Refine
Year of publication
- 2017 (370) (remove)
Document Type
- Article (254)
- Doctoral Thesis (51)
- Postprint (27)
- Other (22)
- Review (12)
- Habilitation Thesis (2)
- Monograph/Edited Volume (1)
- Part of a Book (1)
Is part of the Bibliography
- yes (370)
Keywords
- Arabidopsis (5)
- Arabidopsis thaliana (5)
- ancient DNA (4)
- biogeography (4)
- climate change (4)
- molecularly imprinted polymers (4)
- phylogeny (4)
- synthetic biology (4)
- Actin cytoskeleton (3)
- Apoptosis (3)
Institute
- Institut für Biochemie und Biologie (370) (remove)
The effects of biodiversity on ecosystem functioning generally increase over time, but the underlying processes remain unclear. Using 26 long-term grassland and forest experimental ecosystems, we demonstrate that biodiversity-ecosystem functioning relationships strengthen mainly by greater increases in functioning in high-diversity communities in grasslands and forests. In grasslands, biodiversity effects also strengthen due to decreases in functioning in low-diversity communities. Contrasting trends across grasslands are associated with differences in soil characteristics.
Modeling a secular trend by Monte Carlo simulation of height biased migration in a spatial network
(2017)
Background: In a recent Monte Carlo simulation, the clustering of body height of Swiss military conscripts within a spatial network with characteristic features of the natural Swiss geography was investigated. In this study I examined the effect of migration of tall individuals into network hubs on the dynamics of body height within the whole spatial network. The aim of this study was to simulate height trends. Material and methods: Three networks were used for modeling, a regular rectangular fishing net like network, a real world example based on the geographic map of Switzerland, and a random network. All networks contained between 144 and 148 districts and between 265-307 road connections. Around 100,000 agents were initially released with average height of 170 cm, and height standard deviation of 6.5 cm. The simulation was started with the a priori assumption that height variation within a district is limited and also depends on height of neighboring districts (community effect on height). In addition to a neighborhood influence factor, which simulates a community effect, body height dependent migration of conscripts between adjacent districts in each Monte Carlo simulation was used to re-calculate next generation body heights. In order to determine the direction of migration for taller individuals, various centrality measures for the evaluation of district importance within the spatial network were applied. Taller individuals were favored to migrate more into network hubs, backward migration using the same number of individuals was random, not biased towards body height. Network hubs were defined by the importance of a district within the spatial network. The importance of a district was evaluated by various centrality measures. In the null model there were no road connections, height information could not be delivered between the districts. Results: Due to the favored migration of tall individuals into network hubs, average body height of the hubs, and later, of the whole network increased by up to 0.1 cm per iteration depending on the network model. The general increase in height within the network depended on connectedness and on the amount of height information that was exchanged between neighboring districts. If higher amounts of neighborhood height information were exchanged, the general increase in height within the network was large (strong secular trend). The trend in the homogeneous fishnet like network was lowest, the trend in the random network was highest. Yet, some network properties, such as the heteroscedasticity and autocorrelations of the migration simulation models differed greatly from the natural features observed in Swiss military conscript networks. Autocorrelations of district heights for instance, were much higher in the migration models. Conclusion: This study confirmed that secular height trends can be modeled by preferred migration of tall individuals into network hubs. However, basic network properties of the migration simulation models differed greatly from the natural features observed in Swiss military conscripts. Similar network-based data from other countries should be explored to better investigate height trends with Monte Carlo migration approach.
Behavioral thermoregulation is an important mechanism allowing ectotherms to respond to thermal variations. Its efficiency might become imperative for securing activity budgets under future climate change. For diurnal lizards, thermal microhabitat variability appears to be of high importance, especially in hot deserts where vegetation is highly scattered and sensitive to climatic fluctuations. We investigated the effects of a shading gradient from vegetation on body temperatures and activity timing for two diurnal, terrestrial desert lizards, Ctenotus regius, and Morethia boulengeri, and analyzed their changes under past, present, and future climatic conditions. Both species’ body temperatures and activity timing strongly depended on the shading gradient provided by vegetation heterogeneity. At high temperatures, shaded locations provided cooling temperatures and increased diurnal activity. Conversely, bushes also buffered cold temperature by saving heat. According to future climate change scenarios, cooler microhabitats might become beneficial to warm‐adapted species, such as C. regius, by increasing the duration of daily activity. Contrarily, warmer microhabitats might become unsuitable for less warm‐adapted species such as M. boulengeri for which midsummers might result in a complete restriction of activity irrespective of vegetation. However, total annual activity would still increase provided that individuals would be able to shift their seasonal timing towards spring and autumn. Overall, we highlight the critical importance of thermoregulatory behavior to buffer temperatures and its dependence on vegetation heterogeneity. Whereas studies often neglect ecological processes when anticipating species’ responses to future climate change the strongest impact of a changing climate on terrestrial ectotherms in hot deserts is likely to be the loss of shaded microhabitats rather than the rise in temperature itself. We argue that conservation strategies aiming at addressing future climate changes should focus more on the cascading effects of vegetation rather than on shifts of species distributions predicted solely by climatic envelopes.
This is a cumulative dissertation comprising three original studies (one published, one in revision, one submitted; Effective December 2017) investigating how reptile species in arid Australia respond to various climatic parameters at different spatial scales and analysing the two potential main underlying mechanisms: thermoregulatory behaviour and species interactions. This dissertation combines extensive individual-based field data across trophic levels, selected field experiments, statistical analyses, and predictive modelling techniques. Mechanisms and processes detected in this dissertation can now be used to predict potential future changes in the community of arid-zone lizards. This knowledge will help improving our fundamental understanding of the consequences of global change and thereby prevent biodiversity loss in a vulnerable ecosystem.
The transition from hunting and gathering to farming involved profound cultural and technological changes. In Western and Central Europe, these changes occurred rapidly and synchronously after the arrival of early farmers of Anatolian origin [1-3], who largely replaced the local Mesolithic hunter-gatherers [1, 4-6]. Further east, in the Baltic region, the transition was gradual, with little or no genetic input from incoming farmers [7]. Here we use ancient DNA to investigate the relationship between hunter-gatherers and farmers in the Lower Danube basin, a geographically intermediate area that is characterized by a rapid Neolithic transition but also by the presence of archaeological evidence that points to cultural exchange, and thus possible admixture, between hunter-gatherers and farmers. We recovered four human paleogenomes (1.13 to 4.13 coverage) from Romania spanning a time transect between 8.8 thousand years ago (kya) and 5.4 kya and supplemented them with two Mesolithic genomes (1.73- and 5.33) from Spain to provide further context on the genetic background of Mesolithic Europe. Our results show major Western hunter-gatherer (WHG) ancestry in a Romanian Eneolithic sample with a minor, but sizeable, contribution from Anatolian farmers, suggesting multiple admixture events between hunter-gatherers and farmers. Dietary stableisotope analysis of this sample suggests a mixed terrestrial/ aquatic diet. Our results provide support for complex interactions among hunter-gatherers and farmers in the Danube basin, demonstrating that in some regions, demic and cultural diffusion were not mutually exclusive, but merely the ends of a continuum for the process of Neolithization.
The aim of the study was to investigate the correlation of heights of conscripts living in neighboring districts in Poland. The study used 10% of a nationally representative sample of 26,178 males 18.5-19.5 years old examined during the National survey of Polish conscripts conducted in 2001. The sample represented all regions and social strata of the country and included 354 different districts within 16 voivodships (provinces). Analyses were performed with the R statistical software. A small but significant correlation (0.24, p < 0.0001) was observed for height between 1st order neighboring districts. Correlations decreased with increased distances between neighboring districts, but remained significant for 7th node neighbors (0.18, p < 0.0001). Regarding voivodships (provinces), average height showed a geographical trend from the northwest (relatively tall) to the southeast (relatively short), and the correlation was stronger for first order neighboring provinces (0.796, p < 0.001). This study revealed clusters of tall people and short people, providing a support for hypothesis of the community effect in height. Small correlations between 1st order neighbors than in another country (Switzerland) may be associated with differences in geography, since in Poland there are no natural barriers (e.g., mountains) and road infrastructure is well-developed.
The oomycete Plasmopara obducens was first described on wild Impatiens noli-tangere in Germany in 1877. About 125 years later the first occurrence of P. obducens on cultivated I. walleriana in the United Kingdom was reported, and a worldwide epidemic followed. Although this pathogen is a major threat for ornamental busy lizzy, the identity of the pathogen remained unconfirmed and the high host specificity observed for the genus Plasmopara cast doubts regarding its determination as P. obducens. In this study, using multigene phylogenies and morphological investigation, it is revealed that P. obducens on I. noli-tangere is not the conspecific with the pathogen affecting I. walleriana and another ornamental balsam, I. balsamina. As a consequence, the new names P. destructor and P. velutina are introduced for the pathogens of I. walleriana and I. balsamina, respectively.
The Western honey bee (Apis mellifera) is widely used as commercial pollinator in worldwide agriculture and, therefore, plays an important role in global food security. Among the parasites and pathogens threatening health and survival of honey bees are two species of microsporidia, Nosema apis and Nosema ceranae. Nosema ceranae is considered an emerging pathogen of the Western honey bee. Reports on the spread of N. ceranae suggested that this presumably highly virulent species is replacing its more benign congener N. apis in the global A. mellifera population. We here present a 12 year longitudinal cohort study on the prevalence of N. apis and N. ceranae in Northeast Germany. Between 2005 and 2016, a cohort of about 230 honey bee colonies originating from 23 apiaries was sampled twice a year (spring and autumn) resulting in a total of 5,600 bee samples which were subjected to microscopic and molecular analysis for determining the presence of infections with N. apis or/and N. ceranae. Throughout the entire study period, both N. apis- and N. ceranae-infections could be diagnosed within the cohort. Logistic regression analysis of the prevalence data demonstrated a significant increase of N. ceranae-infections over the last 12 years, both in autumn (reflecting the development during the summer) and in spring (reflecting the development over winter) samples. Cell culture experiments confirmed that N. ceranae has a higher proliferative potential than N. apis at 27. and 33 degrees C potentially explaining the increase in N. ceranae prevalence during summer. In autumn, characterized by generally low infection prevalence, this increase was accompanied by a significant decrease in N. apis- infection prevalence. In contrast, in spring, the season with a higher prevalence of infection, no significant decrease of N. apis infections despite a significant increase in N. ceranae infections could be observed. Therefore, our data do not support a general advantage of N. ceranae over N. apis and an overall replacement of N. apis by N. ceranae in the studied honey bee population.
Many lakes exhibit seasonal stratification, during which they develop strong thermal and chemical gradients. An expansion of depth-integrated monitoring programs has provided insight into the importance of organic carbon processing that occurs below the upper mixed layer. However, the chemical and physical drivers of metabolism and metabolic coupling remain unresolved, especially in the metalimnion. In this depth zone, sharp gradients in key resources such as light and temperature co-occur with dynamic physical conditions that influence metabolic processes directly and simultaneously hamper the accurate tracing of biological activity. We evaluated the drivers of metalimnetic metabolism and its associated uncertainty across 10 stratified lakes in Europe and North America. We hypothesized that the metalimnion would contribute highly to whole-lake functioning in clear oligotrophic lakes, and that metabolic rates would be highly variable in unstable polymictic lakes. Depth-integrated rates of gross primary production (GPP) and ecosystem respiration (ER) were modelled from diel dissolved oxygen curves using a Bayesian approach. Metabolic estimates were more uncertain below the epilimnion, but uncertainty was not consistently related to lake morphology or mixing regime. Metalimnetic rates exhibited high day-to-day variability in all trophic states, with the metalimnetic contribution to daily whole-lake GPP and ER ranging from 0% to 87% and < 1% to 92%, respectively. Nonetheless, the metalimnion of low-nutrient lakes contributed strongly to whole-lake metabolism on average, driven by a collinear combination of highlight, low surface-water phosphorous concentration and high metalimnetic volume. Consequently, a single-sensor approach does not necessarily reflect whole-ecosystem carbon dynamics in stratified lakes.
Extreme weather events can pervasively influence ecosystems. Observations in lakes indicate that severe storms in particular can have pronounced ecosystem-scale consequences, but the underlying mechanisms have not been rigorously assessed in experiments. One major effect of storms on lakes is the redistribution of mineral resources and plankton communities as a result of abrupt thermocline deepening. We aimed at elucidating the importance of this effect by mimicking in replicated large enclosures (each 9 m in diameter, ca. 20 m deep, ca. 1300 m 3 in volume) a mixing event caused by a severe natural storm that was previously observed in a deep clear-water lake. Metabolic rates were derived from diel changes in vertical profiles of dissolved oxygen concentrations using a Bayesian modelling approach, based on high-frequency measurements. Experimental thermocline deepening stimulated daily gross primary production (GPP) in surface waters by an average of 63% for > 4 weeks even though thermal stratification re-established within 5 days. Ecosystem respiration (ER) was tightly coupled to GPP, exceeding that in control enclosures by 53% over the same period. As GPP responded more strongly than ER, net ecosystem productivity (NEP) of the entire water column was also increased. These protracted increases in ecosystem metabolism and autotrophy were driven by a proliferation of inedible filamentous cyanobacteria released from light and nutrient limitation after they were entrained from below the thermocline into the surface water. Thus, thermocline deepening by a single severe storm can induce prolonged responses of lake ecosystem metabolism independent of other storm-induced effects, such as inputs of terrestrial materials by increased catchment run-off. This highlights that future shifts in frequency, severity or timing of storms are an important component of climate change, whose impacts on lake thermal structure will superimpose upon climate trends to influence algal dynamics and organic matter cycling in clear-water lakes. Keywords: climate variability, ecosystem productivity, extreme events, gross primary production, mesocosm, respiration stratified lakes
The appearance of the first leaf from the coleoptile in wheat seedlings (Triticum aestivum L.) coincides with the development of seedling susceptibility to water deficiency on the fifth day following imbibition. In dehydrated wheat seedlings, an increase in the protein carbonyl group has been observed. The coincidence of higher protein carbonylation levels with development of dehydration intolerance drew our attention. To gain more insight into the molecular basis of wheat drought tolerance, the seedling profiles of carbonylated proteins were analysed and compared. Two-dimensional gel electrophoresis (2D-PAGE) and mass spectrometry (MALDI-TOF and LC-MS/MS) were used to indicate and identify differential carbonylated proteins. Among the protein spots with at least a two-fold change in protein abundance in dehydrated seedlings in relation to control (well-watered) plants during the tolerant phase of growth, 19 carbonylated proteins increased and 18 carbonylated proteins decreased in abundance. Among 26 differentially expressed carbonylated proteins in sensitive seedlings, the abundance of 10 protein spots increased while that of 16 proteins decreased upon dehydration. We have demonstrated a link between protein carbonylation and seedling sensitivity to dehydration. The analysis of carbonylated protein profiles clearly showed that proteins with a potential role in the maintenance of dehydration tolerance in wheat seedlings are mainly linked to energy production, anti-fungal and/or insecticidal activity, or to the regulation of both protein synthesis and degradation.
