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The central rift of the Red Sea has 25 brine pools with different physical and geochemical characteristics. Atlantis II (ATIID), Discovery Deeps (DD) and Chain Deep (CD) are characterized by high salinity, temperature and metal content. Several studies reported microbial communities in these brine pools, but few studies addressed the brine pool sediments. Therefore, sediment cores were collected from ATIID, DD, CD brine pools and an adjacent brine-influenced site. Sixteen different lithologic sediment sections were subjected to shotgun DNA pyrosequencing to generate 1.47 billion base pairs (1.47 x 10(9) bp). We generated sediment-specific reads and attempted to annotate all reads. We report the phylogenetic and biochemical uniqueness of the deepest ATIID sulfur-rich brine pool sediments. In contrary to all other sediment sections, bacteria dominate the deepest ATIID sulfur-rich brine pool sediments. This decrease in virus-to-bacteria ratio in selected sections and depth coincided with an overrepresentation of mobile genetic elements. Skewing in the composition of viruses-to-mobile genetic elements may uniquely contribute to the distinct microbial consortium in sediments in proximity to hydrothermally active vents of the Red Sea and possibly in their surroundings, through differential horizontal gene transfer.
Microthlaspi is a predominantly Eurasian genus which also occurs in the northernmost parts of Africa (Maghreb). The most widespread species of the genus is M. perfoliatum, which can be found from Sweden to Algeria and from Portugal to China. The other species are thought to have much more confined distribution ranges, often covering only a few hundred kilometres. This is also believed for the diploid M. erraticum, which was recently re-appraised as a taxon independent from the tetra- to hexaploid M. perfoliatum. Previously, M. erraticum was believed to be present only in Central Europe, from the East of France to Slovenia. In order to gain a deeper understanding of the ecology, evolution and migration history of Microthlaspi it was the focus of the current study to investigate, if M. erraticum is present in habitats outside Central Europe, but with microclimates similar to Central Europe. It is demonstrated that M. erraticum is much more widespread than previously thought, while other lineages apart from M. perfoliatum s.str. and M. erraticum seem to have restricted distribution ranges. The latter species was observed from the Alps and their foreland, the Balkans, the mountainous areas around the Black Sea, Southern Siberia, as well as the Altai and Tien Shan mountains. This demonstrates a widespread occurrence of this easily-overlooked species. (C) 2016 Elsevier GmbH. All rights reserved.
Responses to pathogens, including host transcriptional reprogramming, require partially antagonistic signalling pathways dependent on the phytohormones salicylic (SA) and jasmonic (JA) acids. However, upstream factors modulating the interplay of these pathways are not well characterized. Here, we identify the transcription factor ANAC032 from Arabidopsis thaliana as one such regulator in response to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000 (Pst). ANAC032 directly represses MYC2 activation upon Pst attack, resulting in blockage of coronatine-mediated stomatal reopening which restricts entry of bacteria into plant tissue. Furthermore, ANAC032 activates SA signalling by repressing NIMIN1, a key negative regulator of SA-dependent defence. Finally, ANAC032 reduces expression of JA-responsive genes, including PDF1.2A. Thus, ANAC032 enhances resistance to Pst by generating an orchestrated transcriptional output towards key SA- and JA-signalling genes coordinated through direct binding of ANAC032 to the MYC2, NIMIN1 and PDF1.2A promoters.
Dromedaries have been fundamental to the development of human societies in arid landscapes and for long-distance trade across hostile hot terrains for 3,000 y. Today they continue to be an important livestock resource in marginal agro-ecological zones. However, the history of dromedary domestication and the influence of ancient trading networks on their genetic structure have remained elusive. We combined ancient DNA sequences of wild and early-domesticated dromedary samples from arid regions with nuclear microsatellite and mitochondrial genotype information from 1,083 extant animals collected across the species’ range. We observe little phylogeographic signal in the modern population, indicative of extensive gene flow and virtually affecting all regions except East Africa, where dromedary populations have remained relatively isolated. In agreement with archaeological findings, we identify wild dromedaries from the southeast Arabian Peninsula among the founders of the domestic dromedary gene pool. Approximate Bayesian computations further support the “restocking from the wild” hypothesis, with an initial domestication followed by introgression from individuals from wild, now-extinct populations. Compared with other livestock, which show a long history of gene flow with their wild ancestors, we find a high initial diversity relative to the native distribution of the wild ancestor on the Arabian Peninsula and to the brief coexistence of early-domesticated and wild individuals. This study also demonstrates the potential to retrieve ancient DNA sequences from osseous remains excavated in hot and dry desert environments.
From dark to light
(2016)
Translation is a central cellular process and is optimized for speed and fidelity. The speed of translation of a single codon depends on the concentration of aminoacyl-tRNAs. Here, we used microarray-based approaches to analyze the charging levels of tRNAs in Escherichia coli growing at different growth rates. Strikingly, we observed a non-uniform aminoacylation of tRNAs in complex media. In contrast, in minimal medium, the level of aminoacyl-tRNAs is more uniform and rises to approximately 60%. Particularly, the charging level of tRNA(Ser), tRNA(Cys), tRNA(Thr) and tRNA(His) is below 50% in complex medium and their aminoacylation levels mirror the degree that amino acids inhibit growth when individually added to minimal medium. Serine is among the most toxic amino acids for bacteria and tRNAs(Ser) exhibit the lowest charging levels, below 10%, at high growth rate although intracellular serine concentration is plentiful. As a result some serine codons are among the most slowly translated codons. A large fraction of the serine is most likely degraded by L-serine-deaminase, which competes with the seryl-tRNA-synthetase that charges the tRNAs(Ser). These results indicate that the level of aminoacylation in complex media might be a competition between charging for translation and degradation of amino acids that inhibit growth.
