@article{GebertSteffanDewenterMorettoetal.2019, author = {Gebert, Friederike and Steffan-Dewenter, Ingolf and Moretto, Philippe and Peters, Marcell K.}, title = {Climate rather than dung resources predict dung beetle abundance and diversity along elevational and land use gradients on Mt. Kilimanjaro}, series = {Journal of Biogeography}, volume = {47}, journal = {Journal of Biogeography}, number = {2}, doi = {10.1111/jbi.13710}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-204701}, pages = {371 -- 381}, year = {2019}, abstract = {Aim: While elevational gradients in species richness constitute some of the best depicted patterns in ecology, there is a large uncertainty concerning the role of food resource availability for the establishment of diversity gradients in insects. Here, we analysed the importance of climate, area, land use and food resources for determining diversity gradients of dung beetles along extensive elevation and land use gradients on Mt. Kilimanjaro, Tanzania. Location: Mt. Kilimanjaro, Tanzania. Taxon: Scarabaeidae (Coleoptera). Methods: Dung beetles were recorded with baited pitfall traps at 66 study plots along a 3.6 km elevational gradient. In order to quantify food resources for the dung beetle community in form of mammal defecation rates, we assessed mammalian diversity and biomass with camera traps. Using a multi-model inference framework and path analysis, we tested the direct and indirect links between climate, area, land use and mammal defecation rates on the species richness and abundance of dung beetles. Results: We found that the species richness of dung beetles declined exponentially with increasing elevation. Human land use diminished the species richness of functional groups exhibiting complex behaviour but did not have a significant influence on total species richness. Path analysis suggested that climate, in particular temperature and to a lesser degree precipitation, were the most important predictors of dung beetle species richness while mammal defecation rate was not supported as a predictor variable. Main conclusions: Along broad climatic gradients, dung beetle diversity is mainly limited by climatic factors rather than by food resources. Our study points to a predominant role of temperature-driven processes for the maintenance and origination of species diversity of ectothermic organisms, which will consequently be subject to ongoing climatic changes.}, language = {en} } @article{RequierPailletLarocheetal.2019, author = {Requier, Fabrice and Paillet, Yoan and Laroche, Fabienne and Rutschmann, Benjamin and Zhang, Jie and Lombardi, Fabio and Svoboda, Miroslav and Steffan-Dewenter, Ingolf}, title = {Contribution of European forests to safeguard wild honeybee populations}, series = {Conservation Letters}, volume = {13}, journal = {Conservation Letters}, number = {2}, doi = {10.1111/conl.12693}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-204407}, pages = {e12693}, year = {2019}, abstract = {Abstract Recent studies reveal the use of tree cavities by wild honeybee colonies in European forests. This highlights the conservation potential of forests for a highly threatened component of the native entomofauna in Europe, but currently no estimate of potential wild honeybee population sizes exists. Here, we analyzed the tree cavity densities of 106 forest areas across Europe and inferred an expected population size of wild honeybees. Both forest and management types affected the density of tree cavities. Accordingly, we estimated that more than 80,000 wild honeybee colonies could be sustained in European forests. As expected, potential conservation hotspots were identified in unmanaged forests, and, surprisingly, also in other large forest areas across Europe. Our results contribute to the EU policy strategy to halt pollinator declines and reveal the potential of forest areas for the conservation of so far neglected wild honeybee populations in Europe.}, language = {en} } @article{PattschullWalzGruendletal.2019, author = {Pattschull, Grit and Walz, Susanne and Gr{\"u}ndl, Marco and Schwab, Melissa and R{\"u}hl, Eva and Baluapuri, Apoorva and Cindric-Vranesic, Anita and Kneitz, Susanne and Wolf, Elmar and Ade, Carsten P. and Rosenwald, Andreas and von Eyss, Bj{\"o}rn and Gaubatz, Stefan}, title = {The Myb-MuvB complex is required for YAP-dependent transcription of mitotic genes}, series = {Cell Reports}, volume = {27}, journal = {Cell Reports}, number = {12}, doi = {10.1016/j.celrep.2019.05.071}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-202039}, pages = {3533-3546}, year = {2019}, abstract = {YAP and TAZ, downstream effectors of the Hippo pathway, are important regulators of proliferation. Here, we show that the ability of YAP to activate mitotic gene expression is dependent on the Myb-MuvB (MMB) complex, a master regulator of genes expressed in the G2/M phase of the cell cycle. By carrying out genome-wide expression and binding analyses, we found that YAP promotes binding of the MMB subunit B-MYB to the promoters of mitotic target genes. YAP binds to B-MYB and stimulates B-MYB chromatin association through distal enhancer elements that interact with MMB-regulated promoters through chromatin looping. The cooperation between YAP and B-MYB is critical for YAP-mediated entry into mitosis. Furthermore, the expression of genes coactivated by YAP and B-MYB is associated with poor survival of cancer patients. Our findings provide a molecular mechanism by which YAP and MMB regulate mitotic gene expression and suggest a link between two cancer-relevant signaling pathways.}, language = {en} } @article{RaheemTawfikeAbdelmohsenetal.2019, author = {Raheem, Dotsha J. and Tawfike, Ahmed F. and Abdelmohsen, Usama R. and Edrada-Ebel, RuAngelie and Fitzsimmons-Thoss, Vera}, title = {Application of metabolomics and molecular networking in investigating the chemical profile and antitrypanosomal activity of British bluebells (\(Hyacinthoides\) \(non-scripta\))}, series = {Scientific Reports}, volume = {9}, journal = {Scientific Reports}, doi = {10.1038/s41598-019-38940-w}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-224935}, pages = {2547, 1-13}, year = {2019}, abstract = {Bulb, leaf, scape and flower samples of British bluebells (Hyacinthoides non-scripta) were collected regularly for one growth period. Methanolic extracts of freeze-dried and ground samples showed antitrypanosomal activity, giving more than 50\% inhibition, for 20 out of 41 samples. High-resolution mass spectrometry was used in the dereplication of the methanolic extracts of the different plant parts. The results revealed differences in the chemical profile with bulb samples being distinctly different from all aerial parts. High molecular weight metabolites were more abundant in the flowers, shoots and leaves compared to smaller molecular weight ones in the bulbs. The anti-trypanosomal activity of the extracts was linked to the accumulation of high molecular weight compounds, which were matched with saponin glycosides, while triterpenoids and steroids occurred in the inactive extracts. Dereplication studies were employed to identify the significant metabolites via chemotaxonomic filtration and considering their previously reported bioactivities. Molecular networking was implemented to look for similarities in fragmentation patterns between the isolated saponin glycoside at m/z 1445.64 [M + formic-H](-) equivalent to C64H104O33 and the putatively found active metabolite at m/z 1283.58 [M + formic-H](-) corresponding to scillanoside L-1. A combination of metabolomics and bioactivity-guided approaches resulted in the isolation of a norlanostane-type saponin glycoside with antitrypanosoma I activity of 98.9\% inhibition at 20 mu M.}, language = {en} } @article{RiesSanderDeoletal.2019, author = {Ries, Lena K. and Sander, Bodo and Deol, Kirandeep K. and Letzelter, Marie-Annick and Strieter, Eric Robert and Lorenz, Sonja}, title = {Analysis of ubiquitin recognition by the HECT ligase E6AP provides insight into its linkage specificity}, series = {Journal of Biological Chemistry}, volume = {294}, journal = {Journal of Biological Chemistry}, number = {15}, doi = {10.1074/jbc.RA118.007014}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-226207}, pages = {6113-6129}, year = {2019}, abstract = {Deregulation of the HECT-type ubiquitin ligase E6AP (UBE3A) is implicated in human papilloma virus-induced cervical tumorigenesis and several neurodevelopmental disorders. Yet the structural underpinnings of activity and specificity in this crucial ligase are incompletely understood. Here, we unravel the determinants of ubiquitin recognition by the catalytic domain of E6AP and assign them to particular steps in the catalytic cycle. We identify a functionally critical interface that is specifically required during the initial formation of a thioester-linked intermediate between the C terminus of ubiquitin and the ligase-active site. This interface resembles the one utilized by NEDD4-type enzymes, indicating that it is widely conserved across HECT ligases, independent of their linkage specificities. Moreover, we uncover surface regions in ubiquitin and E6AP, both in the N- and C-terminal portions of the catalytic domain, that are important for the subsequent reaction step of isopeptide bond formation between two ubiquitin molecules. We decipher key elements of linkage specificity, including the C-terminal tail of E6AP and a hydrophilic surface region of ubiquitin in proximity to the acceptor site Lys-48. Intriguingly, mutation of Glu-51, a single residue within this region, permits formation of alternative chain types, thus pointing to a key role of ubiquitin in conferring linkage specificity to E6AP. We speculate that substrate-assisted catalysis, as described previously for certain RING-associated ubiquitin-conjugating enzymes, constitutes a common principle during linkage-specific ubiquitin chain assembly by diverse classes of ubiquitination enzymes, including HECT ligases.}, language = {en} } @article{PanzerBrychBatschaueretal.2019, author = {Panzer, Sabine and Brych, Annika and Batschauer, Alfred and Terpitz, Ulrich}, title = {Opsin 1 and Opsin 2 of the corn smut fungus ustilago maydis are green light-driven proton pumps}, series = {Frontiers in Microbiology}, volume = {10}, journal = {Frontiers in Microbiology}, doi = {10.3389/fmicb.2019.00735}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-201453}, pages = {735}, year = {2019}, abstract = {In fungi, green light is absorbed by rhodopsins, opsin proteins carrying a retinal molecule as chromophore. The basidiomycete Ustilago maydis, a fungal pathogen that infects corn plants, encodes three putative photoactive opsins, called ops1 (UMAG_02629), ops2 (UMAG_00371), and ops3 (UMAG_04125). UmOps1 and UmOps2 are expressed during the whole life cycle, in axenic cultures as well as in planta, whereas UmOps3 was recently shown to be absent in axenic cultures but highly expressed during plant infection. Here we show that expression of UmOps1 and UmOps2 is induced by blue light under control of white collar 1 (Wco1). UmOps1 is mainly localized in the plasma membrane, both when expressed in HEK cells and U. maydis sporidia. In contrast, UmOps2 was mostly found intracellularly in the membranes of vacuoles. Patch-clamp studies demonstrated that both rhodopsins are green light-driven outward rectifying proton pumps. UmOps1 revealed an extraordinary pH dependency with increased activity in more acidic environment. Also, UmOps1 showed a pronounced, concentration-dependent enhancement of pump current caused by weak organic acids (WOAs), especially by acetic acid and indole-3-acetic acid (IAA). In contrast, UmOps2 showed the typical behavior of light-driven, outwardly directed proton pumps, whereas UmOps3 did not exhibit any electrogenity. With this work, insights were gained into the localization and molecular function of two U. maydis rhodopsins, paving the way for further studies on the biological role of these rhodopsins in the life cycle of U. maydis.}, language = {en} } @article{SchartlKneitzVolkoffetal.2019, author = {Schartl, Manfred and Kneitz, Susanne and Volkoff, Helene and Adolfi, Mateus and Schmidt, Cornelia and Fischer, Petra and Minx, Patrick and Tomlinson, Chad and Meyer, Axel and Warren, Wesley C.}, title = {The piranha genome provides molecular insight associated to its unique feeding behavior}, series = {Genome Biology and Evolution}, volume = {11}, journal = {Genome Biology and Evolution}, number = {8}, doi = {10.1093/gbe/evz139}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-202218}, pages = {2099-2106}, year = {2019}, abstract = {The piranha enjoys notoriety due to its infamous predatory behavior but much is still not understood about its evolutionary origins and the underlying molecular mechanisms for its unusual feeding biology. We sequenced and assembled the red-bellied piranha (Pygocentrus nattereri) genome to aid future phenotypic and genetic investigations. The assembled draft genome is similar to other related fishes in repeat composition and gene count. Our evaluation of genes under positive selection suggests candidates for adaptations of piranhas' feeding behavior in neural functions, behavior, and regulation of energy metabolism. In the fasted brain, we find genes differentially expressed that are involved in lipid metabolism and appetite regulation as well as genes that may control the aggression/boldness behavior of hungry piranhas. Our first analysis of the piranha genome offers new insight and resources for the study of piranha biology and for feeding motivation and starvation in other organisms.}, language = {en} } @article{PoppRamirezZavalaSchwanfelderetal.2019, author = {Popp, Christina and Ram{\´i}rez-Zavala, Bernardo and Schwanfelder, Sonja and Kr{\"u}ger, Ines and Morschh{\"a}user, Joachim}, title = {Evolution of fluconazole-resistant Candida albicans strains by drug-induced mating competence and parasexual recombination}, series = {mBio}, volume = {10}, journal = {mBio}, number = {1}, doi = {10.1128/mBio.02740-18}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-200901}, pages = {e02740-18}, year = {2019}, abstract = {The clonal population structure of Candida albicans suggests that (para)sexual recombination does not play an important role in the lifestyle of this opportunistic fungal pathogen, an assumption that is strengthened by the fact that most C. albicans strains are heterozygous at the mating type locus (MTL) and therefore mating-incompetent. On the other hand, mating might occur within clonal populations and allow the combination of advantageous traits that were acquired by individual cells to adapt to adverse conditions. We have investigated if parasexual recombination may be involved in the evolution of highly drug-resistant strains exhibiting multiple resistance mechanisms against fluconazole, an antifungal drug that is commonly used to treat infections by C. albicans. Growth of strains that were heterozygous for MTL and different fluconazole resistance mutations in the presence of the drug resulted in the emergence of derivatives that had become homozygous for the mutated allele and the mating type locus and exhibited increased drug resistance. When MTLa/a and MTLα/α cells of these strains were mixed in all possible combinations, we could isolate mating products containing the genetic material from both parents. The initial mating products did not exhibit higher drug resistance than their parental strains, but further propagation under selective pressure resulted in the loss of the wild-type alleles and increased fluconazole resistance. Therefore, fluconazole treatment not only selects for resistance mutations but also promotes genomic alterations that confer mating competence, which allows cells in an originally clonal population to exchange individually acquired resistance mechanisms and generate highly drug-resistant progeny.