@phdthesis{Riedel2007, author = {Riedel, Alexander}, title = {Untersuchungen zur endogenen MHC-Klasse-II-restringierten Pr{\"a}sentation nukle{\"a}rer Antigene}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-25183}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2007}, abstract = {Die endogene Pr{\"a}sentation von intrazellul{\"a}ren Antigenen auf Major-Histokompatibilit{\"a}tskomplex Klasse-II (MHC-II) -Molek{\"u}len ist von entscheidender Bedeutung f{\"u}r eine Reihe von immunologischen Prozessen. Die mechanistischen Grundlagen dieses Pr{\"a}sentationsweges sind aber noch weitgehend unverstanden. Ziel dieser Arbeit war es, einen Beitrag zum molekularen Verst{\"a}ndnis der Abl{\"a}ufe zu leisten, die an der endogenen Pr{\"a}sentation nukle{\"a}rer Antigene auf MHC-II-Molek{\"u}len beteiligt sind. Dazu sollte am Beispiel des nukle{\"a}r lokalisierten Modellantigens Neomycin-Phosphotransferase II (NucNeoR) sowie des viralen Kernantigens Epstein-Barr-virus nuclear antigen 3C (EBNA3C) und entsprechender antigenspezifischer MHC-II-restringierter CD4+ T-Zellen die verantwortlichen Pr{\"a}sentationswege in professionell und nicht-professionell antigenpr{\"a}sentierenden Zellen untersucht werden. In beiden Zellsystemen wurde NucNeoR {\"u}ber einen endogenen Pr{\"a}sentationsweg und nicht {\"u}ber die Freisetzung und Wiederaufnahme als exogenes Protein auf MHC-II-Molek{\"u}len pr{\"a}sentiert. Durch die Verwendung chemischer Inhibitoren konnte eine Beteiligung der Autophagie an der endogenen Antigenpr{\"a}sentation nachgewiesen werden. Da Autophagie ausschließlich im Zytoplasma stattfindet, wurde nach m{\"o}glichen Eintrittspforten f{\"u}r nukle{\"a}re Proteine in diesen Abbauweg gesucht. F{\"u}r die Autophagie-abh{\"a}ngige Pr{\"a}sentation von NucNeoR war weder ein CRM1-vermittelter aktiver Export des Antigens aus dem Kern ins Zytoplasma, noch eine Aufl{\"o}sung der Kernmembran im Rahmen der Zellteilung und der dadurch bedingten Durchmischung nukle{\"a}rer und zytoplasmatischer Bestandteile notwendig. Mit Hilfe eines konditionalen Antigenexpressionsystems und der Auftrennung antigenexprimierender Zellen nach Zellzyklusphasen konnte eine verst{\"a}rkte Antigenpr{\"a}sentation in der G1/0-Phase nachgewiesen werden, die mit fortschreitendem Zellzyklus immer mehr abnahm. Die Antigenpr{\"a}sentation korrelierte dabei mit der ebenfalls im Laufe des Zellzyklus abnehmenden Transkriptions- bzw. Translationsrate des Antigens, aber nicht mit der absoluten Menge an Antigen in den Zellen. Bei abgeschalteter Antigentranskription dagegen korrelierte die Antigenpr{\"a}sentation mit der MHC-II-Oberfl{\"a}chenexpression, die von der G1/0- bis hin zur G2/M-Phase kontinuierlich zunahm. Eine {\"a}hnliche Korrelation von Antigentranskription/ Antigentranslation und Autophagie-abh{\"a}ngiger Antigenpr{\"a}sentation wurde auch f{\"u}r EBNA3C und die zytoplasmatisch lokalisierte NeoR-Variante beobachtet. Diese Ergebnisse identifizieren die Autophagie-abh{\"a}ngige Pr{\"a}sentation neusynthetisierter Proteine als den verantwortlichen molekularen Mechanismus f{\"u}r die endogene Pr{\"a}sentation der untersuchten nukle{\"a}ren Antigene auf MHC-II-Molek{\"u}len. Durch die Kopplung von Translation und autophagischem Abbau erlangen Proteine unabh{\"a}ngig von ihrer subzellul{\"a}ren Lokalisation Zugang zu diesem Pr{\"a}sentationsweg und erweitern so das Spektrum der intrazellul{\"a}ren Antigene, die einer CD4+ T-Zell{\"u}berwachung unterliegen.