Our team aims to understand the assembly and functioning of two macrocomplexes conserved in the envelope of Gram negative bacteria and involved in cell adaptation and survival: the Tol and Ton systems.

Both Tol and Ton systems harness the energy produced from the proton motive force generated at the inner membrane to transduce it at the outer membrane. These complexes are involved in functions that are essential for bacterial survival. The Ton system mediates the active uptake across the outer membrane of metal, cyanocobalamin and carbohydrates. The Tol system has a more complex function, as it is involved in outer membrane integrity, in the correct localization of some polar factors and chemoreceptors, in outer-membrane phospholipid homeostasis, and in the constriction of the outer membrane during cell division.

Both Tol and Ton systems can be parasitized to open a gate for bacteriotoxins (called colicins), which result in bacterial cell death. These systems are also the target of filamentous phages during the infection process, such as the M13 phage targeting E. coli, and the CTX phage which is specific of the human pathogen Vibrio cholerae.

To obtain an integrate view of the inner membrane Tol and Ton complexes, we developpe an interdisciplinary study in collaboration with different laboratories, mixing genetic, biochemical and microscopy techniques.

1 – Structural and functional analysis of the Tol and Ton complexes.

Structural and functional studies on the Tol and Ton complexes isolated from various Gram negative bacteria are investigated in collaboration with international teams. More specifically, we are interested in the interactions occuring between the complex subunits in vivo, the structural study of soluble domain by NMR, the structural and dynamic studies of the complexes by EPR and the high-resolution structure of the membrane complexes.

The first structure/function results of such motor have been obtained with the E. coli Ton membrane complex. We have characterized the channels formed by the ExbB, ExbB/ExbD complexes and obtained the X-ray and cryo-EM high resolution structures of the ExbB/ExbD subcomplex.

2 – Dynamic study of the Tol-Pal proteins

We have started to develop the in vivo imaging techniques of the Tol-Pal proteins in different background strains using fluorescence microscopy. In collaboration, we use high resolution fluorescent microscopy and microfluidic device to follow the dynamic of the Tol-Pal proteins. The analyses are performed in relation with cell division, environmental conditions and using specific mutations.

3 – Involvement of the Tol-Pal machinery in the adaptation and the pathogenic conversion of Vibrio cholerae

The Tol system is the entrance gate for a family of bacterial viruses named filamentous phages. For some bacterial species such as V. cholerae, infection by these phages can convert the bacterium to a pathogen.

In this project, we aim identify the determinants of phage-host specificity and to identify the sequence of molecular events involved during phage transport across the periplasm. The expected results will shed new light on the mechanism of phage-dependent pathogenic conversion in bacteria, and open new perspectives in the engineering of hybrid phages designed to deliver molecules of interest in bacteria.

