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From its first use in the field of biochemistry, instrumental analysis offered a variety of invaluable tools for the comprehensive description of biological systems. Multi-selective methods that aim to cover as many endogenous compounds as possible in biological samples use different analytical platforms and include methods like gene expression profile and metabolite profile analysis. The enormous amount of data generated in application of profiling methods needs to be evaluated in a manner appropriate to the question under investigation. The new field of system biology rises to the challenge to develop strategies for collecting, processing, interpreting, and archiving this vast amount of data; to make those data available in form of databases, tools, models, and networks to the scientific community. On the background of this development a multi-selective method for the determination of phytohormones was developed and optimised, complementing the profile analyses which are already in use (Chapter I). The general feasibility of a simultaneous analysis of plant metabolites and phytohormones in one sample set-up was tested by studies on the analytical robustness of the metabolite profiling protocol. The recovery of plant metabolites proved to be satisfactory robust against variations in the extraction protocol by using common extraction procedures for phytohormones; a joint extraction of metabolites and hormones from plant tissue seems practicable (Chapter II). Quantification of compounds within the context of profiling methods requires particular scrutiny (Chapter II). In Chapter III, the potential of stable-isotope in vivo labelling as normalisation strategy for profiling data acquired with mass spectrometry is discussed. First promising results were obtained for a reproducible quantification by stable-isotope in vivo labelling, which was applied in metabolomic studies. In-parallel application of metabolite and phytohormone analysis to seedlings of the model plant Arabidopsis thaliana exposed to sulfate limitation was used to investigate the relationship between the endogenous concentration of signal elements and the ‘metabolic phenotype’ of a plant. An automated evaluation strategy was developed to process data of compounds with diverse physiological nature, such as signal elements, genes and metabolites – all which act in vivo in a conditional, time-resolved manner (Chapter IV). Final data analysis focussed on conditionality of signal-metabolome interactions.
This work explores the equilibrium structure and thermodynamic phase behavior of complexes formed by charged polymer chains (polyelectrolytes) and oppositely charged spheres (macroions). Polyelectrolyte-macroion complexes form a common pattern in soft-matter physics, chemistry and biology, and enter in numerous technological applications as well. From a fundamental point of view, such complexes are interesting in that they combine the subtle interplay between electrostatic interactions and elastic as well as entropic effects due to conformational changes of the polymer chain, giving rise to a wide range of structural properties. This forms the central theme of theoretical studies presented in this thesis, which concentrate on a number of different problems involving strongly coupled complexes, i.e. complexes that are characterized by a large adsorption energy and small chain fluctuations. In the first part, a global analysis of the structural phase behavior of a single polyelectrolyte-macroion complex is presented based on a dimensionless representation, yielding results that cover a wide range of realistic system parameters. Emphasize is made on the interplay between the effects due to the polyelectrolytes chain length, salt concentration and the macroion charge as well as the mechanical chain persistence length. The results are summarized into generic phase diagrams characterizing the wrapping-dewrapping behavior of a polyelectrolyte chain on a macroion. A fully wrapped chain state is typically obtained at intermediate salt concentrations and chain lengths, where the amount of polyelectrolyte charge adsorbed on the macroion typically exceeds the bare macroion charge leading thus to a highly overcharged complex. Perhaps the most striking features occur when a single long polyelectrolyte chain is complexed with many oppositely charged spheres. In biology, such complexes form between DNA (which carries the cell's genetic information) and small oppositely charged histone proteins serving as an efficient mechanism for packing a huge amount of DNA into the micron-size cell nucleus in eucaryotic cells. The resultant complex fiber, known as the chromatin fiber, appears with a diameter of 30~nm under physiological conditions. Recent experiments indicate a zig-zag spatial arrangement for individual DNA-histone complexes (nucleosome core particles) along the chromatin fiber. A numerical method is introduced in this thesis based on a simple generic chain-sphere cell model that enables one to investigate the mechanism of fiber formation on a systematic level by incorporating electrostatic and elastic contributions. As will be shown, stable complex fibers exhibit an impressive variety of structures including zig-zag, solenoidal and beads-on-a-string patterns, depending on system parameters such as salt concentration, sphere charge as well as the chain contour length (per sphere). The present results predict fibers of compact zig-zag structure within the physiologically relevant regime with a diameter of about 30~nm, when DNA-histone parameters are adopted. In the next part, a numerical method is developed in order to investigate the role of thermal fluctuations on the structure and thermodynamic phase behavior of polyelectrolyte-macroion complexes. This is based on a saddle-point approximation, which allows to describe the experimentally observed reaction (or complexation) equilibrium in a dilute solution of polyelectrolytes and macroions on a systematic level. This equilibrium is determined by the entropy loss a single polyelectrolyte chain suffers as it binds to an oppositely charged macroion. This latter quantity can be calculated from the spectrum of polyelectrolyte fluctuations around a macroion, which is determined by means of a normal-mode analysis. Thereby, a stability phase diagram is obtained, which exhibits qualitative agreement with experimental findings. At elevated complex concentrations, one needs to account for the inter-complex interactions as well. It will be shown that at small separations, complexes undergo structural changes in such a way that positive patches from one complex match up with negative patches on the other. Furthermore, one of the polyelectrolyte chains may bridge between the two complexes. These mechanisms lead to a strong inter-complex attraction. As a result, the second virial coefficient associated with the inter-complex interaction becomes negative at intermediate salt concentrations in qualitative agreement with recent experiments on solutions of nucleosome core particles.
