selection of data for subsequent analysis, and the automation of steps that were once done manually for single-gene analyses. Even though genome or transcriptome data is available for representatives of most bilaterian phyla, some enigmatic taxa still have an uncertain position in the animal tree of life. This is especially true for myzostomids, a group of symbiotic ( or parasitic) protostomes that are either placed with annelids or flatworms.
Methodology: Based on similarity criteria, Illumina-based transcriptome sequences of one myzostomid were compared to protein sequences of one additional myzostomid and 29 reference metazoa and clustered into gene families. These families were then used to investigate the phylogenetic position of Myzostomida using different approaches: Alignments of 989 sequence families were concatenated, and the resulting superalignment was analyzed under a Maximum Likelihood criterion. We also used all 1,878 gene trees with at least one myzostomid sequence for a supertree approach: the individual gene trees were computed and then reconciled into a species tree using gene tree parsimony.
Conclusions: Superalignments require strictly orthologous genes, and both the gene selection and the widely varying amount of data available for different taxa in our dataset may cause anomalous placements and low bootstrap support. In contrast, gene tree parsimony is designed to accommodate multilocus gene families and therefore allows a much more comprehensive data set to be analyzed. Results of this supertree approach showed a well-resolved phylogeny, in which myzostomids were part of the annelid radiation, and major bilaterian taxa were found to be monophyletic.
In this field experiment we investigate the impact of land use induced savanna degradation on movement behaviour of the spotted sand lizard (Pedioplanis l. lineoocellata) in the southern Kalahari. Foraging behaviour of lizards was tested in a factorial design (low vs. high prey availability) in degraded and non-degraded habitats.
An interaction between habitat structure and prey availability affected movement behaviour. In degraded habitats with low prey availability and in non-degraded habitats with high prey availability the spotted sand lizard moved more like an active forager. In contrast, in degraded habitats with high prey availability and in non-degraded habitats with low prey availability lizards moved like sit-and-wait foragers. Interestingly, the behavioural flexibility of the spotted sand lizard seems to buffer extreme conditions and negative effects of land use impacts.
Aim The study and prediction of speciesenvironment relationships is currently mainly based on species distribution models. These purely correlative models neglect spatial population dynamics and assume that species distributions are in equilibrium with their environment. This causes biased estimates of species niches and handicaps forecasts of range dynamics under environmental change. Here we aim to develop an approach that statistically estimates process-based models of range dynamics from data on species distributions and permits a more comprehensive quantification of forecast uncertainties.
Innovation We present an approach for the statistical estimation of process-based dynamic range models (DRMs) that integrate Hutchinson's niche concept with spatial population dynamics. In a hierarchical Bayesian framework the environmental response of demographic rates, local population dynamics and dispersal are estimated conditional upon each other while accounting for various sources of uncertainty. The method thus: (1) jointly infers species niches and spatiotemporal population dynamics from occurrence and abundance data, and (2) provides fully probabilistic forecasts of future range dynamics under environmental change. In a simulation study, we investigate the performance of DRMs for a variety of scenarios that differ in both ecological dynamics and the data used for model estimation.
Main conclusions Our results demonstrate the importance of considering dynamic aspects in the collection and analysis of biodiversity data. In combination with informative data, the presented framework has the potential to markedly improve the quantification of ecological niches, the process-based understanding of range dynamics and the forecasting of species responses to environmental change. It thereby strengthens links between biogeography, population biology and theoretical and applied ecology.
There is growing consensus that the growth of herbivorous consumers is frequently limited by more than one nutrient simultaneously. This understanding, however, is based primarily on theoretical considerations and the applicability of existing concepts of co-limitation has rarely been tested experimentally. Here, we assessed the suitability of two contrasting concepts of resource limitation, i.e. Liebigs minimum rule and the multiple limitation hypothesis, to describe nutrient-dependent growth responses of a freshwater herbivore (Daphnia magna) in a system with two potentially limiting nutrients (cholesterol and eicosapentaenoic acid). The results indicated that these essential nutrients interact, and do not strictly follow Liebigs minimum rule, which consistently overestimates growth at co-limiting conditions and thus is not applicable to describe multiple nutrient limitation of herbivorous consumers. We infer that the outcome of resource-based modelling approaches assessing herbivore population dynamics strongly depends on the applied concept of co-limitation.
This article gives an overview of the current status of knowledge concerning the role of nanoparticles (inorganic and organic) in deep geological host rocks and the potential influence of these nanoparticles on radionuclide migration in far-field systems. The manuscript is not intended to be a full review paper or overview paper concerning nanoparticles, here the intention is to refer to recent publications but to highlight the progress made in the 6th framework project IP FUNMIG (Fundamental processes of radionuclide migration) and the open literature over the past 5 a concerning the process understanding of nanoparticle related issues in the three host rock formations investigated, namely: claystones, crystalline rocks and salt rock overburden. The results show inter alia that the inorganic nanoparticle concentration in deep groundwaters of advection dominated systems rarely exceeds 1 mg L (1) and is expected to be in the ng L (1) range in diffusion controlled systems. For organic nanoparticles DOC concentrations up to tens of milligrams in diffusion-controlled indurated clays with molecular sizes mostly <500 Da have been found. Fulvic acid type organics have been identified in crystalline environments and plastic Clay formations (Boom Clay) with molecular sizes <= 300 kDa. Additional sources of inorganic nanoparticles from the repository near-field (compacted bentonite) were identified and the initial erosion rates were determined. The results indicate under stagnant conditions similar to 38 mg cm (2) a (1) for bi-distilled water, similar to 20 mg cm (2) a (1) for glacial melt water (Grimsel groundwater) and very low rates similar to 0.02 mg cm (2) a (1) for 5 mM CaCl2 contact water. The low critical coagulation concentration (CCC) indicative for purely diffusion controlled coagulation of 1 mM L (1) Ca2+ found in bentonite nanoparticle stability analysis matches the low nanoparticle mobilization from compacted bentonite found in these systems.
We tested, in the laboratory, the influence of light intensity, temperature, and phosphorus (P) supply on fatty acid (FA) concentrations of four freshwater algae: the green algae Scenedesmus quadricauda (Turpin) Breb. and Chlamydomonas globosa J. Snow, the cryptophyte Cryptomonas ovata Ehrenb., and the diatom Cyclotella meneghiniana Kutz. We investigated the main and interactive effects of two variables on algal FA concentrations (i.e., light intensity and P supply or temperature and P supply). Interactive effects of light intensity and P supply were most pronounced in C. meneghiniana, but were also found in S. quadricauda and C. ovata. Changes in several saturated and unsaturated FA concentrations with light were more distinct in the low-P treatments than in the high-P treatments. Interactive effects of temperature and P supply on various FA concentrations were observed in all four species, but there was no consistent pattern. In lake ecosystems, P limitation often coincides with high light intensities and temperatures in summer. Therefore, it is important to examine how combinations of these environmental conditions affect FA concentrations of primary producers that are important sources of FAs for higher trophic levels.
We studied the spreading of feature-based attention from attended to ignored motion fields (linear, circular, and combinations). When observers attended one of two superimposed motion fields on one side of the visual midline, sub-threshold priming by an ignored motion field was altered significantly on the opposite side of the midline. This attentional spreading was observed only when attended and ignored motion fields conformed to a complex global flow, not when they shared the same linear motion. These findings corroborate an earlier study (Festman & Braun, 2010), which obtained similar results with a complementary methodology. We conclude that feature-based attention is more complex than hitherto appreciated in that it spreads preferentially in an object-specific manner.
The vesicle-inducing protein in plastids (VIPP1) was suggested to play a role in thylakoid membrane formation via membrane vesicles. As this functional assignment is under debate, we investigated the function of VIPP1 in Chlamydomonas reinhardtii. Using immunofluorescence, we localized VIPP1 to distinct spots within the chloroplast. In VIPP1-RNA interference/artificial microRNA cells, we consistently observed aberrant, prolamellar body-like structures at the origin of multiple thylakoid membrane layers, which appear to coincide with the immunofluorescent VIPP1 spots and suggest a defect in thylakoid membrane biogenesis. Accordingly, using quantitative shotgun proteomics, we found that unstressed vipp1 mutant cells accumulate 14 to 20% less photosystems, cytochrome b(6)f complex, and ATP synthase but 30% more light-harvesting complex II than control cells, while complex assembly, thylakoid membrane ultrastructure, and bulk lipid composition appeared unaltered. Photosystems in vipp1 mutants are sensitive to high light, which coincides with a lowered midpoint potential of the Q(A)/Q(A)(-) redox couple and increased thermosensitivity of photosystem II (PSII), suggesting structural defects in PSII. Moreover, swollen thylakoids, despite reduced membrane energization, in vipp1 mutants grown on ammonium suggest defects in the supermolecular organization of thylakoid membrane complexes. Overall, our data suggest a role of VIPP1 in the biogenesis/assembly of thylakoid membrane core complexes, most likely by supplying structural lipids.
