TY - JOUR A1 - Wu, Yu A1 - Pons, Valérie A1 - Goudet, Amélie A1 - Panigai, Laetitia A1 - Fischer, Annette A1 - Herweg, Jo-Ana A1 - Kali, Sabrina A1 - Davey, Robert A. A1 - Laporte, Jérôme A1 - Bouclier, Céline A1 - Yousfi, Rahima A1 - Aubenque, Céline A1 - Merer, Goulven A1 - Gobbo, Emilie A1 - Lopez, Roman A1 - Gillet, Cynthia A1 - Cojean, Sandrine A1 - Popoff, Michel R. A1 - Clayette, Pascal A1 - Le Grand, Roger A1 - Boulogne, Claire A1 - Tordo, Noël A1 - Lemichez, Emmanuel A1 - Loiseau, Philippe M. A1 - Rudel, Thomas A1 - Sauvaire, Didier A1 - Cintrat, Jean-Christophe A1 - Gillet, Daniel A1 - Barbier, Julien T1 - ABMA, a small molecule that inhibits intracellular toxins and pathogens by interfering with late endosomal compartments JF - Scientific Reports N2 - Intracellular pathogenic microorganisms and toxins exploit host cell mechanisms to enter, exert their deleterious effects as well as hijack host nutrition for their development. A potential approach to treat multiple pathogen infections and that should not induce drug resistance is the use of small molecules that target host components. We identifed the compound 1-adamantyl (5-bromo-2-methoxybenzyl) amine (ABMA) from a cell-based high throughput screening for its capacity to protect human cells and mice against ricin toxin without toxicity. This compound efciently protects cells against various toxins and pathogens including viruses, intracellular bacteria and parasite. ABMA provokes Rab7-positive late endosomal compartment accumulation in mammalian cells without affecting other organelles (early endosomes, lysosomes, the Golgi apparatus, the endoplasmic reticulum or the nucleus). As the mechanism of action of ABMA is restricted to host-endosomal compartments, it reduces cell infection by pathogens that depend on this pathway to invade cells. ABMA may represent a novel class of broad-spectrum compounds with therapeutic potential against diverse severe infectious diseases. KW - biology KW - antimicrobials KW - high-throughput screening KW - infectious diseases Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-173170 VL - 7 ER - TY - JOUR A1 - Blättner, Sebastian A1 - Das, Sudip A1 - Paprotka, Kerstin A1 - Eilers, Ursula A1 - Krischke, Markus A1 - Kretschmer, Dorothee A1 - Remmele, Christian W. A1 - Dittrich, Marcus A1 - Müller, Tobias A1 - Schuelein-Voelk, Christina A1 - Hertlein, Tobias A1 - Mueller, Martin J. A1 - Huettel, Bruno A1 - Reinhardt, Richard A1 - Ohlsen, Knut A1 - Rudel, Thomas A1 - Fraunholz, Martin J. T1 - Staphylococcus aureus Exploits a Non-ribosomal Cyclic Dipeptide to Modulate Survival within Epithelial Cells and Phagocytes JF - PLoS Pathogens N2 - Community-acquired (CA) Staphylococcus aureus cause various diseases even in healthy individuals. Enhanced virulence of CA-strains is partly attributed to increased production of toxins such as phenol-soluble modulins (PSM). The pathogen is internalized efficiently by mammalian host cells and intracellular S. aureus has recently been shown to contribute to disease. Upon internalization, cytotoxic S. aureus strains can disrupt phagosomal membranes and kill host cells in a PSM-dependent manner. However, PSM are not sufficient for these processes. Here we screened for factors required for intracellular S. aureus virulence. We infected escape reporter host cells with strains from an established transposon mutant library and detected phagosomal escape rates using automated microscopy. We thereby, among other factors, identified a non-ribosomal peptide synthetase (NRPS) to be required for efficient phagosomal escape and intracellular survival of S. aureus as well as induction of host cell death. By genetic complementation as well as supplementation with the synthetic NRPS product, the cyclic dipeptide phevalin, wild-type phenotypes were restored. We further demonstrate that the NRPS is contributing to virulence in a mouse pneumonia model. Together, our data illustrate a hitherto unrecognized function of the S. aureus NRPS and its dipeptide product during S. aureus infection. KW - cell death KW - cytotoxicity KW - Staphylococcus aureus KW - host cells KW - neutrophils KW - macrophages KW - transposable elements KW - epithelial cells Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-180380 VL - 12 IS - 9 ER - TY - JOUR A1 - Subbarayal, Prema A1 - Karunakaran, Karthika A1 - Winkler, Ann-Cathrin A1 - Rother, Marion