Protected areas are arguably the most important instrument of biodiversity conservation. To keep them fit under climate change, their management needs to be adapted to address related direct and indirect changes. In our study we focus on the adaptation of conservation management planning, evaluating management plans of 60 protected areas throughout Germany with regard to their climate change-robustness. First, climate change-robust conservation management was defined using 11 principles and 44 criteria, which followed an approach similar to sustainability standards. We then evaluated the performance of individual management plans concerning the climate change-robustness framework. We found that climate change-robustness of protected areas hardly exceeded 50 percent of the potential performance, with most plans ranking in the lower quarter. Most Natura 2000 protected areas, established under conservation legislation of the European Union, belong to the sites with especially poor performance, with lower values in smaller areas. In general, the individual principles showed very different rates of accordance with our principles, but similarly low intensity. Principles with generally higher performance values included holistic knowledge management, public accountability and acceptance as well as systemic and strategic coherence. Deficiencies were connected to dealing with the future and uncertainty. Lastly, we recommended the presented principles and criteria as essential guideposts that can be used as a checklist for working towards more climate change-robust planning.
Cellular membranes constantly experience remodeling, as exemplified by morphological changes during endo- and exocytosis. Regulation of membrane morphology is essential for these processes. In this work, we attempt to establish a regulation path based on the use of photoswitches exhibiting conformational changes in model membranes, namely, giant unilamellar vesicles (GUVs). The mechanism of the changes in the GUVs’ morphology caused by isomerization of the photosensitive molecules has been previously explored but still remains elusive. We examine the morphological reshaping of GUVs in the presence of the photoswitch o-tetrafluoroazobenzene (F-azo) and show that the mechanism behind the resulting morphological changes involves both an increase in the membrane area and generation of a positive spontaneous curvature. First, we characterize the partitioning of F-azo in a single-component membrane using both experimental and computational approaches. The partition coefficient calculated from molecular dynamic simulations agrees with experimental data obtained with size-exclusion chromatography. Then, we implement the approach of vesicle electrodeformation in order to assess the increase in the membrane area, which is observed as a result of the conformational change of F-azo. Finally, the local and the effective membrane spontaneous curvatures were estimated from the observed shapes of vesicles exhibiting outward budding. We then extend the application of the F-azo to multicomponent lipid membranes, which exhibit a coexistence of domains in different liquid phases due to a miscibility gap between the lipids. We perform initial experiments to investigate whether F-azo can be employed to modulate the lateral lipid packing and organization. We observe either complete mixing of the domains or the appearing of disordered domains within the domains of more ordered phase. The type of behavior observed in response to the photoisomerization of F-azo was dependent on the used lipid composition. We believe that the findings introduced here will have an impact in understanding and controlling both lipid phase modulation and regulation of the membrane morphology in membrane systems.
Inactivation of thermogenic UCP1 as a historical contingency in multiple placental mammal clades
(2017)
Comparison of the dissociation kinetics of rapid-acting insulins lispro, aspart, glulisine and human insulin under physiologically relevant conditions. Dissociation kinetics after dilution were monitored directly in terms of the average molecular mass using combined static and dynamic light scattering. Changes in tertiary structure were detected by near-UV circular dichroism. Glulisine forms compact hexamers in formulation even in the absence of Zn2+. Upon severe dilution, these rapidly dissociate into monomers in less than 10 s. In contrast, in formulations of lispro and aspart, the presence of Zn2+ and phenolic compounds is essential for formation of compact R6 hexamers. These slowly dissociate in times ranging from seconds to one hour depending on the concentration of phenolic additives. The disadvantage of the long dissociation times of lispro and aspart can be diminished by a rapid depletion of the concentration of phenolic additives independent of the insulin dilution. This is especially important in conditions similar to those after subcutaneous injection, where only minor dilution of the insulins occurs. Knowledge of the diverging dissociation mechanisms of lispro and aspart compared to glulisine will be helpful for optimizing formulation conditions of rapid-acting insulins.
The phylogenetic structure of communities (PSC) reveals how evolutionary history affects community assembly processes. However, there are important knowledge gaps on PSC patterns for annual communities and there is a need for studies along environmental gradients in dry ecosystems where several processes shape PSC. Here, we investigated the PSC of annual plants along an aridity gradient in Israel, including eight years, two spatial scales, the effects of shrubs on understory, and the phylogenetic signal of important traits. Increasing drought stress led to overdispersed PSC at the drier end of the gradient, indicating that species were less related than expected by chance. This was supported at a smaller spatial scale, where within the drier sites, communities in open- more arid- habitats were more overdispersed than those under nurse shrubs. Interestingly, some key traits related to drought resistance were not conserved in the phylogeny. Together, our findings suggested that while habitat filtering selected for drought resistance strategies, these strategies evolved independently along multiple contrasting evolutionary lineages. Our comprehensive PSC study provides strong evidence for the interacting effects of habitat filtering and plant- plant interactions, particularly highlighting that the conservative evolution of traits should not be assumed in future interpretations of PSC patterns.
Planktotrons
(2017)
We established a new indoor mesocosm facility, 12 fully controlled Planktotrons, designed to conduct marine and freshwater experiments for biodiversity and food web approaches using natural or artificial, benthic or planktonic communities. The Planktotrons are a unique and custom-tailored facility allowing long-term experiments. Wall growth can be inhibited by a rotating gate paddle with silicone lips. Additionally, temperature and light intensity are individually controllable for each Planktotron and the large volume (600 L) enables high-frequency or volume-intense measurements. In a pilot freshwater experiment various trophic levels of a pelagic food web were maintained for up to 90 d. First, an artificially assembled phytoplankton community of 11 species was inoculated in all Planktotrons. After 22 d, two ciliates were added to all, and three Daphnia species were added to six Planktotrons. After 72 d, dissolved organic matter (DOM, an alkaline soil extract) was added as an external disturbance to six of the 12 Planktotrons, involving three Planktotrons stocked with Daphnia and three without, respectively. We demonstrate the suitability of the Planktotrons for food web and biodiversity research. Variation among replicated Planktotrons (n=3 minimum) did not differ from other laboratory systems and field experiments. We investigated population dynamics and interactions among the different trophic levels, and found them affected by the sequence of ciliate and Daphnia addition and the disturbance caused by addition of DOM.
Immunodeficient mice are crucial models to evaluate the efficacy of monoclonal antibodies (mAbs). When studying mAb pharmacokinetics (PK), protection from elimination by binding to the neonatal Fc receptor (FcRn) is known to be a major process influencing the unspecific clearance of endogenous and therapeutic IgG. The concentration of endogenous IgG in immunodeficient mice, however is reduced, and this effect on the FcRn protection mechanism and subsequently on unspecific mAb clearance is unknown, yet of great importance for the interpretation of mAb PK data. We used a PBPK modelling approach to elucidate the influence of altered endogenous IgG concentrations on unspecific mAb clearance. To this end, we used PK data in immunodeficient mice, i.e. nude and severe combined immunodeficiency mice. To avoid impact of target-mediated clearance processes, we focussed on mAbs without affinity to a target antigen in these mice. In addition, intravenous immunoglobulin (IVIG) data of immunocompetent mice was used to study the impact of increased total IgG concentrations on unspecific therapeutic antibody clearance. The unspecific clearance is linear, whenever therapeutic IgG concentrations, i.e. mAb and IVIG concentrations are lower than FcRn; it can be non-linear if therapeutic IgG concentrations are larger than FcRn and endogenous IgG concentrations (e.g., under IVIG therapy). Unspecific mAb clearance of immunodeficient mice is effectively linear (under mAb doses as typically used in human). Studying the impact of reduced endogenous IgG concentrations on unspecific mAb clearance is of great relevance for the extrapolation to clinical species, e.g., when predicting mAb PK in immunosuppressed cancer patients.
Assembly of iron sulfur (FeS) clusters is an important process in living cells. The initial sulfur mobilization step for FeS cluster biosynthesis is catalyzed by L-cysteine desulfurase NFS1, a reaction that is localized in mitochondria in humans. In humans, the function of NFS1 depends on the ISD11 protein, which is required to stabilize its structure. The NFS1/ISD11 complex further interacts with scaffold protein ISCU and regulator protein frataxin, thereby forming a quaternary complex for FeS cluster formation. It has been suggested that the role of ISD11 is not restricted to its role in stabilizing the structure of NFS1, because studies of single-amino acid variants of ISD11 additionally demonstrated its importance for the correct assembly of the quaternary complex. In this study, we are focusing on the N-terminal region of ISD11 to determine the role of N-terminal amino acids in the formation of the complex with NFS1 and to reveal the mitochondria) targeting sequence for subcellular localization. Our in vitro studies with the purified proteins and in vivo studies in a cellular system show that the first 10 N-terminal amino acids of ISD11 are indispensable for the activity of NFS1 and especially the conserved "LYR" motif is essential for the role of ISD11 in forming a stable and active complex with NFS1.
Chytridiomycota, often referred to as chytrids, can be virulent parasites with the potential to inflict mass mortalities on hosts, causing e.g. changes in phytoplankton size distributions and succession, and the delay or suppression of bloom events. Molecular environmental surveys have revealed an unexpectedly large diversity of chytrids across a wide range of aquatic ecosystems worldwide. As a result, scientific interest towards fungal parasites of phytoplankton has been gaining momentum in the past few years. Yet, we still know little about the ecology of chytrids, their life cycles, phylogeny, host specificity and range. Information on the contribution of chytrids to trophic interactions, as well as co-evolutionary feedbacks of fungal parasitism on host populations is also limited. This paper synthesizes ideas stressing the multifaceted biological relevance of phytoplankton chytridiomycosis, resulting from discussions among an international team of chytrid researchers. It presents our view on the most pressing research needs for promoting the integration of chytrid fungi into aquatic ecology.
Aldehyde oxidases (AOXs) are molybdo-flavoenzymes characterized by broad substrate specificity, oxidizing aromatic/aliphatic aldehydes into the corresponding carboxylic acids and hydroxylating various heteroaromatic rings. The enzymes use oxygen as the terminal electron acceptor and produce reduced oxygen species during turnover. The physiological function of mammalian AOX isoenzymes is still unclear, however, human AOX (hAOX1) is an emerging enzyme in phase-I drug metabolism. Indeed, the number of xenobiotics acting as hAOX1 substrates is increasing. Further, numerous single-nucleotide polymorphisms (SNPs) have been identified within the hAOX1 gene. SNPs are a major source of inter-individual variability in the human population, and SNP-based amino acid exchanges in hAOX1 reportedly modulate the catalytic function of the enzyme in either a positive or negative fashion. In this report we selected ten novel SNPs resulting in amino acid exchanges in proximity to the FAD site of hAOX1 and characterized the purified enzymes after heterologous expression in Escherichia coli. The hAOX1 variants were characterized carefully by quantitative differences in their ability to produce superoxide radical. ROS represent prominent key molecules in physiological and pathological conditions in the cell. Our data reveal significant alterations in superoxide anion production among the variants. In particular the SNP-based amino acid exchange L438V in proximity to the isoalloxanzine ring of the FAD cofactor resulted in increased rate of superoxide radical production of 75%. Considering the high toxicity of the superoxide in the cell, the hAOX1-L438V SNP variant is an eventual candidate for critical or pathological roles of this natural variant within the human population.
In this work the human AOX1 was characterized and detailed aspects regarding the expression, the enzyme kinetics and the production of reactive oxygen species (ROS) were investigated. The hAOX1 is a cytosolic enzyme belonging to the molybdenum hydroxylase family. Its catalytically active form is a homodimer with a molecular weight of 300 kDa. Each monomer (150 kDa) consists of three domains: a N-terminal domain (20 kDa) containing two [2Fe-2S] clusters, a 40 kDa intermediate domain containing a flavin adenine dinucleotide (FAD), and a C-terminal domain (85 kDa) containing the substrate binding pocket and the molybdenum cofactor (Moco). The hAOX1 has an emerging role in the metabolism and pharmacokinetics of many drugs, especially aldehydes and N- heterocyclic compounds.
In this study, the hAOX1 was hetereogously expressed in E. coli TP1000 cells, using a new codon optimized gene sequence which improved the expressed protein yield of around 10-fold compared to the previous expression systems for this enzyme. To increase the catalytic activity of hAOX1, an in vitro chemical sulfuration was performed to favor the insertion of the equatorial sulfido ligand at the Moco with consequent increased enzymatic activity of around 10-fold. Steady-state kinetics and inhibition studies were performed using several substrates, electron acceptors and inhibitors. The recombinant hAOX1 showed higher catalytic activity when molecular oxygen was used as electron acceptor. The highest turn over values were obtained with phenanthridine as substrate. Inhibition studies using thioridazine (phenothiazine family), in combination with structural studies performed in the group of Prof. M.J. Romão, Nova Universidade de Lisboa, showed a new inhibition site located in proximity of the dimerization site of hAOX1. The inhibition mode of thioridazine resulted in a noncompetitive inhibition type. Further inhibition studies with loxapine, a thioridazine-related molecule, showed the same type of inhibition. Additional inhibition studies using DCPIP and raloxifene were carried out.
Extensive studies on the FAD active site of the hAOX1 were performed. Twenty new hAOX1 variants were produced and characterized. The hAOX1 variants generated in this work were divided in three groups: I) hAOX1 single nucleotide polymorphisms (SNP) variants; II) XOR- FAD loop hAOX1 variants; III) additional single point hAOX1 variants. The hAOX1 SNP variants G46E, G50D, G346R, R433P, A439E, K1231N showed clear alterations in their catalytic activity, indicating a crucial role of these residues into the FAD active site and in relation to the overall reactivity of hAOX1.
Furthermore, residues of the bovine XOR FAD flexible loop (Q423ASRREDDIAK433) were introduced in the hAOX1. FAD loop hAOX1 variants were produced and characterized for their stability and catalytic activity. Especially the variants hAOX1 N436D/A437D/L438I, N436D/A437D/L438I/I440K and Q434R/N436D/A437D/L438I/I440K showed decreased catalytic activity and stability. hAOX1 wild type and variants were tested for reactivity toward NADH but no reaction was observed.
Additionally, the hAOX1 wild type and variants were tested for the generation of reactive oxygen species (ROS). Interestingly, one of the SNP variants, hAOX1 L438V, showed a high ratio of superoxide prodction. This result showed a critical role for the residue Leu438 in the mechanism of oxygen radicals formation by hAOX1. Subsequently, further hAOX1 variants having the mutated Leu438 residue were produced. The variants hAOX1 L438A, L438F and L438K showed superoxide overproduction of around 85%, 65% and 35% of the total reducing equivalent obtained from the substrate oxidation.
