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QuestionThe empirical evidence of root herbivory effects on plant community composition and co-existence is contradictory. This originates from difficulties connected to below-ground research and confinement of experimental studies to a small range of environmental conditions. Here we suggest coupling experimental data with an individual-based model to overcome the limitations inherent in either approach. To demonstrate this, we investigated the consequences of root herbivory, as experimentally observed on individual plants, on plant competition and co-existence in a population and community context under different root herbivory intensities (RHI), fluctuating and constant root herbivore activity and grazing along a resource gradient. LocationBerlin, Germany, glasshouse; Potsdam, Germany, high performance cluster computer. MethodsThe well-established community model IBC-Grass was adapted to allow for a flexible species parameterization and to include annual species. Experimentally observed root herbivory effects on performance of eight common grassland plant species were incorporated into the model by altering plant growth rates. We then determined root herbivore effects on plant populations, competitive hierarchy and consequences for co-existence and community diversity. ResultsRoot herbivory reduced individual biomass, but temporal fluctuation allowed for compensation of herbivore effects. Reducing resource availability strongly shifted competitive hierarchies, with, however, more similar hierarchies along the gradient under root herbivory, pointing to reduced ecological species differences. Consequently, negative effects on co-existence and diversity prevailed, with the exception of a few positive effects on co-existence of selected species pairs. Temporal fluctuation alleviated but did not remove negative root herbivore effects, despite of the stronger influence of intra- compared to interspecific competition. Grazing in general augmented co-existence. Most interestingly, grazing interacted with RHI and resource availability by promoting positive effects of root herbivory. ConclusionsThrough integrating experimental data on the scale of individual plants with a simulation model we verified that root herbivory could affect plant competition with consequences for species co-existence. Our approach demonstrates the benefit that accrues when empirical and modelling approaches are brought more closely together, and that gathering data on distinct processes and under specific conditions, combined with appropriate models, can be used to answer challenging research questions in a more general way.
The spherical vaterite CaCO3 microcrystals are nowadays widely used as sacrificial templates for fabrication of various microcarriers made of biopolymers (e.g., proteins, nucleic acids, enzymes) due to porous structure and mild template elimination conditions. Here, we demonstrated for the first time that polymer microcarriers with tuned internal nanoarchitecture can be designed by employing the CaCO3 crystals of controlled porosity. The layer-by-layer deposition has been utilized to assemble shell-like (hollow) and matrix-like (filled) polymer capsules due to restricted and free polymer diffusion through the crystal pores, respectively. The crystal pore size in the range of few tens of nanometers can be adjusted without any additives by variation of the crystal preparation temperature in the range 745 degrees C. The temperature-mediated growth mechanism is explained by the Ostwald ripening of nanocrystallites forming the crystal secondary structure. Various techniques including SEM, AFM, CLSM, Raman microscopy, nitrogen adsorptiondesorption, and XRD have been employed for crystal and microcapsule analysis. A three-dimensional model is introduced to describe the crystal internal structure and predict the pore cutoff and available surface for the pore diffusing molecules. Inherent biocompatibility of CaCO3 and a possibility to scale the porosity in the size range of typical biomacromolecules make the CaCO3 crystals extremely attractive tools for template assisted designing tailor-made biopolymer-based architectures in 2D to 3D targeted at drug delivery and other bioapplications.
Citizen science has been gaining momentum in the United States and Europe, where citizens are literate and often interested in science. However, in developing countries, which have a dire need for environmental data, such programs are slow to emerge, despite the large and untapped human resources in close proximity to areas of high biodiversity and poorly known floras and faunas. Thus, we propose that the parataxonomist and paraecologist approach, which originates from citizen-based science, is well suited to rural areas in developing countries. Being a paraecologist or a parataxonomist is a vocation and entails full-time employment underpinned by extensive training, whereas citizen science involves the temporary engagement of volunteers. Both approaches have their merits depending on the context and objectives of the research. We examined 4 ongoing paraecologist or parataxonomist programs in Costa Rica, India, Papua New Guinea, and southern Africa and compared their origins, long-term objectives, implementation strategies, activities, key challenges, achievements, and implications for resident communities. The programs supported ongoing research on biodiversity assessment, monitoring, and management, and participants engaged in non-academic capacity development in these fields. The programs in Southern Africa related to specific projects, whereas the programs in Costa Rica, India, and Papua New Guinea were designed for the long term, provided sufficient funding was available. The main focus of the paraecologists’ and parataxonomists’ activities ranged from collection and processing of specimens (Costa Rica and Papua New Guinea) or of socioeconomic and natural science data (India and Southern Africa) to communication between scientists and residents (India and Southern Africa). As members of both the local land user and research communities, paraecologists and parataxonomists can greatly improve the flow of biodiversity information to all users, from local stakeholders to international academia.
