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We investigated the effects of temperature (4 degrees C, 8 degrees C, and 12 degrees C) on structural and storage dynamics, as measured by changes in fatty acids (FA) associated with cell membrane phospholipids (PL) and triacylglycerols (TAG), respectively, as well as on body weight and survival of a freshwater calanoid copepod (Eudiaptomus gracilis) during fasting (10 d) and refeeding (10 d) with two algae of differing nutritional quality (Cryptomonas ozolinii and Scenedesmus obliquus). Fasting led to 50% loss in body weight, a near total depletion of TAG, and a drastic decrease of the polyunsaturated FA (PUFA) in TAG and PL, indicating their preferential utilization and alterations in membrane function, respectively. Higher temperatures accelerated the decrease of body weight and of PUFA in PL and TAG, and decreased survival. After 10 d of refeeding, copepods partially recovered their initial lipid stores and cell membrane composition. The effects of food quality were temperature dependent: Cryptomonas promoted better recovery (i.e., return to or close to the levels at the beginning of the experiment) of both body weight and TAG at only the two higher temperatures (8 degrees C and 12 degrees C), whereas no recovery was observed at 4 degrees C. Higher temperatures and refeeding on Cryptomonas also had a positive, but minor, influence on the recovery of membrane FA composition. Survival differed among treatments but was lowest at the intermediate temperature (8 degrees C) for both diets. We conclude that temperature changes on the order of 4-8 degrees C significantly influence TAG and PL during fasting periods and interact with food quality to determine the extent of recovery in copepod lipids.
A rise in temperature will intensify the feeding links involving ectotherms in food webs. However, it is unclear how the effects will quantitatively differ between the plant-herbivore and herbivore-carnivore interface. To test how warming could differentially affect rates of herbivory and carnivory, we studied trophic interaction strength in a food chain comprised of green algae, herbivorous rotifers and carnivorous rotifers at 10, 15, 20 and 25 degrees C. We found significant warming-induced changes in feeding by both herbivorous and carnivorous rotifers, but these responses occurred at different parts of the entire temperature gradient. The strongest response of the per capita herbivore's ingestion rate occurred due to an increase in temperature from 15 to 20 degrees C (1.9 fold: from 834 to 1611 algal cells per h(-1)) and of the per capita carnivore's ingestion rate from 20 to 25 degrees C (1.6 fold: from 1.5 to 2.5 prey h(-1)). Handling time, an important component of a consumer's functional response, significantly decreased from 15 to 20 degrees C in herbivorous rotifers. In contrast, it decreased from 20 to 25 degrees C in carnivorous rotifers. Attack rates significantly and strongly increased from 10 to 25 degrees C in the herbivorous animals, but not at all in the carnivores. Our results exemplify how the relative forces of top-down control exerted by herbivores and carnivores may strongly shift under global warming. But warming, and its magnitude, are not the only issue: If our results would prove to be representative, shifts in ectotherm interactions will quantitatively differ when a 5 degrees C increase starts out from a low, intermediate or high initial temperature. This would imply that warming could have different effects on the relative forces of carnivory and herbivory in habitats differing in average temperature, as would exist at different altitudes and latitudes.
VMP1-deficient Chlamydomonas exhibits severely aberrant cell morphology and disrupted cytokinesies
(2014)
Background: The versatile Vacuole Membrane Protein 1 (VMP1) has been previously investigated in six species. It has been shown to be essential in macroautophagy, where it takes part in autophagy initiation. In addition, VMP1 has been implicated in organellar biogenesis; endo-, exo- and phagocytosis, and protein secretion; apoptosis; and cell adhesion. These roles underly its proven involvement in pancreatitis, diabetes and cancer in humans.
Results: In this study we analyzed a VMP1 homologue from the green alga Chlamydomonas reinhardtii. CrVMP1 knockdown lines showed severe phenotypes, mainly affecting cell division as well as the morphology of cells and organelles. We also provide several pieces of evidence for its involvement in macroautophagy.
Correct assessment of risks and costs of foraging is vital for the fitness of foragers. Foragers should avoid predation risk and balance missed opportunities. In risk-heterogeneous landscapes animals prefer safer locations over riskier, constituting a landscape of fear. Risk-uniform landscapes do not offer this choice, all locations are equally risky. Here we investigate the effects of predation risk in patches, travelling risk between patches, and missed social opportunities on foraging decisions in risk-uniform and risk-heterogeous landscapes. We investigated patch leaving decisions of 20 common voles (M. arvalis) in three experimental landscapes: safe risk-uniform, risky risk-uniform and risk-heterogeneous. We varied both the predation risk level and the predation risk distribution between two patches experimentally and in steps, assuming that our manipulation consequently yield different distributions and levels of risk while foraging, risk while travelling, and costs of missed, social opportunities (MSOCs). We measured mean GUDs (giving-up density of food left in the patch) for both patches as a measure of foraging gain, and delta GUD, the differences among patches, as a measure of the spatial distribution of foraging effort over a period of six hours. Distribution of foraging effort was most even in the safe risk-uniform landscapes and least even in the risk-heterogeneous landscape, with risky risk-uniform landscapes in between. Foraging gain was higher in the safe than in the two riskier landscapes (both uniform and heterogeneous). Results supported predictions for the effects of risk in foraging patches and while travelling between patches, however predictions for the effects of missed social opportunities were not met in this short term experiment. Thus, both travelling and foraging risk contribute to distinct patterns observable high risk, risk-uniform landscapes.
Background: The forelimb-specific gene tbx5 is highly conserved and essential for the development of forelimbs in zebrafish, mice, and humans. Amongst birds, a single order, Dinornithiformes, comprising the extinct wingless moa of New Zealand, are unique in having no skeletal evidence of forelimb-like structures.
Results: To determine the sequence of tbx5 in moa, we used a range of PCR-based techniques on ancient DNA to retrieve all nine tbx5 exons and splice sites from the giant moa, Dinornis. Moa Tbx5 is identical to chicken Tbx5 in being able to activate the downstream promotors of fgf10 and ANF. In addition we show that missexpression of moa tbx5 in the hindlimb of chicken embryos results in the formation of forelimb features, suggesting that Tbx5 was fully functional in wingless moa. An alternatively spliced exon 1 for tbx5 that is expressed specifically in the forelimb region was shown to be almost identical between moa and ostrich, suggesting that, as well as being fully functional, tbx5 is likely to have been expressed normally in moa since divergence from their flighted ancestors, approximately 60 mya.
The potential of ecological models for supporting environmental decision making is increasingly acknowledged. However, it often remains unclear whether a model is realistic and reliable enough. Good practice for developing and testing ecological models has not yet been established. Therefore, TRACE, a general framework for documenting a model's rationale, design, and testing was recently suggested. Originally TRACE was aimed at documenting good modelling practice. However, the word 'documentation' does not convey TRACE's urgency. Therefore, we re-define TRACE as a tool for planning, performing, and documenting good modelling practice. TRACE documents should provide convincing evidence that a model was thoughtfully designed, correctly implemented, thoroughly tested, well understood, and appropriately used for its intended purpose. TRACE documents link the science underlying a model to its application, thereby also linking modellers and model users, for example stakeholders, decision makers, and developers of policies. We report on first experiences in producing TRACE documents. We found that the original idea underlying TRACE was valid, but to make its use more coherent and efficient, an update of its structure and more specific guidance for its use are needed. The updated TRACE format follows the recently developed framework of model 'evaludation': the entire process of establishing model quality and credibility throughout all stages of model development, analysis, and application. TRACE thus becomes a tool for planning, documenting, and assessing model evaludation, which includes understanding the rationale behind a model and its envisaged use. We introduce the new structure and revised terminology of TRACE and provide examples. (C) 2014 Elsevier B.V. All rights reserved.
Current chemical risk assessment procedures may result in imprecise estimates of risk due to sometimes arbitrary simplifying assumptions. As a way to incorporate ecological complexity and improve risk estimates, mechanistic effect models have been recommended. However, effect modeling has not yet been extensively used for regulatory purposes, one of the main reasons being uncertainty about which model type to use to answer specific regulatory questions. We took an individual-based model (IBM), which was developed for risk assessment of soil invertebrates and includes avoidance of highly contaminated areas, and contrasted it with a simpler, more standardized model, based on the generic metapopulation matrix model RAMAS. In the latter the individuals within a sub-population are not treated as separate entities anymore and the spatial resolution is lower. We explored consequences of model aggregation in terms of assessing population-level effects for different spatial distributions of a toxic chemical. For homogeneous contamination of the soil, we found good agreement between the two models, whereas for heterogeneous contamination, at different concentrations and percentages of contaminated area, RAMAS results were alternatively similar to IBM results with and without avoidance, and different food levels. This inconsistency is explained on the basis of behavioral responses that are included in the IBM but not in RAMAS. Overall, RAMAS was less sensitive than the IBM in detecting population-level effects of different spatial patterns of exposure. We conclude that choosing the right model type for risk assessment of chemicals depends on whether or not population-level effects of small-scale heterogeneity in exposure need to be detected. We recommend that if in doubt, both model types should be used and compared. Describing both models following the same standard format, the ODD protocol, makes them equally transparent and understandable. The simpler model helps to build up trust for the more complex model and can be used for more homogeneous exposure patterns. The more complex model helps detecting and understanding the limitations of the simpler model and is needed to ensure ecological realism for more complex exposure scenarios. (C) 2013 Elsevier B.V. All rights reserved.
Current environmental risk assessment (ERA) of chemicals for aquatic invertebrates relies on standardized laboratory tests in which toxicity effects on individual survival, growth and reproduction are measured. Such tests determine the threshold concentration of a chemical below which no population-level effects are expected. How well this procedure captures effects on individuals and populations, however, remains an open question. Here we used mechanistic effect models, combining individual-level reproduction and survival models with an individual-based population model (IBM), to understand the individuals' responses and extrapolate them to the population level. We used a toxicant (Dispersogen A) for which adverse effects on laboratory populations were detected at the determined threshold concentration and thus challenged the conservatism of the current risk assessment method. Multiple toxicity effects on reproduction and survival were reported, in addition to effects on the F1 generation. We extrapolated commonly tested individual toxicity endpoints, reproduction and survival, to the population level using the IBM. Effects on reproduction were described via regression models. To select the most appropriate survival model, the IBM was run assuming either stochastic death (SD) or individual tolerance (IT). Simulations were run for different scenarios regarding the toxicant's effects: survival toxicity, reproductive toxicity, or survival and reproductive toxicity. As population-level endpoints, we used population size and structure and extinction risk. We found that survival represented as SD explained population dynamics better than IT. Integrating toxicity effects on both reproduction and survival yielded more accurate predictions of population effects than considering isolated effects. To fully capture population effects observed at high toxicant concentrations, toxicity effects transmitted to the F1 generation had to be integrated. Predicted extinction risk was highly sensitive to the assumptions about individual-level effects. Our results demonstrate that the endpoints used in current standard tests may not be sufficient for assessing the risk of adverse effects on populations. A combination of laboratory population experiments with mechanistic effect models is a powerful tool to better understand and predict effects on both individuals and populations. Mechanistic effect modelling thus holds great potential to improve the accuracy of ERA of chemicals in the future. (C) 2013 The Authors. Published by Elsevier B.V. All rights reserved.
Division of labor is a hallmark of social insects. In honey bees, division of labor involves transition of female workers from one task to the next. The most distinct tasks are nursing (providing food for the brood) and foraging (collecting pollen and nectar). The brain mechanisms regulating this form of behavioral plasticity have largely remained elusive. Recently, it was suggested that division of labor is based on nutrition-associated signaling pathways. One highly conserved gene associated with food-related behavior across species is the foraging gene, which encodes a cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG). Our analysis of this gene reveals the presence of alternative splicing in the honey bee. One isoform is expressed in the brain. Expression of this isoform is most pronounced in the mushroom bodies, the subesophageal ganglion, and the corpora allata. Division of labor and sucrose responsiveness in honey bees correlate significantly with foraging gene expression in distinct brain regions. Activating PKG selectively increases sucrose responsiveness in nurse bees to the level of foragers, whereas the same treatment does not affect responsiveness to light. These findings demonstrate a direct link between PKG signaling in distinct brain areas and division of labor. Furthermore, they demonstrate that the difference in sensory responsiveness between nurse bees and foragers can be compensated for by activating PKG. Our findings on the function of PKG in regulating specific sensory responsiveness and social organization offer valuable indications for the function of the cGMP/PKG pathway in many other insects and vertebrates. J. Comp. Neurol. 522:1786-1799, 2014. (c) 2013 Wiley Periodicals, Inc.
In Germany, active bat rabies surveillance was conducted between 1993 and 2012. A total of 4546 oropharyngeal swab samples from 18 bat species were screened for the presence of EBLV-1- , EBLV-2- and BBLV-specific RNA. Overall, 0 center dot 15% of oropharyngeal swab samples tested EBLV-1 positive, with the majority originating from Eptesicus serotinus. Interestingly, out of seven RT-PCR-positive oropharyngeal swabs subjected to virus isolation, viable virus was isolated from a single serotine bat (E. serotinus). Additionally, about 1226 blood samples were tested serologically, and varying virus neutralizing antibody titres were found in at least eight different bat species. The detection of viral RNA and seroconversion in repeatedly sampled serotine bats indicates long-term circulation of the virus in a particular bat colony. The limitations of random-based active bat rabies surveillance over passive bat rabies surveillance and its possible application of targeted approaches for future research activities on bat lyssavirus dynamics and maintenance are discussed.
The non-random chromosomal translocations t(10;11)(p13;q23) and t(10;11)(p13;q14-21) result in leukemogenic fusion proteins comprising the coiled coil domain of the transcription factor AF10 and the proteins MLL or CALM, respectively, and subsequently cause certain types of acute leukemia. The AF10 coiled-coil domain, which is crucial for the leukemogenic effect, has been shown to interact with GAS41, a protein previously identified as the product of an amplified gene in glioblastoma. Using sequential synthetic peptides, we mapped the potential AF10/GAS41 interaction site, which was subsequently be used as scaffold for a library targeting the AF10 coiled-coil domain. Using phage display, we selected a peptide that binds the AF10 coiled-coil domain with higher affinity than the respective coiled-coil region of wild-type GAS41, as demonstrated by phage ELISA, CD, and PCAs. Furthermore, we were able to successfully deploy the inhibitory peptide in a mammalian cell line to lower the expression of Hoxa genes that have been described to be overexpressed in these leukemias. This work dissects molecular determinants mediating AF10-directed interactions in leukemic fusions comprising the N-terminal parts of the proteins MLL or CALM and the C-terminal coiled-coil domain of AF10. Furthermore, it outlines the first steps in recognizing and blocking the leukemia-associated AF10 interaction in histiocytic lymphoma cells and therefore, may have significant implications in future diagnostics and therapeutics. Copyright (c) 2014 European Peptide Society and John Wiley & Sons, Ltd.