A1 - Gonzalez, Erik A1 - Meyer, Thomas F. A1 - Rudel, Thomas T1 - EphrinA2 Receptor (EphA2) Is an Invasion and Intracellular Signaling Receptor for Chlamydia trachomatis JF - PLoS Pathogens N2 - The obligate intracellular bacterium Chlamydia trachomatis invades into host cells to replicate inside a membrane-bound vacuole called inclusion. Multiple different host proteins are recruited to the inclusion and are functionally modulated to support chlamydial development. Invaded and replicating Chlamydia induces a long-lasting activation of the PI3 kinase signaling pathway that is required for efficient replication. We identified the cell surface tyrosine kinase EphrinA2 receptor (EphA2) as a chlamydial adherence and invasion receptor that induces PI3 kinase (PI3K) activation, promoting chlamydial replication. Interfering with binding of C. trachomatis serovar L2 (Ctr) to EphA2, downregulation of EphA2 expression or inhibition of EphA2 activity significantly reduced Ctr infection. Ctr interacts with and activates EphA2 on the cell surface resulting in Ctr and receptor internalization. During chlamydial replication, EphA2 remains active accumulating around the inclusion and interacts with the p85 regulatory subunit of PI3K to support the activation of the PI3K/Akt signaling pathway that is required for normal chlamydial development. Overexpression of full length EphA2, but not the mutant form lacking the intracellular cytoplasmic domain, enhanced PI3K activation and Ctr infection. Despite the depletion of EphA2 from the cell surface, Ctr infection induces upregulation of EphA2 through the activation of the ERK pathway, which keeps the infected cell in an apoptosis-resistant state. The significance of EphA2 as an entry and intracellular signaling receptor was also observed with the urogenital C. trachomatis-serovar D. Our findings provide the first evidence for a host cell surface receptor that is exploited for invasion as well as for receptor-mediated intracellular signaling to facilitate chlamydial replication. In addition, the engagement of a cell surface receptor at the inclusion membrane is a new mechanism by which Chlamydia subverts the host cell and induces apoptosis resistance. KW - membrane proteins KW - chlamydia infection KW - chlamydia trachomatis KW - chlamydia KW - HeLa cells KW - apoptosis KW - host cells KW - membrane receptor signaling Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-125566 VL - 11 IS - 4 ER - TY - JOUR A1 - Herweg, Jo-Ana A1 - Hansmeier, Nicole A1 - Otto, Andreas A1 - Geffken, Anna C. A1 - Subbarayal, Prema A1 - Prusty, Bhupesh K. A1 - Becher, Dörte A1 - Hensel, Michael A1 - Schaible, Ulrich E. A1 - Rudel, Thomas A1 - Hilbi, Hubert T1 - Purification and proteomics of pathogen-modified vacuoles and membranes JF - Frontiers in Cellular and Infection Microbiology N2 - Certain pathogenic bacteria adopt an intracellular lifestyle and proliferate in eukaryotic host cells. The intracellular niche protects the bacteria from cellular and humoral components of the mammalian immune system, and at the same time, allows the bacteria to gain access to otherwise restricted nutrient sources. Yet, intracellular protection and access to nutrients comes with a price, i.e., the bacteria need to overcome cell-autonomous defense mechanisms, such as the bactericidal endocytic pathway. While a few bacteria rupture the early phagosome and escape into the host cytoplasm, most intracellular pathogens form a distinct, degradation-resistant and replication-permissive membranous compartment. Intracellular bacteria that form unique pathogen vacuoles include Legionella, Mycobacterium, Chlamydia, Simkania, and Salmonella species. In order to understand the formation of these pathogen niches on a global scale and in a comprehensive and quantitative manner, an inventory of compartment-associated host factors is required. To this end, the intact pathogen compartments need to be isolated, purified and biochemically characterized. Here, we review recent progress on the isolation and purification of pathogen-modified vacuoles and membranes, as well as their proteomic characterization by mass spectrometry and different validation approaches. These studies provide the basis for further investigations on the specific mechanisms of pathogen-driven compartment formation. KW - spectrometry-based proteomics KW - Mycobacterium tuberculosis