scholarly journals Genome context influences evolutionary flexibility of nearly identical type III effectors in two phytopathogenic Pseudomonads

2021 ◽  
Author(s):  
David A Baltrus ◽  
Qian Feng ◽  
Brian H Kvitko
Keyword(s):  
Pseudomonas Syringae ◽  
Evolutionary Dynamics ◽  
Effector Proteins ◽  
In Planta ◽  
Genomic Context ◽  
Type Iii Effectors ◽  
Type Iii ◽  
Genome Context ◽  
The Impact ◽  
Multiple Type

Integrative Conjugative Elements (ICEs) are replicons that can insert and excise from chromosomal locations in a site specific manner, can conjugate across strains, and which often carry a variety of genes useful for bacterial growth and survival under specific conditions. Although ICEs have been identified and vetted within certain clades of the agricultural pathogen Pseudomonas syringae, the impact of ICE carriage and transfer across the entire P. syringae species complex remains underexplored. Here we identify and vet an ICE (PmaICE-DQ) from P. syringae pv. maculicola ES4326, a strain commonly used for laboratory virulence experiments, demonstrate that this element can excise and conjugate across strains, and contains loci encoding multiple type III effector proteins. Moreover, genome context suggests that another ICE (PmaICE-AOAB) is highly similar in comparison with and found immediately adjacent to PmaICE-DQ within the chromosome of strain ES4326, and also contains multiple type III effectors. Lastly, we present passage data from in planta experiments that suggests that genomic plasticity associated with ICEs may enable strains to more rapidly lose type III effectors that trigger R-gene mediated resistance in comparison to strains where nearly isogenic effectors are not present in ICEs. Taken together, our study sheds light on a set of ICE elements from P. syringae pv. maculicola ES4326 and highlights how genomic context may lead to different evolutionary dynamics for shared virulence genes between strains.

scholarly journals Multiple Xanthomonas euvesicatoria Type III Effectors Inhibit flg22-Triggered Immunity

2016 ◽  
Vol 29 (8) ◽  
pp. 651-660 ◽  
Author(s):  
Georgy Popov ◽  
Malou Fraiture ◽  
Frederic Brunner ◽  
Guido Sessa
Keyword(s):  
Pseudomonas Syringae ◽  
Map Kinases ◽  
Molecular Mechanisms ◽  
Inducible Promoter ◽  
Effector Proteins ◽  
Host Cells ◽  
In Planta ◽  
Callose Deposition ◽  
Type Iii ◽  
Xanthomonas Euvesicatoria

Xanthomonas euvesicatoria is the causal agent of bacterial spot disease in pepper and tomato. X. euvesicatoria bacteria interfere with plant cellular processes by injecting effector proteins into host cells through the type III secretion (T3S) system. About 35 T3S effectors have been identified in X. euvesicatoria 85-10, and a few of them were implicated in suppression of pattern-triggered immunity (PTI). We used an Arabidopsis thaliana pathogen-free protoplast–based assay to identify X. euvesicatoria 85-10 effectors that interfere with PTI signaling induced by the bacterial peptide flg22. Of 33 tested effectors, 17 inhibited activation of a PTI-inducible promoter. Among them, nine effectors also interfered with activation of an abscisic acid–inducible promoter. However, effectors that inhibited flg22-induced signaling did not affect phosphorylation of mitogen-activated protein (MAP) kinases acting downstream of flg22 perception. Further investigation of selected effectors revealed that XopAJ, XopE2, and XopF2 inhibited activation of a PTI-inducible promoter by the bacterial peptide elf18 in Arabidopsis protoplasts and by flg22 in tomato protoplasts. The effectors XopF2, XopE2, XopAP, XopAE, XopH, and XopAJ inhibited flg22-induced callose deposition in planta and enhanced disease symptoms caused by attenuated Pseudomonas syringae bacteria. Finally, selected effectors were found to localize to various plant subcellular compartments. These results indicate that X. euvesicatoria bacteria utilize multiple T3S effectors to suppress flg22-induced signaling acting downstream or in parallel to MAP kinase cascades and suggest they act through different molecular mechanisms.

scholarly journals Two Novel Type III-Secreted Proteins of Xanthomonas campestris pv. vesicatoria Are Encoded within the hrp Pathogenicity Island

