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Biological invasions are a major threat to natural biodiversity; hence, understanding the mechanisms underlying invasibility (i.e., the susceptibility of a community to invasions by new species) is crucial. Invasibility of a resident community may be affected by a complex but hitherto hardly understood interplay of (1) productivity of the habitat, (2) diversity, (3) herbivory, and (4) the characteristics of both invasive and resident species. Using experimental phytoplankton microcosms, we investigated the effect of nutrient supply and species diversity on the invasibility of resident communities for two functionally different invaders in the presence or absence of an herbivore. With increasing nutrient supply, increased herbivore abundance indicated enhanced phytoplankton biomass production, and the invasion success of both invaders showed a unimodal pattern. At low nutrient supply (i.e., low influence of herbivory), the invasibility depended mainly on the competitive abilities of the invaders, whereas at high nutrient supply, the susceptibility to herbivory dominated. This resulted in different optimum nutrient levels for invasion success of the two species due to their individual functional traits. To test the effect of diversity on invasibility, a species richness gradient was generated by random selection from a resident species pool at an intermediate nutrient level. Invasibility was not affected by species richness; instead, it was driven by the functional traits of the resident and/or invasive species mediated by herbivore density. Overall, herbivory was the driving factor for invasibility of phytoplankton communities, which implies that other factors affecting the intensity of herbivory (e.g., productivity or edibility of primary producers) indirectly influence invasions.
Mountain gazelles (Gazella gazella) rank among the most critically endangered mammals on the Arabian Peninsula. Past conservation efforts have been plagued by confusion about the phylogenetic relationship among various 'phenotypically discernable' populations, and even the question of species boundaries was far from being certain. This lack of knowledge has had a direct impact on conservation measures, especially ex situ breeding programmes, hampering the assignment of captive stocks to potential conservation units. Here, we provide a phylogenetic framework, based on the analysis of mtDNA sequences (360 bp cytochrome b and 213 bp Control Region) of 126 individuals collected from the wild throughout the Arabian Peninsula and from captive stocks. Our analyses revealed two reciprocally monophyletic genetic lineages within the presumed species Gazella gazella: one 'northern clade' on the Golan Heights (Israel/Syrian border) and one genetically diverse larger clade from the rest of the Arabian Peninsula including the Arava Valley (Negev, Israel). Applying the Strict Phylogenetic Species Concept (sensu Mishler & Theriot, 2000) allows assigning species status to these two major clades.
In most mammals, females are philopatric while males disperse in order to avoid inbreeding. We investigated social structure in a solitary ungulate, the bushbuck Tragelaphus sylvaticus in Queen Elizabeth National Park, Uganda by combining behavioural and molecular data. We correlated spatial and social vicinity of individual females with a relatedness score obtained from mitochondrial DNA analysis. Presumed clan members shared the same haplotype, showed more socio-positive interactions and had a common home range. Males had a higher haplotype diversity than females. All this suggests the presence of a matrilineal structure in the study population. Moreover, we tested natal dispersal distances between male and female yearlings and used control region sequences to confirm that females remain in their natal breeding areas whereas males disperse. In microsatellite analysis, males showed a higher genetic variability than females. The impoverished genetic variability of females at both molecular marker sets is consistent with a philopatric and matrilineal structure, while the higher degree of genetic variability of males is congruent with a higher dispersal rate expected in this sex. Evidence even for male long-distance dispersal is brought about by one male carrying a haplotype of a different subspecies, previously not described to occur in this area.
Using molecular genetic methods and an ancient DNA approach, we studied population and species succession of rotifers of the genus Brachionus in the Kenyan alkaline-saline crater lake Sonachi since the beginning of the 19th century as well as distribution of Brachionus haplotypes in recent and historic sediments of other lakes of the East African Rift System. The sediment core record of Lake Sonachi displays haplotypes of a distinct evolutionary lineage in all increments. Populations were dominated by a single mitochondrial haplotype for a period of 150 years, and two putatively intraspecific turnovers in dominance occurred. Both changes are concordant with major environmental perturbations documented by a profound visible change in sediment composition of the core. The first change was very abrupt and occurred after the deposition of volcanic ash at the beginning of the 19th century. The second change coincides with a major lake level lowstand during the 1940s. It was preceded by a period of successively declining lake level, in which two other haplotypes appeared in the lake. One of these putatively belongs to another species documented in historical and recent Kenyan lake sediments. The analysis of plankton population dynamics through historical time can reveal patterns of population persistence and turnover in relation to environmental changes.
In this study, we have used fragments of three mitochondrial genes (Control Region, CR; transfer RNA for methionine, tRNA-Met; NADH dehydrogenase subunit 2, ND2 for a total of 1066 bp) to reconstruct the phylogeographic history of the endemic Philippine bulbul (Hypsipetes philippinus) at the scale of the central area of the Philippine archipelago. The study includes two of the five recognized subspecies (guimarasensis and mindorensis), 7 populations and 58 individuals. Multiple phylogenetic and network analyses support the existence of two reciprocally monophyletic maternal lineages corresponding to the two named subspecies. Molecular clock estimates indicate that the split between the two subspecies is consistent with the Pleistocene geological history of the archipelago. Patterns of relationships within guimarasensis are biogeographically less clear. Here, a combination of vicariance and dispersal needs to be invoked to reconcile the molecular data with the geographical origin of samples. In particular, the two islands Boracay and Negros host mitochondrial lineages that do not form monophyletic clusters. Our genetic data suggest multiple independent colonization events for these locations.
This work presents the development of entropy-elastic gelatin based networks in the form of films or scaffolds. The materials have good prospects for biomedical applications, especially in the context of bone regeneration. Entropy-elastic gelatin based hydrogel films with varying crosslinking densities were prepared with tailored mechanical properties. Gelatin was covalently crosslinked above its sol gel transition, which suppressed the gelatin chain helicity. Hexamethylene diisocyanate (HDI) or ethyl ester lysine diisocyanate (LDI) were applied as chemical crosslinkers, and the reaction was conducted either in dimethyl sulfoxide (DMSO) or water. Amorphous films were prepared as measured by Wide Angle X-ray Scattering (WAXS), with tailorable degrees of swelling (Q: 300-800 vol. %) and wet state Young’s modulus (E: 70 740 kPa). Model reactions showed that the crosslinking reaction resulted in a combination of direct crosslinks (3-13 mol.-%), grafting (5-40 mol.-%), and blending of oligoureas (16-67 mol.-%). The knowledge gained with this bulk material was transferred to the integrated process of foaming and crosslinking to obtain porous 3-D gelatin-based scaffolds. For this purpose, a gelatin solution was foamed in the presence of a surfactant, Saponin, and the resulting foam was fixed by chemical crosslinking with a diisocyanate. The amorphous crosslinked scaffolds were synthesized with varied gelatin and HDI concentrations, and analyzed in the dry state by micro computed tomography (µCT, porosity: 65±11–73±14 vol.-%), and scanning electron microscopy (SEM, pore size: 117±28–166±32 µm). Subsequently, the work focused on the characterization of the gelatin scaffolds in conditions relevant to biomedical applications. Scaffolds showed high water uptake (H: 630-1680 wt.-%) with minimal changes in outer dimension. Since a decreased scaffold pore size (115±47–130±49 µm) was revealed using confocal laser scanning microscopy (CLSM) upon wetting, the form stability could be explained. Shape recoverability was observed after removal of stress when compressing wet scaffolds, while dry scaffolds maintained the compressed shape. This was explained by a reduction of the glass transition temperature upon equilibration with water (dynamic mechanical analysis at varied temperature (DMTA)). The composition dependent compression moduli (Ec: 10 50 kPa) were comparable to the bulk micromechanical Young’s moduli, which were measured by atomic force microscopy (AFM). The hydrolytic degradation profile could be adjusted, and a controlled decrease of mechanical properties was observed. Partially-degraded scaffolds displayed an increase of pore size. This was likely due to the pore wall disintegration during degradation, which caused the pores to merge. The scaffold cytotoxicity and immunologic responses were analyzed. The porous scaffolds enabled proliferation of human dermal fibroblasts within the implants (up to 90 µm depth). Furthermore, indirect eluate tests were carried out with L929 cells to quantify the material cytotoxic response. Here, the effect of the sterilization method (Ethylene oxide sterilization), crosslinker, and surfactant were analyzed. Fully cytocompatible scaffolds were obtained by using LDI as crosslinker and PEO40 PPO20-PEO40 as surfactant. These investigations were accompanied by a study of the endotoxin material contamination. The formation of medical-grade materials was successfully obtained (<0.5 EU/mL) by using low-endotoxin gelatin and performing all synthetic steps in a laminar flow hood.