}, subject = {Autophagie}, language = {de} } @article{FraunholzSinha2012, author = {Fraunholz, Martin and Sinha, Bhanu}, title = {Intracellular staphylococcus aureus: Live-in and let die}, series = {Frontiers in Cellular and Infection Microbiology}, volume = {2}, journal = {Frontiers in Cellular and Infection Microbiology}, number = {43}, doi = {10.3389/fcimb.2012.00043}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-123374}, year = {2012}, abstract = {Staphylococcus aureus uses a plethora of virulence factors to accommodate a diversity of niches in its human host. Aside from the classical manifestations of S. aureus-induced diseases, the pathogen also invades and survives within mammalian host cells. The survival strategies of the pathogen are as diverse as strains or host cell types used. S. aureus is able to replicate in the phagosome or freely in the cytoplasm of its host cells. It escapes the phagosome of professional and non-professional phagocytes, subverts autophagy, induces cell death mechanisms such as apoptosis and pyronecrosis, and even can induce anti-apoptotic programs in phagocytes. The focus of this review is to present a guide to recent research outlining the variety of intracellular fates of S. aureus.}, language = {en} } @article{OttoHahlbrockEichetal.2016, author = {Otto, Christoph and Hahlbrock, Theresa and Eich, Kilian and Karaaslan, Ferdi and J{\"u}rgens, Constantin and Germer, Christoph-Thomas and Wiegering, Armin and K{\"a}mmerer, Ulrike}, title = {Antiproliferative and antimetabolic effects behind the anticancer property of fermented wheat germ extract}, series = {BMC Complementary and Alternative Medicine}, volume = {16}, journal = {BMC Complementary and Alternative Medicine}, number = {160}, doi = {10.1186/s12906-016-1138-5}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-146013}, year = {2016}, abstract = {Background Fermented wheat germ extract (FWGE) sold under the trade name Avemar exhibits anticancer activity in vitro and in vivo. Its mechanisms of action are divided into antiproliferative and antimetabolic effects. Its influcence on cancer cell metabolism needs further investigation. One objective of this study, therefore, was to further elucidate the antimetabolic action of FWGE. The anticancer compound 2,6-dimethoxy-1,4-benzoquinone (DMBQ) is the major bioactive compound in FWGE and is probably responsible for its anticancer activity. The second objective of this study was to compare the antiproliferative properties in vitro of FWGE and the DMBQ compound. Methods The IC\(_{50}\) values of FWGE were determined for nine human cancer cell lines after 24 h of culture. The DMBQ compound was used at a concentration of 24 μmol/l, which is equal to the molar concentration of DMBQ in FWGE. Cell viability, cell cycle, cellular redox state, glucose consumption, lactic acid production, cellular ATP levels, and the NADH/NAD\(^+\) ratio were measured. Results The mean IC\(_{50}\) value of FWGE for the nine human cancer cell lines tested was 10 mg/ml. Both FWGE (10 mg/ml) and the DMBQ compound (24 μmol/l) induced massive cell damage within 24 h after starting treatment, with changes in the cellular redox state secondary to formation of intracellular reactive oxygen species. Unlike the DMBQ compound, which was only cytotoxic, FWGE exhibited cytostatic and growth delay effects in addition to cytotoxicity. Both cytostatic and growth delay effects were linked to impaired glucose utilization which influenced the cell cycle, cellular ATP levels, and the NADH/NAD\(^+\) ratio. The growth delay effect in response to FWGE treatment led to induction of autophagy. Conclusions FWGE and the DMBQ compound both induced oxidative stress-promoted cytotoxicity. In addition, FWGE exhibited cytostatic and growth delay effects associated with impaired glucose utilization which led to autophagy, a possible previously unknown mechanism behind the influence of FWGE on cancer cell metabolism.