The tomato Prf complex is a molecular trap for bacterial effectors based on Pto transphosphorylation

The bacteria Pseudomonas syringae is a pathogen of many crop species and one of the model pathogens for studying plant and bacterial arms race coevolution. In the current model, plants perceive bacteria pathogens via plasma membrane receptors, and recognition leads to the activation of general defenses. In turn, bacteria inject proteins called effectors into the plant cell to prevent the activation of immune responses. AvrPto and AvrPtoB are two such proteins that inhibit multiple plant kinases. The tomato plant has reacted to these effectors by the evolution of a cytoplasmic resistance complex. This complex is compromised of two proteins, Prf and Pto kinase, and is capable of recognizing the effector proteins. How the Pto kinase is able to avoid inhibition by the effector proteins is currently unknown. Our data shows how the tomato plant utilizes dimerization of resistance proteins to gain advantage over the faster evolving bacterial pathogen. Here we illustrate that oligomerisation of Prf brings into proximity two Pto kinases allowing them to avoid inhibition by the effectors by transphosphorylation and to activate immune responses

Phevamine A, a small molecule that suppresses plant immune responses

Bacterial plant pathogens cause significant crop damage worldwide. They invade plant cells by producing a variety of virulence factors, including small-molecule toxins and phytohormone mimics. Virulence of the model pathogen Pseudomonas syringae pv. tomato DC3000 (Pto) is regulated in part by the sigma factor HrpL. Our study of the HrpL regulon identified an uncharacterized, three-gene operon in Pto that is controlled by HrpL and related to the Erwinia hrp-associated systemic virulence (hsv) operon. Here, we demonstrate that the hsv operon contributes to the virulence of Pto on Arabidopsis thaliana and suppresses bacteria-induced immune responses. We show that the hsv-encoded enzymes in Pto synthesize a small molecule, phevamine A. This molecule consists of L-phenylalanine, L-valine, and a modified spermidine, and is different from known small molecules produced by phytopathogens. We show that phevamine A suppresses a potentiation effect of spermidine and L-arginine on the reactive oxygen species burst generated upon recognition of bacterial flagellin. The hsv operon is found in the genomes of divergent bacterial genera, including ∼37% of P. syringae genomes, suggesting that phevamine A is a widely distributed virulence factor in phytopathogens. Our work identifies a small-molecule virulence factor and reveals a mechanism by which bacterial pathogens overcome plant defense. This work highlights the power of omics approaches in identifying important small molecules in bacteria–host interactions

Design of synthetic bacterial communities for predictable plant phenotypes

Specific members of complex microbiota can influence host phenotypes, depending on both the abiotic environment and the presence of other microorganisms. Therefore, it is challenging to define bacterial combinations that have predictable host phenotypic outputs. We demonstrate that plant–bacterium binary-association assays inform the design of small synthetic communities with predictable phenotypes in the host. Specifically, we constructed synthetic communities that modified phosphate accumulation in the shoot and induced phosphate starvation–responsive genes in a predictable fashion. We found that bacterial colonization of the plant is not a predictor of the plant phenotypes we analyzed. Finally, we demonstrated that characterizing a subset of all possible bacterial synthetic communities is sufficient to predict the outcome of untested bacterial consortia. Our results demonstrate that it is possible to infer causal relationships between microbiota membership and host phenotypes and to use these inferences to rationally design novel communities

Design of synthetic bacterial communities for predictable plant phenotypes

Specific members of complex microbiota can influence host phenotypes, depending on both the abiotic environment and the presence of other microorganisms. Therefore, it is challenging to define bacterial combinations that have predictable host phenotypic outputs. We demonstrate that plant-bacterium binary-association assays inform the design of small synthetic communities with predictable phenotypes in the host. Specifically, we constructed synthetic communities that modified phosphate accumulation in the shoot and induced phosphate starvation-responsive genes in a predictable fashion. We found that bacterial colonization of the plant is not a predictor of the plant phenotypes we analyzed. Finally, we demonstrated that characterizing a subset of all possible bacterial synthetic communities is sufficient to predict the outcome of untested bacterial consortia. Our results demonstrate that it is possible to infer causal relationships between microbiota membership and host phenotypes and to use these inferences to rationally design novel communitie

Phevamine A, a small molecule that suppresses plant immune responses

Bacterial plant pathogens cause significant crop damage worldwide. They invade plant cells by producing a variety of virulence factors, including small-molecule toxins and phytohormone mimics. Virulence of the model pathogen Pseudomonas syringae pv. tomato DC3000 (Pto) is regulated in part by the sigma factor HrpL. Our study of the HrpL regulon identified an uncharacterized, three-gene operon in Pto that is controlled by HrpL and related to the Erwinia hrp-associated systemic virulence (hsv) operon. Here, we demonstrate that the hsv operon contributes to the virulence of Pto on Arabidopsis thaliana and suppresses bacteria-induced immune responses. We show that the hsv-encoded enzymes in Pto synthesize a small molecule, phevamine A. This molecule consists of L-phenylalanine, L-valine, and a modified spermidine, and is different from known small molecules produced by phytopathogens. We show that phevamine A suppresses a potentiation effect of spermidine and L-arginine on the reactive oxygen species burst generated upon recognition of bacterial flagellin. The hsv operon is found in the genomes of divergent bacterial genera, including ∼37% of P. syringae genomes, suggesting that phevamine A is a widely distributed virulence factor in phytopathogens. Our work identifies a small-molecule virulence factor and reveals a mechanism by which bacterial pathogens overcome plant defense. This work highlights the power of omics approaches in identifying important small molecules in bacteria-host interactions

