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1. Comparative roadmaps of reprogramming and oncogenic transformation identify Bcl11b and Atoh8 as broad regulators of cellular plasticity

Author

Huyghe, A., Furlan, G., Schroeder, J., Cascales, E., Trajkova, A., Ruel, M., Stüder, F., Larcombe, M., Yang Sun, Y. Bo, Mugnier, F., De Matteo, L., Baygin, A., Wang, J., Yu, Y., Rama, N., Gibert, B., Kielbassa, J., Tonon, L., Wajda, P., Gadot, N., Brevet, M., Siouda, M., Mulligan, P., Dante, R., Liu, P., Gronemeyer, H., Mendoza-Parra, M., Polo, J. M., and Lavial, F.

Published
2022
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2. AB1219 HOW TO TAILOR THE ANTI-OSTEOPOROSIS TREATMENT IN PATIENTS WITH ADVANCED LIVER DISEASE? VARIATIONS OF RENAL FUNCTION BY CREATININE AND CYSTATIN C

Author

Rodríguez Alvear, C., primary, Buendía Sánchez, N., additional, Garín Cascales, E., additional, López González, M. D. C., additional, Perea Martínez, E., additional, Avilés, A., additional, Calabuig, I., additional, Peral, M. L., additional, Bernabéu, P., additional, Martínez-Sanchis, A., additional, Esteve Vives, J. J., additional, Vela-Casasempere, P., additional, Jovani, V., additional, and Andrés, M., additional

Published
2023
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6. Protein Interactome Analysis of the Type IX Secretion System Identifies PorW as the Missing Link between the PorK/N Ring Complex and the Sov Translocon

Author

Khursigara, CM, Gorasia, DG, Silva, IL, Butler, CA, Chabalier, M, Doan, T, Cascales, E, Veith, PD, Reynolds, EC, Khursigara, CM, Gorasia, DG, Silva, IL, Butler, CA, Chabalier, M, Doan, T, Cascales, E, Veith, PD, and Reynolds, EC

Abstract

The type IX secretion system (T9SS) transports cargo proteins through the outer membrane of Bacteroidetes and attaches them to the cell surface for functions including pathogenesis, gliding motility, and degradation of carbon sources. The T9SS comprises at least 20 different proteins and includes several modules: the trans-envelope core module comprising the PorL/M motor and the PorK/N ring, the outer membrane Sov translocon, and the cell attachment complex. However, the spatial organization of these modules is unknown. We have characterized the protein interactome of the Sov translocon in Porphyromonas gingivalis and identified Sov-PorV-PorA as well as Sov-PorW-PorN-PorK to be novel networks. PorW also interacted with PGN_1783 (PorD), which was required for maximum secretion efficiency. The identification of PorW as the missing link completes a continuous interaction network from the PorL/M motor to the Sov translocon, providing a pathway for cargo delivery and energy transduction from the inner membrane to the secretion pore. IMPORTANCE The T9SS is a newly identified protein secretion system of the Fibrobacteres-Chlorobi-Bacteroidetes superphylum used by pathogens associated with diseases of humans, fish, and poultry for the secretion and cell surface attachment of virulence factors. The T9SS comprises three known modules: (i) the trans-envelope core module comprising the PorL/M motor and the PorK/N ring, (ii) the outer membrane Sov translocon, and (iii) the cell surface attachment complex. The spatial organization and interaction of these modules have been a mystery. Here, we describe the protein interactome of the Sov translocon in the human pathogen Porphyromonas gingivalis and have identified PorW as the missing link which bridges PorN with Sov and so completes a continuous interaction network from the PorL/M motor to the Sov translocon, providing, for the first time, a pathway for cargo delivery and energy transduction from the inner membrane to the secretion

Published
2022

7. Comparative roadmaps of reprogramming and oncogenic transformation identify Bcl11b and Atoh8 as broad regulators of cellular plasticity

Author

Huyghe, A, Furlan, G, Schroeder, J, Cascales, E, Trajkova, A, Ruel, M, Studer, F, Larcombe, M, Yang Sun, YB, Mugnier, F, De Matteo, L, Baygin, A, Wang, J, Yu, Y, Rama, N, Gibert, B, Kielbassa, J, Tonon, L, Wajda, P, Gadot, N, Brevet, M, Siouda, M, Mulligan, P, Dante, R, Liu, P, Gronemeyer, H, Mendoza-Parra, M, Polo, JM, Lavial, F, Huyghe, A, Furlan, G, Schroeder, J, Cascales, E, Trajkova, A, Ruel, M, Studer, F, Larcombe, M, Yang Sun, YB, Mugnier, F, De Matteo, L, Baygin, A, Wang, J, Yu, Y, Rama, N, Gibert, B, Kielbassa, J, Tonon, L, Wajda, P, Gadot, N, Brevet, M, Siouda, M, Mulligan, P, Dante, R, Liu, P, Gronemeyer, H, Mendoza-Parra, M, Polo, JM, and Lavial, F

Abstract

Coordinated changes of cellular plasticity and identity are critical for pluripotent reprogramming and oncogenic transformation. However, the sequences of events that orchestrate these intermingled modifications have never been comparatively dissected. Here, we deconvolute the cellular trajectories of reprogramming (via Oct4/Sox2/Klf4/c-Myc) and transformation (via Ras/c-Myc) at the single-cell resolution and reveal how the two processes intersect before they bifurcate. This approach led us to identify the transcription factor Bcl11b as a broad-range regulator of cell fate changes, as well as a pertinent marker to capture early cellular intermediates that emerge simultaneously during reprogramming and transformation. Multiomics characterization of these intermediates unveiled a c-Myc/Atoh8/Sfrp1 regulatory axis that constrains reprogramming, transformation and transdifferentiation. Mechanistically, we found that Atoh8 restrains cellular plasticity, independent of cellular identity, by binding a specific enhancer network. This study provides insights into the partitioned control of cellular plasticity and identity for both regenerative and cancer biology.