}, language = {en} } @article{SchmidtHaywardCoelhoetal.2019, author = {Schmidt, Thomas S. B. and Hayward, Matthew R. and Coelho, Luiis P. and Li, Simone S. and Costea, Paul I. and Voigt, Anita Y. and Wirbel, Jakob and Maistrenko, Oleksandr M. and Alves, Renato J. C. and Bergsten, Emma and de Beaufort, Carine and Sobhani, Iradj and Heintz-Buschart, Anna and Sunagawa, Shinichi and Zeller, Georg and Wilmes, Paul and Bork, Peer}, title = {Extensive transmission of microbes along the gastrointestinal tract}, series = {eLife}, volume = {8}, journal = {eLife}, doi = {10.7554/eLife.42693}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-228954}, pages = {e42693, 1-18}, year = {2019}, abstract = {The gastrointestinal tract is abundantly colonized by microbes, yet the translocation of oral species to the intestine is considered a rare aberrant event, and a hallmark of disease. By studying salivary and fecal microbial strain populations of 310 species in 470 individuals from five countries, we found that transmission to, and subsequent colonization of, the large intestine by oral microbes is common and extensive among healthy individuals. We found evidence for a vast majority of oral species to be transferable, with increased levels of transmission in colorectal cancer and rheumatoid arthritis patients and, more generally, for species described as opportunistic pathogens. This establishes the oral cavity as an endogenous reservoir for gut microbial strains, and oral-fecal transmission as an important process that shapes the gastrointestinal microbiome in health and disease.}, subject = {Barrier}, language = {en} } @article{SrivastavaBencurovaGuptaetal.2019, author = {Srivastava, Mugdha and Bencurova, Elena and Gupta, Shishir K. and Weiss, Esther and L{\"o}ffler, J{\"u}rgen and Dandekar, Thomas}, title = {Aspergillus fumigatus challenged by human dendritic cells: metabolic and regulatory pathway responses testify a tight battle}, series = {Frontiers in Cellular and Infection Microbiology}, volume = {9}, journal = {Frontiers in Cellular and Infection Microbiology}, doi = {10.3389/fcimb.2019.00168}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-201368}, pages = {168}, year = {2019}, abstract = {Dendritic cells (DCs) are antigen presenting cells which serve as a passage between the innate and the acquired immunity. Aspergillosis is a major lethal condition in immunocompromised patients caused by the adaptable saprophytic fungus Aspergillus fumigatus. The healthy human immune system is capable to ward off A. fumigatus infections however immune-deficient patients are highly vulnerable to invasive aspergillosis. A. fumigatus can persist during infection due to its ability to survive the immune response of human DCs. Therefore, the study of the metabolism specific to the context of infection may allow us to gain insight into the adaptation strategies of both the pathogen and the immune cells. We established a metabolic model of A. fumigatus central metabolism during infection of DCs and calculated the metabolic pathway (elementary modes; EMs). Transcriptome data were used to identify pathways activated when A. fumigatus is challenged with DCs. In particular, amino acid metabolic pathways, alternative carbon metabolic pathways and stress regulating enzymes were found to be active. Metabolic flux modeling identified further active enzymes such as alcohol dehydrogenase, inositol oxygenase and GTP cyclohydrolase participating in different stress responses in A. fumigatus. These were further validated by qRT-PCR from RNA extracted under these different conditions. For DCs, we outlined the activation of metabolic pathways in response to the confrontation with A. fumigatus. We found the fatty acid metabolism plays a crucial role, along with other metabolic changes. The gene expression data and their analysis illuminate additional regulatory pathways activated in the DCs apart from interleukin regulation. In particular, Toll-like receptor signaling, NOD-like receptor signaling and RIG-I-like receptor signaling were active pathways. Moreover, we identified subnetworks and several novel key regulators such as UBC, EGFR, and CUL3 of DCs to be activated in response to A. fumigatus. In conclusion, we analyze the metabolic and regulatory responses of A. fumigatus and DCs when confronted with each other.}, language = {en} } @article{HeibyGoretzkiJohnsonetal.2019, author = {Heiby, Julia C. and Goretzki, Benedikt and Johnson, Christopher M. and Hellmich, Ute A. and Neuweiler, Hannes}, title = {Methionine in a protein hydrophobic core drives tight interactions required for assembly of spider silk}, series = {Nature Communications}, volume = {10}, journal = {Nature Communications}, doi = {10.1038/s41467-019-12365-5}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-202539}, pages = {4378}, year = {2019}, abstract = {Web spiders connect silk proteins, so-called spidroins, into fibers of extraordinary toughness. The spidroin N-terminal domain (NTD) plays a pivotal role in this process: it polymerizes spidroins through a complex mechanism of dimerization. Here we analyze sequences of spidroin NTDs and find an unusually high content of the amino acid methionine. We simultaneously mutate all methionines present in the hydrophobic core of a spidroin NTD from a nursery web spider's dragline silk to leucine. The mutated NTD is strongly stabilized and folds at the theoretical speed limit. The structure of the mutant is preserved, yet its ability to dimerize is substantially impaired. We find that side chains of core methionines serve to mobilize the fold, which can thereby access various conformations and adapt the association interface for tight binding. Methionine in a hydrophobic core equips a protein with the capacity to dynamically change shape and thus to optimize its function.}, language = {en} } @article{KehrbergerHolzschuh2019, author = {Kehrberger, Sandra and Holzschuh, Andrea}, title = {How does timing of flowering affect competition for pollinators, flower visitation and seed set in an early spring grassland plant?}, series = {Scientific Reports}, volume = {9}, journal = {Scientific Reports}, doi = {10.1038/s41598-019-51916-0}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-202549}, pages = {15593}, year = {2019}, abstract = {Knowledge on how the timing of flowering is related to plant fitness and species interactions is crucial to understand consequences of phenological shifts as they occur under climate change. Early flowering plants may face advantages of low competition for pollinators and disadvantages of low pollinator abundances and unfavourable weather conditions. However, it is unknown how this trade-off changes over the season and how the timing affects reproductive success. On eight grasslands we recorded intra-seasonal changes in pollinators, co-flowering plants, weather conditions, flower visitation rates, floral longevity and seed set of Pulsatilla vulgaris. Although bee abundances and the number of pollinator-suitable hours were low at the beginning of the season, early flowers of P. vulgaris received higher flower visitation rates and estimated total number of bee visits than later flowers, which was positively related to seed set. Flower visitation rates decreased over time and with increasing number of co-flowering plants, which competed with P. vulgaris for pollinators. Low interspecific competition for pollinators seems to be a major driver for early flowering dates. Thus, non-synchronous temporal shifts of co-flowering plants as they may occur under climate warming can be expected to strongly affect plant-pollinator interactions and the fitness of the involved plants.}, language = {en} } @article{HornMitesserHovestadtetal.2019, author = {Horn, Melanie and Mitesser, Oliver and Hovestadt, Thomas and Yoshii, Taishi and Rieger, Dirk and Helfrich-F{\"o}rster, Charlotte}, title = {The circadian clock improves fitness in the fruit fly, Drosophila melanogaster}, series = {Frontiers in Physiology}, volume = {10}, journal = {Frontiers in Physiology}, number = {1374}, issn = {1664-042X}, doi = {10.3389/fphys.2019.01374}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-195738}, year = {2019}, abstract = {It is assumed that a properly timed circadian clock enhances fitness, but only few studies have truly demonstrated this in animals. We raised each of the three classical Drosophila period mutants for >50 generations in the laboratory in competition with wildtype flies. The populations were either kept under a conventional 24-h day or under cycles that matched the mutant's natural cycle, i.e., a 19-h day in the case of pers mutants and a 29-h day for perl mutants. The arrhythmic per0 mutants were grown together with wildtype flies under constant light that renders wildtype flies similar arrhythmic as the mutants. In addition, the mutants had to compete with wildtype flies for two summers in two consecutive years under outdoor conditions. We found that wildtype flies quickly outcompeted the mutant flies under the 24-h laboratory day and under outdoor conditions, but perl mutants persisted and even outnumbered the wildtype flies under the 29-h day in the laboratory. In contrast, pers and per0 mutants did not win against wildtype flies under the 19-h day and constant light, respectively. Our results demonstrate that wildtype flies have a clear fitness advantage in terms of fertility and offspring survival over the period mutants and - as revealed for perl mutants - this advantage appears maximal when the endogenous period resonates with the period of the environment. However, the experiments indicate that perl and pers persist at low frequencies in the population even under the 24-h day. This may be a consequence of a certain mating preference of wildtype and heterozygous females for mutant males and time differences in activity patterns between wildtype and mutants.}, language = {en} } @article{DuquePoelmanSteffanDewenter2019, author = {Duque, Laura and Poelman, Erik H. and Steffan-Dewenter, Ingolf}, title = {Plant-mediated effects of ozone on herbivores depend on exposure duration and temperature}, series = {Scientific Reports}, volume = {9}, journal = {Scientific Reports}, doi = {10.1038/s41598-019-56234-z}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-202805}, pages = {19891}, year = {2019}, abstract = {Abiotic stress by elevated tropospheric ozone and temperature can alter plants' metabolism, growth, and nutritional value and modify the life cycle of their herbivores. We investigated how the duration of exposure of Sinapis arvensis plants to high ozone and temperature levels affect the life cycle of the large cabbage white, Pieris brassicae. Plants were exposed to ozone-clean (control) or ozone-enriched conditions (120 ppb) for either 1 or 5 days and were afterwards kept in a greenhouse with variable temperature conditions. When given the choice, P. brassicae butterflies laid 49\% fewer eggs on ozone-exposed than on control plants when the exposure lasted for 5 days, but showed no preference when exposure lasted for 1 day. The caterpillars took longer to hatch on ozone-exposed plants and at lower ambient temperatures. The ozone treatment had a positive effect on the survival of the eggs. Ozone decreased the growth of caterpillars reared at higher temperatures on plants exposed for 5 days, but not on plants exposed for 1 day. Overall, longer exposure of the plants to ozone and higher temperatures affected the life cycle of the herbivore more strongly. With global warming, the indirect impacts of ozone on herbivores are likely to become more common.}, language = {en} } @article{WestermannVenturiniSellinetal.2019, author = {Westermann, Alexander J. and Venturini, Elisa and Sellin, Mikael E. and F{\"o}rstner, Konrad U. and Hardt, Wolf-Dietrich and Vogel, J{\"o}rg}, title = {The major RNA-binding protein ProQ impacts virulence gene expression in Salmonella enterica serovar Typhimurium}, series = {mBio}, volume = {10}, journal = {mBio}, number = {1}, doi = {10.1128/mBio.02504-18}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-177722}, pages = {e02504-18}, year = {2019}, abstract = {FinO domain proteins such as ProQ of the model pathogen Salmonella enterica have emerged as a new class of major RNA-binding proteins in bacteria. ProQ has been shown to target hundreds of transcripts, including mRNAs from many virulence regions, but its role, if any, in bacterial pathogenesis has not been studied. Here, using a Dual RNA-seq approach to profile ProQ-dependent gene expression changes as Salmonella infects human cells, we reveal dysregulation of bacterial motility, chemotaxis, and virulence genes which is accompanied by altered MAPK (mitogen-activated protein kinase) signaling in the host. Comparison with the other major RNA chaperone in Salmonella, Hfq, reinforces the notion that these two global RNA-binding proteins work in parallel to ensure full virulence. Of newly discovered infection-associated ProQ-bound small noncoding RNAs (sRNAs), we show that the 3′UTR-derived sRNA STnc540 is capable of repressing an infection-induced magnesium transporter mRNA in a ProQ-dependent manner. Together, this comprehensive study uncovers the relevance of ProQ for Salmonella pathogenesis and highlights the importance of RNA-binding proteins in regulating bacterial virulence programs. IMPORTANCE The protein ProQ has recently been discovered as the centerpiece of a previously overlooked "third domain" of small RNA-mediated control of gene expression in bacteria. As in vitro work continues to reveal molecular mechanisms, it is also important to understand how ProQ affects the life cycle of bacterial pathogens as these pathogens infect eukaryotic cells. Here, we have determined how ProQ shapes Salmonella virulence and how the activities of this RNA-binding protein compare with those of Hfq, another central protein in RNA-based gene regulation in this and other bacteria. To this end, we apply global transcriptomics of pathogen and host cells during infection. In doing so, we reveal ProQ-dependent transcript changes in key virulence and host immune pathways. Moreover, we differentiate the roles of ProQ from those of Hfq during infection, for both coding and noncoding transcripts, and provide an important resource for those interested in ProQ-dependent small RNAs in enteric bacteria.}, language = {en} } @article{JaślanDreyerLuetal.2019, author = {Jaślan, Dawid and Dreyer, Ingo and Lu, Jinping and O'Malley, Ronan and Dindas, Julian and Marten, Irene and Hedrich, Rainer}, title = {Voltage-dependent gating of SV channel TPC1 confers vacuole excitability}, series = {Nature Communications}, volume = {10}, journal = {Nature Communications}, doi = {10.1038/s41467-019-10599-x}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-202029}, pages = {2659}, year = {2019}, abstract = {In contrast to the plasma membrane, the vacuole membrane has not yet been associated with electrical excitation of plants. Here, we show that mesophyll vacuoles from Arabidopsis sense and control the membrane potential essentially via the K\(^+\)-permeable TPC1 and TPK channels. Electrical stimuli elicit transient depolarization of the vacuole membrane that can last for seconds. Electrical excitability is suppressed by increased vacuolar Ca\(^{2+}\) levels. In comparison to wild type, vacuoles from the fou2 mutant, harboring TPC1 channels insensitive to luminal Ca\(^{2+}\), can be excited fully by even weak electrical stimuli. The TPC1-loss-of-function mutant tpc1-2 does not respond to electrical stimulation at all, and the loss of TPK1/TPK3-mediated K\(^{+}\) transport affects the duration of TPC1-dependent membrane depolarization. In combination with mathematical modeling, these results show that the vacuolar K\(^+\)-conducting TPC1 and TPK1/TPK3 channels act in concert to provide for Ca\(^{2+}\)- and voltage-induced electrical excitability to the central organelle of plant cells.}, language = {en} } @article{BoetzlKonleKrauss2019, author = {Boetzl, Fabian A. and Konle, Antonia and Krauss, Jochen}, title = {Aphid cards - useful model for assessing predation rates or bias prone nonsense?}, series = {Journal of Applied Entomology}, volume = {144}, journal = {Journal of Applied Entomology}, number = {1-2}, doi = {10.1111/jen.12692}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-204798}, pages = {74-80}, year = {2019}, abstract = {Predation on pest organisms is an essential ecosystem function supporting yields in modern agriculture. However, assessing predation rates is intricate, and they can rarely be linked directly to predator densities or functions. We tested whether sentinel prey aphid cards are useful tools to assess predation rates in the field. Therefore, we looked at aphid cards of different sizes on the ground level as well as within the vegetation. Additionally, by trapping ground-dwelling predators, we examined whether obtained predation rates could be linked to predator densities and traits. Predation rates recorded with aphid cards were independent of aphid card size. However, predation rates on the ground level were three times higher than within the vegetation. We found both predatory carabid activity densities as well as community weighted mean body size to be good predictors for predation rates. Predation rates obtained from aphid cards are stable over card type and related to predator assemblages. Aphid cards, therefore, are a useful, efficient method for rapidly assessing the ecosystem function predation. Their use might especially be recommended for assessments on the ground level and when time and resource limitations rule out more elaborate sentinel prey methods using exclosures with living prey animals.