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.
Achieving cooperation in natural resource management is always a challenge when incentives exist for an individual to maximise her short term benefits at the cost of a group. We study a public good social dilemma in water infrastructure provision on land reform farms in Namibia. In the context of the Namibian land reform, arbitrarily mixed groups of livestock farmers have to share the operation and maintenance of water infrastructure. Typically, water is mainly used for livestock production, and livestock numbers are subject to high fluctuations due to the given environmental conditions. Our paper assesses how alternative payment systems with differing congruence of provision and appropriation support the cooperation in the group given the ever-changing equilibria. In a first step, we conducted an exploratory overview of the social-ecological system of central Namibian land reform projects. The Social Ecological System (SES) Framework served as a guideline for this assessment (Ostrom 2009). Taking the complexity of the cooperation situation into account, in the second step we designed a role-play that is based on a social-ecological simulation model. The role-play simulates the real-life decision situations of land reform beneficiaries wherein equilibria are permanently changing. This approach helped us to not only better understand the cooperation challenges of Namibian land reform beneficiaries, but also supported stakeholders in their decision making and institution building. Our study provides evidence to support that land reform beneficiaries increase their contributions as they own more livestock and as other group members increase their payments. Nevertheless, only groups with relatively homogeneous livestock endowments manage to agree on payment rules. Interestingly, the dominant rule is an "equal payment per farmer" and not a "payment per head of livestock", though the latter would imply a higher congruence of provision and appropriation.
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.
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.
Null mutations in the human IQCB1/NPHP5 (nephrocystin-5) gene that encodes NPHP5 are the most frequent cause of Senior-LOken syndrome, a ciliopathy that is characterized by Leber congenital amaurosis and nephronophthisis. We generated germline Nphp5-knockout mice by placing a -Geo gene trap in intron 4, thereby truncating NPHP5 at Leu87 and removing all known functional domains. At eye opening, Nphp5(-/-) mice exhibited absence of scotopic and photopic electroretinogram responses, a phenotype that resembles Leber congenital amaurosis. Outer segment transmembrane protein accumulation in Nphp5(-/-) endoplasmic reticulum was evident as early as postnatal day (P)6. EGFP-CETN2, a centrosome and transition zone marker, identified basal bodies in Nphp5(-/-) photoreceptors, but without fully developed transition zones. Ultrastructure of P6 and 10 Nphp5(-/-) photoreceptors revealed aberrant transition zones of reduced diameter. Nphp5(-/-) photoreceptor degeneration was complete at 1 mo of age but was delayed significantly in Nphp5(-/-);Nrl(-/-) (cone only) retina. Nphp5(-/-) mouse embryonic fibroblast developed normal cilia, and Nphp5(-/-) kidney histology at 1 yr of age showed no significant pathology. Results establish that nephrocystin-5 is essential for photoreceptor outer segment formation but is dispensable for kidney and mouse embryonic fibroblast ciliary formation.
Synthetic biology aims at designing and engineering organisms. The engineering process typically requires the establishment of suitable DNA constructs generated through fusion of multiple protein coding and regulatory sequences. Conventional cloning techniques, including those involving restriction enzymes and ligases, are often of limited scope, in particular when many DNA fragments must be joined or scar-free fusions are mandatory. Overlap-based-cloning methods have the potential to overcome such limitations. One such method uses seamless ligation cloning extract (SLiCE) prepared from Escherichia coli cells for straightforward and efficient in vitro fusion of DNA fragments. Here, we systematically characterized extracts prepared from the unmodified E. coli strain DH10B for SLiCE-mediated cloning and determined DNA sequence-associated parameters that affect cloning efficiency. Our data revealed the virtual absence of length restrictions for vector backbone (up to 13.5 kbp) and insert (90 bp to 1.6 kbp). Furthermore, differences in GC content in homology regions are easily tolerated and the deletion of unwanted vector sequences concomitant with targeted fragment insertion is straightforward. Thus, SLiCE represents a highly versatile DNA fusion method suitable for cloning projects in virtually all molecular. and synthetic biology projects. (C) 2016 Elsevier Inc. All rights reserved.
