scholarly journals Characterization of Regulatory Pathways in Xylella fastidiosa: Genes and Phenotypes Controlled by gacA

2009 ◽  
Vol 75 (8) ◽  
pp. 2275-2283 ◽  
Author(s):  
Xiang Yang Shi ◽  
C. Korsi Dumenyo ◽  
Rufina Hernandez-Martinez ◽  
Hamid Azad ◽  
Donald A. Cooksey
Keyword(s):  
Pseudomonas Syringae ◽  
Biofilm Formation ◽  
Xylella Fastidiosa ◽  
Cell Aggregation ◽  
Toxin Gene ◽  
Tomato Dc3000 ◽  
Disease Symptoms ◽  
Regulatory Pathways ◽  
Physiological Processes

ABSTRACT The xylem-limited, insect-transmitted bacterium Xylella fastidiosa causes Pierce's disease in grapes through cell aggregation and vascular clogging. GacA controls various physiological processes and pathogenicity factors in many gram-negative bacteria, including biofilm formation in Pseudomonas syringae pv. tomato DC3000. Cloned gacA of X. fastidiosa was found to restore the hypersensitive response and pathogenicity in gacA mutants of P. syringae pv. tomato DC3000 and Erwinia amylovora. A gacA mutant of X. fastidiosa (DAC1984) had significantly reduced abilities to adhere to a glass surface, form biofilm, and incite disease symptoms on grapevines, compared with the parent (A05). cDNA microarray analysis identified 7 genes that were positively regulated by GacA, including xadA and hsf, predicted to encode outer membrane adhesion proteins, and 20 negatively regulated genes, including gumC and an antibacterial polypeptide toxin gene, cvaC. These results suggest that GacA of X. fastidiosa regulates many factors, which contribute to attachment and biofilm formation, as well as some physiological processes that may enhance the adaptation and tolerance of X. fastidiosa to environmental stresses and the competition within the host xylem.

scholarly journals Identification and Characterization of a Well-Defined Series of Coronatine Biosynthetic Mutants of Pseudomonas syringae pv. tomato DC3000

2004 ◽  
Vol 17 (2) ◽  
pp. 162-174 ◽  
Author(s):  
David M. Brooks ◽  
Gustavo Hernández-Guzmán ◽  
Andrew P. Kloek ◽  
Francisco Alarcón-Chaidez ◽  
Aswathy Sreedharan ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
High Performance ◽  
Tomato Dc3000 ◽  
Disease Symptoms ◽  
Coronafacic Acid ◽  
Tn5 Mutants ◽  
Cor Genes ◽  
Identification And Characterization

To identify Pseudomonas syringae pv. tomato genes involved in pathogenesis, we carried out a screen for Tn5 mutants of P. syringae pv. tomato DC3000 with reduced virulence on Arabidopsis thaliana. Several mutants defining both known and novel virulence loci were identified. Six mutants contained insertions in biosynthetic genes for the phytotoxin coronatine (COR). The P. syringae pv. tomato DC3000 COR genes are chromosomally encoded and are arranged in two separate clusters, which encode enzymes responsible for the synthesis of coronafacic acid (CFA) or coronamic acid (CMA), the two defined intermediates in COR biosynthesis. High-performance liquid chromatography fractionation and exogenous feeding studies confirmed that Tn5 insertions in the cfa and cma genes disrupt CFA and CMA biosynthesis, respectively. All six COR biosynthetic mutants were significantly impaired in their ability to multiply to high levels and to elicit disease symptoms on A. thaliana plants. To assess the relative contributions of CFA, CMA, and COR in virulence, we constructed and characterized cfa6 cmaA double mutant strains. These exhibited virulence phenotypes on A. thalliana identical to those observed for the cmaA or cfa6 single mutants, suggesting that reduced virulence of these mutants on A. thaliana is caused by the absence of the intact COR toxin. This is the first study to use biochemically and genetically defined COR mutants to address the role of COR in pathogenesis.

scholarly journals Characterization of Regulatory Pathways in Xylella fastidiosa: Genes and Phenotypes Controlled by algU

2007 ◽  
Vol 73 (21) ◽  
pp. 6748-6756 ◽  
Author(s):  
Xiang Yang Shi ◽  
C. Korsi Dumenyo ◽  
Rufina Hernandez-Martinez ◽  
Hamid Azad ◽  
Donald A. Cooksey
Keyword(s):  
Microarray Analysis ◽  
Biofilm Formation ◽  
Xylella Fastidiosa ◽  
Bacterial Pathogens ◽  
Regulatory Gene ◽  
Cell Aggregation ◽  
Wild Type ◽  
Regulatory Pathways ◽  
In The Wild ◽  
Bacterial Mutants