The transition from juvenility through maturation to senescence is a complex process that involves the regulation of longevity. Here, we identify JUNGBRUNNEN1 (JUB1), a hydrogen peroxide (H2O2)-induced NAC transcription factor, as a central longevity regulator in Arabidopsis thaliana. JUB1 overexpression strongly delays senescence, dampens intracellular H2O2 levels, and enhances tolerance to various abiotic stresses, whereas in jub1-1 knockdown plants, precocious senescence and lowered abiotic stress tolerance are observed. A JUB1 binding site containing a RRYGCCGT core sequence is present in the promoter of DREB2A, which plays an important role in abiotic stress responses. JUB1 transactivates DREB2A expression in mesophyll cell protoplasts and transgenic plants and binds directly to the DREB2A promoter. Transcriptome profiling of JUB1 overexpressors revealed elevated expression of several reactive oxygen species-responsive genes, including heat shock protein and glutathione S-transferase genes, whose expression is further induced by H2O2 treatment. Metabolite profiling identified elevated Pro and trehalose levels in JUB1 overexpressors, in accordance with their enhanced abiotic stress tolerance. We suggest that JUB1 constitutes a central regulator of a finely tuned control system that modulates cellular H2O2 level and primes the plants for upcoming stress through a gene regulatory network that involves DREB2A.
Plant community assembly in temperate forests along gradients of soil fertility and disturbance
(2012)
Plant community assembly from a regional pool is largely driven by two mechanisms: environmental filtering and niche partitioning, which result in trait convergence or divergence, respectively. Although empirical evidence for both assembly mechanisms exists, the environmental conditions and traits where each of the two assembly patterns is prevalent remain unclear. We studied community assembly mechanisms in herb layer communities of temperate forest patches in NW Germany, looking at distributions of competitive and reproductive traits along gradients of soil fertility and disturbance. We also examined how community assembly patterns changed over a time span of two decades. Canopy height converged toward taller species with increasing soil fertility and increasing light availability. Most reproductive traits diverged with an increasing degree of disturbance and with increasing fertility. Comparisons over time indicated that disturbance events induced the coexistence of species with different reproductive strategies and also selected for tall species as a result of enhanced competitive pressure. Our study demonstrates that in accordance with existing hypotheses, competitive traits (e.g., canopy height) can be convergent in favorable environments. However, this convergence is associated with a divergence of traits related to other challenges (e.g., reproduction), indicating that true functional redundancy within communities does not exist. Moreover, our study shows that the expected divergence of reproductive traits at disturbed sites can be accompanied by a convergence of other traits (e.g., canopy height), indicating that several assembly mechanisms can operate simultaneously.
The uptake of potassium ions (K+) accompanied by an acidification of the apoplasm is a prerequisite for stomatal opening. The acidification (approximately 2-2.5 pH units) is perceived by voltage-gated inward potassium channels (K-in) that then can open their pores with lower energy cost. The sensory units for extracellular pH in stomatal K-in channels are proposed to be histidines exposed to the apoplasm. However, in the Arabidopsis thaliana stomatal K-in channel KAT1, mutations in the unique histidine exposed to the solvent (His(267)) do not affect the pH dependency. We demonstrate in the present study that His(267) of the KAT1 channel cannot sense pH changes since the neighbouring residue Phe(266) shifts its pK(a) to undetectable values through a cation-pi interaction. Instead, we show that Glu(240) placed in the extracellular loop between transmembrane segments S5 and S6 is involved in the extracellular acid activation mechanism. Based on structural models we propose that this region may serve as a molecular link between the pH- and the voltage-sensor. Like Glu(240), several other titratable residues could contribute to the pH-sensor of KAT1, interact with each other and even connect such residues far away from the voltage-sensor with the gating machinery of the channel.
Background: Mediterranean temporary water bodies are important reservoirs of biodiversity and host a unique assemblage of diapausing aquatic invertebrates. These environments are currently vanishing because of increasing human pressure. Chirocephalus kerkyrensis is a fairy shrimp typical of temporary water bodies in Mediterranean plain forests and has undergone a substantial decline in number of populations in recent years due to habitat loss. We assessed patterns of genetic connectivity and phylogeographic history in the seven extant populations of the species from Albania, Corfu Is. (Greece), Southern and Central Italy.
Methodology/Principal Findings: We analyzed sequence variation at two mitochondrial DNA genes (Cytochrome Oxidase I and 16s rRNA) in all the known populations of C. kerkyrensis. We used multiple phylogenetic, phylogeographic and coalescence-based approaches to assess connectivity and historical demography across the whole distribution range of the species. C. kerkyrensis is genetically subdivided into three main mitochondrial lineages; two of them are geographically localized (Corfu Is. and Central Italy) and one encompasses a wide geographic area (Albania and Southern Italy). Most of the detected genetic variation (approximate to 81%) is apportioned among the aforementioned lineages.
Conclusions/Significance: Multiple analyses of mismatch distributions consistently supported both past demographic and spatial expansions with the former predating the latter; demographic expansions were consistently placed during interglacial warm phases of the Pleistocene while spatial expansions were restricted to cold periods. Coalescence methods revealed a scenario of past isolation with low levels of gene flow in line with what is already known for other co-distributed fairy shrimps and suggest drift as the prevailing force in promoting local divergence. We recommend that these evolutionary trajectories should be taken in proper consideration in any effort aimed at protecting Mediterranean temporary water bodies.
Ellenberg indicator values are widely used ecological tools to elucidate relationships between vegetation and environment in ecological research and environmental planning. However, they are mainly deduced from expert knowledge on plant species and are thus subject of ongoing discussion. We researched if Ellenberg indicator values can be directly extracted from the vegetation biomass itself. Mean Ellenberg "moisture" (mF) and "nitrogen" (mN) values of 141 grassland plots were related to nutrient concentrations, fibre fractions and spectral information of the aboveground biomass. We developed calibration models for the prediction of mF and mN using spectral characteristics of biomass samples with near-infrared reflectance spectroscopy (NIRS). Prediction goodness was evaluated with internal cross-validations and with an external validation data set. NIRS could accurately predict Ellenberg mN, and with less accuracy Ellenberg mF. Predictions were not more precise for cover-weighted Ellenberg values compared with un-weighted values. Both Ellenberg mN and mF showed significant and strong correlations with some of the nutrient and fibre concentrations in the biomass. Against expectations, Ellenberg mN was more closely related to phosphorus than to nitrogen concentrations, suggesting that this value rather indicates productivity than solely nitrogen. To our knowledge we showed for the first time that mean Ellenberg indicator values could be directly predicted from the aboveground biomass, which underlines the usefulness of the NIRS technology for ecological studies, especially in grasslands ecosystems.
1. Poikilothermic animals incorporate more polyunsaturated fatty acids (PUFAs) into their cellular membranes as temperature declines, suggesting an increased sensitivity to PUFA limitation in cool conditions. To test this we raised Daphnia magna at different temperatures and investigated the effect of varying dietary PUFA on life history parameters (i.e. growth, reproduction) and the PUFA composition of body tissue and eggs.
2. Upon a PUFA-rich diet (Cryptomonas sp.) females showed higher concentrations of several omega 3 PUFAs in their body tissue at 15 degrees C than at 20 degrees C and 25 degrees C, indicating a greater structural requirement for omega 3 PUFAs at low temperature. Their eggs had an equal but higher concentration of omega 3 PUFAs than their body tissue.
3. In a life history experiment at 15 and 20 degrees C we supplemented a diet of a PUFA-free cyanobacterium with the omega 3 PUFA eicosapentaenoic acid (EPA). The growth of D. magna was more strongly EPA limited at low temperature. A greater requirement for structural EPA at 15 degrees C was indicated by a steeper increase in somatic EPA content with dietary EPA compared to 20 degrees C.
4. At 20 degrees C the development of eggs to successful hatching was high when EPA was supplied to the mothers. At 15 degrees C the hatching success was generally poor, despite of a higher maternal provision of EPA to eggs, compared to that at 20 degrees C, suggesting that EPA alone was insufficient for proper neonatal development at the low temperature. The growth of offspring from mothers raised at 20 degrees C without EPA supplementation was very low, indicating that the negative effects of EPA deficiency can be carried on to the next generation.
