@article{BeckerOelschlaegerWullaertetal.2013,
  author    = {Becker, Svetlana and Oelschlaeger, Tobias A. and Wullaert, Andy and Pasparakis, Manolis and Wehkamp, Jan and Stange, Eduard F. and Gersemann, Michael},
  title     = {Bacteria Regulate Intestinal Epithelial Cell Differentiation Factors Both In Vitro and In Vivo},
  series = {PLoS ONE},
  volume    = {8},
  journal   = {PLoS ONE},
  number    = {2},
  doi       = {10.1371/journal.pone.0055620},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-131168},
  pages     = {e55620},
  year      = {2013},
  abstract  = {Background: The human colon harbours a plethora of bacteria known to broadly impact on mucosal metabolism and function and thought to be involved in inflammatory bowel disease pathogenesis and colon cancer development. In this report, we investigated the effect of colonic bacteria on epithelial cell differentiation factors in vitro and in vivo. As key transcription factors we focused on Hes1, known to direct towards an absorptive cell fate, Hath1 and KLF4, which govern goblet cell. Methods: Expression of the transcription factors Hes1, Hath1 and KLF4, the mucins Muc1 and Muc2 and the defensin HBD2 were measured by real-time PCR in LS174T cells following incubation with several heat-inactivated E. coli strains, including the probiotic E. coli Nissle 1917+/- flagellin, Lactobacilli and Bifidobacteria. For protein detection Western blot experiments and chamber-slide immunostaining were performed. Finally, mRNA and protein expression of these factors was evaluated in the colon of germfree vs. specific pathogen free vs. conventionalized mice and colonic goblet cells were counted. Results: Expression of Hes1 and Hath1, and to a minor degree also of KLF4, was reduced by E. coli K-12 and E. coli Nissle 1917. In contrast, Muc1 and HBD2 expression were significantly enhanced, independent of the Notch signalling pathway. Probiotic E. coli Nissle 1917 regulated Hes1, Hath1, Muc1 and HBD2 through flagellin. In vivo experiments confirmed the observed in vitro effects of bacteria by a diminished colonic expression of Hath1 and KLF4 in specific pathogen free and conventionalized mice as compared to germ free mice whereas the number of goblet cells was unchanged in these mice. Conclusions: Intestinal bacteria influence the intestinal epithelial differentiation factors Hes1, Hath1 and KLF4, as well as Muc1 and HBD2, in vitro and in vivo. The induction of Muc1 and HBD2 seems to be triggered directly by bacteria and not by Notch.},
  language  = {en}
}
@article{EhrigKilincChenetal.2013,
  author    = {Ehrig, Klaas and Kilinc, Mehmet O. and Chen, Nanhai G. and Stritzker, Jochen and Buckel, Lisa and Zhang, Qian and Szalay, Aladar A.},
  title     = {Growth inhibition of different human colorectal cancer xenografts after a single intravenous injection of oncolytic vaccinia virus GLV-1h68},
  series = {Journal of Translational Medicine},
  volume    = {11},
  journal   = {Journal of Translational Medicine},
  number    = {79},
  doi       = {10.1186/1479-5876-11-79},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-129619},
  year      = {2013},
  abstract  = {Background: Despite availability of efficient treatment regimens for early stage colorectal cancer, treatment regimens for late stage colorectal cancer are generally not effective and thus need improvement. Oncolytic virotherapy using replication-competent vaccinia virus (VACV) strains is a promising new strategy for therapy of a variety of human cancers. Methods: Oncolytic efficacy of replication-competent vaccinia virus GLV-1h68 was analyzed in both, cell cultures and subcutaneous xenograft tumor models. Results: In this study we demonstrated for the first time that the replication-competent recombinant VACV GLV-1h68 efficiently infected, replicated in, and subsequently lysed various human colorectal cancer lines (Colo 205, HCT-15, HCT-116, HT-29, and SW-620) derived from patients at all four stages of disease. Additionally, in tumor xenograft models in athymic nude mice, a single injection of intravenously administered GLV-1h68 significantly inhibited tumor growth of two different human colorectal cell line tumors (Duke's type A-stage HCT-116 and Duke's type C-stage SW-620), significantly improving survival compared to untreated mice. Expression of the viral marker gene ruc-gfp allowed for real-time analysis of the virus infection in cell cultures and in mice. GLV-1h68 treatment was well-tolerated in all animals and viral replication was confined to the tumor. GLV-1h68 treatment elicited a significant up-regulation of murine immune-related antigens like IFN-γ, IP-10, MCP-1, MCP-3, MCP-5, RANTES and TNF-γ and a greater infiltration of macrophages and NK cells in tumors as compared to untreated controls. Conclusion: The anti-tumor activity observed against colorectal cancer cells in these studies was a result of direct viral oncolysis by GLV-1h68 and inflammation-mediated innate immune responses. The therapeutic effects occurred in tumors regardless of the stage of disease from which the cells were derived. Thus, the recombinant vaccinia virus GLV-1h68 has the potential to treat colorectal cancers independently of the stage of progression.},
