scholarly journals PopP1, a New Member of the YopJ/AvrRxv Family of Type III Effector Proteins, Acts as a Host-Specificity Factor and Modulates Aggressiveness of Ralstonia solanacearum

2002 ◽  
Vol 15 (10) ◽  
pp. 1058-1068 ◽  
Author(s):  
Muriel Lavie ◽  
Edwin Shillington ◽  
Cédric Eguiluz ◽  
Nigel Grimsley ◽  
Christian Boucher
Keyword(s):  
Host Specificity ◽  
Ralstonia Solanacearum ◽  
Effector Proteins ◽  
Avirulence Gene ◽  
Type Iii ◽  
Secretion Pathway ◽  
Plant Infection ◽  
Hrp Genes ◽  
Specificity Factor ◽  
Bacterial Effectors

A functional analysis of an 11-kb-long region of the genome of the plant-pathogenic bacterium Ralstonia solanacearum, previously identified as an alternative codon usage region (ACUR), reveals that it was probably acquired through horizontal gene transfer. This ACUR encodes an insertion sequence and eight potential proteins, one of which is partially homologous with a host-specificity factor from a plant-pathogenic Erwinia sp., and another, PopP1, which is homologous to members of the YopJ/AvrRxv family of type III-secreted bacterial effectors controlling interaction between bacteria and their hosts. The analysis of mutants affecting all except one of the genes identified in the ACUR showed that only the popP1-deficient strain had an altered phenotype in plant infection tests. This mutant strain became pathogenic to a Petunia line that is resistant to the wild-type strain. Therefore, popP1 behaves as a typical avirulence gene. We demonstrate that PopP1 protein is secreted and that secretion of this protein requires a functional type III-secretion pathway. In contrast to the structural genes for other type III-secreted proteins identified in R. solanacearum, transcription of the popP1 gene is not coregulated with transcription of hrp genes but is constitutive.

scholarly journals Pseudolipasin A Is a Specific Inhibitor for Phospholipase A2 Activity of Pseudomonas aeruginosa Cytotoxin ExoU

2006 ◽  
Vol 75 (3) ◽  
pp. 1089-1098 ◽  
Author(s):  
Vincent T. Lee ◽  
Stefan Pukatzki ◽  
Hiromi Sato ◽  
Eriya Kikawada ◽  
Anastasia A. Kazimirova ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type Iii Secretion ◽  
Specific Inhibitor ◽  
Effector Proteins ◽  
Host Cells ◽  
Cytotoxic Effects ◽  
Type Iii ◽  
Secretion Pathway ◽  
Phospholipase A2 Activity

ABSTRACT A number of bacterial pathogens utilize the type III secretion pathway to deliver effector proteins directly into the host cell cytoplasm. Certain strains of Pseudomonas aeruginosa associated with acute infections express a potent cytotoxin, exoenzyme U (ExoU), that is delivered via the type III secretion pathway directly into contacting host cells. Once inside the mammalian cell, ExoU rapidly lyses the intoxicated cells via its phospholipase A2 (PLA2) activity. A high-throughput cell-based assay was developed to screen libraries of compounds for those capable of protecting cells against the cytotoxic effects of ExoU. A number of compounds were identified in this screen, including one group that blocks the intracellular activity of ExoU. In addition, these compounds specifically inhibited the PLA2 activity of ExoU in vitro, whereas eukaryotic secreted PLA2 and cytosolic PLA2 were not inhibited. This novel inhibitor of ExoU-specific PLA2 activity, named pseudolipasin A, may provide a new lead for virulence factor-based therapeutic design.

scholarly journals PopF1 and PopF2, Two Proteins Secreted by the Type III Protein Secretion System of Ralstonia solanacearum, Are Translocators Belonging to the HrpF/NopX Family

2006 ◽  
Vol 188 (13) ◽  
pp. 4903-4917 ◽  
Author(s):  
Damien Meyer ◽  
Sébastien Cunnac ◽  
Mareva Guéneron ◽  
Céline Declercq ◽  
Frédérique Van Gijsegem ◽  
...  
Keyword(s):  
Protein Secretion ◽  
Ralstonia Solanacearum ◽  
Xanthomonas Campestris ◽  
Pathogenic Bacteria ◽  
Secretion System ◽  
Effector Proteins ◽  
Experimental Conditions ◽  
Type Iii ◽  
Protein Secretion System ◽  
Type Iii Protein Secretion