}, language = {en} } @article{SauerJuranekMarksetal.2019, author = {Sauer, Markus and Juranek, Stefan A. and Marks, James and De Magis, Alessio and Kazemier, Hinke G and Hilbig, Daniel and Benhalevy, Daniel and Wang, Xiantao and Hafner, Markus and Paeschke, Katrin}, title = {DHX36 prevents the accumulation of translationally inactive mRNAs with G4-structures in untranslated regions}, series = {Nature Communications}, volume = {10}, journal = {Nature Communications}, number = {2421}, doi = {10.1038/s41467-019-10432-5}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-227486}, pages = {1-15}, year = {2019}, abstract = {Translation efficiency can be affected by mRNA stability and secondary structures, including G-quadruplex structures (G4s). The highly conserved DEAH-box helicase DHX36/RHAU resolves G4s on DNA and RNA in vitro, however a systems-wide analysis of DHX36 targets and function is lacking. We map globally DHX36 binding to RNA in human cell lines and find it preferentially interacting with G-rich and G4-forming sequences on more than 4500 mRNAs. While DHX36 knockout (KO) results in a significant increase in target mRNA abundance, ribosome occupancy and protein output from these targets decrease, suggesting that they were rendered translationally incompetent. Considering that DHX36 targets, harboring G4s, preferentially localize in stress granules, and that DHX36 KO results in increased SG formation and protein kinase R (PKR/EIF2AK2) phosphorylation, we speculate that DHX36 is involved in resolution of rG4 induced cellular stress.}, language = {en} } @phdthesis{SoaresMachado2019, author = {Soares Machado, J{\´e}ssica}, title = {Dosimetry-based Assessment of Radiation-associated Cancer risk for \(^9\)\(^9\)\(^m\)Tc-MAG3 Scans in Infants and Optimization of Administered Activities for \(^6\)\(^8\)Ga-labelled Peptides in Children and Adolescents}, doi = {10.25972/OPUS-19264}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-192640}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {In 2006, 0.18 Mio pediatric nuclear medicine diagnostic exams were performed worldwide. However, for most of the radiopharmaceuticals used data on biokinetics and, as a consequence on dosimetry, are missing or have not been made publicly available. Therefore, most of the dosimetry assessments presented today for diagnostic agents in children and adolescents rely on the biokinetics data of adults. Even for one of the most common nuclear medicine exams for this patient group, renal scintigraphy with 99mTc-MAG3 for assessing renal function measured data on biokinetics is available only from a study performed on four children of different ages. In particular, renal scans are among the most frequent exams performed on infants and toddlers. Due to the young age, this patient group can be classified as a risk group with a higher probability of developing stochastic radiation effects compared to adults. As there are only limited data on biokinetics and dosimetry in this patient group, the aim of this study is to reassess the dosimetry and the associated radiation risk for a larger number of infants undergoing 99mTc-MAG3 renal scans based on a retrospective analysis of existing patient data. Data were collected retrospectively from 34 patients younger than 20 months with normal (20 patients) and abnormal renal function (14 patients) undergoing 99mTc-MAG3 scans. The patient-specific organ activity was estimated based on a retrospective calibration which was performed based on a set of two 3D-printed infant kidneys (newborns: 8.6 ml; 1-year-old: 23.4 ml) filled with known activities. Both phantoms were scanned at different positions along the anteroposterior axis inside a water phantom, providing depth- and size-dependent attenuation correction factors for planar imaging. Time-activity curves were determined by drawing kidney, bladder, and whole body regions-of-interest for each patient, and subsequently applying the calibration factor for conversion of counts to activity. Patient-specific time-integrated activity coefficients were obtained by integrating the organ-specific time-activity curves. Absorbed and effective dose coefficients for each patient were assessed with OLINDA/EXM for the provided newborn and 1-year-old phantom. Based on absorbed dose values, the radiation risk estimation was performed individually for each of the 34 patients with the National Cancer Institute's Radiation Risk Assessment Tool. The patients' organ-specific mean absorbed dose coefficients for the patients with normal renal function were 0.04±0.03 mGy/MBq for the kidneys and 0.27±0.24 mGy/MBq for the bladder. This resulted in a mean effective dose coefficient of 0.02±0.02 mSv/MBq. Based on the dosimetry results, the evaluation of the excess lifetime risk (ELR) for the development of radiation-induced cancer showed that the group of newborns has an ELR of 16.8 per 100,000 persons, which is higher in comparison with the 1-year-old group with an ELR of 14.7 per 100,000 persons. With regard to the 14 patients with abnormal renal function, the mean values for the organ absorbed dose coefficients for the patients were: 0.40±0.34 mGy/MBq for the kidneys and 0.46±0.37 mGy/MBq for the bladder. The corresponding effective dose coefficients (mSv/MBq) was: 0.05±0.02 mSv/MBq. The mean ELR (per 100,000 persons) for developing cancer from radiation exposure for patients with abnormal renal function was 29.2±18.7 per 100,000 persons. As a result, the radiation-associated stochastic risk increases with the organ doses, taking age- and gender-specific influences into account. Overall, the lifetime radiation risk associated with the 99mTc-MAG3 scans is very low in comparison to the general population risk for developing cancer. Furthermore, due to the increasing demand for PET-scans in children and adolescents with 68Ga-labelled peptides, in this work published data sets for those compounds were analyzed to derive recommendations for the administered activities in children and adolescents. The recommendation for the activities to be administered were based on the weight-independent effective dose model, proposed by the EANM Pediatric Dosage Card for application in pediatric nuclear medicine. The aim was to derive recommendations on administered activities for obtaining age-independent effective doses. Consequently, the corresponding weight-dependent effective dose coefficients were rescaled according to the formalism of the EANM dosage card, to determine the radiopharmaceutical class of 68Ga-labeled peptides ("multiples"), and to calculate the baseline activities based on the biokinetics of these compounds and an upper limit of the administered activity of 185 MBq for an adult. Analogous to 18F-fluoride, a minimum activity of 14 MBq is recommended. As a result, for those pediatric nuclear medicine applications involving 68Ga-labeled peptides, new values for the EANM dosage card were proposed and implemented based on the results derived in this work. Overall, despite the low additional radiation-related cancer risk, all efforts should be undertaken to optimize administered activities in children and adolescents for obtaining sufficient diagnostic information with minimal associated radiation risk.}, subject = {Biokinetics}, language = {en} } @phdthesis{Romanov2019, author = {Romanov, Natalie}, title = {Characterizing Variation of Protein Complexes and Functional Modules on a Temporal Scale and across Individuals}, doi = {10.25972/OPUS-16813}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-168139}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {A fundamental question in current biology concerns the translational mechanisms leading from genetic variability to phenotypes. Technologies have evolved to the extent that they can efficiently and economically determine an individual's genomic composition, while at the same time big data on clinical profiles and diagnostics have substantially accumulated. Genome-wide association studies linking genomic loci to certain traits, however, remain limited in their capacity to explain the cellular mechanisms that underlie the given association. For most associations, gene expression has been blamed; yet given that transcript and protein abundance oftentimes do not correlate, that finding does not necessarily decrypt the underlying mechanism. Thus, the integration of further information is crucial to establish a model that could prove more accurate in predicting genotypic effects on the human organism. In this work we describe the so-called proteotype as a feature of the cell that could provide a substantial link between genotype and phenotype. Rather than looking at the proteome as a set of independent molecules, we demonstrate a consistent modular architecture of the proteome that is driven by molecular cooperativity. Functional modules, especially protein complexes, can be further interrogated for differences between individuals and tackled as imprints of genetic and environmental variability. We also show that subtle stoichiometric changes of protein modules could have broader effects on the cellular system, such as the transport of specific molecular cargos. The presented work also delineates to what extent temporal events and processes influence the stoichiometry of protein complexes and functional modules. The re-wiring of the glycolytic pathway for example is illustrated as a potential cause for an increased Warburg effect during the ageing of the human bone marrow. On top of analyzing protein abundances we also interrogate proteome dynamics in terms of stability and solubility transitions during the short temporal progression of the cell cycle. One of our main observations in the thesis encompass the delineation of protein complexes into respective sub-complexes according to distinct stability patterns during the cell cycle. This has never been demonstrated before, and is functionally relevant for our understanding of the dis- and assembly of large protein modules. The insights presented in this work imply that the proteome is more than the sum of its parts, and primarily driven by variability in entire protein ensembles and their cooperative nature. Analyzing protein complexes and functional modules as molecular reflections of genetic and environmental variations could indeed prove to be a stepping stone in closing the gap between genotype and phenotype and customizing clinical treatments in the future.}, subject = {Proteotype}, language = {en} } @phdthesis{ALHijailan2019, author = {AL-Hijailan, Reem Saud}, title = {Establishment of endothelialized cardiac tissue using human induced pluripotent stem cells generated cardiomyocytes}, doi = {10.25972/OPUS-17397}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-173979}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Cardiovascular diseases are considered the leading cause of death worldwide according to the World Health Organization. Heart failure is the last stage of most of these diseases, where loss of myocardium leads to architectural and functional decline. The definitive treatment option for patients with CVDs is organ or tissue transplantation, which relies on donor availability. Therefore, generating an autologous bioengineered myocardium or heart could overcome this limitation. In addition, generating cardiac patches will provide ventricular wall support and enable reparative stem cells delivery to damaged areas. Although many hurdles still exist, a good number of researches have attempted to create an engineered cardiac tissue which can induce endogenous cardiac repair by replacing damaged myocardium. The present study provided cardiac patches in two models, one by a detergent coronary perfusion decellularization protocol that was optimized, and the other that resulted in a 3D cell-free extracellular matrix with intact architecture and preserved s-glycosaminoglycan and vasculature conduits. Perfusion with 1\% Sodium dodecyle sulfate (SDS) under constant pressure resulted in cell-free porcine scaffold within two and cell-free rat scaffold in 7 days, whereas scaffold perfused with 4\% sodium deoxycholate (SDO) was not able to remove cells completely. Re-reendothelialization of tissue vasculature was obtained by injecting human microvascular endothelial cell and human fibroblast in 2:1 ratio in a dynamic culture. One-week later, CD31 positive cells and endothelium markers were observed, indicating new blood lining. Moreover, functionality test of re-endothelialized tissue revealed improvement in clotting seen in decellularized tissues. When the tissue was ready to be repopulated, porcine induced pluripotent stem cells (PiPSc) were generated by transfected reprogramming of porcine skin fibroblast and then differentiated to cardiac cells following a robust protocol, for an autologous cardiac tissue model. However, due to the limitation in the PiPSc cell number, alternatively, human induced pluripotent stem cells generated cardiac cells were used. For reseeding a coculture of human iPSc generated cardiac cells, human mesenchymal stem cells and human fibroblast in 2:1:1 ratio respectively were used in a dynamic culture for 6-8 weeks. Contractions at different areas of the tissue were recorded at an average beating rate of 67 beats/min. In addition, positive cardiac markers (Troponin T), Fibroblast (vemintin), and mesenchymal stem cells (CD90) were detected. Not only that, but by week 3, MSC started differentiating to cardiac cells progressively until few CD90 positive cells were very few by week 6 with increasing troponin t positive cells in parallel. Electrophysiological and drug studies were difficult to obtain due to tissue thickness and limited assessment sources. However, the same construct was established using small intestine submucosa (SISer) scaffold, which recorded a spontaneous beating rate between 0.88 and 1.2 Hz, a conduction velocity of 23.9 ± 0.74 cm s-1, and a maximal contraction force of 0.453 ± 0.015 mN. Moreover, electrophysiological studies demonstrated a drug-dependent response on beating rate; a higher adrenalin frequency was revealed in comparison to the untreated tissue and isoproterenol administration, whereas a decrease in beating rate was observed with propranolol and untreated tissue. The present study demonstrated the establishment of vascularized cardiac tissue, which can be used for human clinical application.