5. The fatty acid composition of Daphnia sp. in published field studies shows increasing proportions of saturated fatty acids with increasing environmental temperature, whereas omega 3 PUFAs and EPA show no clear pattern, suggesting that variations in dietary PUFA may mask temperature-dependent adjustments in omega 3 PUFA concentrations of cladocerans in nature.
1. Atmospheric nitrogen (N) deposition is expected to change forest understorey plant community composition and diversity, but results of experimental addition studies and observational studies are not yet conclusive. A shortcoming of observational studies, which are generally based on resurveys or sampling along large deposition gradients, is the occurrence of temporal or spatial confounding factors.
2. We were able to assess the contribution of N deposition versus other ecological drivers on forest understorey plant communities by combining a temporal and spatial approach. Data from 1205 (semi-)permanent vegetation plots taken from 23 rigorously selected understorey resurvey studies along a large deposition gradient across deciduous temperate forest in Europe were compiled and related to various local and regional driving factors, including the rate of atmospheric N deposition, the change in large herbivore densities and the change in canopy cover and composition.
3. Although no directional change in species richness occurred, there was considerable floristic turnover in the understorey plant community and a shift in species composition towards more shade-tolerant and nutrient-demanding species. However, atmospheric N deposition was not important in explaining the observed eutrophication signal. This signal seemed mainly related to a shift towards a denser canopy cover and a changed canopy species composition with a higher share of species with more easily decomposed litter.
4. Synthesis. Our multi-site approach clearly demonstrates that one should be cautious when drawing conclusions about the impact of atmospheric N deposition based on the interpretation of plant community shifts in single sites or regions due to other, concurrent, ecological changes. Even though the effects of chronically increased N deposition on the forest plant communities are apparently obscured by the effects of canopy changes, the accumulated N might still have a significant impact. However, more research is needed to assess whether this N time bomb will indeed explode when canopies will open up again.
Bacteriophages use specific tail proteins to recognize host cells. It is still not understood to molecular detail how the signal is transmitted over the tail to initiate infection. We have analysed in vitro DNA ejection in long-tailed siphovirus 9NA and short-tailed podovirus P22 upon incubation with Salmonella typhimurium lipopolysaccharide (LPS). We showed for the first time that LPS alone was sufficient to elicit DNA release from a siphovirus in vitro. Crystal structure analysis revealed that both phages use similar tailspike proteins for LPS recognition. Tailspike proteins hydrolyse LPS O antigen to position the phage on the cell surface. Thus we were able to compare in vitro DNA ejection processes from two phages with different morphologies with the same receptor under identical experimental conditions. Siphovirus 9NA ejected its DNA about 30 times faster than podovirus P22. DNA ejection is under control of the conformational opening of the particle and has a similar activation barrier in 9NA and P22. Our data suggest that tail morphology influences the efficiencies of particle opening given an identical initial receptor interaction event.
Mesenchymal stem cells and glioma cells form a structural as well as a functional syncytium in vitro
(2012)
The interaction of human mesenchymal stem cells (hMSCs) and tumor cells has been investigated in various contexts. HMSCs are considered as cellular treatment vectors based on their capacity to migrate towards a malignant lesion. However, concerns about unpredictable behavior of transplanted hMSCs are accumulating. In malignant gliomas, the recruitment mechanism is driven by glioma-secreted factors which lead to accumulation of both, tissue specific stem cells as well as bone marrow derived hMSCs within the tumor. The aim of the present work was to study specific cellular interactions between hMSCs and glioma cells in vitro. We show, that glioma cells as well as hMSCs differentially express connexins. and that they interact via gap-junctional coupling. Besides this so-called functional syncytium formation, we also provide evidence of cell fusion events (structural syncytium). These complex cellular interactions led to an enhanced migration and altered proliferation of both, tumor and mesenchymal stem cell types in vitro. The presented work shows that glioma cells display signs of functional as well as structural syncytium formation with hMSCs in vitro. The described cellular phenomena provide new insight into the complexity of interaction patterns between tumor cells and host cells. Based on these findings, further studies are warranted to define the impact of a functional or structural syncytium formation on malignant tumors and cell based therapies in vivo.
The enzyme penicillin G acylase (EC 3.5.1.11) catalyzes amide-bond cleavage in benzylpenicillin (penicillin G) to yield 6-aminopenicillanic acid, an intermediate chemical used in the production of semisynthetic penicillins. A thermostable penicillin G acylase from Alcaligenes faecalis (AfPGA) has been crystallized using the hanging-drop vapour-diffusion method in two different space groups: C2221, with unit-cell parameters a = 72.9, b = 86.0, c = 260.2 angstrom, and P41212, with unit-cell parameters a = b = 85.6, c = 298.8 angstrom. Data were collected at 293 K and the structure was determined using the molecular-replacement method. Like other penicillin acylases, AfPGA belongs to the N-terminal nucleophilic hydrolase superfamily, has undergone post-translational processing and has a serine as the N-terminal residue of the beta-chain. A disulfide bridge has been identified in the structure that was not found in the other two known penicillin G acylase structures. The presence of the disulfide bridge is perceived to be one factor that confers higher stability to this enzyme.
The global warming potential of nitrous oxide (N2O) and its long atmospheric lifetime mean its presence in the atmosphere is of major concern, and that methods are required to measure and reduce emissions. Large spatial and temporal variations means, however, that simple extrapolation of measured data is inappropriate, and that other methods of quantification are required. Although process-based models have been developed to simulate these emissions, they often require a large amount of input data that is not available at a regional scale, making regional and global emission estimates difficult to achieve. The spatial extent of organic soils means that quantification of emissions from these soil types is also required, but will not be achievable using a process-based model that has not been developed to simulate soil water contents above field capacity or organic soils. The ECOSSE model was developed to overcome these limitations, and with a requirement for only input data that is readily available at a regional scale, it can be used to quantify regional emissions and directly inform land-use change decisions. ECOSSE includes the major processes of nitrogen (N) turnover, with material being exchanged between pools of SOM at rates modified by temperature, soil moisture, soil pH and crop cover. Evaluation of its performance at site-scale is presented to demonstrate its ability to adequately simulate soil N contents and N2O emissions from cropland soils in Europe. Mitigation scenarios and sensitivity analyses are also presented to demonstrate how ECOSSE can be used to estimate the impact of future climate and land-use change on N2O emissions.
'Biodiversity' is increasingly perceived as an important resource for research and conservation, but also for economy. Conservation, access and sustainable use of biodiversity (genetic resources, species, samples) are negotiated on different political levels, resulting in an internationally binding legal framework. Resulting legislation is binding for all parties involved in biological sampling, i.e. researches and (and in italics) countries, and especially applies for tissue or DNA samples and dervied products thereof. Understanding and awareness of export and import permits for biological samples is increasingly important for biologists to perform research projects legally and timely. Nevertheless, some biologists are still exporting and importing biological samples ignoring or noncompliant with national and international legislation, conventions, and regulations. Resulting difficulties may not only cause serious problems during field work, but may also delay the export, import or exchange of samples. Comprehensive a priori information regarding legal requirements helps to avoid or at least diminish potential problems. We identified four major factors facilitating export/import permits: (1) good personal (mutually trusted) contacts in the country of origin, (2) understanding and compliance with all relevant laws and regulations; (3) access to information regarding knowledge on permits, regulations and laws including their circulation within the researcher communities; and (4) access to consistent and up to date regulations
Biotic homogenization, the decrease in beta diversity among formerly distinct species assemblages, has been recognized as an important form of biotic impoverishment for more than a decade. Although researchers have stressed the importance of the functional dimension to understand its potential ecological consequences, biotic homogenization has mostly been studied at a taxonomic level. Here, we explore the relationship between taxonomic and functional homogenization using data on temperate forest herb layer communities in NW Germany, for which taxonomic homogenization has recently been demonstrated. We quantified beta diversity by partitioning Rao's quadratic entropy. We found a general positive relationship between changes in taxonomic and functional beta diversity. This relationship was stronger if multiple functional traits were taken into account. Averaged across sites, however, taxonomic homogenization was not consistently accompanied by functional homogenization. Depending on the traits considered, taxonomic homogenization occurred also together with functional differentiation or no change in functional beta diversity. The species shifts responsible for changes in beta diversity differed substantially between taxonomic and functional beta diversity measures and also among functional beta diversity measures based on different traits. We discuss likely environmental drivers for species shifts. Our study demonstrates that functional homogenization must be explicitly studied as an independent phenomenon that cannot be inferred from taxonomic homogenization.
Dynamic regulatory on/off minimization for biological systems under internal temporal perturbations
(2012)
Background: Flux balance analysis (FBA) together with its extension, dynamic FBA, have proven instrumental for analyzing the robustness and dynamics of metabolic networks by employing only the stoichiometry of the included reactions coupled with adequately chosen objective function. In addition, under the assumption of minimization of metabolic adjustment, dynamic FBA has recently been employed to analyze the transition between metabolic states.