Bottom-up effects of plant diversity on multitrophic interactions in a biodiversity experiment
(2010)
Penicillin amidase from Alacaligenes faecalis is an attractive biocatalyst for hydrolysis of penicillin G for production of 6-aminopenicillanic acid, which is used in the synthesis of semi-synthetic beta-lactam antibiotics. Recently a mutant of this enzyme with extended C-terminus of the A-chain comprising parts of the connecting linker peptide was constructed. Its turnover number for the hydrolysis of penicillin G was 140 s(-1), about twice of the value for the wild-type enzyme (80 s(-1)). At the same time the specificity constant was improved about three-fold. The wild- type and the mutant enzymes showed similar pH stability suggesting that the linker peptide fragment covalently attached to the A-chain does not alter the electrostatic interactions in the protein core. Although the global stability of A. faecalis wild-type enzyme and the T206GS213G variant does not differ, the presence of the linker fragment stabilizes the domains interface, as evidenced by the monophasic transition of the mutant enzyme from folded to unfolded state during urea-induced denaturation. The high stability and activity of the mutant enzyme provides a rationale to use it as a biocatalyst in the industrial processes, where the enzyme must be more robust to fluctuations in the operational conditions.
The Erysiphales species Phyllactinia hippophaës Thuem. ex S. Blumer was found for the fi rst time on cultivated Sea Buckthorn (Hippophaë rhamnoides L.) near Großkayna (Saxony-Anhalt) in October 2009. This fungus was considered to be extinct in Germany. Intensive searching in Saxony-Anhalt and the Potsdam area (Brandenburg) yielded many additional records, most of them from former brown coal mining areas or in Sea Buckthorn plantations.
Human sulfite oxidase (hSO) was immobilised on SAM-coated silver electrodes under preservation of the native heme pocket structure of the cytochrome b5 (Cyt b5) domain and the functionality of the enzyme. The redox properties and catalytic activity of the entire enzyme were studied by surface enhanced resonance Raman (SERR) spectroscopy and cyclic voltammetry (CV) and compared to the isolated heme domain when possible. It is shown that heterogeneous electron transfer and catalytic activity of hSO sensitively depend on the local environment of the enzyme. Increasing the ionic strength of the buffer solution leads to an increase of the heterogeneous electron transfer rate from 17 s(-1) to 440 s(- 1) for hSO as determined by SERR spectroscopy. CV measurements demonstrate an increase of the apparent turnover rate for the immobilised hSO from 0.85 s(-1) in 100 mM buffer to 5.26 s(-1) in 750 mM buffer. We suggest that both effects originate from the increased mobility of the surface-bound enzyme with increasing ionic strength. In agreement with surface potential calculations we propose that at high ionic strength the enzyme is immobilised via the dimerisation domain to the SAM surface. The flexible loop region connecting the Moco and the Cyt b5 domain allows alternating contact with the Moco interaction site and the SAM surface, thereby promoting the sequential intramolecular and heterogeneous electron transfer from Moco via Cyt b5 to the electrode. At lower ionic strength, the contact time of the Cyt b5 domain with the SAM surface is longer, corresponding to a slower overall electron transfer process.
Background: Cysteine is a component in organic compounds including glutathione that have been implicated in the adaptation of plants to stresses. O-acetylserine (thiol) lyase (OAS-TL) catalyses the final step of cysteine biosynthesis. OAS-TL enzyme isoforms are localised in the cytoplasm, the plastids and mitochondria but the contribution of individual OAS-TL isoforms to plant sulphur metabolism has not yet been fully clarified. Results: The seedling lethal phenotype of the Arabidopsis onset of leaf death3-1 (old3-1) mutant is due to a point mutation in the OAS-A1 gene, encoding the cytosolic OAS-TL. The mutation causes a single amino acid substitution from Gly(162) to Glu(162), abolishing old3-1 OAS-TL activity in vitro. The old3-1 mutation segregates as a monogenic semidominant trait when backcrossed to its wild type accession Landsberg erecta (Ler-0) and the Di-2 accession. Consistent with its semi- dominant behaviour, wild type Ler-0 plants transformed with the mutated old3-1 gene, displayed the early leaf death phenotype. However, the old3-1 mutation segregates in an 11: 4: 1 (wild type: semi-dominant: mutant) ratio when backcrossed to the Colombia-0 and Wassilewskija accessions. Thus, the early leaf death phenotype depends on two semi- dominant loci. The second locus that determines the old3-1 early leaf death phenotype is referred to as odd-ler (for old3 determinant in the Ler accession) and is located on chromosome 3. The early leaf death phenotype is temperature dependent and is associated with increased expression of defence-response and oxidative-stress marker genes. Independent of the presence of the odd-ler gene, OAS-A1 is involved in maintaining sulphur and thiol levels and is required for resistance against cadmium stress. Conclusions: The cytosolic OAS-TL is involved in maintaining organic sulphur levels. The old3-1 mutation causes genome-dependent and independent phenotypes and uncovers a novel function for the mutated OAS- TL in cell death regulation.
Supporting species persistence may involve (re)connecting suitable habitats. However, for many declining species habitat suitability and drivers of establishment are poorly known. We addressed this experimentally for a declining flagship species of dry grasslands in Germany, Armeria maritima subsp. elongata. In three regions, we sowed seeds from each of eight source populations back to their origin and to eight apparently suitable, but currently unoccupied, habitats close to the source populations. Overall, seeds germinated and seedlings established equally well in occupied and potential sites indicating that suitable habitats are available, but lack seed input. Germination and establishment varied among sowing sites. Moreover, seeds from populations of lower current connectivity established less well in new sites, and establishment was more variable among seeds from smaller than from larger populations, possibly reflecting genetic consequences of habitat fragmentation. Further, establishment across different new environments differed between seeds from different populations. As this was neither related to a home-away contrast nor to geographic or environmental distance between sites it could not clearly be attributed to local adaptation. To promote long-term persistence within this dry-grassland meta-population context we suggest increasing the density of suitable habitats and supporting dispersal connecting multiple sites, e.g. by promoting sheep transhumance, to increase current populations and their connectivity, and to colonise suitable habitats with material from different sources. We suggest that sowing experiments with characteristic species, including multiple source populations and multiple recipient sites, should be used regularly to inform connecting efforts in plant conservation.
Representatives of the genus Stentor (Stentoridae, Heterotrichea) are striking ciliates in environmental water samples because of their size (up to 4 mm) and their trumpet-like shape. Important for species identification are the following main characteristics: (1) the presence or absence of endosymbiotic algae (zoochlorellae); (2) the colour of the pigmented cortical granules, and (3) the shape of the macronucleus. The complete small subunit rDNA (SSU rDNA) of 19 further representatives of the genus Stentor was sequenced to examine the phylogenetic relationships within this genus and to determine the taxonomic value of these main characteristics. The detailed phylogenetic analyses yielded a separation of all species possessing a single compact macronucleus from those species with an "elongated" macronucleus (moniliform or vermiform). The data also indicate that the uptake of algae as well as the loss of pigmentation happened independently in different lineages. Furthermore, a high level of intraspecific variation within several species was found. Thus, S. muelleri and S. (sp.) cf. katashimai appear to represent distinct species and S. multiformis is composed of a species complex.
The translation of genetic information according to the sequence of the mRNA template occurs with high accuracy and fidelity. Critical events in each single step of translation are selection of transfer RNA (tRNA), codon reading and tRNA-regeneration for a new cycle. We developed a model that accurately describes the dynamics of single elongation steps, thus providing a systematic insight into the sensitivity of the mRNA translation rate to dynamic environmental conditions. Alterations in the concentration of the aminoacylated tRNA can transiently stall the ribosomes during translation which results, as suggested by the model, in two outcomes: either stress-induced change in the tRNA availability triggers the premature termination of the translation and ribosomal dissociation, or extensive demand for one tRNA species results in a competition between frameshift to an aberrant open-reading frame and ribosomal drop-off. Using the bacterial Escherichia coli system, we experimentally draw parallels between these two possible mechanisms.
