An antibacterial T6SS in Pantoea agglomerans pv. betae delivers a lysozyme‐like effector to antagonize competitors

Abstract

The type VI secretion system (T6SS) is deployed by numerous Gramnegative bacteria to deliver toxic effectors into neighbouring cells. The genome of Pantoea agglomerans pv. betae (Pab) phytopathogenic bacteria contains a gene cluster (T6SS1) predicted to encode a complete T6SS. Using secretion and competition assays, we found that T6SS1 in Pab is a functional antibacterial system that allows this pathogen to outcompete rival plant-associated bacteria found in its natural environment. Computational analysis of the T6SS1 gene cluster revealed that antibacterial effector and immunity proteins are encoded within three genomic islands that also harbour arrays of orphan immunity genes or toxin and immunity cassettes. Functional analyses indicated that VgrG, a specialized antibacterial effector, contains a C-terminal catalytically active glucosaminidase domain that is used to degrade prey peptidoglycan. Moreover, we confirmed that a bicistronic unit at the end of the T6SS1 cluster encodes a novel antibacterial T6SS effector and immunity pair. Together, these results demonstrate that Pab T6SS1 is an antibacterial system delivering a lysozyme-like effector to eliminate competitors, and indicate that this bacterium contains additional novel T6SS effectors.Andrea Carobbi and Simone Di Nepi contributed equally to this work.

T6SS-mediated competitive exclusion amongPantoea agglomeransassociated with plants

“…Across all eleven strains, cluster 1 displays the architectural features observed by Carobbi et al (2022) [29] and Boyer et al (2009) [32] and can be divided into five major subset regions of genes. Cluster 1 starts with a SH3-domain-containing protein (similar to A7P61_RS17235 in P. agglomerans pv.…”

“…The pattern of gene presence and absence in cluster 1 furthermore revealed two variable regions that are typical for genomic loci encoding for effector and immunity gene pairs as previously described for P. agglomerans. Across all eleven strains, cluster 1 displays the architectural features observed by Carobbi et al (2022) [29] and Boyer et al (2009) [32] and can be divided into five major subset regions of genes. Cluster 1 starts with a SH3-domain-containing protein (similar to A7P61_RS17235 in P. agglomerans pv.…”

“…The repertoire of T6SS effector and immunity genes in Pantoea agglomerans strains Components of the T6SS have been previously described in several Pantoea species including P. agglomerans [28][29][30] and Pantoea ananatis [31]. Using the known genes as queries we identified gene clusters that putatively encode T6SS components and examined their conservation among eleven P. agglomerans strains isolated from seeds of wheat cultivars (strains A2, B1, B3, R1, and R5) and landraces (strains T1 to T6).…”

“…Here we number the T6SS clusters according to their gene content similarity to previous studies [28,29]. Most of the genes encoding the core T6SS components were conserved throughout the sampled set across three different genomic regions.…”

“…Our analysis of the T6SS cluster 1 led us to conclude that the cluster extends even beyond the effector/immunity gene pair pse2 / psi2 , which was set at the 3’ end of the T6SS cluster 1 in P. agglomerans pv. betae [29]. We identified downstream of the pse2 / psi2 effector/immunity gene pair several genes that we predict to be part of the cluster.…”