Results: Here, we propose a suite of novel methods for analyzing the dynamics of (internally perturbed) metabolic networks and for quantifying their robustness with limited knowledge of kinetic parameters. Following the biochemically meaningful premise that metabolite concentrations exhibit smooth temporal changes, the proposed methods rely on minimizing the significant fluctuations of metabolic profiles to predict the time-resolved metabolic state, characterized by both fluxes and concentrations. By conducting a comparative analysis with a kinetic model of the Calvin-Benson cycle and a model of plant carbohydrate metabolism, we demonstrate that the principle of regulatory on/off minimization coupled with dynamic FBA can accurately predict the changes in metabolic states.
Conclusions: Our methods outperform the existing dynamic FBA-based modeling alternatives, and could help in revealing the mechanisms for maintaining robustness of dynamic processes in metabolic networks over time.
Background: Detection of immunogenic proteins remains an important task for life sciences as it nourishes the understanding of pathogenicity, illuminates new potential vaccine candidates and broadens the spectrum of biomarkers applicable in diagnostic tools. Traditionally, immunoscreenings of expression libraries via polyclonal sera on nitrocellulose membranes or screenings of whole proteome lysates in 2-D gel electrophoresis are performed. However, these methods feature some rather inconvenient disadvantages. Screening of expression libraries to expose novel antigens from bacteria often lead to an abundance of false positive signals owing to the high cross reactivity of polyclonal antibodies towards the proteins of the expression host. A method is presented that overcomes many disadvantages of the old procedures.
Results: Four proteins that have previously been described as immunogenic have successfully been assessed immunogenic abilities with our method. One protein with no known immunogenic behaviour before suggested potential immunogenicity. We incorporated a fusion tag prior to our genes of interest and attached the expressed fusion proteins covalently on microarrays. This enhances the specific binding of the proteins compared to nitrocellulose. Thus, it helps to reduce the number of false positives significantly. It enables us to screen for immunogenic proteins in a shorter time, with more samples and statistical reliability. We validated our method by employing several known genes from Campylobacter jejuni NCTC 11168.
Conclusions: The method presented offers a new approach for screening of bacterial expression libraries to illuminate novel proteins with immunogenic features. It could provide a powerful and attractive alternative to existing methods and help to detect and identify vaccine candidates, biomarkers and potential virulence-associated factors with immunogenic behaviour furthering the knowledge of virulence and pathogenicity of studied bacteria.zeige weniger
Background: There are alternative substrates to the mitochondrial respiration.
Results: Data-driven model-based analysis renders predictions of alternative substrates to the mitochondrial respiration.
Conclusion: Metabolomics data in conjunction with flux-based models can discriminate among hypotheses based on enzymology alone.
Significance: This analysis provides a basic framework for in silico studies of alternative pathways in metabolism.
The large scale production of a monoclonal anti-progesterone antibody in serum free medium followed by affinity chromatography on protein G lead to a contamination of the antibody sample with a protein of about 14 kDa. This protein was identified by mass spectrometry as secretory leukocyte protease inhibitor (SLPI). This SLPI contamination lead to a failure of the fiber-optic based competitive fluorescence assay to detect progesterone in milk. Purification of the monoclonal antibody using protein A columns circumvented this problem.
Dietary lipid quality affects temperature-mediated reaction norms of a freshwater key herbivore
(2012)
Temperature-mediated plasticity in life history traits strongly affects the capability of ectotherms to cope with changing environmental temperatures. We hypothesised that temperature-mediated reaction norms of ectotherms are constrained by the availability of essential dietary lipids, i.e. polyunsaturated fatty acids (PUFA) and sterols, as these lipids are involved in the homeoviscous adaptation of biological membranes to changing temperatures. A life history experiment was conducted in which the freshwater herbivore Daphnia magna was raised at four different temperatures (10, 15, 20, 25A degrees C) with food sources differing in their PUFA and sterol composition. Somatic growth rates increased significantly with increasing temperature, but differences among food sources were obtained only at 10A degrees C at which animals grew better on PUFA-rich diets than on PUFA-deficient diets. PUFA-rich food sources resulted in significantly higher population growth rates at 10A degrees C than PUFA-deficient food, and the optimum temperature for offspring production was clearly shifted towards colder temperatures with an increased availability of dietary PUFA. Supplementation of PUFA-deficient food with single PUFA enabled the production of viable offspring and significantly increased population growth rates at 10A degrees C, indicating that dietary PUFA are crucial for the acclimation to cold temperatures. In contrast, cumulative numbers of viable offspring increased significantly upon cholesterol supplementation at 25A degrees C and the optimum temperature for offspring production was shifted towards warmer temperatures, implying that sterol requirements increase with temperature. In conclusion, essential dietary lipids significantly affect temperature-mediated reaction norms of ectotherms and thus temperature-mediated plasticity in life history traits is subject to strong food quality constraints.
Salivary gland cells of the blowfly Calliphora vicina have a vacuolar-type H+-ATPase (V-ATPase) that lies in their apical membrane and energizes the secretion of a KCl-rich primary saliva upon stimulation with serotonin (5-hydroxytryptamine). Whether and to what extent V-ATPase contributes to intracellular pH (pH(i)) regulation in unstimulated gland cells is unknown. We used the fluorescent dye BCECF to study intracellular pH(i) regulation microfluorometrically and show that: (1) under resting conditions, the application of Na+-free physiological saline induces an intracellular alkalinization attributable to the inhibition of the activity of a Na+-dependent glutamate transporter; (2) the maintenance of resting pHi is Na+, Cl-, concanamycin A and DIDS sensitive; (3) recovery from an intracellular acid load is Na+ sensitive and requires V-ATPase activity; (4) the Na+/H+ antiporter is not involved in pHi recovery after a NH4Cl prepulse; and (5) at least one Na+-dependent transporter and the V-ATPase maintain recovery from an intracellular acid load. Thus, under resting conditions, the V-ATPase and at least one Na+-dependent transporter maintain normal pH(i) values of pH.7.5. We have also detected the presence of a Na+-dependent glutamate transporter, which seems to act as an acid loader. Despite this not being a common pH(i)-regulating transporter, its activity affects steady-state pH(i) in C. vicina salivary gland cells.
The response of forest plant regeneration to temperature variation along a latitudinal gradient
(2012)
The response of forest herb regeneration from seed to temperature variations across latitudes was experimentally assessed in order to forecast the likely response of understorey community dynamics to climate warming.
Seeds of two characteristic forest plants (Anemone nemorosa and Milium effusum) were collected in natural populations along a latitudinal gradient from northern France to northern Sweden and exposed to three temperature regimes in growth chambers (first experiment). To test the importance of local adaptation, reciprocal transplants were also made of adult individuals that originated from the same populations in three common gardens located in southern, central and northern sites along the same gradient, and the resulting seeds were germinated (second experiment). Seedling establishment was quantified by measuring the timing and percentage of seedling emergence, and seedling biomass in both experiments.
Spring warming increased emergence rates and seedling growth in the early-flowering forb A. nemorosa. Seedlings of the summer-flowering grass M. effusum originating from northern populations responded more strongly in terms of biomass growth to temperature than southern populations. The above-ground biomass of the seedlings of both species decreased with increasing latitude of origin, irrespective of whether seeds were collected from natural populations or from the common gardens. The emergence percentage decreased with increasing home-away distance in seeds from the transplant experiment, suggesting that the maternal plants were locally adapted.
Decreasing seedling emergence and growth were found from the centre to the northern edge of the distribution range for both species. Stronger responses to temperature variation in seedling growth of the grass M. effusum in the north may offer a way to cope with environmental change. The results further suggest that climate warming might differentially affect seedling establishment of understorey plants across their distribution range and thus alter future understorey plant dynamics.
Organisms eating each other are only one of many types of well documented and important interactions among species. Other such types include habitat modification, predator interference and facilitation. However, ecological network research has been typically limited to either pure food webs or to networks of only a few (<3) interaction types. The great diversity of non-trophic interactions observed in nature has been poorly addressed by ecologists and largely excluded from network theory. Herein, we propose a conceptual framework that organises this diversity into three main functional classes defined by how they modify specific parameters in a dynamic food web model. This approach provides a path forward for incorporating non-trophic interactions in traditional food web models and offers a new perspective on tackling ecological complexity that should stimulate both theoretical and empirical approaches to understanding the patterns and dynamics of diverse species interactions in nature.