Current rates of environmental change are exceeding the capacity of many populations to adapt to new conditions and thus avoid demographic collapse and ultimate extinction. In particular, cold-water freshwater fish species are predicted to experience strong selective pressure from climate change and a wide range of interacting anthropogenic stressors in the near future. To implement effective management and conservation measures, it is crucial to quantify the maximum rate of change that cold-water freshwater fish populations can withstand. Here, we present a spatially explicit eco-genetic individual-based model, inSTREAM-Gen, to predict the eco-evolutionary dynamics of stream-dwelling trout under anthropogenic environmental change. The model builds on a well-tested demographic model, which includes submodels of river dynamics, bioenergetics, and adaptive habitat selection, with a new genetic module that allows exploration of genetic and life-history adaptations to new environments. The genetic module models the transmission of two key traits, size at emergence and maturity size threshold. We parameterized the model for a brown trout (Salmo trutta L.) population at the warmest edge of its range to validate it and analyze its sensitivity to parameters under contrasting thermal profiles. To illustrate potential applications of the model, we analyzed the population's demographic and evolutionary dynamics under scenarios of (1) climate change-induced warming, and (2) warming plus flow reduction resulting from climate and land use change, compared to (3) a baseline of no environmental change. The model predicted severe declines in density and biomass under climate warming. These declines were lower than expected at range margins because of evolution towards smaller size at both emergence and maturation compared to the natural evolution under the baseline conditions. Despite stronger evolutionary responses, declining rates were substantially larger under the combined warming and flow reduction scenario, leading to a high probability of population extinction over contemporary time frames. Therefore, adaptive responses could not prevent extinction under high rates of environmental change. Our model demonstrates critical elements of next generation ecological modelling aiming at predictions in a changing world as it accounts for spatial and temporal resource heterogeneity, while merging individual behaviour and bioenergetics with microevolutionary adaptations.
OPTIMIZATION OF THE BIOSORPTION OF Cr3+, Cd2+ AND Pb2+ USING A NEW BIOWASTE: Zea mays SEED CHAFF
(2016)
This study highlights the potential use of yellow Zea mays seed chaff (YZMSC) biomass as a biosorbent for the removal of Cr3+, Cd2+ and Pb2+ ions from aqueous solutions. Fourier transformed Infrared analysis of the biomass suggests that YZMSC biomass is basically composed of cellulose and methyl cellulose. The biosorption capacities, q(max), of YZMSC biomass for Cr3+, Cd2+ and Pb2+ are 14.68, 121.95 and 384.62 mg/g respectively. Biosorption equilibrium was achieved at 20, 30 and 60 min for Cr3+, Cd2+ and Pb2+ respectively. YZMSC biomass was found to have higher biosorption capacity and overall kinetic rate of uptake for Pb2+ than for Cd2+ and Cr3+. However, Cr3+ had better initial kinetic rate of uptake by the biomass than Pb2+ and Cd2+. The Freundlich equilibrium isotherm model was found to describe equilibrium data better than Langmuir model suggesting that biosorption of these metal ions could be on more than one active site on the surface of YZMSC biomass. Kinetic study predicted the pseudo-second kinetic model as being able to better describe kinetic data obtained than either modified pseudo-first order or Bangham kinetic models. Biosorption of Cr3+, Cd2+ and Pb2+ onto YZMSC biomass was endothermic in nature with large positive entropy values. Biosorption of these metal ions onto YZMSC biomass was observed to be feasible and spontaneous above 283 K. Optimization of biomass weight for the removal of these metal ions suggest that 384 kg, 129 kg and 144 kg of YZMSC biomass is required for the removal of 95% of Cr3+, Cd2+ and Pb2+ metal ions respectively from 100 mg/L of metal ions in 10 tonnes of aqueous solutions.
Herein we present an efficient synthesis of a biomimetic probe with modular construction that can be specifically bound by the mannose binding FimH protein - a surface adhesion protein of E. coli bacteria. The synthesis combines the new and interesting DBD dye with the carbohydrate ligand mannose via a Click reaction. We demonstrate the binding to E. coli bacteria over a large concentration range and also present some special characteristics of those molecules that are of particular interest for the application as a biosensor. In particular, the mix-and-measure ability and the very good photo-stability should be highlighted here.
Environmental stress is detrimental to cell viability and requires an adequate reprogramming of cellular activities to maximize cell survival. We present a global analysis of the response of Escherichia coli to acute heat and osmotic stress. We combine deep sequencing of total mRNA and ribosome-protected fragments to provide a genome-wide map of the stress response at transcriptional and translational levels. For each type of stress, we observe a unique subset of genes that shape the stress-specific response. Upon temperature upshift, mRNAs with reduced folding stability up-and downstream of the start codon, and thus with more accessible initiation regions, are translationally favoured. Conversely, osmotic upshift causes a global reduction of highly translated transcripts with high copy numbers, allowing reallocation of translation resources to not degraded and newly synthesized mRNAs.
Src1 is a Protein of the Inner Nuclear Membrane Interacting with the Dictyostelium Lamin NE81
(2016)
The nuclear envelope (NE) consists of the outer and inner nuclear membrane (INM), whereby the latter is bound to the nuclear lamina. Src1 is a Dictyostelium homologue of the helix-extension-helix family of proteins, which also includes the human lamin-binding protein MAN1. Both endogenous Src1 and GFP-Src1 are localized to the NE during the entire cell cycle. Immuno-electron microscopy and light microscopy after differential detergent treatment indicated that Src1 resides in the INM. FRAP experiments with GFP-Src1 cells suggested that at least a fraction of the protein could be stably engaged in forming the nuclear lamina together with the Dictyostelium lamin NE81. Both a BioID proximity assay and mis-localization of soluble, truncated mRFP-Src1 at cytosolic clusters consisting of an intentionally mis-localized mutant of GFP-NE81 confirmed an interaction of Src1 and NE81. Expression GFP-Src11–646, a fragment C-terminally truncated after the first transmembrane domain, disrupted interaction of nuclear membranes with the nuclear lamina, as cells formed protrusions of the NE that were dependent on cytoskeletal pulling forces. Protrusions were dependent on intact microtubules but not actin filaments. Our results indicate that Src1 is required for integrity of the NE and highlight Dictyostelium as a promising model for the evolution of nuclear architecture.