The results of this work show for the first time a characterization of the FAD active site of the hAOX1, revealing the importance of specific residues involved in the generation of ROS and effecting the overall enzymatic activity of hAOX1. The hAOX1 SNP variants presented here indicate that those allelic variations in humans might cause alterations ROS balancing and clearance of drugs in humans.
We identified Alizarin Red S and other well known fluorescent dyes useful for the online detection of pyrophosphate in enzymatic assays, including the loop mediated isothermal amplification (LAMP) and polymerase chain reaction (PCR) assays. An iterative screening was used for a selected set of compounds to first secure enzyme compatibility, evaluate inorganic pyrophosphate sensitivity in the presence of manganese as quencher and optimize conditions for an online detection. Of the selected dyes, the inexpensive alizarin red S was found to selectively detect pyrophosphate under LAMP and PCR conditions and is superior with respect to its defined red-shifted spectrum, long shelf life and low toxicity. In addition, the newly identified properties may also be useful in other enzymatic assays which do not generate nucleic acids but are based on inorganic pyrophosphate. Finally, we propose that our screening method may provide a blueprint for rapid screening of compounds for detecting inorganic pyrophosphate.
Hintergrund: Etablierte Protein- und Nukleinsäure-basierte Methoden für den spezifischen Pathogennachweis sind nur unter standardisierten Laborbedingungen von geschultem Personal durchführbar und daher mit einem hohen Zeit- und Kostenaufwand verbunden. In der Nukleinsäure-basierten Diagnostik kann durch die Einführung der isothermalen Amplifikation eine schnelle und kostengünstige Alternative zur Polymerase-Kettenreaktion (PCR) verwendet werden. Die Loop-mediated isothermal amplification (LAMP) bietet aufgrund der hohen Amplifikationseffizienz vielfältige Detektionsmöglichkeiten, die sowohl für Schnelltest- als auch für Monitoring-Anwendungen geeignet sind.
Ein wesentliches Ziel dieser Arbeit war die Verbesserung der Anwendbarkeit der LAMP und die Entwicklung einer neuen Methode für den einfachen, schnellen und günstigen Nachweis von Pathogenen mittels alternativer DNA- oder Pyrophosphat-abhängiger Detektionsverfahren. Hier wurden zunächst direkte und indirekte Detektionsmethoden untersucht und darauf aufbauend ein Verfahren entwickelt, mit dem neue Metallionen-abhängige Fluoreszenzfarbstoffe für die selektive Detektion von Pyrophosphat in der LAMP und anderen enzymatischen Reaktionen identifiziert werden können. Als Alternative für die DNA-basierte Detektion in der digitalen LAMP sollten die zuvor etablierten Farbstoffe für den Pyrophosphatnachweis in einer Emulsion getestet werden. Abschließend wurde ein neuer Reaktionsmechanismus für die effiziente Generierung hochmolekularer DNA unter isothermalen Bedingungen als Alternative zur LAMP entwickelt.
Ergebnisse: Für den Nachweis RNA- und DNA-basierter Phythopathogene konnte die Echtzeit- und Endpunktdetektion mit verschiedenen Farbstoffen in einem geschlossenen System etabliert werden. Hier wurde Berberin als DNA-interkalierender Fluoreszenzfarbstoff mit vergleichbarer Sensitivität zu SYBR Green und EvaGreen erfolgreich in der LAMP mit Echtzeitdetektion eingesetzt. Ein Vorteil von Berberin gegenüber den anderen Farbstoffen ist die Toleranz der DNA-Polymerase auch bei hohen Farbstoffkonzentrationen. Berberin kann daher auch in der geschlossenen LAMP-Reaktion ohne zusätzliche Anpassung der Reaktionsbedingungen für die Endpunktdetektion verwendet werden. Darüber hinaus konnte Hydroxynaphtholblau (HNB), das für den kolorimetrischen Endpunktnachweis bekannt ist, erstmals auch für die fluorimetrische Detektion der LAMP in Echtzeit eingesetzt werden. Zusätzlich konnten in der Arbeit weitere Metallionen-abhängige Farbstoffe zur indirekten Detektion der LAMP über das Pyrophosphat identifiziert werden. Dafür wurde eine iterative Methode entwickelt, mit der potenzielle Farbstoffe hinsichtlich ihrer Enzymkompatibilität und ihrer spektralen Eigenschaften bei An- oder Abwesenheit von Manganionen selektiert werden können. Mithilfe eines kombinatorischen Screenings im Mikrotiterplattenformat konnte die komplexe Konzentrationsabhängigkeit zwischen den einzelnen Komponenten für einen fluorimetrischen Verdrängungsnachweis untersucht werden. Durch die Visualisierung des Signal-Rausch-Verhältnis’ als Intensitätsmatrix (heatmap) konnten zunächst Alizarinrot S und Tetrazyklin unter simulierten Reaktionsbedingungen selektiert werden. In der anschließenden enzymatischen LAMP-Reaktion konnte insbesondere Alizarinrot S als günstiger, nicht-toxischer und robuster Fluoreszenzfarbstoff identifiziert werden und zeigte eine Pyrophosphat-abhängige Zunahme der Fluoreszenzintensität. Die zuvor etablierten Farbstoffe (HNB, Calcein und Alizarinrot S) konnten anschließend erfolgreich für die indirekte, fluorimetrische Detektion von Pyrophosphat in einer LAMP-optimierten Emulsion eingesetzt werden. Die Stabilität und Homogenität der generierten Emulsion wurde durch den Zusatz des Emulgators Poloxamer 188 verbessert. Durch die fluoreszenzmikroskopische Analyse der Emulsion war eine eindeutige Diskriminierung der positiven und negativen Tröpfchen vor allem bei Einsatz von Calcein und Alizarinrot S möglich. Aufgrund des komplexen Primer-Designs und der hohen Wahrscheinlichkeit unspezifischer Amplifikation in der LAMP wurde eine neue Bst DNA-Polymerase-abhängige isothermale Amplifikationsreaktion entwickelt. Durch die Integration einer spezifischen Linkerstruktur (abasische Stelle oder Hexaethylenglykol) zwischen zwei Primersequenzen konnte ein bifunktioneller Primer die effiziente Regenerierung der Primerbindungsstellen gewährleisten. Der neue Primer induziert nach der spezifischen Hybridisierung auf dem Templat die Rückfaltung zu einer Haarnadelstruktur und blockiert gleichzeitig die Polymeraseaktivität am Gegenstrang, wodurch eine autozyklische Amplifikation trotz konstanter Reaktionstemperatur möglich ist. Die Effizienz der „Hinge-initiated Primer dependent Amplification“ (HIP) konnte abschließend durch die Verkürzung der Distanz zwischen einem modifizierten Hinge-Primer und einem PCR-ähnlichen Primer verbessert werden.
Schlussfolgerung: Die LAMP hat sich aufgrund der hohen Robustheit und Effizienz zu einer leistungsfähigen Alternative für die klassische PCR in der molekularbiologischen Diagnostik entwickelt. Unterschiedliche Detektionsverfahren verbessern die Leistungsfähigkeit der qualitativen und quantitativen LAMP für die Feldanwendungen und für die Diagnostik, da die neuen DNA- und Pyrophosphat-abhängigen Nachweismethoden in einer geschlossenen Reaktion eingesetzt werden können und so eine einfache Pathogendiagnostik ermöglichen. Die gezeigten Methoden können darüber hinaus zu einer Kostensenkung und Zeitersparnis gegenüber den herkömmlichen Methoden beitragen. Ein attraktives Ziel stellt die Weiterentwicklung der HIP für den Pathogennachweis als Alternative zur LAMP dar. Hierbei können die neuen LAMP-Detektionsverfahren ebenfalls Anwendung finden. Die Verwendung von Bst DNA-Polymerase-abhängigen Reaktionen ermöglicht darüber hinaus die Integration einer robusten isothermalen Amplifikation in mikrofluidische Systeme. Durch die Kombination der Probenvorbereitung, Amplifikation und Detektion sind zukünftige Anwendungen mit kurzer Analysezeit und geringem apparativen Aufwand insbesondere in der Pathogendiagnostik möglich.
Aerobic anoxygenic phototrophs (AAPs) have been shown to exist in numerous marine and brackish environments where they are hypothesized to play important ecological roles. Despite their potential significance, the study of freshwater AAPs is in its infancy and limited to local investigations. Here, we explore the occurrence, diversity and distribution of AAPs in lakes covering a wide latitudinal gradient: Mongolian and German lakes located in temperate regions of Eurasia, tropical Great East African lakes, and polar permanently ice-covered Antarctic lakes. Our results show a widespread distribution of AAPs in lakes with contrasting environmental conditions and confirm that this group is composed of different members of the Alpha- and Betaproteobacteria. While latitude does not seem to strongly influence AAP abundance, clear patterns of community structure and composition along geographic regions were observed as indicated by a strong macro-geographical signal in the taxonomical composition of AAPs. Overall, our results suggest that the distribution patterns of freshwater AAPs are likely driven by a combination of small-scale environmental conditions (specific of each lake and region) and large-scale geographic factors (climatic regions across a latitudinal gradient).
Engineering artificial networks from modular components is a major challenge in synthetic biology. In the past years, single units, such as switches and oscillators, were successfully constructed and implemented. The effective integration of these parts into functional artificial self-regulated networks is currently on the verge of breakthrough. Here, we describe the design of a modular higher-order synthetic genetic network assembled from two independent self-sustained synthetic units: repressilators coupled via a modified quorum-sensing circuit. The isolated communication circuit and the network of coupled oscillators were analysed in mathematical modelling and experimental approaches. We monitored clustering of cells in groups of various sizes. Within each cluster of cells, cells oscillate synchronously, whereas the theoretical modelling predicts complete synchronization of the whole cellular population to be obtained approximately after 30 days. Our data suggest that self-regulated synchronization in biological systems can occur through an intermediate, long term clustering phase. The proposed artificial multicellular network provides a system framework for exploring how a given network generates a specific behaviour.
The El Nino-Southern Oscillation (ENSO) is the main driver of the interannual variability in eastern African rainfall, with a significant impact on vegetation and agriculture and dire consequences for food and social security. In this study, we identify and quantify the ENSO contribution to the eastern African rainfall variability to forecast future eastern African vegetation response to rainfall variability related to a predicted intensified ENSO. To differentiate the vegetation variability due to ENSO, we removed the ENSO signal from the climate data using empirical orthogonal teleconnection (EOT) analysis. Then, we simulated the ecosystem carbon and water fluxes under the historical climate without components related to ENSO teleconnections. We found ENSO-driven patterns in vegetation response and confirmed that EOT analysis can successfully produce coupled tropical Pacific sea surface temperature-eastern African rainfall teleconnection from observed datasets. We further simulated eastern African vegetation response under future climate change as it is projected by climate models and under future climate change combined with a predicted increased ENSO intensity. Our EOT analysis highlights that climate simulations are still not good at capturing rainfall variability due to ENSO, and as we show here the future vegetation would be different from what is simulated under these climate model outputs lacking accurate ENSO contribution. We simulated considerable differences in eastern African vegetation growth under the influence of an intensified ENSO regime which will bring further environmental stress to a region with a reduced capacity to adapt effects of global climate change and food security.
The El Nino-Southern Oscillation (ENSO) is the main driver of the interannual variability in eastern African rainfall, with a significant impact on vegetation and agriculture and dire consequences for food and social security. In this study, we identify and quantify the ENSO contribution to the eastern African rainfall variability to forecast future eastern African vegetation response to rainfall variability related to a predicted intensified ENSO. To differentiate the vegetation variability due to ENSO, we removed the ENSO signal from the climate data using empirical orthogonal teleconnection (EOT) analysis. Then, we simulated the ecosystem carbon and water fluxes under the historical climate without components related to ENSO teleconnections. We found ENSO-driven patterns in vegetation response and confirmed that EOT analysis can successfully produce coupled tropical Pacific sea surface temperature-eastern African rainfall teleconnection from observed datasets. We further simulated eastern African vegetation response under future climate change as it is projected by climate models and under future climate change combined with a predicted increased ENSO intensity. Our EOT analysis highlights that climate simulations are still not good at capturing rainfall variability due to ENSO, and as we show here the future vegetation would be different from what is simulated under these climate model outputs lacking accurate ENSO contribution. We simulated considerable differences in eastern African vegetation growth under the influence of an intensified ENSO regime which will bring further environmental stress to a region with a reduced capacity to adapt effects of global climate change and food security.
The El Nino-Southern Oscillation (ENSO) is the main driver of the interannual variability in eastern African rainfall, with a significant impact on vegetation and agriculture and dire consequences for food and social security. In this study, we identify and quantify the ENSO contribution to the eastern African rainfall variability to forecast future eastern African vegetation response to rainfall variability related to a predicted intensified ENSO. To differentiate the vegetation variability due to ENSO, we removed the ENSO signal from the climate data using empirical orthogonal teleconnection (EOT) analysis. Then, we simulated the ecosystem carbon and water fluxes under the historical climate without components related to ENSO teleconnections. We found ENSO-driven patterns in vegetation response and confirmed that EOT analysis can successfully produce coupled tropical Pacific sea surface temperature-eastern African rainfall teleconnection from observed datasets. We further simulated eastern African vegetation response under future climate change as it is projected by climate models and under future climate change combined with a predicted increased ENSO intensity. Our EOT analysis highlights that climate simulations are still not good at capturing rainfall variability due to ENSO, and as we show here the future vegetation would be different from what is simulated under these climate model outputs lacking accurate ENSO contribution. We simulated considerable differences in eastern African vegetation growth under the influence of an intensified ENSO regime which will bring further environmental stress to a region with a reduced capacity to adapt effects of global climate change and food security.
The profits of excludability and transferability in redistributive land reform in central Namibia
(2017)
Policies which redistribute property rights to land can improve the well-being of rural households and can have overall growth effects. In many cases, however, land reforms are driven mainly by politically justified objectives. Under such circumstances, little emphasis is placed on whether and, if so, how property rights can increase productivity. Following 18 years of land reform implementation in Namibia, we evaluated 65 beneficiaries in Namibia. We assess to which degree land rights affects their farm income. The study focuses on Namibia’s two main commercial land reform instruments, namely the Farm Unit Resettlement Scheme and the Affirmative Action Loan Scheme. We find evidence that the majority of land reform projects are not profitable. Further, our study confirms the importance of the right to restrict land access compared with the right to transfer. The long-term leasehold contract seemingly provides sufficient incentives to make productive use of the land.
