Analysis of the influence of epitope flanking regions on MHC class I restricted antigen presentation

Peptides presented by MHC class I molecules for CTL recognition are derived mainly from cytosolic proteins. For antigen presentation on the cell surface, epitopes require correct processing by cytosolic and ER proteases, efficient TAP transport and MHC class I binding affinity. The efficiency of epitope generation depends not only on the epitope itself, but also on its flanking regions. In this project, the influence of the C-terminal region of the model epitope SIINFEKL (S8L) from chicken ovalbumin (aa 257-264) on antigen processing has been investigated. S8L is a well characterized epitope presented on the murine MHC class I molecule, H-2Kb. The Flp-In 293Kb cell line was transfected with different constructs each enabling the expression of the S8L sequence with different defined C-terminal flanking regions. The constructs differed at the two first C-terminal positions after the S8L epitope, so called P1’ and P2’. At these sites, all 20 amino acids were exchanged consecutively and tested for their influence on H-2Kb/S8L presentation on the cell surface of the Flp-In 293Kb cells. The detection of this complex was performed by immunostaining and flow cytometry. The prevailing assumption is that proteasomal cleavages are exclusively responsible for the generation of the final C-termini of CTL epitopes. Nevertheless, recent publications showed that TPPII (tripeptidyl peptidase II) is required for the generation of the correct C-terminus of the HLA-A3-restricted HIV epitope Nef(73-82). With this background, the dependence of the S8L generation on proteasomal cleavage of the designed constructs was characterized using proteasomal inhibitors. The results obtained indicate that it is crucial for proteasomal cleavage, which amino acid is flanking the C-terminus of an epitope. Furthermore, partially proteasome independent S8L generation from specific S8L-precursor peptides was observed. Hence, the possibility of other existing endo- or carboxy-peptidases in the cytosol that could be involved in the correct trimming of the C-terminus of antigenic peptides for MHC class I presentation was investigated, performing specific knockdowns and using inhibitors against the target peptidases. In parallel, a purification strategy to identify the novel peptidase was established. The purified peaks showing an endopeptidase activity were further analyzed by mass spectrometry and some potential peptidases (like e.g. Lon) were identified, which have to be further characterized.

Untersuchungen zur Photophysik einzelner konjugierter Polymerketten bei tiefen Temperaturen

Die spektroskopische Untersuchung einzelner konjugierter Polymermoleküle, welche multichromophore Systeme darstellen, bei 1,2K eröffnete den Zugang zu den unterschiedlichen Chromophoren. In Fluoreszenzemissions- wie -anregungsspektren des Poly(2-methoxy-5-(2’-ethylhexyloxy)-rnp-phenylenvinylen) (MEH-PPV) konnten Nullphononenlinien (rein elektronische (0-0)-Übergänge) emittierender Chromophore beobachtet werden, deren Breiten durch das experimentelle Auflösungsvermögen limitiert waren. Dadurch konnte eine obere Grenze für die homogene Linienbreite festgelegt werden. Infolge starker linearer Elektron-Phonon-Kopplung und spektraler Diffusion war die Beobachtung von Nullphononenlinien jedoch auf eine Minderheit der Chromophorernbeschränkt. In geeigneten Probesystemen konnten in Anregungsspektren auch Nullphononenlinien solcher Chromophore identifiziert werden, die als Donoren im intramolekularen Energietransfer fungieren. Aufgrund der dadurch verkürzten Fluoreszenzlebensdauer und der damit verbundenen Linienverbreiterung erlaubte ihre Untersuchung die Ermittlung von Energietransferraten. Eine bei Proben höherer Molmasse auftretenden niederenergetische Subpopulation an Emittern wurde auf Grundlage ihres Fluoreszenzemissions-, -anregungs- und -abklingverhaltens und unter Berücksichtigung theoretischer Arbeiten auf längere chromophore Einheiten zurückgeführt, die sich infolge von Packungseffekten in geordneten Regionen der Polymerkette ausbilden. Die schwächere lineare Elektron-Phonon-Kopplung des leiterartig verknüpften Polypentaphenylen (LPPentP) äußerte sich in einer erhöhten Wahrscheinlichkeit des Auftretens von Nullphononenlinien. Die genaue Bestimmung von Energietransferraten litt allerdings unter einem weiteren signifikanten Beitrag zur Linienbreite neben der Lebensdauer des angeregten Zustandes (möglicherweisernspektrale Diffusion oder schnelle Dephasierungsprozesse).