A complex immune response to flagellin epitope variation in commensal communities

Immune systems restrict microbial pathogens by identifying “non-self” molecules called microbe-associated molecular patterns (MAMPs). It is unclear how immune responses are tuned to or by MAMP diversity present in commensal microbiota. We systematically studied the variability of commensal peptide derivatives of flagellin (flg22), a MAMP detected by plants. We define substantial functional diversity. Most flg22 peptides evade recognition, while others contribute to evasion by manipulating immunity through antagonism and signal modulation. We establish a paradigm of signal integration, wherein the sequential signaling outputs of the flagellin receptor are separable and allow for reprogramming by commensal-derived flg22 epitope variants. Plant-associated communities are enriched for immune evading flg22 epitopes, but upon physiological stress that represses the immune system, immune-activating flg22 epitopes become enriched. The existence of immune-manipulating epitopes suggests that they evolved to either communicate or utilize the immune system for host colonization and thus can influence commensal microbiota community composition

Phevamine A, a small molecule that suppresses plant immune responses

Bacterial plant pathogens cause significant crop damage worldwide. They invade plant cells by producing a variety of virulence factors, including small-molecule toxins and phytohormone mimics. Virulence of the model pathogen Pseudomonas syringae pv. tomato DC3000 (Pto) is regulated in part by the sigma factor HrpL. Our study of the HrpL regulon identified an uncharacterized, three-gene operon in Pto that is controlled by HrpL and related to the Erwinia hrp-associated systemic virulence (hsv) operon. Here, we demonstrate that the hsv operon contributes to the virulence of Pto on Arabidopsis thaliana and suppresses bacteria-induced immune responses. We show that the hsv-encoded enzymes in Pto synthesize a small molecule, phevamine A. This molecule consists of L-phenylalanine, L-valine, and a modified spermidine, and is different from known small molecules produced by phytopathogens. We show that phevamine A suppresses a potentiation effect of spermidine and L-arginine on the reactive oxygen species burst generated upon recognition of bacterial flagellin. The hsv operon is found in the genomes of divergent bacterial genera, including ∼37% of P. syringae genomes, suggesting that phevamine A is a widely distributed virulence factor in phytopathogens. Our work identifies a small-molecule virulence factor and reveals a mechanism by which bacterial pathogens overcome plant defense. This work highlights the power of omics approaches in identifying important small molecules in bacteria–host interactions

Pseudomonas syringae Differentiates into Phenotypically Distinct Subpopulations During Colonization of a Plant Host

© 2016 Society for Applied Microbiology and John Wiley & Sons Ltd Bacterial microcolonies with heterogeneous sizes are formed during colonization of Phaseolus vulgaris by Pseudomonas syringae. Heterogeneous expression of structural and regulatory components of the P. syringae type III secretion system (T3SS), essential for colonization of the host apoplast and disease development, is likewise detected within the plant apoplast. T3SS expression is bistable in the homogeneous environment of nutrient-limited T3SS-inducing medium, suggesting that subpopulation formation is not a response to different environmental cues. T3SS bistability is reversible, indicating a non-genetic origin, and the T3SSHIGH and T3SSLOW subpopulations show differences in virulence. T3SS bistability requires the transcriptional activator HrpL, the double negative regulatory loop established by HrpV and HrpG, and may be enhanced through a positive feedback loop involving HrpA, the main component of the T3SS pilus. To our knowledge, this is the first example of phenotypic heterogeneity in the expression of virulence determinants during colonization of a non-mammalian host

A Novel Role for the TIR Domain in Association with Pathogen-Derived Elicitors

Plant innate immunity is mediated by Resistance (R) proteins, which bear a striking resemblance to animal molecules of similar function. Tobacco N is a TIR-NB-LRR R gene that confers resistance to Tobacco mosaic virus, specifically the p50 helicase domain. An intriguing question is how plant R proteins recognize the presence of pathogen-derived Avirulence (Avr) elicitor proteins. We have used biochemical cell fraction and immunoprecipitation in addition to confocal fluorescence microscopy of living tissue to examine the association between N and p50. Surprisingly, both N and p50 are cytoplasmic and nuclear proteins, and N's nuclear localization is required for its function. We also demonstrate an in planta association between N and p50. Further, we show that N's TIR domain is critical for this association, and indeed, it alone can associate with p50. Our results differ from current models for plant innate immunity that propose detection is mediated solely through the LRR domains of these molecules. The data we present support an intricate process of pathogen elicitor recognition by R proteins involving multiple subcellular compartments and the formation of multiple protein complexes