}, language = {en} } @article{KunzKozjakPavlovic2019, author = {Kunz, Tobias C. and Kozjak-Pavlovic, Vera}, title = {Diverse facets of sphingolipid involvement in bacterial infections}, series = {Frontiers in Cell and Developmental Biology}, volume = {7}, journal = {Frontiers in Cell and Developmental Biology}, number = {203}, doi = {10.3389/fcell.2019.00203}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-201757}, year = {2019}, abstract = {Sphingolipids are constituents of the cell membrane that perform various tasks as structural elements and signaling molecules, in addition to regulating many important cellular processes, such as apoptosis and autophagy. In recent years, it has become increasingly clear that sphingolipids and sphingolipid signaling play a vital role in infection processes. In many cases the attachment and uptake of pathogenic bacteria, as well as bacterial development and survival within the host cell depend on sphingolipids. In addition, sphingolipids can serve as antimicrobials, inhibiting bacterial growth and formation of biofilms. This review will give an overview of our current information about these various aspects of sphingolipid involvement in bacterial infections.}, language = {en} } @article{StojanovićFuchsFiedleretal.2020, author = {Stojanović, Stevan D. and Fuchs, Maximilian and Fiedler, Jan and Xiao, Ke and Meinecke, Anna and Just, Annette and Pich, Andreas and Thum, Thomas and Kunz, Meik}, title = {Comprehensive bioinformatics identifies key microRNA players in ATG7-deficient lung fibroblasts}, series = {International Journal of Molecular Sciences}, volume = {21}, journal = {International Journal of Molecular Sciences}, number = {11}, issn = {1422-0067}, doi = {10.3390/ijms21114126}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-285181}, year = {2020}, abstract = {Background: Deficient autophagy has been recently implicated as a driver of pulmonary fibrosis, yet bioinformatics approaches to study this cellular process are lacking. Autophagy-related 5 and 7 (ATG5/ATG7) are critical elements of macro-autophagy. However, an alternative ATG5/ATG7-independent macro-autophagy pathway was recently discovered, its regulation being unknown. Using a bioinformatics proteome profiling analysis of ATG7-deficient human fibroblasts, we aimed to identify key microRNA (miR) regulators in autophagy. Method: We have generated ATG7-knockout MRC-5 fibroblasts and performed mass spectrometry to generate a large-scale proteomics dataset. We further quantified the interactions between various proteins combining bioinformatics molecular network reconstruction and functional enrichment analysis. The predicted key regulatory miRs were validated via quantitative polymerase chain reaction. Results: The functional enrichment analysis of the 26 deregulated proteins showed decreased cellular trafficking, increased mitophagy and senescence as the major overarching processes in ATG7-deficient lung fibroblasts. The 26 proteins reconstitute a protein interactome of 46 nodes and miR-regulated interactome of 834 nodes. The miR network shows three functional cluster modules around miR-16-5p, miR-17-5p and let-7a-5p related to multiple deregulated proteins. Confirming these results in a biological setting, serially passaged wild-type and autophagy-deficient fibroblasts displayed senescence-dependent expression profiles of miR-16-5p and miR-17-5p. Conclusions: We have developed a bioinformatics proteome profiling approach that successfully identifies biologically relevant miR regulators from a proteomics dataset of the ATG-7-deficient milieu in lung fibroblasts, and thus may be used to elucidate key molecular players in complex fibrotic pathological processes. The approach is not limited to a specific cell-type and disease, thus highlighting its high relevance in proteome and non-coding RNA research.