Projects & Collaborations 13 foundShow per page10 10 20 50 Characterization of accessory proteins and signaling cascades involved in the regulation of the assembly of bacterial Type VI secretion systems Research Project | 1 Project MembersImported from Grants Tool 4701422 The role of type VI secretion systems in infection and competition of intracellular pathogens (BIF Fellowship Hoi Ching Cheung) Research Project | 2 Project MembersThe Type VI Secretion System (T6SS) is a molecular machinery of Gram-negative bacteria which facilitates pathogenesis and interbacterial competition. Understanding the dynamics and function of T6SS is essential for therapeutic insights. Burkholderia is a Gram-negative bacterial genus with species such as B. pseudomallei and B. cenocepacia , which are the causative agents of high mortality diseases such as Melioidosis and Cystic Fibrosis-associated pneumonia in humans. Burkholderia contain one to six different T6SS. One of the systems, T6SS-5, can cause mammalian cells to form multinucleated giant cells (MNGC). These play an important role in virulence because they allow intracellular spread of bacteria. Moreover, Burkholderia uses T6SS-1 to outcompete other bacteria to persist in the host and the environment. In this project, molecular cloning, mammalian cell infection assays and bacterial competition assays, combined with confocal imaging and transcriptomic analysis, will be carried out to determine the dynamics and regulation of T6SS in phagocytic and non-phagocytic mammalian cells and during bacterial competitions. Furthermore, the role of host cell signals in T6SS regulation will be explored. This will provide insight on therapeutic targets for inhibiting the spread and attenuating virulence of Burkholderia infections. Home delivery: Bacterial sRNA mediated treatment of intestinal bowel disease (Freenovation Grant, Miro Plum and Maxim Kolesnikov) Research Project | 3 Project MembersInflammatory bowel disease is characterized by persistent inflammation of the colon and represents an increasing problem worldwide. Existing treatments are broad-acting, unspecific and are limited to temporary relief of symptoms. Moreover, the disease is highly heterogeneous, with a variety of possible causes that range from genetic to environmental. In our project, we will engineer a bacterium that secretes anti-inflammatory RNA in extracellular vesicles. Such a "living pill" may pave the way for further applications that involve RNA dependent regulation. A Death-Dealing Bacterial Nanomachine Research Project | 3 Project MembersType 6 secretion systems (T6SS) are harpoon-like nanomachines that Gram-negative bacterial cells employ to kill other bacterial and eukaryotic cells.Briefly, the T6SS weaponry is tethered to the bacterial cell envelope by a membrane complex, that serves as a platform upon which a baseplate, an extended spring-like sheath and a central spike are assembled. Sheath contraction is biochemically triggered and results in a rapid ejection of the central spike that pierces through a neighboring cell membrane to deliver toxins and other effectors into it. While fluorescence imaging and structural methods have provided deep insight into T6SS structure and function, its destructive mode of action remains unresolved. Here, we will use high-speed atomic force microscope (HS-AFM) imaging, as well as AFM indentation-type force spectroscopy and confocal microscopy (CM) to study the nanomechanical basis by which the T6SS spike punctures bacterial and eukaryotic cell membranes. Mechanisms of subcellular localization of the bacterial Type 6 secretion system assembly Research Project | 1 Project MembersNo Description available NCCR AntiResist: New approaches to combat antibiotic-resistant bacteria Umbrella Project | 32 Project MembersAntibiotics are powerful and indispensable drugs to treat life threatening bacterial infections such as sepsis or pneumonia. Antibiotics also play a central role in many other areas of modern medicine, in particular to protect patients with compromised immunity during cancer therapies, transplantations or surgical interventions. These achievements are now at risk, with the fraction of bacterial pathogens that are resistant to one or more antibiotics steadily increasing. In addition, development of novel antimicrobials lags behind, suffering from inherently high attrition rates in particular for drug candidates against the most problematic Gram-negative bacteria. Together, these factors increasingly limit the options clinicians have for treating bacterial infections. The overarching goal of NCCR AntiResist is to elucidate the physiological properties of bacterial pathogens in infected human patients in order to find new ways of combatting superbugs. Among the many societal, economic, and scientific factors that impact on the development of alternative strategies for antibiotic discovery, our limited understanding of the physiology and heterogeneity of bacterial pathogens in patients ranks highly. Bacteria growing in tissues of patients experience environments very different from standard laboratory conditions, resulting in radically different