Die Qualität von Nutzpflanzen ist von zahlreichen Einflussfaktoren wie beispielsweise Lagerbedingungen und Sorteneigenschaften abhängig. Um Qualitätsmängel zu minimieren und Absatzchancen von Nutzpflanzen zu steigern sind umfangreiche Analysen hinsichtlich ihrer stofflichen Zusammensetzung notwendig. Chromatographische Techniken gekoppelt an ein Massenspektrometer und die Kernspinresonanzspektroskopie wurden dafür bislang verwendet. In der vorliegenden Arbeit wurde ein Gaschromatograph an ein Flugzeitmassenspektrometer (GC-TOF-MS) gekoppelt, um physiologische Prozesse bzw. Eigenschaften (die Schwarzfleckigkeit, die Chipsbräunung, das Physiologische Alter und die Keimhemmung) von Nutzpflanzen aufzuklären. Als Pflanzenmodell wurde dafür die Kartoffelknolle verwendet. Dazu wurden neue analytische Lösungsansätze entwickelt, die eine zielgerichtete Auswertung einer Vielzahl von Proben, die Etablierung einer umfangreichen Referenzspektrenbibliothek und die sichere Archivierung aller experimentellen Daten umfassen. Das Verfahren der Probenvorbereitung wurde soweit modifiziert, dass gering konzentrierte Substanzen mittels GC-TOF-MS analysiert werden können. Dadurch wurde das durch die Probenvorbereitung limitierte Substanzspektrum erweitert. Anhand dieser Lösungsansätze wurden physiologisch relevante Stoffwechselprodukte identifiziert, welche indikativ (klassifizierend) bzw. prädiktiv (vorhersagend) für die physiologischen Prozesse sind. Für die Schwarzfleckigkeitsneigung und die Chipseignung wurde jeweils ein biochemisches Modell zur Vorhersage dieser Prozesse aufgestellt und auf eine Züchtungspopulation übertragen. Ferner wurden für die Schwarzfleckigkeit Stoffwechselprodukte des Respirationsstoffwechsels identifiziert sowie Aminosäuren, Glycerollipide und Phenylpropanoide für das Physiologische Alter als relevant erachtet. Das physiologische Altern konnte durch die Anwendung höherer Temperaturen beschleunigt werden. Durch Anwendung von Keimhemmern (Kümmelöl, Chlorpropham) wurde eine Verzögerung des physiologischen Alterns beobachtet. Die Applikation von Kümmelöl erwies sich dabei als besonders vorteilhaft. Kümmelöl behandelte Knollen wiesen im Vergleich zu unbehandelten Knollen nur Veränderungen im Aminosäure-, Zucker- und Sekundärstoffwechsel auf. Chlorpropham behandelte Knollen wiesen einen ähnlichen Stoffwechsel wie die unbehandelten Knollen auf. Für die bislang noch nicht identifizierten Stoffwechselprodukte wurden im Rahmen dieser Arbeit das Verfahren der „gezielten An-/Abreicherung“, der „gepaarten NMR/GC-TOF-MS Analyse“ und das „Entscheidungsbaumverfahren“ entwickelt. Diese ermöglichen eine Klassifizierung von GC-MS Signalen im Hinblick auf ihre chemische Funktionalität. Das Verfahren der gekoppelten NMR/GC-TOF-MS Analyse erwies sich dabei als besonders erfolgversprechend, da es eine Aufklärung bislang unbekannter gaschromatographischer Signale ermöglicht. In der vorliegenden Arbeit wurden neue Stoffwechselprodukte in der Kartoffelknolle identifiziert, wodurch ein wertvoller Beitrag zur Analytik der Metabolomik geleistet wurde.
This work describes the realization of physically crosslinked networks based on gelatin by the introduction of functional groups enabling specific supramolecular interactions. Molecular models were developed in order to predict the material properties and permit to establish a knowledge-based approach to material design. The effect of additional supramolecular interactions with hydroxyapaptite was then studied in composite materials. The calculated properties are compared to experimental results to validate the models. The models are then further used for the study of physically crosslinked networks. Gelatin was functionalized with desaminotyrosine (DAT) and desaminotyrosyl-tyrosine (DATT) side groups, derived from the natural amino acid tyrosine. These group can potentially undergo to π-π and hydrogen bonding interactions also under physiological conditions. Molecular dynamics (MD) simulations were performed on models with 0.8 wt.-% or 25 wt.-% water content, using the second generation forcefield CFF91. The validation of the models was obtained by the comparison with specific experimental data such as, density, peptide conformational angles and X-ray scattering spectra. The models were then used to predict the supramolecular organization of the polymer chain, analyze the formation of physical netpoints and calculate the mechanical properties. An important finding of simulation was that with the increase of aromatic groups also the number of observed physical netpoints increased. The number of relatively stable physical netpoints, on average zero 0 for natural gelatin, increased to 1 and 6 for DAT and DATT functionalized gelatins respectively. A comparison with the Flory-Rehner model suggested reduced equilibrium swelling by factor 6 of the DATT-functionalized materials in water. The functionalized gelatins could be synthesized by chemoselective coupling of the free carboxylic acid groups of DAT and DATT to the free amino groups of gelatin. At 25 wt.-% water content, the simulated and experimentally determined elastic mechanical properties (e.g. Young Modulus) were both in the order of GPa and were not influenced by the degree of aromatic modification. The experimental equilibrium degree of swelling in water decreased with increasing the number of inserted aromatic functions (from 2800 vol.-% for pure gelatin to 300 vol.-% for the DATT modified gelatin), at the same time, Young’s modulus, elongation at break, and maximum tensile strength increased. It could be show that the functionalization with DAT and DATT influences the chain organization of gelatin based materials together with a controlled drying condition. Functionalization with DAT and DATT lead to a drastic reduction of helical renaturation, that could be more finely controlled by the applied drying conditions. The properties of the materials could then be influenced by application of two independent methods. Composite materials of DAT and DATT functionalized gelatins with hydroxyapatite (HAp) show a drastic reduction of swelling degree. In tensile tests and rheological measurements, the composites equilibrated in water had increased Young’s moduli (from 200 kPa up to 2 MPa) and tensile strength (from 57 kPa up to 1.1 MPa) compared to the natural polymer matrix without affecting the elongation at break. Furthermore, an increased thermal stability from 40 °C to 85 °C of the networks could be demonstrated. The differences of the behaviour of the functionalized gelatins to pure gelatin as matrix suggested an additional stabilizing bond between the incorporated aromatic groups to the hydroxyapatite.
Lake ecosystems across the globe have responded to climate warming of recent decades. However, correctly attributing observed changes to altered climatic conditions is complicated by multiple anthropogenic influences on lakes. This thesis contributes to a better understanding of climate impacts on freshwater phytoplankton, which forms the basis of the food chain and decisively influences water quality. The analyses were, for the most part, based on a long-term data set of physical, chemical and biological variables of a shallow, polymictic lake in north-eastern Germany (Müggelsee), which was subject to a simultaneous change in climate and trophic state during the past three decades. Data analysis included constructing a dynamic simulation model, implementing a genetic algorithm to parameterize models, and applying statistical techniques of classification tree and time-series analysis. Model results indicated that climatic factors and trophic state interactively determine the timing of the phytoplankton spring bloom (phenology) in shallow lakes. Under equally mild spring conditions, the phytoplankton spring bloom collapsed earlier under high than under low nutrient availability, due to a switch from a bottom-up driven to a top-down driven collapse. A novel approach to model phenology proved useful to assess the timings of population peaks in an artificially forced zooplankton-phytoplankton system. Mimicking climate warming by lengthening the growing period advanced algal blooms and consequently also peaks in zooplankton abundance. Investigating the reasons for the contrasting development of cyanobacteria during two recent summer heat wave events revealed that anomalously hot weather did not always, as often hypothesized, promote cyanobacteria in the nutrient-rich lake studied. The seasonal timing and duration of heat waves determined whether critical thresholds of thermal stratification, decisive for cyanobacterial bloom formation, were crossed. In addition, the temporal patterns of heat wave events influenced the summer abundance of some zooplankton species, which as predators may serve as a buffer by suppressing phytoplankton bloom formation. This thesis adds to the growing body of evidence that lake ecosystems have strongly responded to climatic changes of recent decades. It reaches beyond many previous studies of climate impacts on lakes by focusing on underlying mechanisms and explicitly considering multiple environmental changes. Key findings show that climate impacts are more severe in nutrient-rich than in nutrient-poor lakes. Hence, to develop lake management plans for the future, limnologists need to seek a comprehensive, mechanistic understanding of overlapping effects of the multi-faceted human footprint on aquatic ecosystems.