The Influence of Land Use Intensity on the Plant-Associated Microbiome of Dactylis glomerata L.
(2017)
In this study, we investigated the impact of different land use intensities (LUI) on the root-associated microbiome of Dactylis glomerata (orchardgrass). For this purpose, eight sampling sites with different land use intensity levels but comparable soil properties were selected in the southwest of Germany. Experimental plots covered land use levels from natural grassland up to intensively managed meadows. We used 16S rRNA gene based barcoding to assess the plant-associated community structure in the endosphere, rhizosphere and bulk soil of D. glomerata. Samples were taken at the reproductive stage of the plant in early summer. Our data indicated that roots harbor a distinct bacterial community, which clearly differed from the microbiome of the rhizosphere and bulk soil. Our results revealed Pseudomonadaceae, Enterobacteriaceae and Comamonadaceae as the most abundant endophytes independently of land use intensity. Rhizosphere and bulk soil were dominated also by Proteobacteria, but the most abundant families differed from those obtained from root samples. In the soil, the effect of land use intensity was more pronounced compared to root endophytes leading to a clearly distinct pattern of bacterial communities under different LUI from rhizosphere and bulk soil vs. endophytes. Overall, a change of community structure on the plant-soil interface was observed, as the number of shared OTUs between all three compartments investigated increased with decreasing land use intensity. Thus, our findings suggest a stronger interaction of the plant with its surrounding soil under low land use intensity. Furthermore, the amount and quality of available nitrogen was identified as a major driver for shifts in the microbiome structure in all compartments.
Sequelae of prematurity triggered by oxidative stress and free radical-mediated tissue damage have coined the term “oxygen radical disease of prematurity”. Caffeine, a potent free radical scavenger and adenosine receptor antagonist, reduces rates of brain damage in preterm infants. In the present study, we investigated the effects of caffeine on oxidative stress markers, anti-oxidative response, inflammation, redox-sensitive transcription factors, apoptosis, and extracellular matrix following the induction of hyperoxia in neonatal rats. The brain of a rat pups at postnatal Day 6 (P6) corresponds to that of a human fetal brain at 28–32 weeks gestation and the neonatal rat is an ideal model in which to investigate effects of oxidative stress and neuroprotection of caffeine on the developing brain. Six-day-old Wistar rats were pre-treated with caffeine and exposed to 80% oxygen for 24 and 48 h. Caffeine reduced oxidative stress marker (heme oxygenase-1, lipid peroxidation, hydrogen peroxide, and glutamate-cysteine ligase catalytic subunit (GCLC)), promoted anti-oxidative response (superoxide dismutase, peroxiredoxin 1, and sulfiredoxin 1), down-regulated pro-inflammatory cytokines, modulated redox-sensitive transcription factor expression (Nrf2/Keap1, and NFκB), reduced pro-apoptotic effectors (poly (ADP-ribose) polymerase-1 (PARP-1), apoptosis inducing factor (AIF), and caspase-3), and diminished extracellular matrix degeneration (matrix metalloproteinases (MMP) 2, and inhibitor of metalloproteinase (TIMP) 1/2). Our study affirms that caffeine is a pleiotropic neuroprotective drug in the developing brain due to its anti-oxidant, anti-inflammatory, and anti-apoptotic properties.
Sequelae of prematurity triggered by oxidative stress and free radical-mediated tissue damage have coined the term "oxygen radical disease of prematurity". Caffeine, a potent free radical scavenger and adenosine receptor antagonist, reduces rates of brain damage in preterm infants. In the present study, we investigated the effects of caffeine on oxidative stress markers, anti-oxidative response, inflammation, redox-sensitive transcription factors, apoptosis, and extracellular matrix following the induction of hyperoxia in neonatal rats. The brain of a rat pups at postnatal Day 6 (P6) corresponds to that of a human fetal brain at 28-32 weeks gestation and the neonatal rat is an ideal model in which to investigate effects of oxidative stress and neuroprotection of caffeine on the developing brain. Six-day-old Wistar rats were pre-treated with caffeine and exposed to 80% oxygen for 24 and 48 h. Caffeine reduced oxidative stress marker (heme oxygenase-1, lipid peroxidation, hydrogen peroxide, and glutamate-cysteine ligase catalytic subunit (GCLC)), promoted anti-oxidative response (superoxide dismutase, peroxiredoxin 1, and sulfiredoxin 1), down-regulated pro-inflammatory cytokines, modulated redox-sensitive transcription factor expression (Nrf2/Keap1, and NF kappa B), reduced pro-apoptotic effectors (poly (ADP-ribose) polymerase-1 (PARP-1), apoptosis inducing factor (AIF), and caspase-3), and diminished extracellular matrix degeneration (matrix metalloproteinases (MMP) 2, and inhibitor of metalloproteinase (TIMP) 1/2). Our study affirms that caffeine is a pleiotropic neuroprotective drug in the developing brain due to its anti-oxidant, anti-inflammatory, and anti-apoptotic properties.
Trade-offs between functional traits are ubiquitous in nature and can promote species coexistence depending on their shape. Classic theory predicts that convex trade-offs facilitate coexistence of specialized species with extreme trait values (extreme species) while concave trade-offs promote species with intermediate trait values (intermediate species). We show here that this prediction becomes insufficient when the traits translate non-linearly into fitness which frequently occurs in nature, e.g., an increasing length of spines reduces grazing losses only up to a certain threshold resulting in a saturating or sigmoid trait-fitness function. We present a novel, general approach to evaluate the effect of different trade-off shapes on species coexistence. We compare the trade-off curve to the invasion boundary of an intermediate species invading the two extreme species. At this boundary, the invasion fitness is zero. Thus, it separates trait combinations where invasion is or is not possible. The invasion boundary is calculated based on measurable trait-fitness relationships. If at least one of these relationships is not linear, the invasion boundary becomes non-linear, implying that convex and concave trade-offs not necessarily lead to different coexistence patterns. Therefore, we suggest a new ecological classification of trade-offs into extreme-favoring and intermediate-favoring which differs from a purely mathematical description of their shape. We apply our approach to a well-established model of an empirical predator-prey system with competing prey types facing a trade-off between edibility and half-saturation constant for nutrient uptake. We show that the survival of the intermediate prey depends on the convexity of the trade-off. Overall, our approach provides a general tool to make a priori predictions on the outcome of competition among species facing a common trade-off in dependence of the shape of the trade-off and the shape of the trait-fitness relationships.
Background: Severe bacterial infections remain a major challenge in intensive care units because of their high prevalence and mortality. Adequate antibiotic exposure has been associated with clinical success in critically ill patients. The objective of this study was to investigate the target attainment of standard meropenem dosing in a heterogeneous critically ill population, to quantify the impact of the full renal function spectrum on meropenem exposure and target attainment, and ultimately to translate the findings into a tool for practical application. Methods: A prospective observational single-centre study was performed with critically ill patients with severe infections receiving standard dosing of meropenem. Serial blood samples were drawn over 4 study days to determine meropenem serum concentrations. Renal function was assessed by creatinine clearance according to the Cockcroft and Gault equation (CLCRCG). Variability in meropenem serum concentrations was quantified at the middle and end of each monitored dosing interval. The attainment of two pharmacokinetic/pharmacodynamic targets (100% T->MIC, 50% T->4xMIC) was evaluated for minimum inhibitory concentration (MIC) values of 2 mg/L and 8 mg/L and standard meropenem dosing (1000 mg, 30-minute infusion, every 8 h). Furthermore, we assessed the impact of CLCRCG on meropenem concentrations and target attainment and developed a tool for risk assessment of target non-attainment. Results: Large inter-and intra-patient variability in meropenem concentrations was observed in the critically ill population (n = 48). Attainment of the target 100% T->MIC was merely 48.4% and 20.6%, given MIC values of 2 mg/L and 8 mg/L, respectively, and similar for the target 50% T->4xMIC. A hyperbolic relationship between CLCRCG (25-255 ml/minute) and meropenem serum concentrations at the end of the dosing interval (C-8h) was derived. For infections with pathogens of MIC 2 mg/L, mild renal impairment up to augmented renal function was identified as a risk factor for target non-attainment (for MIC 8 mg/L, additionally, moderate renal impairment). Conclusions: The investigated standard meropenem dosing regimen appeared to result in insufficient meropenem exposure in a considerable fraction of critically ill patients. An easy-and free-to-use tool (the MeroRisk Calculator) for assessing the risk of target non-attainment for a given renal function and MIC value was developed.
Populations of several long-distance migratory songbirds in Eurasia are in peril, drastically illustrated by the recent range-wide population collapse in the Yellow-breasted Bunting Emberiza aureola. There are signals of a strong decline also in the Rustic Bunting E. rustica, but no range-wide assessment of population trends in this superabundant and widespread bunting species has yet been undertaken. The conservation status of Rustic Bunting is ‘Least Concern’ on the global IUCN Red List, but it has recently been upgraded to ‘Vulnerable’ on the European Red List. To assess the Rustic Bunting’s global conservation status we compiled, for the first time, population data across its breeding and wintering ranges. The analysis reveals a 75–87% decline in overall population size over the last 30 years and a 32–91% decline over the last 10 years. The trend estimates indicate that the long-term (30-year) range-wide population decline in the Rustic Bunting is of similar magnitude to two well-known examples of declining species within the same genus, the Yellow-breasted Bunting and the Ortolan Bunting E. hortulana. The magnitude of the range-wide population decline over the last 10 years suggests that the Rustic Bunting could be upgraded from ‘Least Concern’ to ‘Vulnerable’ or ‘Endangered’ on the IUCN global Red List. Agricultural intensification in the wintering range and intensified levels of disturbance, including logging and fire, in the breeding range could be important drivers of the range-wide population decline, and persecution could also contribute. Untangling threat factors and their interactions on Rustic Bunting is necessary for conservation, but hampered by our currently limited understanding of the relationships between population dynamics and different threats.
Plastic pollution is a major global concern with several million microplastic particles entering every day freshwater ecosystems via wastewater discharge. Microplastic particles stimulate biofilm formation (plastisphere) throughout the water column and have the potential to affect microbial community structure if they accumulate in pelagic waters, especially enhancing the proliferation of biohazardous bacteria. To test this scenario, we simulated the inflow of treated wastewater into a temperate lake using a continuous culture system with a gradient of concentration of microplastic particles. We followed the effect of microplastics on the microbial community structure and on the occurrence of integrase 1 (intl), a marker associated with mobile genetic elements known as a proxy for anthropogenic effects on the spread of antimicrobial resistance genes. The abundance of intl increased in the plastisphere with increasing microplastic particle concentration, but not in the water surrounding the microplastic particles. Likewise, the microbial community on microplastic was more similar to the original wastewater community with increasing microplastic concentrations. Our results show that microplastic particles indeed promote persistence of typical indicators of microbial anthropogenic pollution in natural waters, and substantiate that their removal from treated wastewater should be prioritised. (C) 2017 Elsevier Ltd. All rights reserved.
Background
A territory as a prerequisite for breeding limits the maximum number of breeders in a given area, and thus lowers the proportion of breeders if population size increases. However, some territorially breeding animals can have dramatic density fluctuations and little is known about the change from density-dependent processes to density-independence of breeding during a population increase or an outbreak. We suggest that territoriality, breeding suppression and its break-down can be understood with an incomplete-control model, developed for social breeders and social suppression.
Results
We studied density dependence in an arvicoline species, the bank vole, known as a territorial breeder with cyclic and non-cyclic density fluctuations and periodically high densities in different parts of its range. Our long-term data base from 38 experimental populations in large enclosures in boreal grassland confirms that breeding rates are density-regulated at moderate densities, probably by social suppression of subordinate potential breeders. We conducted an experiment, were we doubled and tripled this moderate density under otherwise the same conditions and measured space use, mortality, reproduction and faecal stress hormone levels (FGM) of adult females. We found that mortality did not differ among the densities, but the regulation of the breeding rate broke down: at double and triple densities all females were breeding, while at the low density the breeding rate was regulated as observed before. Spatial overlap among females increased with density, while a minimum territory size was maintained. Mean stress hormone levels were higher in double and triple densities than at moderate density.
Conclusions
At low and moderate densities, breeding suppression by the dominant breeders, But above a density-threshold (similar to a competition point), the dominance of breeders could not be sustained (incomplete control). In our experiment, this point was reached after territories could not shrink any further, while the number of intruders continued to increase with increasing density. Probably suppression becomes too costly for the dominants, and increasing number of other breeders reduces the effectiveness of threats. In wild populations, crossing this threshold would allow for a rapid density increase or population outbreaks, enabling territorial species to escape density-dependency.
Pregnancy termination after encountering a strange male, the Bruce effect, is regarded as a counterstrategy of female mammals towards anticipated infanticide. While confirmed in caged rodent pairs, no verification for the Bruce effect existed from experimental field populations of small rodents. We suggest that the effect may be adaptive for breeding rodent females only under specific conditions related to populations with cyclically fluctuating densities. We investigated the occurrence of delay in birth date after experimental turnover of the breeding male under different population composition in bank voles (Myodes glareolus) in large outdoor enclosures: one-male-multiple-females (n = 6 populations/18 females), multiple-males-multiplefemales (n = 15/45), and single-male-single-female (MF treatment, n = 74/74). Most delays were observed in the MF treatment after turnover. Parallel we showed in a laboratory experiment (n = 205 females) that overwintered and primiparous females, the most abundant cohort during population lows in the increase phase of cyclic rodent populations, were more likely to delay births after turnover of the male than year-born and multiparous females. Taken together, our results suggest that the Bruce effect may be an adaptive breeding strategy for rodent females in cyclic populations specifically at low densities in the increase phase, when isolated, overwintered animals associate in MF pairs. During population lows infanticide risk and inbreeding risk may then be higher than during population highs, while also the fitness value of a litter in an increasing population is higher. Therefore, the Bruce effect may be adaptive for females during annual population lows in the increase phases, even at the costs of delaying reproduction.
JUNGBRUNNEN1 Confers Drought Tolerance Downstream of the HD-Zip I Transcription Factor AtHB13
(2017)
Low water availability is the major environmental factor limiting growth and productivity of plants and crops and is therefore considered of high importance for agriculture affected by climate change. Identifying regulatory components controlling the response and tolerance to drought stress is thus of major importance. The NAC transcription factor (TF) JUNGBRUNNEN1 (JUB1) from Arabidopsis thaliana extends leaf longevity under non-stress growth conditions, lowers cellular hydrogen peroxide (H2O2) level, and enhances tolerance against heat stress and salinity. Here, we additionally find that JUB1 strongly increases tolerance to drought stress in Arabidopsis when expressed from both, a constitutive (CaMV 35S) and an abiotic stress-induced (RD29A) promoter. Employing a yeast one-hybrid screen we identified HD-Zip class I TF AtHB13 as an upstream regulator of JUB1. AtHB13 has previously been reported to act as a positive regulator of drought tolerance. AtHB13 and JUB1 thereby establish a joint drought stress control module.