  language  = {en}
}
@article{MielichSuessSchneiderLopez2013,
  author    = {Mielich-S{\"u}ss, Benjamin and Schneider, Johannes and Lopez, Daniel},
  title     = {Overproduction of Flotillin Influences Cell Differentiation and Shape in Bacillus subtilis},
  series = {mBio},
  volume    = {4},
  journal   = {mBio},
  number    = {6},
  doi       = {10.1128/mBio.00719-13},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-129653},
  year      = {2013},
  abstract  = {ABSTRACT Bacteria organize many membrane-related signaling processes in functional microdomains that are structurally and functionally similar to the lipid rafts of eukaryotic cells. An important structural component of these microdomains is the protein flotillin, which seems to act as a chaperone in recruiting other proteins to lipid rafts to facilitate their interaction. In eukaryotic cells, the occurrence of severe diseases is often observed in combination with an overproduction of flotillin, but a functional link between these two phenomena is yet to be demonstrated. In this work, we used the bacterial model Bacillus subtilis as a tractable system to study the physiological alterations that occur in cells that overproduce flotillin. We discovered that an excess of flotillin altered specific signal transduction pathways that are associated with the membrane microdomains of bacteria. As a consequence of this, we detected significant defects in cell division and cell differentiation. These physiological alterations were in part caused by an unusual stabilization of the raft-associated protease FtsH. This report opens the possibility of using bacteria as a working model to better understand fundamental questions related to the functionality of lipid rafts. IMPORTANCE The identification of signaling platforms in the membrane of bacteria that are functionally and structurally equivalent to eukaryotic lipid rafts reveals a level of sophistication in signal transduction and membrane organization unexpected in bacteria. It opens new and promising venues to address intricate questions related to the functionality of lipid rafts by using bacteria as a more tractable system. This is the first report that uses bacteria as a working model to investigate a fundamental question that was previously raised while studying the role of eukaryotic lipid rafts. It also provides evidence of the critical role of these signaling platforms in orchestrating diverse physiological processes in prokaryotic cells.},
  language  = {en}
}
@article{MakgotlhoMarincolaSchaeferetal.2013,
  author    = {Makgotlho, Phuti E. and Marincola, Gabriella and Sch{\"a}fer, Daniel and Liu, Quian and Bae, Taeok and Geiger, Tobias and Wasserman, Elizabeth and Wolz, Christine and Ziebuhr, Wilma and Sinha, Bhanu},
  title     = {SDS Interferes with SaeS Signaling of Staphylococcus aureus Independently of SaePQ},
  series = {PLOS ONE},
  volume    = {8},
  journal   = {PLOS ONE},
  number    = {8},
  issn      = {1932-6203},
  doi       = {10.1371/journal.pone.0071644},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-128469},
  pages     = {e71644},
  year      = {2013},
  abstract  = {The Staphylococcus aureus regulatory saePQRS system controls the expression of numerous virulence factors, including extracellular adherence protein (Eap), which amongst others facilitates invasion of host cells. The saePQRS operon codes for 4 proteins: the histidine kinase SaeS, the response regulator SaeR, the lipoprotein SaeP and the transmembrane protein SaeQ. S. aureus strain Newman has a single amino acid substitution in the transmembrane domain of SaeS (L18P) which results in constitutive kinase activity. SDS was shown to be one of the signals interfering with SaeS activity leading to inhibition of the sae target gene eap in strains with SaeS(L) but causing activation in strains containing SaeS(P). Here, we analyzed the possible involvement of the SaeP protein and saePQ region in SDS-mediated sae/eap expression. We found that SaePQ is not needed for SDS-mediated SaeS signaling. Furthermore, we could show that SaeS activity is closely linked to the expression of Eap and the capacity to invade host cells in a number of clinical isolates. This suggests that SaeS activity might be directly modulated by structurally non-complex environmental signals, as SDS, which possibly altering its kinase/phosphatase activity.},