}, language = {en} } @phdthesis{Nelke2019, author = {Nelke, Lena}, title = {Establishment and optimization of 3-dimensional mamma carcinoma models for therapy simulation and drug testing}, doi = {10.25972/OPUS-17228}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-172280}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Breast cancer is the most common cancer among women worldwide and the second most common cause of cancer death in the developed countries. As the current state of the art in first-line drug screenings is highly ineffective, there is an urgent need for novel test systems that allow for reliable predictions of drug sensitivity. In this study, a tissue engineering approach was used to successfully establish and standardize a 3-dimensional (3D) mamma carcinoma test system that was optimized for the testing of anti-tumour therapies as well as for the investigation of tumour biological issues. This 3D test system is based on the decellularised scaffold of a porcine small intestinal segment and represents the three molecular subsets of oestrogen receptor-positive, HER2/Neu-overexpressing and triple negative breast cancer (TNBC). The characterization of the test system with respect to morphology as well as the expression of markers for epithelial-mesenchymal transition (EMT) and differentiation indicate that the 3D tumour models cultured under static and dynamic conditions reflect tumour relevant features and have a good correlation with in vivo tumour tissue from the corresponding xenograft models. In this respect, the dynamic culture in a flow bioreactor resulted in the generation of tumour models that exhibited best reflection of the morphology of the xenograft material. Furthermore, the proliferation indices of 3D models were significantly reduced compared to 2-dimensional (2D) cell culture and therefore better reflect the in vivo situation. As this more physiological proliferation index prevents an overestimation of the therapeutic effect of cytostatic compounds, this is a crucial advantage of the test system compared to 2D culture. Moreover, it could be shown that the 3D models can recapitulate different tumour stages with respect to tumour cell invasion. The scaffold SISmuc with the preserved basement membrane structure allowed the investigation of invasion over this barrier which tumour cells of epithelial origin have to cross in in vivo conditions during the process of metastasis formation. Additionally, the data obtained from ultrastructural analysis and in situ zymography indicate that the invasion observed is connected to a tumour cell-associated change in the basement membrane in which matrix metalloproteinases (MMPs) are also involved. This features of the model in combination with the mentioned methods of analysis could be used in the future to mechanistically investigate invasive processes and to test anti-metastatic therapy strategies. The validation of the 3D models as a test system with respect to the predictability of therapeutic effects was achieved by the clinically relevant targeted therapy with the monoclonal antibody trastuzumab which induces therapeutic response only in patients with HER2/Neu-overexpressing mamma carcinomas due to its specificity for HER2. While neither in 2D nor in 3D models of all molecular subsets a clear reduction of cell viability or an increase in apoptosis could be observed, a distinct increase in antibody-dependent cell-mediated cytotoxicity (ADCC) was detected only in the HER2/NEU-overexpressing 3D model with the help of an ADCC reporter gene assay that had been adapted for the application in the 3D model in the here presented work. This correlates with the clinical observations and underlines the relevance of ADCC as a mechanism of action (MOA) of trastuzumab. In order to measure the effects of ADCC on the tumour cells in a direct way without the indirect measurement via a reporter gene, the introduction of an immunological component into the models was required. This was achieved by the integration of peripheral blood mononuclear cells (PBMCs), thereby allowing the measurement of the induction of tumour cell apoptosis in the HER2/Neu-overexpressing model. Hence, in this study an immunocompetent model could be established that holds the potential for further testing of therapies from the emergent field of cancer immunotherapies. Subsequently, the established test system was used for the investigation of scientific issues from different areas of application. By the comparison of the sensitivity of the 2D and 3D model of TNBC towards the water-insoluble compound curcumin that was applied in a novel nanoformulation or in a DMSO-based formulation, the 3D test system was successfully applied for the evaluation of an innovative formulation strategy for poorly soluble drugs in order to achieve cancer therapy-relevant concentrations. Moreover, due to the lack of targeted therapies for TNBC, the TNBC model was applied for testing novel treatment strategies. On the one hand, therapy with the WEE1 kinase inhibitor MK 1775 was evaluated as a single agent as well as in combination with the chemotherapeutic agent doxorubicin. This therapy approach did not reveal any distinct benefits in the 3D test system in contrast to testing in 2D culture. On the other hand, a novel therapy approach from the field of cellular immunotherapies was successfully applied in the TNBC 3D model. The treatment with T cells that express a chimeric antigen receptor (CAR) against ROR1 revealed in the static as well as in the dynamic model a migration of T cells into the tumour tissue, an enhanced proliferation of T cells as well as an efficient lysis of the tumour cells via apoptosis and therefore a specific anti-cancer effect of CAR-transduced T cells compared to control T cells. These results illustrate that the therapeutic application of CAR T cells is a promising strategy for the treatment of solid tumours like TNBC and that the here presented 3D models are suitable for the evaluation and optimization of cellular immunotherapies. In the last part of this work, the 3D models were expanded by components of the tumour stroma for future applications. By coculture with fibroblasts, the natural structures of the intestinal scaffold comprising crypts and villi were remodelled and the tumour cells formed tumour-like structures together with the fibroblasts. This tissue model displayed a strong correlation with xenograft models with respect to morphology, marker expression as well as the activation of dermal fibroblasts towards a cancer-associated fibroblast (CAF) phenotype. For the integration of adipocytes which are an essential component of the breast stroma, a coculture with human adipose-derived stromal/stem cells (hASCs) which could be successfully differentiated along the adipose lineage in 3D static as well as dynamic models was established. These models are suitable especially for the mechanistic analysis of the reciprocal interaction between tumour cells and adipocytes due to the complex differentiation process. Taken together, in this study a human 3D mamma carcinoma test system for application in the preclinical development and testing of anti-tumour therapies as well as in basic research in the field of tumour biology was successfully established. With the help of this modular test system, relevant data can be obtained concerning the efficacy of therapies in tumours of different molecular subsets and different tumour stages as well as for the optimization of novel therapy strategies like immunotherapies. In the future this can contribute to improve the preclinical screening and thereby to reduce the high attrition rates in pharmaceutical industry as well as the amount of animal experiments.}, subject = {Brustkrebs}, language = {en} } @article{GrebinykPrylutskaGrebinyketal.2019, author = {Grebinyk, Anna and Prylutska, Svitlana and Grebinyk, Sergii and Prylutskyy, Yuriy and Ritter, Uwe and Matyshevska, Olga and Dandekar, Thomas and Frohme, Marcus}, title = {Complexation with C\(_{60}\) fullerene increases doxorubicin efficiency against leukemic cells in vitro}, series = {Nanoscale Research Letters}, volume = {14}, journal = {Nanoscale Research Letters}, number = {61}, doi = {10.1186/s11671-019-2894-1}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-228257}, year = {2019}, abstract = {Conventional anticancer chemotherapy is limited because of severe side effects as well as a quickly evolving multidrug resistance of the tumor cells. To address this problem, we have explored a C\(_{60}\) fullerene-based nanosized system as a carrier for anticancer drugs for an optimized drug delivery to leukemic cells.Here, we studied the physicochemical properties and anticancer activity of C\(_{60}\) fullerene noncovalent complexes with the commonly used anticancer drug doxorubicin. C\(_{60}\)-Doxorubicin complexes in a ratio 1:1 and 2:1 were characterized with UV/Vis spectrometry, dynamic light scattering, and high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The obtained analytical data indicated that the 140-nm complexes were stable and could be used for biological applications. In leukemic cell lines (CCRF-CEM, Jurkat, THP1 and Molt-16), the nanocomplexes revealed 3.5 higher cytotoxic potential in comparison with the free drug in a range of nanomolar concentrations. Also, the intracellular drug's level evidenced C\(_{60}\) fullerene considerable nanocarrier function.The results of this study indicated that C\(_{60}\) fullerene-based delivery nanocomplexes had a potential value for optimization of doxorubicin efficiency against leukemic cells.}, language = {en} } @article{GrubisicHaimBhusaletal.2019, author = {Grubisic, Maja and Haim, Abraham and Bhusal, Pramod and Dominoni, Davide M. and Gabriel, Katharina M. A. and Jechow, Andreas and Kupprat, Franziska and Lerner, Amit and Marchant, Paul and Riley, William and Stebelova, Katarina and van Grunsven, Roy H. A. and Zeman, Michal and Zubidat, Abed E. and H{\"o}lker, Franz}, title = {Light Pollution, Circadian Photoreception, and Melatonin in Vertebrates}, series = {Sustainability}, volume = {11}, journal = {Sustainability}, number = {22}, issn = {2071-1050}, doi = {10.3390/su11226400}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-193095}, year = {2019}, abstract = {Artificial light at night (ALAN) is increasing exponentially worldwide, accelerated by the transition to new efficient lighting technologies. However, ALAN and resulting light pollution can cause unintended physiological consequences. In vertebrates, production of melatonin—the "hormone of darkness" and a key player in circadian regulation—can be suppressed by ALAN. In this paper, we provide an overview of research on melatonin and ALAN in vertebrates. We discuss how ALAN disrupts natural photic environments, its effect on melatonin and circadian rhythms, and different photoreceptor systems across vertebrate taxa. We then present the results of a systematic review in which we identified studies on melatonin under typical light-polluted conditions in fishes, amphibians, reptiles, birds, and mammals, including humans. Melatonin is suppressed by extremely low light intensities in many vertebrates, ranging from 0.01-0.03 lx for fishes and rodents to 6 lx for sensitive humans. Even lower, wavelength-dependent intensities are implied by some studies and require rigorous testing in ecological contexts. In many studies, melatonin suppression occurs at the minimum light levels tested, and, in better-studied groups, melatonin suppression is reported to occur at lower light levels. We identify major research gaps and conclude that, for most groups, crucial information is lacking. No studies were identified for amphibians and reptiles and long-term impacts of low-level ALAN exposure are unknown. Given the high sensitivity of vertebrate melatonin production to ALAN and the paucity of available information, it is crucial to research impacts of ALAN further in order to inform effective mitigation strategies for human health and the wellbeing and fitness of vertebrates in natural ecosystems.}, language = {en} } @article{HeydarianYangSchweinlinetal.2019, author = {Heydarian, Motaharehsadat and Yang, Tao and Schweinlin, Matthias and Steinke, Maria and Walles, Heike and Rudel, Thomas and Kozjak-Pavlovic, Vera}, title = {Biomimetic human tissue model for long-term study of Neisseria gonorrhoeae infection}, series = {Frontiers in Microbiology}, volume = {10}, journal = {Frontiers in Microbiology}, number = {1740}, issn = {1664-302X}, doi = {10.3389/fmicb.2019.01740}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-197912}, year = {2019}, abstract = {Gonorrhea is the second most common sexually transmitted infection in the world and is caused by Gram-negative diplococcus Neisseria gonorrhoeae. Since N. gonorrhoeae is a human-specific pathogen, animal infection models are only of limited use. Therefore, a suitable in vitro cell culture model for studying the complete infection including adhesion, transmigration and transport to deeper tissue layers is required. In the present study, we generated three independent 3D tissue models based on porcine small intestinal submucosa (SIS) scaffold by co-culturing human dermal fibroblasts with human colorectal carcinoma, endometrial epithelial, and male uroepithelial cells. Functional analyses such as transepithelial electrical resistance (TEER) and FITC-dextran assay indicated the high barrier integrity of the created monolayer. The histological, immunohistochemical, and ultra-structural analyses showed that the 3D SIS scaffold-based models closely mimic the main characteristics of the site of gonococcal infection in human host including the epithelial monolayer, the underlying connective tissue, mucus production, tight junction, and microvilli formation. We infected the established 3D tissue models with different N. gonorrhoeae strains and derivatives presenting various phenotypes regarding adhesion and invasion. The results indicated that the disruption of tight junctions and increase in interleukin production in response to the infection is strain and cell type-dependent. In addition, the models supported bacterial survival and proved to be better suitable for studying infection over the course of several days in comparison to commonly used Transwell® models. This was primarily due to increased resilience of the SIS scaffold models to infection in terms of changes in permeability, cell destruction and bacterial transmigration. In summary, the SIS scaffold-based 3D tissue models of human mucosal tissues represent promising tools for investigating N. gonorrhoeae infections under close-to-natural conditions.}, language = {en} } @article{GrebinykPrylutskaChepurnaetal.2019, author = {Grebinyk, Anna and Prylutska, Svitlana and Chepurna, Oksana and Grebinyk, Sergii and Prylutskyy, Yuriy and Ritter, Uwe and Ohulchanskyy, Tymish Y. and Matyshevska, Olga and Dandekar, Thomas and Frohme, Marcus}, title = {Synergy of chemo- and photodynamic therapies with C\(_{60}\) Fullerene-Doxorubicin nanocomplex}, series = {Nanomaterials}, volume = {9}, journal = {Nanomaterials}, number = {11}, issn = {2079-4991}, doi = {10.3390/nano9111540}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-193140}, year = {2019}, abstract = {A nanosized drug complex was explored to improve the efficiency of cancer chemotherapy, complementing it with nanodelivery and photodynamic therapy. For this, nanomolar amounts of a non-covalent nanocomplex of Doxorubicin (Dox) with carbon nanoparticle C\(_{60}\) fullerene (C\(_{60}\)) were applied in 1:1 and 2:1 molar ratio, exploiting C\(_{60}\) both as a drug-carrier and as a photosensitizer. The fluorescence microscopy analysis of human leukemic CCRF-CEM cells, in vitro cancer model, treated with nanocomplexes showed Dox's nuclear and C\(_{60}\)'s extranuclear localization. It gave an opportunity to realize a double hit strategy against cancer cells based on Dox's antiproliferative activity and C\(_{60}\)'s photoinduced pro-oxidant activity. When cells were treated with 2:1 C\(_{60}\)-Dox and irradiated at 405 nm the high cytotoxicity of photo-irradiated C\(_{60}\)-Dox enabled a nanomolar concentration of Dox and C\(_{60}\) to efficiently kill cancer cells in vitro. The high pro-oxidant and pro-apoptotic efficiency decreased IC\(_{50}\) 16, 9 and 7 × 10\(^3\)-fold, if compared with the action of Dox, non-irradiated nanocomplex, and C\(_{60}\)'s photodynamic effect, correspondingly. Hereafter, a strong synergy of therapy arising from the combination of C\(_{60}\)-mediated Dox delivery and C\(_{60}\) photoexcitation was revealed. Our data indicate that a combination of chemo- and photodynamic therapies with C\(_{60}\)-Dox nanoformulation provides a promising synergetic approach for cancer treatment.}, language = {en} } @article{VillalobosWieseImhoffetal.2019, author = {Villalobos, Alvaro S. and Wiese, Jutta and Imhoff, Johannes F. and Dorador, Cristina and Keller, Alexander and Hentschel, Ute}, title = {Systematic affiliation and genome analysis of Subtercola vilae DB165T with particular emphasis on cold adaptation of an isolate from a high-altitude cold volcano lake}, series = {Microorganisms}, volume = {7}, journal = {Microorganisms}, number = {4}, issn = {2076-2607}, doi = {10.3390/microorganisms7040107}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-197394}, year = {2019}, abstract = {Among the Microbacteriaceae the species of Subtercola and Agreia form closely associated clusters. Phylogenetic analysis demonstrated three major phylogenetic branches of these species. One of these branches contains the two psychrophilic species Subtercola frigoramans and Subtercola vilae, together with a larger number of isolates from various cold environments. Genomic evidence supports the separation of Agreia and Subtercola species. In order to gain insight into the ability of S. vilae to adapt to life in this extreme environment, we analyzed the genome with a particular focus on properties related to possible adaptation to a cold environment. General properties of the genome are presented, including carbon and energy metabolism, as well as secondary metabolite production. The repertoire of genes in the genome of S. vilae DB165\(^T\) linked to adaptations to the harsh conditions found in Llullaillaco Volcano Lake includes several mechanisms to transcribe proteins under low temperatures, such as a high number of tRNAs and cold shock proteins. In addition, S. vilae DB165\(^T\) is capable of producing a number of proteins to cope with oxidative stress, which is of particular relevance at low temperature environments, in which reactive oxygen species are more abundant. Most important, it obtains capacities to produce cryo-protectants, and to combat against ice crystal formation, it produces ice-binding proteins. Two new ice-binding proteins were identified which are unique to S. vilae DB165\(^T\). These results indicate that S. vilae has the capacity to employ different mechanisms to live under the extreme and cold conditions prevalent in Llullaillaco Volcano Lake.