Members

Past Members / Alumni

Technical Staff
  • Bastien SERRANNO (Engineer, 2015-2017)
  • Hervé CELIA (Engineer CNRS, 2008-2018), now at NIDDK, NIH ()
  • Marthe GAVIOLI (TCE, retired in 2010)
Ph.D students
  • Mélissa PETITI (2015-2018) – Assemblage et dynamique in vivo du système Tol-Pal
  • Xiang ZHANG (2007-2010) – Organisation fonctionnelle d’un moteur moléculaire Tol et d’une protéine active contre une toxine bactérienne
  • Nadir SEDDIKI (codirection Dr Susan Buchanan, NIH, Bethesda) (2007-2010) – Import of macromolecules: Structural studies of the pesticin toxin and of an engineered variant
  • Aurélie Barnéoud-Arnoulet (2007-2010)- Mécanismes d’importation des collicines: détournement des fonctions physiologiques des protéines FtsH et FkpA
  • Emilie GOEMAERE (2004-2008) – Le complexe membranaire Tol d’E.coli: un modèle pour l’étude de l’organisation fonctionnelle d’un moteur moléculaire bacterien
  • Stephanie POMMIER (2002-2005) – Le système Tol-Pal d’E. coli
Post-Doctorants
  • Laetitia HOUOT (2010-2012)
  • Xiang ZHANG (2011)
Master 2
  • Ambre MOREAU (2020)- Mécanisme moléculaire d’infection par les phages filamenteux: réseau d’interactions protéiques mis en jeu.
  • Melvin RENAULT (2019) – Etude de la localisation des moteurs Tol et Ton et de l’interchangeabilité des moteurs chez E. coli.
  • Poutoum SAMIRE (2018) – Importance du moteur énergisé TolQRA et de la flexibilité de TolA dans l’import des phages filamenteux.
  • Nicolas GOMEZ (2016) – Analyse fonctionnelle du système Tol-Pal en fonction de l’état redox de TolA
  • Laura FRADALE (2015) – Analyse transcriptionnelle et fonctionnelle du système Tol-Pal chez Vibrio cholera
  • Melissa PETITI (2014) – Etude du système Tol-Pal par mutagénèse dirigée et double hybride bactérien
  • Perinne VAN OVERTVELT (2011)
  • Xiang ZHANG (2007)
  • Aurélie BARNEOUD-ARNOULET (2007)
Master 1
  • Megan CHARLES-ALFRED (2019)
  • Emeline LEMARIE (2019)
  • Melvin RENAULT (2018)
  • Poutoum SAMIRE (2017)
  • Raphaël RACHEDI (2017)
  • Pauline BACCELLI (2014)
  • Marie DESCAMPS (2014)
  • Alexandra LIPSKI (2006)
Undergraduates
  • Jennifer GONCALVES DA SILVA (UE Labo in vivo, L2, 2019)
  • Myriam BOUSSOUFA (UE Labo in vivo, L2, 2019)
  • Defne AKSOY (UE Labo in vivo, L2, 2019)
  • Celia JONAS (UE Labo in vivo, L2, 2019)
  • Victoria SGOLUPPI, L3 (2018)
  • Faniry RABOANARIVOLA, L3 (2017)
  • Jad MOUALLEM, UE Labo in vivo L1 (2017)
  • Megan CHARLES-ALFRED, L3 (2017)
  • Ahmed NASRAOUI, UE Labo in vivo L1 (2016)
  • Clara FERNANDEZ, L2 (2016)
  • Laura BASSET, ESIL-Polytech (2015)
  • Marie-Sarah ROUVIERE, UE Labo in vivo L1 (2015)
  • Axel DEJARDIN, BTS (2014, 2015)
  • Lia FRIEIER, ERASMUS L3 (2013)
  • Audrey FAVARO, ESIL-Polytech (2012)
  • Clement NOEL, ESIL-Polytech (2012)
  • Stéphanie ROLLERO, ESIL-Polytech (2010)
  • Sarah DOCHE, ESIL-Polytech (2009)
  • Nathalie MIARD, INSA Toulouse (2007)
  • Patrick ROCKENFELLER, ERASMUS (2005)

Publications

2019
Connected partner-switches control the life style of Pseudomonas aeruginosa through RpoS regulation

Sophie Bouillet, Moly Ba, Laetitia Houot, Chantal Iobbi-Nivol, Christophe Bordi

Scientific Reports 9 (2019)10.1038/s41598-019-42653-5

2019
Tol Energy-Driven Localization of Pal and Anchoring to the Peptidoglycan Promote Outer-Membrane Constriction

Melissa Petiti, Bastien Serrano, Laura Faure, Roland Lloubes, Tam Mignot, Denis Duché

Journal of Molecular Biology 431:3275-3288 (2019)10.1016/j.jmb.2019.05.039

Show complete publications list

2019
Decoupling filamentous phage uptake and energy of the TolQRA motor in Escherichia coli .

Poutoum Samire, Bastien Serrano, Denis Duché, Emeline Lemarie, Roland Lloubes, Laetitia Houot

Journal of Bacteriology (2019)10.1128/JB.00428-19

2017
Electrostatic interactions between the CTX phage minor coat protein and the bacterial host receptor TolA drive the pathogenic conversion of Vibrio cholerae

Laetitia Houot, Romain Navarro, Matthieu Nouailler, Denis Duché, Francoise Guerlesquin, Roland Lloubès

Journal of Biological Chemistry 292:13584-13598 (2017)10.1074/jbc.M117.786061

2016
(1)H, (15)N and (13)C resonance assignments of the C-terminal domain of Vibrio cholerae TolA protein.

Romain Navarro, Olivier Bornet, Laetitia Houot, Roland Lloubes, Francoise Guerlesquin, Matthieu Nouailler

Biomol NMR Assign (2016)10.1007/s12104-016-9690-y

2015
Virulence from vesicles: Novel mechanisms of host cell injury by Escherichia coli O104:H4 outbreak strain

Lisa Kunsmann, Christian Rüter, Andreas Bauwens, Lilo Greune, Malte Glüder, Björn Kemper, Angelika Fruth, Sun Nyunt Wai, Xiaohua He, Roland Lloubes, M. Alexander Schmidt, Ulrich Dobrindt, Alexander Mellmann, Helge Karch, Martina Bielaszewska