Published
2022

10. The Azospirillum brasilense Type VI secretion system promotes cell aggregation, biocontrol protection against phytopathogens and attachment to the microalgae Chlorella sorokiniana

Author

Cassan, Fabricio, Coniglio, Anahí, Amavizca, Edgar, Maroniche, Guillermo, Cascales, E., Bashan, Yoav, De‐bashan, Luz, National University of Río Cuarto = Universidad Nacional de Río Cuarto (UNRC), Auburn University (AU), Universidad Nacional de Mar del Plata [Mar del Plata] (UNMdP), Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), and CASCALES, ERIC

Subjects
[SDV] Life Sciences [q-bio], [SDV]Life Sciences [q-bio]
Abstract

International audience; The plant-growth-promoting bacterium Azospirillum brasilense is able to associate with the microalgae Chlorella sorokiniana. Attachment of A. brasilense increases the metabolic performances of the microalgae. Recent genome analyses have revealed that the A. brasilense Az39 genome contains two complete sets of genes encoding type VI secretion systems (T6SS), including the T6SS1 that is induced by the indole-3-acetic acid (IAA) phytohormone. The T6SS is a multiprotein machine, widespread in Gram-negative bacteria, that delivers protein effectors in both prokaryotic and eukaryotic cells. Here we show that the A. brasilense T6SS is required for Chlorella-Azospirillum synthetic mutualism. Our data demonstrate that the T6SS is an important determinant to promote production of lipids, carbohydrates and photosynthetic pigments by the microalgae. We further show that this is likely due to the role of the T6SS during the attachment stage and for the production of IAA phytohormones. Finally, we demonstrate that the A. brasilense T6SS provides antagonistic activities against a number of plant pathogens such as Agrobacterium, Pectobacterium, Dickeya and Ralstonia species in vitro, suggesting that, in addition to promoting growth, A. brasilense might confer T6SS-dependent bio-control protection to microalgae and plants against bacterial pathogens.

Published
2021

11. Anchoring the T6SS to the Cell Wall: Crystal Structure of the Peptidoglycan Binding Domain of the TagL Accessory Protein

Author

Nguyen, Van Son, Spinelli, Silvia, Cascales, E., Roussel, Alain, Cambillau, Christian, Leone, Philippe, Architecture et fonction des macromolécules biologiques (AFMB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), ANR-20-CE14-0006,Gly-Pro-PH,Cibler le métabolisme des fibroblastes adventitiels activés pour traiter l'hypertension pulmonaire(2020), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), CASCALES, ERIC, and Cibler le métabolisme des fibroblastes adventitiels activés pour traiter l'hypertension pulmonaire - - Gly-Pro-PH2020 - ANR-20-CE14-0006 - AAPG2020 - VALID

Subjects
Polymers, [SDV]Life Sciences [q-bio], Membrane Protein Complexes, Cell Membranes, DNA cloning, Secretion Systems, Pathology and Laboratory Medicine, Biochemistry, Sequence alignment, Microbial Physiology, Medicine and Health Sciences, Macromolecular Structure Analysis, Bacterial Physiology, Materials, Crystallography, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Physics, Type VI Secretion Systems, Condensed Matter Physics, [SDV] Life Sciences [q-bio], Chemistry, Macromolecules, Physical Sciences, Crystal Structure, Medicine, Pathogens, Cellular Structures and Organelles, Research Article, Cell Binding, Protein Structure, Cell Physiology, Virulence Factors, Science, Materials Science, Peptidoglycan, Microbiology, Cell Walls, Bacterial Proteins, Protein Domains, membrane complex, Gram-Negative Bacteria, [CHIM.CRIS]Chemical Sciences/Cristallography, Solid State Physics, Molecular Biology, Biology and Life Sciences, Membrane Proteins, Proteins, Protein Complexes, Bacteriology, Cell Biology, Peptidoglycans, Polymer Chemistry, Outer Membrane Proteins, Type VI secretion, cell wall, Molecular structure
Abstract

International audience; The type VI secretion system (T6SS) is a widespread mechanism of protein delivery into target cells, present in more than a quarter of all sequenced Gram-negative bacteria. The T6SS constitutes an important virulence factor, as it is responsible for targeting effectors in both prokaryotic and eukaryotic cells. The T6SS comprises a tail structure tethered to the cell envelope via a trans-envelope complex. In most T6SS, the membrane complex is anchored to the cell wall by the TagL accessory protein. In this study, we report the first crystal structure of a peptidoglycan-binding domain of TagL. The fold is conserved with members of the OmpA/Pal/MotB family, and more importantly, the peptidoglycan binding site is conserved. This structure further exemplifies how proteins involved in anchoring to the cell wall for 2 different cellular functions rely on an interaction network with peptidoglycan strictly conserved.