}, language = {en} } @article{LiuKinoshitaAdolfietal.2019, author = {Liu, Ruiqi and Kinoshita, Masato and Adolfi, Mateus C. and Schartl, Manfred}, title = {Analysis of the role of the Mc4r system in development, growth, and puberty of medaka}, series = {Frontiers in Endocrinology}, volume = {10}, journal = {Frontiers in Endocrinology}, doi = {10.3389/fendo.2019.00213}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-201472}, pages = {213}, year = {2019}, abstract = {In mammals the melanocortin 4 receptor (Mc4r) signaling system has been mainly associated with the regulation of appetite and energy homeostasis. In fish of the genus Xiphophorus (platyfish and swordtails) puberty onset is genetically determined by a single locus, which encodes the mc4r. Wild populations of Xiphophorus are polymorphic for early and late-maturing individuals. Copy number variation of different mc4r alleles is responsible for the difference in puberty onset. To answer whether this is a special adaptation of the Mc4r signaling system in the lineage of Xiphophorus or a more widely conserved mechanism in teleosts, we studied the role of Mc4r in reproductive biology of medaka (Oryzias latipes), a close relative to Xiphophorus and a well-established model to study gonadal development. To understand the potential role of Mc4r in medaka, we characterized the major features of the Mc4r signaling system (mc4r, mrap2, pomc, agrp1). In medaka, all these genes are expressed before hatching. In adults, they are mainly expressed in the brain. The transcript of the receptor accessory protein mrap2 co-localizes with mc4r in the hypothalamus in adult brains indicating a conserved function of modulating Mc4r signaling. Comparing growth and puberty between wild-type and mc4r knockout medaka revealed that absence of Mc4r does not change puberty timing but significantly delays hatching. Embryonic development of knockout animals is retarded compared to wild-types. In conclusion, the Mc4r system in medaka is involved in regulation of growth rather than puberty.}, language = {en} } @article{YangRajeeveRudeletal.2019, author = {Yang, Manli and Rajeeve, Karthika and Rudel, Thomas and Dandekar, Thomas}, title = {Comprehensive Flux Modeling of Chlamydia trachomatis Proteome and qRT-PCR Data Indicate Biphasic Metabolic Differences Between Elementary Bodies and Reticulate Bodies During Infection}, series = {Frontiers in Microbiology}, volume = {10}, journal = {Frontiers in Microbiology}, number = {2350}, issn = {1664-302X}, doi = {10.3389/fmicb.2019.02350}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-189434}, year = {2019}, abstract = {Metabolic adaptation to the host cell is important for obligate intracellular pathogens such as Chlamydia trachomatis (Ct). Here we infer the flux differences for Ct from proteome and qRT-PCR data by comprehensive pathway modeling. We compare the comparatively inert infectious elementary body (EB) and the active replicative reticulate body (RB) systematically using a genome-scale metabolic model with 321 metabolites and 277 reactions. This did yield 84 extreme pathways based on a published proteomics dataset at three different time points of infection. Validation of predictions was done by quantitative RT-PCR of enzyme mRNA expression at three time points. Ct's major active pathways are glycolysis, gluconeogenesis, glycerol-phospholipid (GPL) biosynthesis (support from host acetyl-CoA) and pentose phosphate pathway (PPP), while its incomplete TCA and fatty acid biosynthesis are less active. The modeled metabolic pathways are much more active in RB than in EB. Our in silico model suggests that EB and RB utilize folate to generate NAD(P)H using independent pathways. The only low metabolic flux inferred for EB involves mainly carbohydrate metabolism. RB utilizes energy -rich compounds to generate ATP in nucleic acid metabolism. Validation data for the modeling include proteomics experiments (model basis) as well as qRT-PCR confirmation of selected metabolic enzyme mRNA expression differences. The metabolic modeling is made fully available here. Its detailed insights and models on Ct metabolic adaptations during infection are a useful modeling basis for future studies.}, language = {en} } @article{HuppRosenkranzBonfigetal.2019, author = {Hupp, Sabrina and Rosenkranz, Maaria and Bonfig, Katharina and Pandey, Chandana and Roitsch, Thomas}, title = {Noninvasive Phenotyping of Plant-Pathogen Interaction: Consecutive In Situ Imaging of Fluorescing Pseudomonas syringae, Plant Phenolic Fluorescence, and Chlorophyll Fluorescence in Arabidopsis Leaves}, series = {Frontiers in Plant Science}, volume = {10}, journal = {Frontiers in Plant Science}, number = {1239}, issn = {1664-462X}, doi = {10.3389/fpls.2019.01239}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-189425}, year = {2019}, abstract = {Plant-pathogen interactions have been widely studied, but mostly from the site of the plant secondary defense. Less is known about the effects of pathogen infection on plant primary metabolism. The possibility to transform a fluorescing protein into prokaryotes is a promising phenotyping tool to follow a bacterial infection in plants in a noninvasive manner. In the present study, virulent and avirulent Pseudomonas syringae strains were transformed with green fluorescent protein (GFP) to follow the spread of bacteria in vivo by imaging Pulse-Amplitude-Modulation (PAM) fluorescence and conventional binocular microscopy. The combination of various wavelengths and filters allowed simultaneous detection of GFP-transformed bacteria, PAM chlorophyll fluorescence, and phenolic fluorescence from pathogen-infected plant leaves. The results show that fluorescence imaging allows spatiotemporal monitoring of pathogen spread as well as phenolic and chlorophyll fluorescence in situ, thus providing a novel means to study complex plant-pathogen interactions and relate the responses of primary and secondary metabolism to pathogen spread and multiplication. The study establishes a deeper understanding of imaging data and their implementation into disease screening.}, language = {en} } @article{DoerkPeterlongoMannermaaetal.2019, author = {D{\"o}rk, Thilo and Peterlongo, Peter and Mannermaa, Arto and Bolla, Manjeet K. and Wang, Qin and Dennis, Joe and Ahearn, Thomas and Andrulis, Irene L. and Anton-Culver, Hoda and Arndt, Volker and Aronson, Kristan J. and Augustinsson, Annelie and Beane Freeman, Laura E. and Beckmann, Matthias W. and Beeghly-Fadiel, Alicia and Behrens, Sabine and Bermisheva, Marina and Blomqvist, Carl and Bogdanova, Natalia V. and Bojesen, Stig E. and Brauch, Hiltrud and Brenner, Hermann and Burwinkel, Barbara and Canzian, Federico and Chan, Tsun L. and Chang-Claude, Jenny and Chanock, Stephen J. and Choi, Ji-Yeob and Christiansen, Hans and Clarke, Christine L. and Couch, Fergus J. and Czene, Kamila and Daly, Mary B. and dos-Santos-Silva, Isabel and Dwek, Miriam and Eccles, Diana M. and Ekici, Arif B. and Eriksson, Mikael and Evans, D. Gareth and Fasching, Peter A. and Figueroa, Jonine and Flyger, Henrik and Fritschi, Lin and Gabrielson, Marike and Gago-Dominguez, Manuela and Gao, Chi and Gapstur, Susan M. and Garc{\´i}a-Closas, Montserrat and Garc{\´i}a-S{\´a}enz, Jos{\´e} A. and Gaudet, Mia M. and Giles, Graham G. and Goldberg, Mark S. and Goldgar, David E. and Guen{\´e}l, Pascal and Haeberle, Lothar and Haimann, Christopher A. and H{\aa}kansson, Niclas and Hall, Per and Hamann, Ute and Hartman, Mikael and Hauke, Jan and Hein, Alexander and Hillemanns, Peter and Hogervorst, Frans B. L. and Hooning, Maartje J. and Hopper, John L. and Howell, Tony and Huo, Dezheng and Ito, Hidemi and Iwasaki, Motoki and Jakubowska, Anna and Janni, Wolfgang and John, Esther M. and Jung, Audrey and Kaaks, Rudolf and Kang, Daehee and Kapoor, Pooja Middha and Khusnutdinova, Elza and Kim, Sung-Won and Kitahara, Cari M. and Koutros, Stella and Kraft, Peter and Kristensen, Vessela N. and Kwong, Ava and Lambrechts, Diether and Le Marchand, Loic and Li, Jingmei and Lindstr{\"o}m, Sara and Linet, Martha and Lo, Wing-Yee and Long, Jirong and Lophatananon, Artitaya and Lubiński, Jan and Manoochehri, Mehdi and Manoukian, Siranoush and Margolin, Sara and Martinez, Elena and Matsuo, Keitaro and Mavroudis, Dimitris and Meindl, Alfons and Menon, Usha and Milne, Roger L. and Mohd Taib, Nur Aishah and Muir, Kenneth and Mulligan, Anna Marie and Neuhausen, Susan L. and Nevanlinna, Heli and Neven, Patrick and Newman, William G. and Offit, Kenneth and Olopade, Olufunmilayo I. and Olshan, Andrew F. and Olson, Janet E. and Olsson, H{\aa}kan and Park, Sue K. and Park-Simon, Tjoung-Won and Peto, Julian and Plaseska-Karanfilska, Dijana and Pohl-Rescigno, Esther and Presneau, Nadege and Rack, Brigitte and Radice, Paolo and Rashid, Muhammad U. and Rennert, Gad and Rennert, Hedy S. and Romero, Atocha and Ruebner, Matthias and Saloustros, Emmanouil and Schmidt, Marjanka K. and Schmutzler, Rita K. and Schneider, Michael O. and Schoemaker, Minouk J. and Scott, Christopher and Shen, Chen-Yang and Shu, Xiao-Ou and Simard, Jaques and Slager, Susan and Smichkoska, Snezhana and Southey, Melissa C. and Spinelli, John J. and Stone, Jennifer and Surowy, Harald and Swerdlow, Anthony J. and Tamimi, Rulla M. and Tapper, William J. and Teo, Soo H. and Terry, Mary Beth and Toland, Amanda E. and Tollenaar, Rob A. E. M. and Torres, Diana and Torres-Mej{\´i}a, Gabriela and Troester, Melissa A. and Truong, Th{\´e}r{\`e}se and Tsugane, Shoichiro and Untch, Michael and Vachon, Celine M. and van den Ouweland, Ans M. W. and van Veen, Elke M. and Vijai, Joseph and Wendt, Camilla and Wolk, Alicja and Yu, Jyh-Cherng and Zheng, Wei and Ziogas, Argyrios and Ziv, Elad and Dunnig, Alison and Pharaoh, Paul D. P. and Schindler, Detlev and Devilee, Peter and Easton, Douglas F.}, title = {Two truncating variants in FANCC and breast cancer risk}, series = {Scientific Reports}, volume = {9}, journal = {Scientific Reports}, organization = {ABCTB Investigators, NBCS Collaborators}, doi = {10.1038/s41598-019-48804-y}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-222838}, year = {2019}, abstract = {Fanconi anemia (FA) is a genetically heterogeneous disorder with 22 disease-causing genes reported to date. In some FA genes, monoallelic mutations have been found to be associated with breast cancer risk, while the risk associations of others remain unknown. The gene for FA type C, FANCC, has been proposed as a breast cancer susceptibility gene based on epidemiological and sequencing studies. We used the Oncoarray project to genotype two truncating FANCC variants (p.R185X and p.R548X) in 64,760 breast cancer cases and 49,793 controls of European descent. FANCC mutations were observed in 25 cases (14 with p.R185X, 11 with p.R548X) and 26 controls (18 with p.R185X, 8 with p.R548X). There was no evidence of an association with the risk of breast cancer, neither overall (odds ratio 0.77, 95\%CI 0.44-1.33, p = 0.4) nor by histology, hormone receptor status, age or family history. We conclude that the breast cancer risk association of these two FANCC variants, if any, is much smaller than for BRCA1, BRCA2 or PALB2 mutations. If this applies to all truncating variants in FANCC it would suggest there are differences between FA genes in their roles on breast cancer risk and demonstrates the merit of large consortia for clarifying risk associations of rare variants.}, language = {en} } @article{ElMeseryRosenthalRauertWunderlichetal.2019, author = {El-Mesery, Mohamed and Rosenthal, Tina and Rauert-Wunderlich, Hilka and Schreder, Martin and St{\"u}hmer, Thorsten and Leich, Ellen and Schlosser, Andreas and Ehrenschwender, Martin and Wajant, Harald and Siegmund, Daniela}, title = {The NEDD8-activating enzyme inhibitor MLN4924 sensitizes a TNFR1+ subgroup of multiple myeloma cells for TNF-induced cell death}, series = {Cell Death \& Disease}, volume = {10}, journal = {Cell Death \& Disease}, doi = {10.1038/s41419-019-1860-2}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-226666}, year = {2019}, abstract = {The NEDD8-activating enzyme (NAE) inhibitor MLN4924 inhibits cullin-RING ubiquitin ligase complexes including the SKP1-cullin-F-box E3 ligase βTrCP. MLN4924 therefore inhibits also the βTrCP-dependent activation of the classical and the alternative NFĸB pathway. In this work, we found that a subgroup of multiple myeloma cell lines (e.g., RPMI-8226, MM.1S, KMS-12BM) and about half of the primary myeloma samples tested are sensitized to TNF-induced cell death by MLN4924. This correlated with MLN4924-mediated inhibition of TNF-induced activation of the classical NFκB pathway and reduced the efficacy of TNF-induced TNFR1 signaling complex formation. Interestingly, binding studies revealed a straightforward correlation between cell surface TNFR1 expression in multiple myeloma cell lines and their sensitivity for MLN4924/TNF-induced cell death. The cell surface expression levels of TNFR1 in the investigated MM cell lines largely correlated with TNFR1 mRNA expression. This suggests that the variable levels of cell surface expression of TNFR1 in myeloma cell lines are decisive for TNF/MLN4924 sensitivity. Indeed, introduction of TNFR1 into TNFR1-negative TNF/MLN4924-resistant KMS-11BM cells, was sufficient to sensitize this cell line for TNF/MLN4924-induced cell death. Thus, MLN4924 might be especially effective in myeloma patients with TNFR1+ myeloma cells and a TNFhigh tumor microenvironment.}, language = {en} } @article{KrebsBehrmannKalogirouetal.2019, author = {Krebs, Markus and Behrmann, Christoph and Kalogirou, Charis and Sokolakis, Ioannis and Kneitz, Susanne and Kruithof-de Julio, Marianna and Zoni, Eugenio and Rech, Anne and Schilling, Bastian and K{\"u}bler, Hubert and Spahn, Martin and Kneitz, Burkhard}, title = {miR-221 Augments TRAIL-mediated apoptosis in prostate cancer cells by inducing endogenous TRAIL expression and targeting the functional repressors SOCS3 and PIK3R1}, series = {BioMed Research International}, volume = {2019}, journal = {BioMed Research International}, doi = {10.1155/2019/6392748}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-202480}, pages = {6392748}, year = {2019}, abstract = {miR-221 is regarded as an oncogene in many malignancies, and miR-221-mediated resistance towards TRAIL was one of the first oncogenic roles shown for this small noncoding RNA. In contrast, miR-221 is downregulated in prostate cancer (PCa), thereby implying a tumour suppressive function. By using proliferation and apoptosis assays, we show a novel feature of miR-221 in PCa cells: instead of inducing TRAIL resistance, miR-221 sensitized cells towards TRAIL-induced proliferation inhibition and apoptosis induction. Partially responsible for this effect was the interferon-mediated gene signature, which among other things contained an endogenous overexpression of the TRAIL encoding gene TNFSF10. This TRAIL-friendly environment was provoked by downregulation of the established miR-221 target gene SOCS3. Moreover, we introduced PIK3R1 as a target gene of miR-221 in PCa cells. Proliferation assays showed that siRNA-mediated downregulation of SOCS3 and PIK3R1 mimicked the effect of miR-221 on TRAIL sensitivity. Finally, Western blotting experiments confirmed lower amounts of phospho-Akt after siRNA-mediated downregulation of PIK3R1 in PC3 cells. Our results further support the tumour suppressing role of miR-221 in PCa, since it sensitises PCa cells towards TRAIL by regulating the expression of the oncogenes SOCS3 and PIK3R1. Given the TRAIL-inhibiting effect of miR-221 in various cancer entities, our results suggest that the influence of miR-221 on TRAIL-mediated apoptosis is highly context- and entity-dependent.