  language  = {en}
}
@article{HertleinSturmJakobetal.2013,
  author    = {Hertlein, Tobias and Sturm, Volker and Jakob, Peter and Ohlsen, Knut},
  title     = {\(^{19}\)F Magnetic Resonance Imaging of Perfluorocarbons for the Evaluation of Response to Antibiotic Therapy in a Staphylococcus aureus Infection Model},
  series = {PLoS ONE},
  volume    = {8},
  journal   = {PLoS ONE},
  number    = {5},
  doi       = {10.1371/journal.pone.0064440},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-130113},
  pages     = {e64440},
  year      = {2013},
  abstract  = {Background The emergence of antibiotic resistant bacteria in recent decades has highlighted the importance of developing new drugs to treat infections. However, in addition to the design of new drugs, the development of accurate preclinical testing methods is essential. In vivo imaging technologies such as bioluminescence imaging (BLI) or magnetic resonance imaging (MRI) are promising approaches. In a previous study, we showed the effectiveness of \(^{19}\)F MRI using perfluorocarbon (PFC) emulsions for detecting the site of Staphylococcus aureus infection. In the present follow-up study, we investigated the use of this method for in vivo visualization of the effects of antibiotic therapy. Methods/Principal findings Mice were infected with S. aureus Xen29 and treated with 0.9\% NaCl solution, vancomycin or linezolid. Mock treatment led to the highest bioluminescence values during infection followed by vancomycin treatment. Counting the number of colony-forming units (cfu) at 7 days post-infection (p.i.) showed the highest bacterial burden for the mock group and the lowest for the linezolid group. Administration of PFCs at day 2 p.i. led to the accumulation of \(^{19}\)F at the rim of the abscess in all mice (in the shape of a hollow sphere), and antibiotic treatment decreased the \(^{19}\)F signal intensity and volume. Linezolid showed the strongest effect. The BLI, cfu, and MRI results were comparable. Conclusions \(^{19}\)F-MRI with PFCs is an effective non-invasive method for assessing the effects of antibiotic therapy in vivo. This method does not depend on pathogen specific markers and can therefore be used to estimate the efficacy of antibacterial therapy against a broad range of clinically relevant pathogens, and to localize sites of infection.},
  language  = {en}
}
@article{SchulteWestermannVogel2013,
  author    = {Schulte, Leon N. and Westermann, Alexander J. and Vogel, J{\"o}rg},
  title     = {Differential activation and functional specialization of miR-146 and miR-155 in innate immune sensing},
  series = {Nucleic Acids Research},
  volume    = {41},
  journal   = {Nucleic Acids Research},
  number    = {1},
  doi       = {10.1093/nar/gks1030},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-129765},
  pages     = {542-553},
  year      = {2013},
  abstract  = {Many microRNAs (miRNAs) are co-regulated during the same physiological process but the underlying cellular logic is often little understood. The conserved, immunomodulatory miRNAs miR-146 and miR-155, for instance, are co-induced in many cell types in response to microbial lipopolysaccharide (LPS) to feedback-repress LPS signalling through Toll-like receptor TLR4. Here, we report that these seemingly co-induced regulatory RNAs dramatically differ in their induction behaviour under various stimuli strengths and act non-redundantly through functional specialization; although miR-146 expression saturates at sub-inflammatory doses of LPS that do not trigger the messengers of inflammation markers, miR-155 remains tightly associated with the pro-inflammatory transcriptional programmes. Consequently, we found that both miRNAs control distinct mRNA target profiles; although miR-146 targets the messengers of LPS signal transduction components and thus downregulates cellular LPS sensitivity, miR-155 targets the mRNAs of genes pervasively involved in pro-inflammatory transcriptional programmes. Thus, miR-155 acts as a broad limiter of pro-inflammatory gene expression once the miR-146 dependent barrier to LPS triggered inflammation has been breached. Importantly, we also report alternative miR-155 activation by the sensing of bacterial peptidoglycan through cytoplasmic NOD-like receptor, NOD2. We predict that dosedependent responses to environmental stimuli may involve functional specialization of seemingly coinduced miRNAs in other cellular circuitries as well.},