Scientific Reports 1-18 (2015)10.1038/srep13252

2013
Non classical secretion systems.

Roland Lloubes, Alain Bernadac, Laetitia Houot, Stéphanie Pommier

Res. Microbiol. 164:655-63 (2013)10.1016/j.resmic.2013.03.015

2013
Mannitol and the mannitol-specific enzyme IIB subunit activate Vibrio cholerae biofilm formation.

Patrick Ymele-Leki, Laetitia Houot, Paula I Watnick

Applied and Environmental Microbiology 79:4675-83 (2013)10.1128/AEM.01184-13

2012
Characterization of colicin M and its orthologs targeting bacterial cell wall peptidoglycan biosynthesis.

Hélène Barreteau, Meriem El Ghachi, Aurélie Barnéoud-Arnoulet, Emmanuelle Sacco, Thierry Touzé, Denis Duché, Fabien Gérard, Mark Brooks, Delphine Patin, Ahmed Bouhss, Didier Blanot, Herman Tilbeurgh, Michel Arthur, Roland Lloubes, Dominique Mengin-Lecreulx

Microb. Drug Resist. 18:222-9 (2012)10.1089/mdr.2011.0230

2012
A bacterial two-hybrid genome fragment library for deciphering regulatory networks of the opportunistic pathogen Pseudomonas aeruginosa.

Laetitia Houot, Adeline Fanni, Sophie Bentzmann, Christophe Bordi

Microbiology (Reading, Engl.) 158:1964-71 (2012)10.1099/mic.0.057059-0

2012
Energetics of colicin import revealed by genetic cross-complementation between the Tol and Ton systems.

Roland Lloubes, Emilie Goemaere, Xiang Zhang, E. Cascales, Denis Duché

Biochem. Soc. Trans. 40:1480-5 (2012)10.1042/BST20120181

2012
Colicin M, a peptidoglycan lipid-II-degrading enzyme: potential use for antibacterial means?

Thierry Touzé, Hélène Barreteau, Meriem El Ghachi, Ahmed Bouhss, Aurélie Barnéoud-Arnoulet, Delphine Patin, Emmanuelle Sacco, Didier Blanot, Michel Arthur, Denis Duché, Roland Lloubes, Dominique Mengin-Lecreulx

Biochem. Soc. Trans. 40:1522-7 (2012)10.1042/BST20120189

2012
The C-tail anchored TssL subunit, an essential protein of the enteroaggregative Escherichia coli Sci-1 Type VI secretion system, is inserted by YidC.

Abdelrahim Zoued, Roland Lloubes, Laure Journet, E. Cascales, Marie-Stéphanie Aschtgen

MicrobiologyOpen 1:71-82 (2012)10.1002/mbo3.9

2011
Regulation of type VI secretion gene clusters by sigma54 and cognate enhancer binding proteins.

Christophe S Bernard, Yannick R Brunet, Marthe Gavioli, Roland Lloubes, E. Cascales

Journal of Bacteriology 193:2158-67 (2011)10.1128/JB.00029-11

2011
An epigenetic switch involving overlapping fur and DNA methylation optimizes expression of a type VI secretion gene cluster

Yannick R Brunet, Christophe S Bernard, Marthe Gavioli, Roland Lloubes, E. Cascales

PLoS Genetics 7:e1002205 (2011)10.1371/journal.pgen.1002205

2011
The anaerobe-specific orange protein complex of Desulfovibrio vulgaris hildenborough is encoded by two divergent operons coregulated by ?54 and a cognate transcriptional regulator.

Anouchka Fiévet, Laetitia My, E. Cascales, Mireille Ansaldi, Sofia R Pauleta, Isabel Moura, Zorah Dermoun, Christophe S Bernard, Alain Dolla, Corinne Aubert

Journal of Bacteriology 193:3207-19 (2011)10.1128/JB.00044-11

2011
Mapping the interactions between Escherichia coli TolQ transmembrane segments.

Xiang Y-Z Zhang, Emilie L Goemaere, Nadir Seddiki, Hervé Celia, Marthe Gavioli, E. Cascales, Roland Lloubes

Journal of Biological Chemistry 286:11756-64 (2011)10.1074/jbc.M110.192773

2010
Toxicity of the Colicin M Catalytic Domain Exported to the Periplasm Is FkpA Independent

Aurélie Barnéoud-Arnoulet, Hélène Barreteau, Thierry Touze, Dominique Mengin-Lecreulx, Roland Lloubes, Denis Duché

Journal of Bacteriology 192:5212-5219 (2010)10.1128/JB.00431-10

2010
Deciphering the Catalytic Domain of Colicin M, a Peptidoglycan Lipid II-degrading Enzyme

Hélène Barreteau, Ahmed Bouhss, Fabien Gérard, Denis Duché, Boubekeur Boussaid, Didier Blanot, Roland Lloubes, Dominique Mengin-Lecreulx, Thierry Touze