The genome can be considered the blueprint for an organism. Composed of DNA, it harbours all organism-specific instructions for the synthesis of all structural components and their associated functions. The role of carriers of actual molecular structure and functions was believed to be exclusively assumed by proteins encoded in particular segments of the genome, the genes. In the process of converting the information stored genes into functional proteins, RNA – a third major molecule class – was discovered early on to act a messenger by copying the genomic information and relaying it to the protein-synthesizing machinery. Furthermore, RNA molecules were identified to assist in the assembly of amino acids into native proteins. For a long time, these - rather passive - roles were thought to be the sole purpose of RNA. However, in recent years, new discoveries have led to a radical revision of this view. First, RNA molecules with catalytic functions - thought to be the exclusive domain of proteins - were discovered. Then, scientists realized that much more of the genomic sequence is transcribed into RNA molecules than there are proteins in cells begging the question what the function of all these molecules are. Furthermore, very short and altogether new types of RNA molecules seemingly playing a critical role in orchestrating cellular processes were discovered. Thus, RNA has become a central research topic in molecular biology, even to the extent that some researcher dub cells as “RNA machines”. This thesis aims to contribute towards our understanding of RNA-related phenomena by applying Bioinformatics means. First, we performed a genome-wide screen to identify sites at which the chemical composition of DNA (the genotype) critically influences phenotypic traits (the phenotype) of the model plant Arabidopsis thaliana. Whole genome hybridisation arrays were used and an informatics strategy developed, to identify polymorphic sites from hybridisation to genomic DNA. Following this approach, not only were genotype-phenotype associations discovered across the entire Arabidopsis genome, but also regions not currently known to encode proteins, thus representing candidate sites for novel RNA functional molecules. By statistically associating them with phenotypic traits, clues as to their particular functions were obtained. Furthermore, these candidate regions were subjected to a novel RNA-function classification prediction method developed as part of this thesis. While determining the chemical structure (the sequence) of candidate RNA molecules is relatively straightforward, the elucidation of its structure-function relationship is much more challenging. Towards this end, we devised and implemented a novel algorithmic approach to predict the structural and, thereby, functional class of RNA molecules. In this algorithm, the concept of treating RNA molecule structures as graphs was introduced. We demonstrate that this abstraction of the actual structure leads to meaningful results that may greatly assist in the characterization of novel RNA molecules. Furthermore, by using graph-theoretic properties as descriptors of structure, we indentified particular structural features of RNA molecules that may determine their function, thus providing new insights into the structure-function relationships of RNA. The method (termed Grapple) has been made available to the scientific community as a web-based service. RNA has taken centre stage in molecular biology research and novel discoveries can be expected to further solidify the central role of RNA in the origin and support of life on earth. As illustrated by this thesis, Bioinformatics methods will continue to play an essential role in these discoveries.
In dieser Arbeit wurden die Möglichkeiten und Grenzen für Zirkulardichroismus-Messungen mit Synchrotronstrahlung untersucht. Dazu wurde ein Messaufbau für Zirkulardichroismus-Messungen an zwei Strahlrohren am Berliner Elektronenspeicherring für Synchrotronstrahlung eingesetzt, die für Messungen im Bereich des ultravioletten Lichts geeignet sind. Eigenschaften der Strahlrohre und des Messaufbau wurden in einigen wichtigen Punkten mit kommerziellen Zirkulardichroismus-Spektrometern verglichen. Der Schwerpunkt lag auf der Ausdehnung des zugänglichen Wellenlängenbereichs unterhalb von 180 nm zur Untersuchung des Zirkulardichroismus von Proteinen in diesem Bereich. In diesem Bereich ist es nicht nur die Lichtquelle sondern vor allem die Absorption des Lichts durch Wasser, die den Messbereich bei der Messung biologischer Proben in wässriger Lösung einschränkt. Es wurden Bedingungen gefunden, unter denen der Messbereich auf etwa 160 nm, in einigen Fällen bis auf 130 nm ausgedehnt werden konnte. Dazu musste die Pfadlänge deutlich reduziert werden und verschieden Probenküvetten wurden getestet. Der Einfluss der dabei auftretenden Spannungsdoppelbrechung in den Probenküvetten auf das Messsignal konnte mit einem alternativen Messaufbau deutlich reduziert werden. Systematische Fehler im Messsignal und auftretende Strahlenschäden begrenzen jedoch die Zuverlässigkeit der gemessenen Spektren. Bei Proteinfilmen schränkt die Absorption von Wasser den Messbereich kaum ein. Es wurden jedoch meist deutliche Unterschiede zwischen den Spektren von Proteinfilmen und den Spektren von Proteinen in wässriger Lösung festgestellt. Solange diese Unterschiede nicht minimiert werden können, stellen Proteinfilme keine praktikable Alternative zu Messungen in wässriger Lösung dar.
Pektatlyase (Pel-15) aus dem alkalophilen Bodenbakterium Bacillus spec. KSM-P15 ist mit 197 Aminosäuren eines der kleinsten, bekannten β-3-Solenoidproteine. Sie spaltet Polygalakturonsäurederivate in einem Ca2+-abhängigen β-Eliminierungsprozess. Wie bei allen Proteinen dieser Enzymfamilie ist auch die Polypeptidkette von Pel-15 zu einer einsträngigen, rechtsgängigen, parallelen β-Helix aufgewunden. In diesem Strukturmotiv enthält jede Windung drei β-Stränge, die jeweils durch flexible Schleifenbereiche miteinander verbunden sind. Insgesamt acht Windungen stapeln sich in Pel-15 übereinander und bilden entlang der Helixachse flächige, parallele β-Faltblätter aus. Im Bereich dieser β-Faltblätter existiert ein ausgedehntes Netzwerk von Wasserstoffbrückenbindungen, durch das der hydrophobe Kern, der sich im Inneren der β-Helix befindet, vom umgebenden Lösungsmittel abgeschirmt wird. Besondere Abschlussstrukturen an beiden Enden der β-Helix, wie sie typischerweise bei anderen Ver-tretern dieser Strukturklasse ausgeprägt werden, sind in Pel-15 nicht zu beobachten. Stattdessen sind die terminalen Bereiche der β-Helix über Salzbrücken und hydrophobe Seitenkettenkontakte stabilisiert. In der vorliegenden Dissertation wurde die Pektatlyase Pel-15 hinsichtlich ihres Faltungsgleichgewichtes, ihrer enzymatischen Aktivität und der Kinetik ihrer Strukturbildung charakterisiert. In eine evolutionär konservierte Helixwindung wurden destabilisierende Mutationen eingeführt, und deren Auswirkungen mittels spektroskopischer Methoden analysiert. Die Ergebnisse zeigen, dass Pel-15 in Gegenwart des Denaturierungsmittels Guanidiniumhydrochlorid einen hyperfluoreszenten Gleichgewichtsustand (HF) populiert, der nach Messungen von Faltungs- und Entfaltungskinetiken ein konformationelles Ensemble aus den Zuständen HFslow und HFfast darstellt. Diese HF-Zustände sind durch eine hohe Aktivierungsbarriere voneinander getrennt. In Rückfaltungsexperimenten populieren nur etwa 80 % der faltenden Moleküle den Zwischenzustand HFslow, der mit einer Zeitkonstante von ca. 100 s zu HFfast weiterreagiert. Die Denaturierungsmittelabhängigkeit dieser Reaktion ist sehr gering, was eine trans-/cis-Prolylisomerisierung als geschwindigkeitslimitierenden Schritt nahelegt. Die Existenz eines cis-Peptides in der nativen Struktur macht es erforderlich, den denaturierten Zustand als ein Ensemble kinetisch separierter Konformationen, kurz: DSE, zu betrachten, das durch die Spezies Ufast und Uslow populiert wird. Nach dem in dieser Arbeit aufgestellten „Minimalmodell der Pel-15 Faltung“ stehen die HF-Spezies (HFslow, HFfast) mit den Konformationen des DSE in einem thermodynamischen Kreisprozess. Das Modell positioniert HFfast und die native Konformation N auf die „native Seite“ der Aktivierungsbarriere und trägt damit der Tatsache Rechnung, dass die Gleichgewichtseinstellung zwischen diesen Spezies zu schnell ist, um mit manuellen Techniken erfasst zu werden. Die hochaffine Bindung von Ca2+ (Kd = 10 μM) verschiebt sich das Faltungsgleichgewicht bereits in Gegenwart von 1 mM CaCl2 soweit auf die Seite des nativen Zustandes, das HFfast nicht länger nachweisbar ist. Entgegen anfänglicher Vermutungen kommt einer lokalen, evolutionär konservierten Disulfidbrücke im Zentrum der β-Helix eine wichtige Stabilisierungsfunktion zu. Die Disulfidbrücke befindet sich in einem kurzen Schleifenbereich der β-Helix nahe dem aktiven Zentrum. Obwohl ihr Austausch gegen die Reste Val und Ala die freie Stabilisierungsenthalpie des Proteins um ca. 10 kJ/mol reduziert, lässt die Struktur im Bereich der Mutationsstelle keine gravierende Veränderung erkennen. Auch die katalytisch relevante Ca2+-Bindungsaffinität bleibt unbeeinflusst; dennoch zeigen Enzymaktivitätstests für VA-Mutanten eine Reduktion der enzymatischen Aktivität um fast 50 % an. Die evolutionär konservierte Helixwindung im Allgemeinen und die in ihr enthaltene Disulfidbrücke im Besonderen müssen nach den vorliegenden Ergebnissen also eine zentrale Funktion sowohl für die Struktur des katalytischen Zentrums als auch für die Strukturbildung der β-Helix während der Faltungsreaktion besitzen. Die Ergebnisse dieser Arbeit finden in mehreren Punkten Anklang an Faltungseigenschaften, die für andere β -Helixproteine beschrieben wurden. Vor allem aber prädestinieren sie Pel-15 als ein neues, β-helikales Modellprotein. Aufgrund seiner einfachen Topologie, seiner niedrigen Windungszahl und seiner hohen thermodynamischen Stabilität ist Pel-15 sehr gut geeignet, die Determinanten von Stabilität und Strukturbildung des parallelen β-Helix-Motivs in einer Auflösung zu studieren, die aufgrund der Komplexität bestehender β-helikaler Modellsysteme bislang nicht zur Verfügung stand.