ABSTRACT Ralstonia solanacearum GMI1000 is a gram-negative plant pathogen which contains an hrp gene cluster which codes for a type III protein secretion system (TTSS). We identified two novel Hrp-secreted proteins, called PopF1 and PopF2, which display similarity to one another and to putative TTSS translocators, HrpF and NopX, from Xanthomonas spp. and rhizobia, respectively. They also show similarities with TTSS translocators of the YopB family from animal-pathogenic bacteria. Both popF1 and popF2 belong to the HrpB regulon and are required for the interaction with plants, but PopF1 seems to play a more important role in virulence and hypersensitive response (HR) elicitation than PopF2 under our experimental conditions. PopF1 and PopF2 are not necessary for the secretion of effector proteins, but they are required for the translocation of AvrA avirulence protein into tobacco cells. We conclude that PopF1 and PopF2 are type III translocators belonging to the HrpF/NopX family. The hrpF gene of Xanthomonas campestris pv. campestris partially restored HR-inducing ability to popF1 popF2 mutants of R. solanacearum, suggesting that translocators of R. solanacearum and Xanthomonas are functionally conserved. Finally, R. solanacearum strain UW551, which does not belong to the same phylotype as GMI1000, also possesses two putative translocator proteins. However, although one of these proteins is clearly related to PopF1 and PopF2, the other seems to be different and related to NopX proteins, thus showing that translocators might be variable in R. solanacearum.

scholarly journals Genome-Wide Identification of a Large Repertoire of Ralstonia solanacearum Type III Effector Proteins by a New Functional Screen

2010 ◽  
Vol 23 (3) ◽  
pp. 251-262 ◽  
Author(s):  
Takafumi Mukaihara ◽  
Naoyuki Tamura ◽  
Masaki Iwabuchi
Keyword(s):  
Ralstonia Solanacearum ◽  
Type Iii Secretion ◽  
Plant Cells ◽  
Amino Acid Sequences ◽  
Effector Proteins ◽  
Type Iii ◽  
Functional Screen ◽  
Genome Wide ◽  
Large Repertoire ◽  
Insertion Mutants

The gram-negative plant-pathogenic bacterium Ralstonia solanacearum utilizes the hypersensitive response and pathogenicity (Hrp) type III secretion system (T3SS) to cause disease in plants. To determine the entire repertoire of effector proteins possessed by R. solanacearum RS1000, we constructed a transposon carrying a calmodulin-dependent adenylate cyclase reporter that can be used to specifically detect rip (Ralstonia protein injected into plant cells) genes by monitoring the cAMP level in plant leaves inoculated with insertion mutants. From the new functional screen using this transposon, we identified 38 new Rip proteins translocated into plant cells via the Hrp T3SS. In addition, most of the 34 known effectors of RS1000 could be detected by the screen, except for three effectors that appear to be small in size or only weakly expressed. Finally, we identified 72 Rips in RS1000, which include 68 effector proteins classified into over 50 families and four extracellular components of the Hrp T3SS. Interestingly, one-third of the effectors are specific to R. solanacearum. Many effector proteins contain various repeated amino acid sequences or known enzyme motifs. We also show that most of the R. solanacearum effector proteins, but not Hrp extracellular components, require an Hrp-associated protein, HpaB, for their effective translocation into plant cells.

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 A Pseudomonas syringae pv. tomato DC3000 Hrp (Type III Secretion) Deletion Mutant Expressing the Hrp System of Bean Pathogen P. syringae pv. syringae 61 Retains Normal Host Specificity for Tomato

2003 ◽  
Vol 16 (1) ◽  
pp. 43-52 ◽  
Author(s):  
Derrick E. Fouts ◽  
Jorge L. Badel ◽  
Adela R. Ramos ◽  
Ryan A. Rapp ◽  
Alan Collmer
Keyword(s):  
Host Range ◽  
Resistance Gene ◽  
Host Specificity ◽  
Pseudomonas Syringae ◽  
Type Iii Secretion ◽  
Secretion System ◽  
Effector Proteins ◽  
Tomato Dc3000 ◽  
Tomato Plants ◽  
Type Iii

