The Experts below are selected from a list of 243 Experts worldwide ranked by ideXlab platform
Badreddine Douzi - One of the best experts on this subject based on the ideXlab platform.
-
Unraveling the Self-Assembly of the Pseudomonas aeruginosa XcpQ Secretin Periplasmic Domain Provides New Molecular Insights into Type II Secretion System Secreton Architecture and Dynamics
mBio, 2017Co-Authors: Badreddine Douzi, Nhung Trinh, Sandra Michel-souzy, Aline Desmyter, Genevieve Ball, Pascale Barbier, Artemis Kosta, Eric Durand, Katrina Forest, Christian CambillauAbstract:The type II Secretion system (T2SS) releases large folded exoproteins across the envelope of many Gram-negative pathogens. This Secretion Process therefore requires specific gating, interacting, and dynamics properties mainly operated by a bipartite outer membrane channel called secretin. We have a good understanding of the structure-function relationship of the pore-forming C-terminal domain of secretins. In contrast, the high flexibility of their periplasmic N-terminal domain has been an obstacle in obtaining the detailed structural information required to uncover its molecular function. InPseudomonas aeruginosa, the Xcp T2SS plays an important role in bacterial virulence by its capacity to deliver a large panel of toxins and degradative enzymes into the surrounding environment. Here, we revealed that the N-terminal domain of XcpQ secretin spontaneously self-assembled into a hexamer of dimers independently of its C-terminal domain. Furthermore, and by using multidisciplinary approaches, we elucidate the structural organization of the XcpQ N domain and demonstrate that secretin flexibility at interdimer interfaces is mandatory for its function.IMPORTANCEBacterial secretins are large homooligomeric proteins constituting the outer membrane pore-forming element of several envelope-embedded nanomachines essential in bacterial survival and pathogenicity. They comprise a well-defined membrane-embedded C-terminal domain and a modular periplasmic N-terminal domain involved in substrate recruitment and connection with inner membrane components. We are studying the XcpQ secretin of the T2SS present in the pathogenic bacteriumPseudomonas aeruginosaOur data highlight the ability of the XcpQ N-terminal domain to spontaneously oligomerize into a hexamer of dimers. Furtherin vivoexperiments revealed that this domain adopts different conformations essential for the T2SS Secretion Process. These findings provide new insights into the functional understanding of bacterial T2SS secretins.
-
Deciphering the Xcp Pseudomonas aeruginosa type II Secretion machinery through multiple interactions with substrates.
Journal of Biological Chemistry, 2011Co-Authors: Badreddine Douzi, Genevieve Ball, Christian Cambillau, Mariella Tegoni, Romé VoulhouxAbstract:The type II Secretion system enables gram-negative bacteria to secrete exoproteins into the extracellular milieu. We performed biophysical and biochemical experiments to identify systematic interactions between Pseudomonas aeruginosa Xcp type II Secretion system components and their substrates. We observed that three Xcp components, XcpP(C), the secretin XcpQ(D), and the pseudopilus tip, directly and specifically interact with secreted exoproteins. We established that XcpP(C), in addition to its interaction with the substrate, likely shields the entire periplasmic portion of the secreton. It can therefore be considered as the recruiter of the machinery. Moreover, the direct interaction observed between the substrate and the pseudopilus tip validates the piston model hypothesis, in which the pseudopilus pushes the substrate through the secretin pore during the Secretion Process. All together, our results allowed us to propose a model of the different consecutive steps followed by the substrate during the type II Secretion Process.
Ulises Urzúa - One of the best experts on this subject based on the ideXlab platform.