microbial physiology and population heterogeneity compared to conditions generally used for antibacterial discovery. There is currently no systematic strategy to overcome this fundamental problem. This has resulted in: (i) suboptimal screens that identify new antibiotics, which do not target the special properties of bacteria growing within the patient; (ii) an inability to properly evaluate the efficacy of non-conventional antibacterial strategies; (iii) missed opportunities for entirely new treatment strategies. This NCCR utilizes patient samples from ongoing clinical studies and establishes a unique multidisciplinary network of clinicians, biologists, engineers, chemists, computational scientists and drug developers that will overcome this problem. We are excited to merge these disciplines in order to determine the properties of pathogens infecting patients, establish conditions in the lab that reproduce these properties and utilize these in-vitro models for antimicrobial discovery and development. In addition, clinical-trial networks and the pharmaceutical industry have major footprints in antimicrobial R&D. Exploiting synergies between these players has great potential for making transformative progress in this critical field of human health. This NCCR maintains active collaborations with Biotech SMEs and large pharmaceutical companies with the goal to: accelerate antibiotic discovery by providing relevant read-outs for early prioritization of compounds; enable innovative screens for non-canonical strategies such as anti-virulence inhibitors and immunomodulators; identify new antibacterial strategies that effectively combat bacteria either by targeting refractory subpopulations or by synergizing with bacterial stresses imposed by the patients' own immune system. This NCCR proposes a paradigm shift in antibiotic discovery by investigating the physiology of bacterial pathogens in human patients. This knowledge will be used to develop assays for molecular analyses and drug screening under relevant conditions and to accelerate antibacterial discovery, improve treatment regimens, and uncover novel targets for eradicating pathogens. Through this concerted effort, this NCCR will make a crucial and unique contribution to winning the race against superbugs. Defining the molecular basis of dynamic localization of type VI secretion system assembly in Pseudomonas aeruginosa (BIF Fellowship Maxim Kolesnikov) Research Project | 2 Project MembersThe multifunctional type VI secretion system (T6SS) allows bacteria to deliver effectors to both eukaryotic and prokaryotic cells and thus to gain advantage during bacterial competition as well as pathogenesis. T6SS can be conceptualized as a speargun. Its mode of action is similar to contractile phage tails attached to the cytosolic side of the cell envelope. Upon contraction of T6SS sheath, a needle is protruded from the cell with great force. Knowledge of T6SS assembly regulation remains limited. The opportunistic pathogen Pseudomonas aeruginosa , which encodes three T6SS, targets both prokaryotic and eukaryotic cells by localizing T6SS assembly in response to membrane perturbation, believed to be sensed via a dedicated sensor module. Using P. aeruginosa as a model, the molecular mechanism of signaling in T6SS assembly will be determined by structural, biochemical and microscopy approaches. This work will provide fundamental insights into signaling mechanisms and T6SS assembly in an important human pathogen. Efficient High Resolution Cryo EM Microscopy of large assemblies, membrane proteins and cellular structures Research Project | 3 Project MembersNovel developments in instrumentation and software for cryo electron microscopy have resulted in a "resolution revolution" and have transformed this technique into the most efficient method for high-resolution structure determination of larger biological macromolecules. We are applying cryo electron microscopy to study biological macromolecular assemblies, such as mTOR complexes, highly-regulated multienzymes of human metabolism, polyketide synthases and bacterial Type 6 secretion systems as well as membrane proteins. Understanding the architecture, dynamics and functional mechanisms of these complex target molecules is of highest relevance for fundamental molecular biology, with direct relevance also for cancer therapy and the fight against antibiotic resistance. Despite all progress in electron microscopy methods, extensive sample screening is required for highest resolution analysis. In particular, highly dynamic assemblies and transient complexes affected by conformational and compositional heterogeneity remain a challenge for structural analysis. Here, sample quality often limits the final resolution of structure determination and consequently the quality of mechanistic insights. Often, careful sample stability assays, stabilization by specific or unspecific crosslinking and exhaustive testing of sample vitrification and grid preparation are required to obtain useful results. Here, we request funding for a highly efficient FEG electron microscope with direct electron detector and multi-sample