  language  = {en}
}
@article{MorschhaeuserRamirezZavalaWeyleretal.2013,
  author    = {Morschh{\"a}user, Joachim and Ram{\´i}rez-Zavala, Bernardo and Weyler, Michael and Gildor, Tsvia and Schmauch, Christian and Kornitzer, Daniel and Arkowitz, Robert},
  title     = {Activation of the Cph1-Dependent MAP Kinase Signaling Pathway Induces White-Opaque Switching in Candida albicans},
  series = {PLoS Pathogens},
  journal   = {PLoS Pathogens},
  doi       = {10.1371/journal.ppat.1003696},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-97281},
  year      = {2013},
  abstract  = {Depending on the environmental conditions, the pathogenic yeast Candida albicans can undergo different developmental programs, which are controlled by dedicated transcription factors and upstream signaling pathways. C. albicans strains that are homozygous at the mating type locus can switch from the normal yeast form (white) to an elongated cell type (opaque), which is the mating-competent form of this fungus. Both white and opaque cells use the Ste11-Hst7-Cek1/Cek2 MAP kinase signaling pathway to react to the presence of mating pheromone. However, while opaque cells employ the transcription factor Cph1 to induce the mating response, white cells recruit a different downstream transcription factor, Tec1, to promote the formation of a biofilm that facilitates mating of opaque cells in the population. The switch from the white to the opaque cell form is itself induced by environmental signals that result in the upregulation of the transcription factor Wor1, the master regulator of white-opaque switching. To get insight into the upstream signaling pathways controlling the switch, we expressed all C. albicans protein kinases from a tetracycline-inducible promoter in a switching-competent strain. Screening of this library of strains showed that a hyperactive form of Ste11 lacking its N-terminal domain (Ste11ΔN467) efficiently stimulated white cells to switch to the opaque phase, a behavior that did not occur in response to pheromone. Ste11ΔN467-induced switching specifically required the downstream MAP kinase Cek1 and its target transcription factor Cph1, but not Cek2 and Tec1, and forced expression of Cph1 also promoted white-opaque switching in a Wor1-dependent manner. Therefore, depending on the activation mechanism, components of the pheromone-responsive MAP kinase pathway can be reconnected to stimulate an alternative developmental program, switching of white cells to the mating-competent opaque phase.},
  language  = {en}
}
@article{SharmaDugarHerbigetal.2013,
  author    = {Sharma, Cynthia M. and Dugar, Gaurav and Herbig, Alexander and F{\"o}rstner, Konrad U. and Heidrich, Nadja and Reinhardt, Richard and Nieselt, Kay},
  title     = {High-Resolution Transcriptome Maps Reveal Strain-Specific Regulatory Features of Multiple Campylobacter jejuni Isolates},
  series = {PLoS Genetics},
  journal   = {PLoS Genetics},
  doi       = {10.1371/journal.pgen.1003495},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-96610},
  year      = {2013},
  abstract  = {Campylobacter jejuni is currently the leading cause of bacterial gastroenteritis in humans. Comparison of multiple Campylobacter strains revealed a high genetic and phenotypic diversity. However, little is known about differences in transcriptome organization, gene expression, and small RNA (sRNA) repertoires. Here we present the first comparative primary transcriptome analysis based on the differential RNA-seq (dRNA-seq) of four C. jejuni isolates. Our approach includes a novel, generic method for the automated annotation of transcriptional start sites (TSS), which allowed us to provide genome-wide promoter maps in the analyzed strains. These global TSS maps are refined through the integration of a SuperGenome approach that allows for a comparative TSS annotation by mapping RNA-seq data of multiple strains into a common coordinate system derived from a whole-genome alignment. Considering the steadily increasing amount of RNA-seq studies, our automated TSS annotation will not only facilitate transcriptome annotation for a wider range of pro- and eukaryotes but can also be adapted for the analysis among different growth or stress conditions. Our comparative dRNA-seq analysis revealed conservation of most TSS, but also single-nucleotide-polymorphisms (SNP) in promoter regions, which lead to strain-specific transcriptional output. Furthermore, we identified strain-specific sRNA repertoires that could contribute