The amyloid precursor-like protein 1 (APLP1) is a type I transmembrane protein that plays a role in synaptic adhesion and synaptogenesis. Past investigations indicated that APLP1 is involved in the formation of protein-protein complexes that bridge the junctions between neighboring cells. Nevertheless, APLP1-APLP1 trans interactions have never been directly observed in higher eukaryotic cells. Here, we investigated APLP1 interactions and dynamics directly in living human embryonic kidney cells using fluorescence fluctuation spectroscopy techniques, namely cross-correlation scanning fluorescence correlation spectroscopy and number and brightness analysis. Our results show that APLP1 forms homotypic trans complexes at cell-cell contacts. In the presence of zinc ions, the protein forms macroscopic clusters, exhibiting an even higher degree of trans binding and strongly reduced dynamics. Further evidence from giant plasma membrane vesicles suggests that the presence of an intact cortical cytoskeleton is required for zinc-induced cis multimerization. Subsequently, large adhesion platforms bridging interacting cells are formed through APLP1-APLP1 trans interactions. Taken together, our results provide direct evidence that APLP1 functions as a neuronal zinc-dependent adhesion protein and allow a more detailed understanding of the molecular mechanisms driving the formation of APLP1 adhesion platforms.
Direct visualization of APLP1 cell-cell adhesion platforms via fluorescence fluctuation spectroscopy
(2017)
Single molecule RNA fluorescent in situ hybridization (smFISH) enables gene transcription to be assessed at the cellular level. In this point of view article, we describe our recent smFISH research in the model plant Arabidopsis thaliana and discuss how this technique could further knowledge of plant gene transcription in the future.
Background: Leishmania tarentolae, a unicellular eukaryotic protozoan, has been established as a novel host for recombinant protein production in recent years. Current protocols for protein expression in Leishmania are, however, time consuming and require extensive lab work in order to identify well-expressing cell lines. Here we established an alternative protein expression work-flow that employs recently engineered infrared fluorescence protein (IFP) as a suitable and easy-to-handle reporter protein for recombinant protein expression in Leishmania. As model proteins we tested three proteins from the plant Arabidopsis thaliana, including a NAC and a type-B ARR transcription factor.
Results: IFP and IFP fusion proteins were expressed in Leishmania and rapidly detected in cells by deconvolution microscopy and in culture by infrared imaging of 96-well microtiter plates using small cell culture volumes (2 mu L - 100 mu L). Motility, shape and growth of Leishmania cells were not impaired by intracellular accumulation of IFP. In-cell detection of IFP and IFP fusion proteins was straightforward already at the beginning of the expression pipeline and thus allowed early pre-selection of well-expressing Leishmania clones. Furthermore, IFP fusion proteins retained infrared fluorescence after electrophoresis in denaturing SDS-polyacrylamide gels, allowing direct in-gel detection without the need to disassemble cast protein gels. Thus, parameters for scaling up protein production and streamlining purification routes can be easily optimized when employing IFP as reporter.
Conclusions: Using IFP as biosensor we devised a protocol for rapid and convenient protein expression in Leishmania tarentolae. Our expression pipeline is superior to previously established methods in that it significantly reduces the hands-on-time and work load required for identifying well-expressing clones, refining protein production parameters and establishing purification protocols. The facile in-cell and in-gel detection tools built on IFP make Leishmania amenable for high-throughput expression of proteins from plant and animal sources.
The pyranopterin dithiolene (PDT) ligand is an integral component of the molybdenum cofactor (Moco) found in all molybdoenzymes with the sole exception of nitrogenase. However, the roles of the PDT in catalysis are still unknown. The PDT is believed to be bound to the proteins by an extensive hydrogen bonding network, and it has been suggested that these interactions may function to fine-tune Moco for electron- and atom-transfer reactivity in catalysis. Here, we use resonance Raman (rR) spectroscopy to probe Moco-protein interactions using heavy-atom congeners of lumazine, molecules that bind tightly to both wild-type xanthine dehydrogenase (wt-XDH) and its Q102G and Q197A variants following enzymatic hydroxylation to the corresponding violapterin product molecules. The resulting enzyme-product complexes possess intense near-IR absorption, allowing high-quality rR spectra to be collected on wt-XDH and the Q102G and Q197A variants. Small negative frequency shifts relative to wt-XDH are observed for the low-frequency Moco vibrations. These results are interpreted in the context of weak hydrogen-bonding and/or electrostatic interactions between Q102 and the -NH2 terminus of the PDT, and between Q197 and the terminal oxo of the Mo equivalent to O group. The Q102A, Q102G, Q197A, and Q197E variants do not appreciably affect the kinetic parameters k(red) and k(red)/K-D, indicating that a primary role for these glutamine residues is to stabilize and coordinate Moco in the active site of XO family enzymes but to not directly affect the catalytic throughput. Raman frequency shifts between wt-XDH and its Q102G variant suggest that the changes in the electron density at the Mo ion that accompany Mo oxidation during electron-transfer regeneration of the catalytically competent active site are manifest in distortions at the distant PDT amino terminus. This implies a primary role for the PDT as a conduit for facilitating enzymatic electron-transfer reactivity in xanthine oxidase family enzymes.
Comparing mitogenomic timetrees for two African savannah primate genera (Chlorocebus and Papio)
(2017)
Complete mitochondrial (mtDNA) genomes have proved to be useful in reconstructing primate phylogenies with higher resolution and confidence compared to reconstructions based on partial mtDNA sequences. Here, we analyse complete mtDNA genomes of African green monkeys (genus Chlorocebus), a widely distributed primate genus in Africa representing an interesting phylogeographical model for the evolution of savannah species. Previous studies on partial mtDNA sequences revealed nine major clades, suggesting several cases of para- and polyphyly among Chlorocebus species. However, in these studies, phylogenetic relationships among several clades were not resolved, and divergence times were not estimated. We analysed complete mtDNA genomes for ten Chlorocebus samples representing major mtDNA clades to find stronger statistical support in the phylogenetic reconstruction than in the previous studies and to estimate divergence times. Our results confirmed para- and polyphyletic relationships of most Chlorocebus species, while the support for the phylogenetic relationships between the mtDNA clades increased compared to the previous studies. Our results indicate an initial west-east division in the northern part of the Chlorocebus range with subsequent divergence into north-eastern and southern clades. This phylogeographic scenario contrasts with that for another widespread African savannah primate genus, the baboons (Papio), for which a dispersal from southern Africa into East and West Africa was suggested.
Die Hybridomtechnik zur Produktion von monoklonalen Antikörpern ermöglichte einen großen Schritt in der Entwicklung von Immunoassays für die biochemische Forschung und klinische Diagnostik. Auch die Produktion von Antikörpern gegen niedermolekulare Analyten, Haptene, typische Targets in der Lebensmittel- und Umweltanalytik, erlangte in den letzten Jahren eine immer größere Bedeutung. Im Zuge der Durchführung der Hybridomtechnik werden tausende Antikörper-sezernierende und nicht-sezernierende Zellen generiert. Die Selektion der wenigen antigenselektiven Hybridomzellen zählt dabei zu den herausforderndsten Schritten für die Antikörpergewinnung. Bisherige Selektionsverfahren, wie die Limiting-Dilution-Klonierung in Verbindung mit Enzyme-linked Immunosorbent Assays (ELISAs), garantieren keine Monoklonalität und erlauben nur das Screening von einigen wenigen Zellklonen. Hingegen ermöglichen Hochdurchsatz-Selektionsmethoden, wie die Fluoreszenz-aktivierte Zellsortierung (FACS), einen sehr hohen Probendurchsatz. Eine Einzelzellablage garantiert hierbei Monoklonalität. Jedoch sind die dafür erforderlichen Zellmarkierungen oftmals zellschädigend oder aufwendig zu generieren. Auch ist bisher noch keine Markierungsmethode bekannt, die es ermöglicht, Hapten-selektive Hybridomzellen durchflusszytometrisch zu analysieren und eine FACS-Selektion durchzuführen.
Aus diesem Grund wurden in dieser Arbeit zwei Zellmarkierungsmethoden entwickelt, die dies ermöglichen sollten. Die membranständigen Antikörper von Hybridomzellen sollten entweder direkt oder indirekt immunfluoreszenz-markiert und dadurch für die Durchflusszytometrie und FACS-Selektion zugänglich gemacht werden. Die direkte Markierung wurde mittels eines Hapten-Fluorophor-Konjugats durchgeführt. Sie ermöglichte erstmalig den Anteil an Haptenselektiven Hybridomzellen in einer Hybridomzelllinie zu überprüfen. Dies konnte für zwei Hapten-selektive Hybridomzelllinien, die Antikörper gegen das Hormon 17β-Estradiol und das Cardenolid Digoxigenin bilden, gezeigt werden. Durchflusszytometrie und ELISAs lieferten vergleichbare Ergebnisse. Zellen, die Hapten-selektiv markiert werden konnten, sezernierten ebenfalls Hapten-selektive Antikörper. Des Weiteren konnte die direkte Markierung dazu genutzt werden, zwei Mykotoxin-selektive Hybridomzelllinien, welche Antikörper gegen Aflatoxin und Zearalenon bilden, auf Monoklonalität zu testen. Dies ist mittels ELISA nicht möglich. Die Markierungsmethode eignete sich jedoch nur für fixierte Hybridomzellen. Eine Markierung von lebenden Zellen konnte weder durchflusszytometrisch noch mittels konfokaler Laser-Scanning-Mikroskopie gezeigt werden.
Dies gelang erst mit einer neu entwickelten indirekten Immunfluoreszenzmarkierung. Dabei wurden die Zellen zunächst mit einem Hapten-Peroxidase-Konjugat inkubiert, gefolgt von einem Fluorophor-markierten anti-HRP-Antikörper-Konjugat. Dies wurde für zwei Analyten, das Hormon Estron und das Antiepileptikum Carbamazepin, gezeigt. Die indirekte Markierung wurde erfolgreich dazu verwendet, Carbamazepin-selektive Hybridomzellen aus einem Fusionsansatz für die monoklonale Antikörperproduktion auszusortieren. Damit wurde erstmalig eine Zellmarkierungsmethode entwickelt, die eine Hochdurchsatz-Selektion lebender Hybridomzellen aus einem Fusionsansatz ermöglicht. Sie ist nicht zellschädigend und kann zusätzlich zur Selektion Hapten-selektiver Plasmazellen verwendet werden.
Rubisco catalyses the first step of CO2 assimilation into plant biomass. Despite its crucial role, it is notorious for its low catalytic rate and its tendency to fix O2 instead of CO2, giving rise to a toxic product that needs to be recycled in a process known as photorespiration. Since almost all our food supply relies on Rubisco, even small improvements in its specificity for CO2 could lead to an improvement of photosynthesis and ultimately, crop yield. In this work, we attempted to improve photosynthesis by decreasing photorespiration with an artificial CCM based on a fusion between Rubisco and a carbonic anhydrase (CA).
A preliminary set of plants contained fusions between one of two CAs, bCA1 and CAH3, and the N- or C-terminus of RbcL connected by a small flexible linker of 5 amino acids. Subsequently, further fusion proteins were created between RbcL C-terminus and bCA1/CAH3 with linkers of 14, 23, 32, and 41 amino acids. The transplastomic tobacco plants carrying fusions with bCA1 were able to grow autotrophically even with the shortest linkers, albeit at a low rate, and accumulated very low levels of the fusion protein. On the other hand, plants carrying fusions with CAH3 were autotrophic only with the longer linkers. The longest linker permitted nearly wild-type like growth of the plants carrying fusions with CAH3 and increased the levels of fusion protein, but also of smaller degradation products.
The fusion of catalytically inactive CAs to RbcL did not cause a different phenotype from the fusions with catalytically active CAs, suggesting that the selected CAs were not active in the fusion with RbcL or their activity did not have an effect on CO2 assimilation. However, fusions to RbcL did not abolish RbcL catalytic activity, as shown by the autotrophic growth, gas exchange and in vitro activity measurements. Furthermore, Rubisco carboxylation rate and specificity for CO2 was not altered in some of the fusion proteins, suggesting that despite the defect in RbcL folding or assembly caused by the fusions, the addition of 60-150 amino acids to RbcL does not affect its catalytic properties. On the contrary, most growth defects of the plants carrying RbcL-CA fusions are related to their reduced Rubisco content, likely caused by impaired RbcL folding or assembly. Finally, we found that fusions with RbcL C-terminus were better tolerated than with the N-terminus, and increasing the length of the linker relieved the growth impairment imposed by the fusion to RbcL. Together, the results of this work constitute considerable relevant findings for future Rubisco engineering.
Nutrition bioassays in which polyunsaturated fatty acids (PUFA)-deficient diets were supplemented with free long-chain PUFA (>= C20) consistently revealed positive effects on somatic growth and fecundity of Daphnia. However, free PUFA are hardly available in natural diets. In general, PUFA are bound to other lipids, especially to phospholipids and triglycerides. Here, we evaluate the potential of free and phospholipid-bound dietary eicosapentaenoic acid (EPA) to support somatic growth and fecundity of Daphnia magna. In a growth experiment, supplementation of a C20 PUFA-deficient diet with free or phospholipid-bound EPA improved somatic growth rates of D. magna equally. However, the increase in fecundity was significantly more pronounced when phospholipid-bound EPA was provided. Free and phospholipid-bound EPA were provided in the same concentrations in our experiment, suggesting that the allocation to reproduction-related processes is affected differently by phospholipid-bound PUFA and free PUFA. Our finding stresses the need to consider the distribution of dietary PUFA in different lipid classes to gain a better understanding of how PUFA influence life history traits of Daphnids in the field.
Reciprocal selection between aphids, their protective endosymbionts, and the parasitoid wasps that prey upon them offers an opportunity to study the basis of their coevolution. We investigated adaptation to symbiont‐conferred defense by rearing the parasitoid wasp Lysiphlebus fabarum on aphids (Aphis fabae) possessing different defensive symbiont strains (Hamiltonella defensa). After ten generations of experimental evolution, wasps showed increased abilities to parasitize aphids possessing the H. defensa strain they evolved with, but not aphids possessing the other strain. We show that the two symbiont strains encode different toxins, potentially creating different targets for counter‐adaptation. Phenotypic and behavioral comparisons suggest that neither life‐history traits nor oviposition behavior differed among evolved parasitoid lineages. In contrast, comparative transcriptomics of adult female wasps identified a suite of differentially expressed genes among lineages, even when reared in a common, symbiont‐free, aphid host. In concurrence with the specificity of each parasitoid lineages’ infectivity, most differentially expressed parasitoid transcripts were also lineage‐specific. These transcripts are enriched with putative venom toxins and contain highly expressed, potentially defensive viral particles. Together, these results suggest that wild populations of L. fabarum employ a complicated offensive arsenal with sufficient genetic variation for wasps to adapt rapidly and specifically to their hosts’ microbial defenses.