-
sat0175 alterations of vamp2 and sintaxin 2 in salivary acinar cells modify the Secretion Process in sjogren s syndrome patients
Annals of the Rheumatic Diseases, 2013Co-Authors: Marianela Sánchez, Sergio Aguilera, María-josé Barrera, Cecilia Alliende, Verónica Bahamondes, Isabel Castro, Sergio González, Claudio Molina, Cecilia Leyton, Ulises UrzúaAbstract:Background Sjogren’s syndrome patients (SS) have oral and ocular dryness attributed to alterations in the quantity and quality of saliva and tears (1). We have demonstrated that disruption of cell-cell and cell-extracellular matrix (ECM) interactions that could modify the secretory pathway (2). Salivary acinar cells of SS-patients display alterations in their cell-polarity, affecting the correct localization and function of proteins composing the secretory machinery. SNARE-complexes are responsible for the Secretion of proteins which are essential components of saliva. VAMP-2 and STX2 are SNARE-proteins that participate in the constitutive and regulated Secretion. VAMP-2 is located in secretory granules next to the basolateral plasma membrane and it is involved in vesicular traffic towards the apical and basolateral poles, while STX2 is exclusively located in the apical plasma membrane (3). Objectives To determine the expression and localization of VAMP-2 and STX2, as well as their ability to form SNARE complexes under basal conditions in salivary acinar cells of SS-patients. Methods In labial salivary glands from SS-patients (n=27) and control subjects (n=17) mRNA and proteins levels of SNARE complex components were determined by real-time PCR and Western blotting, respectively. Protein localization was evaluated by immunofluorescence and confocal microscopy. Results In controls and SS-patients, STX2 and VAMP-2 mRNA levels remained unchanged. STX2 protein levels were found significantly augmented in SS-patients, while VAMP-2 protein levels did not change. In SS-patients, VAMP-2 showed a significant increase in apical staining, while STX2 showed decreased apical staining and apico-basal redistribution. Interestingly, increased formation of SNARE complexes containing VAMP-2 or STX2 in a manner independent of external secretory stimuli was detected in SS-patients. Conclusions In SS-patients, VAMP-2 was relocated to the apical cytoplasm; a change that might compensate the significant decrease of VAMP-8 observed in the apical region, as reported previously (4). Apical relocalization of VAMP-2 has been reported in VAMP-8 knockout mice, associated to a significant increase of VAMP-2 protein levels. Regardless of VAMP-2 relocalization, these mice show a decreased the secretory response to secretagogues (5). Conversely, STX2 protein levels increased significantly and showed a strong relocation from apical to basal plasma membrane. These findings suggest a probable loss of exocytic ability by the apical pole of acinar cells that may affect the destination of Secretion proteins, thus impairing physiological production of saliva. Furthermore, a higher quantity of fusion complexes containing the studied proteins has been detected in SS-patients. The ectopic formation of SNARE-complexes in the basal domain of acinar cells is probably enhanced. FONDECYT #1120062, 1080006 and CONICYT PhD fellowship (BMJ). References Fox RI. Lancet. 2005;366:321-31 Ewert P. et al. Arthritis Rheum 2010;62:1280–9. Gaisano HY et al. Gastroenterology. 1996;111:1661-9 Barrera MJ et al. J. Autoimmunity, 2012 in press Wang CC et al. Dev Cell. 2004;7:359-71. Disclosure of Interest None Declared
-
SAT0175 Alterations of VAMP2 and SINTAXIN-2 in salivary acinar cells modify the Secretion Process in sjögren’s syndrome patients
Annals of the Rheumatic Diseases, 2013Co-Authors: Marianela Sánchez, Sergio Aguilera, María-josé Barrera, Cecilia Alliende, Verónica Bahamondes, Isabel Castro, Sergio González, Claudio Molina, Cecilia Leyton, Ulises UrzúaAbstract:Background Sjogren’s syndrome patients (SS) have oral and ocular dryness attributed to alterations in the quantity and quality of saliva and tears (1). We have demonstrated that disruption of cell-cell and cell-extracellular matrix (ECM) interactions that could modify the secretory pathway (2). Salivary acinar cells of SS-patients display alterations in their cell-polarity, affecting the correct localization and function of proteins composing the secretory machinery. SNARE-complexes are responsible for the Secretion of proteins which are essential components of saliva. VAMP-2 and STX2 are SNARE-proteins that participate in the constitutive and regulated Secretion. VAMP-2 is located in secretory granules next to the basolateral plasma membrane and it is involved in vesicular traffic towards the apical and basolateral poles, while STX2 is exclusively located in the apical plasma membrane (3). Objectives To determine the expression and localization of VAMP-2 and STX2, as well as their ability to form SNARE complexes under basal conditions in salivary acinar cells of SS-patients. Methods In labial salivary glands from SS-patients (n=27) and control subjects (n=17) mRNA and proteins levels of SNARE complex components were determined by real-time PCR and Western blotting, respectively. Protein localization was evaluated by immunofluorescence and confocal microscopy. Results In controls and SS-patients, STX2 and VAMP-2 mRNA levels remained unchanged. STX2 protein levels were found significantly augmented in SS-patients, while VAMP-2 protein levels did not change. In SS-patients, VAMP-2 showed a significant increase in apical staining, while STX2 showed decreased apical staining and apico-basal redistribution. Interestingly, increased formation of SNARE complexes containing VAMP-2 or STX2 in a manner independent of external secretory stimuli was detected in SS-patients. Conclusions In SS-patients, VAMP-2 was relocated to the apical cytoplasm; a change that might compensate the significant decrease of VAMP-8 observed in the apical region, as reported previously (4). Apical relocalization of VAMP-2 has been reported in VAMP-8 knockout mice, associated to a significant increase of VAMP-2 protein levels. Regardless of VAMP-2 relocalization, these mice show a decreased the secretory response to secretagogues (5). Conversely, STX2 protein levels increased significantly and showed a strong relocation from apical to basal plasma membrane. These findings suggest a probable loss of exocytic ability by the apical pole of acinar cells that may affect the destination of Secretion proteins, thus impairing physiological production of saliva. Furthermore, a higher quantity of fusion complexes containing the studied proteins has been detected in SS-patients. The ectopic formation of SNARE-complexes in the basal domain of acinar cells is probably enhanced. FONDECYT #1120062, 1080006 and CONICYT PhD fellowship (BMJ). References Fox RI. Lancet. 2005;366:321-31 Ewert P. et al. Arthritis Rheum 2010;62:1280–9. Gaisano HY et al. Gastroenterology. 1996;111:1661-9 Barrera MJ et al. J. Autoimmunity, 2012 in press Wang CC et al. Dev Cell. 2004;7:359-71. Disclosure of Interest None Declared
Christian Cambillau - One of the best experts on this subject based on the ideXlab platform.