KW - Chlamydia KW - Salmonella KW - bacterium Legionella pneumophila KW - endocytic multivesicular bodies KW - phagosome maturation arrest KW - III secretion system KW - endoplasmic reticulum KW - Chlamydia trachomatis KW - Simkania negevensis KW - intracellular bacteria KW - host pathogen interactions KW - immuno-magnetic purification KW - Legionella KW - Mycobacterium KW - Simkania KW - pathogen vacuole Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-151823 VL - 5 IS - 48 ER - TY - JOUR A1 - Siegl, Christine A1 - Prusty, Bhupesh K. A1 - Karunakaran, Karthika A1 - Wischhusen, Jörg A1 - Rudel, Thomas T1 - Tumor Suppressor p53 Alters Host Cell Metabolism to Limit Chlamydia trachomatis Infection JF - Cell Reports N2 - Obligate intracellular bacteria depend entirely on nutrients from the host cell for their reproduction. Here, we show that obligate intracellular Chlamydia downregulate the central tumor suppressor p53 in human cells. This reduction of p53 levels is mediated by the PI3K-Akt signaling pathway, activation of HDM2, and subsequent proteasomal degradation of p53. The stabilization of p53 in human cells severely impaired chlamydial development and caused the loss of infectious particle formation. DNA-damage-induced p53 interfered with chlamydial development through downregulation of the pentose phosphate pathway (PPP). Increased expression of the PPP key enzyme glucose-6-phosphate dehydrogenase rescued the inhibition of chlamydial growth induced by DNA damage or stabilized p53. Thus, downregulation of p53 is a key event in the chlamydial life cycle that reprograms the host cell to create a metabolic environment supportive of chlamydial growth. KW - chlamydia trachomatis KW - tumor Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-118200 SN - 2211-1247 VL - 9 IS - 3 ER - TY - JOUR A1 - Baur, Stefanie A1 - Rautenberg, Maren A1 - Faulstich, Manuela A1 - Grau, Timo A1 - Severin, Yannik A1 - Unger, Clemens A1 - Hoffmann, Wolfgang H. A1 - Rudel, Thomas A1 - Autenrieth, Ingo B. A1 - Weidenmaier, Christopher T1 - A Nasal Epithelial Receptor for Staphylococcus aureus WTA Governs Adhesion to Epithelial Cells and Modulates Nasal Colonization JF - PLOS PATHOGENS N2 - Nasal colonization is a major risk factor for S. aureus infections. The mechanisms responsible for colonization are still not well understood and involve several factors on the host and the bacterial side. One key factor is the cell wall teichoic acid (WTA) of S. aureus, which governs direct interactions with nasal epithelial surfaces. We report here the first receptor for the cell wall glycopolymer WTA on nasal epithelial cells. In several assay systems this type F-scavenger receptor, termed SREC-I, bound WTA in a charge dependent manner and mediated adhesion to nasal epithelial cells in vitro. The impact of WTA and SREC-I interaction on epithelial adhesion was especially pronounced under shear stress, which resembles the conditions found in the nasal cavity. Most importantly, we demonstrate here a key role of the WTA-receptor interaction in a cotton rat model of nasal colonization. When we inhibited WTA mediated adhesion with a SREC-I antibody, nasal colonization in the animal model was strongly reduced at the early onset of colonization. More importantly, colonization stayed low over an extended period of 6 days. Therefore we propose targeting of this glycopolymer-receptor interaction as a novel strategy to prevent or control S. aureus nasal colonization. KW - SREC-I KW - clumping factor-B KW - scavender receptor KW - teichoic acids KW - surface proteins KW - cotton rats KW - carriage KW - determinant KW - infections KW - expression Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-116280 SN - 1553-7374 VL - 10 IS - 5 ER - TY - JOUR A1 - Volceanov, Larisa A1 - Herbst, Katharina A1 - Biniossek, Martin A1 - Schilling, Oliver A1 - Haller, Dirk A1 - Nölke, Thilo A1 - Subbarayal, Prema A1 - Rudel, Thomas A1 - Zieger, Barbara A1 - Häcker, Georg T1 - Septins Arrange F-Actin-Containing Fibers on the Chlamydia trachomatis Inclusion and Are Required for Normal Release of the Inclusion by Extrusion JF - MBIO N2 - Chlamydia trachomatis is an obligate intracellular human pathogen that grows inside a membranous, cytosolic vacuole termed an inclusion. Septins are a group of 13 GTP-binding proteins that assemble into oligomeric complexes and that can form higher-order