Main characteristics of the emission from elliptical galaxies

Elliptical galaxies are one of the most characteristic objects we can find in the sky. In order to unveil their properties, such as their structure or chemical composition, one must study their spectral emission. In fact they seem to behave rather differently when observed with different eyes. This is because their light is mainly brought by two different components: optical radiation arises from its stars, while the X emission is primarly due to a halo of extremely hot gas in which ellipticals seem to be embedded. After a brief classification, the two main processes linked to these phenomena will be described, together with the informations we can collect thanks to them. Eventually, we will take a quick look at the other regions of the electromagnetic spectrum.

Global modeling of pollutant transport and deposition from anthropogenic emission hot spots on global scale

Urban centers significantly contribute to anthropogenic air pollution, although they cover only a minor fraction of the Earth's land surface. Since the worldwide degree of urbanization is steadily increasing, the anthropogenic contribution to air pollution from urban centers is expected to become more substantial in future air quality assessments. The main objective of this thesis was to obtain a more profound insight in the dispersion and the deposition of aerosol particles from 46 individual major population centers (MPCs) as well as the regional and global influence on the atmospheric distribution of several aerosol types. For the first time, this was assessed in one model framework, for which the global model EMAC was applied with different representations of aerosol particles. First, in an approach with passive tracers and a setup in which the results depend only on the source location and the size and the solubility of the tracers, several metrics and a regional climate classification were used to quantify the major outflow pathways, both vertically and horizontally, and to compare the balance between pollution export away from and pollution build-up around the source points. Then in a more comprehensive approach, the anthropogenic emissions of key trace species were changed at the MPC locations to determine the cumulative impact of the MPC emissions on the atmospheric aerosol burdens of black carbon, particulate organic matter, sulfate, and nitrate. Ten different mono-modal passive aerosol tracers were continuously released at the same constant rate at each emission point. The results clearly showed that on average about five times more mass is advected quasi-horizontally at low levels than exported into the upper troposphere. The strength of the low-level export is mainly determined by the location of the source, while the vertical transport is mainly governed by the lifting potential and the solubility of the tracers. Similar to insoluble gas phase tracers, the low-level export of aerosol tracers is strongest at middle and high latitudes, while the regions of strongest vertical export differ between aerosol (temperate winter dry) and gas phase (tropics) tracers. The emitted mass fraction that is kept around MPCs is largest in regions where aerosol tracers have short lifetimes; this mass is also critical for assessing the impact on humans. However, the number of people who live in a strongly polluted region around urban centers depends more on the population density than on the size of the area which is affected by strong air pollution. Another major result was that fine aerosol particles (diameters smaller than 2.5 micrometer) from MPCs undergo substantial long-range transport, with about half of the emitted mass being deposited beyond 1000 km away from the source. In contrast to this diluted remote deposition, there are areas around the MPCs which experience high deposition rates, especially in regions which are frequently affected by heavy precipitation or are situated in poorly ventilated locations. Moreover, most MPC aerosol emissions are removed over land surfaces. In particular, forests experience more deposition from MPC pollutants than other land ecosystems. In addition, it was found that the generic treatment of aerosols has no substantial influence on the major conclusions drawn in this thesis. Moreover, in the more comprehensive approach, it was found that emissions of black carbon, particulate organic matter, sulfur dioxide, and nitrogen oxides from MPCs influence the atmospheric burden of various aerosol types very differently, with impacts generally being larger for secondary species, sulfate and nitrate, than for primary species, black carbon and particulate organic matter. While the changes in the burdens of sulfate, black carbon, and particulate organic matter show an almost linear response for changes in the emission strength, the formation of nitrate was found to be contingent upon many more factors, e.g., the abundance of sulfuric acid, than only upon the strength of the nitrogen oxide emissions. The generic tracer experiments were further extended to conduct the first risk assessment to obtain the cumulative risk of contamination from multiple nuclear reactor accidents on the global scale. For this, many factors had to be taken into account: the probability of major accidents, the cumulative deposition field of the radionuclide cesium-137, and a threshold value that defines contamination. By collecting the necessary data and after accounting for uncertainties, it was found that the risk is highest in western Europe, the eastern US, and in Japan, where on average contamination by major accidents is expected about every 50 years.