to differential gene regulation among strains. In addition, we identified a novel minimal CRISPR-system in Campylobacter of the type-II CRISPR subtype, which relies on the host factor RNase III and a trans-encoded sRNA for maturation of crRNAs. This minimal system of Campylobacter, which seems active in only some strains, employs a unique maturation pathway, since the crRNAs are transcribed from individual promoters in the upstream repeats and thereby minimize the requirements for the maturation machinery. Overall, our study provides new insights into strain-specific transcriptome organization and sRNAs, and reveals genes that could modulate phenotypic variation among strains despite high conservation at the DNA level.},
  language  = {en}
}
@phdthesis{Schillig2013,
  author    = {Schillig, Rebecca},
  title     = {Funktionelle Analyse der Zink-Cluster-Transkriptionsfaktorfamilie von Candida albicans durch artifizielle Aktivierung},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-79608},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2013},
  abstract  = {Der Hefepilz Candida albicans geh{\"o}rt zu den opportunistischen Infektionserregern. Er ist Teil der nat{\"u}rlichen Mikroflora der Schleimh{\"a}ute des Gastrointestinal- und Urogenitaltraktes des Menschen. Bei St{\"o}rungen des nat{\"u}rlichen Gleichgewichts dieser Flora kann es zu oberfl{\"a}chlichen Mykosen, z. B. der oropharyngealen Candidiasis (Mundsoor), kommen. Besonders immunsupprimierte Patienten, wie AIDS-Patienten, leiden h{\"a}ufig unter immer wiederkehrenden Infektionen, die mitunter auch zu schwerwiegenden Infektionsverl{\"a}ufen, bis hin zu lebensbedrohlichen systemischen Mykosen f{\"u}hren k{\"o}nnen. Zur Therapie solcher Erkrankungen werden oft Ergosterolbiosyntheseinhibitoren, wie Fluconazol, eingesetzt. Besonders bei wiederkehrenden Infektionen und wiederholender Therapie ist C. albicans in der Lage, gegen diese h{\"a}ufig verabreichten Antimykotika Resistenzen zu entwickeln. Hierbei spielen Zink-Cluster-Transkriptionsfaktoren eine zentrale Rolle. Zink-Cluster-Proteine geh{\"o}ren zu einer pilzspezifischen Familie von Transkriptionsfaktoren, die ein großes Spektrum an zellul{\"a}ren Prozessen regulieren. Die gut charakterisierten Regulatoren Upc2, Tac1 und Mrr1 geh{\"o}ren zu den Zink-Cluster-Transkriptionsfaktoren, die maßgeblich zur Resistenzentwicklung von C. albicans beitragen. Upc2 kontrolliert die Expression vieler Ergosterolbiosynthesegene, besonders die von ERG11, welches f{\"u}r die Zielstruktur des g{\"a}ngigen Antimykotikums Fluconazol kodiert. Tac1 und Mrr1 hingegen regulieren die Expression von Multidrug-Effluxpumpen, den ABC-Transportern CDR1 und CDR2 bzw. dem Major Facilitator MDR1. Gain-of-function-Mutationen in diesen Transkriptionsfaktoren resultieren in einer konstitutiven {\"U}berexpression ihrer Zielgene und sind verantwortlich f{\"u}r die Resistenz vieler klinischer Isolate. In dieser Arbeit wurde gezeigt, dass die Fusion von Mrr1 mit der Gal4-Aktivierungsdom{\"a}ne von Saccharomyces cerevisiae zu einem konstitutiv aktiven Hybridtranskriptionsfaktor f{\"u}hrte, der eine MDR1-{\"U}berexpression bewirkte und Fluconazolresistenz vermittelte. Dieses Hybridprotein vermittelte sogar eine h{\"o}here Resistenz als ein Mrr1 mit nat{\"u}rlich vorkommenden gain-of-function-Mutationen. Analoge Fusionen mit Tac1 und Upc2 resultierten ebenfalls in einer konstitutiven Aktivierung dieser Transkriptionsfaktoren, die einen starken Anstieg der Fluconazolresistenz zur Folge hatte. Daraus ergab sich die Schlussfolgerung, dass dies eine generelle Methode sein k{\"o}nnte, die Zink-Cluster-Transkriptionsfaktoren k{\"u}nstlich zu aktivieren und so ihre biologischen Funktionen zu offenbaren, ohne die genauen Bedingungen f{\"u}r ihre Aktivit{\"a}t zu kennen. Deshalb wurde auf der Basis dieser Strategie eine Bibliothek von C.-albicans-St{\"a}mmen konstruiert, in der alle 82 putativen Zink-Cluster-Transkriptionsfaktoren in dieser m{\"o}glicherweise hyperaktiven Form exprimiert werden. Untersuchungen dieser Bibliothek offenbarten neue Transkriptionsfaktoren, die Fluconazolresistenz vermittelten, aber auch noch unbekannte Regulatoren der Morphogenese und andere Ph{\"a}notypen konnten beobachtet werden. Um einen tieferen Einblick in die Funktionsweise zu bekommen, wurden die Transkriptionsprofile der vier Transkriptionsfaktoren ermittelt, die in ihrer hyperaktiven Form die h{\"o}chste Fluconazolresistenz bewirkten. Dabei stellte sich heraus, dass die zwei k{\"u}nstlich aktivierten (*) Regulatoren ZCF34* und ZNC1* die Expression der wichtigsten Multidrug-Effluxpumpe CDR1 stark hochregulierten. Der