filaments. We report here that the septins SEPT2, -9, -11, and probably -7 form fibrillar structures around the chlamydial inclusion. Colocalization studies suggest that these septins combine with F actin into fibers that encase the inclusion. Targeting the expression of individual septins by RNA interference (RNAi) prevented the formation of septin fibers as well as the recruitment of actin to the inclusion. At the end of the developmental cycle of C. trachomatis, newly formed, infectious elementary bodies are released, and this release occurs at least in part through the organized extrusion of intact inclusions. RNAi against SEPT9 or against the combination of SEPT2/7/9 substantially reduced the number of extrusions from a culture of infected HeLa cells. The data suggest that a higher-order structure of four septins is involved in the recruitment or stabilization of the actin coat around the chlamydial inclusion and that this actin recruitment by septins is instrumental for the coordinated egress of C. trachomatis from human cells. The organization of F actin around parasite-containing vacuoles may be a broader response mechanism of mammalian cells to the infection by intracellular, vacuole-dwelling pathogens. IMPORTANCE Chlamydia trachomatis is a frequent bacterial pathogen throughout the world, causing mostly eye and genital infections. C. trachomatis can develop only inside host cells; it multiplies inside a membranous vacuole in the cytosol, termed an inclusion. The inclusion is covered by cytoskeletal "coats" or "cages," whose organization and function are poorly understood. We here report that a relatively little-characterized group of proteins, septins, is required to organize actin fibers on the inclusion and probably through actin the release of the inclusion. Septins are a group of GTP-binding proteins that can organize into heteromeric complexes and then into large filaments. Septins have previously been found to be involved in the interaction of the cell with bacteria in the cytosol. Our observation that they also organize a reaction to bacteria living in vacuoles suggests that they have a function in the recognition of foreign compartments by a parasitized human cell. KW - mammalian septins KW - host-cells KW - binding KW - proteins KW - organization KW - cytoskeleton KW - cytokinesis KW - mechanisms KW - expression KW - protease Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-115421 SN - 2150-7511 VL - 5 IS - 5 ER - TY - JOUR A1 - Remmele, Christian W. A1 - Xian, Yibo A1 - Albrecht, Marco A1 - Faulstich, Michaela A1 - Fraunholz, Martin A1 - Heinrichs, Elisabeth A1 - Dittrich, Marcus T. A1 - Müller, Tobias A1 - Reinhardt, Richard A1 - Rudel, Thomas T1 - Transcriptional landscape and essential genes of Neisseria gonorrhoeae N2 - The WHO has recently classified Neisseria gonorrhoeae as a super-bacterium due to the rapid spread of antibiotic resistant derivatives and an overall dramatic increase in infection incidences. Genome sequencing has identified potential genes, however, little is known about the transcriptional organization and the presence of non-coding RNAs in gonococci. We performed RNA sequencing to define the transcriptome and the transcriptional start sites of all gonococcal genes and operons. Numerous new transcripts including 253 potentially non-coding RNAs transcribed from intergenic regions or antisense to coding genes were identified. Strikingly, strong antisense transcription was detected for the phase-variable opa genes coding for a family of adhesins and invasins in pathogenic Neisseria, that may have regulatory functions. Based on the defined transcriptional start sites, promoter motifs were identified. We further generated and sequenced a high density Tn5 transposon library to predict a core of 827 gonococcal essential genes, 133 of which have no known function. Our combined RNA-Seq and Tn-Seq approach establishes a detailed map of gonococcal genes and defines the first core set of essential gonococcal genes. KW - Neisseria gonorrhoeae Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-113676 ER - TY - JOUR A1 - Rudel, Thomas A1 - Prusty, Bhupesh K. A1 - Siegl, Christine A1 - Gulve, Nitish A1 - Mori, Yasuko T1 - GP96 Interacts with HHV-6 during Viral Entry and Directs It for Cellular Degradation N2 - CD46 and CD134 mediate attachment of Human Herpesvirus 6A (HHV-6A) and HHV-6B to host