Maria A Sainz-Polo, Julian Parkhill, Amy K. Cain, Laura M. Nolan, Alain Filloux, Despoina A. I. Mavridou, Gordon Dougan, David Albesa-Jové, R. Christopher D. Furniss, Thomas Clamens, Eleni Manoli, Medical Research Council (UK), European Commission, Biotechnology and Biological Sciences Research Council (UK), Ministerio de Economía y Competitividad (España), Fundación Biofísica Bizkaia, Eusko Jaurlaritza, Cain, Amy K., Furniss, R. Christopher D., Albesa-Jové, David, Parkhill, Julian, Mavridou, Despoina A. I., Filloux, Alain, Imperial College London, Cain, Amy K [0000-0002-4230-6572], Furniss, R Christopher D [0000-0002-5806-5099], Albesa-Jové, David [0000-0003-2904-8203], Parkhill, Julian [0000-0002-7069-5958], Mavridou, Despoina A I [0000-0002-7449-1151], Filloux, Alain [0000-0003-1307-0289], Apollo - University of Cambridge Repository, and Mavridou, Despoina AI [0000-0002-7449-1151]
The Type VI secretion system (T6SS) is a bacterial nanomachine that delivers toxic effectors to kill competitors or subvert some of their key functions. Here, we use transposon directed insertion-site sequencing to identify T6SS toxins associated with the H1-T6SS, one of the three T6SS machines found in Pseudomonas aeruginosa. This approach identified several putative toxin-immunity pairs, including Tse8-Tsi8. Full characterization of this protein pair demonstrated that Tse8 is delivered by the VgrG1a spike complex into prey cells where it targets the transamidosome, a multiprotein complex involved in protein synthesis in bacteria that lack either one, or both, of the asparagine and glutamine transfer RNA synthases. Biochemical characterization of the interactions between Tse8 and the transamidosome components GatA, GatB and GatC suggests that the presence of Tse8 alters the fine-tuned stoichiometry of the transamidosome complex, and in vivo assays demonstrate that Tse8 limits the ability of prey cells to synthesize proteins. These data expand the range of cellular components targeted by the T6SS by identifying a T6SS toxin affecting protein synthesis and validate the use of a transposon directed insertion site sequencing-based global genomics approach to expand the repertoire of T6SS toxins in T6SS-encoding bacteria., L.M.N. was supported by Medical Research Council (MRC) Grant MR/N023250/1 and a Marie Curie Fellowship (PIIF-GA-2013-625318). A.F. was supported by MRC Grants MR/K001930/1 and MR/N023250/1 and Biotechnology and Biological Sciences Research Council (BBSRC) Grant BB/N002539/1. R.C.D.F. and D.A.I.M. were supported by the MRC Career Development Award MR/M009505/1. D.A.-J. acknowledges support by the MINECO Contract CTQ2016-76941-R, Fundación Biofísica Bizkaia, the Basque Excellence Research Centre (BERC) program and IT709-13 of the Basque Government, and Fundación BBVA. M.A.S.-P. was supported by the MINECO under the “Juan de la Cierva Postdoctoral program” (position FJCI-2015-25725).