Src1 is a Protein of the Inner Nuclear Membrane Interacting with the Dictyostelium Lamin NE81
(2016)
The nuclear envelope (NE) consists of the outer and inner nuclear membrane (INM), whereby the latter is bound to the nuclear lamina. Src1 is a Dictyostelium homologue of the helix-extension-helix family of proteins, which also includes the human lamin-binding protein MAN1. Both endogenous Src1 and GFP-Src1 are localized to the NE during the entire cell cycle. Immuno-electron microscopy and light microscopy after differential detergent treatment indicated that Src1 resides in the INM. FRAP experiments with GFP-Src1 cells suggested that at least a fraction of the protein could be stably engaged in forming the nuclear lamina together with the Dictyostelium lamin NE81. Both a BioID proximity assay and mis-localization of soluble, truncated mRFP-Src1 at cytosolic clusters consisting of an intentionally mis-localized mutant of GFP-NE81 confirmed an interaction of Src1 and NE81. Expression GFP-Src11–646, a fragment C-terminally truncated after the first transmembrane domain, disrupted interaction of nuclear membranes with the nuclear lamina, as cells formed protrusions of the NE that were dependent on cytoskeletal pulling forces. Protrusions were dependent on intact microtubules but not actin filaments. Our results indicate that Src1 is required for integrity of the NE and highlight Dictyostelium as a promising model for the evolution of nuclear architecture.
Individuals within populations often differ substantially in habitat use, the ecological consequences of which can be far reaching. Stable isotope analysis provides a convenient and often cost effective means of indirectly assessing the habitat use of individuals that can yield valuable insights into the spatiotemporal distribution of foraging specialisations within a population. Here we use the stable isotope ratios of southern sea lion (Otaria flavescens) pup vibrissae at the Falkland Islands, in the South Atlantic, as a proxy for adult female habitat use during gestation. A previous study found that adult females from one breeding colony (Big Shag Island) foraged in two discrete habitats, inshore (coastal) or offshore (outer Patagonian Shelf). However, as this species breeds at over 70 sites around the Falkland Islands, it is unclear if this pattern is representative of the Falkland Islands as a whole. In order to characterize habitat use, we therefore assayed carbon (delta C-13) and nitrogen (delta N-15) ratios from 65 southern sea lion pup vibrissae, sampled across 19 breeding colonies at the Falkland Islands. Model-based clustering of pup isotope ratios identified three distinct clusters, representing adult females that foraged inshore, offshore, and a cluster best described as intermediate. A significant difference was found in the use of inshore and offshore habitats between West and East Falkland and between the two colonies with the largest sample sizes, both of which are located in East Falkland. However, habitat use was unrelated to the proximity of breeding colonies to the Patagonian Shelf, a region associated with enhanced biological productivity. Our study thus points towards other factors, such as local oceanography and its influence on resource distribution, playing a prominent role in inshore and offshore habitat use.
A new sequestrate Lactarius species was found in a humid evergreen tropical rainforest dominated by Fabaceae of the subfamily Caesalpinioideae in Cameroon, Central Africa. It is described here as new to science and is named Lactarius megalopterus, referring to its spore ornamentation of extraordinarily high wings. Anatomical characters and molecular systematic analyses confirm its relationship to Lactarius subgenus Plinthogali. Phylogenetic analyses based on two nuclear DNA regions revealed its close relationship to Lactarius angiocarpus, which is also an angiocarpous species from Zambia in Africa. Molecular studies have shown that tuber-like, sequestrate sporocarps evolved independently in several lineages of Basidiomycota. The findings of sequestrate fungi in tropical rainforests raise questions regarding the evolutionary benefit of enclosing the spore-producing hymenium. The enclosure of spore-producing tissue has often been associated with the protection of the delicate hymenium against desiccation in arid habitats or against frost in cold habitats. However, these cannot be the selective factors in warm and humid areas like the tropics. This controversy is exemplarily studied and discussed in the family of Russulaceae, especially in the genus Lactarius. Characters shown by the angiocarpous sporocarp of the new Lactarius, such as thick-walled statismospores, an aromatic smell and mild taste, can be interpreted as adaptations to endozoochorous spore dispersal by mammals. Therefore, here we prefer the alternative hypothesis that sequestrate sporocarps are the result of adaptation to endozoochorous spore dispersal.
Characterization of the Clp protease complex and identification of putative substrates in N. tabacum
(2016)
The interplay between soil structure, roots, and microbiota as a determinant of plant-soil feedback
(2016)
Plant-soil feedback (PSF) can influence plant community structure via changes in the soil microbiome. However, how these feedbacks depend on the soil environment remains poorly understood. We hypothesized that disintegrating a naturally aggregated soil may influence the outcome of PSF by affecting microbial communities. Furthermore, we expected plants to differentially interact with soil structure and the microbial communities due to varying root morphology. We carried out a feedback experiment with nine plant species (five forbs and four grasses) where the training phase consisted of aggregated versus disintegrated soil. In the feedback phase, a uniform soil was inoculated in a fully factorial design with soil washings from conspecific- versus heterospecific-trained soil that had been either disintegrated or aggregated. This way, the effects of prior soil structure on plant performance in terms of biomass production and allocation were examined. In the training phase, soil structure did not affect plant biomass. But on disintegrated soil, plants with lower specific root length (SRL) allocated more biomass aboveground. PSF in the feedback phase was negative overall. With training on disintegrated soil, conspecific feedback was positively correlated with SRL and significantly differed between grasses and forbs. Plants with higher SRL were likely able to easily explore the disintegrated soil with smaller pores, while plants with lower SRL invested in belowground biomass for soil exploration and seemed to be more susceptible to fungal pathogens. This suggests that plants with low SRL could be more limited by PSF on disintegrated soils of early successional stages. This study is the first to examine the influence of soil structure on PSF. Our results suggest that soil structure determines the outcome of PSF mediated by SRL. We recommend to further explore the effects of soil structure and propose to include root performance when working with PSF.