cell, respectively. But many cell types interfere with viral infection through rapid degradation of viral DNA. Hence, not all cells expressing these receptors are permissive to HHV-6 DNA replication and production of infective virions suggesting the involvement of additional factors that influence HHV-6 propagation. Here, we used a proteomics approach to identify other host cell proteins necessary for HHV-6 binding and entry. We found host cell chaperone protein GP96 to interact with HHV-6A and HHV-6B and to interfere with virus propagation within the host cell. In human peripheral blood mononuclear cells (PBMCs), GP96 is transported to the cell surface upon infection with HHV-6 and interacts with HHV-6A and -6B through its C-terminal end. Suppression of GP96 expression decreased initial viral binding but increased viral DNA replication. Transient expression of human GP96 allowed HHV-6 entry into CHO-K1 cells even in the absence of CD46. Thus, our results suggest an important role for GP96 during HHV-6 infection, which possibly supports the cellular degradation of the virus. KW - host cells KW - immunoprecipitation KW - HeLa cells KW - antibodies KW - cell binding KW - viral transmission and infection KW - viral entry KW - flow cytometry Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-111068 ER - TY - JOUR A1 - Rudel, Thomas A1 - Krohne, George A1 - Prusty, Bhupesh K. T1 - Reactivation of Chromosomally Integrated Human Herpesvirus-6 by Telomeric Circle Formation N2 - More than 95% of the human population is infected with human herpesvirus-6 (HHV-6) during early childhood and maintains latent HHV-6 genomes either in an extra-chromosomal form or as a chromosomally integrated HHV-6 (ciHHV-6). In addition, approximately 1% of humans are born with an inheritable form of ciHHV-6 integrated into the telomeres of chromosomes. Immunosuppression and stress conditions can reactivate latent HHV-6 replication, which is associated with clinical complications and even death. We have previously shown that Chlamydia trachomatis infection reactivates ciHHV-6 and induces the formation of extra-chromosomal viral DNA in ciHHV-6 cells. Here, we propose a model and provide experimental evidence for the mechanism of ciHHV-6 reactivation. Infection with Chlamydia induced a transient shortening of telomeric ends, which subsequently led to increased telomeric circle (t-circle) formation and incomplete reconstitution of circular viral genomes containing single viral direct repeat (DR). Correspondingly, short t-circles containing parts of the HHV-6 DR were detected in cells from individuals with genetically inherited ciHHV-6. Furthermore, telomere shortening induced in the absence of Chlamydia infection also caused circularization of ciHHV-6, supporting a t-circle based mechanism for ciHHV-6 reactivation. Author Summary: Human herpesviruses (HHVs) can reside in a lifelong non-infectious state displaying limited activity in their host and protected from immune responses. One possible way by which HHV-6 achieves this state is by integrating into the telomeric ends of human chromosomes, which are highly repetitive sequences that protect the ends of chromosomes from damage. Various stress conditions can reactivate latent HHV-6 thus increasing the severity of multiple human disorders. Recently, we have identified Chlamydia infection as a natural cause of latent HHV-6 reactivation. Here, we have sought to elucidate the molecular mechanism of HHV-6 reactivation. HHV-6 efficiently utilizes the well-organized telomere maintenance machinery of the host cell to exit from its inactive state and initiate replication to form new viral DNA. We provide experimental evidence that the shortening of telomeres, as a consequence of interference with telomere maintenance, triggers the release of the integrated virus from the chromosome. Our data provide a mechanistic basis to understand HHV-6 reactivation scenarios, which in light of the high prevalence of HHV-6 infection and the possibility of chromosomal integration of other common viruses like HHV-7 have important medical consequences for several million people worldwide. KW - chlamydia infection KW - circular DNA KW - telomeres KW - polymerase chain reaction KW - DNA electrophoresis KW - chromosomes KW - southern hybridization KW - DNA hybridization Y1 - 2013 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-111380 ER -