Characterization of the Clp protease complex and identification of putative substrates in N. tabacum
(2016)
Monoclonal antibodies are highly valuable tools in biomedicine but the generation by hybridoma technology is very time-consuming and elaborate. In order to circumvent the consisting drawbacks an in vitro immunization approach was established by which murine as well as human monoclonal antibodies against a viral coat protein could be developed. The in vitro immunization process was performed by isolation of murine hematopoietic stem cells or human monocytes and an in vitro differentiation into immature dendritic cells. After antigen loading the cells were co-cultivated with naive T and B lymphocytes for three days in order to obtain antigen-specific B lymphocytes in culture, followed by fusion with murine myeloma cells or human/murine heteromyeloma cells. Antigen-specific hybridomas were selected and the generated antibodies were purified and characterized in this study by ELISA, western blot, gene sequencing, affinity measurements. Further the characteristics were compared to a monoclonal antibody against the same target generated by conventional hybridoma technology. Isotype detection revealed a murine IgM and a human IgG4 antibody in comparison to an IgG1 for the conventionally generated antibody. The antibodies derived from in vitro immunization showed indeed a lower affinity for the antigen as compared to the conventionally generated one, which is probably based on the significantly shorter B cell maturation (3 days) during the immunization process. Nevertheless, they were suitable for building up a sandwich based detection system. Therefore, the in vitro immunization approach seems to be a good and particularly fast alternative to conventional hybridoma technology.
Carbon and nutrient cycling in kettle hole sediments depending on hydrological dynamics: a review
(2016)
Kettle holes as a specific group of isolated, small lentic freshwater systems (LFS) often are (i) hot spots of biogeochemical cycling and (ii) exposed to frequent sediment desiccation and rewetting. Their ecological functioning is greatly determined by immanent carbon and nutrient transformations. The objective of this review is to elucidate effects of a changing hydrological regime (i.e., dry-wet cycles) on carbon and nutrient cycling in kettle hole sediments. Generally, dry-wet cycles have the potential to increase C and N losses as well as P availability. However, their duration and frequency are important controlling factors regarding direction and intensity of biogeochemical and microbiological responses. To evaluate drought impacts on sediment carbon and nutrient cycling in detail requires the context of the LFS hydrological history. For example, frequent drought events induce physiological adaptation of exposed microbial communities and thus flatten metabolic responses, whereas rare events provoke unbalanced, strong microbial responses. Different potential of microbial resilience to drought stress can irretrievably change microbial communities and functional guilds, gearing cascades of functional responses. Hence, dry-wet events can shift the biogeochemical cycling of organic matter and nutrients to a new equilibrium, thus affecting the dynamic balance between carbon burial and mineralization in kettle holes.
Pancreatic secretory zymogen-granule membrane glycoprotein 2 (GP2) has been identified to be a major autoantigenic target in Crohn’s disease patients. It was discussed recently that a long and a short isoform of GP2 exists whereas the short isoform is often detected by GP2-specific autoantibodies. In the outcome of inflammatory bowel diseases, these GP2-specific autoantibodies are discussed as new serological markers for diagnosis and therapeutic monitoring. To investigate this further, camelid nanobodies were generated by phage display and selected against the short isoform of GP2 in order to isolate specific tools for the discrimination of both isoforms. Nanobodies are single domain antibodies derived from camelid heavy chain only antibodies and characterized by a high stability and solubility. The selected candidates were expressed, purified and validated regarding their binding properties in different enzyme-linked immunosorbent assays formats, immunofluorescence, immunohistochemistry and surface plasmon resonance spectroscopy. Four different nanobodies could be selected whereof three recognize the short isoform of GP2 very specifically and one nanobody showed a high binding capacity for both isoforms. The KD values measured for all nanobodies were between 1.3 nM and 2.3 pM indicating highly specific binders suitable for the application as diagnostic tool in inflammatory bowel disease.
Organism growth can be limited either by a single resource or by multiple resources simultaneously (co-limitation). Efforts to characterise co-limitation have generated two influential approaches. One approach uses limitation scenarios of factorial growth assays to distinguish specific types of co-limitation; the other uses growth responses spanned over a continuous, multi-dimensional resource space to characterise different types of response surfaces. Both approaches have been useful in investigating particular aspects of co-limitation, but a synthesis is needed to stimulate development of this recent research area. We address this gap by integrating the two approaches, thereby presenting a more general framework of co-limitation. We found that various factorial (co-)limitation scenarios can emerge in different response surface types based on continuous availabilities of essential or substitutable resources. We tested our conceptual co-limitation framework on data sets of published and unpublished studies examining the limitation of two herbivorous consumers in a two-dimensional resource space. The experimental data corroborate the predictions, suggesting a general applicability of our co-limitation framework to generalist consumers and potentially also to other organisms. The presented framework might give insight into mechanisms that underlie co-limitation responses and thus can be a seminal starting point for evaluating co-limitation patterns in experiments and nature.
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.