2002 ◽  
Vol 184 (5) ◽  
pp. 1340-1348 ◽  
Author(s):  
Laurent Noël ◽  
Frank Thieme ◽  
Dirk Nennstiel ◽  
Ulla Bonas
Keyword(s):  
Gene Cluster ◽  
Pseudomonas Syringae ◽  
Xanthomonas Campestris ◽  
Pathogenicity Island ◽  
Effector Proteins ◽  
In Planta ◽  
Type Iii ◽  
Reverse Transcription Pcr ◽  
New Genes ◽  
Type Iii Protein Secretion

ABSTRACT The Hrp type III protein secretion system (TTSS) is essential for pathogenicity of gram-negative plant pathogen Xanthomonas campestris pv. vesicatoria. cDNA-amplified fragment length polymorphism and reverse transcription-PCR analyses identified new genes, regulated by key hrp regulator HrpG, in the regions flanking the hrp gene cluster. Sequence analysis revealed genes encoding HpaG, a predicted leucine-rich repeat-containing protein, the lysozyme-like HpaH protein, and XopA and XopD, which are similar in sequence to Hpa1 from Xanthomonas oryzae pv. oryzae and PsvA from Pseudomonas syringae, respectively. XopA and XopD (Xanthomonas outer proteins) are secreted by the Xanthomonas Hrp TTSS and thus represent putative effector proteins. Mutations in xopA, but not in xopD, resulted in reduced bacterial growth in planta and delayed plant reactions in susceptible and resistant host plants. Since the xopD promoter contains a putative hrp box, which is characteristic of hrpL-regulated genes in P. syringae and Erwinia spp., the gene was probably acquired by horizontal gene transfer. Interestingly, the regions flanking the hrp gene cluster also contain insertion sequences and genes for a putative transposase and a tRNAArg. These features suggest that the hrp gene cluster of X. campestris pv. vesicatoria is part of a pathogenicity island.

scholarly journals The Majority of the Type III Effector Inventory of Pseudomonas syringae pv. tomato DC3000 Can Suppress Plant Immunity

2009 ◽  
Vol 22 (9) ◽  
pp. 1069-1080 ◽  
Author(s):  
Ming Guo ◽  
Fang Tian ◽  
Yashitola Wamboldt ◽  
James R. Alfano
Keyword(s):  
Pseudomonas Syringae ◽  
Plant Immunity ◽  
In Planta ◽  
Tomato Dc3000 ◽  
Type Iii Effector ◽  
Type Iii Effectors ◽  
Type Iii ◽  
Molecular Pattern ◽  
Effector Triggered Immunity ◽  
Type Iii Protein Secretion

The Pseudomonas syringae type III protein secretion system (T3SS) and the type III effectors it injects into plant cells are required for plant pathogenicity and the ability to elicit a hypersensitive response (HR). The HR is a programmed cell death that is associated with effector-triggered immunity (ETI). A primary function of P. syringae type III effectors appears to be the suppression of ETI and pathogen-associated molecular pattern–triggered immunity (PTI), which is induced by conserved molecules on microorganisms. We reported that seven type III effectors from P. syringae pv. tomato DC3000 were capable of suppressing an HR induced by P. fluorescens(pHIR11) and have now tested 35 DC3000 type III effectors in this assay, finding that the majority of them can suppress the HR induced by HopA1. One newly identified type III effector with particularly strong HR suppression activity was HopS2. We used the pHIR11 derivative pLN1965, which lacks hopA1, in related assays and found that a subset of the type III effectors that suppressed HopA1-induced ETI also suppressed an ETI response induced by AvrRpm1 in Arabidopsis thaliana. A. thaliana plants expressing either HopAO1 or HopF2, two type III effectors that suppressed the HopA1-induced HR, were reduced in the flagellin-induced PTI response as well as PTI induced by other PAMPs and allowed enhanced in planta growth of P. syringae. Collectively, our results suggest that the majority of DC3000 type III effectors can suppress plant immunity. Additionally, the construct pLN1965 will likely be a useful tool in determining whether other type III effectors or effectors from other types of pathogens can suppress either ETI, PTI, or both.