Defining species by their climatic niche is the simple and intuitive principle underlying Bioclimatic Envelope Model (BEM) predictions for climate change effects. However, these correlative models are often criticised for neglecting many ecological processes. Here, we apply the same niche principle to entire communities within a medium/long-term climate manipulation study, where ecological processes are inherently included. In a nine generation study in Israel, we manipulated rainfall (Drought -30%; Irrigation +30%; Control natural rainfall) at two sites which differ chiefly in rainfall quantity and variability. We analysed community responses to the manipulations by grouping species based on their climatic rainfall niche. These responses were compared to analyses based on single species, total densities, and commonly used taxonomic groupings. Climate Niche Groups yielded clear and consistent results: within communities, those species distributed in drier regions performed relatively better in the drought treatment, and those from wetter climates performed better when irrigated. In contrast, analyses based on other principles revealed little insight into community dynamics. Notably, most relationships were weaker at the drier, more variable site, suggesting that enhanced adaptation to variability may buffer climate change impacts. We provide robust experimental evidence that using climatic niches commonly applied in BEMs is a valid approach for eliciting community changes in response to climate change. However, we also argue that additional empirical information needs to be gathered using experiments in situ to correctly assess community vulnerability. Climatic Niche Groups used in this way, may therefore provide a powerful tool and directional testing framework to generalise and compare climate change impacts across habitats. (C) 2016 The Authors. Published by Elsevier GmbH.
Warming and eutrophication are two of the most important global change stressors for natural ecosystems, but their interaction is poorly understood. We used a dynamic model of complex, size-structured food webs to assess interactive effects on diversity and network structure. We found antagonistic impacts: Warming increases diversity in eutrophic systems and decreases it in oligotrophic systems. These effects interact with the community size structure: Communities of similarly sized species such as parasitoid-host systems are stabilized by warming and destabilized by eutrophication, whereas the diversity of size-structured predator-prey networks decreases strongly with warming, but decreases only weakly with eutrophication. Nonrandom extinction risks for generalists and specialists lead to higher connectance in networks without size structure and lower connectance in size-structured communities. Overall, our results unravel interactive impacts of warming and eutrophication and suggest that size structure may serve as an important proxy for predicting the community sensitivity to these global change stressors.
Background/Aims: To analyze the duration of treatment with antipsychotics in German dementia patients. Methods: This study included patients aged 60 years and over with dementia who received a first-time antipsychotic prescription by psychiatrists between 2009 and 2013. The main outcome measure was the treatment rate for more than 6 months following the index date. Results: A total of 12,979 patients with dementia (mean age 82 years, 52.1% living in nursing homes) were included. After 2 years of follow-up, 54.8%, 57.2%, 61.1%, and 65.4% of patients aged 60 - 69, 70 - 79, 80 - 89, and 90 - 99 years, respectively, received antipsychotic prescriptions. 63.9% of subjects living in nursing homes and 55.0% of subjects living at home also continued their treatment (p-value < 0.001). Conclusion: The percentage of dementia patients treated with anti psychotics is very high.
There has been a long-standing controversy regarding the effect of chemical denaturants on the dimensions of unfolded and intrinsically disordered proteins: A wide range of experimental techniques suggest that polypeptide chains expand with increasing denaturant concentration, but several studies using small-angle X-ray scattering (SAXS) have reported no: such increase of the radius of gyration (R-g). This inconsistency challenges our current understanding of the mechanism of chemical denaturants, which are widely employed to investigate protein folding and stability. Here, we use a combination Of single-molecule Forster resonance energy transfer (FRET), SAXS, dynamic light scattering (DLS), and two-focus fluorescence correlation spectroscopy (2f-FCS) to characterize the denaturant dependence of the unfolded state of the spectrin domain R17 and the intrinsically disordered protein ACTR in two different denaturants. Standard analysis of the primary data clearly indicates an expansion of the unfolded state with increasing denaturant concentration irrespective of the protein, denaturant, or experimental method used. This is the first case in which SAXS and FRET have yielded even qualitatively consistent results regarding expansion in denaturant when applied to the same proteins. To more directly illustrate this self-consistency, we used both SAXS and FRET data in a Bayesian procedure to refine structural ensembles representative of the observed unfolded state. This analysis demonstrates that both of these experimental probes are compatible with a common ensemble of protein configurations for each denaturant concentration. Furthermore, the resulting ensembles reproduce the trend of increasing hydrodynamic radius, with denaturant concentration obtained by 2f-FCS,and DLS. We were thus able to reconcile the results from all four experimental techniques quantitatively, to obtain a comprehensive structural picture of denaturant;induced unfolded state expansion, and to identify the Most likely sources of earlier discrepancies.
The use of high-frequency sensors on profiling buoys to investigate physical, chemical, and biological processes in lakes is
increasing rapidly. Profiling buoys with automated winches and sensors that collect high-frequency chlorophyll fluorescence
(ChlF) profiles in 11 lakes in the Global Lake Ecological Observatory Network (GLEON) allowed the study of the vertical
and temporal distribution of ChlF, including the formation of subsurface chlorophyll maxima (SSCM). The effectiveness of 3
methods for sampling phytoplankton distributions in lakes, including (1) manual profiles, (2) single-depth buoys, and (3)
profiling buoys were assessed. High-frequency ChlF surface data and profiles were compared to predictions from the
Plankton Ecology Group (PEG) model. The depth-integrated ChlF dynamics measured by the profiling buoy data revealed a
greater complexity that neither conventional sampling nor the generalized PEG model captured. Conventional sampling
techniques would have missed SSCM in 7 of 11 study lakes. Although surface-only ChlF data underestimated average water
column ChlF, at times by nearly 2-fold in 4 of the lakes, overall there was a remarkable similarity between surface and mean
water column data. Contrary to the PEG model’s proposed negligible role for physical control of phytoplankton during the
growing season, thermal structure and light availability were closely associated with ChlF seasonal depth distribution. Thus,
an extension of the PEG model is proposed, with a new conceptual framework that explicitly includes physical metrics to
better predict SSCM formation in lakes and highlight when profiling buoys are especially informative.