Background: Rodents of the genus Rattus are among the most pervasive and successful invasive species, causing major vicissitudes in native ecological communities. A broad and flexible generalist diet has been suggested as key to the invasion success of Rattus spp. Here, we use an indirect approach to better understand foraging niche width, plasticity, and overlap within and between introduced Rattus spp. in anthropogenic habitats and natural humid forests of Madagascar. Results: Based on stable carbon and nitrogen isotope values measured in hair samples of 589 individual rodents, we found that Rattus rattus had an extremely wide foraging niche, encompassing the isotopic space covered by a complete endemic forest-dwelling Malagasy small mammal community. Comparisons of Bayesian standard ellipses, as well as (multivariate) mixed-modeling analyses, revealed that the stable isotope niche of R. rattus tended to change seasonally and differed between natural forests and anthropogenic habitats, indicating plasticity in feeding niches. In co-occurrence, R. rattus and Rattus norvegicus partitioned feeding niches. Isotopic mismatch of signatures of individual R. rattus and the habitat in which they were captured, indicate frequent dispersal movements for this species between natural forest and anthropogenic habitats. Conclusions: Since R. rattus are known to transmit a number of zoonoses, potentially affecting communities of endemic small mammals, as well as humans, these movements presumably increase transmission potential. Our results suggest that due to their generalist diet and potential movement between natural forest and anthropogenic habitats, Rattus spp. might affect native forest-dependent Malagasy rodents as competitors, predators, and disease vectors. The combination of these effects helps explain the invasion success of Rattus spp. and the detrimental effects of this genus on the endemic Malagasy rodent fauna.
1. Given fundamental energetic trade-offs among growth, maintenance and reproduction, individual differences in energy saving should have consequences for survival and reproductive success. Many endotherms use periodic heterothermy to reduce energy and water requirements and individual variation in heterothermy should have fitness consequences. However, attempts to disentangle individual- and population-level variation in heterothermy are scarce. 2. Here, we quantified patterns of heterothermy of 55 free-ranging eastern chipmunks (Tamias striatus), food-hoarding hibernators. Over five hibernation periods, we obtained a total of 7108 daily individual heterothermy indices (median: 118 per individual). 3. Based on an individual reaction norm approach, we found that the use of heterothermy was repeatable and varied among individuals of the same population under similar environmental conditions. This among-individual variation had consequences for winter survival and reproductive success. Individuals using less heterothermy at the beginning of the winter had decreased survival in resource-rich but not in resource-poor years and higher reproductive success in the subsequent breeding season. 4. These results support the hypothesis that fluctuating selection maintains heterothermic diversity and suggest that individualized ecophysiology can contribute to a more thorough understanding of the evolution of energy-saving strategies in endotherms.
While branched polyglycerol (PG)-based molecules are well established as hydrophilic particles, the capacity of utilizing PG in bulk materials and opportunities arising by their further surface functionalization have only recently been considered. Here we investigated how the mold used in PG network synthesis may affect surface composition and how the permeability of substances through PG can be controlled by altering network structure, i.e. introducing 20mol% oligoethylene glycol (OEG) bifunctional spacer molecules. Overall, PG-based bulk network materials were shown to be tailorable, hydrophilic, low swelling and relatively stiff polyether-based materials, with low impact of salt onto material properties. Based on these features, but also on the principal capacity of free hydroxyl groups to be used for functionalization reactions, these materials may be an interesting platform for medical and technical applications, e.g. as diffusion-rate controlling membrane in aqueous environment. Copyright (c) 2016 John Wiley & Sons, Ltd.
Changes in body height throughout extended historic periods are very complex and dynamic processes. Thispilot study aimed to investigate the pattern of longitudinal height z-scores changes in children before and after entering kindergarten. In summer 2016, we measured height and weight of 32 children from 4 groups of two kindergartens aged 3–6 years. All ages were centered according to the age of entry into the kindergarten. For each child we determined mean z-scores for height before and after entering the kindergarten, and assessed the variances for each kindergarten group. Twenty-two children targeted in height z-scores towards average height of their respective kindergarten group, 10 children did not. Due to the small numbers, the convergence in height variance however, remained insignificant (chi-squared independence test, p = 0.127). Additional studies with larger sample sizes are needed to confirm this pilot study.
Since its discovery over two decades ago as an important cell death regulator in Arabidopsis thaliana, the role of LESION SIMULATING DISEASE 1 (LSD1) has been studied intensively within both biotic and abiotic stress responses as well as with respect to plant fitness regulation. However, its molecular mode of action remains enigmatic. Here, we demonstrate that nucleo-cytoplasmic LSD1 interacts with a broad range of other proteins that are engaged in various molecular pathways such as ubiquitination, methylation, cell cycle control, gametogenesis, embryo development and cell wall formation. The interaction of LSD1 with these partners is dependent on redox status, as oxidative stress significantly changes the quantity and types of LSD1-formed complexes. Furthermore, we show that LSD1 regulates the number and size of leaf mesophyll cells and affects plant vegetative growth. Importantly, we also reveal that in addition to its function as a scaffold protein, LSD1 acts as a transcriptional regulator. Taken together, our results demonstrate that LSD1 plays a dual role within the cell by acting as a condition-dependent scaffold protein and as a transcription regulator.
Neisseria gonorrhoeae is the causative organism of gonorrhoea, a sexually transmitted disease that globally accounts for an estimated 80 to 100 million new infections per year. Increasing resistances to all common antibiotics used for N. gonorrhoeae treatment pose the risk of an untreatable disease. Further knowledge of ways of infection and host immune response are needed to understand the pathogen-host interaction and to discover new treatment alternatives against this disease. Therefore, detailed information about immunogenic proteins and their properties like epitope sites could advance further research in this area. In this work, we investigated immunogenic proteins of N. gonorrhoeae for linear epitopes by microarrays. Dominant linear epitopes were identified for eleven of the nineteen investigated proteins with three polyclonal rabbit antibodies from different immunisations. Identified linear epitopes were further examined for non-specific binding with antibodies to Escherichia coli and the closely related pathogen Neisseria meningitidis. On top of that, amino acids crucial for the antibody epitope binding were detected by microarray based alanine scans.
Polyunsaturated fatty acids (PUFA), especially long-chain (i.e., >= 20 carbons) polyunsaturated fatty acids (LC-PUFA), are fundamental to the health and survival of marine and terrestrial organisms. Therefore, it is imperative that we gain a better understanding of their origin, abundance, and transfer between and within these ecosystems. We evaluated the natural variation in PUFA distribution and abundance that exists between and within these ecosystems by amassing and analyzing, using multivariate and analysis of variance (ANOVA) methods, >3000 fatty acid (FA) profiles from marine and terrestrial organisms. There was a clear dichotomy in LC-PUFA abundance between organisms in marine and terrestrial ecosystems, mainly driven by the C-18 PUFA in terrestrial organisms and omega-3 (n-3) LC-PUFA in marine organisms. The PUFA content of an organism depended on both its biome (marine vs terrestrial) and taxonomic group. Within the marine biome, the PUFA content varied among taxonomic groups. PUFA content of marine organisms was dependent on both geographic zone (i.e., latitude, and thus broadly related to temperature) and trophic level (a function of diet). The contents of n-3 LC-PUFA were higher in polar and temperate marine organisms than those from the tropics. Therefore, we conclude that, on a per capita basis, high latitude marine organisms provide a disproportionately large global share of these essential nutrients to consumers, including terrestrial predators. Our analysis also hints at how climate change, and other anthropogenic stressors, might act to negatively impact the global distribution and abundance of n-3 LC-PUFA within marine ecosystems and on the terrestrial consumers that depend on these subsidies.
Near the end of the Pleistocene epoch, populations of the woolly mammoth (Mammuthus primigenius) were distributed across parts of three continents, from western Europe and northern Asia through Beringia to the Atlantic seaboard of North America. Nonetheless, questions about the connectivity and temporal continuity of mammoth populations and species remain unanswered. We use a combination of targeted enrichment and high-throughput sequencing to assemble and interpret a data set of 143 mammoth mitochondrial genomes, sampled from fossils recovered from across their Holarctic range. Our dataset includes 54 previously unpublished mitochondrial genomes and significantly increases the coverage of the Eurasian range of the species. The resulting global phylogeny confirms that the Late Pleistocene mammoth population comprised three distinct mitochondrial lineages that began to diverge ~1.0–2.0 million years ago (Ma). We also find that mammoth mitochondrial lineages were strongly geographically partitioned throughout the Pleistocene. In combination, our genetic results and the pattern of morphological variation in time and space suggest that male-mediated gene flow, rather than large-scale dispersals, was important in the Pleistocene evolutionary history of mammoths.
There is the tendency to assume that endangered species have been both genetically and demographically healthier in the past, so that any genetic erosion observed today was caused by their recent decline. The Iberian lynx (Lynx pardinus) suffered a dramatic and continuous decline during the 20th century, and now shows extremely low genome- and species-wide genetic diversity among other signs of genomic erosion. We analyze ancient (N = 10), historical (N = 245), and contemporary (N = 172) samples with microsatellite and mitogenome data to reconstruct the species' demography and investigate patterns of genetic variation across space and time. Iberian lynx populations transitioned from low but significantly higher genetic diversity than today and shallow geographical differentiation millennia ago, through a structured metapopulation with varying levels of diversity during the last centuries, to two extremely genetically depauperate and differentiated remnant populations by 2002. The historical subpopulations show varying extents of genetic drift in relation to their recent size and time in isolation, but these do not predict whether the populations persisted or went finally extinct. In conclusion, current genetic patterns were mainly shaped by genetic drift, supporting the current admixture of the two genetic pools and calling for a comprehensive genetic management of the ongoing conservation program. This study illustrates how a retrospective analysis of demographic and genetic patterns of endangered species can shed light onto their evolutionary history and this, in turn, can inform conservation actions.
Pollen-based quantitative reconstructions of past climate variables is a standard palaeoclimatic approach. Despite knowing that the spatial extent of the calibration-set affects the reconstruction result, guidance is lacking as to how to determine a suitable spatial extent of the pollen-climate calibration-set. In this study, past mean annual precipitation (P-ann) during the Holocene (since 11.5 cal ka BP) is reconstructed repeatedly for pollen records from Qinghai Lake (36.7 degrees N, 100.5 degrees E; north-east Tibetan Plateau), Gonghai Lake (38.9 degrees N, 112.2 degrees E; north China) and Sihailongwan Lake (42.3 degrees N, 126.6 degrees E; north-east China) using calibration-sets of varying spatial extents extracted from the modern pollen dataset of China and Mongolia (2559 sampling sites and 168 pollen taxa in total). Results indicate that the spatial extent of the calibration-set has a strong impact on model performance, analogue quality and reconstruction diagnostics (absolute value, range, trend, optimum). Generally, these effects are stronger with the modern analogue technique (MAT) than with weighted averaging partial least squares (WA-PLS). With respect to fossil spectra from northern China, the spatial extent of calibration-sets should be restricted to radii between ca. 1000 and 1500 km because small-scale calibration-sets (<800 km radius) will likely fail to include enough spatial variation in the modern pollen assemblages to reflect the temporal range shifts during the Holocene, while too broad a scale calibration-set (>1500 km radius) will include taxa with very different pollen-climate relationships. (C) 2017 Elsevier Ltd. All rights reserved.
Macroecology and biogeography are concerned with understanding biodiversity patterns across space and time. In the past, the two disciplines have addressed this question mainly with correlative approaches, despite frequent calls for more mechanistic explanations. Recent advances in computational power, theoretical understanding, and statistical tools are, however, currently facilitating the development of more system-oriented, mechanistic models. We review these models, identify different model types and theoretical frameworks, compare their processes and properties, and summarize emergent findings. We show that ecological (physiology, demographics, dispersal, biotic interactions) and evolutionary processes, as well as environmental and human-induced drivers, are increasingly modelled mechanistically; and that new insights into biodiversity dynamics emerge from these models. Yet, substantial challenges still lie ahead for this young research field. Among these, we identify scaling, calibration, validation, and balancing complexity as pressing issues. Moreover, particular process combinations are still understudied, and so far models tend to be developed for specific applications. Future work should aim at developing more flexible and modular models that not only allow different ecological theories to be expressed and contrasted, but which are also built for tight integration with all macroecological data sources. Moving the field towards such a ‘systems macroecology’ will test and improve our understanding of the causal pathways through which eco-evolutionary processes create diversity patterns across spatial and temporal scales.
In Escherichia coli, two different systems that are important for the coordinate formation of Fe–S clusters have been identified, namely, the ISC and SUF systems. The ISC system is the housekeeping Fe–S machinery, which provides Fe–S clusters for numerous cellular proteins. The IscS protein of this system was additionally revealed to be the primary sulfur donor for several sulfur-containing molecules with important biological functions, among which are the molybdenum cofactor (Moco) and thiolated nucleosides in tRNA. Here, we show that deletion of central components of the ISC system in addition to IscS leads to an overall decrease in Fe–S cluster enzyme and molybdoenzyme activity in addition to a decrease in the number of Fe–S-dependent thiomodifications of tRNA, based on the fact that some proteins involved in Moco biosynthesis and tRNA thiolation are Fe–S-dependent. Complementation of the ISC deficient strains with the suf operon restored the activity of Fe–S-containing proteins, including the MoaA protein, which is involved in the conversion of 5′GTP to cyclic pyranopterin monophosphate in the fist step of Moco biosynthesis. While both systems share a high degree of similarity, we show that the function of their respective l-cysteine desulfurase IscS or SufS is specific for each cellular pathway. It is revealed that SufS cannot play the role of IscS in sulfur transfer for the formation of 2-thiouridine, 4-thiouridine, or the dithiolene group of molybdopterin, being unable to interact with TusA or ThiI. The results demonstrate that the role of the SUF system is exclusively restricted to Fe–S cluster assembly in the cell.