Polymer multicomponent coatings such as multilayers mimic an extracellular, matrix (ECM) that attracts significant attention for the use of the multilayers as functional supports for advanced cell culture and tissue engineering. Herein, biodegradation and molecular transport in hyaluronan/polylysine multilayers coated with gold nanoparticles were described. Nanoparticle coating acts as a semipermeable barrier that governs molecular transport into/from the multilayers, and makes them biodegradation-resistant. Model protein lysozyme (mimics of ECM-soluble signals) diffuses into the multilayers as fast- and, slow-diffusing populations existing in an equilibrium,. Such a. composite system may have high potential to be exploited as degradation-resistant drug-delivery platforms suitable for cell-based applications.
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.
Understanding interactions of bacterial surface polysaccharides with receptor protein scaffolds is important for the development of antibiotic therapies. The corresponding protein recognition domains frequently form low-affinity complexes with polysaccharides that are difficult to address with experimental techniques due to the conformational flexibility of the polysaccharide. In this work, we studied the tailspike protein (TSP) of the bacteriophage Sf6. Sf6TSP binds and hydrolyzes the high-rhamnose, serotype Y O-antigen polysaccharide of the Gram-negative bacterium Shigella flexneri (S. flexneri) as a first step of bacteriophage infection. Spectroscopic analyses and enzymatic cleavage assays confirmed that Sf6TSP binds long stretches of this polysaccharide. Crystal structure analysis and saturation transfer difference (STD) NMR spectroscopy using an enhanced method to interpret the data permitted the detailed description of affinity contributions and flexibility in an Sf6TSP-octasaccharide complex. Dodecasaccharide fragments corresponding to three repeating units of the O-antigen in complex with Sf6TSP were studied computationally by molecular dynamics simulations. They showed that distortion away from the low-energy solution conformation found in the octasaccharide complex is necessary for ligand binding. This is in agreement with a weak-affinity functional polysaccharide protein contact that facilitates correct placement and thus hydrolysis of the polysaccharide close to the catalytic residues. Our simulations stress that the flexibility of glycan epitopes together with a small number of specific protein contacts provide the driving force for Sf6TSP-polysaccharide complex formation in an overall weak-affinity interaction system.
Background
Non-typhoid Salmonella Typhimurium (S. Typhimurium) accounts for a high number of registered salmonellosis cases, and O-serotyping is one important tool for monitoring epidemiology and spread of the disease. Moreover, variations in glucosylated O-antigens are related to immunogenicity and spread in the host. However, classical autoagglutination tests combined with the analysis of specific genetic markers cannot always reliably register phase variable glucose modifications expressed on Salmonella O-antigens and additional tools to monitor O-antigen glucosylation phenotypes of S. Typhimurium would be desirable.
Results
We developed a test for the phase variable O-antigen glucosylation state of S. Typhimurium using the tailspike proteins (TSP) of Salmonella phages 9NA and P22. We used this ELISA like tailspike adsorption (ELITA) assay to analyze a library of 44 Salmonella strains. ELITA was successful in discriminating strains that carried glucose 1-6 linked to the galactose of O-polysaccharide backbone (serotype O1) from non-glucosylated strains. This was shown by O-antigen compositional analyses of the respective strains with mass spectrometry and capillary electrophoresis. The ELITA test worked rapidly in a microtiter plate format and was highly O-antigen specific. Moreover, TSP as probes could also detect glucosylated strains in flow cytometry and distinguish multiphasic cultures differing in their glucosylation state.
Conclusions
Tailspike proteins contain large binding sites with precisely defined specificities and are therefore promising tools to be included in serotyping procedures as rapid serotyping agents in addition to antibodies. In this study, 9NA and P22TSP as probes could specifically distinguish glucosylation phenotypes of Salmonella on microtiter plate assays and in flow cytometry. This opens the possibility for flow sorting of cell populations for subsequent genetic analyses or for monitoring phase variations during large scale O-antigen preparations necessary for vaccine production.
Background: Non-typhoid Salmonella Typhimurium (S. Typhimurium) accounts for a high number of registered salmonellosis cases, and O-serotyping is one important tool for monitoring epidemiology and spread of the disease. Moreover, variations in glucosylated O-antigens are related to immunogenicity and spread in the host. However, classical autoagglutination tests combined with the analysis of specific genetic markers cannot always reliably register phase variable glucose modifications expressed on Salmonella O-antigens and additional tools to monitor O-antigen glucosylation phenotypes of S. Typhimurium would be desirable. Results: We developed a test for the phase variable O-antigen glucosylation state of S. Typhimurium using the tailspike proteins (TSP) of Salmonella phages 9NA and P22. We used this ELISA like tailspike adsorption (ELITA) assay to analyze a library of 44 Salmonella strains. ELITA was successful in discriminating strains that carried glucose 1-6 linked to the galactose of O-polysaccharide backbone (serotype O1) from non-glucosylated strains. This was shown by O-antigen compositional analyses of the respective strains with mass spectrometry and capillary electrophoresis. The ELITA test worked rapidly in a microtiter plate format and was highly O-antigen specific. Moreover, TSP as probes could also detect glucosylated strains in flow cytometry and distinguish multiphasic cultures differing in their glucosylation state. Conclusions: Tailspike proteins contain large binding sites with precisely defined specificities and are therefore promising tools to be included in serotyping procedures as rapid serotyping agents in addition to antibodies. In this study, 9NA and P22TSP as probes could specifically distinguish glucosylation phenotypes of Salmonella on microtiter plate assays and in flow cytometry. This opens the possibility for flow sorting of cell populations for subsequent genetic analyses or for monitoring phase variations during large scale O-antigen preparations necessary for vaccine production.