Connective ties in discourse: Three ERP studies on causal, temporal and concessive connective ties and their influence on language processing. Questions In four experiments the influence of lexical connectives such as " darum", therefore, " danach", afterwards, and " trotzdem", nevertheless, on the processing of short two-sentence discourses was examined and compared to the processing of deictical sentential adverbs such as " gestern", yesterday, and " lieber", rather. These latter words do not have the property of signaling a certain discourse relation between two sentences, as connective ties do. Three questions were central to the work: * Do the processing contrasts found between connective and non-connective elements extend to connective ties and deictical sentential adverbs (experiments 2 and 3)? * Does the semantic content of the connective ties play the primary role, i.e is the major distinction to be made indeed between connective and non-connective or instead between causal, temporal and concessive? * When precisely is the information provided by connective ties used? There is some evidence that connective ties can have an immediate influence on the integration of subsequent elements, but the end of the second sentences appears to play an important role as well: experiments 2, 3, and 4. Conclusions First of all, the theoretical distinction between connective and non-connective elements does indeed have " cognitive reality" . This has already been shown in previous studies. The present studies do however show, that there is also a difference between one-place discourse elements (deictical sentential adverbs) and two-place discourse elements, namely connective ties, since all experiments examining this contrast found evidence for qualitatively and quantitatively different processing (experiments 1, 2, and 3). Secondly, the semantic type of the connective ties also plays a role. This was not shown for the LAN, found for all connective ties when compared to non-connective elements, and consequently interpreted as a more abstract reflection of the integration of connective ties. There was also no difference between causal and temporal connective ties before the end of the discourses in experiment 3. However, the N400 found for incoherent discourses in experiment 2, larger for connective incoherent than non-connective incoherent discourses, as well as the P3b found for concessive connective ties in the comparison between causal and concessive connective ties gave reason to assume that the semantic content of connective ties is made use of in incremental processing, and that the relation signaled by the connective tie is the one that readers attempt to construct. Concerning when the information provided by connective ties is used, it appears as if connectivity is generally and obligatorily taken at face value. As long as the meaning of a connective tie did not conflict with a preferred canonical discourse relation, there were no differences found for varying connective discourses (experiment 3). However, the fact that concessive connective ties announce the need for a more complex text representation was recognized and made use of immediately (experiment 4). Additionally, a violation of the discourse relation resulted in more difficult semantic integration if a connective tie was present (experiment 2). It is therefore concluded here that connective ties influence processing immediately. This claim has to be modified somewhat, since the sentence-final elements suggested that connective ties trigger different integration processes than non-connective elements. It seems as if the answer to the question of when connective ties are processed is neither exclusively immediately nor exclusively afterwards, but that both viewpoints are correct. It is suggested here that before the end of a discourse economy plays a central role in that a canonical relation is assumed unless there is evidence to the contrary. A connective tie could have the function of reducing the dimensions evaluated in a discourse to the one signaled by the connective tie. At the end of the discourse the representation is evaluated and verified, and an integrated situation model constructed. Here, the complexity of the different discourse relations that connective ties can signal, is expressed.
