Gaming the competition in microbial cell–cell interactions

Abstract

Cell (2013) 152 4, 884–894. doi:; DOI: 10.1016/j.cell.2013.01.042

It helps to pay attention to an opponent’s prior actions when playing strategy games, be they chess, poker or rock-paper-scissors. In a thought provoking paper by Basler et al (2013), published in Cell, we have a chance to watch bacteria ‘game’ each other in a deadly match of life or death involving the deployment of a potent weapon called the Type VI Secretion System (T6SS; Basler et al, 2013). The results establish a paradigm for inter-bacterial competition based on defensive counterattack with temporal and spatial precision.

T6SS organelles are encoded by about a quarter of all Gram-negative bacteria, including pathogens, commensals and environmental organisms. Contributions to virulence and host-cell injury have been established for a handful of these (Pukatzki et al, 2006; French et al, 2011), but it has now become apparent that this secretion system frequently targets bacterial cells as well (Hood et al, 2010; MacIntyre et al, 2010). In this case, the system has been shown to kill ‘prey’ bacterial cells by forceful transfer of toxic effector proteins with muramindase or lipase activity (Russell et al, 2011; Dong et al, 2013). The T6SS organelle is a dynamic structure that is evolutionarily and functionally related to the contractile tails of bacteriophages (Figure 1), although it assembles in the cytoplasm and envelope of bacterial cells (Leiman et al, 2009; Basler et al, 2012). The firing of the apparatus correlates with contraction of a tail sheath-like structure and the sudden ejection of a phage tail tube-like structure tipped with a poison spike (Pukatzki et al, 2007; Basler et al, 2012). An ATPase called ClpV recognizes the contracted sheath and remodels it to re-cock the fired T6SS (Bonemann et al, 2009; Basler and Mekalanos, 2012). Biogenesis and cycling of the organelle is energetically expensive, so it is no surprise that expression and assembly are tightly regulated. Nonetheless, few of us would have guessed that the deployment of such a weapon might be regulated by the T6SS-mediated attack of a heterologous bacteria cell. In homage to Robert Axelrod’s treatise ‘The Evolution of Cooperation’ and the winning strategy of Anatol Rapoport for the ‘Prisoner’s Dilemma’ game (Axelrod and Hamilton, 1981), Mekalanos and colleagues entitled their paper ‘Tit-for-tat: Type VI secretion counterattack in bacterial cell-cell interactions’.

Figure 1.

A model for T6SS aiming by P. aeruginosa in a duel with V. cholerae. T6SS attack from V. cholerae is sensed by the P. aeruginosa TagQRST signal-transduction system (yellow), which activates the PpkA kinase to phosphorylate Fha1 (red box). Phosphorylated Fha1 directs assembly of a T6SS apparatus near the point of attack. Contraction of the outer sheath (violet) fires the inner tube/spike complex (dark green), positioning the counterattack at the point of contact with the aggressor cell. Effector proteins delivered by the T6SS, such as the Tse1 peptidoglycan amidase, contribute to destruction of the target V. cholerae. Disassembly of the fired apparatus facilitated by the ClpV ATPase and dephosphorylation of Fha1 allows redeployment to new sites.

Previously, Basler and Mekalanos (2012) reported a phenomenon they termed ‘T6SS duelling’, in which sister cells of Pseudomonas aeruginosa could be observed in pitched battles when a fluorescent ClpV–GFP fusion protein was used to monitor the position of T6SS organelle firing. No apparent harm comes to sister cells in such duelling behaviour, as they encode immunity proteins to their own T6SS effectors (Hood et al, 2010). However, their stunning videos suggested that spatial and temporal T6SS activity in one cell could induce corresponding T6SS activity in other cells in direct contact. These observations prompted the present study, which addresses the regulatory mechanism and biological consequences of what appears to be a strategy for prey selection during interspecies interactions.

In their most recent report, Basler et al (2013) found that P. aeruginosa efficiently kills two other bacterial species, Vibrio cholerae and Acinetobacter baylyi, only if these organisms also deploy functional T6SS organelles. Imaging cellular mixtures confirmed that T6SS-competent V. cholerae cells were targeted for delivery of the P. aeruginosa toxic effector protein Tse1 30-fold more frequently than T6SS-defective V. cholerae mutants, even when all three strains were mixed together at the same time. This suggested that P. aeruginosa is capable of assembling, aiming and firing its T6SS apparatus at aggressive V. cholerae, while sparing peaceful bystanders. The lack of collateral damage in the midst of ongoing battle reveals an unanticipated precision of the P. aeruginosa T6SS counterattack, begging a molecular explanation. Basler et al (2013) went on to show that a signal-transduction system, TagQRST–PpkA–PppA, is involved in sensing T6SS attack, presumably by detecting penetration of the P. aeruginosa cell surface. This membrane-associated complex was previously shown to mediate phosphorylation of a scaffold protein, Fha1, which controls T6SS organelle assembly (Mougous et al, 2007). It now appears that phosphorylated Fha1 also functions to position the T6SS apparatus at the precise point of engagement with an attacking bacterium. Furthermore, through cycles of phosphorylation and dephosphorylation, the signalling system facilitates spatial repositioning. Aiming T6SS counterattack in response to initial heterologous T6SS attack dramatically affects the accuracy of prey selection by P. aeruginosa. In contrast, other bacterial species such as V. cholerae and A. baylyi kill Escherichia coli K12 with their T6SS apparatus with apparently no requirement that this prey species is armed with a functional T6SS.

As with any paradigm-shifting study, the Basler et al (2013) report raises as many questions as it answers. How, exactly, does T6SS attack trigger the duelling counterattack response in P. aeruginosa? According to available data, the signal may be physical and positional (e.g., localized membrane perturbation) rather than enzymatic or chemical. What is the mechanism of T6SS organelle localization in P. aeruginosa, and how does spatially defined duelling differ, biologically and mechanistically, from the apparently random placement of sequential T6SS assembly sites by V. cholerae (Basler et al, 2012)? In a broader context, does the tit-for-tat response also characterize T6SS control in other bacterial species? As the ‘Prisoner’s Dilemma’ involves a defined reward system for which ‘tit-for-tat’ is the winning strategy, one wonders if evolutionary fitness is the reward P. aeruginosa receives by adopting this tactic in contrast to less discriminating (and more energetically costly) deployment of T6SS organelles?

Early results that contributed to this study were presented by John Mekalanos to a packed house at the President’s Forum of the 2012 General Meeting of the American Society for Microbiology, held in San Francisco last June. The tit-for-tat story was a highlight of the meeting and it clearly inspired many microbiologists to view bacterial cell–cell interactions in a new light. These discoveries point to a new mechanism that could determine, in part, the composition of the complex communities that make up the commensal microbiota, as well as microbial biofilms that characterize pathological environments such as the inflammatory milieu of the cystic fibrosis lung, acne pustules or the chronic inflammatory bowel. Pharmacological control of T6SS deployment may provide a new tool for controlling the composition of polymicrobial communities to promote health and mitigate disease.