Fire prone Mediterranean-type vegetation systems like those in the Mediterranean Basin and South-Western Australia are global hot spots for plant species diversity. To ensure management programs act to maintain these highly diverse plant communities, it is necessary to get a profound understanding of the crucial mechanisms of coexistence. In the current literature several mechanisms are discussed. The objective of my thesis is to systematically explore the importance of potential mechanisms for maintaining multi-species, fire prone vegetation by modelling. The model I developed is spatially-explicit, stochastic, rule- and individual-based. It is parameterised on data of population dynamics collected over 18 years in the Mediterranean-type shrublands of Eneabba, Western Australia. From 156 woody species of the area seven plant traits have been identified to be relevant for this study: regeneration mode, annual maximum seed production, seed size, maximum crown diameter, drought tolerance, dispersal mode and seed bank type. Trait sets are used for the definition of plant functional types (PFTs). The PFT dynamics are simulated annual by iterating life history processes. In the first part of my thesis I investigate the importance of trade-offs for the maintenance of high diversity in multi-species systems with 288 virtual PFTs. Simulation results show that the trade-off concept can be helpful to identify non-viable combinations of plant traits. However, the Shannon Diversity Index of modelled communities can be high despite of the presence of ‘supertypes’. I conclude, that trade-offs between two traits are less important to explain multi-species coexistence and high diversity than it is predicted by more conceptual models. Several studies show, that seed immigration from the regional seed pool is essential for maintaining local species diversity. However, systematical studies on the seed rain composition to multi-species communities are missing. The results of the simulation experiments, as presented in part two of this thesis, show clearly, that without seed immigration the local species community found in Eneabba drifts towards a state with few coexisting PFTs. With increasing immigration rates the number of simulated coexisting PFTs and Shannon diversity quickly approaches values as also observed in the field. Including the regional seed input in the model is suited to explain more aggregated measures of the local plant community structure such as species richness and diversity. Hence, the seed rain composition should be implemented in future studies. In the third part of my thesis I test the sensitivity of Eneabba PFTs to four different climate change scenarios, considering their impact on both local and regional processes. The results show that climate change clearly has the potential to alter the number of dispersed seeds for most of the Eneabba PFTs and therefore the source of the ‘immigrants’ at the community level. A classification tree analysis shows that, in general, the response to climate change was PFT-specific. In the Eneabba sand plains sensitivity of a PFT to climate change depends on its specific trait combination and on the scenario of environmental change i.e. development of the amount of rainfall and the fire frequency. This result emphasizes that PFT-specific responses and regional process seed immigration should not be ignored in studies dealing with the impact of climate change on future species distribution. The results of the three chapters are finally analysed in a general discussion. The model is discussed and improvements and suggestions are made for future research. My work leads to the following conclusions: i) It is necessary to support modelling with empirical work to explain coexistence in species-rich plant communities. ii) The chosen modelling approach allows considering the complexity of coexistence and improves the understanding of coexistence mechanisms. iii) Field research based assumptions in terms of environmental conditions and plant life histories can relativise the importance of more hypothetic coexistence theories in species-rich systems. In consequence, trade-offs can play a lower role than predicted by conceptual models. iv) Seed immigration is a key process for local coexistence. Its alteration because of climate change should be considered for prognosis of coexistence. Field studies should be carried out to get data on seed rain composition.
The primary auditory cortex (AI) of adult Pteronotus parnellii features a foveal representation of the second harmonic constant frequency (CF2) echolocation call component. In the corresponding Doppler-shifted constant frequency (DSCF) area, the 61 kHz range is over-represented for extraction of frequency-shift information in CF2 echoes. To assess to which degree AI postnatal maturation depends on active echolocation or/and reflects ongoing cochlear maturation, cortical neurons were recorded in juveniles up to postnatal day P29, before the bats are capable of active foraging.At P1-2, neurons in posterior AI are tuned sensitively to low frequencies (22-45 dB SPL, 28-35 kHz). Within the prospective DSCF area, neurons had insensitive responses (>60 dB SPL) to frequencies <40 kHz and lacked sensitive tuning curve tips. Up to P10, when bats do not yet actively echolocate, tonotopy is further developed and DSCF neurons respond to frequencies of 51-57 kHz with maximum tuning sharpness (Q(10dB)) of 57. Between P11 and 20, the frequency representation in AI includes higher frequencies anterior and dorsal to the DSCF area. More multipeaked neurons (33%) are found than at older age. In the oldest group, DSCF neurons are tuned to frequencies close to 61 kHz with Q(10dB) values <= 212, and threshold sensitivity, tuning sharpness and cortical latencies are adult-like. The data show that basic aspects of cortical tonotopy are established before the bats actively echolocate. Maturation of tonotopy, increase of tuning sharpness, and upward shift in the characteristic frequency of DSCF neurons appear to strongly reflect cochlear maturation.
The centromeric histone H3 variant (CenH3) serves to target the kinetochore to the centromeres and thus ensures correct chromosome segregation during mitosis and meiosis. The Dictyostelium H3-like variant H3v1 was identified as the CenH3 ortholog. Dictyostelium CenH3 has an extended N-terminal domain with no similarity to any other known proteins and a histone fold domain at its C-terminus. Within the histone fold, alpha-helix 2 (alpha 2) and an extended loop 1 (L1) have been shown to be required for targeting CenH3 to centromeres. Compared to other known and putative CenH3 histones, Dictyostelium CenH3 has a shorter L1, suggesting that the extension is not an obligatory feature. Through ChIP analysis and fluorescence microscopy of live and fixed cells, we provide here the first survey of centromere structure in amoebozoa. The six telocentric centromeres were found to mostly consist of all the DIRS-1 elements and to associate with H3K9me3. During interphase, the centromeres remain attached to the centrosome forming a single CenH3-containing cluster. Loading of Dictyostelium CenH3 onto centromeres occurs at the G2/prophase transition, in contrast to the anaphase/ telophase loading of CenH3 observed in metazoans. This suggests that loading during G2/prophase is the ancestral eukaryotic mechanism and that anaphase/telophase loading of CenH3 has evolved more recently after the amoebozoa diverged from the animal linage.
A major aim in ecology is identifying determinants of invasiveness. We performed a meta-analysis of 117 field or experimental-garden studies that measured pair-wise trait differences of a total of 125 invasive and 196 non-invasive plant species in the invasive range of the invasive species. We tested whether invasiveness is associated with performance-related traits (physiology, leaf-area allocation, shoot allocation, growth rate, size and fitness), and whether such associations depend on type of study and on biogeographical or biological factors. Overall, invasive species had significantly higher values than non-invasive species for all six trait categories. More trait differences were significant for invasive vs. native comparisons than for invasive vs. non-invasive alien comparisons. Moreover, for comparisons between invasive species and native species that themselves are invasive elsewhere, no trait differences were significant. Differences in physiology and growth rate were larger in tropical regions than in temperate regions. Trait differences did not depend on whether the invasive alien species originates from Europe, nor did they depend on the test environment. We conclude that invasive alien species had higher values for those traits related to performance than non-invasive species. This suggests that it might become possible to predict future plant invasions from species traits.
Aims: Factors limiting distributions of species are fundamental to ecology and evolution but have rarely been addressed experimentally for multiple species. The conspicuous linear distribution patterns of plant species confined to river corridors in the Central European lowlands constitute an especially long-standing distribution puzzle. We experimentally tested our novel hypothesis that the tolerance of species to river corridor conditions is independent of the degree of confinement to river corridor habitats, but that species not confined to river corridors are better able to take advantage of the more benign non-river corridor conditions. Methods: We grew 42 herbaceous species differing in their confinement to river corridors in a common garden experiment on loamy soil typical for river corridor areas and sandy soil typical for non-river corridor areas, and with and without a flooding period. For a subset of species, we grew plants of both river corridor and non-river corridor origin to test for adaptation to river corridor conditions. Important findings: Species more confined to river corridor areas benefited less from the more benign non-flooded and non-river corridor soil conditions than species of wider distributional range did. For subsets of 7 and 12 widespread species, the response to flooding and soil origin, respectively, did not differ between plants from river corridor sites and plants from other sites, suggesting that the habitat tolerance of widespread species is clue to phenotypic plasticity rather than to local adaptation. Overall, we found clear support for our novel hypothesis that species not confined to river corridors are more able to take advantage of the more benign non-river corridor conditions. Our study provides a general hypothesis on differences between species confined to stressful habitats and widespread species out for test in further multispecies comparative experiments.
