Abstract
The type VI secretion system (T6SS) is a widespread bacterial nanoweapon used for delivery of toxic proteins into cell targets and contributes to virulence, anti-inflammatory processes, and interbacterial competition. In the model phytopathogenic bacterium Pseudomonas syringae pv. tomato (Pst) DC3000, two T6SS gene clusters, HSI-I and HSI-II, were identified, but their functions remain unclear. We previously reported that hcp2, located in HSI-II, is involved in competition with enterobacteria and yeast. Here, we demonstrated that interbacterial competition of Pst DC3000 against several Gram-negative plant-associated bacteria requires mainly HSI-II activity. By means of a systematic approach using in-frame deletion mutants for each gene in the HSI-II cluster, we identified genes indispensable for Hcp2 expression, Hcp2 secretion and interbacterial competition ability. Deletion of PSPTO_5413 only affected growth in interbacterial competition assays but not Hcp2 secretion, which suggests that PSPTO_5413 might be a putative effector. Moreover, PSPTO_5424, encoding a putative σ54-dependent transcriptional regulator, positively regulated the expression of all three operons in HSI-II. Our discovery that the HSI-II gene cluster gives Pst DC3000 the ability to compete with other plant-associated bacteria could help in understanding a possible mechanism of how phytopathogenic bacteria maintain their ecological niches.
Highlights
Natural habitats do not always provide suitable conditions for microorganisms to survive; bacteria have evolved various secretion systems to adapt to changing environments by translocating effector proteins or DNA into the surrounding milieu or target cells (Costa et al, 2015)
Whether T6SS is required for Pst DC3000 to survive in its natural niche full of other plant-associated bacteria and how Hcp secretion island 1 (HSI-I) and HSIII gene clusters are involved in T6SS apparatus assembly and effector delivery in Pst DC3000 remain unknown
The GFP signal intensity was decreased to various degrees when the bacterial competitors D. dadantii CAS9, E. coli MG1655, Psph 1448a, Pss 61, Psta 11528, Pst T1, and X. euvesicatoria XvT147 were co-incubated with Pst DC3000 wild type and HSI-I versus incubation alone or co-incubation with HSI-II and HSI-I/II
Summary
Natural habitats do not always provide suitable conditions for microorganisms to survive; bacteria have evolved various secretion systems to adapt to changing environments by translocating effector proteins or DNA into the surrounding milieu or target cells (Costa et al, 2015). Functional and structural analyses indicated that T6SS could assemble into a tubular structure similar to a contractile phage tail, in which an Hcp inner tube capped by VgrG/PAAR spike proteins and wrapped by a VipA/B ( known as HsiB/C, or TssB/C) sheath is polymerized on a membrane-associated baseplate complex (Lossi et al, 2013). A conformational change in the baseplate complex stimulates sheath contraction, leading to propelling of an Hcp/VgrG/PAAR puncturing device through cell membranes for effector secretion (Basler et al, 2012; Shneider et al, 2013; Zoued et al, 2016). The phage tail-like structure is stabilized by a membrane core complex containing TssL, M, and J-like proteins, and this complex could function as a channel for inner tube passage after sheath contraction (Brunet et al, 2015). To quickly and efficiently respond to the environmental stimuli, T6SS in a subset of bacterial species can be regulated by threonine phosphorylation and posttranslational regulation via serine/threonine kinase PpkA, phosphatase PphA, and a forkhead-associated (FHA) domain-containing protein, Fha (Mougous et al, 2007; Lin et al, 2014)
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