Published
2021

12. The Type IX Secretion System and Its Role in Bacterial Function and Pathogenesis

Author

Veith, PD, Glew, MD, Gorasia, DG, Cascales, E, Reynolds, EC, Veith, PD, Glew, MD, Gorasia, DG, Cascales, E, and Reynolds, EC

Abstract

Porphyromonas, Tannerella, and Prevotella species found in severe periodontitis use the Type IX Secretion System (T9SS) to load their outer membrane surface with an array of virulence factors. These virulence factors are then released on outer membrane vesicles (OMVs), which penetrate the host to dysregulate the immune response to establish a positive feedback loop of chronic, inflammatory destruction of the tooth's supporting tissues. In this review, we present the latest information on the molecular architecture of the T9SS and provide mechanistic insight into its role in secretion and attachment of cargo proteins to produce a virulence coat on cells and OMVs. The recent molecular structures of the T9SS motor comprising PorL and PorM as well as the secretion pore Sov, together with advances in the overall interactome, have provided insight into the possible mechanisms of secretion. We propose the presence of PorL/M motors arranged in a circle at the inner membrane with bent periplasmic rotors interacting with the PorN protein. At the outer membrane, we envisage a slide carousel model where the PorN protein is driven around a circular track composed of PorK. Cargo proteins are transported by PorN to PorW and the Sov translocon just as slides are rotated to the projection window. Secreted proteins are proposed to then be shuttled along highways consisting of the PorV shuttle protein to an array of attachment complexes distributed around the cell. The cell surface attachment of cargo is a hallmark of the T9SS, and in Porphyromonas gingivalis and Tannerella forsythia, this attachment is achieved via covalent bonding to a linking sugar synthesized by the Wbp/Vim pathway. The cell-surface attached cargo are enriched on OMVs, which are then released from the cell.

Published
2021

13. Fur -Dam regulatory interplay at an internal promoter of the enteroaggregative Escherichia coli Type VI secretion sci1 gene cluster

Author

Brunet, Yannick R, Bernard, Christophe S, Cascales, E., Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and CASCALES, ERIC

Subjects
[SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, ComputingMilieux_MISCELLANEOUS
Abstract

International audience

Published
2020

14. Combined Body Mass Index and Waist-to-Height Ratio and Its Association with Lifestyle and Health Factors among Spanish Children: The PASOS Study

Author

Bibiloni, Maria del Mar, Gallardo Alfaro, Laura, Gómez, S.F., Warnberg, Julia, Oses, Maddi, Gonzalez Gross, Marcela, Gusi, Narcis, Aznar Laín, Susana, Marín Cascales, E., González Valeiro, Miguel, Serra Majem, Lluis, Terrados, Nicolás, Segu, Marta, Lassale, Camille, Homs, Clara, Benavente Marín, Juan Carlos, Labayen, Idoia, Augusto G. Zapico, G. Zapico, Augusto, Sánchez Gómez, Jesús, Jiménez Zazo, Fabio, Alcaraz, Pedro E., Sevilla Sanchez, Marta, Herrera Ramos, Estefanía, Pulgar, Susana, Sistac, Clara, Schröder, Helmut, Bouzas, C., Tur, Josep A., Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa. ISFOOD - Institute for Innovation and Sustainable Development in Food Chain, Universidad Pública de Navarra. Departamento de Ciencias de la Salud, and Nafarroako Unibertsitate Publikoa. Osasun Zientziak Saila

Subjects
Male, lifestyle, Pediatric Obesity, Adolescent, Child Behavior, Mothers, PASOS, Adolescents, Mediterranean diet, children, adolescents, Article, Body Mass Index, Estilo de vida, Screen Time, Risk Factors, Prevalence, Humans, TX341-641, Nenos, Child, Exercise, Life Style, Adolescentes, Children, Waist-Height Ratio, Nutrition and Dietetics, Anthropometry, Nutrition. Foods and food supply, Lifestyle, Dieta mediterránea, Cross-Sectional Studies, Nutrición, Spain, Educational Status, Female, Food Science
Abstract

Background and Aims: The World Health Organization recommended simultaneous measurement of body mass index (BMI) and waist circumference (WC) and suggested joint use to predict disease risks. The aim of this study was to assess the prevalence of BMI and waist-to-height ratio (WHtR) categories among Spanish children and adolescents, as well as their associations with several lifestyle factors. Methods: Cross-sectional analysis of 8–16-year-old children and adolescents (n = 3772) were included in the PASOS nationwide representative study. Children/adolescents and their mothers/female caregivers answered a questionnaire on lifestyle and health factors. Child/adolescent anthropometrics were measured. Four combined BMI-WHtR disease risk categories were built. Results: A third of participants showed combined BMI-WHtR categories with high disease risk (12.3% ‘increased risk’, 9.7% ‘high risk’, 14.3% ‘very high risk’). Participants in the ‘very high risk’ group were less likely to be females (odds ratio 0.63; 95% CI: 0.52–0.76) and adolescents (0.60; 95% CI: 0.49–0.72), to practice ≥60 min/day of moderate-vigorous physical activity (MVPA) (0.73; 95% CI: 0.57–0.93), and to watch

Published
2022

15. The Type IX Secretion System and Its Role in Bacterial Function and Pathogenesis.

Author

Veith, P.D., Glew, M.D., Gorasia, D.G., Cascales, E., and Reynolds, E.C.

Abstract

Porphyromonas, Tannerella, and Prevotella species found in severe periodontitis use the Type IX Secretion System (T9SS) to load their outer membrane surface with an array of virulence factors. These virulence factors are then released on outer membrane vesicles (OMVs), which penetrate the host to dysregulate the immune response to establish a positive feedback loop of chronic, inflammatory destruction of the tooth's supporting tissues. In this review, we present the latest information on the molecular architecture of the T9SS and provide mechanistic insight into its role in secretion and attachment of cargo proteins to produce a virulence coat on cells and OMVs. The recent molecular structures of the T9SS motor comprising PorL and PorM as well as the secretion pore Sov, together with advances in the overall interactome, have provided insight into the possible mechanisms of secretion. We propose the presence of PorL/M motors arranged in a circle at the inner membrane with bent periplasmic rotors interacting with the PorN protein. At the outer membrane, we envisage a slide carousel model where the PorN protein is driven around a circular track composed of PorK. Cargo proteins are transported by PorN to PorW and the Sov translocon just as slides are rotated to the projection window. Secreted proteins are proposed to then be shuttled along highways consisting of the PorV shuttle protein to an array of attachment complexes distributed around the cell. The cell surface attachment of cargo is a hallmark of the T9SS, and in Porphyromonas gingivalis and Tannerella forsythia, this attachment is achieved via covalent bonding to a linking sugar synthesized by the Wbp/Vim pathway. The cell-surface attached cargo are enriched on OMVs, which are then released from the cell. [ABSTRACT FROM AUTHOR]

