Identification of Tse8 as a Type VI secretion system toxin from Pseudomonas aeruginosa that targets the bacterial transamidosome to inhibit protein synthesis in prey cells

Abstract

The Type VI secretion system (T6SS) is a bacterial nanomachine which delivers toxic effectors to kill competitors or subvert some of their key functions. Here we use tra nsposon d irected i nsertion-site s equencing (TraDIS) 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 tRNA 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 proteins and validate the use of a TraDIS-based global genomics approach to expand the repertoire of T6SS toxins in T6SS-encoding bacteria.

Recruitment of Mobile Genetic Elements for Diverse Cellular Functions in Prokaryotes

“…but many function as toxins that exert anti-bacterial or anti-eukaryotic activity ( Cherrak et al, 2019 ). One such toxin apparently evolved from the phage tail tip protein (M. Iyer et al, 2021 ), whereas numerous other T6SS toxins possess cognate antitoxins ( Nolan et al, 2021 ), suggesting that TA systems were recruited for inter-species competition. Other proteins secreted via phage tails perform non-competitive roles ( Russell et al, 2014 ).…”

Core and accessory effectors of type VI secretion systems contribute differently to the intraspecific diversity of Pseudomonas aeruginosa

“…The detection of T6SS components in the sputum of CF patients and recent reports of T6SSmediated colonisation resistance to Burkholderia in the CF lung demonstrate the clinical relevance of this secretion system 4,20,21 . Each T6SS effector is (i) translocated by one of the three types of T6SSs (H1, H2, and H3) 9,[22][23][24][25][26][27][28] , (ii) associated with the secretion machinery by Hcp, VgrG or PAAR-domains [29][30][31][32] , and (iii) targeting nutrient uptake, prokaryotes and/or eukaryotic cells 1,9,14,25,33 . To our best knowledge, more T6SS effectors are known and characterised in depth for P. aeruginosa than for any other species.…”

A type IVB secretion system adapted for bacterial killing, biofilm invasion and biocontrol

“…We therefore reasoned that it should be possible to identify the genetic elements required for self-protection by transposon sequencing. This approach has previously been employed to identify E-I pairs of T6SSs in V. cholerae and P. aeruginosa 58,59 . To this end, we generated a saturated transposon insertion library in the IsoF wild type strain.…”