ABSTRACT Many virulence genes in plant bacterial pathogens are coordinately regulated by “global” regulatory genes. Conducting DNA microarray analysis of bacterial mutants of such genes, compared with the wild type, can help to refine the list of genes that may contribute to virulence in bacterial pathogens. The regulatory gene algU, with roles in stress response and regulation of the biosynthesis of the exopolysaccharide alginate in Pseudomonas aeruginosa and many other bacteria, has been extensively studied. The role of algU in Xylella fastidiosa, the cause of Pierce's disease of grapevines, was analyzed by mutation and whole-genome microarray analysis to define its involvement in aggregation, biofilm formation, and virulence. In this study, an algU::nptII mutant had reduced cell-cell aggregation, attachment, and biofilm formation and lower virulence in grapevines. Microarray analysis showed that 42 genes had significantly lower expression in the algU::nptII mutant than in the wild type. Among these are several genes that could contribute to cell aggregation and biofilm formation, as well as other physiological processes such as virulence, competition, and survival.

scholarly journals Localization and characterization of Xylella fastidiosa haemagglutinin adhesins

Microbiology ◽  
2010 ◽  
Vol 156 (7) ◽  
pp. 2172-2179 ◽  
Author(s):  
Tanja M. Voegel ◽  
Jeremy G. Warren ◽  
Ayumi Matsumoto ◽  
Michele M. Igo ◽  
Bruce C. Kirkpatrick
Keyword(s):  
Molecular Mass ◽  
Outer Membrane ◽  
Biofilm Formation ◽  
Xylella Fastidiosa ◽  
Membrane Vesicles ◽  
Cell Aggregation ◽  
Pathogenic Bacterium ◽  
Plant Pathogenic Bacterium ◽  
Gram Negative

Xylella fastidiosa is a Gram-negative, xylem-inhabiting, plant-pathogenic bacterium responsible for several important diseases including Pierce's disease (PD) of grapevines. The bacteria form biofilms in grapevine xylem that contribute to the occlusion of the xylem vessels. X. fastidiosa haemagglutinin (HA) proteins are large afimbrial adhesins that have been shown to be crucial for biofilm formation. Little is known about the mechanism of X. fastidiosa HA-mediated cell–cell aggregation or the localization of the adhesins on the cell. We generated anti-HA antibodies and show that X. fastidiosa HAs are present in the outer membrane and secreted both as soluble proteins and in membrane vesicles. Furthermore, the HA pre-proteins are processed from the predicted molecular mass of 360 kDa to a mature 220 kDa protein. Based on this information, we are evaluating a novel form of potential resistance against PD by generating HA-expressing transgenic grapevines.

scholarly journals The awr Gene Family Encodes a Novel Class of Ralstonia solanacearum Type III Effectors Displaying Virulence and Avirulence Activities

2012 ◽  
Vol 25 (7) ◽  
pp. 941-953 ◽  
Author(s):  
Montserrat Solé ◽  
Crina Popa ◽  
Oriane Mith ◽  
Kee Hoon Sohn ◽  
Jonathan D. G. Jones ◽  
...  
Keyword(s):  
Gene Family ◽  
Pseudomonas Syringae ◽  
Ralstonia Solanacearum ◽  
Tomato Dc3000 ◽  
Type Iii Effectors ◽  
Type Iii ◽  
New Gene ◽  
Transient Overexpression ◽  
Effector Recognition

We present here the characterization of a new gene family, awr, found in all sequenced Ralstonia solanacearum strains and in other bacterial pathogens. We demonstrate that the five paralogues in strain GMI1000 encode type III-secreted effectors and that deletion of all awr genes severely impairs its capacity to multiply in natural host plants. Complementation studies show that the AWR (alanine-tryptophan-arginine tryad) effectors display some functional redundancy, although AWR2 is the major contributor to virulence. In contrast, the strain devoid of all awr genes (Δawr1-5) exhibits enhanced pathogenicity on Arabidopsis plants. A gain-of-function approach expressing AWR in Pseudomonas syringae pv. tomato DC3000 proves that this is likely due to effector recognition, because AWR5 and AWR4 restrict growth of this bacterium in Arabidopsis. Transient overexpression of AWR in nonhost tobacco species caused macroscopic cell death to varying extents, which, in the case of AWR5, shows characteristics of a typical hypersensitive response. Our work demonstrates that AWR, which show no similarity to any protein with known function, can specify either virulence or avirulence in the interaction of R. solanacearum with its plant hosts.