It has been predicted that Europe will experience a rise in temperature of 2.2-5.3 A degrees C within this century. This increase in temperature may lead to vegetation change along altitudinal gradients. To test whether vegetation composition has already changed in the recent decade due to current warming (and other concomitant environmental changes), we recorded plant species composition in 1995 and 2005/2006 in Swiss pre-alpine fen meadows (800-1,400 m a.s.l.). Despite no obvious changes in the management of these fens, overall, plant species richness (cumulative number of plant species at five plots per site) significantly increased over this period. This was mainly due to an increase in the number of thermophilous, rich-soil-indicator and shade-indicator species, which corresponded to increased community productivity and shading within the vegetation layer. In contrast, fen specialists significantly declined in species numbers. The strongest species shifts occurred at the lowest sites, which overall had a higher colonization rate by new species than did sites at higher altitudes. Vegetation change along the altitudinal gradient was also affected by different types of land management: early-flowering species and species with low habitat specificity had high colonization rates in grazed fens, especially at low altitudes.
Hidden diversity in diatoms of Kenyan Lake Naivasha a genetic approach detects temporal variation
(2012)
This study provides insights into the morphological and genetic diversity in diatoms occurring in core sediments from tropical lakes in Kenya. We developed a genetic survey technique specific for diatoms utilizing a short region (7667 bp) of the ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL) gene as genetic barcode. Our analyses (i) validated the use of rbcL as a barcoding marker for diatoms, applied to sediment samples, (ii) showed a significant correlation between the results obtained by morphological and molecular data and (iii) indicated temporal variation in diatom assemblages on the inter- and intra-specific level. Diatom assemblages from a short core from Lake Naivasha show a drastic shift over the last 200 years, as littoral species (e.g. Navicula) are replaced by more planktonic ones (e.g. Aulacoseira). Within that same period, we detected periodic changes in the respective frequencies of distinct haplotype groups of Navicula, which coincide with wet and dry periods of Lake Naivasha between 1820 and 1938 AD. Our genetic analyses on historical lake sediments revealed inter- and intra-specific variation in diatoms, which is partially hidden behind single morphotypes. The occurrence of particular genetic lineages is probably correlated with environmental factors.
Neuronal computation of object distance from echo delay is an essential task that echolocating bats must master for spatial orientation and the capture of prey. In the dorsal auditory cortex of bats, neurons specifically respond to combinations of short frequency-modulated components of emitted call and delayed echo. These delay-tuned neurons are thought to serve in target range calculation. It is unknown whether neuronal correlates of active space perception are established by experience-dependent plasticity or by innate mechanisms. Here we demonstrate that in the first postnatal week, before onset of echolocation and flight, dorsal auditory cortex already contains functional circuits that calculate distance from the temporal separation of a simulated pulse and echo. This innate cortical implementation of a purely computational processing mechanism for sonar ranging should enhance survival of juvenile bats when they first engage in active echolocation behaviour and flight.
Translatome and metabolome effects triggered by gibberellins during rosette growth in Arabidopsis
(2012)
Although gibberellins (GAs) are well known for their growth control function, little is known about their effects on primary metabolism. Here the modulation of gene expression and metabolic adjustment in response to changes in plant (Arabidopsis thaliana) growth imposed on varying the gibberellin regime were evaluated. Polysomal mRNA populations were profiled following treatment of plants with paclobutrazol (PAC), an inhibitor of GA biosynthesis, and gibberellic acid (GA(3)) to monitor translational regulation of mRNAs globally. Gibberellin levels did not affect levels of carbohydrates in plants treated with PAC and/or GA(3). However, the tricarboxylic acid cycle intermediates malate and fumarate, two alternative carbon storage molecules, accumulated upon PAC treatment. Moreover, an increase in nitrate and in the levels of the amino acids was observed in plants grown under a low GA regime. Only minor changes in amino acid levels were detected in plants treated with GA(3) alone, or PAC plus GA(3). Comparison of the molecular changes at the transcript and metabolite levels demonstrated that a low GA level mainly affects growth by uncoupling growth from carbon availability. These observations, together with the translatome changes, reveal an interaction between energy metabolism and GA-mediated control of growth to coordinate cell wall extension, secondary metabolism, and lipid metabolism.
Metabolic networks are characterized by complex interactions and regulatory mechanisms between many individual components. These interactions determine whether a steady state is stable to perturbations. Structural kinetic modeling (SKM) is a framework to analyze the stability of metabolic steady states that allows the study of the system Jacobian without requiring detailed knowledge about individual rate equations. Stability criteria can be derived by generating a large number of structural kinetic models (SK-models) with randomly sampled parameter sets and evaluating the resulting Jacobian matrices. Until now, SKM experiments applied univariate tests to detect the network components with the largest influence on stability. In this work, we present an extended SKM approach relying on supervised machine learning to detect patterns of enzyme-metabolite interactions that act together in an orchestrated manner to ensure stability. We demonstrate its application on a detailed SK-model of the Calvin-Benson cycle and connected pathways. The identified stability patterns are highly complex reflecting that changes in dynamic properties depend on concerted interactions between several network components. In total, we find more patterns that reliably ensure stability than patterns ensuring instability. This shows that the design of this system is strongly targeted towards maintaining stability. We also investigate the effect of allosteric regulators revealing that the tendency to stability is significantly increased by including experimentally determined regulatory mechanisms that have not yet been integrated into existing kinetic models.
Background: Flux coupling analysis (FCA) has become a useful tool in the constraint-based analysis of genome-scale metabolic networks. FCA allows detecting dependencies between reaction fluxes of metabolic networks at steady-state. On the one hand, this can help in the curation of reconstructed metabolic networks by verifying whether the coupling between reactions is in agreement with the experimental findings. On the other hand, FCA can aid in defining intervention strategies to knock out target reactions.
Results: We present a new method F2C2 for FCA, which is orders of magnitude faster than previous approaches. As a consequence, FCA of genome-scale metabolic networks can now be performed in a routine manner.
Conclusions: We propose F2C2 as a fast tool for the computation of flux coupling in genome-scale metabolic networks. F2C2 is freely available for non-commercial use at https://sourceforge.net/projects/f2c2/files/.
A lamin in lower eukaryotes?
(2012)
Lamins are the major components of the nuclear lamina and serve not only as a mechanical support, but are also involved in chromatin organization, epigenetic regulation, transcription and mitotic events. Despite these universal tasks, lamins have so far been found only in metazoans. Yet, recently we have identified Dictyostelium NE81 as the first lamin-like protein in a lower eukaryote. Based on the current knowledge, we draw a model for nuclear envelope organization in Dictyostelium in this Extra View and we review the experimental data that justified this classification. Furthermore we provide unpublished data underscoring the requirement of posttranslational CaaX-box processing for proper protein localization at the nuclear envelope. Sequence comparison of NE81 sequences from four Dictyostelia with bona fide lamins illustrates the evolutional relationship between these proteins. Under certain conditions these usually unicellular social amoebae congregate to form a multicellular body. We propose that the evolution of the lamin-like NE81 went along with the invention of multicellularity.
Aldehyde oxidase (AO) is a complex molybdo-flavoprotein that belongs to the xanthine oxidase family. AO is active as a homodimer, and each 150-kDa monomer binds two distinct [2Fe2S] clusters, FAD, and the molybdenum cofactor. AO has an important role in the metabolism of drugs based on its broad substrate specificity oxidizing aromatic aza-heterocycles, for example, N-1-methylnicotinamide and N-methylphthalazinium, or aldehydes, such as benzaldehyde, retinal, and vanillin. Sequencing the 35 coding exons of the human AOX1 gene in a sample of 180 Italian individuals led to the identification of relatively frequent, synonymous, missense and nonsense single-nucleotide polymorphisms (SNPs). Human aldehyde oxidase (hAOX1) was purified after heterologous expression in Escherichia coli. The recombinant protein was obtained with a purity of 95% and a yield of 50 mu g/l E. coli culture. Site-directed mutagenesis of the hAOX1 cDNA allowed the purification of protein variants bearing the amino acid changes R802C, R921H, N1135S, and H1297R, which correspond to some of the identified SNPs. The hAOX1 variants were purified and compared with the wild-type protein relative to activity, oligomerization state, and metal content. Our data show that the mutation of each amino acid residue has a variable impact on the ability of hAOX1 to metabolize selected substrates. Thus, the human population is characterized by the presence of functionally inactive hAOX1 allelic variants as well as variants encoding enzymes with different catalytic activities. Our results indicate that the presence of these allelic variants should be considered for the design of future drugs.