The cytoskeleton is an essential component of living cells. It is composed of different types of protein filaments that form complex, dynamically rearranging, and interconnected networks. The cytoskeleton serves a multitude of cellular functions which further depend on the cell context. In animal cells, the cytoskeleton prominently shapes the cell's mechanical properties and movement. In plant cells, in contrast, the presence of a rigid cell wall as well as their larger sizes highlight the role of the cytoskeleton in long-distance intracellular transport. As it provides the basis for cell growth and biomass production, cytoskeletal transport in plant cells is of direct environmental and economical relevance. However, while knowledge about the molecular details of the cytoskeletal transport is growing rapidly, the organizational principles that shape these processes on a whole-cell level remain elusive.
This thesis is devoted to the following question: How does the complex architecture of the plant cytoskeleton relate to its transport functionality? The answer requires a systems level perspective of plant cytoskeletal structure and transport. To this end, I combined state-of-the-art confocal microscopy, quantitative digital image analysis, and mathematically powerful, intuitively accessible graph-theoretical approaches.
This thesis summarizes five of my publications that shed light on the plant cytoskeleton as a transportation network: (1) I developed network-based frameworks for accurate, automated quantification of cytoskeletal structures, applicable in, e.g., genetic or chemical screens; (2) I showed that the actin cytoskeleton displays properties of efficient transport networks, hinting at its biological design principles; (3) Using multi-objective optimization, I demonstrated that different plant cell types sustain cytoskeletal networks with cell-type specific and near-optimal organization; (4) By investigating actual transport of organelles through the cell, I showed that properties of the actin cytoskeleton are predictive of organelle flow and provided quantitative evidence for a coordination of transport at a cellular level; (5) I devised a robust, optimization-based method to identify individual cytoskeletal filaments from a given network representation, allowing the investigation of single filament properties in the network context. The developed methods were made publicly available as open-source software tools.
Altogether, my findings and proposed frameworks provide quantitative, system-level insights into intracellular transport in living cells. Despite my focus on the plant cytoskeleton, the established combination of experimental and theoretical approaches is readily applicable to different organisms. Despite the necessity of detailed molecular studies, only a complementary, systemic perspective, as presented here, enables both understanding of cytoskeletal function in its evolutionary context as well as its future technological control and utilization.
Arabidopsis FORGETTER1 mediates stress-induced chromatin memory through nucleosome remodeling
(2016)
Plants as sessile organisms can adapt to environmental stress to mitigate its adverse effects. As part of such adaptation they maintain an active memory of heat stress for several days that promotes a more efficient response to recurring stress. We show that this heat stress memory requires the activity of the FORGETTER1 (FGT1) locus, with fgt1 mutants displaying reduced maintenance of heat-induced gene expression. FGT1 encodes the Arabidopsis thaliana orthologue of Strawberry notch (Sno), and the protein globally associates with the promoter regions of actively expressed genes in a heat-dependent fashion. FGT1 interacts with chromatin remodelers of the SWI/ SNF and ISWI families, which also display reduced heat stress memory. Genomic targets of the BRM remodeler overlap significantly with FGT1 targets. Accordingly, nucleosome dynamics at loci with altered maintenance of heat-induced expression are affected in fgt1. Together, our results suggest that by modulating nucleosome occupancy, FGT1 mediates stress-induced chromatin memory.
Investigation of novel proteins and polysaccharides associated with coccoliths of Emiliania huxleyi
(2016)
Aerobic Anoxygenic Phototrophic (AAP) bacteria are bacteriochlorophyll (BChl) a -containing organisms which use light energy to supplement their predominantly heterotrophic metabolism. Here, we investigated mortality and growth rates of AAP bacteria in three different freshwater lakes in Central Europe: the mountain lake Plesne, the oligo-mesotrophic Lake Stechlin and the forest pond Huntov. The mortality of AAP bacteria was estimated from diel changes of BChl a fluorescence. Net and gross growth rates were calculated from the increases in AAP cell numbers. The gross growth rates of AAP bacteria ranged from 0.38 to 5.6 d(-1), with the highest values observed during summer months. Simultaneously, the rapidly growing AAP cells have to cope with an intense grazing pressure by both zooplankton and protists. The presented results document that during the day, gross growth usually surpased mortality. Our results indicate that AAP bacteria utilize light energy under natural conditions to maintain rapid growth rates, which are balanced by a generally intense grazing pressure.
Seit der Einführung von Antibiotika in die medizinische Behandlung von bakteriellen Infektionskrankheiten existiert ein Wettlauf zwischen der Evolution von Bakterienresistenzen und der Entwicklung wirksamer Antibiotika. Während bis in die 80er Jahre verstärkt an neuen Antibiotika geforscht wurde, gewinnen multiresistente Keime heute zunehmend die Oberhand. Um einzelne Pathogene erfolgreich nachzuweisen und zu bekämpfen, ist ein grundlegendes Wissen über den Erreger unumgänglich. Bakterielle Proteine, die bei einer Infektion vorrangig vom Immunsystem prozessiert und präsentiert werden, könnten für die Entwicklung von Impfstoffen oder gezielten Therapeutika nützlich sein. Auch für die Diagnostik wären diese immundominanten Proteine interessant. Allerdings herrscht ein Mangel an Wissen über spezifische Antigene vieler pathogener Bakterien, die eine eindeutige Diagnostik eines einzelnen Erregers erlauben würden.