Environmental change is likely to have a strong impact on biodiversity, and many species may shift their distribution in response. In this study, we aimed at projecting the availability of suitable habitat for an endangered amphibian species, the Fire-bellied toad Bombina bombina, in Brandenburg (north-eastern Germany). We modelled a potential habitat distribution map based on (1) a database with 10,581 presence records for Bombina from the years 1990 to 2009, (2) current estimates for ecogeographical variables (EGVs) and (3) the future projection of these EGVs according to the statistical regional model, respectively, the soil and water integrated model, applying the maximum entropy approach (Maxent). By comparing current and potential future distributions, we evaluated the projected change in distribution of suitable habitats and identified the environmental variables most associated with habitat suitability that turned out to be climatic variables related to the hydrological cycle. Under the applied scenario, our results indicate increasing habitat suitability in many areas and an extended range of suitable habitats. However, even if the environmental conditions in Brandenburg may change as predicted, it is questionable whether the Fire-bellied toad will truly benefit, as dispersal abilities of amphibian species are limited and strongly influenced by anthropogenic disturbances, that is, intensive agriculture, habitat destruction and fragmentation. Furthermore, agronomic pressure is likely to increase on productive areas with fertile soils and high water retention capacities, indeed those areas suitable for B. bombina. All these changes may affect temporary pond hydrology as well as the reproductive success and breeding phenology of toads.
The amount of terrestrial particulate organic matter (t-POM) entering lakes is predicted to increase as a result of climate change. This may especially alter the structure and functioning of ecosystems in small, shallow lakes which can rapidly shift from a clear-water, macrophyte-dominated into a turbid, phytoplankton-dominated state. We used the integrative ecosystem model PCLake to predict how rising t-POM inputs affect the resilience of the clear-water state. PCLake links a pelagic and benthic food chain with abiotic components by a number of direct and indirect effects. We focused on three pathways (zoobenthos, zooplankton, light availability) by which elevated t-POM inputs (with and without additional nutrients) may modify the critical nutrient loading thresholds at which a clear-water lake becomes turbid and vice versa. Our model results show that (1) increased zoobenthos biomass due to the enhanced food availability results in more benthivorous fish which reduce light availability due to bioturbation, (2) zooplankton biomass does not change, but suspended t-POM reduces the consumption of autochthonous particulate organic matter which increases the turbidity, and (3) the suspended t-POM reduces the light availability for submerged macrophytes. Therefore, light availability is the key process that is indirectly or directly changed by t-POM input. This strikingly resembles the deteriorating effect of terrestrial dissolved organic matter on the light climate of lakes. In all scenarios, the resilience of the clear-water state is reduced thus making the turbid state more likely at a given nutrient loading. Therefore, our study suggests that rising t-POM input can add to the effects of climate warming making reductions in nutrient loadings even more urgent.
The design and selection of peptides targeting cellular proteins is challenging and often yields candidates with undesired properties. Therefore we deployed a new selection system based on the twin-arginine translocase (TAT) pathway of Escherichia coli, named hitchhiker translocation (HiT) selection. A pool of alpha-helix encoding sequences was designed and selected for interference with the coiled coil domain (CC) of a melanoma-associated basic-helix-loop-helix-leucine-zipper (bHLHLZ) protein, the microphthalmia associated transcription factor (MITF). One predominant sequence (iM10) was enriched during selection and showed remarkable protease resistance, high solubility and thermal stability while maintaining its specificity. Furthermore, it exhibited nanomolar range affinity towards the target peptide. A mutation screen indicated that target-binding helices of increased homodimer stability and improved expression rates were preferred in the selection process. The crystal structure of the iM10/MITF-CC heterodimer (2.1 angstrom) provided important structural insights and validated our design predictions. Importantly, iM10 did not only bind to the MITF coiled coil, but also to the markedly more stable HLHLZ domain of MITF. Characterizing the selected variants of the semi-rational library demonstrated the potential of the innovative bacterial selection approach. (C) 2014 Elsevier Inc. All rights reserved.
Recent advances in high-throughput omics techniques render it possible to decode the function of genes by using the "guilt-by-association" principle on biologically meaningful clusters of gene expression data. However, the existing frameworks for biological evaluation of gene clusters are hindered by two bottleneck issues: (1) the choice for the number of clusters, and (2) the external measures which do not take in consideration the structure of the analyzed data and the ontology of the existing biological knowledge. Here, we address the identified bottlenecks by developing a novel framework that allows not only for biological evaluation of gene expression clusters based on existing structured knowledge, but also for prediction of putative gene functions. The proposed framework facilitates propagation of statistical significance at each of the following steps: (1) estimating the number of clusters, (2) evaluating the clusters in terms of novel external structural measures, (3) selecting an optimal clustering algorithm, and (4) predicting gene functions. The framework also includes a method for evaluation of gene clusters based on the structure of the employed ontology. Moreover, our method for obtaining a probabilistic range for the number of clusters is demonstrated valid on synthetic data and available gene expression profiles from Saccharomyces cerevisiae. Finally, we propose a network-based approach for gene function prediction which relies on the clustering of optimal score and the employed ontology. Our approach effectively predicts gene function on the Saccharomyces cerevisiae data set and is also employed to obtain putative gene functions for an Arabidopsis thaliana data set.
The control of gene expression by transcriptional regulators and other types of functionally relevant DNA transactions such as chromatin remodeling and replication underlie a vast spectrum of biological processes in all organisms. DNA transactions require the controlled interaction of proteins with DNA sequence motifs which are often located in nucleosome-depleted regions (NDRs) of the chromatin. Formaldehyde-assisted isolation of regulatory elements (FAIRE) has been established as an easy-to-implement method for the isolation of NDRs from a number of eukaryotic organisms, and it has been successfully employed for the discovery of new regulatory segments in genomic DNA from, for example, yeast, Drosophila, and humans. Until today, however, FAIRE has only rarely been employed in plant research and currently no detailed FAIRE protocol for plants has been published. Here, we provide a step-by-step FAIRE protocol for NDR discovery in Arabidopsis thaliana. We demonstrate that NDRs isolated from plant chromatin are readily amenable to quantitative polymerase chain reaction and next-generation sequencing. Only minor modification of the FAIRE protocol will be needed to adapt it to other plants, thus facilitating the global inventory of regulatory regions across species.
Redox modulation of protein activity by thioredoxins (TRXs) plays a key role in cellular regulation. Thioredoxin z (TRX z) and its interaction partner fructokinase-like protein 1 (FLN1) represent subunits of the plastid-encoded RNA polymerase (PEP), suggesting a role of both proteins in redox regulation of chloroplast gene expression. Loss of TRX z or FLN1 expression generates a PEP-deficient phenotype and renders the plants incapable to grow autotrophically. This study shows that PEP function in trx z and fln1 plants can be restored by complementation with redox-inactive TRX z C106S and FLN1 C(105/106)A protein variants, respectively. The complemented plants showed wild-type levels of chloroplast gene expression and were restored in photosynthetic capacity, indicating that redox regulation of PEP through TRX z/FLN1 per se is not essential for autotrophic growth. Promoter-reporter gene studies indicate that TRX z and FLN1 are expressed during early phases of leaf development while expression ceases at maturation. Taken together, our data support a model in which TRX z and FLN1 are essential structural components of the PEP complex and their redox activity might only play a role in the fine tuning of PEP function.
In the genome of Drosophila melanogaster, four genes coding for aldehyde oxidases (AOX1-4) were identified on chromosome 3. Phylogenetic analysis showed that the AOX gene cluster evolved via independent duplication events in the vertebrate and invertebrate lineages. The functional role and the substrate specificity of the distinct Drosophila AOX enzymes is unknown. Two loss-of-function mutant alleles in this gene region, low pyridoxal oxidase (Po-lpo) and aldehyde oxidase-1 (Aldox-1(n1)) are associated with a phenotype characterized by undetectable AOX enzymatic activity. However, the genes involved and the corresponding mutations have not yet been identified. In this study we characterized the activities, substrate specificities and expression profiles of the four AOX enzymes in D. melanogaster. We show that the Po-lpo-associated phenotype is the consequence of a structural alteration of the AOX1 gene. We identified an 11-bp deletion in the Po-lpo allele, resulting in a frame-shift event, which removes the molybdenum cofactor domain of the encoded enzyme. Furthermore, we show that AOX2 activity is detectable only during metamorphosis and characterize a Minos-AOX2 insertion in this developmental gene that disrupts its activity. We demonstrate that the Aldox-1(n1) phenotype maps to the AOX3 gene and AOX4 activity is not detectable in our assays.
Evolution of the nucleus
(2014)
The nucleus represents a major evolutionary transition. As a consequence of separating translation from transcription many new functions arose, which likely contributed to the remarkable success of eukaryotic cells. Here we will consider what has recently emerged on the evolutionary histories of several key aspects of nuclear biology; the nuclear pore complex, the lamina, centrosomes and evidence for prokaryotic origins of relevant players.
Bat activity is often concentrated near linear and edge landscape structures such as hedgerows, but information about seasonal and species-specific bat activity near hedges is scarce despite their abundance in the cultural landscapes of central Europe. Exact knowledge on animals' habitat use, however, is key to effective landscape planning to avoid human-wildlife-conflicts, such as the construction of wind turbines in areas with high bat activity that may result in bat fatalities. We measured bat activity in relation to distance to hedgerows in an agricultural landscape in northeastern Germany. We recorded bat echolocation calls at ground level at 0, 50, 100 and 200 m distances from hedges at five sites during three nights in spring (April to June) and three nights in summer (July to October) at each site. For all bat species we found the overall activity to be similar between seasons, with the highest activity near the hedges, but with considerable variation in species-specific spatial activity patterns between spring and summer. While the genus Myotis and Pipistrellus pipistrellus were mostly active close to the hedges at a similar intensity over the entire study period (i.e. 84% and 86% of all bat passes, respectively), Nyctalus noctula and Pipistrellus nathusii showed generally less pronounced concentration of activity near the hedges, and increased activity away from the hedges in summer. Similarly, Pipistrellus pygmaeus showed decreased activity away from the hedges during both seasons, but with reduced activity near the hedges in summer. The observed behavioural changes in activity in relation to distance to hedgerows are likely due to migration or the bats foraging for different prey between seasons. Our findings are highly relevant for landscape planning and distance recommendations for the construction of wind turbines linked to their potential threat for bats.
We investigated the systems response of metabolism and growth after an increase in irradiance in the nonsaturating range in the algal model Chlamydomonas reinhardtii. In a three-step process, photosynthesis and the levels of metabolites increased immediately, growth increased after 10 to 15 min, and transcript and protein abundance responded by 40 and 120 to 240 min, respectively. In the first phase, starch and metabolites provided a transient buffer for carbon until growth increased. This uncouples photosynthesis from growth in a fluctuating light environment. In the first and second phases, rising metabolite levels and increased polysome loading drove an increase in fluxes. Most Calvin-Benson cycle (CBC) enzymes were substrate-limited in vivo, and strikingly, many were present at higher concentrations than their substrates, explaining how rising metabolite levels stimulate CBC flux. Rubisco, fructose-1,6-biosphosphatase, and seduheptulose-1,7-bisphosphatase were close to substrate saturation in vivo, and flux was increased by posttranslational activation. In the third phase, changes in abundance of particular proteins, including increases in plastidial ATP synthase and some CBC enzymes, relieved potential bottlenecks and readjusted protein allocation between different processes. Despite reasonable overall agreement between changes in transcript and protein abundance (R-2 = 0.24), many proteins, including those in photosynthesis, changed independently of transcript abundance.
A major goal of island biogeography is to understand how island communities are assembled over time. However, we know little about the influence of variable area and ecological opportunity on island biotas over geological time-scales. Islands have limited life spans, and it has been posited that insular diversity patterns should rise and fall with an island's ontogeny. The potential of phylogenies to inform us of island ontogenetic stage remains unclear, as we lack a phylogenetic framework that focuses on islands rather than clades. Here, we present a parsimonious island-centric model that integrates phylogeny and ontogeny into island biogeography and can incorporate a negative feedback of diversity on species origination. This framework allows us to generate predictions about species richness and phylogenies on islands of different ages. We find that peak richness lags behind peak island area, and that endemic species age increases with island age on volcanic islands. When diversity negatively affects rates of immigration and cladogenesis, our model predicts speciation slowdowns on old islands. Importantly, we find that branching times of in situ radiations can be informative of an island's ontogenetic stage. This novel framework provides a quantitative means of uncovering processes responsible for island biogeography patterns using phylogenies.
Dendritic polyglycerol-poly(ethylene glycol)-based polymer networks for biosensing application
(2014)
This work describes the formation of a new dendritic polyglycerol-poly(ethylene glycol)-based 3D polymer network as a matrix for immobilization of the redox enzyme periplasmatic aldehyde oxidoreductase to create an electrochemical biosensor. The novel network is built directly on the gold surface, where it simultaneously stabilizes the enzyme for up to 4 days. The prepared biosensors can be used for amperometric detection of benzaldehyde in the range of 0.8-400 mu M.
Site-directed spin labeling of the unnatural amino acid p-acetylphenylalanine (p-AcPhe) using oxime based coupling chemistry is successfully applied to investigate human sulfite oxidase (hSO), a protein containing an essential cysteine residue, which impedes the use of thiol based coupling chemistry. The protein was found to be sensitive toward typical reaction conditions of oxime coupling, namely, acidic reaction conditions and elevated temperatures. Thus, coupling at neutral pH and room temperature is mandatory. Three catalysts described in the literature to accelerate the reaction rate have been tested. Best spin labeling efficiencies were observed for p-methoxyaniline, while the other catalysts described in the literature to have even better performance for oxime coupling at neutral pH were substantially less active or led to precipitation of the protein. A clear correlation of spin labeling efficiency with the local environment of the residue is found, shedding some light on the importance of the sterically demanding reaction complex between p-AcPhe, the aniline catalyst, and the spin label for the reaction rate. The analysis of the line shape has shown that its interpretation in terms of local environment is more challenging as compared to the well-established spin labels based on cysteine chemistry. To this end the results presented here indicate that the larger steric demand of the spin labeled p-AcPhe can induce structural effects instead of reporting on them.
Glucan, water dikinase (GWD) is a key enzyme of starch metabolism but the physico-chemical properties of starches isolated from GWD-deficient plants and their implications for starch metabolism have so far not been described. Transgenic Arabidopsis thaliana plants with reduced or no GWD activity were used to investigate the properties of starch granules. In addition, using various in vitro assays, the action of recombinant GWD, -amylase, isoamylase and starch synthase 1 on the surface of native starch granules was analysed. The internal structure of granules isolated from GWD mutant plants is unaffected, as thermal stability, allomorph, chain length distribution and density of starch granules were similar to wild-type. However, short glucan chain residues located at the granule surface dominate in starches of transgenic plants and impede GWD activity. A similarly reduced rate of phosphorylation by GWD was also observed in potato tuber starch fractions that differ in the proportion of accessible glucan chain residues at the granule surface. A model is proposed to explain the characteristic morphology of starch granules observed in GWD transgenic plants. The model postulates that the occupancy rate of single glucan chains at the granule surface limits accessibility to starch-related enzymes.
Aim Growth is both a matter of amplitude and tempo. We aimed to develop references for body height, body weight and body mass index (BMI) with respect to tempo of maturity. Methods Data obtained from the German KiGGS study (2003-2006) on body height, body weight and presence or absence of the menarche were re-analysed in 3776 girls, aged 10-17years. We developed smoothed centiles for BMI-, body-height- and body-weight-for-age using the LMS method for premenarcheal and postmenarcheal girls. Results Body height, body weight and BMI differed significantly between premenarcheal and postmenarcheal girls. On average, postmenarcheal girls aged 11-17years were 5.3cm taller and 9.7kg heavier, and their BMI was 2.9kg/m2 higher than in premenarcheal girls of the same calendar age. Conclusion Adolescent BMI rises with calendar age and biological age. New reference charts for adolescent girls aged 10-18years were generated to be inserted into the currently used references to avoid misclassifying underweight and overweight pubertal girls.
