Molecular Dissection of the Interface between the Type VI Secretion TssM Cytoplasmic Domain and the TssG Baseplate Component.
The type VI secretion system (T6SS) is a multiprotein complex that catalyses toxin secretion through the bacterial cell envelope of various Gram-negative bacteria including important human pathogens. This machine uses a bacteriophage-like contractile tail to puncture the prey cell and inject harmful toxins. The T6SS tail comprises an inner tube capped by the cell-puncturing spike and wrapped by the contractile sheath. This structure is built on an assembly platform, the baseplate, which is anchored to the bacterial cell envelope by the TssJLM membrane complex (MC). This MC serves as both a tail docking station and a channel for the passage of the inner tube. The TssM transmembrane protein is a key component of the MC as it connects the inner and outer membranes. In this study, we define the TssM topology, highlighting a large but poorly studied 35-kDa cytoplasmic domain, TssMCyto, located between two transmembrane segments. Protein-protein interaction assays further show that TssMCytooligomerises and makes contacts with several baseplate components. Using computer predictions, we delineate two subdomains in TssMCyto, including a nucleotide triphosphatase (NTPase) domain, followed by a 110-aa extension. Finally, site-directed mutagenesis coupled to functional assays reveals the contribution of these subdomains and conserved motifs to the interaction with T6SS partners and to the function of the secretion apparatus.
Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Aix-Marseille Université, CNRS - UMR 7255, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France.
Laureen
L
Logger
Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Aix-Marseille Université, CNRS - UMR 7255, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France.
Marie-Stéphanie
MS
Aschtgen
Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Aix-Marseille Université, CNRS - UMR 7255, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France.
Marie
M
Guérin
Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Aix-Marseille Université, CNRS - UMR 7255, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France. Electronic address: cascales@imm.cnrs.fr.
Eric
E
Cascales
Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Aix-Marseille Université, CNRS - UMR 7255, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France. Electronic address: edurand@imm.cnrs.fr.
Eric
E
Durand
D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D025941:Protein Interaction Mapping; D055503:Protein Multimerization; D000069376:Type VI Secretion Systems / Q000378:metabolism*
D029968:Escherichia coli Proteins; D008565:Membrane Proteins; C000620726:TssG protein, E coli; C000600302:TssM protein, E coli; D000069376:Type VI Secretion Systems
bacterial competition; membrane complex; protein secretion; protein transport; type VI secretion