Journal of Biological Chemistry 285:12378-12389 (2010)10.1074/jbc.M109.093583

2010
Determinants of the Proton Selectivity of the Colicin A Channel

Stephen Slatin, Daniel Baty, Denis Duché

Biochemistry 49:4786-4793 (2010)10.1021/bi100122g

2010
Channel Domain of Colicin A Modifies the Dimeric Organization of Its Immunity Protein

Xiang Y-Z Zhang, Roland Lloubes, Denis Duché

Journal of Biological Chemistry 285:38053-38061 (2010)10.1074/jbc.M110.144071

2008
Immunity Protein Protects Colicin E2 from OmpT Protease

Denis Duché, Mohamed Issouf, Roland Lloubes

Journal of Biochemistry 145:95-101 (2008)10.1093/jb/mvn149

2007
Colicin biology.

E. Cascales, Susan Buchanan, Denis Duché, Colin Kleanthous, Roland Lloubes, Kathleen Postle, Margaret Riley, Stephen Slatin, Danièle Cavard

Microbiology and Molecular Biology Reviews 71:158-229 (2007)10.1128/MMBR.00036-06

2006
Release of Immunity Protein Requires Functional Endonuclease Colicin Import Machinery

Denis Duché, Aurélie Frenkian, Valérie Prima, Roland Lloubes

Journal of Bacteriology 188:8593-8600 (2006)10.1128/JB.00941-06

2005
Combining high-field EPR with site-directed spin labeling reveals unique information on proteins in action

K. Möbius, A. Savitsky, C. Wegener, M. Plato, M. Fuchs, A. Schnegg, A. Dubinskii, Y. Grishin, I. Grigor'Ev, M. Kühn, Denis Duché, H. Zimmermann, H.-J. Steinhoff

Magnetic Resonance in Chemistry 43:S4-S19 (2005)10.1002/mrc.1690

2004
Spontaneous Refolding of the Pore-Forming Colicin A Toxin upon Membrane Association As Studied by X-Band and W-Band High-Field Electron Paramagnetic Resonance Spectroscopy †

Anton Savitsky, Martin Kühn, Denis Duché, Klaus Möbius, Heinz-Jürgen Steinhoff

Journal of Physical Chemistry B 108:9541-9548 (2004)10.1021/jp036397l

2004
Gating Movements of Colicin A and Colicin Ia Are Different

S.L. Slatin, Denis Duché, P.K. Kienker, D. Baty

Journal of Membrane Biology 202:73-83 (2004)10.1007/s00232-004-0720-9

2002
Analysis of the Escherichia coli Tol-Pal and TonB systems by periplasmic production of Tol, TonB, colicin, or phage capsid soluble domains.

Emmanuelle Bouveret, Laure Journet, Anne Walburger, E. Cascales, Hélène Bénédetti, Roland Lloubes

Biochimie 84:413-21 (2002)

2001
The C-terminal half of the colicin A pore-forming domain is active in vivo and in vitro11Edited by I. B. Holland

Angèle Nardi, Stephen Slatin, Daniel Baty, Denis Duché

Journal of Molecular Biology 307:1293-1303 (2001)10.1006/jmbi.2001.4524

1999
Integration of the colicin A pore-forming domain into the cytoplasmic membrane of Escherichia coli 1 1Edited by I. B. Holland

Denis Duché, Yves Corda, Vincent Géli, Daniel Baty

Journal of Molecular Biology 285:1965-1975 (1999)10.1006/jmbi.1998.2423

1998
Intracellular Immunization of Prokaryotic Cells against a Bacteriotoxin

Patrick Chames, Jacques Fieschi, Daniel Baty, Denis Duché

Journal of Bacteriology 180:514-518 (1998)10.1128/JB.180.3.514-518.1998

1995
Quantification of group A colicin import sites.

Denis Duché, L. Letellier, V. Geli, H. BÉnÉdetti, D. Baty

Journal of Bacteriology 177:4935-4939 (1995)10.1128/jb.177.17.4935-4939.1995

1994
1994
The role of electrostatic charge in the membrane insertion of colicin A. Calculation and mutation

Jeremy Lakey, Michael Parker, J González-Mañas, Denis Duché, Gert Vriend, Daniel Baty, Franc Pattus, Juan Gonzalez-Manas, Denis Duché

European Journal of Biochemistry 220:155-163 (1994)10.1111/j.1432-1033.1994.tb18610.x

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Collaborations

  • National
    • T. Mignot, LCB UMR7283, Marseille
  • International
    • S. Buchanan, NIDDK, NIH, Bethesda, MD, USA

Funding

  • ANR MeMox – Projet ANR- 18-CE11-0027 (2018-2021)
  • ANR BacMolMot – Projet ANR-14-CE09-0023 (2014-2017)
  • FRM Postdoctoral fellowship (2010-2012)
  • ANR Sodatol – Projet ANR- 07-BLAN-067

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