Background: Natural accessions of Arabidopsis thaliana are characterized by a high level of phenotypic variation that can be used to investigate the extent and mode of selection on the primary metabolic traits. A collection of 54 A. thaliana natural accession-derived lines were subjected to deep genotyping through Single Feature Polymorphism (SFP) detection via genomic DNA hybridization to Arabidopsis Tiling 1.0 Arrays for the detection of selective sweeps, and identification of associations between sweep regions and growth-related metabolic traits. Results: A total of 1,072,557 high-quality SFPs were detected and indications for 3,943 deletions and 1,007 duplications were obtained. A significantly lower than expected SFP frequency was observed in protein-, rRNA-, and tRNA-coding regions and in non- repetitive intergenic regions, while pseudogenes, transposons, and non-coding RNA genes are enriched with SFPs. Gene families involved in plant defence or in signalling were identified as highly polymorphic, while several other families including transcription factors are depleted of SFPs. 198 significant associations between metabolic genes and 9 metabolic and growth-related phenotypic traits were detected with annotation hinting at the nature of the relationship. Five significant selective sweep regions were also detected of which one associated significantly with a metabolic trait. Conclusions: We generated a high density polymorphism map for 54 A. thaliana accessions that highlights the variability of resistance genes across geographic ranges and used it to identify selective sweeps and associations between metabolic genes and metabolic phenotypes. Several associations show a clear biological relationship, while many remain requiring further investigation.
In einer Wochenstubengesellschaft der Rauhhautfledermaus (Pipistrellus nathusii), in der Schorfheide im Norden Brandenburgs, wurde ein starkes Vorkommen der Roten Waldameise (Formica rufa) festgestellt. Die Fledermäuse waren offensichtlich durch die Anwesenheit der Ameisen nicht beunruhigt. Der ungewöhnlich enge Kontakt zwischen Ameisen und Fledermäusen führte zur Überlegung, dass es regelmässige Interaktionen zwischen Ameisen und Fledermäusen geben könnte, die beiden Seiten Vorteile bieten. Vergleichbare Beziehungen wurden bisher hauptsächlich bei Vögeln als anting beschrieben. Interaktionen zum gegenseitigen Vorteil sind zwischen Fledermäusen und Ameisen bisher kaum beachtet worden, so dass weitere Untersuchungen notwendig sind.
Bei der Untersuchung von Ektoparasiten in verschiedenen Winterqartieren von Fledermäusen im Nordosten Brandenburgs wurde das Artenspektrum an Flöhen, Lausfliegen und Milben erfasst. Die häufigsten Parasiten waren die sogenannten "Winterflöhe", die mit drei Arten (Nycteridopsylla eusarca, N. longiceps und N. pentactena) präsent waren. Bei der Anzahl dieser Flöhe in den Winterquartieren wurden extreme Unterschiede festgestellt. In einzelnen Quartieren kann es zu einem relativ hohen Besatz an Flöhen kommen. Solche Populationszentren könnten für das regionale Überleben der seltenen Ektoparasitenarten eine große Rolle spielen. In diesen Quartieren werden neben den Hauptwirten auch andere überwinternde Fledermausarten als Blutspender genutzt. An den Wasserfledermäusen gab es regelmäßig Funde der Fledermausfliege Nycteribia kolenatii. Im Gegensatz zu den Flöhen und dieser Fledermausfliegenart traten die meisten anderen Parasiten nur sporadisch auf. Bei der Untersuchung wurden drei für Brandenburg neue Ektoparasitenarten festgestellt. Es handelt sich dabei um eine Fledermausfliege: Basilia nana sowie um zwei kleine Milbenarten der Familie Macronyssidae: Macronyssus cyclaspis und Macronyssus ellipticus.
A biosensor, based on a redoxactive osmium polymer and sulfite oxidase on screen-printed electrodes, is presented here as a promising method for the detection of sulfite. A catalytic oxidative current was generated when a sample containing sulfite was pumped over the carbon screen-printed electrode modified with osmium redox polymer wired sulfite oxidase. A stationary value was reached after approximately 50 s and a complete measurement lasted no more than 3 min. The electrode polarized at -0.1 V (vs. Ag vertical bar AgCl 1M KCl) permits minimizing the influence of interfering substances, since these compounds can be unspecific oxidized at higher potentials. Because of the good stability of the protein film on the electrode surface, a well functioning biosensor-flow system was possible to construct. The working stability and reproducibility were further enhanced by the addition of bovine serum albumin generating a more long-term stable and biocompatible protein environment. The optimized biosensor showed a stable signal for more than a week of operation and a coefficient of variation of 4.8% for 12 successive measurements. The lower limit of detection of the sensor was 0.5 mu M sulfite and the response was linear until 100 mu M. The high sensitivity permitted a 1:500 dilution of wine samples. The immobilization procedure and the operational conditions granted minimized interferences. Additionally, repeating the immobilization procedure to form several layers of wired SO further increased the sensitivity of such a sensor. Finally. the applicability of the developed sulfite biosensor was tested on real samples, such as white and red wines.
Synthetic Biology is advanced by many users and relies on the assembly of genetic elements to devices, systems and finally genomes. SynBioWave is a software suite that enables multiple distributed users to analyze and construct genetic parts in real-time collaboration. It builds on Google Wave and provides an extensible robot-robot-user communication framework, a menu driven user interface, biological data handling including DAS and an internal database communication. We demonstrate its use by implementing robots for gene-data retrieval, manipulation and display. The initial development of SynBioWave demonstrates the power of the underlying Google Wave protocol for Synthetic Biology and lays the foundation for continuous and user-friendly extensions. Specialized wave-robots with a manageable set of capabilities will divide and conquer the complex task of creating a genome in silico.
Invertebrate herbivores are ubiquitous in most terrestrial ecosystems, and theory predicts that their impact on plant community biomass should depend on diversity and productivity of the associated plant communities. To elucidate general patterns in the relationship between invertebrate herbivory, plant diversity, and productivity, we carried out a long-term herbivore exclusion experiment at multiple grassland sites in a mountainous landscape of central Germany. Over a period of five years, we used above-and belowground insecticides as well as a molluscicide to manipulate invertebrate herbivory at 14 grassland sites, covering a wide range of plant species diversity (13-38 species/m(2)) and aboveground plant productivity (272-1125 g.m(-2).yr(-1)), where plant species richness and productivity of the sites were not significantly correlated. Herbivore exclusion had significant effects on the plant communities: it decreased plant species richness and evenness, and it altered plant community composition. In particular, exclusion of belowground herbivores promoted grasses at the expense of herbs. In contrast to our expectation, herbivore effects on plant community biomass were not influenced by productivity. However, effect size of invertebrate herbivores was negatively correlated with plant diversity of the grasslands: the effect of herbivory on biomass tended to be negative at sites of high diversity and positive at sites of low diversity. In general, the effects of aboveground herbivores were relatively small as compared to belowground herbivores, which were important drivers of plant community composition. Our study is the first to show that variation in the effects of invertebrate herbivory on plant communities across a landscape is significantly influenced by plant species richness.
In freshwater systems, many abiotic and biotic factors determine the natural fluctuation of Daphnia spec. populations: climatic and water quality parameters, quantitative and qualitative food quality and quantity, predation, and humic substances. Many factors/stressors act in concert. In this contribution, we supplied Daphnia magna with two different diets (chlorococcal alga Pseudokirchneriella subcapitata and baker's yeast) fed ad libitum and exposed it to an environmentally realistic concentration of humic substances (HSs). Exposure to HSs caused a transcriptionally controlled stress response with studied genes; cat and hsp60, for the latter partial sequences have been identified. Furthermore, the exposure to HSs reduced the antioxidant capacity. Yet, a much stronger oxidative stress is caused by feeding yeast, which reduced the anti-oxidative capacity to values of approximately 50% of the green algal diet. This reduction is most likely due to the yeast's cell wall to resist digestion rather than to the elemental ratio or deficiency in long-chained unsaturated fatty acids, because both diets were deficient in fatty acids with back bones of more than 20 C-atoms. We assume that the biochemical machinery in the gut continuously activated oxygen to cleave the yeast's cell wall and, hence, reduced the antioxidative capacity of the animals. Neither the analyzed oxidant, H2O2, nor the antioxidants, total apparent ascorbic acid nor free proline, reflected the oxidative stress situations properly. In addition to the stress, HS exposure extended the mean lifespan of algae-fed D. magna, but at the expense of offspring numbers; so did also the pure yeast diet as compared to the algae diet. The first lifespan extension can be explained by the potential of HSs to block the pathway via the insulin-like growth factor 1 (IGF), whereas the second matches the, in aging papers, well described, but mechanistically poorly understood caloric restriction. Yeast-fed animals, exposed to HSs changed the energy allocation by reducing life span, but increasing offspring numbers. With the lifespan and offspring numbers, ecologically relevant parameters are differently affected by the simultaneous action of two environmentally relevant stressors.