Isolation of Ralstonia solanacearum hrpB constitutive mutants and secretion analysis of hrpB-regulated gene products that share homology with known type III effectors and enzymes

Microbiology ◽  
2005 ◽  
Vol 151 (9) ◽  
pp. 2873-2884 ◽  
Author(s):  
Naoyuki Tamura ◽  
Yukio Murata ◽  
Takafumi Mukaihara
Keyword(s):  
Pseudomonas Syringae ◽  
Ralstonia Solanacearum ◽  
Type Iii Secretion ◽  
Effector Proteins ◽  
Gene Products ◽  
Nudix Hydrolase ◽  
Type Iii Effectors ◽  
Type Iii ◽  
Extracellular Secretion ◽  
Culture Supernatants

The Hrp type III secretion system (TTSS) is essential for the pathogenicity of the Gram-negative plant pathogen Ralstonia solanacearum. To examine the secretion of type III effector proteins via the Hrp TTSS, a screen was done of mutants constitutively expressing the hrpB gene, which encodes an AraC-type transcriptional activator for the hrp regulon. A mutant was isolated that in an hrp-inducing medium expresses several hrpB-regulated genes 4·9–83-fold higher than the wild-type. R. solanacearum Hrp-secreted outer proteins PopA and PopC were secreted at high levels into the culture supernatants of the hrpB constitutive (hrpB c) mutant. Using hrpB c mutants, the extracellular secretion of several hrpB-regulated (hpx) gene products that share homology with known type III effectors and enzymes was examined. Hpx23, Hpx24 and Hpx25, which are similar in sequence to Pseudomonas syringae pv. tomato effector proteins HopPtoA1, HolPtoR and HopPtoD1, are also secreted via the Hrp TTSS in R. solanacearum. The secretion of two hpx gene products that share homology with known enzymes, glyoxalase I (Hpx19) and Nudix hydrolase (Hpx26), was also examined. Hpx19 is accumulated inside the cell, but interestingly, Hpx26 is secreted outside the cell as an Hrp-secreted outer protein, suggesting that Hpx19 functions intracellularly but Hpx26 is a novel effector protein of R. solanacearum.

scholarly journals The Pseudomonas syringae HopPtoV Protein Is Secreted in Culture and Translocated into Plant Cells via the Type III Protein Secretion System in a Manner Dependent on the ShcV Type III Chaperone

2004 ◽  
Vol 186 (11) ◽  
pp. 3621-3630 ◽  
Author(s):  
Misty D. Wehling ◽  
Ming Guo ◽  
Zheng Qing Fu ◽  
James R. Alfano
Keyword(s):  
Pseudomonas Syringae ◽  
Protein Secretion ◽  
Plant Cells ◽  
Secretion System ◽  
Effector Proteins ◽  
In Planta ◽  
Type Iii ◽  
Protein Secretion System ◽  
Type Iii Protein Secretion ◽  
Plant Pathogenesis

ABSTRACT The bacterial plant pathogen Pseudomonas syringae depends on a type III protein secretion system and the effector proteins that it translocates into plant cells to cause disease and to elicit the defense-associated hypersensitive response on resistant plants. The availability of the P. syringae pv. tomato DC3000 genome sequence has resulted in the identification of many novel effectors. We identified the hopPtoV effector gene on the basis of its location next to a candidate type III chaperone (TTC) gene, shcV, and within a pathogenicity island in the DC3000 chromosome. A DC3000 mutant lacking ShcV was unable to secrete detectable amounts of HopPtoV into culture supernatants or translocate HopPtoV into plant cells, based on an assay that tested whether HopPtoV-AvrRpt2 fusions were delivered into plant cells. Coimmunoprecipitation and Saccharomyces cerevisiae two-hybrid experiments showed that ShcV and HopPtoV interact directly with each other. The ShcV binding site was delimited to an N-terminal region of HopPtoV between amino acids 76 and 125 of the 391-residue full-length protein. Our results demonstrate that ShcV is a TTC for the HopPtoV effector. DC3000 overexpressing ShcV and HopPtoV and DC3000 mutants lacking either HopPtoV or both ShcV and HopPtoV were not significantly impaired in disease symptoms or bacterial multiplication in planta, suggesting that HopPtoV plays a subtle role in pathogenesis or that other effectors effectively mask the contribution of HopPtoV in plant pathogenesis.

scholarly journals The HopZ Family of Pseudomonas syringae Type III Effectors Require Myristoylation for Virulence and Avirulence Functions in Arabidopsis thaliana

2008 ◽  
Vol 190 (8) ◽  
pp. 2880-2891 ◽  
Author(s):  
Jennifer D. Lewis ◽  
Wasan Abada ◽  
Wenbo Ma ◽  
David S. Guttman ◽  
Darrell Desveaux
Keyword(s):  
Arabidopsis Thaliana ◽  
Pseudomonas Syringae ◽  
Hypersensitive Response ◽  
Pathogenic Bacteria ◽  
Functional Characterization ◽  
Defense Responses ◽  
Effector Proteins ◽  
Type Iii Effectors ◽  
Type Iii ◽  
Virulence Protein