Habitat loss poses a severe threat to biodiversity. While many studies yield valuable information on how specific species cope with such environmental modification, the mechanistic understanding of how interacting species or whole communities are affected by habitat loss is still poor. Individual movement plays a crucial role for the space use characteristics of species, since it determines how individuals perceive and use their heterogeneous environment. At the community level, it is therefore essential to include individual movement and how it is influenced by resource sharing into the investigation of consequences of habitat loss. To elucidate the effects of foraging movement on communities in face of habitat loss, we here apply a recently published spatially-explicit and individual-based model of home range formation. This approach allows predicting the individual size distribution (ISD) of mammal communities in simulation landscapes that vary in the amount of suitable habitat. We apply three fundamentally different foraging movement approaches (central place forager (CPF), patrolling forager (PF) and body mass dependent nomadic forager (BNF)). Results show that the efficiency of the different foraging strategies depends on body mass, which again affects community structure in face of habitat loss. CPF is only efficient for small animals, and therefore yields steep ISD exponents on which habitat loss has little effect (due to a movement limitation of body mass). PF and particularly BNF are more efficient for larger animals, resulting in less steep ISDs with higher mass maxima, both showing a threshold behaviour with regard to loss of suitable habitat. These findings represent a new way of explaining observed extinction thresholds, and therefore indicate the importance of individual space use characterized by physiology and behaviour, i.e. foraging movement, for communities and their response to habitat loss. Findings also indicate the necessity to incorporate the crucial role of movement into future conservation efforts of terrestrial communities.
Biodetection formats, such as DNA and antibody microarrays, are valuable tools in the life sciences, but for some applications, the detection limits are insufficient. A straightforward strategy to obtain signal amplification is the rolling circle amplification (RCA), an easy, isothermal, and enzymatic nucleic acid synthesis that has already been employed successfully to increase the signal yield for several single-analyte and multiplexing assays in conjunction with hybridization probes. Here, we systematically investigated the parameters responsible for the RCA driven signal amplification with fluorescent labels, such as the type of fluorophore chosen, labeling strategy, composition of reaction solution, and number of handling steps. In labeling strategies, post-synthetic labeling via a Cy3-hybridization probe was compared to the direct incorporation of fluorescent Cy3-dUTP and DY-555-dUTP into the nascent strand during synthesis. With our direct labeling protocol, the assay's runtime and handling steps could be reduced while the signal yield was increased. These features are very attractive for many detection formats but especially for point-of-care diagnostic kits that need to be simple enough to be performed by scientifically untrained personnel.
The present study established the way in which plastidial alpha-glucan phosphorylase (Pho1) synthesizes maltodextrin (MD) which can be the primer for starch biosynthesis in rice endosperm. The synthesis of MD by Pho1 was markedly accelerated by branching enzyme (BE) isozymes, although the greatest effect was exhibited by the presence of branching isozyme I (BEI) rather than by isozyme IIa (BEIIa) or isozyme IIb (BEIIb). The enhancement of the activity of Pho1 by BE was not merely due to the supply of a non-reducing ends. At the same time, Pho1 greatly enhanced the BE activity, possibly by generating a branched carbohydrate substrate which is used by BE with a higher affinity. The addition of isoamylase to the reaction mixture did not prevent the concerted action of Pho1 and BEI. Furthermore, in the product, the branched structure was, at least to some extent, maintained. Based on these results we propose that the interaction between Pho1 and BE is not merely due to chain-elongating and chain-branching reactions, but occurs in a physically and catalytically synergistic manner by each activating the mutual capacity of the other, presumably forming a physical association of Pho1, BEI and branched MDs. This close interaction might play a crucial role in the synthesis of branched MDs and the branched MDs can act as a primer for the biosynthesis of amylopectin molecules.
Parenchyma cells from tubers of Solanum tuberosum L. convert several externally supplied sugars to starch but the rates vary largely. Conversion of glucose 1-phosphate to starch is exceptionally efficient. In this communication, tuber slices were incubated with either of four solutions containing equimolar [U-C-14]glucose 1-phosphate, [U-C-14]sucrose, [U-C-14]glucose 1-phosphate plus unlabelled equimolar sucrose or [U-C-14]sucrose plus unlabelled equimolar glucose 1-phosphate. C-14-incorporation into starch was monitored. In slices from freshly harvested tubers each unlabelled compound strongly enhanced C-14 incorporation into starch indicating closely interacting paths of starch biosynthesis. However, enhancement disappeared when the tubers were stored. The two paths (and, consequently, the mutual enhancement effect) differ in temperature dependence. At lower temperatures, the glucose 1-phosphate-dependent path is functional, reaching maximal activity at approximately 20 degrees C but the flux of the sucrose-dependent route strongly increases above 20 degrees C. Results are confirmed by in vitro experiments using [U-C-14]glucose 1-phosphate or adenosine-[U-C-14]glucose and by quantitative zymograms of starch synthase or phosphorylase activity. In mutants almost completely lacking the plastidial phosphorylase isozyme(s), the glucose 1-phosphate-dependent path is largely impeded. Irrespective of the size of the granules, glucose 1-phosphate-dependent incorporation per granule surface area is essentially equal. Furthermore, within the granules no preference of distinct glucosyl acceptor sites was detectable. Thus, the path is integrated into the entire granule biosynthesis. In vitro C-14-incorporation into starch granules mediated by the recombinant plastidial phosphorylase isozyme clearly differed from the in situ results. Taken together, the data clearly demonstrate that two closely but flexibly interacting general paths of starch biosynthesis are functional in potato tuber cells.
To identify genetic variants associated with head circumference in infancy, we performed a meta-analysis of seven genome-wide association studies (GWAS) (N = 10,768 individuals of European ancestry enrolled in pregnancy and/or birth cohorts) and followed up three lead signals in six replication studies (combined N = 19,089). rs7980687 on chromosome 12q24 (P = 8.1 x 10(-9)) and rs1042725 on chromosome 12q15 (P = 2.8 x 10(-10)) were robustly associated with head circumference in infancy. Although these loci have previously been associated with adult height(1), their effects on infant head circumference were largely independent of height (P = 3.8 x 10(-7) for rs7980687 and P = 1.3 x 10(-7) for rs1042725 after adjustment for infant height). A third signal, rs11655470 on chromosome 17q21, showed suggestive evidence of association with head circumference (P = 3.9 x 10(-6)). SNPs correlated to the 17q21 signal have shown genome-wide association with adult intracranial volume(2), Parkinson's disease and other neurodegenerative diseases(3-5), indicating that a common genetic variant in this region might link early brain growth with neurological disease in later life.
We tested the utility of a 230 base pair intron fragment of the highly conserved nuclear gene Elongation Factor 1-alpha (EF1-alpha) as a proper marker to reconstruct the phylogeography of the marine amphipod Pontogammarus maeoticus (Sowinsky, 1894) from the Caspian and Black Seas. As a prerequisite for further analysis, we confirmed by Southern blot analysis that EF1-alpha is encoded at a single locus in P. maeoticus. We included 15 populations and 60 individuals in the study. Both the phylogeny of the 27 unique alleles found and population genetic analyses revealed a significant differentiation between populations from the aforementioned sea basins. Our results are in remarkable agreement with recent studies on a variety of species from the same area, which invariably support a major phylogeographic break between the Caspian and Black Seas. We thus conclude that our EF1-alpha intron is an informative marker for phylogeographic studies in amphipods at the shallow population level.
Using the carcass sinking rate and density determined in laboratory for several freshwater zooplankton species, we developed a model of zooplankton carcass sinking as affected by turbulence and stratification. The model was subsequently used to estimate the residence time of zooplankton carcasses in the water column of Lake Stechlin, a typical temperate dimictic lake in northeastern Germany. The residence time varied among the different species and was strongly affected by thermal stratification. At the peak of summer stratification, the carcasses stayed up to 5 d in the 70 m-deep water column before reaching the lake bottom. Residence time was long enough that zooplankton carcasses could serve as an important matter and energy source for bacteria in the lake's pelagic zone and hence have the potential to significantly affect aquatic carbon and nutrient cycling. The proposed model of sinking rates, based on physically sound relationships, can be easily applied to other passively sinking particles, and be integrated into large ecosystem models.
We studied two pathways that involve the transfer of persulfide sulfur in humans, molybdenum cofactor biosynthesis and tRNA thiolation. Investigations using human cells showed that the two-domain protein MOCS3 is shared between both pathways. MOCS3 has an N-terminal adenylation domain and a C-terminal rhodanese-like domain. We showed that MOCS3 activates both MOCS2A and URM1 by adenylation and a subsequent sulfur transfer step for the formation of the thiocarboxylate group at the C terminus of each protein. MOCS2A and URM1 are beta-grasp fold proteins that contain a highly conserved C-terminal double glycine motif. The role of the terminal glycine of MOCS2A and URM1 was examined for the interaction and the cellular localization with MOCS3. Deletion of the C-terminal glycine of either MOCS2A or URM1 resulted in a loss of interaction with MOCS3. Enhanced cyan fluorescent protein and enhanced yellow fluorescent protein fusions of the proteins were constructed, and the fluorescence resonance energy transfer efficiency was determined by the decrease in the donor lifetime. The cellular localization results showed that extension of the C terminus with an additional glycine of MOCS2A and URM1 altered the localization of MOCS3 from the cytosol to the nucleus.