The individual functional traits of different species play a key role for ecosystem function in aquatic and terrestrial systems. We modeled a multispecies predator-prey system with functionally different predator and prey species based on observations of the community dynamics of ciliates and their algal prey in Lake Constance. The model accounted for differences in predator feeding preferences and prey susceptibility to predation, and for the respective trade-offs. A low food demand of the predator was connected to a high food selectivity, and a high growth rate of the prey was connected to a high vulnerability to grazing. The data and the model did not show standard uniform predator- prey cycles, but revealed both complex dynamics and a coexistence of predator and prey at high biomass levels. These dynamics resulted from internally driven alternations in species densities and involved compensatory dynamics between functionally different species. Functional diversity allowed for ongoing adaptation of the predator and prey communities to changing environmental conditions such as food composition and grazing pressure. The trade-offs determined whether compensatory or synchronous dynamics occurred which influence the variability at the community level. Compensatory dynamics were promoted by a joint carrying capacity linking the different prey species which is particularly relevant at high prey biomasses, i.e., when grazers are less efficient. In contrast, synchronization was enhanced by the coupling of the different predator and prey species via common feeding links, e.g., by a high grazing pressure of a nonselective predator. The communities had to be functionally diverse in terms of their trade-offs and their traits to yield compensatory dynamics. Rather similar predator species tended to cycle synchronously, whereas profoundly different species did not coexist. Compensatory dynamics at the community level thus required intermediately strong tradeoffs for functional traits in both predators and their prey.
CsMan26 from Caldicellulosiruptor strain Rt8.B4 is a modular beta-mannanase consisting of two N-terminal family 27 carbohydrate-binding modules (CBMs), followed by a family 35 CBM and a family 26 glycoside hydrolase catalytic module (mannanase). A functional dissection of the full-length CsMan26 and a comprehensive characterisation of the truncated derivatives were undertaken to evaluate the role of the CBMs. Limited proteolysis was used to define biochemically the boundaries of the different structural modules in CsMan26. The full-length CsMan26 and three truncated derivatives were produced in Escherichia coli, purified and characterised. The systematic removal of the CBMs resulted in a decrease in the optimal temperature for activity and in the overall thermostability of the derivatives. Kinetic experiments indicated that the presence of the mannan-specific family 27 CBMs increased the affinity of the enzyme towards the soluble galactomannan substrate but this was accompanied by lower catalytic efficiency. The full-length CsMan26 and its truncated derivatives were unable to hydrolyse mannooligosaccharides with degree of polymerisation (DP) of three or less. The major difference in the hydrolysis pattern of larger mannooligosaccharides (DP > 3) by the derivatives was determined by their abilities to further hydrolyse the intermediate sugar mannotetraose.
COR15A and COR15B form a tandem repeat of highly homologous genes in Arabidopsis thaliana. Both genes are highly cold induced and the encoded proteins belong to the Pfam LEA_4 group (group 3) of the late embryogenesis abundant (LEA) proteins. Both proteins were predicted to be intrinsically disordered in solution. Only COR15A has previously been characterized and it was shown to be localized in the soluble stroma fraction of chloroplasts. Ectopic expression of COR15A in Arabidopsis resulted in increased freezing tolerance of both chloroplasts after freezing and thawing of intact leaves and of isolated protoplasts frozen and thawed in vitro. In the present study we have generated recombinant mature COR15A and COR15B for a comparative study of their structure and possible function as membrane protectants. CD spectroscopy showed that both proteins are predominantly unstructured in solution and mainly a-helical after drying. Both proteins showed similar effects on the thermotropic phase behavior of dry liposomes. A decrease in the gel to liquid-crystalline phase transition temperature depended on both the unsaturation of the fatty acyl chains and lipid headgroup structure. FTIR spectroscopy indicated no strong interactions between the proteins and the lipid phosphate and carbonyl groups, but significant interactions with the galactose headgroup of the chloroplast lipid monogalactosyldiacylglycerol. These findings were rationalized by modeling the secondary structure of COR15A and COR15B. Helical wheel projection indicated the presence of amphipathic a-helices in both proteins. The helices lacked a clear separation of positive and negative charges on the hydrophilic face, but contained several hydroxylated amino acids.
Serotonin plays a key role in modulating various physiological and behavioral processes in both protostomes and deuterostomes. The vast majority of serotonin receptors belong to the superfamily of G-protein-coupled receptors. We report the cloning of a cDNA from the honeybee (Am5-ht1A) sharing high similarity with members of the 5-HT1 receptor class. Activation of Am5-HT1A by serotonin inhibited the production of cAMP in a dose-dependent manner (EC50 = 16.9 nM). Am5-HT1A was highly expressed in brain regions known to be involved in visual information processing. Using in vivo pharmacology, we could demonstrate that Am5-HT1A receptor ligands had a strong impact on the phototactic behavior of individual bees. The data presented here mark the first comprehensive study-from gene to behavior-of a 5-HT1A receptor in the honeybee, paving the way for the eventual elucidation of additional roles of this receptor subtype in the physiology and behavior of this social insect.
Malignant gliomas are a fatal disease lacking sufficient possibilities for early diagnosis and chemical markers to detect remission or relapse. The recruitment of progenitor cells such as mesenchymal stem cells (MSC) is a main feature of gliomas. Stromal cell-derived factor-1 (SDF-1), a chemokine produced in glioma cell lines, enhances migration in MSC and has been associated with cell survival and apoptosis in gliomas. Therefore, this study was performed to evaluate (i) whether SDF-1 and its receptors are expressed in human malignant gliomas in situ and (ii) if SDF-1 might potentially play a role in recruiting MSCs into human glioma. In glioblastoma tissue, immunohistochemistry revealed that SDF-1 and its receptor CXCR4 are expressed in regions of angiogenesis and necrosis, and qPCR showed that SDF-1 is elevated. Public expression data indicated that CXCR4 was upregulated. The latter data also illustrate that SDF-1 could be up- or downregulated in glioma compared to normal brain in a transcript-specific manner. In plasma, SDF-1 is elevated in glioma patients. The level is reduced by both dexamethasone intake and surgery. Dexamethasone also decreased SDF-1 production in cells in vitro. The undirected migration of human MSC (hMSC) was not enhanced by the addition of SDF-1. However, SDF-1 stimulated directed invasion of hMSC in a dose-dependent manner. Taken together, we show that SDF-1 is a potent chemoattractant of progenitor cells such as hMSCs and that its expression is elevated in glioma tissue, which results in elevated SDF-1 levels in the patient's plasma samples with concomittant decrease after tumor resection. The fact that elevated SDF-1 plasma levels are significantly decreased after tumor surgery could be a first hint that SDF-1 might act as tumor marker for malignant gliomas in order to detect disease progression or remission, respectively.
Lake morphometry and wind exposure may shape the plankton community structure in acidic mining lakes
(2010)
Acidic mining lakes (pH <3) are specific habitats exhibiting particular chemical and biological characteristics. The species richness is low and mixotrophy and omnivory are common features of the plankton food web in such lakes. The plankton community structure of mining lakes of different morphometry and mixing type but similar chemical characteristics (Lake 130, Germany and Lake Langau, Austria) was investigated. The focus was laid on the species composition, the trophic relationship between the phago-mixotrophic flagellate Ochromonas sp. and bacteria and the formation of a deep chlorophyll maximum along a vertical pH-gradient. The shallow wind-exposed Lake 130 exhibited a higher species richness than Lake Langau. This increase in species richness was made up mainly by mero-planktic species, suggesting a strong benthic/littoral - pelagic coupling. Based on the field data from both lakes, a nonlinear, negative relation between bacteria and Ochromonas biomass was found, suggesting that at an Ochromonas biomass below 50 mu g CL-1. the grazing pressure on bacteria is low and with increasing Ochromonas biomass bacteria decline. Furthermore, in Lake Langau, a prominent deep chlorophyll maximum was found with chlorophyll concentrations ca. 50 times higher than in the epilimnion which was build up by the euglenophyte Lepocinclis sp. We conclude that lake morphometry, and specific abiotic characteristics such as mixing behaviour influence the community structure in these mining lakes.