Published
2022
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16. In vivo TssA proximity labeling reveals temporal interactions during Type VI secretion 1 biogenesis and TagA, a protein that stops and holds the sheath

Author

Santin, Yoann, Doan, Thierry, Lebrun, Régine, Espinosa, Leon, Journet, Laure, Cascales, E., Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), Plateforme Protéomique [Marseille], Institut de Microbiologie de la Méditerranée (IMM), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de chimie bactérienne (LCB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), ANR-14-CE14-0006,B-War,Guerre bactérienne: Architecture et Fonction du Système de Sécrétion de Type VI(2014), ANR-17-CE11-0039,T6-PLATFORM,Une approche multidisciplinaire et intégrative pour comprendre l'assemblage, la structure et la dynamique d'une plateforme de queue contractile(2017), and Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
[SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, [SDV]Life Sciences [q-bio], [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology
Abstract

International audience; The Type VI secretion system (T6SS) is a multiprotein weapon used by bacteria to destroy competitor cells. The T6SS contractile sheath wraps an effector-loaded syringe that is injected into the target cell. This tail structure assembles onto the baseplate that is docked to the membrane complex. In entero-aggregative Escherichia coli TssA plays a central role at each stage of the T6SS assembly pathway by stabilizing the baseplate and coordinating the polymerization of the tail. Here we adapted an assay based on APEX2-dependent biotinylation to identify the proximity partners of TssA in vivo. By using stage-blocking mutations, we define the temporal contacts of TssA during T6SS biogenesis. This proteomic mapping approach also revealed an additional partner of TssA, TagA. We show that TagA is a cytosolic protein tightly associated with the membrane. Analyses of sheath dynamics further demonstrate that TagA captures the distal end of the sheath to stop its polymerization and to maintain it under the extended conformation.

Published
2018

17. Chapter 16. Fusion reporter approaches to monitor transmembrane helix interactions in bacterial membranes Running head: protein-protein interaction in the membrane

Author

Logger, Laureen, Zoued, Abdelrahim, Cascales, E., Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and CASCALES, ERIC

Subjects
cI repressor, BACTH, GALLEX, [SDV]Life Sciences [q-bio], helix, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, heterotypic, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, [SDV] Life Sciences [q-bio], protein-protein interaction, TOXCAT, one-hybrid, transmembrane segment, helix interaction, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, membrane protein, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, two-hybrid, [SDV.MP.BAC] Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], homotypic, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM]
Abstract

International audience; In transenvelope multiprotein machines such as bacterial secretion systems, protein-protein interactions not only occur between soluble domains but are also mediated by helix-helix contacts in the inner membrane. Here, we describe genetic assays commonly used to test interactions between transmembrane α-helices in their native membrane environment. These assays are based on the reconstitution of dimeric regulators allowing the expression of reporter genes. We provide detailed protocols for the TOXCAT and GALLEX assays used to monitor homotypic and heterotypic transmembrane helix-helix interactions.

Published
2017

18. Chapter 12. Measure of peptidoglycan hydrolase activity Running head: peptidoglycan remodelling enzymes

Author

Cascales, E., Santin, Yoann, Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), CASCALES, ERIC, and Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
[SDV]Life Sciences [q-bio], remazol blue, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, peptidoglycan, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, [SDV] Life Sciences [q-bio], [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, cell wall, lytic transglycosylase, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, localized degradation, [SDV.MP.BAC] Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM]
Abstract

International audience; Most of the gene clusters encoding multiprotein complexes of the bacterial cell envelope, such as conjugation and secretion systems, Type IV pili and flagella, bear a gene encoding an enzyme with peptidoglycan hydrolase activity. These enzymes are usually glycoside hydrolases that cleave the glycan chains of the peptidoglycan. Their activities are spatially controlled to avoid cell lysis and to create localized rearrangement of the cell wall. This is assured by interaction with structural subunits of the apparatus. Here, we describe protocols to test the peptidoglycan hydrolase activity of these proteins in vitro and in solution.

Published
2017

19. Probing inner membrane protein topology by proteolysis running head: Protease accessibility

Author

Vincent, Maxence, Cascales, E., CASCALES, ERIC, Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU)

Subjects
proteolysis, topology, polytopic, [SDV]Life Sciences [q-bio], bitopic, proteinase K, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, protease, trans-membrane segment, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, insertion, [SDV] Life Sciences [q-bio], [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, membrane protein, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.MP.BAC] Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, carboxypeptidase Y, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], inner membrane
Abstract

International audience; Inner membrane proteins are inserted into the membrane via α-helices. These helices are not only membrane anchors but usually mediate specific interactions with membrane protein partners or participate to energetic processes. The number, location and orientation of these helices is referred as topology. Bitopic membrane proteins that consist of a single membrane-embedded domain connecting two soluble domains are distinguished from polytopic ones that consist to multiple-spanning TMH connected by extramembrane domains. Defining inner membrane protein topology could be achieved by different methods. Here, we describe a protease accessibility assay that allows to define topology based on digestion profiles.

