Project description:Secretion systems are used as weapons by a variety of Gram-negative bacteria. Among them the Type VI Secretion System (T6SS) gained more interest throughout the last years. The system functions as a molecular nano-weapon: it is used in inter-kingdom competition by various bacteria to deliver toxic effectors in target cells. Here we describe the role of the T6SS in Photorhabdus laumondii subsp. laumondii strain DJC, an entomopathogenic biocontrol agent able to live in different environmental niches, such as in symbiosis with nematodes and in the rhizosphere on plant roots. Using bioinformatic and protein motif analyses we identified four T6SS gene clusters (T6SS-1, T6SS-2, T6SS-3 and T6SS-4) and multiple orphan T6SS related genes in the genome of P. laumondii. Furthermore, we highlighted 11 T6SS effector-immunity pairs, including three undescribed membrane disrupting effectors, each with putatively different antibacterial activities. By label-free mass spectrometry of P. laumondii wild type cells and respective T6SS-deficient strains, we could point out a cross-link between T6SS and other Photorhabdus’ virulence related mechanisms such as PVCs, T3SS and pyocins. Furthermore, a change in motility as well as in the secondary metabolism was observed upon T6SS-deficiency. Here, we shed light on the T6SS in P. laumondii DJC and suggesting a cross-link of various virulence mechanisms, which could help to gain knowledge on T6SS and better figure out the Photorhabdus ability to live in polymicrobial environments.
Project description:In Gram-negative bacteria, resistance-nodulation-division (RND)-type effluxeffluxpumps, particularly AcrAB-TolC, play a critical role in mediating resistance to antimicrobial agents and toxic metabolites, contributing to multidrug resistance. Photorhabdus laumondii is an entomopathogenic bacterium that has garnered significant interest due to its production of bioactive specialized metabolites with anti-inflammatory, antimicrobial, and scavenger deterrent properties. In previous work, we demonstrated that AcrAB confers self-resistance to stilbenes in P. laumondii TT01. Here, we explore the pleiotropic effects of AcrAB in this bacterium. RNA sequencing of ΔacrA compared to wild type revealed growth-phase-specific gene regulation, with stationary-phase cultures showing significant downregulation of genes involved in stilbenes, fatty acid, and anthraquinone pigment biosynthesis, as well as genes related to cellular clumping and fimbrial pilin formation. Genes encoding putative LuxR regulators, type VI secretion systems, two-partner secretion systems, and contact-dependent growth inhibition systems were upregulated in ΔacrA. Additionally, exponential-phase cultures revealed reduced expression of genes related to motility in ΔacrA. The observed transcriptional changes were consistent with phenotypic assays, demonstrating that the ΔacrA mutant had altered bioluminescence and defective orange pigmentation due to disrupted anthraquinone production. These findings confirm the role of stilbenes as signaling molecules involved in gene expression, thereby shaping these phenotypes. Furthermore, we showed that AcrAB contributes to swarming and swimming motilities independently of stilbenes. Collectively, these results highlight that disrupting acrAB causes transcriptional and metabolic dysregulation in P. laumondii, likely by impeding the export of key signaling molecules such as stilbenes, which may serve as a ligand for global transcriptional regulators.