In this study, we report the genetic population structure of the Fire-bellied toad Bombina bombina in Brandenburg (East Germany) in the context of conservation. We analysed 298 samples originating from 11 populations in Brandenburg using mitochondrial control region sequences and six polymorphic microsatellite loci. For comparison, we included one population each from Poland and Ukraine into our analysis. Within Brandenburg, we detected a moderate variability in the mitochondrial control region (19 different haplotypes) and at microsatellite loci (9-12 alleles per locus). These polymorphisms revealed a clear population structure among toads in Brandenburg, despite a relatively high overall population density and the moderate size of single populations (100-2000 individuals). The overall genetic population structure is consistent with a postglacial colonization from South East-Europe and a subsequent population expansion. Based on genetic connectivity, we infer Management Units (MUs) as targets for conservation. Our genetic survey identified MUs, within which human infrastructure is currently preventing any genetic exchange. We also detect an unintentional translocation from South East to North West Brandenburg, presumably in the course of fish stocking activities. Provided suitable conservation measures are taken, Brandenburg should continue to harbor large populations of this critically endangered species.
Mechanistic understanding of consumer-resource dynamics is critical to predicting the effects of global change on ecosystem structure, function and services. Such understanding is severely limited by mechanistic models inability to reproduce the dynamics of multiple populations interacting in the field. We surpass this limitation here by extending general consumer-resource network theory to the complex dynamics of a specific ecosystem comprised by the seasonal biomass and production patterns in a pelagic food web of a large, well-studied lake. We parameterised our allometric trophic network model of 24 guilds and 107 feeding relationships using the lakes food web structure, initial spring biomasses and body-masses. Adding activity respiration, the detrital loop, minimal abiotic forcing, prey resistance and several empirically observed rates substantially increased the model's fit to the observed seasonal dynamics and the size-abundance distribution. This process illuminates a promising approach towards improving food-web theory and dynamic models of specific habitats.
Fourteen microsatellite markers were isolated and characterized for the endangered Visayan tarictic hornbill (Penelopides panini, Aves: Bucerotidae). In an analysis of 76 individuals, the number of alleles per locus varied from one to 12. Expected and observed heterozygosity ranged from 0.00 to 0.87 and from 0.00 to 0.89, respectively. All primers also amplify microsatellite loci in Luzon tarictic hornbill (Penelopides manillae), Mindanao tarictic hornbill (Penelopides affinis), the critically endangered Walden's hornbill (Aceros waldeni) and the near-threatened writhed hornbill (Aceros leucocephalus). Two loci which are monomorphic in P. panini were found polymorphic in at least one of the other species. These 14 new microsatellite markers specifically developed for two genera of Philippine hornbills, in combination with those already available for the hornbill genera Buceros and Bucorvus, comprise a reasonable number of loci to genetically analyse wild and captive populations of these and probably other related, often endangered hornbills.
The plant genome encodes at least two distinct and evolutionary conserved plastidial starch-related dikinases that phosphorylate a low percentage of glucosyl residues at the starch granule surface. Esterification of starch favours the transition of highly ordered a-glucans to a less ordered state and thereby facilitates the cleavage of interglucose bonds by hydrolases. Metabolically most important is the phosphorylation at position C6, which is catalysed by the glucan, water dikinase (GWD). The reactions mediated by recombinant wild-type GWD from Arabidopsis thaliana (AtGWD) and from Solanum tuberosum (StGWD) were studied. Two mutated proteins lacking the conserved histidine residue that is indispensible for glucan phosphorylation were also included. The wild-type GWDs consume approximately 20% more ATP than is required for glucan phosphorylation. Similarly, although incapable of phosphorylating a-glucans, the two mutated dikinase proteins are capable of degrading ATP. Thus, consumption of ATP and phosphorylation of a-glucans are not strictly coupled processes but, to some extent, occur as independent phosphotransfer reactions. As revealed by incubation of the GWDs with [gamma-33P]ATP, the consumption of ATP includes the transfer of the gamma-phosphate group to the GWD protein but this autophosphorylation does not require the conserved histidine residue. Thus, the GWD proteins possess two vicinal phosphorylation sites, both of which are transiently phosphorylated. Following autophosphorylation at both sites, native dikinases flexibly use various terminal phosphate acceptors, such as water, alpha-glucans, AMP and ADP. A model is presented describing the complex phosphotransfer reactions of GWDs as affected by the availability of the various acceptors.
The reasons for the apparent dominance of the toxic cyanobacterium Microcystis sp., reflected by its massive blooms in many fresh water bodies, are poorly understood. We show that in addition to a large array of secondary metabolites, some of which are toxic to eukaryotes, Microcystis sp. secretes large amounts of fibrous exopolysaccharides that form extremely long fibres several millimetres in length. This phenomenon was detected in field and laboratory cultures of various Microcystis strains. In addition, we have identified and characterized three of the proteins associated with the fibres and the genes encoding them in Microcystis sp. PCC 7806 but were unable to completely delete them from its genome. Phylogenetic analysis of the most abundant one, designated IPF-469, showed its presence only in cyanobacteria. Its closest relatives were detected in Synechocystis sp. PCC 6803 and in Cyanothece sp. strains; in the latter the genomic organization of the IPF-469 was highly conserved. IPF-469 and the other two proteins identified here, a haloperoxidase and a haemolysin-type calcium-binding protein, may be part of the fibres secretion pathway. The biological role of the fibres in Microcystis sp. is discussed.
The centrosome consists of a layered core structure surrounded by a microtubule-nucleating corona. A tight linkage through the nuclear envelope connects the cytosolic centrosome with the clustered centromeres within the nuclear matrix. At G2/M the corona dissociates, and the core structure duplicates, yielding two spindle poles. CP148 is a novel coiled coil protein of the centrosomal corona. GFP-CP148 exhibited cell cycle-dependent presence and absence at the centrosome, which correlates with dissociation of the corona in prophase and its reformation in late telophase. During telophase, GFP-CP148 formed cytosolic foci, which coalesced and joined the centrosome. This explains the hypertrophic appearance of the corona upon strong overexpression of GFP-CP148. Depletion of CP148 by RNAi caused virtual loss of the corona and disorganization of interphase microtubules. Surprisingly, formation of the mitotic spindle and astral microtubules was unaffected. Thus, microtubule nucleation complexes associate with centrosomal core components through different means during interphase and mitosis. Furthermore, CP148 RNAi caused dispersal of centromeres and altered Sun1 distribution at the nuclear envelope, suggesting a role of CP148 in the linkage between centrosomes and centromeres. Taken together, CP148 is an essential factor for the formation of the centrosomal corona, which in turn is required for centrosome/centromere linkage.
The response of species diversity to dispersal capability is inherently scale-dependent: increasing dispersal capability is expected to increase diversity at the local scale, while decreasing diversity at the metacommunity scale. However, these expectations are based on model formulations that neglect dispersal limitation and species segregation at the local scale. We developed a unifying framework of dispersaldiversity relationships and tested the generality of these expectations. For this purpose we used a spatially-explicit neutral model with various combinations of survey area (local scale) and landscape size (metacommunity scale). Simulations were conducted using landscapes of finite and of conceptually infinite size. We analyzed the scale-dependence of dispersal-diversity relationships for exponentially-bounded versus fat-tailed dispersal kernels, several levels of speciation rate and contrasting assumptions on recruitment at short dispersal distances. We found that the ratio of survey area to landscape size is a major determinant of dispersaldiversity relationships. With increasing survey-to-landscape area ratio the dispersaldiversity relationship switches from monotonically increasing through a U-shaped pattern (with a local minimum) to a monotonically decreasing pattern. Therefore, we provide a continuous set of dispersaldiversity relationships, which contains the response shapes reported previously as extreme cases. We suggest the mean dispersal distance with the minimum of species diversity (minimizing dispersal distance) for a certain scenario as a key characteristic of dispersaldiversity relationships. We show that not only increasing mean dispersal distances, but also increasing variances of dispersal can enhance diversity at the local scale, given a diverse species pool at the metacommunity scale. In conclusion, the response of diversity to variations of dispersal capability at spatial scales of interest, e.g. conservation areas, can differ more widely than expected previously. Therefore, land use and conservation activities, which manipulate dispersal capability, need to consider the landscape context and potential species pools carefully.