Antarctic cloud spectral emission from ground-based measurements, a focus on far infrared signatures

The present work belongs to the PRANA project, the first extensive field campaign of observation of atmospheric emission spectra covering the Far InfraRed spectral region, for more than two years. The principal deployed instrument is REFIR-PAD, a Fourier transform spectrometer used by us to study Antarctic cloud properties. A dataset covering the whole 2013 has been analyzed and, firstly, a selection of good quality spectra is performed, using, as thresholds, radiance values in few chosen spectral regions. These spectra are described in a synthetic way averaging radiances in selected intervals, converting them into BTs and finally considering the differences between each pair of them. A supervised feature selection algorithm is implemented with the purpose to select the features really informative about the presence, the phase and the type of cloud. Hence, training and test sets are collected, by means of Lidar quick-looks. The supervised classification step of the overall monthly datasets is performed using a SVM. On the base of this classification and with the help of Lidar observations, 29 non-precipitating ice cloud case studies are selected. A single spectrum, or at most an average over two or three spectra, is processed by means of the retrieval algorithm RT-RET, exploiting some main IR window channels, in order to extract cloud properties. Retrieved effective radii and optical depths are analyzed, to compare them with literature studies and to evaluate possible seasonal trends. Finally, retrieval output atmospheric profiles are used as inputs for simulations, assuming two different crystal habits, with the aim to examine our ability to reproduce radiances in the FIR. Substantial mis-estimations are found for FIR micro-windows: a high variability is observed in the spectral pattern of simulation deviations from measured spectra and an effort to link these deviations to cloud parameters has been performed.

Proteins encoded in genomic regions associated with immune-mediated disease physically interact and suggest underlying biology

Genome-wide association studies (GWAS) have defined over 150 genomic regions unequivocally containing variation predisposing to immune-mediated disease. Inferring disease biology from these observations, however, hinges on our ability to discover the molecular processes being perturbed by these risk variants. It has previously been observed that different genes harboring causal mutations for the same Mendelian disease often physically interact. We sought to evaluate the degree to which this is true of genes within strongly associated loci in complex disease. Using sets of loci defined in rheumatoid arthritis (RA) and Crohn's disease (CD) GWAS, we build protein-protein interaction (PPI) networks for genes within associated loci and find abundant physical interactions between protein products of associated genes. We apply multiple permutation approaches to show that these networks are more densely connected than chance expectation. To confirm biological relevance, we show that the components of the networks tend to be expressed in similar tissues relevant to the phenotypes in question, suggesting the network indicates common underlying processes perturbed by risk loci. Furthermore, we show that the RA and CD networks have predictive power by demonstrating that proteins in these networks, not encoded in the confirmed list of disease associated loci, are significantly enriched for association to the phenotypes in question in extended GWAS analysis. Finally, we test our method in 3 non-immune traits to assess its applicability to complex traits in general. We find that genes in loci associated to height and lipid levels assemble into significantly connected networks but did not detect excess connectivity among Type 2 Diabetes (T2D) loci beyond chance. Taken together, our results constitute evidence that, for many of the complex diseases studied here, common genetic associations implicate regions encoding proteins that physically interact in a preferential manner, in line with observations in Mendelian disease.