}, language = {en} } @article{MolinasGonzalezCastroGonzalezMegiasetal.2019, author = {Molinas-Gonz{\´a}lez, Carlos R. and Castro, Jorge and Gonz{\´a}lez-Meg{\´i}as, Adela and Leverkus, Alexandro B.}, title = {Effects of post-fire deadwood management on soil macroarthropod communities}, series = {Forests}, volume = {10}, journal = {Forests}, number = {11}, issn = {1999-4907}, doi = {10.3390/f10111046}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-193079}, year = {2019}, abstract = {Dead wood comprises a vast amount of biological legacies that set the scene for ecological regeneration after wildfires, yet its removal is the most frequent management strategy worldwide. Soil-dwelling organisms are conspicuous, and they provide essential ecosystem functions, but their possible affection by different post-fire management strategies has so far been neglected. We analyzed the abundance, richness, and composition of belowground macroarthropod communities under two contrasting dead-wood management regimes after a large wildfire in the Sierra Nevada Natural and National Park (Southeast Spain). Two plots at different elevation were established, each containing three replicates of two experimental treatments: partial cut, where trees were cut and their branches lopped off and left over the ground, and salvage logging, where all the trees were cut, logs were piled, branches were mechanically masticated, and slash was spread on the ground. Ten years after the application of the treatments, soil cores were extracted from two types of microhabitat created by these treatments: bare-soil (in both treatments) and under-logs (in the partial cut treatment only). Soil macroarthropod assemblages were dominated by Hemiptera and Hymenoptera (mostly ants) and were more abundant and richer in the lowest plot. The differences between dead-wood treatments were most evident at the scale of management interventions: abundance and richness were lowest after salvage logging, even under similar microhabitats (bare-soil). However, there were no significant differences between microhabitat types on abundance and richness within the partial cut treatment. Higher abundance and richness in the partial cut treatment likely resulted from higher resource availability and higher plant diversity after natural regeneration. Our results suggest that belowground macroarthropod communities are sensitive to the manipulation of dead-wood legacies and that management through salvage logging could reduce soil macroarthropod recuperation compared to other treatments with less intense management even a decade after application.}, language = {en} } @article{LatifiValbuena2019, author = {Latifi, Hooman and Valbuena, Ruben}, title = {Current trends in forest ecological applications of three-dimensional remote sensing: Transition from experimental to operational solutions?}, series = {Forests}, volume = {10}, journal = {Forests}, number = {10}, issn = {1999-4907}, doi = {10.3390/f10100891}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-193282}, year = {2019}, abstract = {The alarming increase in the magnitude and spatiotemporal patterns of changes in composition, structure and function of forest ecosystems during recent years calls for enhanced cross-border mitigation and adaption measures, which strongly entail intensified research to understand the underlying processes in the ecosystems as well as their dynamics. Remote sensing data and methods are nowadays the main complementary sources of synoptic, up-to-date and objective information to support field observations in forest ecology. In particular, analysis of three-dimensional (3D) remote sensing data is regarded as an appropriate complement, since they are hypothesized to resemble the 3D character of most forest attributes. Following their use in various small-scale forest structural analyses over the past two decades, these sources of data are now on their way to be integrated in novel applications in fields like citizen science, environmental impact assessment, forest fire analysis, and biodiversity assessment in remote areas. These and a number of other novel applications provide valuable material for the Forests special issue "3D Remote Sensing Applications in Forest Ecology: Composition, Structure and Function", which shows the promising future of these technologies and improves our understanding of the potentials and challenges of 3D remote sensing in practical forest ecology worldwide.}, language = {en} } @phdthesis{Derakhshani2019, author = {Derakhshani, Shaghayegh}, title = {Measles virus infection enhances dendritic cell migration in a 3D environment}, doi = {10.25972/OPUS-18918}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-189182}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {The respiratory system is amongst the most important compartments in the human body. Due to its connection to the external environment, it is one of the most common portals of pathogen entry. Airborne pathogens like measles virus (MV) carried in liquid droplets exhaled from the infected individuals via a cough or sneeze enter the body from the upper respiratory tract and travel down to the lower respiratory tract and reach the alveoli. There, pathogens are captured by the resident dendritic cells (DCs) or macrophages and brought to the lymph node where immune responses or, as in case of MV, dissemination via the hematopoietic cell compartment are initiated. Basic mechanisms governing MV exit from the respiratory tract, especially virus transmission from infected immune cells to the epithelial cells have not been fully addressed before. Considering the importance of these factors in the viral spread, a complex close-to-in-vivo 3D human respiratory tract model was generated. This model was established using de-cellularized porcine intestine tissue as a biological scaffold and H358 cells as targets for infection. The scaffold was embedded with fibroblast cells, and later on, an endothelial cell layer seeded at the basolateral side. This provided an environment resembling the respiratory tract where MV infected DCs had to transmigrate through the collagen scaffold and transmit the virus to epithelial cells in a Nectin-4 dependent manner. For viral transmission, the access of infected DCs to the recipient epithelial cells is an essential prerequisite and therefore, this important factor which is reflected by cell migration was analyzed in this 3D system. The enhanced motility of specifically MV-infected DCs in the 3D models was observed, which occurred independently of factors released from the other cell types in the models. Enhanced motility of infected DCs in 3D collagen matrices suggested infection-induced cytoskeletal remodeling, as also verified by detection of cytoskeletal polarization, uropod formation. This enforced migration was sensitive to ROCK inhibition revealing that MV infection induces an amoeboid migration mode in DCs. In support of this, the formation of podosome structures and filopodia, as well as their activity, were reduced in infected DCs and retained in their uninfected siblings. Differential migration modes of uninfected and infected DCs did not cause differential maturation, which was found to be identical for both populations. As an underlying mechanism driving this enforced migration, the role of sphingosine kinase (SphK) and sphingosine-1-phosphate (S1P) was studied in MV-exposed cultures. It was shown in this thesis that MV-infection increased S1P production, and this was identified as a contributing factor as inhibition sphingosine kinase activity abolished enforced migration of MV-infected DCs. These findings revealed that MV infection induces a fast push-and-squeeze amoeboid mode of migration, which is supported by SphK/S1P axis. However, this push-and-squeeze amoeboid migration mode did not prevent the transendothelial migration of MV-infected DCs. Altogether, this 3D system has been proven to be a suitable model to study specific parameters of mechanisms involved in infections in an in vivo-like conditions.}, subject = {Dendritische Zelle}, language = {en} } @article{HovestadtThomasMitesseretal.2019, author = {Hovestadt, Thomas and Thomas, Jeremy A. and Mitesser, Oliver and Sch{\"o}nrogge, Karsten}, title = {Multiple host use and the dynamics of host-switching in host-parasite systems}, series = {Insect Conservation and Diversity}, volume = {12}, journal = {Insect Conservation and Diversity}, number = {6}, doi = {10.1111/icad.12374}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-204747}, pages = {511-522}, year = {2019}, abstract = {The link between multi-host use and host switching in host-parasite interactions is a continuing area of debate. Lycaenid butterflies in the genus Maculinea, for example, exploit societies of different Myrmica ant species across their ranges, but there is only rare evidence that they simultaneously utilise multiple hosts at a local site, even where alternative hosts are present. We present a simple population-genetic model accounting for the proportion of two alternative hosts and the fitness of parasite genotypes on each host. In agreement with standard models, we conclude that simultaneous host use is possible whenever fitness of heterozygotes on alternative hosts is not too low. We specifically focus on host-shifting dynamics when the frequency of hosts changes. We find that (i) host shifting may proceed so rapidly that multiple host use is unlikely to be observed, (ii) back and forth transition in host use can exhibit a hysteresis loop, (iii) the parasites' host use may not be proportional to local host frequencies and be restricted to the rarer host under some conditions, and (iv) that a substantial decline in parasite abundance may typically precede a shift in host use. We conclude that focusing not just on possible equilibrium conditions but also considering the dynamics of host shifting in non-equilibrium situations may provide added insights into host-parasite systems.}, language = {en} } @phdthesis{Wilde2019, author = {Wilde, Sabrina}, title = {Einsatz von mechanistischen Biomarkern zur Charakterisierung und Bewertung von \(in\) \(vitro\) Genotoxinen}, doi = {10.25972/OPUS-18278}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-182782}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Die verf{\"u}gbaren in vitro Genotoxizit{\"a}tstests weisen hinsichtlich ihrer Spezifit{\"a}t und ihres Informationsgehalts zum vorliegenden Wirkmechanismus (Mode of Action, MoA) Einschr{\"a}nkungen auf. Um diese M{\"a}ngel zu {\"u}berwinden, wurden in dieser Arbeit zwei Ziele verfolgt, die zu der Entwicklung und Etablierung neuer in vitro Methoden zur Pr{\"u}fung auf Genotoxizit{\"a}t in der Arzneimittelentwicklung beitragen. 1. Etablierung und Bewertung einer neuen in vitro Genotoxizit{\"a}tsmethode (MultiFlow Methode) Die MultiFlow Methode basiert auf DNA-schadensassoziierten Proteinantworten von γH2AX (DNA-Doppelstrangbr{\"u}che), phosphorylierten H3 (S10) (mitotische Zellen), nukle{\"a}ren Protein p53 (Genotoxizit{\"a}t) und cleaved PARP1 (Apoptose) in TK6-Zellen. Insgesamt wurden 31 Modellsubstanzen mit dem MultiFlow Assay und erg{\"a}nzend mit dem etablierten Mikrokerntest (MicroFlow MNT), auf ihre F{\"a}higkeit verschiedene MoA-Gruppen (Aneugene/Klastogene/Nicht-Genotoxine) zu differenzieren, untersucht. Die Performance der „neuen" gegen{\"u}ber der „alten" Methode f{\"u}hrte zu einer verbesserten Sensitivit{\"a}t von 95\% gegen{\"u}ber 90\%, Spezifit{\"a}t von 90\% gegen{\"u}ber 72\% und einer MoA-Klassifizierungsrate von 85\% gegen{\"u}ber 45\% (Aneugen vs. Klastogen). 2. Identifizierung mechanistischer Biomarker zur Klassifizierung genotoxischer Substanzen Die Analyse 67 ausgew{\"a}hlter DNA-schadensassoziierter Gene in der QuantiGene Plex Methode zeigte, dass mehrere Gene gleichzeitig zur MoA-Klassifizierung beitragen k{\"o}nnen. Die Kombination der h{\"o}chstrangierten Marker BIK, KIF20A, TP53I3, DDB2 und OGG1 erm{\"o}glichte die beste Identifizierungsrate der Modellsubstanzen. Das synergetische Modell kategorisierte 16 von 16 Substanzen korrekt in Aneugene, Klastogene und Nicht-Genotoxine. Unter Verwendung der Leave-One-Out-Kreuzvalidierung wurde das Modell evaluiert und erreichte eine Sensitivit{\"a}t, Spezifit{\"a}t und Pr{\"a}diktivit{\"a}t von 86\%, 83\% und 85\%. Ergebnisse der traditionellen qPCR Methode zeigten, dass Genotoxizit{\"a}t mit TP53I3, Klastogenit{\"a}t mit ATR und RAD17 und oxidativer Stress mit NFE2L2 detektiert werden kann. Durch die Untersuchungen von posttranslationalen Modifikationen unter Verwendung der High-Content-Imaging-Technologie wurden mechanistische Assoziationen f{\"u}r BubR1 (S670) und pH3 (S28) mit Aneugenit{\"a}t, 53BP1 (S1778) und FANCD2 (S1404) mit Klastogenit{\"a}t, p53 (K373) mit Genotoxizit{\"a}t und Nrf2 (S40) mit oxidativem Stress identifiziert. Diese Arbeit zeigt, dass (Geno)toxine unterschiedliche Gen- und Proteinver{\"a}nderungen in TK6-Zellen induzieren, die zur Erfassung mechanistischer Aktivit{\"a}ten und Einteilung (geno)toxischer MoA-Gruppen (Aneugen/Klastogen/ Reaktive Sauerstoffspezies) eingesetzt werden k{\"o}nnen und daher eine bessere Risikobewertung von Wirkstoffkandidaten erm{\"o}glichen.}, subject = {Genotoxizit{\"a}t}, language = {de} } @article{VeyKapsnerFuchsetal.2019, author = {Vey, Johannes and Kapsner, Lorenz A. and Fuchs, Maximilian and Unberath, Philipp and Veronesi, Giulia and Kunz, Meik}, title = {A toolbox for functional analysis and the systematic identification of diagnostic and prognostic gene expression signatures combining meta-analysis and machine learning}, series = {Cancers}, volume = {11}, journal = {Cancers}, number = {10}, issn = {2072-6694}, doi = {10.3390/cancers11101606}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-193240}, year = {2019}, abstract = {The identification of biomarker signatures is important for cancer diagnosis and prognosis. However, the detection of clinical reliable signatures is influenced by limited data availability, which may restrict statistical power. Moreover, methods for integration of large sample cohorts and signature identification are limited. We present a step-by-step computational protocol for functional gene expression analysis and the identification of diagnostic and prognostic signatures by combining meta-analysis with machine learning and survival analysis. The novelty of the toolbox lies in its all-in-one functionality, generic design, and modularity. It is exemplified for lung cancer, including a comprehensive evaluation using different validation strategies. However, the protocol is not restricted to specific disease types and can therefore be used by a broad community. The accompanying R package vignette runs in ~1 h and describes the workflow in detail for use by researchers with limited bioinformatics training.}, language = {en} } @article{HicklHeintzBuschartTrautweinSchultetal.2019, author = {Hickl, Oskar and Heintz-Buschart, Anna and Trautwein-Schult, Anke and Hercog, Rajna and Bork, Peer and Wilmes, Paul and Becher, D{\"o}rte}, title = {Sample preservation and storage significantly impact taxonomic and functional profiles in metaproteomics studies of the human gut microbiome}, series = {Microorganisms}, volume = {7}, journal = {Microorganisms}, number = {9}, issn = {2076-2607}, doi = {10.3390/microorganisms7090367}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-195976}, year = {2019}, abstract = {With the technological advances of the last decade, it is now feasible to analyze microbiome samples, such as human stool specimens, using multi-omic techniques. Given the inherent sample complexity, there exists a need for sample methods which preserve as much information as possible about the biological system at the time of sampling. Here, we analyzed human stool samples preserved and stored using different methods, applying metagenomics as well as metaproteomics. Our results demonstrate that sample preservation and storage have a significant effect on the taxonomic composition of identified proteins. The overall identification rates, as well as the proportion of proteins from Actinobacteria were much higher when samples were flash frozen. Preservation in RNAlater overall led to fewer protein identifications and a considerable increase in the share of Bacteroidetes, as well as Proteobacteria. Additionally, a decrease in the share of metabolism-related proteins and an increase of the relative amount of proteins involved in the processing of genetic information was observed for RNAlater-stored samples. This suggests that great care should be taken in choosing methods for the preservation and storage of microbiome samples, as well as in comparing the results of analyses using different sampling and storage methods. Flash freezing and subsequent storage at -80 °C should be chosen wherever possible.