-
Unraveling the Self-Assembly of the Pseudomonas aeruginosa XcpQ Secretin Periplasmic Domain Provides New Molecular Insights into Type II Secretion System Secreton Architecture and Dynamics
mBio, 2017Co-Authors: Badreddine Douzi, Nhung Trinh, Sandra Michel-souzy, Aline Desmyter, Genevieve Ball, Pascale Barbier, Artemis Kosta, Eric Durand, Katrina Forest, Christian CambillauAbstract:The type II Secretion system (T2SS) releases large folded exoproteins across the envelope of many Gram-negative pathogens. This Secretion Process therefore requires specific gating, interacting, and dynamics properties mainly operated by a bipartite outer membrane channel called secretin. We have a good understanding of the structure-function relationship of the pore-forming C-terminal domain of secretins. In contrast, the high flexibility of their periplasmic N-terminal domain has been an obstacle in obtaining the detailed structural information required to uncover its molecular function. InPseudomonas aeruginosa, the Xcp T2SS plays an important role in bacterial virulence by its capacity to deliver a large panel of toxins and degradative enzymes into the surrounding environment. Here, we revealed that the N-terminal domain of XcpQ secretin spontaneously self-assembled into a hexamer of dimers independently of its C-terminal domain. Furthermore, and by using multidisciplinary approaches, we elucidate the structural organization of the XcpQ N domain and demonstrate that secretin flexibility at interdimer interfaces is mandatory for its function.IMPORTANCEBacterial secretins are large homooligomeric proteins constituting the outer membrane pore-forming element of several envelope-embedded nanomachines essential in bacterial survival and pathogenicity. They comprise a well-defined membrane-embedded C-terminal domain and a modular periplasmic N-terminal domain involved in substrate recruitment and connection with inner membrane components. We are studying the XcpQ secretin of the T2SS present in the pathogenic bacteriumPseudomonas aeruginosaOur data highlight the ability of the XcpQ N-terminal domain to spontaneously oligomerize into a hexamer of dimers. Furtherin vivoexperiments revealed that this domain adopts different conformations essential for the T2SS Secretion Process. These findings provide new insights into the functional understanding of bacterial T2SS secretins.
-
Deciphering the Xcp Pseudomonas aeruginosa type II Secretion machinery through multiple interactions with substrates.
Journal of Biological Chemistry, 2011Co-Authors: Badreddine Douzi, Genevieve Ball, Christian Cambillau, Mariella Tegoni, Romé VoulhouxAbstract:The type II Secretion system enables gram-negative bacteria to secrete exoproteins into the extracellular milieu. We performed biophysical and biochemical experiments to identify systematic interactions between Pseudomonas aeruginosa Xcp type II Secretion system components and their substrates. We observed that three Xcp components, XcpP(C), the secretin XcpQ(D), and the pseudopilus tip, directly and specifically interact with secreted exoproteins. We established that XcpP(C), in addition to its interaction with the substrate, likely shields the entire periplasmic portion of the secreton. It can therefore be considered as the recruiter of the machinery. Moreover, the direct interaction observed between the substrate and the pseudopilus tip validates the piston model hypothesis, in which the pseudopilus pushes the substrate through the secretin pore during the Secretion Process. All together, our results allowed us to propose a model of the different consecutive steps followed by the substrate during the type II Secretion Process.
Romé Voulhoux - One of the best experts on this subject based on the ideXlab platform.
-
Deciphering the Xcp Pseudomonas aeruginosa type II Secretion machinery through multiple interactions with substrates.