The trafficking and delivery of sulfur to cofactors and nucleosides is a highly regulated and conserved process among all organisms. All sulfur transfer pathways generally have an L-cysteine desulfurase as an initial sulfur mobilizing enzyme in common, which serves as a sulfur donor for the biosynthesis of sulfur-containing biomolecules like iron sulfur (Fe-S) clusters, thiamine, biotin, lipoic acid, the molybdenum cofactor (Moco), and thiolated nucleosides in tRNA. The human L-cysteine desulfurase NFS1 and the Escherichia coli homologue IscS share a level of amino acid sequence identity of similar to 60%. While E. coli IscS has a versatile role in the cell and was shown to have numerous interaction partners, NFS1 is mainly localized in mitochondria with a crucial role in the biosynthesis of Fe-S clusters. Additionally, NFS1 is also located in smaller amounts in the cytosol with a role in Moco biosynthesis and mcm(5)s(2)U34 thio modifications of nucleosides in tRNA. NFS1 and IscS were conclusively shown to have different interaction partners in their respective organisms. Here, we used functional complementation studies of an E. coli iscS deletion strain with human NFS1 to dissect their conserved roles in the transfer of sulfur to a specific target protein. Our results show that human NFS1 and E. coli IscS share conserved binding sites for proteins involved in Fe-S cluster assembly like IscU, but not with proteins for tRNA thio modifications or Moco biosynthesis. In addition, we show that human NFS1 was almost fully able to complement the role of IscS in Moco biosynthesis when its specific interaction partner protein MOCS3 from humans was also present.
Can't remember to forget you
(2017)
In nature plants are exposed to frequent changes in their abiotic and biotic environment. While some environmental cues are used to gauge the environment and align growth and development, others are beyond the regularly encountered spectrum of a species and trigger stress responses. Such stressful conditions provide a potential threat to survival and integrity. Plants adapt to extreme environmental conditions through physiological adaptations that are usually transient and are maintained until stressful environments subside. It is increasingly appreciated that in some cases environmental cues activate a stress memory that persists for some time after the extreme condition has subsided. Recent research has shown that this stress-induced environmental memory is mediated by epigenetic and chromatin-based mechanisms and both histone methylation and nucleosome occupancy are associated with it.
Plant functional traits reflect individual and community ecological strategies. They allow the detection of directional changes in community dynamics and ecosystemic processes, being an additional tool to assess biodiversity than species richness. Analysis of functional patterns in plant communities provides mechanistic insight into biodiversity alterations due to anthropogenic activity. Although studies have consi-dered of either anthropogenic management or nutrient availability on functional traits in temperate grasslands, studies combining effects of both drivers are scarce. Here, we assessed the impacts of management intensity (fertilization, mowing, grazing), nutrient stoichiometry (C, N, P, K), and vegetation composition on community-weighted means (CWMs) and functional diversity (Rao's Q) from seven plant traits in 150 grasslands in three regions in Germany, using data of 6 years. Land use and nutrient stoichiometry accounted for larger proportions of model variance of CWM and Rao's Q than species richness and productivity. Grazing affected all analyzed trait groups; fertilization and mowing only impacted generative traits. Grazing was clearly associated with nutrient retention strategies, that is, investing in durable structures and production of fewer, less variable seed. Phenological variability was increased. Fertilization and mowing decreased seed number/mass variability, indicating competition-related effects. Impacts of nutrient stoichiometry on trait syndromes varied. Nutrient limitation (large N:P, C:N ratios) promoted species with conservative strategies, that is, investment in durable plant structures rather than fast growth, fewer seed, and delayed flowering onset. In contrast to seed mass, leaf-economics variability was reduced under P shortage. Species diversity was positively associated with the variability of generative traits. Synthesis. Here, land use, nutrient availability, species richness, and plant functional strategies have been shown to interact complexly, driving community composition, and vegetation responses to management intensity. We suggest that deeper understanding of underlying mechanisms shaping community assembly and biodiversity will require analyzing all these parameters.
Tailed bacteriophages specific for Gram‐negative bacteria encounter lipopolysaccharide (LPS) during the first infection steps. Yet, it is not well understood how biochemistry of these initial interactions relates to subsequent events that orchestrate phage adsorption and tail rearrangements to initiate cell entry. For many phages, long O‐antigen chains found on the LPS of smooth bacterial strains serve as essential receptor recognized by their tailspike proteins (TSP). Many TSP are depolymerases and O‐antigen cleavage was described as necessary step for subsequent orientation towards a secondary receptor. However, O‐antigen specific host attachment must not always come along with O‐antigen degradation. In this issue of Molecular Microbiology Prokhorov et al. report that coliphage G7C carries a TSP that deacetylates O‐antigen but does not degrade it, whereas rough strains or strains lacking O‐antigen acetylation remain unaffected. Bacteriophage G7C specifically functionalizes its tail by attaching the deacetylase TSP directly to a second TSP that is nonfunctional on the host's O‐antigen. This challenges the view that bacteriophages use their TSP only to clear their way to a secondary receptor. Rather, O‐antigen specific phages may employ enzymatically active TSP as a tool for irreversible LPS membrane binding to initiate subsequent infection steps.
The homodinuclear ruthenium(II) complex [{Ru(l-N4Me2)}(2)(-tape)](PF6)(4) {[1](PF6)(4)} (l-N4Me2=N,N-dimethyl-2,11-diaza[3.3](2,6)-pyridinophane, tape=1,6,7,12-tetraazaperylene) can store one or two electrons in the energetically low-lying * orbital of the bridging ligand tape. The corresponding singly and doubly reduced complexes [{Ru(l-N4Me2)}(2)(-tape(.-))](PF6)(3) {[2](PF6)(3)} and [{Ru(l-N4Me2)}(2)(-tape(2-))](PF6)(2) {[3](PF6)(2)}, respectively, were electrochemically generated, successfully isolated and fully characterized by single-crystal X-ray crystallography, spectroscopic methods and magnetic susceptibility measurements. The singly reduced complex [2](PF6)(3) contains the -radical tape(.-) and the doubly reduced [3](PF6)(2) the diamagnetic dianion tape(2-) as bridging ligand, respectively. Nucleophilic aromatic substitution at the bridging tape in [1](4+) by two sulfite units gave the complex [{Ru(l-N4Me2)}(2){-tape-(SO3)(2)}](2+) ([4](2+)). Complex dication [4](2+) was exploited as a redox mediator between an anaerobic homogenous reaction solution of an enzyme system (sulfite/sulfite oxidase) and the electrode via participation of the low-energy *-orbital of the disulfonato-substituted bridging ligand tape-(SO3)(2)(2-) (E-red1=-0.1V versus Ag/AgCl/1m KCl in water).
Background
Much of the organismal variation we observe in nature is due to differences in organ size. The observation that even closely related species can show large, stably inherited differences in organ size indicates a strong genetic component to the control of organ size. Despite recent progress in identifying factors controlling organ growth in plants, our overall understanding of this process remains limited, partly because the individual factors have not yet been connected into larger regulatory pathways or networks. To begin addressing this aim, we have studied the upstream regulation of expression of BIG BROTHER (BB), a central growth-control gene in Arabidopsis thaliana that prevents overgrowth of organs. Final organ size and BB expression levels are tightly correlated, implying the need for precise control of its expression. BB expression mirrors proliferative activity, yet the gene functions to limit proliferation, suggesting that it acts in an incoherent feedforward loop downstream of growth activators to prevent over-proliferation.
Results
To investigate the upstream regulation of BB we combined a promoter deletion analysis with a phylogenetic footprinting approach. We were able to narrow down important, highly conserved, cis-regulatory elements within the BB promoter. Promoter sequences of other Brassicaceae species were able to partially complement the A. thaliana bb-1 mutant, suggesting that at least within the Brassicaceae family the regulatory pathways are conserved.
Conclusions
This work underlines the complexity involved in precise quantitative control of gene expression and lays the foundation for identifying important upstream regulators that determine BB expression levels and thus final organ size.
The actin cytoskeleton is an essential intracellular filamentous structure that underpins cellular transport and cytoplasmic streaming in plant cells. However, the system-level properties of actin-based cellular trafficking remain tenuous, largely due to the inability to quantify key features of the actin cytoskeleton. Here, we developed an automated image-based, network-driven framework to accurately segment and quantify actin cytoskeletal structures and Golgi transport. We show that the actin cytoskeleton in both growing and elongated hypocotyl cells has structural properties facilitating efficient transport. Our findings suggest that the erratic movement of Golgi is a stable cellular phenomenon that might optimize distribution efficiency of cell material. Moreover, we demonstrate that Golgi transport in hypocotyl cells can be accurately predicted from the actin network topology alone. Thus, our framework provides quantitative evidence for system-wide coordination of cellular transport in plant cells and can be readily applied to investigate cytoskeletal organization and transport in other organisms.
Late embryogenesis abundant (LEA) proteins are related to cellular dehydration tolerance. Most LEA proteins are predicted to have no stable secondary structure in solution, i.e., to be intrinsically disordered proteins (IDPs), but they may acquire alpha-helical structure upon drying. In the model plant Arabidopsis thaliana, the LEA proteins COR15A and COR15B are highly induced upon cold treatment and are necessary for the plants to attain full freezing tolerance. Freezing leads to increased intracellular crowding due to dehydration by extracellular ice crystals. In vitro, crowding by high glycerol concentrations induced partial folding of COR15 proteins. Here, we have extended these investigations to two related proteins, LEA11 and LEA25. LEA25 is much longer than LEA11 and COR15A, but shares a conserved central sequence domain with the other two proteins. We have created two truncated versions of LEA25 (2H and 4H) to elucidate the structural and functional significance of this domain. Light scattering and CD spectroscopy showed that all five proteins were largely unstructured and monomeric in dilute solution. They folded in the presence of increasing concentrations of trifluoroethanol and glycerol. Additional folding was observed in the presence of glycerol and membranes. Fourier transform infra red spectroscopy revealed an interaction of the LEA proteins with membranes in the dry state leading to a depression in the gel to liquid-crystalline phase transition temperature. Liposome stability assays revealed a cryoprotective function of the proteins. The C- and N-terminal extensions of LEA25 were important in cryoprotection, as the central domain itself (2H, 4H) only provided a low level of protection.
Intrinsically Disordered Stress Protein COR15A Resides at the Membrane Surface during Dehydration
(2017)
Plants from temperate climate zones are able to increase their freezing tolerance during exposure to low, above zero temperatures in a process termed cold acclimation. During this process, several cold-regulated (COR) proteins are accumulated in the cells. One of them is COR15A, a small, intrinsically disordered protein that contributes to leaf freezing tolerance by stabilizing cellular membranes. The isolated protein folds into amphipathic a-helices in response to increased crowding conditions, such as high concentrations of glycerol. Although there is evidence for direct COR15A-membrane interactions, the orientation and depth of protein insertion were unknown. In addition, although folding due to high osmolyte concentrations had been established, the folding response of the protein under conditions of gradual dehydration had not been investigated. Here we show, using Fourier transform infrared spectroscopy, that COR15A starts to fold into a-helices already under mild dehydration conditions (97% relative humidity (RH), corresponding to freezing at -3 degrees C) and that folding gradually increases with decreasing RH. Neutron diffraction experiments at 97 and 75% RH established that the presence of COR15A had no significant influence on the structure of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) membranes. However, using deuterated POPC we. could clearly establish that COR15A interacts with the membranes and penetrates below the headgroup region into the upper part of the fatty acyl chain region. This localization is in agreement with our hypothesis that COR15A-membrane interaction is at least, in part, driven by a hydrophobic interaction between the lipids and the hydrophobic face of the amphipathic protein alpha-helix.
Head and neck squamous cell carcinomas (HNSCC) exhibiting resistance to the EGFR-targeting drug cetuximab poses a challenge to their effective clinical management. Here, we report a specific mechanism of resistance in this setting based upon the presence of a single nucleotide polymorphism encoding EGFR-K-521 (K-allele), which is expressed in > 40% of HNSCC cases. Patients expressing the K-allele showed significantly shorter progressionfree survival upon palliative treatment with cetuximab plus chemotherapy or radiation. In several EGFR-mediated cancer models, cetuximab failed to inhibit downstream signaling or to kill cells harboring a high K-allele frequency. Cetuximab affinity for EGFR-K-521 was reduced slightly, but ligand-mediated EGFR acti-vation was intact. We found a lack of glycan sialyation on EGFR-K-521 that associated with reduced protein stability, suggesting a structural basis for reduced cetuximab efficacy. CetuGEX, an antibody with optimized Fc glycosylation targeting the same epitope as cetuximab, restored HNSCC sensitivity in a manner associated with antibody-dependent cellular cytotoxicity rather than EGFR pathway inhibition. Overall, our results highlight EGFR-K-521 expression as a key mechanism of cetuximab resistance to evaluate prospectively as a predictive biomarker in HNSCC patients. Further, they offer a preclinical rationale for the use of ADCC-optimized antibodies to treat tumors harboring this EGFR isoform.
Oceanic island biogeography through the lens of the general dynamic model: assessment and prospect
(2017)
1. During the last couple of decades, invasive species have become a worldwide problem in many freshwater systems. Besides higher plants and animals, microbes, in particular the potentially toxic cyanobacterium Cylindrospermopsis raciborskii, has attracted increasing attention, due to its spread towards temperate zones of the northern and southern hemisphere. A number of advantageous functional traits and a high intraspecific plasticity have been suggested to explain its invasion success. 2. The aim of this study was to examine intraspecific functional trait variability in 12 different isolates of C.raciborskii originating from different lakes in an invaded region in Northeast Germany. We measured growth rate, C:N:P ratios, chlorophyll-a content and the abundance of heterocysts under nutrient-replete and phosphorus-limited conditions. Moreover, the isolate-specific morphology and grazing losses by an herbivorous rotifer, as a top-down force, were studied. 3. DNA fingerprinting revealed that all isolates were genetically different. C.raciborskii exhibited a large variability in all measured traits among isolates. The C:P, N:P and Chl-a:C ratios differed by a factor of two or more. The trait variability among isolates was higher under nutrient-replete conditions, except for the C:P ratio, which varied most during phosphorus limitation. The susceptibility to grazing, calculated as maximum ingestion rates of the rotifer Brachionus calyciflorus on C.raciborskii, varied most among isolates, but was not related to any of the measured physiological or morphological traits, i.e. no trade-off was found. 4. Ecological and genetic clustering did not match, indicating that the genetic relationship based on DNA fingerprinting did not cover ecological differences. 5. Our results show a high trait variability within locally occurring and partly co-occurring C.raciborskii isolates. No overall trade-offs between the measured functional traits were found. This demonstrates the ecological relevance of linking multiple traits, e.g. competitive and consumptive. Furthermore, this study emphasises the importance of analysing more than one strain of a species, as different strains show different trait values potentially relevant for their invasibility and the field of general trait-based ecology.