loader for sample screening and analysis. This instrument shall permit high-resolution sample analysis and characterization crucial for the detection of e.g. presence of small protein binding partners, conformational homogeneity, as well as successful trapping and stabilization of specific biological states of the target molecules. Overall, this instrument will uniquely contribute to moving cellular biology analysis to molecular resolution. EMBO Young Investigator Programme Award Research Project | 1 Project MembersMy lab studies structure, function and dynamics of the Type VI secretion system (T6SS). This nanomachine is evolutionarily related to contractile phage tails and delivers proteins into both bacterial and eukaryotic cells. We use various biochemical, genetic and imaging techniques to understand the key molecular mechanisms underlying the assembly, substrate delivery, and regulation of T6SS function during bacterial cell-cell interactions. Cryo-Electron Microscopy in the ZMB of the University of Basel Research Project | 6 Project MembersMit diesem Antrag erbitten wir finanzielle Unterstützung zur Anschaffung eines Elektronenmikroskops für die Untersuchungen von tief-gefrorenen biologischen Proben in der Elektronenmikroskopie Service Facility "BioEM Lab" der Universität Basel. Das BioEM Lab wurde in Frühjahr 2016 als Nachfolger des Zentrum für Mikroskopie (ZMB) der Uni Basel gegründet. Das BioEM Lab bietet seinen Kunden moderne Elektronenmikroskopie Strukturuntersuchungen als Service an. Hierzu gehört unter anderem auch die hoch-auflösende Strukturbestimmung von einzelnen Protein Partikeln, welche im tief-gefrorenen Kryo-Zustand (-190 ºC) mit dem beantragten Elektronenmikroskop fotografiert werden sollen. Durch Computer Bildverarbeitung wird anschliessend die detaillierte 3D Struktur bis hin zu atomarer Auflösung ermittelt. Das ZMB hat bisher diese moderne Methode des "Cryo-EM" nicht anboten. Mit Inbetriebnahme des hier beantragten neuen Gerätes kann das BioEM Lab als Nachfolger des ZMB nun auch diese revolutionäre Methode anbieten. Die Kunden des BioEM Labs sind vorwiegend aus dem Life Sciences Bereich, und schliessen Forschergruppen des Biozentrums der Uni Basel, sowie aus dem weiteren Raum Basel, Schweiz, und benachbartes Ausland mit ein. Mit dem Gerät werden somit zum Beispiel bakterielle Proteine untersucht, um zu verstehen, wie manche gefährlichen Erreger die menschlichen Wirtszellen manipulieren um zu chronischen Infektionen zu führen. Ein Verständnis dieses Mechanismus ist nötig, um gezielte Behandlungsstrategien entwickeln zu können. Andere Beispiele von untersuchten Proben stammen aus der Erforschung der Parkinson'schen Krankheit, oder aus der Nano-Biologie. 12 12
Characterization of accessory proteins and signaling cascades involved in the regulation of the assembly of bacterial Type VI secretion systems Research Project | 1 Project MembersImported from Grants Tool 4701422
The role of type VI secretion systems in infection and competition of intracellular pathogens (BIF Fellowship Hoi Ching Cheung) Research Project | 2 Project MembersThe Type VI Secretion System (T6SS) is a molecular machinery of Gram-negative bacteria which facilitates pathogenesis and interbacterial competition. Understanding the dynamics and function of T6SS is essential for therapeutic insights. Burkholderia is a Gram-negative bacterial genus with species such as B. pseudomallei and B. cenocepacia , which are the causative agents of high mortality diseases such as Melioidosis and Cystic Fibrosis-associated pneumonia in humans. Burkholderia contain one to six different T6SS. One of the systems, T6SS-5, can cause mammalian cells to form multinucleated giant cells (MNGC). These play an important role in virulence because they allow intracellular spread of bacteria. Moreover, Burkholderia uses T6SS-1 to outcompete other bacteria to persist in the host and the environment. In this project, molecular cloning, mammalian cell infection assays and bacterial competition assays, combined with confocal imaging and transcriptomic analysis, will be carried out to determine the dynamics and regulation of T6SS in phagocytic and non-phagocytic mammalian cells and during bacterial competitions. Furthermore, the role of host cell signals in T6SS regulation will be explored. This will provide insight on therapeutic targets for inhibiting the spread and attenuating virulence of Burkholderia infections.
Home delivery: Bacterial sRNA mediated treatment of intestinal bowel disease (Freenovation Grant, Miro Plum and Maxim Kolesnikov) Research Project | 3 Project MembersInflammatory bowel disease is characterized by persistent inflammation of the colon and represents an increasing problem worldwide. Existing treatments are broad-acting, unspecific and are limited to temporary relief of symptoms. Moreover, the disease is highly heterogeneous, with a variety of possible causes that range from genetic to environmental. In our project, we will engineer a bacterium that secretes anti-inflammatory RNA in extracellular vesicles. Such a "living pill" may pave the way for further applications that involve RNA dependent regulation.