}, language = {en} } @article{OelschlaegelWeissSadanSalpeteretal.2020, author = {Oelschlaegel, Diana and Weiss Sadan, Tommy and Salpeter, Seth and Krug, Sebastian and Blum, Galia and Schmitz, Werner and Schulze, Almut and Michl, Patrick}, title = {Cathepsin inhibition modulates metabolism and polarization of tumor-associated macrophages}, series = {Cancers}, volume = {12}, journal = {Cancers}, number = {9}, issn = {2072-6694}, doi = {10.3390/cancers12092579}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-213040}, year = {2020}, abstract = {Stroma-infiltrating immune cells, such as tumor-associated macrophages (TAM), play an important role in regulating tumor progression and chemoresistance. These effects are mostly conveyed by secreted mediators, among them several cathepsin proteases. In addition, increasing evidence suggests that stroma-infiltrating immune cells are able to induce profound metabolic changes within the tumor microenvironment. In this study, we aimed to characterize the impact of cathepsins in maintaining the TAM phenotype in more detail. For this purpose, we investigated the molecular effects of pharmacological cathepsin inhibition on the viability and polarization of human primary macrophages as well as its metabolic consequences. Pharmacological inhibition of cathepsins B, L, and S using a novel inhibitor, GB111-NH\(_2\), led to changes in cellular recycling processes characterized by an increased expression of autophagy- and lysosome-associated marker genes and reduced adenosine triphosphate (ATP) content. Decreased cathepsin activity in primary macrophages further led to distinct changes in fatty acid metabolites associated with increased expression of key modulators of fatty acid metabolism, such as fatty acid synthase (FASN) and acid ceramidase (ASAH1). The altered fatty acid profile was associated with an increased synthesis of the pro-inflammatory prostaglandin PGE\(_2\), which correlated with the upregulation of numerous NF\(_k\)B-dependent pro-inflammatory mediators, including interleukin-1 (IL-1), interleukin-6 (IL-6), C-C motif chemokine ligand 2 (CCL2), and tumor necrosis factor-alpha (TNFα). Our data indicate a novel link between cathepsin activity and metabolic reprogramming in macrophages, demonstrated by a profound impact on autophagy and fatty acid metabolism, which facilitates a pro-inflammatory micromilieu generally associated with enhanced tumor elimination. These results provide a strong rationale for therapeutic cathepsin inhibition to overcome the tumor-promoting effects of the immune-evasive tumor micromilieu.}, language = {en} } @article{YangHeydarianKozjakPavlovicetal.2020, author = {Yang, Tao and Heydarian, Motaharehsadat and Kozjak-Pavlovic, Vera and Urban, Manuela and Harbottle, Richard P. and Rudel, Thomas}, title = {Folliculin Controls the Intracellular Survival and Trans-Epithelial Passage of Neisseria gonorrhoeae}, series = {Frontiers in Cellular and Infection Microbiology}, volume = {10}, journal = {Frontiers in Cellular and Infection Microbiology}, number = {422}, issn = {2235-2988}, doi = {10.3389/fcimb.2020.00422}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-211372}, year = {2020}, abstract = {Neisseria gonorrhoeae, a Gram-negative obligate human pathogenic bacterium, infects human epithelial cells and causes sexually transmitted diseases. Emerging multi-antibiotic resistant gonococci and increasing numbers of infections complicate the treatment of infected patients. Here, we used an shRNA library screen and next-generation sequencing to identify factors involved in epithelial cell infection. Folliculin (FLCN), a 64 kDa protein with a tumor repressor function was identified as a novel host factor important for N. gonorrhoeae survival after uptake. We further determined that FLCN did not affect N. gonorrhoeae adherence and invasion but was essential for its survival in the cells by modulating autophagy. In addition, FLCN was also required to maintain cell to cell contacts in the epithelial layer. In an infection model with polarized cells, FLCN inhibited the polarized localization of E-cadherin and the transcytosis of gonococci across polarized epithelial cells. In conclusion, we demonstrate here the connection between FLCN and bacterial infection and in particular the role of FLCN in the intracellular survival and transcytosis of gonococci across polarized epithelial cell layers.