}, language = {en} } @phdthesis{Beck2019, author = {Beck, Sebastian}, title = {Using optogenetics to influence the circadian clock of \(Drosophila\) \(melanogaster\)}, doi = {10.25972/OPUS-18495}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-184952}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Almost all life forms on earth have adapted to the most impactful and most predictable recurring change in environmental condition, the cycle of day and night, caused by the axial rotation of the planet. As a result many animals have evolved intricate endogenous clocks, which adapt and synchronize the organisms' physiology, metabolism and behaviour to the daily change in environmental conditions. The scientific field researching these endogenous clocks is called chronobiology and has steadily grown in size, scope and relevance since the works of the earliest pioneers in the 1960s. The number one model organism for the research of circadian clocks is the fruit fly, Drosophila melanogaster, whose clock serves as the entry point to understanding the basic inner workings of such an intricately constructed endogenous timekeeping system. In this thesis it was attempted to combine the research on the circadian clock with the techniques of optogenetics, a fairly new scientific field, launched by the discovery of Channelrhodopsin 2 just over 15 years ago. Channelrhodopsin 2 is a light-gated ion channel found in the green alga Chlamydomonas reinhardtii. In optogenetics, researches use these light-gated ion channels like Channelrhodopsin 2 by heterologously expressing them in cells and tissues of other organisms, which can then be stimulated by the application of light. This is most useful when studying neurons, as these channels provide an almost non-invasive tool to depolarize the neuronal plasma membranes at will. The goal of this thesis was to develop an optogenetic tool, which would be able to influence and phase shift the circadian clock of Drosophila melanogaster upon illumination. A phase shift is the adaptive response of the circadian clock to an outside stimulus that signals a change in the environmental light cycle. An optogenetic tool, able to influence and phase shift the circadian clock predictably and reliably, would open up many new ways and methods of researching the neuronal network of the clock and which neurons communicate to what extent, ultimately synchronizing the network. The first optogenetic tool to be tested in the circadian clock of Drosophila melanogaster was ChR2-XXL, a channelrhodopsin variant with dramatically increased expression levels and photocurrents combined with a prolonged open state. The specific expression of ChR2-XXL and of later constructs was facilitated by deploying the three different clock-specific GAL4-driver lines, clk856-gal4, pdf-gal4 and mai179-gal4. Although ChR2-XXL was shown to be highly effective at depolarizing neurons, these stimulations proved to be unable to significantly phase shift the circadian clock of Drosophila. The second series of experiments was conducted with the conceptually novel optogenetic tools Olf-bPAC and SthK-bPAC, which respectively combine a cyclic nucleotide-gated ion channel (Olf and SthK) with the light-activated adenylyl-cyclase bPAC. These tools proved to be quite useful when expressed in the motor neurons of instar-3 larvae of Drosophila, paralyzing the larvae upon illumination, as well as affecting body length. This way, these new tools could be precisely characterized, spawning a successfully published research paper, centered around their electrophysiological characterization and their applicability in model organisms like Drosophila. In the circadian clock however, these tools caused substantial damage, producing severe arrhythmicity and anomalies in neuronal development. Using a temperature-sensitive GAL80-line to delay the expression until after the flies had eclosed, yielded no positive results either. The last series of experiments saw the use of another new series of optogenetic tools, modelled after the Olf-bPAC, with bPAC swapped out for CyclOp, a membrane-bound guanylyl-cyclase, coupled with less potent versions of the Olf. This final attempt however also ended up being unsuccessful. While these tools could efficiently depolarize neuronal membranes upon illumination, they were ultimately unable to stimulate the circadian clock in way that would cause it to phase shift. Taken together, these mostly negative results indicate that an optogenetic manipulation of the circadian clock of Drosophila melanogaster is an extremely challenging subject. As light already constitutes the most impactful environmental factor on the circadian clock, the combination of chronobiology with optogenetics demands the parameters of the conducted experiments to be tuned with an extremely high degree of precision, if one hopes to receive positive results from these types of experiments at all.}, subject = {Chronobiologie}, language = {en} } @article{Puetz2019, author = {P{\"u}tz, Stephanie M.}, title = {Mbt/PAK4 together with SRC modulates N-Cadherin adherens junctions in the developing Drosophila eye}, series = {Biology Open}, volume = {8}, journal = {Biology Open}, doi = {10.1242/bio.038406}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-200898}, pages = {bio038406}, year = {2019}, abstract = {Tissue morphogenesis is accompanied by changes of adherens junctions (AJ). During Drosophila eye development, AJ reorganization includes the formation of isolated N-Cadherin AJ between photoreceptors R3/R4. Little is known about how these N-Cadherin AJ are established and maintained. This study focuses on the kinases Mbt/PAK4 and SRC, both known to alter E-Cadherin AJ across phyla. Drosophila p21-activated kinase Mbt and the non-receptor tyrosine kinases Src64 and Src42 regulate proper N-Cadherin AJ. N-Cadherin AJ elongation depends on SRC kinase activity. Cell culture experiments demonstrate binding of both Drosophila SRC isoforms to N-Cadherin and its subsequent tyrosine phosphorylation. In contrast, Mbt stabilizes but does not bind N-Cadherin in vitro. Mbt is required in R3/R4 for zipping the N-Cadherin AJ between these cells, independent of its kinase activity and Cdc42-binding. The mbt phenotype can be reverted by mutations in Src64 and Src42. Because Mbt neither directly binds to SRC proteins nor has a reproducible influence on their kinase activity, the conclusion is that Mbt and SRC signaling converge on N-Cadherin. N-Cadherin AJ formation during eye development requires a proper balance between the promoting effects of Mbt and the inhibiting influences of SRC kinases.}, language = {en} } @phdthesis{Breitenbach2019, author = {Breitenbach, Tim}, title = {A mathematical optimal control based approach to pharmacological modulation with regulatory networks and external stimuli}, doi = {10.25972/OPUS-17436}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-174368}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {In this work models for molecular networks consisting of ordinary differential equations are extended by terms that include the interaction of the corresponding molecular network with the environment that the molecular network is embedded in. These terms model the effects of the external stimuli on the molecular network. The usability of this extension is demonstrated with a model of a circadian clock that is extended with certain terms and reproduces data from several experiments at the same time. Once the model including external stimuli is set up, a framework is developed in order to calculate external stimuli that have a predefined desired effect on the molecular network. For this purpose the task of finding appropriate external stimuli is formulated as a mathematical optimal control problem for which in order to solve it a lot of mathematical methods are available. Several methods are discussed and worked out in order to calculate a solution for the corresponding optimal control problem. The application of the framework to find pharmacological intervention points or effective drug combinations is pointed out and discussed. Furthermore the framework is related to existing network analysis tools and their combination for network analysis in order to find dedicated external stimuli is discussed. The total framework is verified with biological examples by comparing the calculated results with data from literature. For this purpose platelet aggregation is investigated based on a corresponding gene regulatory network and associated receptors are detected. Furthermore a transition from one to another type of T-helper cell is analyzed in a tumor setting where missing agents are calculated to induce the corresponding switch in vitro. Next a gene regulatory network of a myocardiocyte is investigated where it is shown how the presented framework can be used to compare different treatment strategies with respect to their beneficial effects and side effects quantitatively. Moreover a constitutively activated signaling pathway, which thus causes maleficent effects, is modeled and intervention points with corresponding treatment strategies are determined that steer the gene regulatory network from a pathological expression pattern to physiological one again.}, subject = {Bioinformatik}, language = {en} } @article{MatosMachadoSchartletal.2019, author = {Matos, Isa and Machado, Miguel P. and Schartl, Manfred and Coelho, Maria Manuela}, title = {Allele-specific expression variation at different ploidy levels in Squalius alburnoides}, series = {Scientific Reports}, volume = {9}, journal = {Scientific Reports}, doi = {10.1038/s41598-019-40210-8}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-200910}, pages = {3688}, year = {2019}, abstract = {Allopolyploid plants are long known to be subject to a homoeolog expression bias of varying degree. The same phenomenon was only much later suspected to occur also in animals based on studies of single selected genes in an allopolyploid vertebrate, the Iberian fish Squalius alburnoides. Consequently, this species became a good model for understanding the evolution of gene expression regulation in polyploid vertebrates. Here, we analyzed for the first time genome-wide allele-specific expression data from diploid and triploid hybrids of S. alburnoides and compared homoeolog expression profiles of adult livers and of juveniles. Co-expression of alleles from both parental genomic types was observed for the majority of genes, but with marked homoeolog expression bias, suggesting homoeolog specific reshaping of expression level patterns in hybrids. Complete silencing of one allele was also observed irrespective of ploidy level, but not transcriptome wide as previously speculated. Instead, it was found only in a restricted number of genes, particularly ones with functions related to mitochondria and ribosomes. This leads us to hypothesize that allelic silencing may be a way to overcome intergenomic gene expression interaction conflicts, and that homoeolog expression bias may be an important mechanism in the achievement of sustainable genomic interactions, mandatory to the success of allopolyploid systems, as in S. alburnoides.}, language = {en} } @phdthesis{Jannasch2019, author = {Jannasch, Maren Annika}, title = {In vitro Fremdk{\"o}rpermodellsysteme zur Vorhersage von biomaterialinduzierten Immunreaktionen}, doi = {10.25972/OPUS-16289}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-162893}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Die Implantation eines Medizinprodukts in den menschlichen K{\"o}rper ruft eine Immunreaktion hervor, die zur fibr{\"o}sen Einkapselung f{\"u}hren kann. Makrophagen in direktem Kontakt mit der Oberfl{\"a}che des Implantats erfassen sensorisch den Fremdk{\"o}rper und {\"u}bersetzten das Signal in die Freisetzung zahlreicher l{\"o}slicher Mediatoren. Das generierte Entz{\"u}ndungsmilieu moduliert die Heilungsreaktion und kann zur Anreicherung von Fibroblasten sowie zur Erh{\"o}hung der Matrixsyntheserate in der Wundumgebung f{\"u}hren. Eine dichte fibr{\"o}se Kapsel um ein Medizinprodukt beeintr{\"a}chtigt den Ersatz von K{\"o}rperstrukturen, das Unterst{\"u}tzen physiologischer K{\"o}rperfunktionen sowie die Effizienz einer medizinischen Therapie. Zur Identifizierung potenzieller Biomaterialkandidaten mit optimalen Eigenschaften ist jedoch eine evidenzbasierte Entscheidungsfindung notwendig und diese wiederum muss durch geeignete Testmethoden unterst{\"u}tzt werden. Zur Erfassung lokaler Effekte nach Implantation eines Biomaterials begr{\"u}ndet die Komplexi-t{\"a}t der ablaufenden Fremdk{\"o}rperreaktion die Anwendung von Tiermodellen als Goldstandard. Die Eingliederung von in vitro Modellsystemen in standardisierte Testverfahren scheitert oft an der Verf{\"u}gbarkeit validierter, verl{\"a}sslicher und reproduzierbarer Methoden. Demnach ist kein standardisiertes in vitro Testverfahren beschrieben, das die komplexen dreidimensionalen Gewebsstrukturen w{\"a}hrend einer Fremdk{\"o}rperreaktion abbildet und sich zur Testung {\"u}ber l{\"a}ngere Kontaktphasen zwischen Blutkomponenten und Biomaterialien eignet. Jedoch k{\"o}nnen in vitro Testungen kosten- und zeiteffizienter sein und durch die Anwendung humaner Zellen eine h{\"o}here {\"U}bertragbarkeit auf den Menschen aufweisen. Zus{\"a}tzlich adressiert die Pr{\"a}ferenz zu in vitro Testmethoden den Aspekt „Reduzierung" der 3R-Prinzipien „Replacement, Reduction, Refinement" (Ersatz, Reduzierung, Verbesserung) von Russel und Burch (1959) zu einer bewussten und begr{\"u}ndeten Anwendung von Tiermodellen in der Wissenschaft. Ziel von diesem Forschungsvorhaben war die Entwicklung von humanen in vitro Modellsystemen, die den Kontakt zu Blutkomponenten sowie die Reaktion des umliegenden Bindegewebes bei lokaler Implantation eines Biomaterials abbilden. Referenzmaterialien, deren Gewebsantwort nach Implantation in Tiere oder den Menschen bekannt ist, dienten als Validierungskriterium f{\"u}r die entwickelten Modellsysteme. Die Anreicherung von Zellen sowie die Bildung extrazellul{\"a}rer Matrix in der Wundumgebung stellen wichtige Teilprozesse w{\"a}hrend einer Fremdk{\"o}rperreaktion dar. F{\"u}r beide Teilprozesse konnte in einem indirekten zellbasierten Modellsystem der Einfluss einer zellvermittelten Konditionierung wie die Freisetzung von l{\"o}slichen Mediatoren durch materialadh{\"a}rente Makrophagen auf die gerichtete Wanderung von Fibroblasten sowie den Umbau eines dreidimensionalen Bindegewebsmodells aufgezeigt werden. Des Weiteren ließ sich das Freisetzungsprofil von Zytokinen durch materialst{\"a}ndige Makrophagen unter verschiedenen Testbedingungen wie der Kontamination mit LPS, der Oberfl{\"a}chenbehandlung mit humanem Blutplasma und der Gegenwart von IL-4 bestimmen. Die anschließende vergleichende statistische Modellierung der generierten komplexen multifaktoriellen Datenmatrix erm{\"o}glichte die {\"U}bersetzung in eine Biomaterialbewertung. Dieses entwickelte Testverfahren eignete sich einerseits zur Validierung von in vitro Testbedingungen sowie andererseits zur Bewertung von Biomaterialien. Dar{\"u}ber hinaus konnte in einem dreidimensionalen Fremdk{\"o}rpermodell die komplexe dreidimensionale Struktur der extrazellul{\"a}ren Matrix in einer Wunde durch die Kombination unterschiedlicher Zell- und Matrixkomponenten biomimetisch nachgebaut werden. Diese neuartigen dreidimensionalen Fremdk{\"o}rpermodelle erm{\"o}glichten die Testung von Biomaterialien {\"u}ber l{\"a}ngere Testphasen und k{\"o}nnen in anschließenden Studien angewandt werden, um dynamische Prozesse zu untersuchen. Zusammenfassend konnten in dieser Arbeit drei unterschiedliche Teststrategien entwickelt werden, die (I) die Bewertung von Teilprozessen erm{\"o}glichen, (II) die Identifizierung verl{\"a}sslicher Testbedingungen unterst{\"u}tzen und (III) biomimetisch ein Wundgewebe abbilden. Wesentlich ist, dass biomimetisch ein dreidimensionales Gewebemodell entwickelt werden konnte, das eine verl{\"a}ssliche Unterscheidungskapazit{\"a}t zwischen Biomaterialien aufweist.