A Death-Dealing Bacterial Nanomachine Research Project | 3 Project MembersType 6 secretion systems (T6SS) are harpoon-like nanomachines that Gram-negative bacterial cells employ to kill other bacterial and eukaryotic cells.Briefly, the T6SS weaponry is tethered to the bacterial cell envelope by a membrane complex, that serves as a platform upon which a baseplate, an extended spring-like sheath and a central spike are assembled. Sheath contraction is biochemically triggered and results in a rapid ejection of the central spike that pierces through a neighboring cell membrane to deliver toxins and other effectors into it. While fluorescence imaging and structural methods have provided deep insight into T6SS structure and function, its destructive mode of action remains unresolved. Here, we will use high-speed atomic force microscope (HS-AFM) imaging, as well as AFM indentation-type force spectroscopy and confocal microscopy (CM) to study the nanomechanical basis by which the T6SS spike punctures bacterial and eukaryotic cell membranes.
Mechanisms of subcellular localization of the bacterial Type 6 secretion system assembly Research Project | 1 Project MembersNo Description available
NCCR AntiResist: New approaches to combat antibiotic-resistant bacteria Umbrella Project | 32 Project MembersAntibiotics are powerful and indispensable drugs to treat life threatening bacterial infections such as sepsis or pneumonia. Antibiotics also play a central role in many other areas of modern medicine, in particular to protect patients with compromised immunity during cancer therapies, transplantations or surgical interventions. These achievements are now at risk, with the fraction of bacterial pathogens that are resistant to one or more antibiotics steadily increasing. In addition, development of novel antimicrobials lags behind, suffering from inherently high attrition rates in particular for drug candidates against the most problematic Gram-negative bacteria. Together, these factors increasingly limit the options clinicians have for treating bacterial infections. The overarching goal of NCCR AntiResist is to elucidate the physiological properties of bacterial pathogens in infected human patients in order to find new ways of combatting superbugs. Among the many societal, economic, and scientific factors that impact on the development of alternative strategies for antibiotic discovery, our limited understanding of the physiology and heterogeneity of bacterial pathogens in patients ranks highly. Bacteria growing in tissues of patients experience environments very different from standard laboratory conditions, resulting in radically different microbial physiology and population heterogeneity compared to conditions generally used for antibacterial discovery. There is currently no systematic strategy to overcome this fundamental problem. This has resulted in: (i) suboptimal screens that identify new antibiotics, which do not target the special properties of bacteria growing within the patient; (ii) an inability to properly evaluate the efficacy of non-conventional antibacterial strategies; (iii) missed opportunities for entirely new treatment strategies. This NCCR utilizes patient samples from ongoing clinical studies and establishes a unique multidisciplinary network of clinicians, biologists, engineers, chemists, computational scientists and drug developers that will overcome this problem. We are excited to merge these disciplines in order to determine the properties of pathogens infecting patients, establish conditions in the lab that reproduce these properties and utilize these in-vitro models for antimicrobial discovery and development. In addition, clinical-trial networks and the pharmaceutical industry have major footprints in antimicrobial R&D. Exploiting synergies between these players has great potential for making transformative progress in this critical field of human health. This NCCR maintains active collaborations with Biotech SMEs and large pharmaceutical companies with the goal to: accelerate antibiotic discovery by providing relevant read-outs for early prioritization of compounds; enable innovative screens for non-canonical strategies such as anti-virulence inhibitors and immunomodulators; identify new antibacterial strategies that effectively combat bacteria either by targeting refractory subpopulations or by synergizing with bacterial stresses imposed by the patients' own immune system. This NCCR proposes a paradigm shift in antibiotic discovery by investigating the physiology of bacterial pathogens in human patients. This knowledge will be used to develop assays for molecular analyses and drug screening under relevant conditions and to accelerate antibacterial discovery, improve treatment regimens, and uncover novel targets for eradicating pathogens. Through this concerted effort, this NCCR will make a crucial and unique contribution to winning the race against superbugs.
Defining the molecular basis of dynamic localization of type VI secretion system assembly in Pseudomonas aeruginosa (BIF Fellowship Maxim Kolesnikov) Research Project | 2 Project MembersThe multifunctional type VI secretion system (T6SS) allows bacteria to deliver effectors to both eukaryotic and prokaryotic cells and thus to gain advantage during bacterial competition as well as pathogenesis. T6SS can be conceptualized as a speargun. Its mode of action is similar to contractile phage tails attached to the cytosolic side of the cell envelope. Upon contraction of T6SS sheath, a needle is protruded from the cell with great force. Knowledge of T6SS assembly regulation remains limited. The opportunistic pathogen Pseudomonas aeruginosa , which encodes three T6SS, targets both prokaryotic and eukaryotic cells by localizing T6SS assembly in response to membrane perturbation, believed to be sensed via a dedicated sensor module. Using P. aeruginosa as a model, the molecular mechanism of signaling in T6SS assembly will be determined by structural, biochemical and microscopy approaches. This work will provide fundamental insights into signaling mechanisms and T6SS assembly in an important human pathogen.