Daher wurden in dieser Arbeit vier verschiedene Humanpathogene mittels Phage Display untersucht: Neisseria gonorrhoeae, Neisseria meningitidis, Borrelia burgdorferi und Clostridium difficile. Hierfür wurden aus der genomischen DNA der vier Erreger Bibliotheken konstruiert und durch wiederholte Selektion und Amplifikation, dem sogenannten Panning, immunogene Proteine isoliert. Für alle Erreger bis auf C. difficile wurden immunogene Proteine aus den jeweiligen Bibliotheken isoliert. Die identifizierten Proteine von N. meningitidis und B. burgdorferi waren größtenteils bekannt, konnten aber in dieser Arbeit durch Phage Display verifiziert werden. Für N. gonorrhoeae wurden 21 potentiell immunogene Oligopeptide isoliert, von denen sechs Proteine als neue zuvor unbeschriebene Proteine mit immunogenem Charakter identifiziert wurden. Von den Phagen-präsentierten Oligopeptide der 21 immunogenen Proteine wurden Epitopmappings mit verschiedenen polyklonalen Antikörpern durchgeführt, um immunogene Bereiche näher zu identifizieren und zu charakterisieren. Bei zehn Proteinen wurden lineare Epitope eindeutig mit drei polyklonalen Antikörpern identifiziert, von fünf weiteren Proteinen waren Epitope mit mindestens einem Antikörper detektierbar. Für eine weitere Charakterisierung der ermittelten Epitope wurden Alaninscans durchgeführt, die eine detaillierte Auskunft über kritische Aminosäuren für die Bindung des Antikörpers an das Epitop geben.
Ausgehend von dem neu identifizierten Protein mit immunogenem Charakter NGO1634 wurden 26 weitere Proteine aufgrund ihrer funktionellen Ähnlichkeit ausgewählt und mithilfe bioinformatischer Analysen auf ihre Eignung zur Entwicklung einer diagnostischen Anwendung analysiert. Durch Ausschluss der meisten Proteine aufgrund ihrer Lokalisation, Membrantopologie oder unspezifischen Proteinsequenz wurden scFv-Antikörper gegen acht Proteine mittels Phage Display generiert und anschließend als scFv-Fc-Fusionsantikörper produziert und charakterisiert.
Die hier identifizierten Proteine und linearen Epitope könnten einen Ansatzpunkt für die Entwicklung einer diagnostischen oder therapeutischen Anwendung bieten. Lineare Epitopsequenzen werden häufig für die Impfstoffentwicklung eingesetzt, sodass vor allem die in dieser Arbeit bestimmten Epitope von Membranproteinen interessante Kandidaten für weitere Untersuchungen in diese Richtung sind. Durch weitere Untersuchungen könnten möglicherweise unbekannte Virulenzfaktoren entdeckt werden, deren Inhibierung einen entscheidenden Einfluss auf Infektionen haben könnten.
Neisseria gonorrhoeae is one of the most prevalent sexually transmitted diseases worldwide with more than 100 million new infections per year. A lack of intense research over the last decades and increasing resistances to the recommended antibiotics call for a better understanding of gonococcal infection, fast diagnostics and therapeutic measures against N. gonorrhoeae. Therefore, the aim of this work was to identify novel immunogenic proteins as a first step to advance those unresolved problems. For the identification of immunogenic proteins, pHORF oligopeptide phage display libraries of the entire N. gonorrhoeae genome were constructed. Several immunogenic oligopeptides were identified using polyclonal rabbit antibodies against N. gonorrhoeae. Corresponding full-length proteins of the identified oligopeptides were expressed and their immunogenic character was verified by ELISA. The immunogenic character of six proteins was identified for the first time. Additional 13 proteins were verified as immunogenic proteins in N. gonorrhoeae.
The xanthine oxidase (XO) family comprises molybdenum-dependent enzymes that usually form homodimers (or dimers of heterodimers/trimers) organized in three domains that harbor two [2Fe-2S] clusters, one FAD, and a Mo cofactor. In this work, we crystallized an unusual member of the family, the periplasmic aldehyde oxidoreductase PaoABC from Escherichia coli. This is the first example of an E. coli protein containing a molybdopterin-cytosine-dinucleotide cofactor and is the only heterotrimer of the XO family so far structurally characterized. The crystal structure revealed the presence of an unexpected [4Fe-4S] cluster, anchored to an additional 40 residues subdomain. According to phylogenetic analysis, proteins containing this cluster are widely spread in many bacteria phyla, putatively through repeated gene transfer events. The active site of PaoABC is highly exposed to the surface with no aromatic residues and an arginine (PaoC-R440) making a direct interaction with PaoC-E692, which acts as a base catalyst. In order to understand the importance of R440, kinetic assays were carried out, and the crystal structure of the PaoC-R440H variant was also determined.
Functionally diverse communities can adjust their species composition to altered environmental conditions, which may influence food web dynamics. Trait-based aggregate models cope with this complexity by ignoring details about species identities and focusing on their functional characteristics (traits). They describe the temporal changes of the aggregate properties of entire communities, including their total biomasses, mean trait values, and trait variances. The applicability of aggregate models depends on the validity of their underlying assumptions that trait distributions are normal and exhibit small variances. We investigated to what extent this can be expected to work by comparing an innovative model that accounts for the full trait distributions of predator and prey communities to a corresponding aggregate model. We used a food web structure with well-established trade-offs among traits promoting mutual adjustments between prey edibility and predator selectivity in response to selection. We altered the shape of the trade-offs to compare the outcome of the two models under different selection regimes, leading to trait distributions increasingly deviating from normality. Their biomass and trait dynamics agreed very well for stabilizing selection and reasonably well for directional selection, under which different trait values are favored at different times. However, for disruptive selection, the results of the aggregate model strongly deviated from the full trait distribution model that showed bimodal trait distributions with large variances. Hence, the outcome of aggregate models is reliable under ideal conditions but has to be questioned when confronted with more complex selection regimes and trait distributions, which are commonly observed in nature.