The plant pathogenic species Pseudomonas syringae is divided into numerous pathovars based on host specificity. For example, P. syringae pv. tomato DC3000 is pathogenic on tomato and Arabidopsis, whereas P. syringae pv. syringae 61 is pathogenic on bean. The ability of P. syringae strains to elicit the hypersensitive response (HR) in non-hosts or be pathogenic (or parasitic) in hosts is dependent on the Hrp (type III secretion) system and effector proteins this system is thought to inject into plant cells. To test the role of the Hrp system in determining host range, the hrp/hrc gene cluster (hrpK through hrpR) was deleted from DC3000 and complemented in trans with the orthologous cluster from strain 61. Mutant CUCPB5114 expressing the bean pathogen Hrp system on plasmid pCPP2071 retained the ability of wild-type DC3000 to elicit the HR in bean, to grow and cause bacterial speck in tomato, and to elicit a cultivar-specific (gene-for-gene) HR in tomato plants carrying the Pto resistance gene. However, the symptoms produced in compatible tomato plants involved markedly reduced chlorosis, and CUCPB5114(pCPP2071) did not grow or produce symptoms in Arabidopsis Col-0 although it was weakly virulent in NahG Arabidopsis. A hypersensitive-like collapse was produced by CUCPB5114(pCPP2071) in Arabidopsis Col-0 at 1 × 107 CFU/ml, but only if the bacteria also expressed AvrB, which is recognized by the RPM1 resistance gene in Col-0 and confers incompatibility. These observations support the concept that the P. syringae effector proteins, rather than secretion system components, are the primary determinants of host range at both the species and cultivar levels of host specificity.

scholarly journals Identification of novel Ralstonia solanacearum type III effector proteins through translocation analysis of hrpB-regulated gene products

Microbiology ◽  
2009 ◽  
Vol 155 (7) ◽  
pp. 2235-2244 ◽  
Author(s):  
Takafumi Mukaihara ◽  
Naoyuki Tamura
Keyword(s):  
Ralstonia Solanacearum ◽  
Pathogenic Bacteria ◽  
Tandem Repeats ◽  
Plant Cells ◽  
Secretion System ◽  
Effector Proteins ◽  
Gene Products ◽  
Type Iii Effector ◽  
Type Iii ◽  
Effector Genes

The Hrp type III secretion system (TTSS) is essential for the pathogenicity of Ralstonia solanacearum on host plants. Hrp TTSS is a specialized secretion system that injects virulence proteins, the so-called type III effector proteins, into plant cells. In R. solanacearum, the expression of Hrp TTSS-related genes is regulated by an AraC-type transcriptional activator, HrpB. We have identified 30 hrpB-regulated hpx ( hrpB-dependent expression) genes and three well-known hrpB-regulated genes, popA, popB and popC, as candidate effector genes in R. solanacearum strain RS1000. In this study, we newly cloned 11 additional candidate effector genes that share homology with known hpx genes from R. solanacearum RS1000. Using a Cya reporter system, we investigated the translocation of these 44 gene products into plant cells via the Hrp TTSS and identified 34 effector proteins. These include three effector families composed of more than four members, namely the Hpx4, Hpx30 and GALA families. The Hpx30 family effectors are 2200–2500 aa in size and appear to be the largest class of effector proteins among animal- and plant-pathogenic bacteria. Members of this family contain 12–18 tandem repeats of a novel 42 aa motif, designated SKWP repeats.

scholarly journals A Draft Genome Sequence of Pseudomonas syringae pv. tomato T1 Reveals a Type III Effector Repertoire Significantly Divergent from That of Pseudomonas syringae pv. tomato DC3000

2009 ◽  
Vol 22 (1) ◽  
pp. 52-62 ◽  
Author(s):  
Nalvo F. Almeida ◽  
Shuangchun Yan ◽  
Magdalen Lindeberg ◽  
David J. Studholme ◽  
David J. Schneider ◽  
...  
Keyword(s):  
Host Range ◽  
Host Specificity ◽  
Pseudomonas Syringae ◽  
Genome Sequence ◽  
Draft Genome ◽  
Draft Genome Sequence ◽  
Tomato Dc3000 ◽  
Type Iii ◽  
Host Range Determination ◽  
Range Determination

Diverse gene products including phytotoxins, pathogen-associated molecular patterns, and type III secreted effectors influence interactions between Pseudomonas syringae strains and plants, with additional yet uncharacterized factors likely contributing as well. Of particular interest are those interactions governing pathogen-host specificity. Comparative genomics of closely related pathogens with different host specificity represents an excellent approach for identification of genes contributing to host-range determination. A draft genome sequence of Pseudomonas syringae pv. tomato T1, which is pathogenic on tomato but nonpathogenic on Arabidopsis thaliana, was obtained for this purpose and compared with the genome of the closely related A. thaliana and tomato model pathogen P. syringae pv. tomato DC3000. Although the overall genetic content of each of the two genomes appears to be highly similar, the repertoire of effectors was found to diverge significantly. Several P. syringae pv. tomato T1 effectors absent from strain DC3000 were confirmed to be translocated into plants, with the well-studied effector AvrRpt2 representing a likely candidate for host-range determination. However, the presence of avrRpt2 was not found sufficient to explain A. thaliana resistance to P. syringae pv. tomato T1, suggesting that other effectors and possibly type III secretion system–independent factors also play a role in this interaction.

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