Fungal plant pathogens are common in natural communities where they affect plant physiology, plant survival, and biomass production. Conversely, pathogen transmission and infection may be regulated by plant community characteristics such as plant species diversity and functional composition that favor pathogen diversity through increases in host diversity while simultaneously reducing pathogen infection via increased variability in host density and spatial heterogeneity. Therefore, a comprehensive understanding of multi-host multi-pathogen interactions is of high significance in the context of biodiversity-ecosystem functioning. We investigated the relationship between plant diversity and aboveground obligate parasitic fungal pathogen ("pathogens" hereafter) diversity and infection in grasslands of a long-term, large-scale, biodiversity experiment with varying plant species (1-60 species) and plant functional group diversity (1-4 groups). To estimate pathogen infection of the plant communities, we visually assessed pathogen-group presence (i.e., rusts, powdery mildews, downy mildews, smuts, and leaf-spot diseases) and overall infection levels (combining incidence and severity of each pathogen group) in 82 experimental plots on all aboveground organs of all plant species per plot during four surveys in 2006.
Pathogen diversity, assessed as the cumulative number of pathogen groups on all plant species per plot, increased log-linearly with plant species diversity. However, pathogen incidence and severity, and hence overall infection, decreased with increasing plant species diversity. In addition, co-infection of plant individuals by two or more pathogen groups was less likely with increasing plant community diversity. We conclude that plant community diversity promotes pathogen-community diversity while at the same time reducing pathogen infection levels of plant individuals.
The relative contribution of personal and social information to explain individual and collective behavior in different species and contexts is an open question in animal ecology. In particular, there is a major lack of studies combining theoretical and empirical approaches to test the relative relevance of different hypothesized individual behaviors to predict empirical collective patterns. We used an individual-based model to confront three hypotheses about the information transfer between social scavengers (Griffon Vultures, Gyps fulvus) when searching for carrion: (1) Vultures only use personal information during foraging ("nonsocial" hypothesis); (2) they create long chains of vultures by following both other vultures that are flying towards carcasses and vultures that are following other vultures that are flying towards carcasses ("chains of vultures" hypothesis); and (3) vultures are only attracted by other vultures that are sinking vertically to a carcass ("local enhancement" hypothesis). The chains of vultures hypothesis has been used in existing models, but never been confronted with field data. Testing is important, though, because these hypotheses could have different management implications. The model was parameterized to mimic the behavior and the densities of both Griffon Vultures and carcasses in a 10 000-km(2) study area in northeastern Spain. We compared the number of vultures attending simulated carcasses with those attending 25 continuously monitored experimental carcasses in the field. Social hypotheses outperformed the nonsocial hypothesis. The chains of vultures hypothesis overestimated the number of vultures feeding on carcasses; the local enhancement hypothesis fitted closely to the empirical data. Supported by our results, we discuss mechanistic and adaptive considerations that reveal that local enhancement may be the key social mechanism behind collective foraging in this and likely other avian scavengers and/or social birds. It also highlights the current need for more studies confronting alternative models of key behaviors with empirical patterns in order to understand how collective behavior emerges in animal societies.
Background: Protein kinases constitute a particularly large protein family in Arabidopsis with important functions in cellular signal transduction networks. At the same time Arabidopsis is a model plant with high frequencies of gene duplications. Here, we have conducted a systematic analysis of the Arabidopsis kinase complement, the kinome, with particular focus on gene duplication events. We matched Arabidopsis proteins to a Hidden-Markov Model of eukaryotic kinases and computed a phylogeny of 942 Arabidopsis protein kinase domains and mapped their origin by gene duplication.
Results: The phylogeny showed two major clades of receptor kinases and soluble kinases, each of which was divided into functional subclades. Based on this phylogeny, association of yet uncharacterized kinases to families was possible which extended functional annotation of unknowns. Classification of gene duplications within these protein kinases revealed that representatives of cytosolic subfamilies showed a tendency to maintain segmentally duplicated genes, while some subfamilies of the receptor kinases were enriched for tandem duplicates. Although functional diversification is observed throughout most subfamilies, some instances of functional conservation among genes transposed from the same ancestor were observed. In general, a significant enrichment of essential genes was found among genes encoding for protein kinases.
Conclusions: The inferred phylogeny allowed classification and annotation of yet uncharacterized kinases. The prediction and analysis of syntenic blocks and duplication events within gene families of interest can be used to link functional biology to insights from an evolutionary viewpoint. The approach undertaken here can be applied to any gene family in any organism with an annotated genome.
Leaf senescence is an active process with a pivotal impact on plant productivity. It results from extensive signalling cross-talk coordinating environmental factors with intrinsic age-related mechanisms. Although many studies have shown that leaf senescence is affected by a range of external parameters, knowledge about the regulatory systems that govern the interplay between developmental programmes and environmental stress is still vague. Salinity is one of the most important environmental stresses that promote leaf senescence and thus affect crop yield. Improving salt tolerance by avoiding or delaying senescence under stress will therefore play an important role in maintaining high agricultural productivity. Experimental evidence suggests that hydrogen peroxide (H2O2) functions as a common signalling molecule in both developmental and salt-induced leaf senescence. In this study, microarray-based gene expression profiling on Arabidopsis thaliana plants subjected to long-term salinity stress to induce leaf senescence was performed, together with co-expression network analysis for H2O2-responsive genes that are mutually up-regulated by salt induced-and developmental leaf senescence. Promoter analysis of tightly co-expressed genes led to the identification of seven cis-regulatory motifs, three of which were known previously, namely CACGTGT and AAGTCAA, which are associated with reactive oxygen species (ROS)-responsive genes, and CCGCGT, described as a stress-responsive regulatory motif, while the others, namely ACGCGGT, AGCMGNC, GMCACGT, and TCSTYGACG were not characterized previously. These motifs are proposed to be novel elements involved in the H2O2-mediated control of gene expression during salinity stress-triggered and developmental senescence, acting through upstream transcription factors that bind to these sites.
Leaf senescence is a developmentally controlled process, which is additionally modulated by a number of adverse environmental conditions. Nitrogen shortage is a well-known trigger of precocious senescence in many plant species including crops, generally limiting biomass and seed yield. However, leaf senescence induced by nitrogen starvation may be reversed when nitrogen is resupplied at the onset of senescence. Here, the transcriptomic, hormonal, and global metabolic rearrangements occurring during nitrogen resupply-induced reversal of senescence in Arabidopsis thaliana were analysed. The changes induced by senescence were essentially in keeping with those previously described; however, these could, by and large, be reversed. The data thus indicate that plants undergoing senescence retain the capacity to sense and respond to the availability of nitrogen nutrition. The combined data are discussed in the context of the reversibility of the senescence programme and the evolutionary benefit afforded thereby. Future prospects for understanding and manipulating this process in both Arabidopsis and crop plants are postulated.
This study examines a natural, rapid, fivefold increase in dissolved organic carbon (DOC) concentrations in a temperate shallow lake, describing the processes by which increased DOC resulted in anoxic conditions and altered existing carbon cycling pathways. High precipitation for two consecutive years led to rising water levels and the flooding of adjacent degraded peatlands. Leaching from the flooded soils provided an initial increase in DOC concentrations (from a 2010 mean of 12 +/- 1 mg L-1 to a maximum concentration of 53 mg L-1 by June 2012). Increasing water levels, DOC, and phytoplankton concentrations reduced light reaching the sediment surface, eliminating most benthic primary production and promoting anoxia in the hypolimnion. From January to June 2012 there was a sudden increase in total phosphorus (from 57 mg L-1 to 216 mg L-1), DOC (from 24.6 mg L-1 to 53 mg L-1), and iron (from 0.12 mg L-1 to 1.07 mg L-1) concentrations, without any further large fluxes in water levels. We suggest that anoxic conditions at the sediment surface and flooded soils produced a dramatic release of these chemicals that exacerbated brownification and eutrophication, creating anoxic conditions that persisted roughly 6 months below a water depth of 1 m and extended periodically to the water surface. This brownification-anoxia feedback loop resulted in a near-complete loss of macroinvertebrate and fish populations, and increased surface carbon dioxide (CO2) emissions by an order of magnitude relative to previous years.
The teleost v1r-related ora genes are a small, highly conserved olfactory receptor gene family of only six genes, whose direct orthologues can be identified in lineages as far as that of cartilaginous fish. However, no ligands for fish olfactory receptor class A related genes (ORA) had been uncovered so far. Here we have deorphanized the ORA1 receptor using heterologous expression and calcium imaging. We report that zebrafish ORA1 recognizes with high specificity and sensitivity 4-hydroxyphenylacetic acid. The carboxyl group of this compound is required in a particular distance from the aromatic ring, whereas the hydroxyl group in the para-position is not essential, but strongly enhances the binding efficacy. Low concentrations of 4-hydroxyphenylacetic acid elicit increases in oviposition frequency in zebrafish mating pairs. This effect is abolished by naris closure. We hypothesize that 4-hydroxyphenylacetic acid might function as a pheromone for reproductive behavior in zebrafish. ORA1 is ancestral to mammalian V1Rs, and its putative function as pheromone receptor is reminiscent of the role of several mammalian V1Rs as pheromone receptors.
Starch synthase (SS) and branching enzyme (BE) establish the two glycosidic linkages existing in starch. Both enzymes exist as several isoforms. Enzymes derived from several species were studied extensively both in vivo and in vitro over the last years, however, analyses of a functional interaction of SS and BE isoforms are missing so far. Here, we present data from in vitro studies including both interaction of leaf derived and heterologously expressed SS and BE isoforms. We found that SSI activity in native PAGE without addition of glucans was dependent on at least one of the two BE isoforms active in Arabidopsis leaves. This interaction is most likely not based on a physical association of the enzymes, as demonstrated by immunodetection and native PAGE mobility analysis of SSI, BE2, and BE3. The glucans formed by the action of SSI/BEs were analysed using leaf protein extracts from wild type and be single mutants (Atbe2 and Atbe3 mutant lines) and by different combinations of recombinant proteins. Chain length distribution (CLD) patterns of the formed glucans were irrespective of SSI and BE isoforms origin and still independent of assay conditions. Furthermore, we show that all SS isoforms (SSI-SSIV) were able to interact with BEs and form branched glucans. However, only SSI/BEs generated a polymodal distribution of glucans which was similar to CLD pattern detected in amylopectin of Arabidopsis leaf starch. We discuss the impact of the SSI/BEs interplay for the CLD pattern of amylopectin.
Pyranopterin dithiolene distortions relevant to electron transfer in xanthine oxidase/dehydrogenase
(2014)
The reducing substrates 4-thiolumazine and 2,4-dithiolumazine have been used to form Mo-IV-product complexes with xanthine oxidase (XO) and xanthine dehydrogenase. These Mo-IV-product complexes display an intense metal-to-ligand charge-transfer (MLCT) band in the near-infrared region of the spectrum. Optical pumping into this MLCT band yields resonance Raman spectra of the Mo site that are devoid of contributions from the highly absorbing FAD and 2Fe2S clusters in the protein. The resonance Raman spectra reveal in-plane bending modes of the bound product and low-frequency molybdenum dithiolene and pyranopterin dithiolene vibrational modes. This work provides keen insight into the role of the pyranopterin dithiolene in electron-transfer reactivity.
Females may select a mate based on signalling traits that are believed to accurately correlate with heritable aspects of male quality. Anthropogenic actions, in particular chemicals released into the environment, are now disrupting the accuracy of mating signals to convey information about male quality. The long-term prediction for disrupted mating signals is most commonly loss of female preference. Yet, this prediction has rarely been tested using quantitative models. We use agent-based models to explore the effects of rapid disruption of mating signals. In our model, a gene determines survival. Males signal their level of genetic quality via a signal trait, which females use to select a mate. We allowed this system of sexual selection to become established, before introducing a disruption between the male signal trait and quality, which was similar in nature to that induced by exogenous chemicals. Finally, we assessed the capacity of the system to recover from this disruption. We found that within a relatively short time frame, disruption of mating signals led to a lasting loss of female preference. Decreases in mean viability at the population-level were also observed, because sexual-selection acting against newly arising deleterious mutations was relaxed. The ability of the population to recover from disrupted mating signals was strongly influenced by the mechanisms that promoted or maintained genetic diversity in traits under sexual selection. Our simple model demonstrates that environmental perturbations to the accuracy of male mating signals can result in a long-term loss of female preference for those signals within a few generations. What is more, the loss of this preference can have knock-on consequences for mean population fitness.
By combining morphology, ecology, biology, and biogeography with the available molecular (sequence variation of the entire mitochondrial cytochrome b gene; cyt-b) and karyology data, the taxonomy of several species of the Rutilus complex inhabiting southern Europe is revised. Rutilus stoumboudae, new species, is described from Lake Volvi, Greece. It differs from Rutilus rutilus in possessing more total GR and less branched rays in both dorsal and anal fins and in its placement in the cyt-b based phylogeny of the genus. The resurrected genus Leucos Heckel, 1843 (type species Leucos aula, Bonaparte, 1841), which according to molecular data diverged from Rutilus more than 5 million years ago, during the Messinian salinity crisis, includes five species of small size, without spinous tubercles on scales and head in reproductive males, pharyngeal teeth formula 5-5, and all show a preference for still waters. Leucos aula is the Italian species endemic in the Padany-Venetian district: L. basak is widespread in Croatia, Albania, Montenegro and former Yugoslav Republic of Macedonia (FYROM); L. albus, recently described from Lake Skadar, Montenegro, is also found in rivers Moraca and Zeta (Montenegro). L. albus differs from L. basak, its closest relative, in having more scales on the LL and less anal-fin rays; L. panosi is endemic to the western-Greece district, and L. ylikiensis is endemic to lakes Yliki and Paralimni in eastern Greece (introduced in Lake Volvi). Among the nominal species examined, Rutilus karamani, R. ohridanus, R. prespensis and R. prespensis vukovici are all junior synonyms of Leucos basak. Rutilus vegariticus is definitively regarded as junior synonym for R. rutilus. Sarmarutilus n.gen. is a monotypic genus, with Sarmarutilus rubilio as the type species. According to phylogenetic data, Sarmarutilus rubilio is basal to a cluster of species that includes Leucos basak, L. albus, L. aula, L. panosi and L. ylikiensis. Sarmarutilus possibly evolved in pre-Messinian time, in the Lago Mare, entered the Mediterranean area during the Messinian Lago Mare phase of the Mediterranean Sea and survived only in the Tuscany-Latium district. This genus differs from Leucos in having large pearl organs on the central part of head and body scales in mature males and for the habitat preference, being a riverine-adapted species. It differs from Rutilus in pharyngeal teeth formula (5-5 in Sarmarutilus and 6-5 in Rutilus), size (small in Sarmarutilus and large in Rutilus) and for the preferential habitat (riverine vs. still water). Finally, lectotypes for Leucos basak, Leucos aula, and Sarmarutilus rubilio are designated.