}, language = {en} } @article{AuerHuegelschaefferFischeretal.2020, author = {Auer, Daniela and H{\"u}gelsch{\"a}ffer, Sophie D. and Fischer, Annette B. and Rudel, Thomas}, title = {The chlamydial deubiquitinase Cdu1 supports recruitment of Golgi vesicles to the inclusion}, series = {Cellular Microbiology}, volume = {22}, journal = {Cellular Microbiology}, number = {5}, doi = {10.1111/cmi.13136}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-208675}, pages = {e13136}, year = {2020}, abstract = {Chlamydia trachomatis is the main cause of sexually transmitted diseases worldwide. As obligate intracellular bacteria Chlamydia replicate in a membrane bound vacuole called inclusion and acquire nutrients for growth and replication from their host cells. However, like all intracellular bacteria, Chlamydia have to prevent eradication by the host's cell autonomous system. The chlamydial deubiquitinase Cdu1 is secreted into the inclusion membrane, facing the host cell cytosol where it deubiquitinates cellular proteins. Here we show that inactivation of Cdu1 causes a growth defect of C. trachomatis in primary cells. Moreover, ubiquitin and several autophagy receptors are recruited to the inclusion membrane of Cdu1-deficient Chlamydia . Interestingly, the growth defect of cdu1 mutants is not rescued when autophagy is prevented. We find reduced recruitment of Golgi vesicles to the inclusion of Cdu1 mutants indicating that vesicular trafficking is altered in bacteria without active deubiquitinase (DUB). Our work elucidates an important role of Cdu1 in the functional preservation of the chlamydial inclusion surface.}, language = {en} } @article{MainzSarhanRothetal.2022, author = {Mainz, Laura and Sarhan, Mohamed A. F. E. and Roth, Sabine and Sauer, Ursula and Maurus, Katja and Hartmann, Elena M. and Seibert, Helen-Desiree and Rosenwald, Andreas and Diefenbacher, Markus E. and Rosenfeldt, Mathias T.}, title = {Autophagy blockage reduces the incidence of pancreatic ductal adenocarcinoma in the context of mutant Trp53}, series = {Frontiers in Cell and Developmental Biology}, volume = {10}, journal = {Frontiers in Cell and Developmental Biology}, issn = {2296-634X}, doi = {10.3389/fcell.2022.785252}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-266005}, year = {2022}, abstract = {Macroautophagy (hereafter referred to as autophagy) is a homeostatic process that preserves cellular integrity. In mice, autophagy regulates pancreatic ductal adenocarcinoma (PDAC) development in a manner dependent on the status of the tumor suppressor gene Trp53. Studies published so far have investigated the impact of autophagy blockage in tumors arising from Trp53-hemizygous or -homozygous tissue. In contrast, in human PDACs the tumor suppressor gene TP53 is mutated rather than allelically lost, and TP53 mutants retain pathobiological functions that differ from complete allelic loss. In order to better represent the patient situation, we have investigated PDAC development in a well-characterized genetically engineered mouse model (GEMM) of PDAC with mutant Trp53 (Trp53\(^{R172H}\)) and deletion of the essential autophagy gene Atg7. Autophagy blockage reduced PDAC incidence but had no impact on survival time in the subset of animals that formed a tumor. In the absence of Atg7, non-tumor-bearing mice reached a similar age as animals with malignant disease. However, the architecture of autophagy-deficient, tumor-free pancreata was effaced, normal acinar tissue was largely replaced with low-grade pancreatic intraepithelial neoplasias (PanINs) and insulin expressing islet β-cells were reduced. Our data add further complexity to the interplay between Atg7 inhibition and Trp53 status in tumorigenesis.}, language = {en} }