The population status of the harbour porpoise (Phocoena phocoena) in the Baltic area has been a continuous matter of debate. Here we present the by far most comprehensive genetic population structure assessment to date for this region, both with regard to geographic coverage and sample size: 497 porpoise samples from North Sea, Skagerrak, Kattegat, Belt Sea, and Inner Baltic Sea were sequenced at the mitochondrial Control Region and 305 of these specimens were typed at 15 polymorphic microsatellite loci. Samples were stratified according to sample type (stranding vs. by- caught), sex, and season (breeding vs. non-breeding season). Our data provide ample evidence for a population split between the Skagerrak and the Belt Sea, with a transition zone in the Kattegat area. Among other measures, this was particularly visible in significant frequency shifts of the most abundant mitochondrial haplotypes. A particular haplotype almost absent in the North Sea was the most abundant in Belt Sea and Inner Baltic Sea. Microsatellites yielded a similar pattern (i.e., turnover in occurrence of clusters identified by STRUCTURE). Moreover, a highly significant association between microsatellite assignment and unlinked mitochondrial haplotypes further indicates a split between North Sea and Baltic porpoises. For the Inner Baltic Sea, we consistently recovered a small, but significant separation from the Belt Sea population. Despite recent arguments that separation should exceed a predefined threshold before populations shall be managed separately, we argue in favour of precautionary acknowledging the Inner Baltic porpoises as a separate management unit, which should receive particular attention, as it is threatened by various factors, in particular local fishery measures.
Question Which mechanisms promote the maintenance of the protected pioneer grass Corynephorus canescens in a mosaic landscape? Which are the interactive effects of small-scale disturbances, successional stage and year-to-year variation on early establishment probabilities of C. canescens? Location Brandenburg, NE Germany. Methods We measured emergence and survival rates over 3 yr in a sowing-experiment conducted in three successional stages (C. canescens- dominated site, ruderal forb site and pioneer forest) under two different regimes of mechanical ground disturbance (disturbed versus undisturbed control). Results Overall, disturbance led to higher emergence in a humid year and to lower emergence in a very dry year. Apparently, when soil moisture was sufficient, the main factor limiting C. canescens' establishment was competition, while in the dry year, water became the limiting factor. Survival rates were not affected by disturbance. In humid years, C. canescens emerged in higher numbers in open successional stages while in the dry year, emergence rates were higher in late stages, suggesting an important role of late successional stages for the persistence of C. canescens. Conclusions Our results suggest that small-scale disturbances can promote germination of C. canescens. However, disturbances should be carefully planned. The optimal strategy for promoting C. canescens is to apply disturbances just before seed dispersal and not during dry years. At the landscape scale, a mosaic of different vegetation types is beneficial for the protected pioneer grass as facilitation by late-successional species may be an important mechanism for the persistence of C. canescens, especially in dry years.
The interplay between turnover or degradation and ribosome loading of messenger RNA (mRNA) is studied theoretically using a stochastic model that is motivated by recent experimental results. Random mRNA degradation affects the statistics of polysomes, i.e., the statistics of the number of ribosomes per mRNA as extracted from cells. Since ribosome loading of newly created mRNA chains requires some time to reach steady state, a fraction of the extracted mRNA/ ribosome complexes does not represent steady state conditions. As a consequence, the mean ribosome density obtained from the extracted complexes is found to be inversely proportional to the mRNA length. On the other hand, the ribosome density profile shows an exponential decrease along the mRNA for prokaryotes and becomes uniform in eukaryotic cells. Copyright (C) EPLA, 2010
The biogenic amine octopamine and its biological precursor tyramine are thought to be the invertebrate functional homologues of the vertebrate adrenergic transmitters. Octopamine functions as a neuromodulator, neurotransmitter and neurohormone in insect nervous systems and prompts the whole organism to "dynamic action". A growing number of studies suggest a prominent role for octopamine in modulating multiple physiological and behavioural processes in invertebrates, as for example the phase transition in Schistocerca gregaria. Both octopamine and tyramine exert their effects by binding to specific receptor proteins that belong to the superfamily of G protein-coupled receptors. Since these receptors do not appear to be present in vertebrates, they may present very suitable and specific insecticide and acaricide targets. (C) 2010 Elsevier Ltd. All rights reserved.
The biogenic amine octopamine functions as a neuromodulator, neurotransmitter and neurohormone in insect nervous systems. It plays a prominent role in modulating multiple physiological and behavioural processes in invertebrates. Octopamine exerts its effects by binding to specific receptor proteins that belong to the superfamily of G protein-coupled receptors. We found two partial sequences of putative octopamine receptors in the desert locust Schistocerca gregaria (SgOct alpha R and SgOct beta R) and investigated their transcript levels in males and females of both phases and during the transition between long-term solitarious and gregarious locusts. The transcript levels of SgOctaR are the highest in the central nervous system, whereas those of SgOct beta R are the highest in the flight muscles, followed by the central nervous system. Both SgOct alpha R and SgOct beta R show higher transcript levels in long-term gregarious locusts as compared to solitarious ones. The rise of SgOct beta R transcript levels already appears during the first 4 h of gregarisation, during which also the behavioural changes take place.
The thrombin-like serine protease TLBm from Bothrops marajoensis was isolated in one chromatographic step in reverse phase HPLC. Its molecular mass was 33239.95 Da, as based on the determined primary structure and confirmed experimentally by MALDI-TOF mass spectrometry (33332.5 Da) and it contains 12 half-cysteine residues. This TLBm exhibited high specificity for BA rho NA, Michaelis-Menten behavior with K-m 2.3 x 10(-1) M and the V-max 0.52 x 10(-1) nmoles rho-NA/lt/min for this substrate. TLBm also showed ability to coagulate bovine fibrinogen and was inhibited by soybean trypsin inhibitor, EDTA and S(Dm) from the serum of the species Didelphis marsupialis. The primary structure of TLBm showed the presence of His(45), Asp(103) and Ser(228) residues in the corresponding positions of the catalytic triad established in the serine proteases and Ser(228) are inhibited by phenylmethylsulfonyl fluoride (PMSF). Amino acid analysis showed a high content of Asp, Glu, Gly, Set, Ala and Pro as well as 12 half-cysteine residues and calculated pI of 6.47; TLBm presented 285 amino acid residues. In this work, we investigated the ability of TLBm to degrade fibrinogen and we observed that it is able to cause alpha- and beta-chain cleavage. Enzymatic as well as the platelet aggregation activities were strongly inhibited when incubated with PMSF, a specific inhibitor of serine protease. Also, TLBm induced platelet aggregation in washed and platelet-rich plasma, and in both cases, PMSF inhibited its activity.
Current reversal is an intriguing phenomenon that has been central to recent experimental and theoretical investigations of transport based on ratchet mechanism. By considering a system of two interacting ratchets, we demonstrate how the coupling can be used to control the reversals. In particular, we find that current reversal that exists in a single driven ratchet system can ultimately be eliminated with the presence of a second ratchet. For specific coupling strengths a current-reversal free regime has been detected. Furthermore, in the fully synchronized state characterized by the coupling threshold k(th), a specific driving amplitude a(opt) is found for which the transport is optimum.
Calcineurin activity augments cAMP/PKA-dependent activation of V-ATPase in blowfly salivary glands
(2010)
We have examined the role of the Ca2+-dependent protein phosphatase 2B (calcineurin) in the regulation of the vacuolar H+-ATPase (V-ATPase) in blowfly salivary glands. In response to the neurohormone serotonin [5-hydroxytryptamine (5-HT)] and under the mediation of the cAMP/PKA signaling pathway, the secretory cells assemble and activate V-ATPase molecules at the apical membrane. We demonstrate that the inhibition of calcineurin activity by cyclosporin A, by FK- 506, or by prevention of the elevation of Ca2+ diminishes the 5-HT-induced assembly and activation of V-ATPase. The effect of calcineurin on V-ATPase is mediated by the cAMP/PKA signaling pathway, with calcineurin acting upstream of PKA, because 1) cyclosporin A does not influence the 8-(4-chlorophenylthio) adenosine-3',5'-cyclic monophosphate (8-CPT-cAMP)-induced activation of V-ATPase, and 2) the 5-HT-induced rise in cAMP is highly reduced in the presence of cyclosporin A. Moreover, a Ca2+ rise evoked by the sarco(endo) plasmic reticulum Ca2+-ATPase (SERCA) inhibitor cyclopiazonic acid leads to an increase in intracellular cAMP concentration and a calcineurin-mediated PKA- dependent activation of V-ATPase. We propose that calcineurin activity mediates cross talk between the inositol 1,4,5- trisphosphate/Ca2+ and the cAMP/PKA signaling pathways, thereby augmenting the 5-HT-induced rise in cAMP and thus the cAMP/PKA-mediated activation of V-ATPase.