Savanna rangelands worldwide are threatened by shrub encroachment, i.e. the increase of woody plant species at the cost of perennial grasses, causing a strong decline in the productivity of domestic livestock production. Although recent studies indicate that fire might be of great importance for semi-arid and arid savanna dynamics, it is largely not applied in the management of semi-arid rangelands especially with regard to woody plant control. We used the eco-hydrological savanna model EcoHyD to simulate the effects of different fire management strategies on semi-arid savanna vegetation and to assess their longterm suitability for semi-arid rangeland management. Simulation results show that prescribed fires, timed to kill tree seedlings prevented shrub encroachment for a broad range of livestock densities while the possible maximum long-term cattle densities on the simulated semi-arid rangeland in Namibia increased by more than 30%. However, when grazing intensity was too high, fire management failed in preventing shrub encroachment.
Our findings indicate that with regard to fire management a clear distinction between mesic and more arid savannas is necessary: While the frequency of fires is of relevance for mesic savannas, we recommend a fire management focussing on the timing of fire for semi-arid and arid savannas. (C) 2014 Elsevier Ltd. All rights reserved.
Diatom diversity in lakes of northwest Yakutia (Siberia) was investigated by microscopic and genetic analysis of surface and cored lake sediments, to evaluate the use of sedimentary DNA for paleolimnological diatom studies and to identify obscure genetic diversity that cannot be detected by microscopic methods. Two short (76 and 73 bp) and one longer (577 bp) fragments of the ribulose 1,5-bisphosphate carboxylase/oxygenase (rbcL) gene, encoding the large subunit of the rbcL, were used as genetic markers. Diverse morphological assemblages of diatoms, dominated by small benthic fragilarioid taxa, were retrieved from the sediments of each lake. These minute fragilarioid taxa were examined by scanning electron microscopy, revealing diverse morphotypes in Staurosira and Staurosirella from the different lakes. Genetic analyses indicated a dominance of haplotypes that were assigned to fragilarioid taxa and less genetic diversity in other diatom taxa. The long rbcL_577 amplicon identified considerable diversification among haplotypes clustering within the Staurosira/Staurosirella genera, revealing 19 different haplotypes whose spatial distribution appears to be primarily related to the latitude of the lakes, which corresponds to a vegetation and climate gradient. Our rbcL markers are valuable tools for tracking differences between diatom lineages that are not visible in their morphologies. These markers revealed putatively high genetic diversity within the Staurosira/Staurosirella species complex, at a finer scale than is possible to resolve by microscopic determination. The rbcL markers may provide additional reliable information on the diversity of barely distinguishable minute benthic fragilarioids. Environmental sequencing may thus allow the tracking of spatial and temporal diversification in Siberian lakes, especially in the context of diatom responses to recent environmental changes, which remains a matter of controversy.
Animal personalities are by definition stable over time, but to what extent they may change during development and in adulthood to adjust to environmental change is unclear. Animals of temperate environments have evolved physiological and behavioural adaptations to cope with the cyclic seasonal changes. This may also result in changes in personality: suites of behavioural and physiological traits that vary consistently among individuals. Winter, typically the adverse season challenging survival, may require individuals to have shy/cautious personality, whereas during summer, energetically favourable to reproduction, individuals may benefit from a bold/risk-taking personality. To test the effects of seasonal changes in early life and in adulthood on behaviours (activity, exploration and anxiety), body mass and stress response, we manipulated the photoperiod and quality of food in two experiments to simulate the conditions of winter and summer. We used the common voles (Microtus arvalis) as they have been shown to display personality based on behavioural consistency over time and contexts. Summer-born voles allocated to winter conditions at weaning had lower body mass, a higher corticosterone increase after stress and a less active, more cautious behavioural phenotype in adulthood compared to voles born in and allocated to summer conditions. In contrast, adult females only showed plasticity in stress-induced corticosterone levels, which were higher in the animals that were transferred to the winter conditions than to those staying in summer conditions. These results suggest a sensitive period for season-related behavioural plasticity in which juveniles shift over the bold-shy axis.
Objective Wnt signalling pathways regulate proliferation, motility and survival in a variety of human cell types. Dickkopf 1 (DKK1) gene codes for a secreted Wnt inhibitory factor. It functions as tumour suppressor gene in breast cancer and as a pro-apoptotic factor in glioma cells. In this study, we aimed to demonstrate whether the different expression of DKK1 in human glioma-derived cells is dependent on microenvironmental factors like hypoxia and regulated by the intercellular crosstalk with bone-marrow-derived mesenchymal stem cells (bmMSCs).
Methods Glioma cell line U87-MG, three cell lines from human glioblastoma grade IV (glioma-derived mesenchymal stem cells) and three bmMSCs were selected for the experiment. The expression of DKK1 in cell lines under normoxic/hypoxic environment or co-culture condition was measured using real-time PCR and enzyme-linked immunoadsorbent assay. The effect of DKK1 on cell migration and proliferation was evaluated by in vitro wound healing assays and sulphorhodamine assays, respectively.
Results Glioma-derived cells U87-MG displayed lower DKK1 expression compared with bmMSCs. Hypoxia led to an overexpression of DKK1 in bmMSCs and U87-MG when compared to normoxic environment, whereas co-culture of U87-MG with bmMSCs induced the expression of DKK1 in both cell lines. Exogenous recombinant DKK1 inhibited cell migration on all cell lines, but did not have a significant effect on cell proliferation of bmMSCs and glioma cell lines.
Conclusion In this study, we showed for the first time that the expression of DKK1 was hypoxia dependent in human malignant glioma cell lines. The induction of DKK1 by intracellular crosstalk or hypoxia stimuli sheds light on the intense adaption of glial tumour cells to environmental alterations.
Climate change will likely affect population dynamics of numerous plant species by modifying several aspects of the life cycle. Because plant regeneration from seeds may be particularly vulnerable, here we assess the possible effects of climate change on seed characteristics and present an integrated analysis of seven seed traits (nutrient concentrations, samara mass, seed mass, wing length, seed viability, germination percentage, and seedling biomass) of Acer platanoides and A. pseudoplatanus seeds collected along a wide latitudinal gradient from Italy to Norway. Seed traits were analyzed in relation to the environmental conditions experienced by the mother trees along the latitudinal gradient. We found that seed traits of A. platanoides were more influenced by the climatic conditions than those of A. pseudoplatanus. Additionally, seed viability, germination percentage, and seedling biomass of A. platanoides were strongly related to the seed mass and nutrient concentration. While A. platanoides seeds were more influenced by the environmental conditions (generally negatively affected by rising temperatures), compared to A. pseudoplatanus, A. platanoides still showed higher germination percentage and seedling biomass than A. pseudoplatanus. Thus, further research on subsequent life-history stages of both species is needed. The variation in seed quality observed along the climatic gradient highlights the importance of studying the possible impact of climate change on seed production and species demography.
Ecosystem functioning is affected by horizontal (within trophic groups) and vertical (across trophic levels) biodiversity. Theory predicts that the effects of vertical biodiversity depend on consumer specialization. In a microcosm experiment, we investigated ciliate consumer diversity and specialization effects on algal prey biovolume, evenness and composition, and on ciliate biovolume production. The experimental data was complemented by a process-based model further analyzing the ecological mechanisms behind the observed diversity effects. Overall, increasing consumer diversity had no significant effect on prey biovolume or evenness. However, consumer specialization affected the prey community. Specialist consumers showed a stronger negative impact on prey biovolume and evenness than generalists. The model confirmed that this pattern was mainly driven by a single specialist with a high per capita grazing rate, consuming the two most productive prey species. When these were suppressed, the prey assemblage became dominated by a less productive species, consequently decreasing prey biovolume and evenness. Consumer diversity increased consumer biovolume, which was stronger for generalists than for specialists and highest in mixed combinations, indicating that consumer functional diversity, i.e. more diverse feeding strategies, increased resource use efficiency. Overall, our results indicate that consumer diversity effects on prey and consumers strongly depend on species-specific growth and grazing rates, which may be at least equally important as consumer specialization in driving consumer diversity effects across trophic levels.
2. We present a hierarchical model that integrates observations from multiple sources to estimate spatio-temporal abundance trends. The model links annual population densities on a spatial grid to both long-term count data and to opportunistic occurrence records from a citizen science programme. Specific observation models for both data types explicitly account for differences in data structure and quality.
3. We test this novel method in a virtual study with simulated data and apply it to the estimation of abundance dynamics across the range of a butterfly species (Pyronia tithonus) in Great Britain between 1985 and 2004. The application to simulated and real data demonstrates how the hierarchical model structure accommodates various sources of uncertainty which occur at different stages of the link between observational data and the modelled abundance, thereby it accounts for these uncertainties in the inference of abundance variations.
4. We show that by using hierarchical observation models that integrate different types of commonly available data sources, we can improve the estimates of variation in species abundances across space and time. This will improve our ability to detect regional trends and can also enhance the empirical basis for understanding range dynamics.
Transposons are massively abundant in all eukaryotic genomes and are suppressed by epigenetic silencing. Transposon activity contributes to the evolution of species; however, it is unclear how much transposition-induced variation exists at a smaller scale and how transposons are targeted for silencing. Here, we exploited differential silencing of the AtMu1c transposon in the Arabidopsis thaliana accessions Columbia (Col) and Landsberg erecta (Ler). The difference persisted in hybrids and recombinant inbred lines and was mapped to a single expression quantitative trait locus within a 20-kb interval. In Ler only, this interval contained a previously unidentified copy of AtMu1c, which was inserted at the 39 end of a protein-coding gene and showed features of expressed genes. By contrast, AtMu1c(Col) was intergenic and associated with heterochromatic features. Furthermore, we identified widespread natural AtMu1c transposition from the analysis of over 200 accessions, which was not evident from alignments to the reference genome. AtMu1c expression was highest for insertions within 39 untranslated regions, suggesting that this location provides protection from silencing. Taken together, our results provide a species-wide view of the activity of one transposable element at unprecedented resolution, showing that AtMu1c transposed in the Arabidopsis lineage and that transposons can escape epigenetic silencing by inserting into specific genomic locations, such as the 3' end of genes.
Climate change will increasingly affect the natural habitat and diet of polar bears (Ursus maritimus). Understanding the energetic needs of polar bears is therefore important. We developed a theoretical method for estimating polar bear food consumption based on using the highly recalcitrant polychlorinated biphenyl (PCB) congener, 2,2',4,4',55-hexaCB (CB153) in bear adipose tissue as an indicator of food intake. By comparing the CB153 tissue concentrations in wild polar bears with estimates from a purposely designed individual-based model, we identified the possible combinations of field metabolic rates (FMR) and CB153 deposition efficiencies in East Greenland polar bears. Our simulations indicate that if 30% of the CB153 consumed by polar bear individuals were deposited into their adipose tissue, the corresponding FMR would be only two times the basal metabolic rate. In contrast, if the modelled CB153 deposition efficiency were 10%, adult polar bears would require six times more energy than that needed to cover basal metabolism. This is considerably higher than what has been assumed for polar bears in previous studies though it is similar to FMRs found in other marine mammals. An implication of this result is that even relatively small reductions in future feeding opportunities could impact the survival of East Greenland polar bears.
Differential responses of herbivores and herbivory to management in temperate Eeuropean beech
(2014)
Forest management not only affects biodiversity but also might alter ecosystem processes mediated by the organisms, i.e. herbivory the removal of plant biomass by plant-eating insects and other arthropod groups. Aiming at revealing general relationships between forest management and herbivory we investigated aboveground arthropod herbivory in 105 plots dominated by European beech in three different regions in Germany in the sun-exposed canopy of mature beech trees and on beech saplings in the understorey. We separately assessed damage by different guilds of herbivores, i.e. chewing, sucking and scraping herbivores, gall-forming insects and mites, and leaf-mining insects. We asked whether herbivory differs among different forest management regimes (unmanaged, uneven-aged managed, even-aged managed) and among age-classes within even-aged forests. We further tested for consistency of relationships between regions, strata and herbivore guilds. On average, almost 80% of beech leaves showed herbivory damage, and about 6% of leaf area was consumed. Chewing damage was most common, whereas leaf sucking and scraping damage were very rare. Damage was generally greater in the canopy than in the understorey, in particular for chewing and scraping damage, and the occurrence of mines. There was little difference in herbivory among differently managed forests and the effects of management on damage differed among regions, strata and damage types. Covariates such as wood volume, tree density and plant diversity weakly influenced herbivory, and effects differed between herbivory types. We conclude that despite of the relatively low number of species attacking beech; arthropod herbivory on beech is generally high. We further conclude that responses of herbivory to forest management are multifaceted and environmental factors such as forest structure variables affecting in particular microclimatic conditions are more likely to explain the variability in herbivory among beech forest plots.
Pathogens such as viruses and bacteria use their envelope proteins and their adhesin lectins to recognize the glycan residues presented on the cell surface of the target tissues. This principle of recognition is used in a new electrochemical displacement sensor for the protein concanavalin A (ConA). A gold electrode was first modified with a self-assembled monolayer of a thiolated mannose/OEG conjugate and a ferrocene boroxol derivative was pre-assembled as reporter molecule onto the mannose surface. The novel tracer molecule based on a 2-hydroxymethyl phenyl boronic acid derivative binds even at neutral pH to the saccharides which could expand the application towards biological samples (i.e., urine and feces). Upon the binding of ConA, the tracer was displaced and washed away from the sensor surface leading to a decrease in the electrochemical signal. Using square wave voltammetry (SWV), the concentration of ConA in the sample solution could be determined in the dynamic concentration range established from 38 nmol L-1 to 5.76 mu mol L-1 with a reproducible detection limit of 1 mu g mL(-1) (38 nmol L-1) based on the signal-to-noise ratio (S/N=3) with fast response of 15 min. The new reporter molecule showed a reduced non-specific displacement by BSA and ribonuclease A. The sensor was also successfully transferred to the first proof of principle for the detection of Escherichia coli exhibiting a detection limit of approximately 6 x 102 cells/mL Specificity of the displacement by target protein ConA and E. coli was demonstrated since the control proteins (i.e., BSA and RNaseA) and the control E. coli strain, which lack of type 1 fimbriae, were ineffective. (C) 2014 Elsevier B.V. All rights reserved.
Elucidating the genetic basis of morphological changes in evolution remains a major challenge in biology [1-3]. Repeated independent trait changes are of particular interest because they can indicate adaptation in different lineages or genetic and developmental constraints on generating morphological variation [4-6]. In animals, changes to "hot spot" genes with minimal pleiotropy and large phenotypic effects underlie many cases of repeated morphological transitions [4-8]. By contrast, only few such genes have been identified from plants [8-11], limiting cross-kingdom comparisons of the principles of morphological evolution. Here, we demonstrate that the REDUCED COMPLEXITY (RCO) locus [12] underlies more than one naturally evolved change in leaf shape in the Brassicaceae. We show that the difference in leaf margin dissection between the sister species Capsella rubella and Capsella grandiflora is caused by cis-regulatory variation in the homeobox gene RCO-A, which alters its activity in the developing lobes of the leaf. Population genetic analyses in the ancestral C. grandiflora indicate that the more-active C. rubella haplotype is derived from a now rare or lost C. grandiflora haplotype via additional mutations. In Arabidopsis thaliana, the deletion of the RCO-A and RCO-B genes has contributed to its evolutionarily derived smooth leaf margin [12], suggesting the RCO locus as a candidate for an evolutionary hot spot. We also find that temperature-responsive expression of RCO-A can explain the phenotypic plasticity of leaf shape to ambient temperature in Capsella, suggesting a molecular basis for the well-known negative correlation between temperature and leaf margin dissection.