scholarly journals Characterization of the Fur Regulon in Pseudomonas syringae pv. tomato DC3000

2011 ◽  
Vol 193 (18) ◽  
pp. 4598-4611 ◽  
Author(s):  
B. G. Butcher ◽  
P. A. Bronstein ◽  
C. R. Myers ◽  
P. V. Stodghill ◽  
J. J. Bolton ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Tomato Dc3000

scholarly journals Pseudomonas syringae pv. tomato DC3000 CmaL (PSPTO4723), a DUF1330 Family Member, Is Needed To Produce l-allo-Isoleucine, a Precursor for the Phytotoxin Coronatine

2012 ◽  
Vol 195 (2) ◽  
pp. 287-296 ◽  
Author(s):  
Jay N. Worley ◽  
Alistair B. Russell ◽  
Aaron G. Wexler ◽  
Philip A. Bronstein ◽  
Brian H. Kvitko ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Genomic Region ◽  
Tomato Dc3000 ◽  
Content Type ◽  
Type Iii Effectors ◽  
Type Iii ◽  
Coronafacic Acid ◽  
Feeding Experiments ◽  
Leaf Chlorosis

ABSTRACTPseudomonas syringaepv. tomato DC3000 produces the phytotoxin coronatine, a major determinant of the leaf chlorosis associated with DC3000 pathogenesis. The DC3000 PSPTO4723 (cmaL) gene is located in a genomic region encoding type III effectors; however, it promotes chlorosis in the model plantNicotiana benthamianain a manner independent of type III secretion. Coronatine is produced by the ligation of two moieties, coronafacic acid (CFA) and coronamic acid (CMA), which are produced by biosynthetic pathways encoded in separate operons. Cross-feeding experiments, performed inN. benthamianawithcfa,cma, andcmaLmutants, implicate CmaL in CMA production. Furthermore, analysis of bacterial supernatants under coronatine-inducing conditions revealed that mutants lacking either thecmaoperon orcmaLaccumulate CFA rather than coronatine, supporting a role for CmaL in the regulation or biosynthesis of CMA. CmaL does not appear to regulate CMA production, since the expression of proteins with known roles in CMA production is unaltered incmaLmutants. Rather, CmaL is needed for the first step in CMA synthesis, as evidenced by the fact that wild-type levels of coronatine production are restored to a ΔcmaLmutant when it is supplemented with 50 μg/mll-allo-isoleucine, the starting unit for CMA production.cmaLis found in all other sequencedP. syringaestrains with coronatine biosynthesis genes. This characterization of CmaL identifies a critical missing factor in coronatine production and provides a foundation for further investigation of a member of the widespread DUF1330 protein family.

scholarly journals Genome-Wide Gene Expression Analysis of Pseudomonas syringae pv. tomato DC3000 Reveals Overlapping and Distinct Pathways Regulated by hrpL and hrpRS

2006 ◽  
Vol 19 (9) ◽  
pp. 976-987 ◽  
Author(s):  
Lefu Lan ◽  
Xin Deng ◽  
Jianmin Zhou ◽  
Xiaoyan Tang
Keyword(s):  
Pseudomonas Syringae ◽  
Housekeeping Genes ◽  
The Novel ◽  
Tomato Dc3000 ◽  
Regulatory Pathways ◽  
Genome Wide ◽  
Genes Encoding ◽  
Substrate Proteins ◽  
Genome Wide Gene Expression ◽  
Insight Into

Pseudomonas syringae pv. tomato DC3000 is a model pathogen infecting tomato and Arabidopsis plants. Genes encoding the type III secretion system and substrate proteins (collectively called TTSS genes) of this bacterium are induced in plants and in minimal medium (MM). The induction of TTSS genes is mediated by HrpL, an alternative sigma factor recognizing the hrp box in the promoter of TTSS genes. The transcription of hrpL is activated by HrpR and HrpS, two homologous DNA-binding proteins encoded by the hrpRS operon. Microarray analysis was conducted to evaluate the DC3000 genes regulated by hrpL and hrpRS in MM. The analysis identified a number of novel hrpL-activated genes with a putative TTSS-independent function. Genes regulated by hrpL were mostly regulated by hrpRS in the same manner, but a large number of genes regulated by hrpRS were hrpL-independent, indicating that hrpL represents one branch of the regulatory pathways downstream of hrpRS. The induction of the TTSS genes was associated with downregulation of the housekeeping genes, indicating that the activation of the TTSS has a cost on the basic cellular activities. The novel genes and pathways identified by the microarray provide new insight into the bacterial functions coordinating with the TTSS.