Published
2017

23. Priming and polymerization of a bacterial contractile tail structure. 1' 2' 17'

Author

Zoued, Abdelrahim, Durand, Eric, Brunet, Yannick, Spinelli, Silvia, Douzi, Badreddine, Guzzo, Mathilde, Flaugnatti, Nicolas, Legrand, Pierre, Journet, Laure, Fronzes, Rémi, Mignot, Tam, Cambillau, Christian, Cascales, E., Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Architecture et fonction des macromolécules biologiques (AFMB), Institut National de la Recherche Agronomique (INRA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de chimie bactérienne (LCB), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Synchrotron SOLEIL (SSOLEIL), Centre National de la Recherche Scientifique (CNRS), European Institute of Chemistry and Biology, Université Sciences et Technologies - Bordeaux 1, Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Institut National de la Recherche Agronomique (INRA), Université Sciences et Technologies - Bordeaux 1 (UB), and CASCALES, ERIC

Subjects
Models, Molecular, Escherichia coli/*chemistry/ultrastructure, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], [SDV]Life Sciences [q-bio], [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Crystallography, X-Ray, Protein Structure, Tertiary, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], [SDV] Life Sciences [q-bio], Microscopy, Electron, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Microscopy, Fluorescence, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Escherichia coli Proteins/*chemistry/*metabolism/ultrastructure, Type VI Secretion Systems/chemistry/metabolism/ultrastructure, Polymerization, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.BC] Life Sciences [q-bio]/Cellular Biology, [SDV.MP] Life Sciences [q-bio]/Microbiology and Parasitology
Abstract

Zoued, Abdelrahim Durand, Eric Brunet, Yannick R Spinelli, Silvia Douzi, Badreddine Guzzo, Mathilde Flaugnatti, Nicolas Legrand, Pierre Journet, Laure Fronzes, Remi Mignot, Tam Cambillau, Christian Cascales, Eric eng Research Support, Non-U.S. Gov't England Nature. 2016 Mar 3;531(7592):59-63. doi: 10.1038/nature17182. Epub 2016 Feb 24.; International audience; Contractile tails are composed of an inner tube wrapped by an outer sheath assembled in an extended, metastable conformation that stores mechanical energy necessary for its contraction. Contraction is used to propel the rigid inner tube towards target cells for DNA or toxin delivery. Although recent studies have revealed the structure of the Type VI secretion system contractile sheath, the mechanisms by which its polymerization is controlled and coordinated with inner tube assembly remain unsolved. In this study, we report that the starfish-like TssA dodecameric complex interacts with tube and sheath components. Fluorescence microscopy experiments revealed that TssA binds first to the T6SS membrane core complex and then initiates tail polymerization. TssA remains at the tip of the growing structure and incorporates new tube and sheath blocks. Based on these results, we propose that TssA primes and coordinates tail tube and sheath biogenesis.

Published
2016

24. Polyamino-Isoprenic Derivatives Block Intrinsic Resistance of P. aeruginosa to Doxycycline and Chloramphenicol In Vitro

Author

Borselli, Diane, Lieutaud, Aurélie, Thefenne, Hélène, Garnotel, Eric, Pagès, Jean-Marie, Brunel, Jean Michel, Bolla, Jean-Michel, Cascales, E., Transporteurs membranaires, chimioresistance et drug-design (TMCD2), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM), Hôpital d'Instruction des Armées Laveran, Service de Santé des Armées, Centre de Recherche en Cancérologie de Marseille (CRCM), Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU)

Subjects
0301 basic medicine, Antibiotics, lcsh:Medicine, Drug resistance, Pharmacology, medicine.disease_cause, Pathology and Laboratory Medicine, Medicine and Health Sciences, lcsh:Science, Doxycycline, Multidisciplinary, biology, Antimicrobials, Organic Compounds, Monosaccharides, Pseudomonas Aeruginosa, Drugs, 3. Good health, Bacterial Pathogens, Chemistry, Medical Microbiology, Physical Sciences, Efflux, Pathogens, medicine.drug, Research Article, Gram-negative bacteria, medicine.drug_class, 030106 microbiology, Carbohydrates, Microbial Sensitivity Tests, Library Screening, In Vitro Techniques, Research and Analysis Methods, Microbiology, 03 medical and health sciences, Antibiotic resistance, Pseudomonas, Microbial Control, Drug Resistance, Bacterial, medicine, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Molecular Biology Techniques, Microbial Pathogens, Molecular Biology, Gram Negative Bacteria, Molecular Biology Assays and Analysis Techniques, Dose-Response Relationship, Drug, Bacteria, Pseudomonas aeruginosa, Chloramphenicol, lcsh:R, Organic Chemistry, Organisms, Chemical Compounds, Biology and Life Sciences, Bacteriology, biology.organism_classification, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, 030104 developmental biology, Glucose, Antibiotic Resistance, lcsh:Q, Antimicrobial Resistance
Abstract

International audience; Multidrug resistant bacteria have been a worldwide concern for decades. Though new molecules that effectively target Gram-positive bacteria are currently appearing on the market, a gap remains in the treatment of infections caused by Gram-negative bacteria. Therefore, new strategies must be developed against these pathogens. The aim of this study was to select an antibiotic for which a bacterium is naturally resistant and to use an escort molecule to restore susceptibility, similarly to the model of β-lactam/ β-lactamase inhibitors. High-content screening was performed on the reference strain PA01, allowing the selection of four polyamino-isoprenic compounds that acted synergistically with doxycycline. They were assayed against clinical isolates and Multi-Drug-Resistant strains. One of these compounds was able to decrease the MIC of doxycycline on the reference strain, efflux pump overpro-ducers and clinical isolates of P. aeruginosa, to the susceptibility level. Similar results were obtained using chloramphenicol as the antibiotic. Membrane permeation assays and real-time efflux experiments were used to characterize the mechanism of doxycycline potentiation. The results showed that the selected compound strongly decreases the efficiency of glucose triggered efflux associated with a slight destabilization of the outer membrane. According to these data, targeting natural resistance may become an interesting way to combat MDR pathogens and could represent an alternative to already devised strategies.