Interkonversion zellulärer Signaltransduktionsprozesse durch Phosphorylierung und Redoxregulation
(2005)
Die trainingswissenschaftliche Diagnostik in den Kernbereichen Training, Wettkampf und Leistungsfähigkeit ist durch einen hohen Praxisbezug, eine ausgeprägte strukturelle Komplexität und vielseitige Wechselwirkungen der sportwissenschaftlichen Teilgebiete geprägt. Diese Eigenschaften haben in der Vergangenheit dazu geführt, dass zentrale Fragestellungen, wie beispielsweise die Maximierung der sportlichen Leistungsfähigkeit, eine ökonomische Trainingsgestaltung, eine effektive Talentauswahl und -sichtung oder die Modellbildung noch nicht vollständig gelöst werden konnten. Neben den bereits vorhandenen linearen Lösungsansätzen werden in dieser Arbeit Methoden aus dem Bereich der Neuronalen Netzwerke eingesetzt. Diese nichtlinearen Diagnoseverfahren sind besonders geeignet für die Analyse von Prozessabläufen, wie sie beispielsweise im Training vorliegen. Im theoretischen Teil werden zunächst Gemeinsamkeiten, Abhängigkeiten und Unterschiede in den Bereichen Training, Wettkampf und Leistungsfähigkeit untersucht sowie die Brücke zwischen trainingswissenschaftlicher Diagnostik und nichtlinearen Verfahren über die Begriffe der Interdisziplinarität und Integrativität geschlagen. Angelehnt an die Theorie der Neuronalen Netze werden anschließend die Grundlagenmodelle Perzeptron, Multilayer-Perzeptron und Selbstorganisierende Karten theoretisch erläutert. Im empirischen Teil stehen dann die nichtlineare Analyse von personalen Anforderungsstrukturen, Zustände der sportlichen Form und die Prognose sportlichen Talents - allesamt bei jugendlichen Leistungsschwimmerinnen und -schwimmern - im Mittelpunkt. Die nichtlinearen Methoden werden dabei einerseits auf ihre wissenschaftliche Aussagekraft überprüft, andererseits untereinander sowie mit linearen Verfahren verglichen.
In this thesis the magnetohydrodynamic jet formation and the effects of magnetic diffusion on the formation of axisymmetric protostellar jets have been investigated in three different simulation sets. The time-dependent numerical simulations have been performed, using the magnetohydrodynamic ZEUS-3D code.
Strong damped Lyman alpha absorption (DLA) lines seen spectra of distant quasar are believed to arise when the sight line to the quasar goes trough the disc of a galaxy or a proto galaxy. Most of the neutral matter in the universe is contained in these clouds of neutral hydrogen that cause the absorption lines. Hence these DLAs are reservoirs for the formation of stars and galaxies throughout the universe. Despite intensive efforts over more than two decades only few galaxies responsible for the DLAs have been found. The problem is that the galaxies that harbour the neutral clouds are not necessarily bright, and selecting galaxies based on absorption lines could well select different types of galaxies than found in large surveys. If we are to understand how galaxies form out of neutral gas clouds it is essential to locate the galaxies in which DLAs reside. This thesis explores the use of integral field spectroscopy (IFS) to observe quasars known to have strong absorption lines in their spectra. IFS allows us to obtain a spectrum at many spatial points close to the quasar, thus providing images and spectroscopy simultaneously. From the imaging part, we can directly identify objects, and from the spectroscopy we can calculate the distances to the objects. When the distance of the object found in emission matches the distance to the object that cause the DLA line, we have identified the absorbing galaxy. Using this technique, we have showed that we can successfully recover a few DLA galaxies known previously from the literature. In a survey aimed to increase the number of DLA galaxies we have identified eight new candidate DLA galaxies. The projected distances from the candidates to the quasar sight lines indicate that the DLA galaxies have sizes similar to local disc galaxies. Hence our results suggest that large discs may be present when the universe was just 2 billion years old. We furthermore find no differences between the sizes of the very distant DLA galaxies and those that are not so distant. The large sizes imply that their neutral hydrogen masses are also similar to those in local galaxies, but we argue that the DLA galaxies are not necessarily as luminous as the present day disc galaxies. Taking advantage of the three-dimensional view provided by the IFS data, the second part of this thesis investigates extended emission line regions arising in the quasar neighborhood. We find that extended emission line nebulae are common around quasars, and explore the effects that may be the cause. Some quasars are known to be powerful radio emitters while others are not detected at radio wavelengths. We find that significantly larger and brighter emission line nebulae are found around the quasars which have the brightest radio emission, and in particular those that have large radio jets. The existence of the nebulae can be interpreted as an interaction of the radio jet with the surrounding medium, but we can not rule out a scenario where there are density or temperature differences in the surrounding environment. Only for the brightest object, where additional velocity information can be derived from the IFS data, can we argue for an interaction. In conclusion the use of IFS to search for faint emission lines, both from point sources and extended nebulae provides exciting new results within the scientific areas studied here.