Marine habitats harbor a great diversity of microorganism from the three domains of life, only a small fraction of which can be cultivated. Metagenomic approaches are increasingly popular for addressing microbial diversity without culture, serving as sensitive and relatively unbiased methods for identifying and cataloging the diversity of nucleic acid sequences derived from organisms in environmental samples. Aerobic anoxygenic phototrophic bacteria (AAP) play important roles in carbon and energy cycling in aquatic systems. In oceans, those bacteria are widely distributed; however, their abundance and importance are still poorly understood. The aim of this study was to estimate abundance and diversity of AAPs in metagenomes from an upwelling affected coastal bay in Arraial do Cabo, Brazil, using in silico screening for the anoxygenic photosynthesis core genes. Metagenomes from the Global Ocean Sample Expedition (GOS) were screened for comparative purposes. AAPs were highly abundant in the free-living bacterial fraction from Arraial do Cabo: 23.88% of total bacterial cells, compared with 15% in the GOS dataset. Of the ten most AAP abundant samples from GOS, eight were collected close to the Equator where solar irradiation is high year-round. We were able to assign most retrieved sequences to phylo-groups, with a particularly high abundance of Roseobacter in Arraial do Cabo samples. The high abundance of AAP in this tropical bay may be related to the upwelling phenomenon and subsequent picoplankton bloom. These results suggest a link between upwelling and light abundance and demonstrate AAP even in oligotrophic tropical and subtropical environments. Longitudinal studies in the Arraial do Cabo region are warranted to understand the dynamics of AAP at different locations and seasons, and the ecological role of these unique bacteria for biogeochemical and energy cycling in the ocean.
DPF3 (BAF45c) is a member of the BAF chromatin remodeling complex. Two isoforms have been described, namely DPF3a and DPF3b. The latter binds to acetylated and methylated lysine residues of histones. Here, we elaborate on the role of DPF3a and describe a novel pathway of cardiac gene transcription leading to pathological cardiac hypertrophy. Upon hypertrophic stimuli, casein kinase 2 phosphorylates DPF3a at serine 348. This initiates the interaction of DPF3a with the transcriptional repressors HEY, followed by the release of HEY from the DNA. Moreover, BRG1 is bound by DPF3a, and is thus recruited to HEY genomic targets upon interaction of the two components. Consequently, the transcription of downstream targets such as NPPA and GATA4 is initiated and pathological cardiac hypertrophy is established. In human, DPF3a is significantly up-regulated in hypertrophic hearts of patients with hypertrophic cardiomyopathy or aortic stenosis. Taken together, we show that activation of DPF3a upon hypertrophic stimuli switches cardiac fetal gene expression from being silenced by HEY to being activated by BRG1. Thus, we present a novel pathway for pathological cardiac hypertrophy, whose inhibition is a long-term therapeutic goal for the treatment of the course of heart failure.
Background. Retroviral integration into the host germline results in permanent viral colonization of vertebrate genomes. The koala retrovirus (KoRV) is currently invading the germline of the koala (Phascolarctos cinereus) and provides a unique opportunity for studying retroviral endogenization. Previous analysis of KoRV integration patterns in modern koalas demonstrate that they share integration sites primarily if they are related, indicating that the process is currently driven by vertical transmission rather than infection. However, due to methodological challenges, KoRV integrations have not been comprehensively characterized. Results. To overcome these challenges, we applied and compared three target enrichment techniques coupled with next generation sequencing (NGS) and a newly customized sequence-clustering based computational pipeline to determine the integration sites for 10 museum Queensland and New South Wales (NSW) koala samples collected between the 1870s and late 1980s. A secondary aim of this study sought to identify common integration sites across modern and historical specimens by comparing our dataset to previously published studies. Several million sequences were processed, and the KoRV integration sites in each koala were characterized. Conclusions. Although the three enrichment methods each exhibited bias in integration site retrieval, a combination of two methods, Primer Extension Capture and hybridization capture is recommended for future studies on historical samples. Moreover, identification of integration sites shows that the proportion of integration sites shared between any two koalas is quite small.
Heterostyly is a wide-spread floral adaptation to promote outbreeding, yet its genetic basis and evolutionary origin remain poorly understood. In Primula (primroses), heterostyly is controlled by the S-locus supergene that determines the reciprocal arrangement of reproductive organs and incompatibility between the two morphs. However, the identities of the component genes remain unknown. Here, we identify the Primula CYP734A50 gene, encoding a putative brassinosteroid-degrading enzyme, as the G locus that determines the style-length dimorphism. CYP734A50 is only present on the short-styled S-morph haplotype, it is specifically expressed in S-morph styles, and its loss or inactivation leads to long styles. The gene arose by a duplication specific to the Primulaceae lineage and shows an accelerated rate of molecular evolution. Thus, our results provide a mechanistic explanation for the Primula style-length dimorphism and begin to shed light on the evolution of the S-locus as a prime model for a complex plant supergene.
Polyadenylation is a critical 3-end processing step during maturation of pre-mRNAs, and the length of the poly(A) tail affects mRNA stability, nuclear export and translation efficiency. The Arabidopsis thaliana genome encodes three canonical nuclear poly(A) polymerase (PAPS) isoforms fulfilling specialized functions, as reflected by their different mutant phenotypes. While PAPS1 affects several processes, such as the immune response, organ growth and male gametophyte development, the roles of PAPS2 and PAPS4 are largely unknown. Here we demonstrate that PAPS2 and PAPS4 promote flowering in a partially redundant manner. The enzymes act antagonistically to PAPS1, which delays the transition to flowering. The opposite flowering-time phenotypes in paps1 and paps2 paps4 mutants are at least partly due to decreased or increased FLC activity, respectively. In contrast to paps2 paps4 mutants, plants with increased PAPS4 activity flower earlier than the wild-type, concomitant with reduced FLC expression. Double mutant analyses suggest that PAPS2 and PAPS4 act independently of the autonomous pathway components FCA, FY and CstF64. The direct polyadenylation targets of the three PAPS isoforms that mediate their effects on flowering time do not include FLC sense mRNA and remain to be identified. Thus, our results uncover a role for canonical PAPS isoforms in flowering-time control, raising the possibility that modulating the balance of the isoform activities could be used to fine tune the transition to flowering. Significance Statement The length of the poly(A) tail affects mRNA stability, nuclear export and translation efficiency. Arabidopsis has three isoforms of nuclear poly(A) polymerase (PAPS): PAPS1 plays a major role in organ growth and plant defence. Here we show that PAPS2 and PAPS4 redundantly promote flowering and act antagonistically to PAPS1, which delays flowering. We suggest that modulating the activity of these isoforms fine-tunes the transition to flowering.