During heart development, the onset of heartbeat and blood flow coincides with a ballooning of the cardiac chambers. Here, we have used the zebrafish as a vertebrate model to characterize chamber ballooning morphogenesis of the endocardium, a specialized population of endothelial cells that line the interior of the heart. By combining functional manipulations, fate mapping studies, and high-resolution imaging, we show that endocardial growth occurs without an influx of external cells. Instead, endocardial cell proliferation is regulated, both by blood flow and by Bmp signaling, in a manner independent of vascular endothelial growth factor (VEGF) signaling. Similar to myocardial cells, endocardial cells obtain distinct chamber-specific and inner- versus outer-curvature-specific surface area sizes. We find that the hemodynamic-sensitive transcription factor Klf2a is involved in regulating endocardial cell morphology. These findings establish the endocardium as the flow-sensitive tissue in the heart with a key role in adapting chamber growth in response to the mechanical stimulus of blood flow.
The extremophilic green microalga Chlamydomonas acidophila grows in very acidic waters (pH 2.3-3.4), where CO2 is the sole inorganic carbon source. Previous work has revealed that the species can accumulate inorganic carbon (Ci) and exhibits high affinity CO2 utilization under low-CO2 (air-equilibrium) conditions, similar to organisms with an active CO2 concentrating mechanism (CCM), whereas both processes are down-regulated under high CO2 (4.5 % CO2) conditions. Responses of this species to phosphorus (Pi)-limited conditions suggested a contrasting regulation of the CCM characteristics. Therefore, we measured external carbonic anhydrase (CA(ext)) activities and protein expression (CAH1), the internal pH, Ci accumulation, and CO2-utilization in cells adapted to high or low CO2 under Pi-replete and Pi-limited conditions. Results reveal that C. acidophila expressed CA(ext) activity and expressed a protein cross-reacting with CAH1 (the CA(ext) from Chlamydomonas reinhardtii). Although the function of this CA remains unclear, CA(ext) activity and high affinity CO2 utilization were the highest under low CO2 conditions. C. acidophila accumulated Ci and expressed the CAH1 protein under all conditions tested, and C. reinhardtii also contained substantial amounts of CAH1 protein under Pi-limitation. In conclusion, Ci utilization is optimized in C. acidophila under ecologically relevant conditions, which may enable optimal survival in its extreme Ci- and Pi-limited habitat. The exact physiological and biochemical acclimation remains to be further studied.
Patterns of past vegetation changes over time and space can help facilitate better understanding of the interactions among climate, ecosystem, and human impact. Biome changes in China over the last 22,000 yr (calibrated radiocarbon date, a BP) were numerically reconstructed by using a standard approach of pollen-plant functional type-biome assignment (biomization). The biomization procedure involves pollen data from 2434 surface sites and 228 fossil sites with a high quality of pollen count and C-14 dating, 51 natural and three anthropogenic plant functional types (PFTs), as well as 19 natural and one anthropogenic biome. Surface pollen-based reconstruction of modern natural biome patterns is in good agreement (74.4%) with actual vegetation distribution in China. However, modem large-scale anthropogenic biome reconstruction has not been successful based on the current setup of three anthropogenic PFTs (plantation, secondary, and disturbed PFT) because of the limitation of non-species level pollen identification and the difficulty in the clear assignment of disturbed PFTs. The non-anthropogenic biome distributions of 44 time slices at 500-year intervals show large-scale discrepant and changed vegetation patterns from the last glacial maximum (LGM) to the Holocene throughout China. From 22 ka BP to 19 ka BP, temperate grassland, xerophytic shrubland, and desert dominated northern China, whereas cold or cool forests flourished in central China. Warm-temperate evergreen forests were restricted to far southern China, and tropical forests were absent During 18.5 ka BP to 12 ka BP, cold, cool, and dry biomes extended to some parts of northern, westem, and eastern China. Warm-temperate evergreen and mixed forests gradually expanded to occupy the whole of southern China. A slight northward shift of forest biomes occurred from 15 ka BP to 12 lea BP. During 11.5 ka BP to 9 ka BP, temperate grassland and shrubland gradually stretched to northern and western China. Cold and cool forests widely expanded into northern and central China, as well as in the northern margin of South China along with temperate deciduous forest. Since the early mid-Holocene (approximately 8.5 ka BP to 5.5 ka BP), all forest biomes shifted northward at the expense of herbaceous and shrubby biomes. Simultaneously, cold and cool forest biomes occupied the marginal areas of the Tibetan Plateau and the high mountains in western China. During the middle to late Holocene, from 5 ka to the present, temperate grassland and xerophytic shrubland expanded to the south and east, whereas temperate deciduous forests slightly shifted southward. After 3 lea BP, forest biomes were absent in western China and on the Tibetan plateau surface. Dramatic biome shifts from the LGM to the Holocene were observed in the forest-grassland ecotone and transitional zones between temperate and subtropical climates, between subtropical and tropical regions, and in the mountainous margins of the eastern Tibetan Plateau. Evidence showed more human disturbances during the late Holocene. More pollen records and historical documents are therefore further needed to understand fully the human disturbance-induced large-scale forest changes. In addition, more classifications of anthropogenic biome or land cover, more distinct assignment of pollen taxa to anthropogenic PFTs, and more effective numerical and/or mechanistic techniques in building large-scale human disturbances are required. (C) 2014 Elsevier B.V. All rights reserved.
Factors controlling decomposition rates of fine root litter in temperate forests and grasslands
(2014)
Fine root decomposition contributes significantly to element cycling in terrestrial ecosystems. However, studies on root decomposition rates and on the factors that potentially influence them are fewer than those on leaf litter decomposition. To study the effects of region and land use intensity on fine root decomposition, we established a large scale study in three German regions with different climate regimes and soil properties. Methods In 150 forest and 150 grassland sites we deployed litterbags (100 mu m mesh size) with standardized litter consisting of fine roots from European beech in forests and from a lowland mesophilous hay meadow in grasslands. In the central study region, we compared decomposition rates of this standardized litter with root litter collected on-site to separate the effect of litter quality from environmental factors.
Standardized herbaceous roots in grassland soils decomposed on average significantly faster (24 +/- 6 % mass loss after 12 months, mean +/- SD) than beech roots in forest soils (12 +/- 4 %; p < 0.001). Fine root decomposition varied among the three study regions. Land use intensity, in particular N addition, decreased fine root decomposition in grasslands. The initial lignin:N ratio explained 15 % of the variance in grasslands and 11 % in forests. Soil moisture, soil temperature, and C:N ratios of soils together explained 34 % of the variance of the fine root mass loss in grasslands, and 24 % in forests.
Grasslands, which have higher fine root biomass and root turnover compared to forests, also have higher rates of root decomposition. Our results further show that at the regional scale fine root decomposition is influenced by environmental variables such as soil moisture, soil temperature and soil nutrient content. Additional variation is explained by root litter quality.
Eight polymorphic microsatellite loci were developed for the brook lamprey Lampetra planeri through 454 sequencing and their usefulness was tested in 45 individuals of both L. planeri and the river lamprey Lampetra fluviatilis. The number of alleles per loci ranged between two and five; the Italian and Irish populations had a mean expected heterozygosity of 0.388 and 0.424 and a mean observed heterozygosity of 0.418 and 0.411, respectively. (C) 2014 The Fisheries Society of the British Isles
We report herein on the polymer-crystallization-assisted thiol-ene photosynthesis of an amphiphilic comb/graft DNA copolymer, or molecular brush, composed of a hydrophobic poly(2-oxazoline) backbone and hydrophilic short single-stranded nucleic acid grafts. Coupling efficiencies are above 60% and thus higher as compared with the straight solid-phase-supported synthesis of amphiphilic DNA block copolymers. The DNA molecular brushes self-assemble into sub-micron-sized spherical structures in water as evidenced by light scattering as well as atomic force and electron microscopy imaging. The nucleotide sequences remain functional, as assessed by UV and fluorescence spectroscopy subsequent to isoindol synthesis at the surface of the structures. The determination of a vesicular morphology is supported by encapsulation and subsequent spectroscopy monitoring of the release of a water-soluble dye and spectroscopic quantification of the hybridization efficiency (30% in average) of the functional nucleic acid strands engaged in structure formation: about one-half of the nucleotide sequences are available for hybridization, whereas the other half are hindered within the self-assembled structure. Because speciation between complementary and non complementary sequences in the medium could be ascertained by confocal laser scanning microscopy, the stable self-assembled molecular brushes demonstrate the potential for sensing applications.
D-Peptides have been attributed pharmacological advantages over regular L-peptides, yet design rules are largely unknown. Based on a designed coiled coil-like D/L heterotetramer, named L-Base/D-Acid, we generated a library offering alternative residues for interaction with the D-peptide. Phage display selection yielded one predominant peptide, named HelixA, that differed at 13 positions from the scaffold helix. In addition to the observed D-/L-heterotetramers, ratio-dependent intermediate states were detected by isothermal titration calorimetry. Importantly, the formation of the selected HelixA/D-Acid bundle passes through fewer intermediate states than L-Base/D-Acid. Back mutation of HelixA core residues to L-Base (HelixLL) revealed that the residues at e/g-positions are responsible for the different intermediates. Furthermore, a Val-core variant (PeptideVV) was completely devoid of binding D-Acid, whereas an Ile-core helix (HelixII) interacted with D-Acid in a significantly more specific complex than L-Base.
Transfer RNAs (tRNAs) through their abundance and modification pattern significantly influence protein translation. Here, we present a systematic analysis of the tRNAome of Lactococcus lactis. Using the next-generation sequencing approach, we identified 40 tRNAs which carry 16 different post-transcriptional modifications as revealed by mass spectrometry analysis. While small modifications are located in the tRNA body, hypermodified nucleotides are mainly present in the anticodon loop, which through wobbling expand the decoding potential of the tRNAs. Using tRNA-based microarrays, we also determined the dynamics in tRNA abundance upon changes in the growth rate and heterologous protein overexpression stress. With a fourfold increase in the growth rate, the relative abundance of tRNAs cognate to low abundance codons decrease, while the tRNAs cognate to major codons remain mostly unchanged. Significant changes in the tRNA abundances are observed upon protein overexpression stress, which does not correlate with the codon usage of the overexpressed gene but rather reflects the altered expression of housekeeping genes.
Steroidal glycoalkaloids (SGAs) are nitrogen-containing secondary metabolites of the Solanum species, which are known to have large chemical and bioactive diversity in nature. While recent effort and development on LC/MS techniques for SGA profiling have elucidated the main pathways of SGA metabolism in tomato, the problem of peak annotation still remains due to the vast diversity of chemical structure and similar on overlapping of chemical formula. Here we provide a case study of peak classification and annotation approach by integration of species and tissue specificities of SGA accumulation for provision of comprehensive pathways of SGA biosynthesis. In order to elucidate natural diversity of SGA biosynthesis, a total of 169 putative SGAs found in eight tomato accessions (Solanum lycopersicum, S. pimpinellifolium, S. cheesmaniae, S. chmielewskii, S. neorickii, S. peruvianum, S. habrochaites, S. pennellii) and four tissue types were used for correlation analysis. The results obtained in this study contribute annotation and classification of SGAs as well as detecting putative novel biosynthetic branch points. As such this represents a novel strategy for peak annotation for plant secondary metabolites.
Growth of plant organs relies on cell proliferation and expansion. While an increasingly detailed picture about the control of cell proliferation is emerging, our knowledge about the control of cell expansion remains more limited. We demonstrate the internal-motor kinesin AtKINESIN-13A (AtKIN13A) limits cell expansion and cell size in Arabidopsis thaliana, ion atkinl3a mutants forming larger petals with larger cells. The homolog, AtKINESIN-13B, also affects cell expansion and double mutants display growth, gametophytic and early embryonic defects, indicating a redundant role of he two genes. AtKIN13A is known to depolymerize microtubules and influence Golgi motility and distribution. Consistent his function, AtKIN13A interacts genetically with ANGUSTIFOLIA, encoding a regulator of Golgi dynamics. Reduced AtIGN13A activity alters cell wall structure as assessed by Fourier-transformed infrared-spectroscopy and triggers signalling he THESEUS1-dependent cell-wall integrity pathway, which in turn promotes the excess cell expansion in the atkinl3a mutant. Thus, our results indicate that the intracellular activity of AtKIN13A regulates cell expansion and wall architecture via THESEUS1, providing a compelling case of interplay between cell wall integrity sensing and expansion.
The ubiquitous InsP(3)/Ca2+ signalling pathway is modulated by diverse mechanisms, i.e. feedback of Ca2+ and interactions with other signalling pathways. In the salivary glands of the blowfly Calliphora vicina, the hormone serotonin (5-HT) causes a parallel rise in intracellular [Ca2+] and [cAMP] via two types of 5-HT receptors. We have shown recently that cAMP/protein kinase A (PKA) sensitizes InsP(3)-induced Ca2+ release. We have now identified the protein phosphatase that counteracts the effect of PKA on 5-HT-induced InsP(3)/Ca2+ signalling. We demonstrate that (1) tautomycin and okadaic acid, inhibitors of protein phosphatases PP1 and PP2A, have no effect on 5-HT-induced Ca2+ signals; (2) cyclosporin A and FK506, inhibitors of Ca2+/calmodulin-activated protein phosphatase calcineurin, cause an increase in the frequency of 5-HT-induced Ca2+ oscillations; (3) the sensitizing effect of cyclosporin A on 5-HT-induced Ca2+ responses does not involve Ca2+ entry into the cells; (4) cyclosporin A increases InsP(3)-dependent Ca2+ release; (5) inhibition of PKA abolishes the effect of cyclosporin A on the 5-HT-induced Ca2+ responses, indicating that PKA and calcineurin act antagonistically on the InsP(3)/Ca2+ signalling pathway. These findings suggest that calcineurin provides a negative feedback on InsP(3)/Ca2+ signalling in blowfly salivary glands, counteracting the effect of PKA and desensitizing the signalling cascade at higher 5-HT concentrations. (C) 2014 Elsevier Ltd. All rights reserved.
For the first time the direct electron transfer of an enzyme - cellobiose dehydrogenase, CDH - has been coupled with the hexokinase catalyzed competition for glucose in a sensor for ATP. To enhance the signal output for ATP, pyruvate kinase was coimmobilized to recycle ADP by the phosphoenolpyruvate driven reaction. The new sensor overcomes the limit of 1:1 stoichiometry of the sequential or competitive conversion of ATP by effective enzymatic recycling of the analyte. The anodic oxidation of the glucose converting CDH proceeds at electrode potentials below 0 mV vs. Ag vertical bar AgCl thus potentially interfering substances like ascorbic acid or catecholamines do not influence the measuring signal. The combination of direct electron transfer of CDH with the enzymatic recycling results in an interference-free and oxygen-independent measurement of ATP in the lower mu molar concentration range with a lower limit of detection of 63.3 nM (S/N=3).
Objectives: Modern human life style has led to significant decrease in everyday physical activity and bipedal locomotion. It has previously been shown that skeletal robustness (relative elbow breadth) is associated with daily step counts. The aim of the study was to investigate whether also other skeletal measures, particularly pelvic breadth may have changed in recent decades.