Nitrogen is an essential macronutrient for plants and nitrogen fertilizers are indispensable for modern agriculture. Unfortunately, we know too little about how plants regulate their use of soil nitrogen, to maximize fertilizers-N use by crops and pastures. This project took a dual approach, involving forward and reverse genetics, to identify N-regulators in plants, which may prove useful in the future to improve nitrogen-use efficiency in agriculture. To identify nitrogen-regulated transcription factor genes in Arabidopsis that may control N-use efficiency we developed a unique resource for qRT-PCR measurements on all Arabidpsis transcription factor genes. Using closely spaced, gene-specific primer pairs and SYBR® Green to monitor amplification of double-stranded DNA, transcript levels of 83% of all target genes could be measured in roots or shoots of young Arabidopsis wild-type plants. Only 4% of reactions produced non-specific PCR products, and 13% of TF transcripts were undetectable in these organs. Measurements of transcript abundance were quantitative over six orders of magnitude, with a detection limit equivalent to one transcript molecule in 1000 cells. Transcript levels for different TF genes ranged between 0.001-100 copies per cell. Real-time RT-PCR revealed 26 root-specific and 39 shoot-specific TF genes, most of which have not been identified as organ-specific previously. An enlarged and improved version of the TF qRT-PCR platform contains now primer pairs for 2256 Arabidopsis TF genes, representing 53 gene families and sub-families arrayed on six 384-well plates. Set-up of real-time PCR reactions is now fully robotized. One researcher is able to measure expression of all 2256 TF genes in a single biological sample in a just one working day. The Arabidopsis qRT-PCT platform was successfully used to identify 37 TF genes which transcriptionaly responded at the transcriptional level to N-deprivation or to nitrate per se. Most of these genes have not been characterized previously. Further selection of TF genes based on the responses of selected candidates to other macronutrients and abiotic stresses allowed to distinguish between TFs regulated (i) specifically by nitrogen (29 genes) (ii) regulated by general macronutrient or by salt and osmotic stress (6 genes), and (iii) responding to all major macronutrients and to abiotic stresses. Most of the N-regulated TF genes were also regulated by carbon. Further characterization of sixteen selected TF genes, revealed: (i) lack of transcriptional response to organic nitrogen, (ii) two major types of kinetics of induction by nitrate, (iii) specific responses for the majority of the genes to nitrate but not downstream products of nitrate assimilation. All sixteen TF genes were cloned into binary vectors for constitutive and ethanol inducible over expression, and the first generation of transgenic plants were obtained for almost all of them. Some of the plants constitutively over expressing TF genes under control of the 35S promoter revealed visible phenotypes in T1 generation. Homozygous T-DNA knock out lines were also obtained for many of the candidate TF genes. So far, one knock out line revealed a visible phenotype: retardation of flowering time. A forward genetic approach using an Arabidopsis ATNRT2.1 promoter : Luciferase reporter line, resulted in identification of eleven EMS mutant reporter lines affected in induction of ATNRT2.1 expression by nitrate. These lines could by divided in the following classes according to expression of other genes involved in primary nitrogen and carbon metabolism: (i) lines affected exclusively in nitrate transport, (ii) those affected in nitrate transport, acquisition, but also in glycolysis and oxidative pentose pathway, (iii) mutants affected moderately in nitrate transport, oxidative pentose pathway and glycolysis but not in primary nitrate assimilation. Thus, several different N-regulatory genes may have been mutated in this set of mutants. Map-based cloning has begun to identify the genes affected in these mutants.