Molybdoenzymes are widespread in eukaryotic and prokaryotic organisms where they play crucial functions in detoxification reactions in the metabolism of humans and bacteria, in nitrate assimilation in plants and in anaerobic respiration in bacteria. To be fully active, these enzymes require complex molybdenum-containing cofactors, which are inserted into the apoenzymes after folding. For almost all the bacterial molybdoenzymes, molybdenum cofactor insertion requires the involvement of specific chaperones. In this review, an overview on the molybdenum cofactor biosynthetic pathway is given together with the role of specific chaperones dedicated for molybdenum cofactor insertion and maturation. Many bacteria are involved in geochemical cycles on earth and therefore have an environmental impact. The roles of molybdoenzymes in bioremediation and for environmental applications are presented.This review gives an overview of the diverse mechanisms leading to the insertion of the different forms of the molybdenum cofactor into the respective target enzymes and summarizes the roles of different molybdoenzymes in the environment.This review gives an overview of the diverse mechanisms leading to the insertion of the different forms of the molybdenum cofactor into the respective target enzymes and summarizes the roles of different molybdoenzymes in the environment.
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.
Recent technological developments have increased the number of variables being monitored in lakes and reservoirs using automatic high frequency monitoring (AHFM). However, design of AHFM systems and posterior data handling and interpretation are currently being developed on a site-by-site and issue-by-issue basis with minimal standardization of protocols or knowledge sharing. As a result, many deployments become short-lived or underutilized, and many new scientific developments that are potentially useful for water management and environmental legislation remain underexplored. This Critical Review bridges scientific uses of AHFM with their applications by providing an overview of the current AHFM capabilities, together with examples of successful applications. We review the use of AHFM for maximizing the provision of ecosystem services supplied, by lakes and reservoirs (consumptive and non consumptive uses, food production, and recreation), and for reporting lake status in the EU Water Framework Directive. We also highlight critical issues to enhance the application of AHFM, and suggest the establishment of appropriate networks to facilitate knowledge sharing and technological transfer between potential users. Finally, we give advice on how modern sensor technology can successfully be applied on a larger scale to the management of lakes and reservoirs and maximize the ecosystem services they provide.
Arf-like Protein 3 (ARL3) Regulates Protein Trafficking and Ciliogenesis in Mouse Photoreceptors
(2016)
Arf-like protein 3 (ARL3) is a ubiquitous small GTPase expressed in ciliated cells of plants and animals. Germline deletion of Arl3 in mice causes multiorgan ciliopathy reminiscent of Bardet-Biedl or Joubert syndromes. As photoreceptors are elegantly compartmentalized and have cilia, we probed the function of ARL3 (ADP-ribosylation factor (Arf)-like 3 protein) by generating rod photoreceptor-specific (prefix (rod)) and retina-specific (prefix (ret)) Arl3 deletions. In predegenerate (rod)Arl3(-/-) mice, lipidated phototransduction proteins showed trafficking deficiencies, consistent with the role of ARL3 as a cargo displacement factor for lipid-binding proteins. By contrast, (ret)Arl3(-/-) rods and cones expressing Cre recombinase during embryonic development formed neither connecting cilia nor outer segments and degenerated rapidly. Absence of cilia infers participation of ARL3 in ciliogenesis and axoneme formation. Ciliogenesis was rescued, and degeneration was reversed in part by subretinal injection of adeno-associated virus particles expressing ARL3-EGFP. The conditional knock-out phenotypes permitted identification of two ARL3 functions, both in the GTP-bound form as follows: one as a regulator of intraflagellar transport participating in photoreceptor ciliogenesis and the other as a cargo displacement factor transporting lipidated protein to the outer segment. Surprisingly, a farnesylated inositol polyphosphate phosphatase only trafficked from the endoplasmic reticulum to the Golgi, thereby excluding it from a role in photoreceptor cilia physiology.
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.