Characterization of the PvdS-regulated promoter motif in Pseudomonas syringae pv. tomato DC3000 reveals regulon members and insights regarding PvdS function in other pseudomonads

2008 ◽  
Vol 68 (4) ◽  
pp. 871-889 ◽  
Author(s):  
Bryan Swingle ◽  
Deepti Thete ◽  
Monica Moll ◽  
Christopher R. Myers ◽  
David J. Schneider ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Tomato Dc3000 ◽  
Promoter Motif

scholarly journals Knot Formation Caused by Pseudomonas syringae subsp. savastanoi on Olive Plants Is hrp-Dependent

2004 ◽  
Vol 94 (5) ◽  
pp. 484-489 ◽  
Author(s):  
A. Sisto ◽  
M. G. Cipriani ◽  
M. Morea
Keyword(s):  
Sequence Analysis ◽  
Pseudomonas Syringae ◽  
Open Reading Frame ◽  
Gene Encoding ◽  
Disease Symptoms ◽  
Phytopathogenic Bacteria ◽  
Reading Frame ◽  
Nonhost Plant ◽  
Induced Mutant

The virulence of Pseudomonas syringae subsp. savastanoi, which causes hyperplastic symptoms (knots) on olive plants, is associated with secreted phytohormones. We identified a Tn5-induced mutant of P. syringae subsp. savastanoi that did not cause disease symptoms on olive plants although it was still able to produce phytohormones. In addition, the mutant failed to elicit a hypersensitive response in a nonhost plant. Molecular characterization of the mutant revealed that a single Tn5 insertion occurred within an open reading frame encoding a protein 92% identical to the HrcC protein of P. syringae pv. syringae. Moreover, sequence analysis revealed that the gene encoding the HrcC protein in P. syringae subsp. savastanoi was part of an operon that included five genes arranged as in other phytopathogenic bacteria. These results imply that hrp/hrc genes are functional in P. syringae subsp. savastanoi and that they play a key role in the pathogenicity of this plant pathogen.

scholarly journals Protection of Arabidopsis thaliana against Leaf-Pathogenic Pseudomonas syringae by Sphingomonas Strains in a Controlled Model System

2011 ◽  
Vol 77 (10) ◽  
pp. 3202-3210 ◽  
Author(s):  
Gerd Innerebner ◽  
Claudia Knief ◽  
Julia A. Vorholt
Keyword(s):  
Arabidopsis Thaliana ◽  
Pseudomonas Syringae ◽  
Plant Protection ◽  
Severe Disease ◽  
Model System ◽  
Tomato Dc3000 ◽  
Disease Symptoms ◽  
Content Type ◽  
Cell Densities ◽  
Symptom Formation

ABSTRACTDiverse bacterial taxa live in association with plants without causing deleterious effects. Previous analyses of phyllosphere communities revealed the predominance of few bacterial genera on healthy dicotyl plants, provoking the question of whether these commensals play a particular role in plant protection. Here, we tested two of them,MethylobacteriumandSphingomonas, with respect to their ability to diminish disease symptom formation and the proliferation of the foliar plant pathogenPseudomonas syringaepv. tomato DC3000 onArabidopsis thaliana. Plants were grown under gnotobiotic conditions in the absence or presence of the potential antagonists and then challenged with the pathogen. No effect ofMethylobacteriumstrains on disease development was observed. However, members of the genusSphingomonasshowed a striking plant-protective effect by suppressing disease symptoms and diminishing pathogen growth. A survey of differentSphingomonasstrains revealed that most plant isolates protectedA. thalianaplants from developing severe disease symptoms. This was not true forSphingomonasstrains isolated from air, dust, or water, even when they reached cell densities in the phyllosphere comparable to those of the plant isolates. This suggests that plant protection is common among plant-colonizingSphingomonasspp. but is not a general trait conserved within the genusSphingomonas. The carbon source profiling of representative isolates revealed differences between protecting and nonprotecting strains, suggesting that substrate competition plays a role in plant protection bySphingomonas. However, other mechanisms cannot be excluded at this time. In conclusion, the ability to protect plants as shown here in a model system may be an unexplored, common trait of indigenousSphingomonasspp. and may be of relevance under natural conditions.

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