Brain activation during craving for alcohol measured by positron emission tomography

OBJECTIVE: Craving for alcohol is probably involved in acquisition and maintenance of alcohol dependence to a substantial degree. However, the brain substrates and mechanisms that underlie alcohol craving await more detailed elucidation. METHOD: Positron emission tomography was used to map regional cerebral blood flow (CBF) in 21 detoxified patients with alcohol dependence during exposure to alcoholic and non-alcoholic beverages. RESULTS: During the alcohol condition compared with the control condition, significantly increased CBF was found in the ventral putamen. Additionally, activated areas included insula, dorsolateral prefrontal cortex and cerebellum. Cerebral blood flow increase in these regions was related to self-reports of craving assessed in the alcoholic patients. CONCLUSIONS: In this investigation, cue-induced alcohol craving was associated with activation of brain regions particularly involved in brain reward mechanisms, memory and attentional processes. These results are consistent with studies on craving for other addictive substances and may offer strategies for more elaborate studies on the neurobiology of addiction.

Time-course of "off-line" prefrontal rTMS effects--a PET study.

Low-frequency "off-line" repetitive transcranial magnetic stimulation (rTMS) over the course of several minutes has attained considerable attention as a research tool in cognitive neuroscience due to its ability to induce functional disruptions of brain areas. This disruptive rTMS effect is highly valuable for revealing a causal relationship between brain and behavior. However, its influence on remote interconnected areas and, more importantly, the duration of the induced neurophysiological effects, remain unknown. These aspects are critical for a study design in the context of cognitive neuroscience. In order to investigate these issues, 12 healthy male subjects underwent 8 H(2)(15)O positron emission tomography (PET) scans after application of long-train low-frequency rTMS to the right dorsolateral prefrontal cortex (DLPFC). Immediately after the stimulation train, regional cerebral blood flow (rCBF) increases were present under the stimulation site as well as in other prefrontal cortical areas, including the ventrolateral prefrontal cortex (VLPFC) ipsilateral to the stimulation site. The mean increases in rCBF returned to baseline within 9 min. The duration of this unilateral prefrontal rTMS effect on rCBF is of particular interest to those who aim to influence behavior in cognitive paradigms that use an "off-line" approach.

Deficit actual tool use in schizophrenia is linked to structural alterations in key regions of planning and executing of tool use and connecting fibers

Introduction: Schizophrenia patients frequently suffer from complex motor abnormalities including fine and gross motor disturbances, abnormal involuntary movements, neurological soft signs and parkinsonism. These symptoms occur early in the course of the disease, continue in chronic patients and may deteriorate with antipsychotic medication. Furthermore gesture performance is impaired in patients, including the pantomime of tool use. Whether schizophrenia patients would show difficulties of actual tool use has not yet been investigated. Human tool use is complex and relies on a network of distinct and distant brain areas. We therefore aim to test if schizophrenia patients had difficulties in tool use and to assess associations with structural brain imaging using voxel based morphometry (VBM) and tract based spatial statistics (TBSS). Methode: In total, 44 patients with schizophrenia (DSM-5 criteria; 59% men, mean age 38) underwent structural MR imaging and performed the Tool-Use test. The test examines the use of a scoop and a hammer in three conditions: pantomime (without the tool), demonstration (with the tool) and actual use (with a recipient object). T1-weighted images were processed using SPM8 and DTI-data using FSL TBSS routines. To assess structural alterations of impaired tool use we first compared gray matter (GM) volume in VBM and white matter (WM) integrity in TBSS data of patients with and without difficulties of actual tool use. Next we explored correlations of Tool use scores and VBM and TBSS data. Group comparisons were family wise error corrected for multiple tests. Correlations were uncorrected (p < 0.001) with a minimum cluster threshold of 17 voxels (equivalent to a map-wise false positive rate of alpha < 0.0001 using a Monte Carlo procedure). Results: Tool use was impaired in schizophrenia (43.2% pantomime, 11.6% demonstration, 11.6% use). Impairment was related to reduced GM volume and WM integrity. Whole brain analyses detected an effect in the SMA in group analysis. Correlations of tool use scores and brain structure revealed alterations in brain areas of the dorso-dorsal pathway (superior occipital gyrus, superior parietal lobule, and dorsal premotor area) and the ventro-dorsal pathways (middle occipital gyrus, inferior parietal lobule) the action network, as well as the insula and the left hippocampus. Furthermore, significant correlations within connecting fiber tracts - particularly alterations within the bilateral corona radiata superior and anterior as well as the corpus callosum -were associated with Tool use performance. Conclusions: Tool use performance was impaired in schizophrenia, which was associated with reduced GM volume in the action network. Our results are in line with reports of impaired tool use in patients with brain lesions particularly of the dorso-dorsal and ventro-dorsal stream of the action network. In addition an effect of tool use on WM integrity was shown within fiber tracts connecting regions important for planning and executing tool use. Furthermore, hippocampus is part of a brain system responsible for spatial memory and navigation.The results suggest that structural brain alterations in the common praxis network contribute to impaired tool use in schizophrenia.