G(0)W(0) calculations for predicting vertical ionization potentials (IPs) and electron affinities of molecules and clusters are known to show a significant dependence on the density functional theory (DFT) starting point. A number of nonempirical procedures to find an optimal starting point have been proposed, typically based on tuning the amount of HF exchange in the underlying hybrid functional specifically for the system at hand. For the case of pi-conjugated molecular chains, these approaches lead to a significantly different amount of HF exchange for different oligomer sizes. In this study, we analyze if and how strongly this size dependence affects the ability of nonempirical tuning approaches to predict accurate IPs for pi-conjugated molecular chains of increasing chain length. To this end, we employ three different nonempirical tuning procedures for the G(0)W(0) starting point to calculate the IP of polyene oligomers up to 22 repeat units and compare the results to highly accurate coupled-cluster calculations. We find that, despite its size dependence, using an IP-tuned hybrid functional as a starting point for G(0)W(0) yields excellent agreement with the reference data for all chain lengths.
Global effects of income and income inequality on adult height and sexual dimorphism in height
(2017)
Objectives: Average adult height of a population is considered a biomarker of the quality of the health environment and economic conditions. The causal relationships between height and income inequality are not well understood. We analyze data from 169 countries for national average heights of men and women and national-level economic factors to test two hypotheses: (1) income inequality has a greater association with average adult height than does absolute income; and (2) neither income nor income inequality has an effect on sexual dimorphism in height. Methods: Average height data come from the NCD-RisC health risk factor collaboration. Economic indicators are derived from the World Bank data archive and include gross domestic product (GDP), Gross National Income per capita adjusted for personal purchasing power (GNI_ PPP), and income equality assessed by the Gini coefficient calculated by the Wagstaff method. Results: Hypothesis 1 is supported. Greater income equality is most predictive of average height for both sexes. GNI_ PPP explains a significant, but smaller, amount of the variation. National GDP has no association with height. Hypothesis 2 is rejected. With greater average adult height there is greater sexual dimorphism. Conclusions: Findings support a growing literature on the pernicious effects of inequality on growth in height and, by extension, on health. Gradients in height reflect gradients in social disadvantage. Inequality should be considered a pollutant that disempowers people from the resources needed for their own healthy growth and development and for the health and good growth of their children.
Influenza A virus matrix protein 1 (M1) is an essential component involved in the structural stability of the virus and in the budding of new virions from infected cells. A deeper understanding of the molecular basis of virion formation and the budding process is required in order to devise new therapeutic approaches. We performed a detailed investigation of the interaction between M1 and phosphatidylserine (PS) (i.e., its main binding target at the plasma membrane [PM]), as well as the distribution of PS itself, both in model membranes and in living cells. To this end, we used a combination of techniques, including Forster resonance energy transfer (FRET), confocal microscopy imaging, raster image correlation spectroscopy, and number and brightness (N&B) analysis. Our results show that PS can cluster in segregated regions in the plane of the lipid bilayer, both in model bilayers constituted of PS and phosphatidylcholine and in living cells. The viral protein M1 interacts specifically with PS-enriched domains, and such interaction in turn affects its oligomerization process. Furthermore, M1 can stabilize PS domains, as observed in model membranes. For living cells, the presence of PS clusters is suggested by N&B experiments monitoring the clustering of the PS sensor lactadherin. Also, colocalization between M1 and a fluorescent PS probe suggest that, in infected cells, the matrix protein can specifically bind to the regions of PM in which PS is clustered. Taken together, our observations provide novel evidence regarding the role of PS-rich domains in tuning M1-lipid and M1-M1 interactions at the PM of infected cells. IMPORTANCE Influenza virus particles assemble at the plasma membranes (PM) of infected cells. This process is orchestrated by the matrix protein M1, which interacts with membrane lipids while binding to the other proteins and genetic material of the virus. Despite its importance, the initial step in virus assembly (i.e., M1-lipid interaction) is still not well understood. In this work, we show that phosphatidylserine can form lipid domains in physical models of the inner leaflet of the PM. Furthermore, the spatial organization of PS in the plane of the bilayer modulates M1-M1 interactions. Finally, we show that PS domains appear to be present in the PM of living cells and that M1 seems to display a high affinity for them.
Gelatin-based hydrogels offer various biochemical cues that support encapsulated cells and are therefore suitable as cell delivery vehicles in regenerative medicine. However, besides the biochemical signals, biomechanical cues are crucial to ensure an optimal support of encapsulated cells. Hence, we aimed to correlate the cellular response of encapsulated cells to macroscopic and microscopic elastic properties of glycidylmethacrylate (GMA)-functionalized gelatin-based hydrogels. To ensure that different observations in cellular behavior could be attributed to differences in elastic properties, an identical concentration as well as degree of functionalization of biopolymers was utilized to form covalently crosslinked hydrogels. Elastic properties were merely altered by varying the average gelatin-chain length. Hydrogels exhibited an increased degree of swelling and a decreased bulk elastic modulus G with prolonged autoclaving of the starting solution. This was accompanied by an increase of hydrogel mesh size and thus by a reduction of crosslinking density. Tougher hydrogels retained the largest amount of cells; however, they also interfered with cell viability. Softer gels contained a lower cell density, but supported cell elongation and viability. Observed differences could be partially attributed to differences in bulk properties, as high crosslinking densities interfere with diffusion and cell spreading and thus can impede cell viability. Interestingly, a microscopic elastic modulus in the range of native soft tissue supported cell viability and elongation best while ensuring a good cell entrapment. In conclusion, gelatin-based hydrogels providing a soft tissue-like microenvironment represent adequate cell delivery vehicles for tissue engineering approaches. Copyright (c) 2016 John Wiley & Sons, Ltd.
Recent reports of increasing iron (Fe) concentrations in freshwaters are of concern, given the fundamental role of Fe in biogeochemical processes. Still, little is known about the frequency and geographical distribution of Fe trends or about the underlying drivers. We analyzed temporal trends of Fe concentrations across 340 water bodies distributed over 10 countries in northern Europe and North America in order to gain a clearer understanding of where, to what extent, and why Fe concentrations are on the rise. We found that Fe concentrations have significantly increased in 28% of sites, and decreased in 4%, with most positive trends located in northern Europe. Regions with rising Fe concentrations tend to coincide with those with organic carbon (OC) increases. Fe and OC increases may not be directly mechanistically linked, but may nevertheless be responding to common regional-scale drivers such as declining sulfur deposition or hydrological changes. A role of hydrological factors was supported by covarying trends in Fe and dissolved silica, as these elements tend to stem from similar soil depths. A positive relationship between Fe increases and conifer cover suggests that changing land use and expanded forestry could have contributed to enhanced Fe export, although increases were also observed in nonforested areas. We conclude that the phenomenon of increasing Fe concentrations is widespread, especially in northern Europe, with potentially significant implications for wider ecosystem biogeochemistry, and for the current browning of freshwaters.
Glycoproteins adsorbing on an implant upon contact with body fluids can affect the biological response in vitro and in vivo, depending on the type and conformation of the adsorbed biomacromolecules. However, this process is poorly characterized and so far not controllable. Here, protein monolayers of high molecular cohesion with defined density are transferred onto polymeric substrates by the Langmuir-Schaefer (LS) technique and were compared with solution deposition (SO) method. It is hypothesized that on polydimethylsiloxane (PDMS), a substrate with poor cell adhesion capacity, the fibronectin (FN) layers generated by the LS and SO methods will differ in their organization, subsequently facilitating differential stem cell adhesion behavior. Indeed, atomic force microscopy visualization and immunofluorescence images indicated that organization of the FN layer immobilized on PDMS was uniform and homogeneous. In contrast, FN deposited by SO method was rather heterogeneous with appearance of structures resembling protein aggregates. Human mesenchymal stem cells showed reduced absolute numbers of adherent cells, and the vinculin expression seemed to be higher and more homogenously distributed after seeding on PDMS equipped with FN by LS in comparison with PDMS equipped with FN by SO. These divergent responses could be attributed to differences in the availability of adhesion molecule ligands such as the Arg-Gly-Asp (RGD) peptide sequence presented at the interface. The LS method allows to control the protein layer characteristics, including the thickness and the protein orientation or conformation, which can be harnessed to direct stem cell responses to defined outcomes, including migration and differentiation. Copyright (c) 2016 John Wiley & Sons, Ltd.
How AAA(+) chaperones conformationally remodel specific target proteins in an ATP-dependent manner is not well understood. Here, we investigated the mechanism of the AAA(+) protein Rubisco activase (Rca) in metabolic repair of the photosynthetic enzyme Rubisco, a complex of eight large (RbcL) and eight small (RbcS) subunits containing eight catalytic sites. Rubisco is prone to inhibition by tight-binding sugar phosphates, whose removal is catalyzed by Rca. We engineered a stable Rca hexamer ring and analyzed its functional interaction with Rubisco. Hydrogen/deuterium exchange and chemical crosslinking showed that Rca structurally destabilizes elements of the Rubisco active site with remarkable selectivity. Cryo-electron microscopy revealed that Rca docks onto Rubisco over one active site at a time, positioning the C-terminal strand of RbcL, which stabilizes the catalytic center, for access to the Rca hexamer pore. The pulling force of Rca is fine-tuned to avoid global destabilization and allow for precise enzyme repair.
Epigenetic maintenance of gene repression is essential for development. Polycomb complexes are central to this memory, but many aspects of the underlying mechanism remain unclear. LIKE HETEROCHROMATIN PROTEIN 1 (LHP1) binds Polycomb-deposited H3K27me3 and is required for repression of many Polycomb target genes in Arabidopsis. Here we show that LHP1 binds RNA in vitro through the intrinsically disordered hinge region. By independently perturbing the RNA-binding hinge region and H3K27me3 (trimethylation of histone H3 at Lys27) recognition, we found that both facilitate LHP1 localization and H3K27me3 maintenance. Disruption of the RNAbinding hinge region also prevented formation of subnuclear foci, structures potentially important for epigenetic repression.
Intraspecific trait variability plays an important role in species adaptation to climate change. However, it still remains unclear how plants in semi-arid environments respond to increasing aridity. We investigated the intraspecific trait variability of two common Mediterranean annuals (Geropogon hybridus and Crupina crupinastrum) with similar habitat preferences. They were studied along a steep precipitation gradient in Israel similar to the maximum predicted precipitation changes in the eastern Mediterranean basin (i.e. -30% until 2100). We expected a shift from competitive ability to stress tolerance with decreasing precipitation and tested this expectation by measuring key functional traits (canopy and seed release height, specific leaf area, N-and P-leaf content, seed mass). Further, we evaluated generative bet-hedging strategies by different seed traits. Both species showed different responses along the precipitation gradient. C. crupinastrum exhibited only decreased plant height toward saridity, while G. hybridus showed strong trends of generative adaptation to aridity. Different seed trait indices suggest increased bet-hedging of G. hybridus in arid environments. However, no clear trends along the precipitation gradient were observed in leaf traits (specific leaf area and leaf N-/P-content) in both species. Moreover, variance decomposition revealed that most of the observed trait variation (>> 50%) is found within populations. The findings of our study suggest that responses to increased aridity are highly species-specific and local environmental factors may have a stronger effect on intraspecific trait variation than shifts in annual precipitation. We therefore argue that trait-based analyses should focus on precipitation gradients that are comparable to predicted precipitation changes and compare precipitation effects to effects of local environmental factors. (C) 2017 Gesellschaft fur Okologie. Published by Elsevier GmbH. All rights reserved.
Spatial environmental heterogeneity is considered a fundamental factor for the maintenance of plant species richness. However, it still remains unclear whether heterogeneity may also facilitate coexistence at fine grain sizes or whether other processes, like mass effects and source sink dynamics due to dispersal, control species composition and diversity at these scales. In this study, we used two complimentary analyses to identify the role of heterogeneity within 15 m x 15 m plots for the coexistence of species-rich annual communities in a semi-arid environment along a steep precipitation gradient. Specifically, we: (a) analyzed the effect of environmental heterogeneity on species, functional and phylogenetic diversity within microsites (alpha diversity, 0.06 m(2) and 1 m(2)), across microsites (beta diversity), and diversity at the entire plot (gamma diversity); (b) further we used two null models to detect non-random trait and phylogenetic patterns in order to infer assembly processes, i.e. whether co-occurring species tend to share similar traits (trait convergence) or dissimilar traits (trait divergence). In general, our results showed that heterogeneity had a positive effect on community diversity. Specifically, for alpha diversity, the effect was significant for functional diversity, and not significant for either species or phylogenetic diversities. For beta diversity, all three measures of community diversity (species, functional, and phylogenetic) increased significantly, as they also did for gamma diversity, where functional measures were again stronger than for species or phylogenetic measures. In addition, the null model approach consistently detected trait convergence, indicating that species with similar traits tended to co-occur and had high abundances in a given microsite. While null model analysis across the phylogeny partly supported these trait findings, showing phylogenetic underdispersion at the 1m(2) grain size, surprisingly when species abundances in microsites were analyzed they were more evenly distributed across the phylogenetic tress than expected (phylogenetic overdispersion). In conclusion, our results provide compelling support that environmental heterogeneity at a relatively fine scale is an important factor for species co-existence as it positively affects diversity as well as influences species assembly. Our study underlines the need for trait-based approaches conducted at fine grain sizes in order to better understand species coexistence and community assembly. (C) 2017 Elsevier GmbH. All rights reserved.
Background: We investigated height of Norwegian conscripts in view of the hypothesis of a "community effect on height" using autocorrelation analysis of district heights within a time-span of 20 years at the end of the 19th century and correlations between neighboring districts at this time. Material and methods: After digitalizing available body height data of Norwegian draftees in 1877-1878, 1880 (averaged as 1878), and 1895-1897 (averaged as 1896) we calculated the magnitude of autocorrelation of body height within the same municipality at different time points. Furthermore, we generated three different neighborhood networks, (1) based on Euclidean distances, (2) a minimum spanning tree build on those distances, (3) a network founded on real world road connections. The networks were used to determine the correlation between body height of neighboring districts depending on the number of edges required to connect two municipalities. Results: The autocorrelation value for body heights was around r = 0.5 (for all p < 0.001) in the years 1878 and 1896. The correlation between neighboring districts varied in the Euclidean distance based network between 0.47 and 0.27 approximately for both years in a sorted order, descending from nearest (0-50 km) to farthest (150-200 km, for all p < 0.001). First order neighbors in the minimum spanning tree network correlation was 0.36 in 1878 and 0.42 in 1896 (for all p < 0.001). The values of neighbor correlation in the road connection based network ranged in 1878 from 0.42 (first order neighbors) to 0.17 (forth order neighbors, for all p < 0.01) and in 1896 from 0.46 (first order neighbors) to 0.12 (forth order neighbors, for all p < 0.05). Conclusion: This initial study of Norwegian conscript height data from the 19th century showed significant medium sized effects for the within district autocorrelation between 1878 and 1896 as well as medium neighborhood correlation, slightly lower in comparison to a recent study regarding Swiss conscripts. Digitalizing more data from other years in this and later time spans as well as using older road and ship connections instead of the actual road data might stabilize and improve those findings.