Published
2015

26. The Type VI Secretion TssEFGK-VgrG Phage-Like Baseplate Is Recruited to the TssJLM Membrane Complex via Multiple Contacts and Serves As Assembly Platform for Tail Tube/Sheath Polymerization

Author

Brunet, Yannick, Zoued, Abdelrahim, Boyer, Frédéric, Douzi, Badreddine, Cascales, E., Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), Laboratoire d'Ecologie Alpine (LECA), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), Architecture et fonction des macromolécules biologiques (AFMB), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Institut National de la Recherche Agronomique (INRA), ANR-10-JCJC-1303,A Fun T6SS,Assemblage et Fonction des Systèmes de Sécrétion de Type VI bactériens(2010), ANR-14-CE14-0006,B-War,Guerre bactérienne: Architecture et Fonction du Système de Sécrétion de Type VI(2014), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Université Joseph Fourier - Grenoble 1 (UJF)-Université Grenoble Alpes (UGA), Institut National de la Recherche Agronomique (INRA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), and Université Joseph Fourier - Grenoble 1 (UJF)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Centre National de la Recherche Scientifique (CNRS)

Subjects
Fluorescence microscopy, Cytoplasm, lcsh:QH426-470, Sequence Homology, Amino Acid, [SDV]Life Sciences [q-bio], Escherichia coli Proteins, Membrane Proteins, Viral Tail Proteins, Type VI Secretion Systems, Polymerization, lcsh:Genetics, Viral Proteins, Secretion systems, Contractile Proteins, Microscopy, Fluorescence, Multiprotein Complexes, Escherichia coli, Immunoprecipitation, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Bacteriophages, Membrane protein complexes, Cell fusion, ComputingMilieux_MISCELLANEOUS, Research Article
Abstract

The Type VI secretion system (T6SS) is a widespread weapon dedicated to the delivery of toxin proteins into eukaryotic and prokaryotic cells. The 13 T6SS subunits assemble a cytoplasmic contractile structure anchored to the cell envelope by a membrane-spanning complex. This structure is evolutionarily, structurally and functionally related to the tail of contractile bacteriophages. In bacteriophages, the tail assembles onto a protein complex, referred to as the baseplate, that not only serves as a platform during assembly of the tube and sheath, but also triggers the contraction of the sheath. Although progress has been made in understanding T6SS assembly and function, the composition of the T6SS baseplate remains mostly unknown. Here, we report that six T6SS proteins–TssA, TssE, TssF, TssG, TssK and VgrG–are required for proper assembly of the T6SS tail tube, and a complex between VgrG, TssE,-F and-G could be isolated. In addition, we demonstrate that TssF and TssG share limited sequence homologies with known phage components, and we report the interaction network between these subunits and other baseplate and tail components. In agreement with the baseplate being the assembly platform for the tail, fluorescence microscopy analyses of functional GFP-TssF and TssK-GFP fusion proteins show that these proteins assemble stable and static clusters on which the sheath polymerizes. Finally, we show that recruitment of the baseplate to the apparatus requires initial positioning of the membrane complex and contacts between TssG and the inner membrane TssM protein., Author Summary In the environment, bacteria compete for privileged access to nutrients or to a particular niche. Bacteria have therefore evolved mechanisms to eliminate competitors. Among them, the Type VI secretion system (T6SS) is a contractile machine functionally comparable to a crossbow: an inner tube is wrapped by a contractile structure. Upon contraction of this outer sheath, the inner tube is propelled towards the target cell and delivers anti-bacterial effectors. The tubular structure assembles on a protein complex called the baseplate. Here we define the composition of the baseplate, demonstrating that it is composed of five subunits: TssE, TssF, TssG, TssK and VgrG. We further detail the role of the TssF and TssG proteins by defining their localizations and identifying their partners. We show that, in addition to TssE and VgrG that have been shown to share homologies with the bacteriophage gp25 and gp27-gp5 proteins, the TssF and TssG proteins also have homologies with bacteriophage components. Finally, we show that this baseplate is recruited to the TssJLM membrane complex prior to the assembly of the contractile tail structure. This study allows a better understanding of the early events of the assembly pathway of this molecular weapon.

Published
2015

27. Inhibition of type VI secretion by an anti-TssM llama nanobody

Author

Nguyen, van Son, Logger, Laureen, Spinelli, Silvia, Desmyter, Aline, Hang Le, Thi Thu, Kellenberger, Christine, Douzi, Badreddine, Durand, Eric, Roussel, Alain, Cascales, E., Cambillau, Christian, Architecture et fonction des macromolécules biologiques (AFMB), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Institut National de la Recherche Agronomique (INRA), Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), JCJC-1303-03, Institut National de la Recherche Agronomique (INRA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS), and Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Models, Molecular, Protein Conformation, [SDV]Life Sciences [q-bio], Molecular Sequence Data, Antibody Affinity, STRUCTURAL BIOLOGY, PROTEIN, lcsh:Medicine, Bacterial Proteins, Antibody Specificity, [SDV.IDA]Life Sciences [q-bio]/Food engineering, Animals, [SPI.GPROC]Engineering Sciences [physics]/Chemical and Process Engineering, CRYSTAL-STRUCTURE, Amino Acid Sequence, SINGLE-DOMAIN ANTIBODIES, lcsh:Science, FRAGMENTS, lcsh:R, Type VI Secretion Systems, ENTEROAGGREGATIVE ESCHERICHIA-COLI, BACTERIAL TYPE VI, EFFECTORS, Thermodynamics, lcsh:Q, Camelids, New World, Sequence Alignment, SYSTEM, Protein Binding, Research Article
Abstract