Influence of Martian crustal magnetic anomalies on the emission of energetic neutral hydrogen atoms

We analyze the data on hydrogen energetic neutral atoms (ENAs) emissions from the dayside of Mars, recorded by a Neutral Particle Detector of the Analyzer of Space Plasmas and Energetic Atoms aboard Mars Express from 14 March to 9 July 2004. We first identify and analyze events of the ENA flux enhancement coinciding with the presence of the crustal magnetic anomalies on the dayside of Mars. We then backtrace the ENA emissions to the lower altitudes (source region) and build up an average map of the flux intensities in the geographic coordinates with all the available data. The map shows a peak-to-valley ENA flux enhancement of 40%–90% close to the crustal magnetic anomaly regions. These results suggest the influence of the magnetic anomalies on the ENA emission from the dayside of Mars. The enhancement may result from the deviation of the highly directional plasma flow above anomalies toward the detectors such that more charge exchange ENAs would be recorded. Alternatively, higher exospheric densities above the anomalies would also result in an increase of the charge exchange ENA flux.

INCREASED EXPRESSION OF ONE OF TWO ADENOSINE DEAMINASE ALLELES IN A HUMAN CHORIOCARCINOMA CELL LINE FOLLOWING SELECTION WITH ADENINE NUCLEOSIDES

The human choriocarcinoma cell line JEG-3 is heterozygous at the adenosine deaminase (ADA) gene locus. Both allelic genes are under strong but incomplete repression causing a very low level expression of the gene locus. Because cytotoxic adenosine analogues such as 9-(beta)-D arabinofuranosyladenine (ara-A) and 9-(beta)-D xylofuranosyladenine (xyl-A) can be specifically detoxified by the action of ADA, these analogues were used to select for JEG-3 derived cells which had increased ADA expression. When JEG-3 cells were subjected to a multi-step, successively increasing dosage of either ara-A or xyl-A, resistant cells with increased ADA expression were generated. This increased ADA expression in the resistant cells was unstable, so that when the selective pressure was removed, cellular ADA expression would decrease. Subclone analysis of xyl-A resistant cells revealed that compared to parental JEG-3 cells, individual resistant cells had either elevated ADA levels or decreased adenosine kinase (ADK) levels or both. This altered ADA and ADK expression in the resistant cells were found to be independent events. Because of high endogenous tissue conversion factor (TCF) expression in the JEG-3 cells, the allelic nature of the increased ADA expression in most of the resistant cells could not be determined. However, several resistant subcloned cells were found to have lost TCF expression. These TCF('-) cells expressed only the ADA*2 allelic gene product. Cell fusion experiments demonstrated that the ADA*1 allelic gene was intact and functional in the A3-1A7 cell line. Chromosomal analysis of the A3-1A7 cells showed that they had no double-minutes or homogeneously staining chromosomal regions, although a pair of new chromosomes were found in these cells. Segregation analysis of the hybrid cells indicated that an ADA*2 allelic gene was probably located on this new chromosome. The analysis of the A3-1A7 cell line suggested that the expression of only ADA 2 in these cells was the result of possibly a cis-deregulation of the ADA gene locus or more probably an amplification of the ADA*2 allelic gene. Two effective positive selection systems for ADA('+) cells were also developed and tested. These selection systems should eventually lead to the isolation of the ADA gene.^