Methods: We re-analyzed elbow breadth, pelvic breadth (bicristal), and thoracic depth and breadth, of up to 28,975 healthy females and 28,288 healthy males aged 3-18 years from cross-sectional anthropological surveys performed between 1980 and 2012 by the Universities of Potsdam and Berlin, Germany.
Results: Relative elbow breadth (Frame index) significantly decreased in both sexes since 1980 (< 0.001). The trend toward slighter built was even more pronounced in absolute and relative pelvic breadth. In contrast, equivalent changes of parts of the skeletal system that are not involved in bipedal locomotion such as thoracic breadth, thoracic depth, and the thoracic index were absent.
Conclusions: The present investigation confirms the decline in relative elbow breadth in recent decades. Analogue, but even more pronounced changes were detected in pelvic breadth that coincides with the modern decline in upright locomotion. The findings underscore the phenotypic plasticity of humans while adapting to new environmental conditions. (C) 2014Wiley Periodicals, Inc.
Arthropods are incapable of synthesizing sterols de novo and thus require a dietary source to cover their physiological demands. The most prominent sterol in animal tissues is cholesterol, which is an indispensable structural component of cell membranes and serves as precursor for steroid hormones. Instead of cholesterol, plants and algae contain a variety of different phytosterols. Consequently, herbivorous arthropods have to metabolize dietary phytosterols to cholesterol to meet their requirements for growth and reproduction. Here, we investigated sterol-limited growth responses of the freshwater herbivore Daphnia magna by supplementing a sterol-free diet with increasing amounts of 10 different phytosterols and comparing thresholds for sterol-limited growth. In addition, we analyzed the sterol composition of D. magna to explore sterol metabolic constraints and bioconversion capacities. We show that dietary phytosterols strongly differ in their potential to support somatic growth of D. magna. The dietary threshold concentrations obtained by supplementing the different sterols cover a wide range (3.5-34.4 mu g mg C-1) and encompass the one for cholesterol (8.9 mu g mg C-1), indicating that certain phytosterols are more efficient in supporting somatic growth than cholesterol (e.g., fucosterol, brassicasterol) while others are less efficient (e.g., dihydrocholesterol, lathosterol). The dietary sterol concentration gradients revealed that the poor quality of particular sterols can be alleviated partially by increasing dietary concentrations, and that qualitative differences among sterols are most pronounced at low to moderate dietary concentrations. We infer that the dietary sterol composition has to be considered in zooplankton nutritional ecology to accurately assess potential sterol limitations under field conditions.
Intensive land use is a driving force for biodiversity decline in many ecosystems. In semi-natural grasslands, land-use activities such as mowing, grazing and fertilization affect the diversity of plants and arthropods, but the combined effects of different drivers and the chain of effects are largely unknown. In this study we used structural equation modelling to analyse how the arthropod communities in managed grasslands respond to land use and whether these responses are mediated through changes in resource diversity or resource quantity (biomass). Plants were considered resources for herbivores which themselves were considered resources for predators. Plant and arthropod (herbivores and predators) communities were sampled on 141 meadows, pastures and mown pastures within three regions in Germany in 2008 and 2009. Increasing land-use intensity generally increased plant biomass and decreased plant diversity, mainly through increasing fertilization. Herbivore diversity decreased together with plant diversity but showed no response to changes in plant biomass. Hence, land-use effects on herbivore diversity were mediated through resource diversity rather than quantity. Land-use effects on predator diversity were mediated by both herbivore diversity (resource diversity) and herbivore quantity (herbivore biomass), but indirect effects through resource quantity were stronger. Our findings highlight the importance of assessing both direct and indirect effects of land-use intensity and mode on different trophic levels. In addition to the overall effects, there were subtle differences between the different regions, pointing to the importance of regional land-use specificities. Our study underlines the commonly observed strong effect of grassland land use on biodiversity. It also highlights that mechanistic approaches help us to understand how different land-use modes affect biodiversity.
Classic theories of ageing consider extrinsic mortality (EM) a major factor in shaping longevity and ageing, yet most studies of functional ageing focus on species with low EM. This bias may cause overestimation of the influence of senescent declines in performance over condition-dependent mortality on demographic processes across taxa. To simultaneously investigate the roles of functional senescence (FS) and intrinsic, extrinsic and condition-dependent mortality in a species with a high predation risk in nature, we compared age trajectories of body mass (BM) in wild and captive grey mouse lemurs (Microcebus murinus) using longitudinal data (853 individuals followed through adulthood). We found evidence of non-random mortality in both settings. In captivity, the oldest animals showed senescence in their ability to regain lost BM, whereas no evidence of FS was found in the wild. Overall, captive animals lived longer, but a reversed sex bias in lifespan was observed between wild and captive populations. We suggest that even moderately condition-dependent EM may lead to negligible FS in the wild. While high EM may act to reduce the average lifespan, this evolutionary process may be counteracted by the increased fitness of the long-lived, high-quality individuals.
The disease risk and age of onset of Huntington disease (HD) and nine other repeat disorders strongly depend on the expansion of CAG repeats encoding consecutive polyglutamines (polyQ) in the corresponding disease protein. PolyQ length-dependent misfolding and aggregation are the hallmarks of CAG pathologies. Despite intense effort, the overall structure of these aggregates remains poorly understood. Here, we used sensitive time-dependent fluorescent decay measurements to assess the architecture of mature fibrils of huntingtin (Htt) exon 1 implicated in HD pathology. Varying the position of the fluorescent labels in the Htt monomer with expanded 51Q (Htt51Q) and using structural models of putative fibril structures, we generated distance distributions between donors and acceptors covering all possible distances between the monomers or monomer dimensions within the polyQ amyloid fibril. Using Monte Carlo simulations, we systematically scanned all possible monomer conformations that fit the experimentally measured decay times. Monomers with four-stranded 51Q stretches organized into five-layered beta-sheets with alternating N termini of the monomers perpendicular to the fibril axis gave the best fit to our data. Alternatively, the core structure of the polyQ fibrils might also be a zipper layer with antiparallel four-stranded stretches as this structure showed the next best fit. All other remaining arrangements are clearly excluded by the data. Furthermore, the assessed dimensions of the polyQ stretch of each monomer provide structural evidence for the observed polyQ length threshold in HD pathology. Our approach can be used to validate the effect of pharmacological substances that inhibit or alter amyloid growth and structure.
Forest management is known to influence species diversity of various taxa but inconsistent or even contrasting effects are reported for arthropods. Regional differences in management as well as differences in regional species pools might be responsible for these inconsistencies, but, inter-regional replicated studies that account for regional variability are rare. We investigated the effect of forest type on the abundance, diversity, community structure and composition of two important ground-dwelling beetle families, Carabidae and Staphylinidae, in 149 forest stands distributed over three regions in Germany. In particular we focused on recent forestry history, stand age and dominant tree species, in addition to a number of environmental descriptors. Overall management effects on beetle communities were small and mainly mediated by structural habitat parameters such as the cover of forest canopy or the plant diversity on forest stands. The general response of both beetle taxa to forest management was similar in all regions: abundance and species richness of beetles was higher in older than in younger stands and species richness was lower in unmanaged than in managed stands. The abundance ratio of forest species-to-open habitat species differed between regions, but generally increased from young to old stands, from coniferous to deciduous stands and from managed to unmanaged stands. The response of both beetle families to dominant tree species was variable among regions and staphylinid richness varied in the response to recent forestry history. Our results suggest that current forest management practices change the composition of ground-dwelling beetle communities mainly by favoring generalists and open habitat species. To protect important forest beetle communities and thus the ecosystem functions and services provided by them, we suggest to shelter remaining ancient forests and to develop near-to-nature management strategies by prolonging rotation periods and increasing structural diversity of managed forests. Possible geographic variations in the response of beetle communities need to be considered in conservation-orientated forest management strategies. (C) 2014 Elsevier B.V. All rights reserved.
Catalytic bio-chemo and bio-bio tandem oxidation reactions for amide and carboxylic acid synthesis
(2014)
A catalytic toolbox for three different water-based one-pot cascades to convert aryl alcohols to amides and acids and cyclic amines to lactams, involving combination of oxidative enzymes (monoamine oxidase, xanthine dehydrogenase, galactose oxidase and laccase) and chemical oxidants (TBHP or Cul(cat)/H2O2) at mild temperatures, is presented. Mutually compatible conditions were found to afford products in good to excellent yields.
Octopamine is an important neurotransmitter in insects with multiple functions. Here, we investigated the role of this amine in a simple form of learning (habituation) in the fruit fly Drosophila melanogaster. Specifically, we asked if octopamine is necessary for normal habituation of a proboscis extension response (PER) to different sucrose concentrations. In addition, we analyzed the relationship between responsiveness to sucrose solutions applied to the tarsus and habituation of the proboscis extension response in the same individual. The Tyramine-beta-hydroxylase (T beta h) mutant lacks the enzyme catalyzing the final step of octopamine synthesis. This mutant was significantly less responsive to sucrose than controls. The reduced responsiveness directly led to faster habituation. Systemic application of octopamine or induction of octopamine synthesis by T beta h expression in a cluster of octopaminergic neurons within the suboesophageal ganglion restored sucrose responsiveness and habituation of octopamine mutants to control level. Further analyses imply that the reduced sucrose responsiveness of T beta h mutants is related to a lower sucrose preference, probably due to a changed carbohydrate metabolism, since T beta h mutants survived significantly longer under starved conditions. These findings suggest a pivotal role for octopamine in regulating sucrose responsiveness in fruit flies. Further, octopamine indirectly influences non-associative learning and possibly associative appetitive learning by regulating the evaluation of the sweet component of a sucrose reward. (C) 2014 Elsevier Ltd. All rights reserved.
The Great Hungarian Plain was a crossroads of cultural transformations that have shaped European prehistory. Here we analyse a 5,000-year transect of human genomes, sampled from petrous bones giving consistently excellent endogenous DNA yields, from 13 Hungarian Neolithic, Copper, Bronze and Iron Age burials including two to high (similar to 22x) and seven to similar to 1x coverage, to investigate the impact of these on Europe's genetic landscape. These data suggest genomic shifts with the advent of the Neolithic, Bronze and Iron Ages, with interleaved periods of genome stability. The earliest Neolithic context genome shows a European hunter-gatherer genetic signature and a restricted ancestral population size, suggesting direct contact between cultures after the arrival of the first farmers into Europe. The latest, Iron Age, sample reveals an eastern genomic influence concordant with introduced Steppe burial rites. We observe transition towards lighter pigmentation and surprisingly, no Neolithic presence of lactase persistence.
We report on the development of an on-chip RPA (recombinase polymerase amplification) with simultaneous multiplex isothermal amplification and detection on a solid surface. The isothermal RPA was applied to amplify specific target sequences from the pathogens Neisseria gonorrhoeae, Salmonella enterica and methicillin-resistant Staphylococcus aureus (MRSA) using genomic DNA. Additionally, a positive plasmid control was established as an internal control. The four targets were amplified simultaneously in a quadruplex reaction. The amplicon is labeled during on-chip RPA by reverse oligonucleotide primers coupled to a fluorophore. Both amplification and spatially resolved signal generation take place on immobilized forward primers bount to expoxy-silanized glass surfaces in a pump-driven hybridization chamber. The combination of microarray technology and sensitive isothermal nucleic acid amplification at 38 degrees C allows for a multiparameter analysis on a rather small area. The on-chip RPA was characterized in terms of reaction time, sensitivity and inhibitory conditions. A successful enzymatic reaction is completed in < 20 min and results in detection limits of 10 colony-forming units for methicillin-resistant Staphylococcus aureus and Salmonella enterica and 100 colony-forming units for Neisseria gonorrhoeae. The results show this method to be useful with respect to point-of-care testing and to enable simplified and miniaturized nucleic acid-based diagnostics.
The Kv-like (potassium voltage-dependent) K+ channels at the plasma membrane, including the inward-rectifying KAT1 K+ channel of Arabidopsis (Arabidopsis thaliana), are important targets for manipulating K+ homeostasis in plants. Gating modification, especially, has been identified as a promising means by which to engineer plants with improved characteristics in mineral and water use. Understanding plant K+ channel gating poses several challenges, despite many similarities to that of mammalian Kv and Shaker channel models. We have used site-directed mutagenesis to explore residues that are thought to form two electrostatic countercharge centers on either side of a conserved phenylalanine (Phe) residue within the S2 and S3 alpha-helices of the voltage sensor domain (VSD) of Kv channels. Consistent with molecular dynamic simulations of KAT1, we show that the voltage dependence of the channel gate is highly sensitive to manipulations affecting these residues. Mutations of the central Phe residue favored the closed KAT1 channel, whereas mutations affecting the countercharge centers favored the open channel. Modeling of the macroscopic current kinetics also highlighted a substantial difference between the two sets of mutations. We interpret these findings in the context of the effects on hydration of amino acid residues within the VSD and with an inherent bias of the VSD, when hydrated around a central Phe residue, to the closed state of the channel.
Carbohydrate metabolism in plants is tightly linked to photosynthesis and is essential for energy and carbon skeleton supply of the entire organism. Thus, the hexose phosphate pools of the cytosol and the chloroplast represent important metabolic resources that are maintained through action of phosphoglucose isomerase (PGI) and phosphoglucose mutase interconverting glucose 6-phosphate, fructose 6-phosphate, and glucose 1-phosphate. Here, we investigated the impact of disrupted cytosolic PGI (cPGI) function on plant viability and metabolism. Overexpressing an artificial microRNA targeted against cPGI (amiR-cpgi) resulted in adult plants with vegetative tissue essentially free of cPGI activity. These plants displayed diminished growth compared with the wild type and accumulated excess starch in chloroplasts but maintained low sucrose content in leaves at the end of the night. Moreover, amiR-cpgi plants exhibited increased nonphotochemical chlorophyll a quenching during photosynthesis. In contrast to amiR-cpgi plants, viable transfer DNA insertion mutants disrupted in cPGI function could only be identified as heterozygous individuals. However, homozygous transfer DNA insertion mutants could be isolated among plants ectopically expressing cPGI. Intriguingly, these plants were only fertile when expression was driven by the ubiquitin10 promoter but sterile when the seed-specific unknown seed protein promoter or the Cauliflower mosaic virus 35S promoter were employed. These data show that metabolism is apparently able to compensate for missing cPGI activity in adult amiR-cpgi plants and indicate an essential function for cPGI in plant reproduction. Moreover, our data suggest a feedback regulation in amiR-cpgi plants that fine-tunes cytosolic sucrose metabolism with plastidic starch turnover.
Insects and their six-legged relatives (Hexapoda) comprise more than half of all described species and dominate terrestrial and freshwater ecosystems. Understanding the macroevolutionary processes generating this richness requires a historical perspective, but the fossil record of hexapods is patchy and incomplete. Dated molecular phylogenies provide an alternative perspective on divergence times and have been combined with birth-death models to infer patterns of diversification across a range of taxonomic groups. Here we generate a dated phylogeny of hexapod families, based on previously published sequence data and literature derived constraints, in order to identify the broad pattern of macroevolutionary changes responsible for the composition of the extant hexapod fauna. The most prominent increase in diversification identified is associated with the origin of complete metamorphosis, confirming this as a key innovation in promoting insect diversity. Subsequent reductions are recovered for several groups previously identified as having a higher fossil diversity during the Mesozoic. In addition, a number of recently derived taxa are found to have radiated following the development of flowering plant (angiosperm) floras during the mid-Cretaceous. These results reveal that the composition of the modern hexapod fauna is a product of a key developmental innovation, combined with multiple and varied evolutionary responses to environmental changes from the mid Cretaceous floral transition onward.