The most crucial step in data processing from high-throughput sequencing applications is the accurate and sensitive alignment of the sequencing reads to reference genomes or transcriptomes. The accurate detection of insertions and deletions (indels) and errors introduced by the sequencing platform or by misreading of modified nucleotides is essential for the quantitative processing of the RNA-based sequencing (RNA-Seq) datasets and for the identification of genetic variations and modification patterns. We developed a new, fast and accurate algorithm for nucleic acid sequence analysis, FANSe, with adjustable mismatch allowance settings and ability to handle indels to accurately and quantitatively map millions of reads to small or large reference genomes. It is a seed-based algorithm which uses the whole read information for mapping and high sensitivity and low ambiguity are achieved by using short and non-overlapping reads. Furthermore, FANSe uses hotspot score to prioritize the processing of highly possible matches and implements modified Smith-Watermann refinement with reduced scoring matrix to accelerate the calculation without compromising its sensitivity. The FANSe algorithm stably processes datasets from various sequencing platforms, masked or unmasked and small or large genomes. It shows a remarkable coverage of low-abundance mRNAs which is important for quantitative processing of RNA-Seq datasets.
Background: Female sperm storage has evolved independently multiple times among vertebrates to control reproduction in response to the environment. In internally fertilising amphibians, female salamanders store sperm in cloacal spermathecae, whereas among anurans sperm storage in oviducts is known only in tailed frogs. Facilitated through extensive field sampling following historical observations we tested for sperm storing structures in the female urogenital tract of fossorial, tropical caecilian amphibians.
Findings: In the oviparous Ichthyophis cf. kohtaoensis, aggregated sperm were present in a distinct region of the posterior oviduct but not in the cloaca in six out of seven vitellogenic females prior to oviposition. Spermatozoa were found most abundantly between the mucosal folds. In relation to the reproductive status decreased amounts of sperm were present in gravid females compared to pre-ovulatory females. Sperm were absent in females past oviposition.
Conclusions: Our findings indicate short-term oviductal sperm storage in the oviparous Ichthyophis cf. kohtaoensis. We assume that in female caecilians exhibiting high levels of parental investment sperm storage has evolved in order to optimally coordinate reproductive events and to increase fitness.
Complex networks have been successfully employed to represent different levels of biological systems, ranging from gene regulation to protein-protein interactions and metabolism. Network-based research has mainly focused on identifying unifying structural properties, such as small average path length, large clustering coefficient, heavy-tail degree distribution and hierarchical organization, viewed as requirements for efficient and robust system architectures. However, for biological networks, it is unclear to what extent these properties reflect the evolutionary history of the represented systems. Here, we show that the salient structural properties of six metabolic networks from all kingdoms of life may be inherently related to the evolution and functional organization of metabolism by employing network randomization under mass balance constraints. Contrary to the results from the common Markov-chain switching algorithm, our findings suggest the evolutionary importance of the small-world hypothesis as a fundamental design principle of complex networks. The approach may help us to determine the biologically meaningful properties that result from evolutionary pressure imposed on metabolism, such as the global impact of local reaction knockouts. Moreover, the approach can be applied to test to what extent novel structural properties can be used to draw biologically meaningful hypothesis or predictions from structure alone.
The stability of proteins is paramount for their therapeutic and industrial use and, thus, is a major task for protein engineering. Several types of chemical and physical stabilities are desired, and discussion revolves around whether each stability trait needs to be addressed separately and how specific and compatible stabilizing mutations act. We demonstrate a stepwise perturbation-compensation strategy, which identifies mutations rescuing the activity of a truncated TEM beta-lactamase. Analyses relating structural stress with the external stresses of heat, denaturants, and proteases reveal our second-site suppressors as general stability centers that also improve the full-length enzyme. A library of lactamase variants truncated by 15 N-terminal and three C-terminal residues (Bla-N Delta 15C Delta 3) was subjected to activity selection and DNA shuffling. The resulting clone with the best in vivo performance harbored eight mutations, surpassed the full-length wild-type protein by 5.3 degrees C in T-m, displayed significantly higher catalytic activity at elevated temperatures, and showed delayed guanidine-induced denaturation. The crystal structure of this mutant was determined and provided insights into its stability determinants. Stepwise reconstitution of the N- and C-termini increased its thermal, denaturant, and proteolytic resistance successively, leading to a full-length enzyme with a T-m increased by 15.3 degrees C and a half-denaturation concentration shifted from 0.53 to 1.75 M guanidinium relative to that of the wild type. These improvements demonstrate that iterative truncation-optimization cycles can exploit stability-trait linkages in proteins and are exceptionally suited for the creation of progressively stabilized variants and/or downsized proteins without the need for detailed structural or mechanistic information.
1. The polyunsaturated fatty acid eicosapentaenoic acid (EPA) plays an important role in aquatic food webs, in particular at the primary producerconsumer interface where keystone species such as daphnids may be constrained by its dietary availability. Such constraints and their seasonal and interannual changes may be detected by continuous measurements of EPA concentrations. However, such EPA measurements became common only during the last two decades, whereas long-term data sets on plankton biomass are available for many well-studied lakes. Here, we test whether it is possible to estimate EPA concentrations from abiotic variables (light and temperature) and the biomass of prey organisms (e.g. ciliates, diatoms and cryptophytes) that potentially provide EPA for consumers. 2. We used multiple linear regression to relate size- and taxonomically resolved plankton biomass data and measurements of temperature and light intensity to directly measured EPA concentrations in Lake Constance during a whole year. First, we tested the predictability of EPA concentrations from the biomass of EPA-rich organisms (diatoms, cryptophytes and ciliates). Secondly, we included the variables mean temperature and mean light intensity over the sampling depth (020 m) and depth (08 and 820 m) as factors in our model to check for large-scale seasonal- and depth-dependent effects on EPA concentrations. In a third step, we included the deviations of light and temperature from mean values in our model to allow for their potential influence on the biochemical composition of plankton organisms. We used the Akaike Information Criterion to determine the best models. 3. All approaches supported our proposition that the biomasses of specific plankton groups are variables from which seston EPA concentrations can be derived. The importance of ciliates as an EPA source in the seston was emphasised by their high weight in our models, although ciliates are neglected in most studies that link fatty acids to seston taxonomic composition. The large-scale seasonal variability of light intensity and its interaction with diatom biomass were significant predictors of EPA concentrations. The deviation of temperature from mean values, accounting for a depth-dependent effect on EPA concentrations, and its interaction with ciliate biomass were also variables with high predictive power. 4. The best models from the first and second approaches were validated with measurements of EPA concentrations from another year (1997). The estimation with the best model including only biomass explained 80%, and the best model from the second approach including mean temperature and depth explained 87% of the variability in EPA concentrations in 1997. 5. We show that it is possible to predict EPA concentrations reliably from plankton biomass, while the inclusion of abiotic factors led to results that were only partly consistent with expectations from laboratory studies. Our approach of including biotic predictors should be transferable to other systems and allow checking for biochemical constraints on primary consumers.
In the deep, cooler layers of clear, nutrient-poor, stratified water bodies, phytoplankton often accumulate to form a thin band or "deep chlorophyll maximum" (DCM) of ecological importance. Under such conditions, these photosynthetic microorganisms may be close to their physiological compensation points and to the boundaries of their ecological tolerance. To grow and survive any resulting energy limitation, DCM species are thought to exhibit highly specialised or flexible acclimation strategies. In this study, we investigated several of the adaptable ecophysiological strategies potentially employed by one such species, Chlamydomonas acidophila: a motile, unicellular, phytoplanktonic flagellate that often dominates the DCM in stratified, acidic lakes. Physiological and behavioural responses were measured in laboratory experiments and were subsequently related to field observations. Results showed moderate light compensation points for photosynthesis and growth at 22A degrees C, relatively low maintenance costs, a behavioural preference for low to moderate light, and a decreased compensation point for photosynthesis at 8A degrees C. Even though this flagellated alga exhibited a physiologically mediated diel vertical migration in the field, migrating upwards slightly during the day, the ambient light reaching the DCM was below compensation points, and so calculations of daily net photosynthetic gain showed that survival by purely autotrophic means was not possible. Results suggested that strategies such as low-light acclimation, small-scale directed movements towards light, a capacity for mixotrophic growth, acclimation to low temperature, in situ exposure to low O-2, high CO2 and high P concentrations, and an avoidance of predation, could combine to help overcome this energetic dilemma and explain the occurrence of the DCM. Therefore, corroborating the deceptive ecophysiological complexity of this and similar organisms, only a suite of complementary strategies can facilitate the survival of C. acidophila in this DCM.