Simulated Bromoform emission and concentration using the ocean biogeochemistry model MPIOM/HAMOCC forced by 6-hourly NCEP data

Bromoform (CHBr3) is one important precursor of atmospheric reactive bromine species that are involved in ozone depletion in the troposphere and stratosphere. In the open ocean bromoform production is linked to phytoplankton that contains the enzyme bromoperoxidase. Coastal sources of bromoform are higher than open ocean sources. However, open ocean emissions are important because the transfer of tracers into higher altitude in the air, i.e. into the ozone layer, strongly depends on the location of emissions. For example, emissions in the tropics are more rapidly transported into the upper atmosphere than emissions from higher latitudes. Global spatio-temporal features of bromoform emissions are poorly constrained. Here, a global three-dimensional ocean biogeochemistry model (MPIOM-HAMOCC) is used to simulate bromoform cycling in the ocean and emissions into the atmosphere using recently published data of global atmospheric concentrations (Ziska et al., 2013) as upper boundary conditions. Our simulated surface concentrations of CHBr3 match the observations well. Simulated global annual emissions based on monthly mean model output are lower than previous estimates, including the estimate by Ziska et al. (2013), because the gas exchange reverses when less bromoform is produced in non-blooming seasons. This is the case for higher latitudes, i.e. the polar regions and northern North Atlantic. Further model experiments show that future model studies may need to distinguish different bromoform-producing phytoplankton species and reveal that the transport of CHBr3 from the coast considerably alters open ocean bromoform concentrations, in particular in the northern sub-polar and polar regions.

Simulated Methyl iodide emission and concentration using the ocean biogeochemistry model MPIOM/HAMOCC forced by OMIP data

Production pathways of the prominent volatile organic halogen compound methyl iodide (CH3I) are not fully understood. Based on observations, production of CH3I via photochemical degradation of organic material or via phytoplankton production has been proposed. Additional insights could not be gained from correlations between observed biological and environmental variables or from biogeochemical modeling to identify unambiguously the source of methyl iodide. In this study, we aim to address this question of source mechanisms with a three-dimensional global ocean general circulation model including biogeochemistry (MPIOM-HAMOCC (MPIOM - Max Planck Institute Ocean Model HAMOCC - HAMburg Ocean Carbon Cycle model)) by carrying out a series of sensitivity experiments. The simulated fields are compared with a newly available global data set. Simulated distribution patterns and emissions of CH3I differ largely for the two different production pathways. The evaluation of our model results with observations shows that, on the global scale, observed surface concentrations of CH3I can be best explained by the photochemical production pathway. Our results further emphasize that correlations between CH3I and abiotic or biotic factors do not necessarily provide meaningful insights concerning the source of origin. Overall, we find a net global annual CH3I air-sea flux that ranges between 70 and 260 Gg/yr. On the global scale, the ocean acts as a net source of methyl iodide for the atmosphere, though in some regions in boreal winter, fluxes are of the opposite direction (from the atmosphere to the ocean).

Spatial spillover effects in determining China's regional CO2 emission growth : 2007-2010

This paper proposes an alternative input-output based spatial-structural decomposition analysis to elucidate the role of domestic-regional heterogeneity and interregional spillover effects in determining China's regional CO2 emission growth. Our empirical results based on the 2007 and 2010 Chinese interregional input-output tables show that the changes in most regions' final demand scale, final expenditure structure and export scale give positive spatial spillover effects on other regions' CO2 emission growth, the changes in most regions' consumption and export preference help the reduction of other regions' CO2 emissions, the changes in production technology, and investment preference may give positive or negative impacts on other region's CO2 emission growth through domestic supply chains. For some regions, the aggregate spillover effect from other regions may be larger than the intra-regional effect in determining regional emission growth. All these facts can significantly help better and deeper understanding on the driving forces of China's regional CO2 emission growth, thus can enrich the policy implication concerning a narrow definition of "carbon leakage" through domestic-interregional trade, and relevant political consensus about the responsibility sharing between developed and developing regions inside China.