ABSTRACT Pseudomonas syringae utilizes the type III secretion system to translocate effector proteins into plant cells, where they can contribute to the pathogen's ability to infect and cause disease. Recognition of these effectors by resistance proteins induces defense responses that typically include a programmed cell death reaction called the hypersensitive response. The YopJ/HopZ family of type III effector proteins is a common family of effector proteins found in animal- and plant-pathogenic bacteria. The HopZ family in P. syringae includes HopZ1aPsyA2, HopZ1bPgyUnB647, HopZ1cPmaE54326, HopZ2Ppi895A and HopZ3PsyB728a. HopZ1a is predicted to be most similar to the ancestral hopZ allele and causes a hypersensitive response in multiple plant species, including Arabidopsis thaliana. Therefore, it has been proposed that host defense responses have driven the diversification of this effector family. In this study, we further characterized the hypersensitive response induced by HopZ1a and demonstrated that it is not dependent on known resistance genes. Further, we identified a novel virulence function for HopZ2 that requires the catalytic cysteine demonstrated to be required for protease activity. Sequence analysis of the HopZ family revealed the presence of a predicted myristoylation sequence in all members except HopZ3. We demonstrated that the myristoylation site is required for membrane localization of this effector family and contributes to the virulence and avirulence activities of HopZ2 and HopZ1a, respectively. This paper provides insight into the selective pressures driving virulence protein evolution by describing a detailed functional characterization of the diverse HopZ family of type III effectors with the model plant Arabidopsis.

scholarly journals Naturally Occurring Nonpathogenic Isolates of the Plant Pathogen Pseudomonas syringae Lack a Type III Secretion System and Effector Gene Orthologues

2008 ◽  
Vol 190 (8) ◽  
pp. 2858-2870 ◽  
Author(s):  
Toni J. Mohr ◽  
Haijie Liu ◽  
Shuangchun Yan ◽  
Cindy E. Morris ◽  
José A. Castillo ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Type Iii Secretion ◽  
Plant Cells ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Effector Proteins ◽  
Plant Diseases ◽  
In Planta ◽  
Type Iii ◽  
Effector Gene

ABSTRACT Pseudomonas syringae causes plant diseases, and the main virulence mechanism is a type III secretion system (T3SS) that translocates dozens of effector proteins into plant cells. Here we report the existence of a subgroup of P. syringae isolates that do not cause disease on any plant species tested. This group is monophyletic and most likely evolved from a pathogenic P. syringae ancestor through loss of the T3SS. In the nonpathogenic isolate P. syringae 508 the genomic region that in pathogenic P. syringae strains contains the hrp-hrc cluster coding for the T3SS and flanking effector genes is absent. P. syringae 508 was also surveyed for the presence of effector orthologues from the closely related pathogenic strain P. syringae pv. syringae B728a, but none were detected. The absence of the hrp-hrc cluster and effector orthologues was confirmed for other nonpathogenic isolates. Using the AvrRpt2 effector as reporter revealed the inability of P. syringae 508 to translocate effectors into plant cells. Adding a plasmid-encoded T3SS and the P. syringae pv. syringae 61 effector gene hopA1 increased in planta growth almost 10-fold. This suggests that P. syringae 508 supplemented with a T3SS could be used to determine functions of individual effectors in the context of a plant infection, avoiding the confounding effect of other effectors with similar functions present in effector mutants of pathogenic isolates.

scholarly journals Functional Analysis of the Type III Effectors AvrRpt2 and AvrRpm1 of Pseudomonas syringae with the Use of a Single-Copy Genomic Integration System

2001 ◽  
Vol 14 (2) ◽  
pp. 145-155 ◽  
Author(s):  
David S. Guttman ◽  
Jean T. Greenberg
Keyword(s):  
Pseudomonas Syringae ◽  
Modular Design ◽  
Plant Cells ◽  
Single Copy ◽  
Effector Proteins ◽  
Resistance Response ◽  
Integration System ◽  
Type Iii Effectors ◽  
Genomic Integration ◽  
Type Iii