Fungi constitute important and conspicuous components of aquatic microbial communities, but their diversity and functional roles remain poorly characterized. New methods and conceptual frameworks are required to accurately describe their ecological roles, involvement in global cycling processes, and utility for human activities, considering both cultivation independent techniques as well as experiments in laboratory and in natural ecosystems. Here we highlight recent developments and extant knowledge gaps in aquatic mycology, and provide a conceptual model to expose the importance of fungi in aquatic food webs and related biogeochemical processes.
Background: Antiphospholipid antibodies (aPL) can be detected in asymptomatic carriers and infectious patients. The aim was to investigate whether a novel line immunoassay (LIA) differentiates between antiphospholipid syndrome (APS) and asymptomatic aPL+ carriers or patients with infectious diseases (infectious diseases controls (IDC)). Methods: Sixty-one patients with APS (56 primary, 22/56 with obstetric events only, and 5 secondary), 146 controls including 24 aPL+ asymptomatic carriers and 73 IDC were tested on a novel hydrophobic solid phase coated with cardiolipin (CL), phosphatic acid, phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, phosphatidylserine, beta2-glycoprotein I (beta 2GPI), prothrombin, and annexin V. Samples were also tested by anti-CL and anti-beta 2GPI ELISAs and for lupus anticoagulant activity. Human monoclonal antibodies (humoAbs) against human beta 2GPI or PL alone were tested on the same LIA substrates in the absence or presence of human serum, purified human beta 2GPI or after CL-micelle absorption. Results: Comparison of LIA with the aPL-classification assays revealed good agreement for IgG/IgM a beta 2GPI and aCL. Anti-CL and anti-beta 2GPI IgG/IgM reactivity assessed by LIA was significantly higher in patients with APS versus healthy controls and IDCs, as detected by ELISA. IgG binding to CL and beta 2GPI in the LIA was significantly lower in aPL+ carriers and Venereal Disease Research Laboratory test (VDRL) + samples than in patients with APS. HumoAb against domain 1 recognized beta 2GPI bound to the LIA-matrix and in anionic phospholipid (PL) complexes. Absorption with CL micelles abolished the reactivity of a PL-specific humoAb but did not affect the binding of anti-beta 2GPI humoAbs. Conclusions: The LIA and ELISA have good agreement in detecting aPL in APS, but the LIA differentiates patients with APS from infectious patients and asymptomatic carriers, likely through the exposure of domain 1.
Background
Antiphospholipid antibodies (aPL) can be detected in asymptomatic carriers and infectious patients. The aim was to investigate whether a novel line immunoassay (LIA) differentiates between antiphospholipid syndrome (APS) and asymptomatic aPL+ carriers or patients with infectious diseases (infectious diseases controls (IDC)).
Methods
Sixty-one patients with APS (56 primary, 22/56 with obstetric events only, and 5 secondary), 146 controls including 24 aPL+ asymptomatic carriers and 73 IDC were tested on a novel hydrophobic solid phase coated with cardiolipin (CL), phosphatic acid, phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, phosphatidylserine, beta2-glycoprotein I (β2GPI), prothrombin, and annexin V. Samples were also tested by anti-CL and anti-β2GPI ELISAs and for lupus anticoagulant activity. Human monoclonal antibodies (humoAbs) against human β2GPI or PL alone were tested on the same LIA substrates in the absence or presence of human serum, purified human β2GPI or after CL-micelle absorption.
Results
Comparison of LIA with the aPL-classification assays revealed good agreement for IgG/IgM aß2GPI and aCL. Anti-CL and anti-ß2GPI IgG/IgM reactivity assessed by LIA was significantly higher in patients with APS versus healthy controls and IDCs, as detected by ELISA. IgG binding to CL and ß2GPI in the LIA was significantly lower in aPL+ carriers and Venereal Disease Research Laboratory test (VDRL) + samples than in patients with APS. HumoAb against domain 1 recognized β2GPI bound to the LIA-matrix and in anionic phospholipid (PL) complexes. Absorption with CL micelles abolished the reactivity of a PL-specific humoAb but did not affect the binding of anti-β2GPI humoAbs.
Conclusions
The LIA and ELISA have good agreement in detecting aPL in APS, but the LIA differentiates patients with APS from infectious patients and asymptomatic carriers, likely through the exposure of domain 1.
Antibodies against spike proteins of influenza are used as a tool for characterization of viruses and therapeutic approaches. However, development, production and quality control of antibodies is expensive and time consuming. To circumvent these difficulties, three peptides were derived from complementarity determining regions of an antibody heavy chain against influenza A spike glycoprotein. Their binding properties were studied experimentally, and by molecular dynamics simulations. Two peptide candidates showed binding to influenza A/Aichi/2/68 H3N2. One of them, termed PeB, with the highest affinity prevented binding to and infection of target cells in the micromolar region without any cytotoxic effect. PeB matches best the conserved receptor binding site of hemagglutinin. PeB bound also to other medical relevant influenza strains, such as human-pathogenic A/California/7/2009 H1N1, and avian-pathogenic A/MuteSwan/Rostock/R901/2006 H7N1. Strategies to improve the affinity and to adapt specificity are discussed and exemplified by a double amino acid substituted peptide, obtained by substitutional analysis. The peptides and their derivatives are of great potential for drug development as well as biosensing.