International audience; The type VI secretion system (T6SS) is a secretion pathway widespread in Gram-negative bacteria that targets toxins in both prokaryotic and eukaryotic cells. Although most T6SSs identified so far are involved in inter-bacterial competition, a few are directly required for full virulence of pathogens. The T6SS comprises 13 core proteins that assemble a large complex structurally and functionally similar to a phage contractile tail structure anchored to the cell envelope by a trans-membrane spanning stator. The central part of this stator, TssM, is a 1129-amino-acid protein anchored in the inner membrane that binds to the TssJ outer membrane lipoprotein. In this study, we have raised camelid antibodies against the purified TssM periplasmic domain. We report the crystal structure of two specific nanobodies that bind to TssM in the nanomolar range. Interestingly, the most potent nanobody, nb25, competes with the TssJ lipoprotein for TssM binding in vitro suggesting that TssJ and the nb25 CDR3 loop share the same TssM binding site or causes a steric hindrance preventing TssM-TssJ complex formation. Indeed, periplasmic production of the nanobodies displacing the TssM-TssJ interaction inhibits the T6SS function in vivo. This study illustrates the power of nanobodies to specifically target and inhibit bacterial secretion systems.

Published
2015

28. EAEC T6SS TssA-Cterminus

Author

Durand, E., primary, Zoued, A., additional, Spinelli, S., additional, Douzi, B., additional, Brunet, Y.R., additional, Bebeacua, C., additional, Legrand, P., additional, Journet, L., additional, Mignot, T., additional, Cambillau, C., additional, and Cascales, E., additional

Published
2016
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29. Enteroaggregative Escherichia Coli TssA N-terminal fragment

Author

Durand, E., primary, Zoued, A., additional, Spinelli, S., additional, Douzi, B., additional, Brunet, Y.R., additional, Bebeacua, C., additional, Legrand, P., additional, Journet, L., additional, Mignot, T., additional, Cambillau, C., additional, and Cascales, E., additional

Published
2016
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31. Llama nanobody n02 raised against EAEC T6SS TssM

Author

Nguyen, V.S., primary, Desmyter, A., additional, Le, T.T.H., additional, Durand, E., additional, Kellenberger, C., additional, Douzi, B., additional, Spinelli, S., additional, Cascales, E., additional, Cambillau, C., additional, and Roussel, A., additional

Published
2015
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34. Nooks and crannies in type VI secretion regulation

Author

Bernard, C. S., Brunet, Y. R., Gueguen, Erwan, Cascales, E., Microbiologie, adaptation et pathogénie (MAP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), Trafic et signalisation membranaires chez les bactéries (MTSB), and Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL)

Subjects
[SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, [SDV]Life Sciences [q-bio], [SDV.EE.IEO]Life Sciences [q-bio]/Ecology, environment/Symbiosis
Published
2010

35. Agrobacterium tumefaciens oncogenic suppressors inhibit T-DNA and VirE2 protein substrate binding to the VirD4 coupling protein

Author

Cascales, E., Atmakuri, Krishnamohan, Liu, Zhenying, Binns, Andrew, Christie, Peter, Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU)

Subjects
DNA, Bacterial, Recombinant Fusion Proteins, [SDV]Life Sciences [q-bio], Genetic Complementation Test, Gene Expression Regulation, Bacterial, Article, Ion Channels, DNA-Binding Proteins, Plant Leaves, Bacterial Proteins, Agrobacterium tumefaciens, Genes, Tumor Suppressor, ComputingMilieux_MISCELLANEOUS, Plasmids, Protein Binding
Abstract

Agrobacterium tumefaciens uses a type IV secretion (T4S) system composed of VirB proteins and VirD4 to deliver oncogenic DNA (T-DNA) and protein substrates to susceptible plant cells during the course of infection. Here, by use of the Transfer DNA ImmunoPrecipitation (TrIP) assay, we present evidence that the mobilizable plasmid RSF1010 (IncQ) follows the same translocation pathway through the VirB/D4 secretion channel as described previously for the T-DNA. The RSF1010 transfer intermediate and the Osa protein of plasmid pSa (IncW), related in sequence to the FiwA fertility inhibition factor of plasmid RP1 (IncPalpha), render A. tumefaciens host cells nearly avirulent. By use of a semi-quantitative TrIP assay, we show that both of these 'oncogenic suppressor factors' inhibit binding of T-DNA to the VirD4 substrate receptor. Both factors also inhibit binding of the VirE2 protein substrate to VirD4, as shown by coimmunoprecipitation and bimolecular fluorescence complementation assays. Osa fused to the green fluorescent protein (GFP) also blocks T-DNA and VirE2 binding to VirD4, and Osa-GFP colocalizes with VirD4 at A. tumefaciens cell poles. RSF1010 and Osa interfere specifically with VirD4 receptor function and not with VirB channel activity, as shown by (i) TrIP and (ii) a genetic screen for effects of the oncogenic suppressors on pCloDF13 translocation through a chimeric secretion channel composed of the pCloDF13-encoded MobB receptor and VirB channel subunits. Our findings establish that a competing plasmid substrate and a plasmid fertility inhibition factor act on a common target, the T4S receptor, to inhibit docking of DNA and protein substrates to the translocation apparatus.