Journal of Biological Chemistry, 2011Co-Authors: Badreddine Douzi, Genevieve Ball, Christian Cambillau, Mariella Tegoni, Romé VoulhouxAbstract:The type II Secretion system enables gram-negative bacteria to secrete exoproteins into the extracellular milieu. We performed biophysical and biochemical experiments to identify systematic interactions between Pseudomonas aeruginosa Xcp type II Secretion system components and their substrates. We observed that three Xcp components, XcpP(C), the secretin XcpQ(D), and the pseudopilus tip, directly and specifically interact with secreted exoproteins. We established that XcpP(C), in addition to its interaction with the substrate, likely shields the entire periplasmic portion of the secreton. It can therefore be considered as the recruiter of the machinery. Moreover, the direct interaction observed between the substrate and the pseudopilus tip validates the piston model hypothesis, in which the pseudopilus pushes the substrate through the secretin pore during the Secretion Process. All together, our results allowed us to propose a model of the different consecutive steps followed by the substrate during the type II Secretion Process.
Genevieve Ball - One of the best experts on this subject based on the ideXlab platform.
-
Unraveling the Self-Assembly of the Pseudomonas aeruginosa XcpQ Secretin Periplasmic Domain Provides New Molecular Insights into Type II Secretion System Secreton Architecture and Dynamics
mBio, 2017Co-Authors: Badreddine Douzi, Nhung Trinh, Sandra Michel-souzy, Aline Desmyter, Genevieve Ball, Pascale Barbier, Artemis Kosta, Eric Durand, Katrina Forest, Christian CambillauAbstract:The type II Secretion system (T2SS) releases large folded exoproteins across the envelope of many Gram-negative pathogens. This Secretion Process therefore requires specific gating, interacting, and dynamics properties mainly operated by a bipartite outer membrane channel called secretin. We have a good understanding of the structure-function relationship of the pore-forming C-terminal domain of secretins. In contrast, the high flexibility of their periplasmic N-terminal domain has been an obstacle in obtaining the detailed structural information required to uncover its molecular function. InPseudomonas aeruginosa, the Xcp T2SS plays an important role in bacterial virulence by its capacity to deliver a large panel of toxins and degradative enzymes into the surrounding environment. Here, we revealed that the N-terminal domain of XcpQ secretin spontaneously self-assembled into a hexamer of dimers independently of its C-terminal domain. Furthermore, and by using multidisciplinary approaches, we elucidate the structural organization of the XcpQ N domain and demonstrate that secretin flexibility at interdimer interfaces is mandatory for its function.IMPORTANCEBacterial secretins are large homooligomeric proteins constituting the outer membrane pore-forming element of several envelope-embedded nanomachines essential in bacterial survival and pathogenicity. They comprise a well-defined membrane-embedded C-terminal domain and a modular periplasmic N-terminal domain involved in substrate recruitment and connection with inner membrane components. We are studying the XcpQ secretin of the T2SS present in the pathogenic bacteriumPseudomonas aeruginosaOur data highlight the ability of the XcpQ N-terminal domain to spontaneously oligomerize into a hexamer of dimers. Furtherin vivoexperiments revealed that this domain adopts different conformations essential for the T2SS Secretion Process. These findings provide new insights into the functional understanding of bacterial T2SS secretins.
-
aeruginosa Secretion in Pseudomonas Membrane Is Required for Extracellular Assembly of XcpR in the Cytoplasmic
2013Co-Authors: Gérard Michel, Genevieve Ball, Marc Bally, Virginie Chapon-hervé, Sophie BlevesAbstract:geneproducts. XcpR, a putative nucleotide-binding protein, is essential for the Secretion Process across the outermembrane even though the protein contains no hydrophobic sequence that could target or anchor it to thebacterial envelope. For a better understanding of the relationship between XcpR and the other Xcp proteinswhich are located in the envelope, we have studied its subcellular localization. In a wild-type
-
Deciphering the Xcp Pseudomonas aeruginosa type II Secretion machinery through multiple interactions with substrates.
Journal of Biological Chemistry, 2011Co-Authors: Badreddine Douzi, Genevieve Ball, Christian Cambillau, Mariella Tegoni, Romé VoulhouxAbstract:The type II Secretion system enables gram-negative bacteria to secrete exoproteins into the extracellular milieu. We performed biophysical and biochemical experiments to identify systematic interactions between Pseudomonas aeruginosa Xcp type II Secretion system components and their substrates. We observed that three Xcp components, XcpP(C), the secretin XcpQ(D), and the pseudopilus tip, directly and specifically interact with secreted exoproteins. We established that XcpP(C), in addition to its interaction with the substrate, likely shields the entire periplasmic portion of the secreton. It can therefore be considered as the recruiter of the machinery. Moreover, the direct interaction observed between the substrate and the pseudopilus tip validates the piston model hypothesis, in which the pseudopilus pushes the substrate through the secretin pore during the Secretion Process. All together, our results allowed us to propose a model of the different consecutive steps followed by the substrate during the type II Secretion Process.