Wastewater samples from a Swedish chemi-thermo-mechanical pulp (CTMP) mill collected at different purification stages in a wastewater treatment plant (WWTP) were analyzed with an amperometric enzyme-based biosensor array in a flow-injection system. In order to resolve the complex composition of the wastewater, the array consists of several sensing elements which yield a multidimensional response. We used principal component analysis (PCA) to decompose the array's responses, and found that wastewater with different degrees of pollution can be differentiated. With the help of partial least squares regression (PLS-R), we could link the sensor responses to the toxicity parameter, as well as to global organic pollution parameters (COD, BOD, and TOC). From investigating the influences of individual sensors in the array, it was found that the best models were in most cases obtained when all sensors in the array were included in the PLS-R model. We find that fast simultaneous determination of several global environmental parameters characterizing wastewaters is possible with this kind of biosensor array, in particular because of the link between the sensor responses and the biological effect onto the ecosystem into which the wastewater would be released. In conjunction with multivariate data analysis tools, there is strong potential to reduce the total time until a result is yielded from days to a few minutes.
The vascular system is critical for developmental growth, tissue homeostasis and repair but also for tumor development. Bone morphogenetic protein (BMP) signaling has recently emerged as a fundamental pathway of the endothelium by regulating cardiovascular and lymphatic development and by being causative for several vascular dysfunctions. Two vascular disorders have been directly linked to impaired BMP signaling: pulmonary arterial hypertension and hereditary hemorrhagic telangiectasia. Endothelial BMP signaling critically depends on the cellular context, which includes among others vascular heterogeneity, exposure to flow, and the intertwining with other signaling cascades (Notch, WNT, Hippo and hypoxia). The purpose of this review is to highlight the most recent findings illustrating the clear need for reconsidering the role of BMPs in vascular biology. (C) 2015 Elsevier Ltd. All rights reserved.
ATP production requires the establishment of an electrochemical proton gradient across the inner mitochondrial membrane. Mitochondrial uncouplers dissipate this proton gradient and disrupt numerous cellular processes, including vesicular trafficking, mainly through energy depletion. Here we show that Endosidin9 (ES9), a novel mitochondrial uncoupler, is a potent inhibitor of clathrin-mediated endocytosis (CME) in different systems and that ES9 induces inhibition of CME not because of its effect on cellular ATP, but rather due to its protonophore activity that leads to cytoplasm acidification. We show that the known tyrosine kinase inhibitor tyrphostinA23, which is routinely used to block CME, displays similar properties, thus questioning its use as a specific inhibitor of cargo recognition by the AP-2 adaptor complex via tyrosine motif-based endocytosis signals. Furthermore, we show that cytoplasm acidification dramatically affects the dynamics and recruitment of clathrin and associated adaptors, and leads to reduction of phosphatidylinositol 4,5-biphosphate from the plasma membrane.
We compiled global occurrence data sets of 13 congeneric sexual and apomictic species pairs, and used principal components analysis (PCA) and kernel smoothers to compare changes in climatic niche optima, breadths and unfilling/expansion between native and alien ranges. Niche change metrics were compared between sexual and apomictic species. All 26 species showed changes in niche optima and/or breadth and 14 species significantly expanded their climatic niches. However, we found no effect of the reproductive system on niche dynamics. Instead, species with narrower native niches showed higher rates of niche expansion in the alien ranges. Our results suggest that niche shifts are frequent in plant invasions but evolutionary potential may not be of major importance for such shifts. Niche dynamics rather appear to be driven by changes of the realized niche without adaptive change of the fundamental climatic niche.
Microelectrodes modified with electropolymerized plumbagin (PLG) were used for the generation of superoxide radical (O-2(center dot-)) and hydrogen peroxide (H2O2) during oxygen reduction reaction (ORR) in an aqueous medium, specifically in serum-free cell culture media. This is enabled by the specific design of a polymer film on the microelectrode. The generation and diffusion of O-2(center dot-) during electrocatalytic ORR at a positionable PLG polymer-modified microelectrode was followed by fluorescence microscopy with the selective dye 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) and by amperometric detection using a cytochrome c-modified electrode at + 0.13 V. H2O2 production, either by direct oxygen reduction or as product of O-2(center dot-) disproportionation, was monitored by the reaction with Amplex UltraRed. The PLG polymer-modified microelectrodes were used to expose mammalian B6-RPE07 retinal cells to defined local fluxes of reactive oxygen species (ROS), and cellular responses and morphological alterations were observed. The use of a controllable source of ROS opens many possibilities to study how living cells respond to the presence of a certain flux of specific ROS.
The extent of gene flow during the range expansion of non-native species influences the amount of genetic diversity retained in expanding populations. Here, we analyse the population genetic structure of the raccoon dog (Nyctereutes procyonoides) in north-eastern and central Europe. This invasive species is of management concern because it is highly susceptible to fox rabies and an important secondary host of the virus. We hypothesized that the large number of introduced animals and the species’ dispersal capabilities led to high population connectivity and maintenance of genetic diversity throughout the invaded range. We genotyped 332 tissue samples from seven European countries using 16 microsatellite loci. Different algorithms identified three genetic clusters corresponding to Finland, Denmark and a large ‘central’ population that reached from introduction areas in western Russia to northern Germany. Cluster assignments provided evidence of long-distance dispersal. The results of an Approximate Bayesian Computation analysis supported a scenario of equal effective population sizes among different pre-defined populations in the large central cluster. Our results are in line with strong gene flow and secondary admixture between neighbouring demes leading to reduced genetic structuring, probably a result of its fairly rapid population expansion after introduction. The results presented here are remarkable in the sense that we identified a homogenous genetic cluster inhabiting an area stretching over more than 1500km. They are also relevant for disease management, as in the event of a significant rabies outbreak, there is a great risk of a rapid virus spread among raccoon dog populations.