Transkriptionsfaktor mit dem vorl{\"a}ufigen Namen ZCF34 konnte im Verlauf dieser Arbeit als ein wichtiger Regulator f{\"u}r die CDR1-Expression identifiziert werden. Er ist sowohl an der Aktivierung der Expression von CDR1 beteiligt als auch f{\"u}r die basale CDR1-Promotoraktivit{\"a}t notwendig. Aus diesem Grund wurde er in MRR2 (multidrug resistance regulator 2) umbenannt. Mit der Entdeckung eines neuen Regulators der wichtigsten Multidrug-Effluxpumpe von C. albicans wurde ein wichtiger Beitrag zum Verst{\"a}ndnis der Regulation solcher Transporter geleistet. Die {\"U}berexpression dieser Pumpen ist einer der h{\"a}ufigsten Resistenzmechanismen in C. albicans. Auf diesem Wege kann Resistenz gegen strukturell v{\"o}llig unterschiedliche Antimykotika bewirkt werden. Somit stellen sowohl diese Effluxpumpen, als auch deren Regulatoren m{\"o}gliche Angriffsziele f{\"u}r die Entwicklung neuer oder Weiterentwicklung bereits vorhandener Antimykotika dar.},
  subject      = {Candida albicans},
  language  = {de}
}
@phdthesis{Ngwa2013,
  author    = {Ngwa, Che Julius},
  title     = {The mosquito midgut-specific stages of the malaria parasite as targets for transmission blocking interventions},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-83594},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2013},
  abstract  = {Die Tropenkrankheit Malaria, wird durch eine Infektion mit einzelligen Parasiten der Gattung Plasmodium verursacht und durch den Stich der weiblichen Anopheles-M{\"u}cke von Mensch zu Mensch verbreitet. Dabei kann eine erfolgreiche {\"U}bertragung des Parasiten auf den Menschen nur dann stattfinden, wenn der Parasit seine sexuelle Entwicklungsphase im Mitteldarm der M{\"u}cke erfolgreich durchl{\"a}uft. Ziel dieser Arbeit war es daher, die Wechselwirkungen des Malariaparasiten im Mitteldarm der M{\"u}cke in Hinblick auf die Identifizierung m{\"o}glicher neuer transmissionsblockierender Strategien zu untersuchen. Der Zweck von transmissionsblockierende Strategien ist es, der Verbreitung der Malaria durch die M{\"u}cke entgegenzuwirken, indem die Entwicklung des Parasiten in der M{\"u}cke unterbunden und dadurch der Lebenszyklus des Parasiten unterbrochen wird. Der Schwerpunkt der vorliegenden Arbeit lag auf insgesamt drei Aspekten. Der erste Aspekt der Arbeit befasste sich mit der Wechselwirkung zwischen dem Para-siten und der mikrobiellen Darmflora der M{\"u}cke. Dabei sollte der m{\"o}gliche Einfluss des Parasiten auf die Darmflora untersucht werden und weiterf{\"u}hrend die potentielle Verwendung von Darmbakterien als Vehikel f{\"u}r die Herstellung paratransgener M{\"u}cken erforscht werden. Vergleichende16S-rRNA- und DGGE-Analysen an der Darmflora des asiatischen Malariavektors Anopheles stephensi zeigten eine deutliche Reduktion der mikrobiellen Diversit{\"a}t w{\"a}hrend der Entwicklung vom Ei zur adulten M{\"u}cke. Zudem konnte das gram-negative Bakterium Elizabethkingia meningoseptica, das sich stadien- und generations{\"u}bergreifend verbreitet, als dominante Darmspezies bei im Labor aufgezogenen weiblichen und m{\"a}nnlichen An. stephensi festgestellt werden. Die Dominanz von E. meningoseptica wurde zudem nicht durch die Aufnahme von infiziertem Blut oder einer ver{\"a}nderten Nahrung beeinflusst. F{\"u}r die Studien wurde sowohl der humanpathogene Parasit P. falciparum als auch der Nagermalariaerreger P. berghei verwendet. Weiterf{\"u}hrende Versuche zeigten, dass Extrakte von E. meningoseptica antibakterielle, antifungale und antiplasmodiale Aktivit{\"a}ten aufwiesen, die ein m{\"o}glicher Grund f{\"u}r die Dominanz dieser Spezies im Mitteldarm des Vektors waren. Isolate von E. meningoseptica sind im Labor kultivierbar; dadurch stellt das Bakterium einen potentiellen Kandidaten zur Generierung von paratransgenen Anopheles-M{\"u}cken dar. Ein zweites Ziel dieser Arbeit war es, m{\"o}gliche Unterschiede in der Genexpression von P. falciparum darzustellen, die in den ersten 30 Minuten nach dessen {\"U}bertragung auf die M{\"u}cke erfolgen. Dies hatte zum einen zum Zweck, die durch den Wirtswechsel hervorgerufenen Genregulationen besser zu verstehen, und bot zum anderen die M{\"o}glichkeit, neue Proteine zu identifizieren, die als potentielle transmissionsblockierende Ziele genutzt werden k{\"o}nnen. Mittels supression substractive hybridization (SSH) konnten insgesamt 126 Gene identifiziert werden, deren Expression sich w{\"a}hrend der Gametogenese ver{\"a}ndert. Die identifizierten Gene konnten einer Vielzahl von putativen Funktionen wie zum Beispiel in der Signaltransduktion (17,5\%), im Zellzyklus (14,3\%) oder im Zytoskelett (8,7\%) zugeordnet werden. Des Weiteren wurden 7,9\% der Gene eine Funktion in der Proteastase und 6,4\% in metabolischen Prozessen zugeordnet. 