Published
2005

36. Deletion analyses of the peptidoglycan-associated lipoprotein Pal reveals three independent binding sequences including a TolA box

Author

Cascales, E., Lloubès, Roland, Laboratoire d'ingénierie des systèmes macromoléculaires (LISM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Aix Marseille Université (AMU), and Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
[SDV]Life Sciences [q-bio], ComputingMilieux_MISCELLANEOUS
Abstract

International audience

Published
2004

40. New teratological cases in Apoidea (Hymenoptera)

Author

Ornosa Gallego, Concepción, Cascales, E., Ortiz Sánchez, F.J., and Facultad de Biología

Subjects
5 - Ciencias puras y naturales::57 - Biología [CDU], Himenópteros, Teratología
Published
2001

41. A Genomic Island in Salmonella enterica ssp salamae Provides New Insights on the Genealogy of the Locus of Enterocyte Effacement

Author

Cascales, E, Chandry, PS, Gladman, S, Moore, SC, Seemann, T, Crandall, KA, Fegan, N, Cascales, E, Chandry, PS, Gladman, S, Moore, SC, Seemann, T, Crandall, KA, and Fegan, N

Abstract

The genomic island encoding the locus of enterocyte effacement (LEE) is an important virulence factor of the human pathogenic Escherichia coli. LEE typically encodes a type III secretion system (T3SS) and secreted effectors capable of forming attaching and effacing lesions. Although prominent in the pathogenic E. coli such as serotype O157:H7, LEE has also been detected in Citrobacter rodentium, E. albertii, and although not confirmed, it is likely to also be in Shigella boydii. Previous phylogenetic analysis of LEE indicated the genomic island was evolving through stepwise acquisition of various components. This study describes a new LEE region from two strains of Salmonella enterica subspecies salamae serovar Sofia along with a phylogenetic analysis of LEE that provides new insights into the likely evolution of this genomic island. The Salmonella LEE contains 36 of the 41 genes typically observed in LEE within a genomic island of 49, 371 bp that encodes a total of 54 genes. A phylogenetic analysis was performed on the entire T3SS and four T3SS genes (escF, escJ, escN, and escV) to elucidate the genealogy of LEE. Phylogenetic analysis inferred that the previously known LEE islands are members of a single lineage distinct from the new Salmonella LEE lineage. The previously known lineage of LEE diverged between islands found in Citrobacter and those in Escherichia and Shigella. Although recombination and horizontal gene transfer are important factors in the genealogy of most genomic islands, the phylogeny of the T3SS of LEE can be interpreted with a bifurcating tree. It seems likely that the LEE island entered the Enterobacteriaceae through horizontal gene transfer as a single unit, rather than as separate subsections, which was then subjected to the forces of both mutational change and recombination.

Published
2012

49. Differential CheR Affinity for Chemoreceptor C-Terminal Pentapeptides Modulates Chemotactic Responses.

Author

Velando F, Monteagudo-Cascales E, Matilla MA, and Krell T

Subjects
Binding Sites, Methyl-Accepting Chemotaxis Proteins metabolism, Methyl-Accepting Chemotaxis Proteins genetics, Phosphorylation, Histidine Kinase metabolism, Histidine Kinase genetics, Protein Binding, Methyltransferases, Chemotaxis, Bacterial Proteins metabolism, Bacterial Proteins genetics, Escherichia coli Proteins metabolism, Escherichia coli Proteins genetics, Escherichia coli metabolism, Escherichia coli genetics
Abstract

Many chemoreceptors contain a C-terminal pentapeptide at the end of a linker. In Escherichia coli, this pentapeptide forms a high-affinity binding site for CheR and phosphorylated CheB, and its removal interferes with chemoreceptor adaptation. Analysis of chemoreceptors revealed significant variation in their pentapeptide sequences, and bacteria often possess multiple chemoreceptors with differing pentapeptides. To assess whether this sequence variation alters CheR affinity and chemotaxis, we used Pectobacterium atrosepticum SCRI1043 as a model. SCRI1043 has 36 chemoreceptors, with 19 of them containing a C-terminal pentapeptide. We show that the affinity of CheR for the different pentapeptides varies up to 11-fold (K D 90 nM to 1 μM). Pentapeptides with the highest and lowest affinities differ only in a single amino acid. Deletion of the cheR gene abolishes chemotaxis. The replacement of the pentapeptide in the PacC chemoreceptor with those of the highest and lowest affinities significantly reduced chemotaxis to its cognate chemoeffector, L-Asp. Altering the PacC pentapeptide also reduced chemotaxis to L-Ser, but not to nitrate, which are responses mediated by the nontethered PacB and PacN chemoreceptors, respectively. Changes in the pentapeptide sequence thus modulate the response of the cognate receptor and that of another chemoreceptor., (© 2024 John Wiley & Sons Ltd.)

Published
2024
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50. Ubiquitous purine sensor modulates diverse signal transduction pathways in bacteria.

Author

Monteagudo-Cascales E, Gumerov VM, Fernández M, Matilla MA, Gavira JA, Zhulin IB, and Krell T

Subjects
Cyclic GMP metabolism, Cyclic GMP analogs & derivatives, Gene Expression Regulation, Bacterial, Bacteria metabolism, Bacteria genetics, Escherichia coli metabolism, Escherichia coli genetics, Second Messenger Systems, Purines metabolism, Signal Transduction, Bacterial Proteins metabolism, Bacterial Proteins genetics
Abstract

Purines and their derivatives control intracellular energy homeostasis and nucleotide synthesis, and act as signaling molecules. Here, we combine structural and sequence information to define a purine-binding motif that is present in sensor domains of thousands of bacterial receptors that modulate motility, gene expression, metabolism, and second-messenger turnover. Microcalorimetric titrations of selected sensor domains validate their ability to specifically bind purine derivatives, and evolutionary analyses indicate that purine sensors share a common ancestor with amino-acid receptors. Furthermore, we provide experimental evidence of physiological relevance of purine sensing in a second-messenger signaling system that modulates c-di-GMP levels., (© 2024. The Author(s).)

Published
2024
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