Efficient High Resolution Cryo EM Microscopy of large assemblies, membrane proteins and cellular structures Research Project | 3 Project MembersNovel developments in instrumentation and software for cryo electron microscopy have resulted in a "resolution revolution" and have transformed this technique into the most efficient method for high-resolution structure determination of larger biological macromolecules. We are applying cryo electron microscopy to study biological macromolecular assemblies, such as mTOR complexes, highly-regulated multienzymes of human metabolism, polyketide synthases and bacterial Type 6 secretion systems as well as membrane proteins. Understanding the architecture, dynamics and functional mechanisms of these complex target molecules is of highest relevance for fundamental molecular biology, with direct relevance also for cancer therapy and the fight against antibiotic resistance. Despite all progress in electron microscopy methods, extensive sample screening is required for highest resolution analysis. In particular, highly dynamic assemblies and transient complexes affected by conformational and compositional heterogeneity remain a challenge for structural analysis. Here, sample quality often limits the final resolution of structure determination and consequently the quality of mechanistic insights. Often, careful sample stability assays, stabilization by specific or unspecific crosslinking and exhaustive testing of sample vitrification and grid preparation are required to obtain useful results. Here, we request funding for a highly efficient FEG electron microscope with direct electron detector and multi-sample loader for sample screening and analysis. This instrument shall permit high-resolution sample analysis and characterization crucial for the detection of e.g. presence of small protein binding partners, conformational homogeneity, as well as successful trapping and stabilization of specific biological states of the target molecules. Overall, this instrument will uniquely contribute to moving cellular biology analysis to molecular resolution.
EMBO Young Investigator Programme Award Research Project | 1 Project MembersMy lab studies structure, function and dynamics of the Type VI secretion system (T6SS). This nanomachine is evolutionarily related to contractile phage tails and delivers proteins into both bacterial and eukaryotic cells. We use various biochemical, genetic and imaging techniques to understand the key molecular mechanisms underlying the assembly, substrate delivery, and regulation of T6SS function during bacterial cell-cell interactions.
Cryo-Electron Microscopy in the ZMB of the University of Basel Research Project | 6 Project MembersMit diesem Antrag erbitten wir finanzielle Unterstützung zur Anschaffung eines Elektronenmikroskops für die Untersuchungen von tief-gefrorenen biologischen Proben in der Elektronenmikroskopie Service Facility "BioEM Lab" der Universität Basel. Das BioEM Lab wurde in Frühjahr 2016 als Nachfolger des Zentrum für Mikroskopie (ZMB) der Uni Basel gegründet. Das BioEM Lab bietet seinen Kunden moderne Elektronenmikroskopie Strukturuntersuchungen als Service an. Hierzu gehört unter anderem auch die hoch-auflösende Strukturbestimmung von einzelnen Protein Partikeln, welche im tief-gefrorenen Kryo-Zustand (-190 ºC) mit dem beantragten Elektronenmikroskop fotografiert werden sollen. Durch Computer Bildverarbeitung wird anschliessend die detaillierte 3D Struktur bis hin zu atomarer Auflösung ermittelt. Das ZMB hat bisher diese moderne Methode des "Cryo-EM" nicht anboten. Mit Inbetriebnahme des hier beantragten neuen Gerätes kann das BioEM Lab als Nachfolger des ZMB nun auch diese revolutionäre Methode anbieten. Die Kunden des BioEM Labs sind vorwiegend aus dem Life Sciences Bereich, und schliessen Forschergruppen des Biozentrums der Uni Basel, sowie aus dem weiteren Raum Basel, Schweiz, und benachbartes Ausland mit ein. Mit dem Gerät werden somit zum Beispiel bakterielle Proteine untersucht, um zu verstehen, wie manche gefährlichen Erreger die menschlichen Wirtszellen manipulieren um zu chronischen Infektionen zu führen. Ein Verständnis dieses Mechanismus ist nötig, um gezielte Behandlungsstrategien entwickeln zu können. Andere Beispiele von untersuchten Proben stammen aus der Erforschung der Parkinson'schen Krankheit, oder aus der Nano-Biologie.