Treating creationism as a controversial topic within the science and religion issue in the science classroom has been widely discussed in the recent literature. Some researchers have proposed that this topic is best addressed by focusing on sociocognitive conflict. To prepare new learning opportunities for this approach, it is necessary to know the concrete arguments that students use in their discussions on this issue. Therefore, this study aimed to provide a systematic description of these arguments. For this purpose, upper secondary students (N=43) argued for either the acceptance of evolutionary theory or faith in Genesis in a written speech. The study was conducted during their regular biology and religious education classes. Generated arguments were analysed by qualitative content analysis. Three dimensions of the arguments were described: the content (science or religion), the valuation of the argument (positive or negative), and whether the argument consisted of a descriptive or normative argumentation. The results indicate that students found it easier to generate arguments about the scientific side of the issue; however, these arguments were negatively constructed. The results are discussed with regard to implications for educational approaches for teaching controversial issues at the high-school level.
Changes in carbon flow and sink/source activities can affect floral, architectural, and reproductive traits of plants. In potato, overexpression (OE) of the purple acid phosphatase 2 of Arabidopsis (AtPAP2) resulted in earlier flowering, faster growth rate, increased tubers and tuber starch content, and higher photosynthesis rate. There was a significant change in sucrose, glucose and fructose levels in leaves, phloem and sink biomass of the OE lines, consistent with an increased expression of sucrose transporter 1 (StSUT1). Furthermore, the expression levels and enzyme activity of sucrose-phosphate synthase (SPS) were also significantly increased in the OE lines. These findings strongly suggest that higher carbon supply from the source and improved sink strength can improve potato tuber yield. (C) 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
The continuous expansion of nosocomial infections around the globe has become a precarious situation. Key challenges include mounting dissemination of multiple resistances to antibiotics, the easy transmission and the growing mortality rates of hospital-acquired bacterial diseases. Thus, new ways to rapidly detect these infections are vital. Consequently, researchers around the globe pursue innovative approaches for point-of-care devices. In many cases the specific interaction of an antigen and a corresponding antibody is pivotal. However, the knowledge about suitable antigens is lacking. The aim of this study was to identify novel antigens as specific diagnostic markers. Additionally, these proteins might be aptly used for the generation of vaccines to improve current treatment options. Hence, a cDNA-based expression library was constructed and screened via microarrays to detect novel antigens of Klebsiella pneumoniae, a prominent agent of nosocomial infections well-known for its extensive antibiotics resistance, especially by extended-spectrum beta-lactamases (ESBL). After screening 1536 clones, 14 previously unknown immunogenic proteins were identified. Subsequently, each protein was expressed in full-length and its immunodominant character examined by ELISA and microarray analyses. Consequently, six proteins were selected for epitope mapping and three thereof possessed linear epitopes. After specificity analysis, homology survey and 3d structural modelling, one epitope sequence GAVVALSTTFA of KPN_00363, an ion channel protein, was identified harboring specificity for K. pneumoniae. The remaining epitopes showed ambiguous results regarding the specificity for K. pneumoniae. The approach adopted herein has been successfully utilized to discover novel antigens of Campylobacter jejuni and Salmonella enterica antigens before. Now, we have transferred this knowledge to the key nosocomial agent, K. pneumoniae. By identifying several novel antigens and their linear epitope sites, we have paved the way for crucial future research and applications including the design of point-of-care devices, vaccine development and serological screenings for a highly relevant nosocomial pathogen.
African weakly electric fishes (Mormyridae) underwent an outstanding adaptive radiation (about 200 species), putatively owing to their ability to communicate through species-specific weak electric signals. The electric organ discharge (EOD) is produced by muscle-derived electrocytes organized in piles to form an electric organ. Despite the importance of this trait as a prezygotic isolation mechanism, genomic resources remained limited. We present here a first draft of the skeletal muscle and electric organ transcriptomes from the weakly electric fish species Campylomormyrus compressirostris, obtained using the Illumina HiSeq2000 sequencing technology. Approximately 6.8 Gbp of cDNA sequence data were produced from both tissues, resulting in 57268109 raw reads for the skeletal muscle and 46934923 for the electric organ, and assembled de novo into 46143 and 89270 contigs, respectively. About 50% of both transcriptomes were annotated after protein databases search. The two transcriptomes show similar profiles in terms of Gene Ontology categories composition. We identified several candidate genes which are likely to play a central role in the production and evolution of the electric signal. For most of these genes, and for many other housekeeping genes, we were able to obtain the complete or partial coding DNA sequences (CDS), which can be used for the development of primers to be utilized in qRT-PCR experiments. We present also the complete mitochondrial genome and compare it to those available from other weakly electric fish species. Additionally, we located 1671 SSR-containing regions with their flanking sites and designed the relative primers. This study establishes a first step in the development of genomic tools aimed at understanding the role of electric communication during speciation.
Serving many at once: How a database approach can create unity in dynamical ecosystem modelling
(2014)
Simulation modelling in ecology is a field that is becoming increasingly compartmentalized. Here we propose a Database Approach To Modelling (DATM) to create unity in dynamical ecosystem modelling with differential equations. In this approach the storage of ecological knowledge is independent of the language and platform in which the model will be run. To create an instance of the model, the information in the database is translated and augmented with the language and platform specifics. This process is automated so that a new instance can be created each time the database is updated. We describe the approach using the simple Lotka-Volterra model and the complex ecosystem model for shallow lakes PCLake, which we automatically implement in the frameworks OSIRIS, GRIND for MATLAB, ACSL, R, DUFLOW and DELWAQ. A clear advantage of working in a database is the overview it provides. The simplicity of the approach only adds to its elegance. (C) 2014 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-SA license (http://creativecommons.org/licenses/by-nc-sa/3.0/).
In Australia, the rabbit haemorrhagic disease virus (RHDV) has been used since 1996 to reduce numbers of introduced European rabbits (Oryctolagus cuniculus) which have a devastating impact on the native Australian environment. RHDV causes regular, short disease outbreaks, but little is known about how the virus persists and survives between epidemics. We examined the initial spread of RHDV to show that even upon its initial spread, the virus circulated continuously on a regional scale rather than persisting at a local population level and that Australian rabbit populations are highly interconnected by virus-carrying flying vectors. Sequencing data obtained from a single rabbit population showed that the viruses that caused an epidemic each year seldom bore close genetic resemblance to those present in previous years. Together, these data suggest that RHDV survives in the Australian environment through its ability to spread amongst rabbit subpopulations. This is consistent with modelling results that indicated that in a large interconnected rabbit meta-population, RHDV should maintain high virulence, cause short, strong disease outbreaks but show low persistence in any given subpopulation. This new epidemiological framework is important for understanding virus-host co-evolution and future disease management options of pest species to secure Australia's remaining natural biodiversity.
In food webs, herbivores are often constrained by low food quality in terms of mineral and biochemical limitations, which in aquatic ecosystems can co-occur with limited oxygen conditions. As low food quality implies that carbon (C) is available in excess, and therefore a regulation to get rid of excess C is crucial for the performance of consumers, we examined the C pathways (ingestion, feces release, excretion, and respiration) of a planktonic key herbivore (Daphnia magna). We tested whether consumer C pathways increase due to mineral (phosphorus, P) or biochemical (cholesterol and fatty acid) limitations and how these regulations vary when in addition oxygen is low. Under such conditions, at least the capability of the upregulation of respiration may be restricted. Furthermore, we discussed the potential role of the oxygen-transporting protein hemoglobin (Hb) in the regulation of C budgets. Different food quality constraints led to certain C regulation patterns to increase the removal of excess dietary C: P-limited D. magna increased excretion and respiration, while cholesterol-limited Daphnia in addition upregulated the release of feces. In contrast, the regulative effort was low and only feces release increased when D. magna was limited by a long-chain polyunsaturated fatty acid (eicosapentaenoic acid, EPA). Co-limiting oxygen did not always impact the discharge of excess C. We found the food-quality-induced upregulation of respiration was still present at low oxygen. In contrast, higher excretion of excess C was diminished at low oxygen supply. Besides the effect that the Hb concentration increased under low oxygen, our results indicate a low food-quality-induced increase in the Hb content of the animals. Overall, C budgeting is phenotypically plastic towards different (co-) limiting scenarios. These trigger specific regulation responses that could be the result of evolutionary adaptations.
Despite the success of highly active antiretroviral therapy (HAART) in the management of human immunodeficiency virus (HIV)-1 infection, virological failure due to drug resistance development remains a major challenge. Resistant mutants display reduced drug susceptibilities, but in the absence of drug, they generally have a lower fitness than the wild type, owing to a mutation-incurred cost. The interaction between these fitness costs and drug resistance dictates the appearance of mutants and influences viral suppression and therapeutic success. Assessing in vivo viral fitness is a challenging task and yet one that has significant clinical relevance. Here, we present a new computational modelling approach for estimating viral fitness that relies on common sparse cross-sectional clinical data by combining statistical approaches to learn drug-specific mutational pathways and resistance factors with viral dynamics models to represent the host-virus interaction and actions of drug mechanistically. We estimate in vivo fitness characteristics of mutant genotypes for two antiretroviral drugs, the reverse transcriptase inhibitor zidovudine (ZDV) and the protease inhibitor indinavir (IDV). Well-known features of HIV-1 fitness landscapes are recovered, both in the absence and presence of drugs. We quantify the complex interplay between fitness costs and resistance by computing selective advantages for different mutants. Our approach extends naturally to multiple drugs and we illustrate this by simulating a dual therapy with ZDV and IDV to assess therapy failure. The combined statistical and dynamical modelling approach may help in dissecting the effects of fitness costs and resistance with the ultimate aim of assisting the choice of salvage therapies after treatment failure.
Myriapods (e. g., centipedes and millipedes) display a simple homonomous body plan relative to other arthropods. All members of the class are terrestrial, but they attained terrestriality independently of insects. Myriapoda is the only arthropod class not represented by a sequenced genome. We present an analysis of the genome of the centipede Strigamia maritima. It retains a compact genome that has undergone less gene loss and shuffling than previously sequenced arthropods, and many orthologues of genes conserved from the bilaterian ancestor that have been lost in insects. Our analysis locates many genes in conserved macro-synteny contexts, and many small-scale examples of gene clustering. We describe several examples where S. maritima shows different solutions from insects to similar problems. The insect olfactory receptor gene family is absent from S. maritima, and olfaction in air is likely effected by expansion of other receptor gene families. For some genes S. maritima has evolved paralogues to generate coding sequence diversity, where insects use alternate splicing. This is most striking for the Dscam gene, which in Drosophila generates more than 100,000 alternate splice forms, but in S. maritima is encoded by over 100 paralogues. We see an intriguing linkage between the absence of any known photosensory proteins in a blind organism and the additional absence of canonical circadian clock genes. The phylogenetic position of myriapods allows us to identify where in arthropod phylogeny several particular molecular mechanisms and traits emerged. For example, we conclude that juvenile hormone signalling evolved with the emergence of the exoskeleton in the arthropods and that RR-1 containing cuticle proteins evolved in the lineage leading to Mandibulata. We also identify when various gene expansions and losses occurred. The genome of S. maritima offers us a unique glimpse into the ancestral arthropod genome, while also displaying many adaptations to its specific life history.
A novel method that optimizes the screening for antibody-secreting hapten-specific hybridoma cells by using flow cytometry is described. Cell clones specific for five different haptens were analyzed. We selectively double stained and analyzed fixed hybridoma cells with fluorophore-labeled haptens to demonstrate the target-selectivity, and with a fluorophore-labeled anti-mouse IgG antibody to characterize the level of surface expression of membrane-bound IgGs. ELISA measurements with the supernatants of the individual hybridoma clones revealed that antibodies from those cells, which showed the highest fluorescence intensities in the flow cytometric analysis, also displayed the highest affinities for the target antigens. The fluorescence intensity of antibody-producing cells corresponded well with the produced antibodies' affinities toward their respective antigens. Immunohistochemical staining verified the successful double labeling of the cells. Our method makes it possible to perform a high-throughput screening for hybridoma cells, which have both an adequate IgG production rate and a high target affinity. (C) 2014 Elsevier B.V. All rights reserved.
Background: Human populations differ in height. Recent evidence suggests that social networks play an important role in the regulation of adolescent growth and adult height. We further investigated the effect of physical connectedness on height.
Material and methods: We considered Switzerland as a geographic network with 169 nodes (district capitals) and 335 edges (connecting roads) and studied effects of connectedness on height in Swiss conscript from 1884-1891, 1908-1910, and 2004-2009. We also created exponential-family random graph models to separate possible unspecific effects of geographic vicinity.
Results: In 1884-1891, in 1908-1910, and in 2004-2009, 1st, 2nd and 3rd order neighboring districts significantly correlate in height (p<0.01). The correlations depend on the order of connectedness, they decline with increasing distance. Short stature districts tend to have short, tall stature districts tend to have tall neighbors. Random network analyses suggest direct road effects on height. Whereas in 1884-1891, direct road effects were only visible between 1st order neighbors, direct road effects extended to 2nd and 3rd in 1908-1910, and in 2004-2009, also to 4th order neighbors, and might reflect historic improvements in transportation. The spatial correlations did not significantly change when height was controlled for goiter (1884-1889) and for median per capita income (2006), suggesting direct road effects also in goiter-allowed-for height and income-allowed-for height.
Conclusion: Height in a district depends on height of physically connected neighboring districts. The association decreases with increasing distance in the net. The present data suggest that people can be short because their neighbors are short; or tall because their neighbors are tall (community effect on growth). Psycho-biological effects seem to control growth and development within communities that go far beyond our current understanding of growth regulation.
We applied a top-down systems biology approach to understand how Chlamydomonas reinhardtii acclimates to long-term heat stress (HS) and recovers from it. For this, we shifted cells from 25 to 42 degrees C for 24 h and back to 25 degrees C for >= 8 h and monitored abundances of 1856 proteins/protein groups, 99 polar and 185 lipophilic metabolites, and cytological and photosynthesis parameters. Our data indicate that acclimation of Chlamydomonas to long-term HS consists of a temporally ordered, orchestrated implementation of response elements at various system levels. These comprise (1) cell cycle arrest; (2) catabolism of larger molecules to generate compounds with roles in stress protection; (3) accumulation of molecular chaperones to restore protein homeostasis together with compatible solutes; (4) redirection of photosynthetic energy and reducing power from the Calvin cycle to the de novo synthesis of saturated fatty acids to replace polyunsaturated ones in membrane lipids, which are deposited in lipid bodies; and (5) when sinks for photosynthetic energy and reducing power are depleted, resumption of Calvin cycle activity associated with increased photorespiration, accumulation of reactive oxygen species scavengers, and throttling of linear electron flow by antenna uncoupling. During recovery from HS, cells appear to focus on processes allowing rapid resumption of growth rather than restoring pre-HS conditions.