12,7\% der Gene kodierten f{\"u}r zelloberfl{\"a}chenassoziierte Proteine. 11,9\% der Gene hatten anderen Funktionen, w{\"a}hrend 20\% der Gene keine putative Funktion zugeordnet werden konnte. Etwa 40\% der identifizierten Genprodukte waren bisher nicht in Proteomstudien nachgewiesen worden. In weiterf{\"u}hrenden Analysen wurden 34 Gene aus jeder ontologischen Gruppe ausgew{\"a}hlt und deren Expressionsver{\"a}nderung per quantitativer real time RT-PCR im Detail untersucht. F{\"u}r 29 Gene konnte dabei eine Transkriptexpression in Gametozyten nachgewiesen werden. Zudem wiesen 20 Gene eine erh{\"o}hte Expression in Gametozyten im Vergleich asexuellen Stadien auf. Insgesamt zeigten 8 Gene besonders hohe Transkriptlevel in aktivierten Gametozyten, was auf eine Funktion dieser Proteine w{\"a}hrend der {\"U}bertragung des Parasiten auf die M{\"u}cke hindeutet und diese somit potentielle Angriffspunkte f{\"u}r transmissionsblockierende Strategien darstellen k{\"o}nnten. Im letzten Teil dieser Arbeit stand die Untersuchung verschiedener antimikrobieller Substanzen in Bezug auf ihre transmissionsblockierenden Eigenschaften im Vordergrund. Die Substanzen waren entweder direkt aus der H{\"a}molymphe verschiedener Insekten isoliert oder rekombinant in transgenem Tabak exprimiert worden. Dabei wurden die rekombinanten Peptide so ausgew{\"a}hlt, dass sie entweder gegen die Mitteldarmstadien des Parasiten wirken oder m{\"u}ckenspezifische Rezeptoren blockieren, die der Parasit f{\"u}r seine weitere Entwicklung ben{\"o}tigt. Dabei konnte gezeigt werden, dass das antimikrobielle Molek{\"u}l Harmonin, ein Abwehrmolek{\"u}l aus der H{\"a}molymphe des asiatischen Marienk{\"a}fers Harmonia axyridis, antiplasmodiale als auch transmissions-blockierende Eigenschaften besitzt. Harmonin stellt daher eine potentielle Leitstruktur f{\"u}r die Entwicklung neuer Malariawirkstoffe dar},
  subject      = {Malariam{\"u}cke},
  language  = {en}
}
@phdthesis{Dunkel2013,
  author    = {Dunkel, Nico},
  title     = {Regulation of virulence-associated traits of the human fungal pathogen Candida albicans by nitrogen availability},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-83076},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2013},
  abstract  = {Nitrogen-regulated pathogenesis describes the expression of virulence attributes as direct response to the quantity and quality of an available nitrogen source. As consequence of nitrogen availability, the opportunistic human fungal pathogen Candida albicans changes its morphology and secretes aspartic proteases [SAPs], both well characterized virulence attributes. C. albicans, contrarily to its normally non-pathogenic relative Saccharomyces cerevisiae, is able to utilize proteins, which are considered as abundant and important nitrogen source within the human host. To assimilate complex proteinaceous matter, extracellular proteolysis is followed by uptake of the degradation products through dedicated peptide transporters (di-/tripeptide transporters [PTRs] and oligopeptide transporters [OPTs]). The expression of both traits is transcriptionally controlled by Stp1 - the global regulator of protein utilization - in C. albicans. The aim of the present study was to elucidate the regulation of virulence attributes of the pathogenic fungus C. albicans by nitrogen availability in more detail. Within a genome wide binding profile of Stp1, during growth with proteins, more than 600 Stp1 target genes were identified, thereby confirming its role in the usage of proteins, but also other nitrogenous compounds as nitrogen source. Moreover, the revealed targets suggest an involvement of Stp1 in the general adaption to nutrient availability as well as in the environmental stress response. With the focus on protein utilization and nitrogen-regulated pathogenesis, the regulation of the major secreted aspartic protease Sap2 - additionally one of the prime examples of allelic heterogeneity in C. albicans - was investigated in detail. Thereby, the heterogezygous SAP2 promoter helped to identify an unintended genomic alteration as the true cause of a growth defect of a C. albicans mutant. Additionally, the promoter region, which was responsible for the differential activation of the SAP2 alleles, was delimited. Furthermore, general Sap2 induction was demonstrated to be mediated by distinct cis-acting elements that are required for a high or a low activity of SAP2 expression. For the utilization of proteins as nitrogen source it is also crucial