Gram-negative phytopathogenic bacteria require a type III secretion apparatus for pathogenesis, presumably to deliver Avr effector proteins directly into plant cells. To extend previous studies of Avr effectors that employed plasmids encoding Avr proteins, we developed a system that permits the integration of any gene into the Pseudomonas syringae genome in single copy. With this system, we confirmed earlier findings showing that P. syringae pv. maculicola strain PsmES4326 expressing the AvrRpt2 effector induces a resistance response in plants with the cognate R gene, RPS2. Chromosomally located avrRpt2, however, provoked a stronger resistance response than that observed with plasmid-expressed AvrRpt2 in RPS2+ plants. Additionally, chromosomal expression of AvrRpt2 conferred a fitness advantage on P. syringae grown in rps2- plants, aiding in growth within leaves and escape to leaf surfaces that was difficult to detect with plasmid-borne avrRpt2. Finally, with the use of the genomic integration system, we found that a chimeric protein composed of the N terminus of the heterologous AvrRpm1 effector and the C-terminal effector region of AvrRpt2 was delivered to plant cells. Because the C terminus of AvrRpt2 cannot translocate into plant cells on its own, this indicates that the N-terminal region can direct secretion and translocation during an infection, which supports the view that Avr proteins have a modular design. This work establishes a readily manipulatable system to study type III effectors in a biologically realistic context.

scholarly journals Virulence in a Pseudomonas syringae Strain with a Small Repertoire of Predicted Effectors

2013 ◽  
Author(s):  
Kevin L Hockett ◽  
Marc T Nishimura ◽  
Erick Karlsrud ◽  
Kevin Dougherty ◽  
David A Baltrus
Keyword(s):  
Pseudomonas Syringae ◽  
Effector Proteins ◽  
Type Iii Effector ◽  
Type Iii Effectors ◽  
Virulence Proteins ◽  
E Coli ◽  
Type Iii ◽  
Catalytic Sites ◽  
New Type ◽  
Additional Support

stract Both type III effector proteins and non-ribosomal peptide toxins play important roles for Pseudomonas syringae pathogenicity in host plants, but whether and how these virulence pathways interact to promote infection remains unclear. Genomic evidence from one clade of P. syringae suggests a tradeoff between the total number of type III effector proteins and presence of syringomycin, syringopeptin, and syringolin A toxins. Here we report the complete genome sequence from P. syringae CC1557, which contains the lowest number of known type III effectors to date and has also acquired genes similar to sequences encoding syringomycin pathways from other strains. We demonstrate that this strain is pathogenic on Nicotiana benthamiana and that both the type III secretion system and a new type III effector family, hopBJ1, contribute to virulence. We further demonstrate that virulence activity of HopBJ1 is dependent on similar catalytic sites as the E. coli CNF1 toxin. Taken together, our results provide additional support for a negative correlation between type III effector repertoires and the potential to produce syringomycin-like toxins while also highlighting how genomic synteny and bioinformatics can be used to identify and characterize novel virulence proteins.

scholarly journals HopZ4 from Pseudomonas syringae, a Member of the HopZ Type III Effector Family from the YopJ Superfamily, Inhibits the Proteasome in Plants

2014 ◽  
Vol 27 (7) ◽  
pp. 611-623 ◽  
Author(s):  
Suayib Üstün ◽  
Patrick König ◽  
David S. Guttman ◽  
Frederik Börnke
Keyword(s):  
Pseudomonas Syringae ◽  
Sequence Similarity ◽  
Bacterial Species ◽  
Inhibitory Effect ◽  
Catalytic Triad ◽  
Effector Proteins ◽  
In Planta ◽  
High Sequence Similarity ◽  
Type Iii Effector ◽  
Type Iii

The YopJ family of type III effector proteins (T3E) is one of the largest and most widely distributed families of effector proteins, whose members are highly diversified in virulence functions. In the present study, HopZ4, a member of the YopJ family of T3E from the cucumber pathogen Pseudomonas syringae pv. lachrymans is described. HopZ4 shares high sequence similarity with the Xanthomonas T3E XopJ, and a functional analysis suggests a conserved virulence function between these two T3E. As has previously been shown for XopJ, HopZ4 interacts with the proteasomal subunit RPT6 in yeast and in planta to inhibit proteasome activity during infection. The inhibitory effect on the proteasome is dependent on localization of HopZ4 to the plasma membrane as well as on an intact catalytic triad of the effector protein. Furthermore, HopZ4 is able to complement loss of XopJ in Xanthomonas spp., as it prevents precocious host cell death during a compatible Xanthomonas-pepper interaction. The data presented here suggest that different bacterial species employ inhibition of the proteasome as a virulence strategy by making use of conserved T3E from the YopJ family of bacterial effector proteins.

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