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.
Species diversity is changing globally and locally, but the complexity of ecological communities hampers a general understanding of the consequences of animal species loss on ecosystem functioning. High animal diversity increases complementarity of herbivores but also increases feeding rates within the consumer guild. Depending on the balance of these counteracting mechanisms, species-rich animal communities may put plants under top-down control or may release them from grazing pressure. Using a dynamic food-web model with body-mass constraints, we simulate ecosystem functions of 20,000 communities of varying animal diversity. We show that diverse animal communities accumulate more biomass and are more exploitative on plants, despite their higher rates of intra-guild predation. However, they do not reduce plant biomass because the communities are composed of larger, and thus energetically more efficient, plant and animal species. This plasticity of community body-size structure reconciles the debate on the consequences of animal species loss for primary productivity.
Ancient DNA studies have revolutionized the study of extinct species and populations, providing insights on phylogeny, phylogeography, admixture and demographic history. However, inferences on behaviour and sociality have been far less frequent. Here, we investigate the complete mitochondrial genomes of extinct Late Pleistocene cave bears and middle Holocene brown bears that each inhabited multiple geographically proximate caves in northern Spain. In cave bears, we find that, although most caves were occupied simultaneously, each cave almost exclusively contains a unique lineage of closely related haplotypes. This remarkable pattern suggests extreme fidelity to their birth site in cave bears, best described as homing behaviour, and that cave bears formed stable maternal social groups at least for hibernation. In contrast, brown bears do not show any strong association of mitochondrial lineage and cave, suggesting that these two closely related species differed in aspects of their behaviour and sociality. This difference is likely to have contributed to cave bear extinction, which occurred at a time in which competition for caves between bears and humans was likely intense and the ability to rapidly colonize new hibernation sites would have been crucial for the survival of a species so dependent on caves for hibernation as cave bears. Our study demonstrates the potential of ancient DNA to uncover patterns of behaviour and sociality in ancient species and populations, even those that went extinct many tens of thousands of years ago.
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.
analysis
(2016)
The development of ‘omics’ technologies has progressed to address complex biological questions that underlie various plant functions thereby producing copious amounts of data. The need to assimilate large amounts of data into biologically meaningful interpretations has necessitated the development of statistical methods to integrate multidimensional information. Throughout this review, we provide examples of recent outcomes of ‘omics’ data integration together with an overview of available statistical methods and tools.
Maintenance of triplet decoding is crucial for the expression of functional protein because deviations either into the -1 or +1 reading frames are often non-functional. We report here that expression of huntingtin (Htt) exon 1 with expanded CAG repeats, implicated in Huntington pathology, undergoes a sporadic +1 frameshift to generate from the CAG repeat a trans-frame AGC repeat-encoded product. This +1 recoding is exclusively detected in pathological Htt variants, i.e. those with expanded repeats with more than 35 consecutive CAG codons. An atypical +1 shift site, UUC C at the 5 end of CAG repeats, which has some resemblance to the influenza A virus shift site, triggers the +1 frameshifting and is enhanced by the increased propensity of the expanded CAG repeats to form a stem-loop structure. The +1 trans-frame-encoded product can directly influence the aggregation of the parental Htt exon 1.
Over the last decades, the world’s population has been growing at a faster rate, resulting in increased urbanisation, especially in developing countries. More than half of the global population currently lives in urbanised areas with an increasing tendency. The growth of cities results in a significant loss of vegetation cover, soil compaction and sealing of the soil surface which in turn results in high surface runoff during high-intensity storms and causes the problem of accelerated soil water erosion on streets and building grounds. Accelerated soil water erosion is a serious environmental problem in cities as it gives rise to the contamination of aquatic bodies, reduction of ground water recharge and increase in land degradation, and also results in damages to urban infrastructures, including drainage systems, houses and roads. Understanding the problem of water erosion in urban settings is essential for the sustainable planning and management of cities prone to water erosion. However, in spite of the vast existence of scientific literature on water erosion in rural regions, a concrete understanding of the underlying dynamics of urban erosion still remains inadequate for the urban dryland environments.