A detailed knowledge of cell wall heterogeneity and complexity is crucial for understanding plant growth and development. One key challenge is to establish links between polysaccharide-rich cell walls and their phenotypic characteristics. It is of particular interest for some plant material, like cotton fibers, which are of both biological and industrial importance. To this end, we attempted to study cotton fiber characteristics together with glycan arrays using regression based approaches. Taking advantage of the comprehensive microarray polymer profiling technique (CoMPP), 32 cotton lines from different cotton species were studied. The glycan array was generated by sequential extraction of cell wall polysaccharides from mature cotton fibers and screening samples against eleven extensively characterized cell wall probes. Also, phenotypic characteristics of cotton fibers such as length, strength, elongation and micronaire were measured. The relationship between the two datasets was established in an integrative manner using linear regression methods. In the conducted analysis, we demonstrated the usefulness of regression based approaches in establishing a relationship between glycan measurements and phenotypic traits. In addition, the analysis also identified specific polysaccharides which may play a major role during fiber development for the final fiber characteristics. Three different regression methods identified a negative correlation between micronaire and the xyloglucan and homogalacturonan probes. Moreover, homogalacturonan and callose were shown to be significant predictors for fiber length. The role of these polysaccharides was already pointed out in previous cell wall elongation studies. Additional relationships were predicted for fiber strength and elongation which will need further experimental validation.
Background: Kinetic characterization of wild-type xanthine dehydrogenase and variants. Results: Comparison of the pH dependence of both k(red) and k(red)/K-d, as well as k(cat) and k(cat)/K-m. Conclusion: Ionized Glu(232) of wild-type enzyme plays an important role in catalysis by discriminating against the monoanionic form of xanthine. Significance: Examining the contributions of Glu(232) to catalysis is essential for understanding the mechanism of xanthine dehydrogenase.
The kinetic properties of an E232Q variant of the xanthine dehydrogenase from Rhodobacter capsulatus have been examined to ascertain whether Glu(232) in wild-type enzyme is protonated or unprotonated in the course of catalysis at neutral pH. We find that k(red), the limiting rate constant for reduction at high [xanthine], is significantly compromised in the variant, a result that is inconsistent with Glu(232) being neutral in the active site of the wild-type enzyme. A comparison of the pH dependence of both k(red) and k(red)/K-d from reductive half-reaction experiments between wild-type and enzyme and the E232Q variant suggests that the ionized Glu(232) of wild-type enzyme plays an important role in catalysis by discriminating against the monoanionic form of substrate, effectively increasing the pK(a) of substrate by two pH units and ensuring that at physiological pH the neutral form of substrate predominates in the Michaelis complex. A kinetic isotope study of the wild-type R. capsulatus enzyme indicates that, as previously determined for the bovine and chicken enzymes, product release is principally rate-limiting in catalysis. The disparity in rate constants for the chemical step of the reaction and product release, however, is not as great in the bacterial enzyme as compared with the vertebrate forms. The results indicate that the bacterial and bovine enzymes catalyze the chemical step of the reaction to the same degree and that the faster turnover observed with the bacterial enzyme is due to a faster rate constant for product release than is seen with the vertebrate enzyme.
Ribonuclease H2 plays an essential role for genome stability as it removes ribonucleotides misincorporated into genomic DNA by replicative polymerases and resolves RNA/DNA hybrids. Biallelic mutations in the genes encoding the three RNase H2 subunits cause Aicardi-Goutieres syndrome (AGS), an early-onset inflammatory encephalopathy that phenotypically overlaps with the autoimmune disorder systemic lupus erythematosus. Here we studied the intracellular dynamics of RNase H2 in living cells during DNA replication and in response to DNA damage using confocal time-lapse imaging and fluorescence cross-correlation spectroscopy. We demonstrate that the RNase H2 complex is assembled in the cytosol and imported into the nucleus in an RNase H2B-dependent manner. RNase H2 is not only recruited to DNA replication foci, but also to sites of PCNA-dependent DNA repair. By fluorescence recovery after photobleaching, we demonstrate a high mobility and fast exchange of RNase H2 at sites of DNA repair and replication. We provide evidence that recruitment of RNase H2 is not only PCNA-dependent, mediated by an interaction of the B subunit with PCNA, but also PCNA-independent mediated via the catalytic domain of the A subunit. We found that AGS-associated mutations alter complex formation, recruitment efficiency and exchange kinetics at sites of DNA replication and repair suggesting that impaired ribonucleotide removal contributes to AGS pathogenesis.
Phosphoglucomutase (PGM) catalyses the interconversion of glucose 1-phosphate (G1P) and glucose 6-phosphate (G6P) and exists as plastidial (pPGM) and cytosolic (cPGM) isoforms. The plastidial isoform is essential for transitory starch synthesis in chloroplasts of leaves, whereas the cytosolic counterpart is essential for glucose phosphate partitioning and, therefore, for syntheses of sucrose and cell wall components. In Arabidopsis two cytosolic isoforms (PGM2 and PGM3) exist. Both PGM2 and PGM3 are redundant in function as single mutants reveal only small or no alterations compared to wild type with respect to plant primary metabolism. So far, there are no reports of Arabidopsis plants lacking the entire cPGM or total PGM activity, respectively. Therefore, amiRNA transgenic plants were generated and used for analyses of various parameters such as growth, development, and starch metabolism. The lack of the entire cPGM activity resulted in a strongly reduced growth revealed by decreased rosette fresh weight, shorter roots, and reduced seed production compared to wild type. By contrast content of starch, sucrose, maltose and cell wall components were significantly increased. The lack of both cPGM and pPGM activities in Arabidopsis resulted in dwarf growth, prematurely die off, and inability to develop a functional inflorescence. The combined results are discussed in comparison to potato, the only described mutant with lack of total PGM activity.
Background: The identification of brassinosteroid (BR) deficient and BR insensitive mutants provided conclusive evidence that BR is a potent growth-promoting phytohormone. Arabidopsis mutants are characterized by a compact rosette structure, decreased plant height and reduced root system, delayed development, and reduced fertility. Cell expansion, cell division, and multiple developmental processes depend on BR. The molecular and physiological basis of BR action is diverse. The BR signalling pathway controls the activity of transcription factors, and numerous BR responsive genes have been identified. The analysis of dwarf mutants, however, may to some extent reveal phenotypic changes that are an effect of the altered morphology and physiology. This restriction holds particularly true for the analysis of established organs such as rosette leaves. Results: In this study, the mode of BR action was analysed in established leaves by means of two approaches. First, an inhibitor of BR biosynthesis (brassinazole) was applied to 21-day-old wild-type plants. Secondly, BR complementation of BR deficient plants, namely CPD (constitutive photomorphogenic dwarf)-antisense and cbb1 (cabbage1) mutant plants was stopped after 21 days. BR action in established leaves is associated with stimulated cell expansion, an increase in leaf index, starch accumulation, enhanced CO2 release by the tricarboxylic acid cycle, and increased biomass production. Cell number and protein content were barely affected. Conclusion: Previous analysis of BR promoted growth focused on genomic effects. However, the link between growth and changes in gene expression patterns barely provided clues to the physiological and metabolic basis of growth. Our study analysed comprehensive metabolic data sets of leaves with altered BR levels. The data suggest that BR promoted growth may depend on the increased provision and use of carbohydrates and energy. BR may stimulate both anabolic and catabolic pathways.
In winter 2009/10, a sudden under-ice bloom of heterotrophic bacteria occurred in the seasonally ice-covered, temperate, deep, oligotrophic Lake Stechlin (Germany). Extraordinarily high bacterial abundance and biomass were fueled by the breakdown of a massive bloom of Aphanizomenon flos-aquae after ice formation. A reduction in light resulting from snow coverage exerted a pronounced physiological stress on the cyanobacteria. Consequently, these were rapidly colonized, leading to a sudden proliferation of attached and subsequently of free-living heterotrophic bacteria. Total bacterial protein production reached 201 mg C L-1 d(-1), ca. five times higher than spring-peak values that year. Fluorescence in situ hybridization and denaturing gradient gel electrophoresis at high temporal resolution showed pronounced changes in bacterial community structure coinciding with changes in the physiology of the cyanobacteria. Pyrosequencing of 16S rRNA genes revealed that during breakdown of the cyanobacterial population, the diversity of attached and free-living bacterial communities were reduced to a few dominant families. Some of these were not detectable during the early stages of the cyanobacterial bloom indicating that only specific, well adapted bacterial communities can colonize senescent cyanobacteria. Our study suggests that in winter, unlike commonly postulated, carbon rather than temperature is the limiting factor for bacterial growth. Frequent phytoplankton blooms in ice-covered systems highlight the need for year-round studies of aquatic ecosystems including the winter season to correctly understand element and energy cycling through aquatic food webs, particularly the microbial loop. On a global scale, such knowledge is required to determine climate change induced alterations in carbon budgets in polar and temperate aquatic systems.
Para-maleimidophenyl (p-MP) modified gold surfaces have been prepared by one-step electrochemical deposition and used in surface plasmon resonance (SPR) studies. Therefore, a FITC mimotope peptide (MP1, 12 aa), a human mucin 1 epitope peptide (MUC, 9 aa) and a protein with their specific antibodies were used as model systems. The peptides were modified with an N-terminal cysteine for covalent and directed coupling to the maleimido functionalized surface by means of Michael addition. The coupling yield of the peptide, the binding characteristics of antibody and the unspecific adsorption of the analytes were investigated. The results expand the spectrum of biosensors usable with p-MP by widely used SPR and support its potential to be versatile for several electrochemical and optical biosensors. This allows the combination of an electrochemical and optical read-out for a broad variety of biomolecular interactions on the same chip. Copyright (c) 2014 John Wiley & Sons, Ltd.
The honey bee (Apis melliferaL.) has developed into an important ethological model organism for social behaviour and behavioural plasticity. Bees perform a complex age-dependent division of labour with the most pronounced behavioural differences occurring between in-hive bees and foragers. Whereas nurse bees, for example, stay inside the hive and provide the larvae with food, foragers leave the hive to collect pollen and nectar for the entire colony. The biogenic amine octopamine appears to play a major role in division of labour but the molecular mechanisms involved are unknown. We here investigated the role of two characterized octopamine receptors in honey bee division of labour. AmOctR1 codes for a Ca2+-linked octopamine receptor. AmOctR3/4 codes for a cyclic adenosine monophosphate-coupled octopamine receptor. Messenger RNA expression of AmOctR1 in different brain neuropils correlates with social task, whereas expression of AmOctR3/4 changes with age rather than with social role per se. Our results for the first time link the regulatory role of octopamine in division of labour to specific receptors and brain regions. They are an important step forward in our understanding of complex behavioural organization in social groups.
Membranes of eukaryotic cells contain high lipid-order sterol-rich domains that are thought to mediate temporal and spatial organization of cellular processes. Sterols are crucial for execution of cytokinesis, the last stage of cell division, in diverse eukaryotes. The cell plate of higher-plant cells is the membrane structure that separates daughter cells during somatic cytokinesis. Cell-plate formation in Arabidopsis relies on sterol- and DYNAMIN-RELATED PROTEIN1A (DRP1A)-dependent endocytosis. However, functional relationships between lipid membrane order or lipid packing and endocytic machinery components during eukaryotic cytokinesis have not been elucidated. Using ratiometric live imaging of lipid order-sensitive fluorescent probes, we show that the cell plate of Arabidopsis thaliana represents a dynamic, high lipid-order membrane domain. The cell-plate lipid order was found to be sensitive to pharmacological and genetic alterations of sterol composition. Sterols co-localize with DRP1A at the cell plate, and DRP1A accumulates in detergent-resistant membrane fractions. Modifications of sterol concentration or composition reduce cell-plate membrane order and affect DRP1A localization. Strikingly, DRP1A function itself is essential for high lipid order at the cell plate. Our findings provide evidence that the cell plate represents a high lipid-order domain, and pave the way to explore potential feedback between lipid order and function of dynamin-related proteins during cytokinesis.
Influence of experimental soil disturbances on the diversity of plants in agricultural grasslands
(2014)
Disturbance is supposed to play an important role for biodiversity and ecosystem stability as described by the intermediate disturbance hypothesis (IDH), which predicts highest species richness at intermediate levels of disturbances. In this study, we tested the effects of artificial soil disturbances on diversity of annual and perennial vascular plants and bryophytes in a field experiment in 86 agricultural grasslands differing in land use in two regions of Germany. On each grassland, we implemented four treatments: three treatments differing in application time of soil disturbances and one control. One year after experimental disturbance, we recorded vegetation and measured biomass productivity and bare ground. We analysed the disturbance response taking effects of region and land-use-accompanied disturbance regimes into account.
Region and land-use type strongly determined plant species richness. Experimental disturbances had small positive effects on the species richness of annuals, but none on perennials or bryophytes. Bare ground was positively related to species richness of bryophytes. However, exceeding the creation of 12% bare ground further disturbance had a detrimental effect on bryophyte species richness, which corresponds to the IDH. As biomass productivity was unaffected by disturbance our results indicate that the disturbance effect on species richness of annuals was not due to decreased overall productivity, but rather due to short-term lowered inter- and intraspecific competition at the newly created microsites.
Generally, our results highlight the importance of soil disturbances for species richness of annual plants and bryophytes in agricultural grasslands. However, most grasslands were disturbed naturally or by land-use practices and our additional experimental soil disturbances only had a small short-term effect. Overall, total plant diversity in grasslands seemed to be more limited by the availability of propagules rather than by suitable microsites for germination. Thus, nature conservation efforts to increase grassland diversity should focus on overcoming propagule limitation, for instance by additional sowing of seeds, while the creation of additional open patches by disturbance might only be appropriate where natural disturbances are scarce.
For the ecological risk assessment of toxic chemicals, standardized tests on individuals are often used as proxies for population-level effects. Here, we address the utility of one commonly used metric, reproductive output, as a proxy for population-level effects. Because reproduction integrates the outcome of many interacting processes (e.g., feeding, growth, allocation of energy to reproduction), the observed toxic effects in a reproduction test could be due to stress on one of many processes. Although this makes reproduction a robust endpoint for detecting stress, it may mask important population-level consequences if the different physiological processes stress affects are associated with different feedback mechanisms at the population level. We therefore evaluated how an observed reduction in reproduction found in a standard reproduction test translates to effects at the population level if it is caused by hypothetical toxicants affecting different physiological processes (physiological modes of action; PMoA). For this we used two consumer-resource models: the Yodzis-Innes (YI) model, which is mathematically tractable, but requires strong assumptions of energetic equivalence among individuals as they progress through ontogeny, and an individual-based implementation of dynamic energy budget theory (DEB-IBM), which relaxes these assumptions at the expense of tractability. We identified two important feedback mechanisms controlling the link between individual- and population-level stress in the YI model. These mechanisms turned out to also be important for interpreting some of the individual-based model results; for two PMoAs, they determined the population response to stress in both models. In contrast, others stress types involved more complex feedbacks, because they asymmetrically stressed the production efficiency of reproduction and somatic growth. The feedbacks associated with different PMoAs drastically altered the link between individual- and population-level effects. For example, hypothetical stressors with different PMoAs that had equal effects on reproduction had effects ranging from a negligible decline in biomass to population extinction. Thus, reproduction tests alone are of little use for extrapolating toxicity to the population level, but we showed that the ecological relevance of standard tests could easily be improved if growth is measured along with reproduction.