to take up the peptides that are produced by extracellular proteolysis. Therefore, the function and importance of specific peptide transporters was investigated in C. albicans mutants, unable to use peptides as nitrogen source (opt1Δ/Δ opt2Δ/Δ opt3Δ/Δ opt4Δ/Δ opt5Δ/Δ ptr2Δ/Δ ptr22Δ/Δ septuple null mutants). The overexpression of individual transporters in these mutants revealed differential substrate specificities and expanded the specificity of the OPTs to dipeptides, a completely new facet of these transporters. The peptide-uptake deficient mutants were further used to elucidate, whether indeed proteins and peptides are an important in vivo nitrogen source for C. albicans. It was found that during competitive colonization of the mouse intestine these mutants exhibited wild-type fitness, indicating that neither proteins nor peptides are primary nitrogen sources required to efficiently support growth of C. albicans in the mouse gut. Adequate availability of the preferred nitrogen source ammonium represses the utilization of proteins and other alternative nitrogen sources, but also the expression of virulence attributes, like Sap secretion and nitrogen-starvation induced filamentation. In order to discriminate, whether ammonium availability is externally sensed or determined inside the cell by C. albicans, the response to exterior ammonium concentrations of ammonium-uptake deficient mutants (mep1Δ/Δ mep2Δ/Δ null mutants) was investigated. This study showed that presence of an otherwise suppressing ammonium concentration did not inhibit Sap2 proteases secretion and arginine-induced filamentation in these mutants. Conclusively, ammonium availability is primarily determined inside the cell in order to control the expression of virulence traits. In sum, the present work contributes to the current understanding of how C. albicans regulates expression of virulence-associated traits in response to the presence of available nitrogen sources - especially proteins and peptides - in order to adapt its lifestyle within a human host.},
  subject      = {Candida albicans},
  language  = {en}
}
@article{SzalayWeibelHofmannetal.2013,
  author    = {Szalay, Aladar A and Weibel, Stephanie and Hofmann, Elisabeth and Basse-Luesebrink, Thomas Christian and Donat, Ulrike and Seubert, Carolin and Adelfinger, Marion and Gnamlin, Prisca and Kober, Christina and Frentzen, Alexa and Gentschev, Ivaylo and Jakob, Peter Michael},
  title     = {Treatment of malignant effusion by oncolytic virotherapy in an experimental subcutaneous xenograft model of lung cancer},
  series = {Journal of Translational Medicine},
  journal   = {Journal of Translational Medicine},
  doi       = {doi:10.1186/1479-5876-11-106},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-96016},
  year      = {2013},
  abstract  = {Background Malignant pleural effusion (MPE) is associated with advanced stages of lung cancer and is mainly dependent on invasion of the pleura and expression of vascular endothelial growth factor (VEGF) by cancer cells. As MPE indicates an incurable disease with limited palliative treatment options and poor outcome, there is an urgent need for new and efficient treatment options. Methods In this study, we used subcutaneously generated PC14PE6 lung adenocarcinoma xenografts in athymic mice that developed subcutaneous malignant effusions (ME) which mimic pleural effusions of the orthotopic model. Using this approach monitoring of therapeutic intervention was facilitated by direct observation of subcutaneous ME formation without the need of sacrificing mice or special imaging equipment as in case of MPE. Further, we tested oncolytic virotherapy using Vaccinia virus as a novel treatment modality against ME in this subcutaneous PC14PE6 xenograft model of advanced lung adenocarcinoma. Results We demonstrated significant therapeutic efficacy of Vaccinia virus treatment of both advanced lung adenocarcinoma and tumor-associated ME. We attribute the efficacy to the virus-mediated reduction of tumor cell-derived VEGF levels in tumors, decreased invasion of tumor cells into the peritumoral tissue, and to viral infection of the blood vessel-invading tumor cells. Moreover, we showed that the use of oncolytic Vaccinia virus encoding for a single-chain antibody (scAb) against VEGF (GLAF-1) significantly enhanced mono-therapy of oncolytic treatment. Conclusions Here, we demonstrate for the first time that oncolytic virotherapy using tumor-specific Vaccinia virus represents a novel and promising treatment modality for therapy of ME associated with advanced lung cancer.},
  subject      = {Lungenkrebs},
  language  = {en}
}