This study aimed at assessing water erosion and the associated socio-environmental determinants in a typical dryland urban area and used the city of Windhoek, Namibia, as a case study. The study used a multidisciplinary approach to assess the problem of water erosion. This included an in depth literature review on current research approaches and challenges of urban erosion, a field survey method for the quantification of the spatial extent of urban erosion in the dryland city of Windhoek, and face to face interviews by using semi-structured questionnaires to analyse the perceptions of stakeholders on urban erosion.
The review revealed that around 64% of the literatures reviewed were conducted in the developed world, and very few researches were carried out in regions with extreme climate, including dryland regions. Furthermore, the applied methods for erosion quantification and monitoring are not inclusive of urban typical features and they are not specific for urban areas. The reviewed literature also lacked aspects aimed at addressing the issues of climate change and policies regarding erosion in cities. In a field study, the spatial extent and severity of an urban dryland city, Windhoek, was quantified and the results show that nearly 56% of the city is affected by water erosion showing signs of accelerated erosion in the form of rills and gullies, which occurred mainly in the underdeveloped, informal and semi-formal areas of the city. Factors influencing the extent of erosion in Windhoek included vegetation cover and type, socio-urban factors and to a lesser extent slope estimates. A comparison of an interpolated field survey erosion map with a conventional erosion assessment tool (the Universal Soil Loss Equation) depicted a large deviation in spatial patterns, which underlines the inappropriateness of traditional non-urban erosion tools to urban settings and emphasises the need to develop new erosion assessment and management methods for urban environments. It was concluded that measures for controlling water erosion in the city need to be site-specific as the extent of erosion varied largely across the city.
The study also analysed the perceptions and understanding of stakeholders of urban water erosion in Windhoek, by interviewing 41 stakeholders using semi-structured questionnaires. The analysis addressed their understanding of water erosion dynamics, their perceptions with regards to the causes and the seriousness of erosion damages, and their attitudes towards the responsibilities for urban erosion. The results indicated that there is less awareness of the process as a phenomenon, instead there is more awareness of erosion damages and the factors contributing to the damages. About 69% of the stakeholders considered erosion damages to be ranging from moderate to very serious. However, there were notable disparities between the private householders and public authority groups. The study further found that the stakeholders have no clear understanding of their responsibilities towards the management of the control measures and payment for the damages. The private householders and local authority sectors pointed fingers at each other for the responsibilities for erosion damage payments and for putting up prevention measures. The reluctance to take responsibility could create a predicament for areas affected, specifically in the informal settlements where land management is not carried out by the local authority and land is not owned by the occupants.
The study concluded that in order to combat urban erosion, it is crucial to understand diverse dynamics aggravating the process of urbanisation from different scales. Accordingly, the study suggests that there is an urgent need for the development of urban-specific approaches that aim at: (a) incorporating the diverse socio-economic-environmental aspects influencing erosion, (b) scientifically improving natural cycles that influence water storages and nutrients for plants in urbanised dryland areas in order to increase the amount of vegetation cover, (c) making use of high resolution satellite images to improve the adopted methods for assessing urban erosion, (d) developing water erosion policies, and (e) continuously monitoring the impact of erosion and the influencing processes from local, national and international levels.
The amyloid precursor protein (APP) and its paralogs, amyloid precursor-like protein 1 (APLP1) and APLP2, are metalloproteins with a putative role both in synaptogenesis and in maintaining synapse structure. Here, we studied the effect of zinc on membrane localization, adhesion, and secretase cleavage of APP, APLP1, and APLP2 in cell culture and rat neurons. For this, we employed live-cell microscopy techniques, a microcontact printing adhesion assay and ELISA for protein detection in cell culture supernatants. We report that zinc induces the multimerization of proteins of the amyloid precursor protein family and enriches them at cellular adhesion sites. Thus, zinc facilitates the formation of de novo APP and APLP1 containing adhesion complexes, whereas it does not have such influence on APLP2. Furthermore, zinc-binding prevented cleavage of APP and APLPs by extracellular secretases. In conclusion, the complexation of zinc modulates neuronal functions of APP and APLPs by (i) regulating formation of adhesion complexes, most prominently for APLP1, and (ii) by reducing the concentrations of neurotrophic soluble APP/APLP ectodomains.
A common misconception persists that the genomes of toxic and non-toxic cyanobacterial strains are largely conserved with the exception of the presence or absence of the genes responsible for toxin production. Implementation of -omics era technologies has challenged this paradigm, with comparative analyses providing increased insight into the differences between strains of the same species. The implementation of genomic, transcriptomic and proteomic approaches has revealed distinct profiles between toxin-producing and non-toxic strains. Further, metagenomics and metaproteomics highlight the genomic potential and functional state of toxic bloom events over time. In this review, we highlight how these technologies have shaped our understanding of the complex relationship between these molecules, their producers and the environment at large within which they persist.