}, subject = {Biomaterial}, language = {de} } @phdthesis{Yu2019, author = {Yu, Sung-Huan}, title = {Development and application of computational tools for RNA-Seq based transcriptome annotations}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-176468}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {In order to understand the regulation of gene expression in organisms, precise genome annotation is essential. In recent years, RNA-Seq has become a potent method for generating and improving genome annotations. However, this Approach is time consuming and often inconsistently performed when done manually. In particular, the discovery of non-coding RNAs benefits strongly from the application of RNA-Seq data but requires significant amounts of expert knowledge and is labor-intensive. As a part of my doctoral study, I developed a modular tool called ANNOgesic that can detect numerous transcribed genomic features, including non-coding RNAs, based on RNA-Seq data in a precise and automatic fashion with a focus on bacterial and achaeal species. The software performs numerous analyses and generates several visualizations. It can generate annotations of high-Resolution that are hard to produce using traditional annotation tools that are based only on genome sequences. ANNOgesic can detect numerous novel genomic Features like UTR-derived small non-coding RNAs for which no other tool has been developed before. ANNOgesic is available under an open source license (ISCL) at https://github.com/Sung-Huan/ANNOgesic. My doctoral work not only includes the development of ANNOgesic but also its application to annotate the transcriptome of Staphylococcus aureus HG003 - a strain which has been a insightful model in infection biology. Despite its potential as a model, a complete genome sequence and annotations have been lacking for HG003. In order to fill this gap, the annotations of this strain, including sRNAs and their functions, were generated using ANNOgesic by analyzing differential RNA-Seq data from 14 different samples (two media conditions with seven time points), as well as RNA-Seq data generated after transcript fragmentation. ANNOgesic was also applied to annotate several bacterial and archaeal genomes, and as part of this its high performance was demonstrated. In summary, ANNOgesic is a powerful computational tool for RNA-Seq based annotations and has been successfully applied to several species.}, subject = {Genom}, language = {en} } @phdthesis{Memmel2019, author = {Memmel, Simon}, title = {Automatisierte Algorithmen zur Analyse der Migration und der strahleninduzierten DNA-Sch{\"a}den humaner Glioblastomzellen nach kombinierter PI3K/mTOR/Hsp90-Inhibierung}, doi = {10.25972/OPUS-18571}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-185710}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Das hohe invasive Potential und die starke Resistenz gegen Radio-/Chemotherapie von Glioblastoma multiforme (GBM) Zellen machen sie zu dem t{\"o}dlichsten Tumor ihrer Art. Es ist deshalb von großem Interesse die Grundlagen, welche der Migrationsf{\"a}higkeit und DNA Reparatur zu Grunde liegen, besser zu verstehen. Im ersten Teil dieser Arbeit wurden zwei Algorithmen zur automatischen Analyse der Migration in der Einzelzellverfolgung und im Wundheilungsassay modifiziert. Die Auswertung der Daten konnte automatisch und somit schnell, effektiv und mit geringerem Arbeitsaufwand durchgef{\"u}hrt werden. Mit Hilfe dieser automatischen Algorithmen wurde die Migrationsf{\"a}higkeit von zwei GBM-Zelllinien (DK-MG und SNB19) untersucht. Zus{\"a}tzlich wurde die konfokale Laserscanning- sowie die hochaufl{\"o}sende dSTORM-Fluoreszenzmikroskopie verwendet um die, der Zellbewegung zu Grunde liegende, Struktur des F Aktin und der fokalen Adh{\"a}sionskinase (FAK) aufzul{\"o}sen und darzustellen. Unter Anwendung dieser genannten Methoden sind die Effekte des dualen PI3K/mTOR Inhibitors PI-103 alleine und in Kombination mit dem Hsp90 Inhibitor NVP AUY922 mit und ohne Bestrahlung auf die Bewegung untersucht worden. Es konnte festgestellt werden, dass sich beide Zelllinien deutlich in ihrem migratorischem Potential in vitro unterscheiden und zudem auch markante Unterschiede in ihrer Morphologie aufweisen. Die weniger invasiven DK MG-Zellen besitzen eine polarisierte Zellstruktur, wohingegen SNB19-Zellen sich durch multipolare ungerichtete Bewegung auszeichneten. Zudem wurde die Migration, durch PI3K/mTOR Inhibition mit PI-103 bei den DK-MG-Zellen (p53 wt, PTEN wt), sehr effektiv unterdr{\"u}ckt. Wohingegen sich die SNB19-Zellen (p53 mut, PTEN mut) resistent gegen diesen Inhibitor zeigten. Hsp90 Inhibition offenbarte in beiden Zelllinien einen starken inhibitorischen Effekt auf die Migration der Zellen sowie die Reorganisierung des F Aktinskelettes. In der zweiten H{\"a}lfte dieser Arbeit wurde ein Augenmerk auf die DNA-DSB-Reparatur der GBM Zellen nach ionisierender Strahlung gelegt. Zun{\"a}chst wurde eine automatische Analysesoftware „FocAn-3D" entwickelt, mit dessen Hilfe die DNA Doppelstrangbruchreparaturkinetik untersucht werden sollte. Diese Software erm{\"o}glicht es die gesamten Zellkerne mit ihren γH2AX-Foci in 3D-cLSM-Aufnahmen zu untersuchen. Es konnte somit eine Verbesserung der Genauigkeit in der Ausz{\"a}hlung der γH2AX-Foci erreicht werden, welche 2D beschr{\"a}nkter Software verwehrt bleibt. Mit FocAn-3D konnte der gesamte Verlauf der Induktions- und Abbauphase der γH2AX-Foci in DK MG- und SNB19-Zellen mit einem mathematischen Modell ausgewertet und dargestellt werden. Des Weiteren wurde die Nanometerstruktur von γH2AX- und pDNA-PKcs-Foci mittels hochaufl{\"o}sender dSTORM-Mikroskopie untersucht. Konventionelle Mikroskopiemethoden, begrenzt durch das Beugungslimit und einer Aufl{\"o}sung von ~200 nm, konnten die Nanometerstruktur (<100 nm) der Reparaturfoci bisher nicht darstellen. Mit Hilfe der beugungsunbegrenzten dSTORM-Mikroskopie war es m{\"o}glich in DK MG- und SNB19-Zellen die Nanometerstruktur genannten Reparaturproteine in den Foci mit einer Aufl{\"o}sung von bis zu ~20 nm darzustellen. γH2AX-Foci zeigten sich als eine Verteilung aus einzelnen Untereinheiten („Nanofoci") mit einem Durchmesser von ~45 nm. Dies l{\"a}sst die Vermutung zu, dass es sich hier um die elementare Substruktur der Foci und somit der γH2AX enthaltenen Nukleosome handelt. DNA-PK-Foci wiesen hingegen eine diffusere Verteilung auf. Die in dieser Arbeit ermittelten Unterschiede im Migrationsverhalten der Zellen rechtfertigen eine weitere pr{\"a}klinische Untersuchung der verwendeten Inhibitoren als potentielle Zelltherapeutika f{\"u}r die Behandlung von GBM. Zudem konnte sich dSTORM als machtvolles Hilfsmittel, sowohl zur Analyse der Migration zugrundeliegenden Zytoskelettstruktur und der Effekte der Hsp90 Inhibierung, als auch, der Nanostruktur der DNA-DSB-Reparaturfoci herausstellen. Es ist anzunehmen, dass beugungsunbegrenzte Mikroskopiemethoden sich als bedeutende Werkzeuge in der medizinischen und biologischen Erforschung der DNA-Reparaturmechanismen herausstellen werden. Das in dieser Arbeit entwickelte ImageJ Plugin „FocAn-3D" bewies sich ebenfalls als ein vielversprechendes Werkzeug f{\"u}r die Analyse der Reparaturkinetik. Mit Hilfe von „FocAn-3D" sollte es somit m{\"o}glich sein u.a. den Einfluss gezielter Inhibition auf den zeitlichen Verlauf der Induktion und des Abbaus der DNA-Reparaturmaschinerie genauer zu studieren.}, subject = {Glioblastom}, language = {de} } @phdthesis{Kaymak2019, author = {Kaymak, Irem}, title = {Identification of metabolic liabilities in 3D models of cancer}, doi = {10.25972/OPUS-18154}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-181544}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Inefficient vascularisation of solid tumours leads to the formation of oxygen and nutrient gradients. In order to mimic this specific feature of the tumour microenvironment, a multicellular tumour spheroid (SPH) culture system was used. These experiments were implemented in p53 isogenic colon cancer cell lines (HCT116 p53 +/+ and HCT116 p53-/-) since Tp53 has important regulatory functions in tumour metabolism. First, the characteristics of the cells cultured as monolayers and as spheroids were investigated by using RNA sequencing and metabolomics to compare gene expression and metabolic features of cells grown in different conditions. This analysis showed that certain features of gene expression found in tumours are also present in spheroids but not in monolayer cultures, including reduced proliferation and induction of hypoxia related genes. Moreover, comparison between the different genotypes revealed that the expression of genes involved in cholesterol homeostasis is induced in p53 deficient cells compared to p53 wild type cells and this difference was only detected in spheroids and tumour samples but not in monolayer cultures. In addition, it was established that loss of p53 leads to the induction of enzymes of the mevalonate pathway via activation of the transcription factor SREBP2, resulting in a metabolic rewiring that supports the generation of ubiquinone (coenzyme Q10). An adequate supply of ubiquinone was essential to support mitochondrial electron transport and pyrimidine biosynthesis in p53 deficient cancer cells under conditions of metabolic stress. Moreover, inhibition of the mevalonate pathway using statins selectively induced oxidative stress and apoptosis in p53 deficient colon cancer cells exposed to oxygen and nutrient deprivation. This was caused by ubiquinone being required for electron transfer by dihydroorotate dehydrogenase, an essential enzyme of the pyrimidine nucleotide biosynthesis pathway. Supplementation with exogenous nucleosides relieved the demand for electron transfer and restored viability of p53 deficient cancer cells under metabolic stress. Moreover, the mevalonate pathway was also essential for the synthesis of ubiquinone for nucleotide biosynthesis to support growth of intestinal tumour organoids. Together, these findings highlight the importance of the mevalonate pathway in cancer cells and provide molecular evidence for an enhanced sensitivity towards the inhibition of mitochondrial electron transfer in tumour-like metabolic environments.}, subject = {Tumor}, language = {en} } @phdthesis{Gorelashvili2019, author = {Gorelashvili, Maximilian Georg}, title = {Investigation of megakaryopoiesis and the acute phase of ischemic stroke by advanced fluorescence microscopy}, doi = {10.25972/OPUS-18600}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-186002}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {In mammals, anucleate platelets circulate in the blood flow and are primarily responsible for maintaining functional hemostasis. Platelets are generated in the bone marrow (BM) by megakaryocytes (MKs), which mainly reside directly next to the BM sinusoids to release proplatelets into the blood. MKs originate from hematopoietic stem cells and are thought to migrate from the endosteal to the vascular niche during their maturation, a process, which is, despite being intensively investigated, still not fully understood. Long-term intravital two photon microscopy (2PM) of MKs and vasculature in murine bone marrow was performed and mean squared displacement analysis of cell migration was performed. The MKs exhibited no migration, but wobbling-like movement on time scales of 3 h. Directed cell migration always results in non-random spatial distribution. Thus, a computational modelling algorithm simulating random MK distribution using real 3D light-sheet fluorescence microscopy data sets was developed. Direct comparison of real and simulated random MK distributions showed, that MKs exhibit a strong bias to vessel-contact. However, this bias is not caused by cell migration, as non-vessel-associated MKs were randomly distributed in the intervascular space. Furthermore, simulation studies revealed that MKs strongly impair migration of other cells in the bone marrow by acting as large-sized obstacles. MKs are thought to migrate from the regions close to the endosteum towards the vasculature during their maturation process. MK distribution as a function of their localization relative to the endosteal regions of the bones was investigated by light sheet fluorescence microscopy (LSFM). The results show no bone-region dependent distribution of MKs. Taken together, the newly established methods and obtained results refute the model of MK migration during their maturation. Ischemia reperfusion (I/R) injury is a frequent complication of cerebral ischemic stroke, where brain tissue damage occurs despite successful recanalization. Platelets, endothelial cells and immune cells have been demonstrated to affect the progression of I/R injury in experimental mouse models 24 h after recanalization. However, the underlying Pathomechanisms, especially in the first hours after recanalization, are poorly understood. Here, LSFM, 2PM and complemental advanced image analysis workflows were established for investigation of platelets, the vasculature and neutrophils in ischemic brains. Quantitative analysis of thrombus formation in the ipsilateral and contralateral hemispheres at different time points revealed that platelet aggregate formation is minimal during the first 8 h after recanalization and occurs in both hemispheres. Considering that maximal tissue damage already is present at this time point, it can be concluded that infarct progression and neurological damage do not result from platelet aggregated formation. Furthermore, LSFM allowed to confirm neutrophil infiltration into the infarcted hemisphere and, here, the levels of endothelial cell marker PECAM1 were strongly reduced. However, further investigations must be carried out to clearly identify the role of neutrophils and the endothelial cells in I/R injury.}, subject = {Fluoreszenzmikroskopie}, language = {en} } @phdthesis{Fuellsack2019, author = {F{\"u}llsack, Simone Alexandra}, title = {Die Bedeutung von Todesdom{\"a}ne Adapterproteinen f{\"u}r die Signaltransduktion des TNFR1 und der TRAIL Todesrezeptoren}, doi = {10.25972/OPUS-18451}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-184518}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Die NFκB-Signalwege, Apoptose und Nekroptose sind essentielle Prozesse in der Immunantwort. Außerdem sind diese Signalwege Teil der Regulation von Zelldifferenzierung, -proliferation, -tod und Entz{\"u}ndungsreaktionen. Dabei wird zuerst der Rezeptor (TNFR1 oder TRAILR 1/2) aktiviert, die rekrutierten DD-Adapterproteine TRADD, FADD und RIPK1 leiten dann die entsprechende Signalkaskade weiter und bestimmen durch ihre Zusammenwirkung, ob der NFκB-Signalweg, Apoptose oder Nekroptose induziert wird. TNFR1 und TRAILR 1/2 ben{\"o}tigen die DD-Adapterproteine TRADD, FADD und RIPK1 f{\"u}r die Zelltodinduktion, deren konkrete Bedeutung in Bezug auf Rezeptor-Spezifit{\"a}t, Zusammenwirken und Relevanz allerdings noch unklar ist. Um das Zusammenspiel dieser Proteine besser zu verstehen, wurden in dieser Arbeit Nekroptose-kompetente RIPK3-exprimierende HeLa-Zellen verwendet, bei denen die DD-Adapterproteine FADD, TRADD und RIPK1 einzeln oder in Kombination von zweien ausgeknockt wurden. Es stellte sich heraus, dass RIPK1 essentiell f{\"u}r die TNFR1- und TRAILR 1/2-vermittelte Nekroptose-Induktion ist, doch RIPK1 alleine, d.h. ohne FADD- oder TRADD-Mitbeteiligung, nur bei der TNFR1-Nekroptose-Induktion ausreicht. Wiederum inhibiert TRADD die TNFR1- und TRAILR 1/2-induzierte Nekroptose. RIPK1 und TRADD sind aber unverzichtbar f{\"u}r die NFκB-Aktivierung durch TNFR1 oder TRAILR 1/2 und spielen eine wichtige Rolle bei TNFR1-induzierter Apoptose. Andererseits ist FADD alleine ausreichend f{\"u}r die TRAILR 1/2-bezogene Caspase-8 Aktivierung. Zudem ist FADD notwendig f{\"u}r die TRAIL-induzierte NFκB-Signalaktivierung. In Abwesenheit von FADD und TRADD vermittelt RIPK1 die TNF-induzierte Caspase-8 Aktivierung. FADD wird f{\"u}r die TRAIL-induzierte Nekroptose ben{\"o}tigt, aber gegenl{\"a}ufig wirkt die TNF-induzierte Nektroptose in einer Caspase-8 abh{\"a}ngigen und unabh{\"a}ngigen Weise. Zudem sensitiviert TWEAK die TNF- und TRAIL-induzierte Nekroptose. Zusammenfassend wurde in dieser Arbeit die Auswirkung von TNFR1 und TRAILR 1/2 auf die Aktivierung der unterschiedlichen Signalkaskaden untersucht. Des Weiteren wurde gezeigt, in welcher Weise sich das Zusammenspiel von TRADD, FADD und RIPK1 auf die Induktion von NFκB, Apoptose und Nekroptose auswirkt.}, subject = {Signaltransduktion}, language = {de} }